vol.44 no.2 june 2010 pemetreksed Pravi Izbor Pravi Bolnik Pravi Cas Edina histološko usmerjena citostaticna terapija ALIMTA/cisplatin: Zdravljenje prvega reda pri bolnikih z nedrobnocelicnim pljucnim karcinomom, ki nimajo pretežno luskaste histologije Edina kombinirana terapija s signifikantno izboljšanim preživetjem: 12,6 meseca pri bolnikih z adenokarcinomom pljuc1 SKRAJŠAN POVZETEK GLAVNIH ZNACILNOSTI ZDRAVILA Ime zdravila ALIMTA 100 mg prašek za koncentrat za raztopino za infundiranje in ALIMTA 500 mg prašek za koncentrat za raztopino za infundiranje Kakovostna in kolicinska sestava ALIMTA 100 mg: vsaka viala vsebuje 100 mg pemetrekseda (v obliki dinatrijevega pemetrekseda). Po pripravi vsebuje vsaka viala 25 mg/ml pemetrekseda. Pomožne snovi: Vsaka viala vsebuje približno 11 mg natrija. Manitol, klorovodikova kislina, natrijev hidroksid. ALIMTA 500 mg: vsaka viala vsebuje 500 mg pemetrekseda (v obliki dinatrijevega pemetrekseda). Po pripravi vsebuje vsaka viala 25 mg/ml pemetrekseda. Pomožne snovi: Vsaka viala vsebuje približno 54 mg natrija. Manitol, klorovodikova kislina, natrijev hidroksid. Terapevtske indikacije: ALIMTA je v kombinaciji s cisplatinom indicirana za zdravljenje bolnikov z neresektabilnim malignim plevralnim mezoteliomom, ki jih še nismo zdravili s kemoterapijo. ALIMTA je v kombinaciji s cisplatinom indicirana kot zdravljenje prvega izbora za bolnike z lokalno napredovalim ali metastatskim nedrobnocelicnim pljucnim karcinomom, ki nima pretežno luskaste celicne histologije. ALIMTA je indicirana kot monoterapija za zdravljenje lokalno napredovalega ali metastatskega nedrobnocelicnega pljucnega karcinoma, ki nima pretežno luskaste celicne histologije pri bolnikih, pri katerih bolezen ni napredovala neposredno po ke­moterapiji na osnovi platine. Zdravljenje prvega izbora naj bo platinasta dubleta z gemcitabinom, paklitakselom ali docetakselom. ALIMTA je indicirana kot monoterapija za zdravljenje drugega izbora bolnikov z lokalno napredovalim ali metastatskim nedrobnocelicnim pljucnim karcinomom, ki nima pretežno luskaste celicne histologije. Odmerjanje in nacin uporabe: ALIMTO smemo dajati le pod nadzorom zdravnika, usposobljenega za uporabo kemoterapije za zdravljenje raka. ALIMTA v kombinaciji s cisplatinom Priporoceni odmerek ALIMTE je 500 mg/m2 telesne površine (TP), dan kot intravenska infuzija v 10 minutah prvi dan vsakega 21-dnevnega ciklusa. Priporoceni odmerek cisplatina je 75 mg/m2 TP, infundiran v dveh urah približno 30 minut po zakljucku infuzije pemetrekseda prvi dan vsakega 21 dnevnega ciklusa. Priporoceni odmerek cisplatina je 75 mg/m2 TP, infundiran v dveh urah približno 30 minut po zakljucku infuzije pemetrekseda prvi dan vsakega 21 dnevnega ciklusa. Bolniki morajo prejeti zadostno antiemeticno zdravljenje, pred in/ali po prejemanju cisplatina jih moramo tudi ustrezno hidrirati. ALIMTA kot samostojno zdravilo Priporoceni odmerek ALIMTE je 500 mg/m2 TP, dan kot intravenska infuzija v 10 minutah prvi dan vsakega 21 dnevnega ciklusa. Režim premedikacije Da zmanjšamo incidenco in resnost kožnih reakcij, dajemo kortikosteroid dan pred dajan­jem pemetrekseda, na dan dajanja pemetrekseda in naslednji dan. Kortikosteroid naj ustreza 4 mg deksametazona, danega peroralno dvakrat dnevno. Za zmanjšanje toksicnosti morajo bolniki dnevno jemati tudi peroralno folno kislino ali multivitaminski pripravek, ki jo vsebuje (350 do 1000 mikrogramov). V sedmih dneh pred prvim odmerkom pemetrek­seda morajo vzeti vsaj pet odmerkov folne kisline, odmerjanje pa morajo nadaljevati ves cas zdravljenja in še 21 dni po zadnjem odmerku pemetrekseda. Bolniki morajo prejeti tudi intramuskularno injekcijo vitamina B12 (1000 mikrogramov) v tednu pred prvim odmerkom pemetrekseda in enkrat vsake tri cikluse zatem. Kasnejše injekcije vitamina B12 lahko dajemo isti dan kot pemetreksed. Kontraindikacije: Preobcutljivost za zdravilno ucinkovino ali katerokoli pomožno snov. Dojenje. Socasno cepljenje proti rumeni mrzlici. Posebna opozorila in previdnostni ukrepi: Pemetreksed lahko zavre delovanje kostnega mozga, kar se kaže kot nevtropenija, trombocitopenija in anemija (ali pancitopenija). Pri bolnikih, ki pred zdravljenjem niso prejemali kortikosteroidov, so porocali o kožnih reakcijah. Uporabe pemetrekseda pri bolnikih z ocistkom kreatinina < 45 ml/min ne priporocamo. Bolniki z blagim do zmernim popušcanjem delovanja ledvic naj se izogibajo jemanju nesteroidnih protivnetnih zdravil (NSAID), denimo, ibuprofena in acetilsalicilne kisline 2 dni pred dajanjem pemetrekseda, na dan dajanja in še 2 dni po dajanju pemetrekseda. Vsi bolniki, ki jih lahko zdravimo s pemetreksedom, naj se izogibajo jemanju NSAID-ov z dolgimi razpolovnimi casi izlocanja vsaj 5 dni pred dajanjem pemetrekseda, na dan dajanja in še vsaj 2 dni po dajanju pemetrekseda. Porocali so o resnih ledvicnih primerih, vkljucno z akutno ledvicno odpovedjo, s pemetreksedom samim ali v povezavi z drugimi kemoterapevtiki. Pri bolnikih s klinicno pomembno tekocino tretjega prostora moramo razmisliti o drenaži izliva pred dajanjem pemetrekseda. Kot posledico toksicnosti pemetrekseda v kombinaciji s cisplatinom za prebavila so opažali hudo dehidracijo, zato moramo bolnike pred prejemanjem terapije in/ali po njej ustrezno hidrirati, prejeti morajo zadostno antiemeticno zdravljenje. Obcasno so v klinicnih študijah pemetrekseda, obicajno ob socasnem dajanju z drugo citotoksicno ucinkovino, porocali o resnih srcnožilnih dogodkih, vkljucno z miokardnim infarktom in možganskožilnimi dogodki. Odsvetujemo uporabo živih oslabljenih cepiv. Spolno zrelim moškim odsvetujemo zaploditev otroka v casu zdravljenja in še 6 mesecev zatem. Priporocamo ukrepe prosti zanositvi ali vzdržnost. Zaradi možnosti, da zdravljenje s pemetreksedom povzroci trajno neplodnost, naj se moški pred zacetkom zdravljenja posvetujejo o shranjevanju semena. Ženske v rodni dobi morajo v casu zdravljenja s pemetreksedom uporabljati ucinkovito kontracepcijo. Porocali so o primerih radiacijske pljucnice pri bolnikih, ki so jih zdravili z radiacijo pred, med ali po zdravljenju s pemetreksedom. Porocali so o radiacijskem izpušcaju pri bolnikih, ki so se zdravili z radioterapijo pred tedni ali leti. Zdravilo Alimta 500 mg vsebuje približno 54 mg natrija na vi-alo. Pomembno za bolnike, ki so na dieti z nadzorovanim vnosom natrija. Medsebojno delovanje z drugimi zdravili in druge oblike interakcij: Socasno dajanje nefrotoksicnih zdravil (denimo, aminoglikozidov, diuretikov zanke, spojin platine, ciklosporina) lahko potencialno povzroci zakasnjeni ocistek pemetrekseda. Socasno dajanje snovi, ki se tudi izlocajo s tubulno sekrecijo (denimo, probenecid, penicilin), lahko potencialno povzroci zakasnjeni ocistek pemetrekseda. Pri bolnikih z normalnim delovanjem ledvic lahko visoki odmerki nesteroidnih protivnetnih zdravil (NSAID-i, denimo, ibuprofen) in acetilsalicilna kislina v visokih odmerkih zmanjšajo eliminacijo pemetrekseda in tako lahko povecajo pojavnost neželenih ucinkov pemetrekseda. Pri bolnikih z blagim do zmernim popušcanjem delovanja ledvic se moramo izogibati socasnemu dajanju pemetrekseda z NSAID-i (denimo, ibuprofenom) ali acetilsalicilne kisline v visokih odmerkih 2 dni pred dajanjem pemetrekseda, na dan dajanja in še 2 dni po dajanju pemetrekseda. Socasnemu dajanju NSAID-ov z daljšimi razpolovnimi casi s pemetreksedom se moramo izogibati vsaj 5 dni pred dajanjem pemetrekseda, na dan dajanja in še vsaj 2 dni po dajanju pemetrekseda.Velika razlicnost med posamezniki v koagulacijskem statusu v casu bolezni ter možnost medsebojnega delovanja med peroralnimi antikoagulacijskimi ucinkovinami ter kemoter­apijo proti raku zahtevata povecano pogostnost spremljanja INR. Kontraindicirana socasna uporaba: Cepivo proti rumeni mrzlici: tveganje za smrtno generalizirano bolezen po cepljenju. Odsvetovana socasna uporaba: Živa oslabljena cepiva (razen proti rumeni mrzlici): tveganje za sistemsko, potencialno smrtno bolezen. Neželeni ucinki Klinicne študije malignega plevralnega mezotelioma Zelo pogosto: znižani nevtrofilci/granulociti, znižani levkociti, znižan hemoglobin, znižani trombociti, nevropatija-senzoricna, diareja, bruhanje, stomatitis/faringitis, slabost, anoreksija, zaprtje, izpušcaj, alopecija, povišan kreatinin, znižan ocistek kreatinina, utrujenost. Pogosti: dehidracija, motnje okusa, konjuktivitis, dispepsija. Klinicne študije nedrobnocelicnega pljucnega karcinoma - ALIMTA monoterapija, zdravljenje 2. izbora: Zelo pogosti: znižan nevtrofilci/granulociti, znižani levkocitit, znižan hemoglobin, diareja, bruhanje, stomatitis/faringitis, slabost, anoreksija, izpušcaj/lušcenje, utrujenost. Pogosti: znižani trombociti, zaprtje, povišanje SGPT (ALT), povišanje SGOT (AST), srbenje, alopecija, povišana telesna temperatura. Klinicne študije nedrobnocelicnega pljucnega karcinoma - ALIMTA v kombinaciji s cisplatinom, zdravljenje 1. izbora: Zelo pogosti: znižan hemoglobin, znižani nevtrofilci/granulociti, znižani levkocitit, znižani trombociti, slabost, bruhanje, anoreksija, zaprtje, stomatitis/faringitis, diareja brez kolostomije, alopecija, izpušcaj/lušcenje, povišan kreatinin. Pogosti: nevropatija-senzoricna, motnje okusa, dispepsija/zgaga. Klinicne študije nedrobnocelicnega pljucnega karcinoma - ALIMTA monoterapija, vzdrževalno zdravljenje: Zelo pogosti: znižan hemoglobin, slabost, anoreksija, utrujenost, izpušcaj/lušcenje, utrujenost. Pogosti: infekcija, znižani levkociti, znižani nevtrofilci, nevropatija-senzoricna, bruhanje, mukozitis/stomatitis, diareja, povišanje ALT (SGPT), povišanje AST (SGOT). Obcasno so v klinicnih študijah pemetrekseda porocali o primerih resnih srcnožilnih in možganskožilnih dogodkov, vkljucno z miokardnim infarktom, angino pektoris, cerebrovaskularnim insultom in prehodnimi ishemicnimi atakami; primerih kolitisa ter o primerih intersticijske pljucnice z respiratorno insuficience, primerih edema in o ezofagitisu/radiacijskem ezofagitisu. Redkeje pa o primerih potencialno resnega hepatitisa in pancitopenije. Po uvedbi zdravila na trg so porocali o primerih akutne odpovedi ledvic s pemetreksedom samim ali v povezavi z drugimi kemoterapevtiki, primerih radiacijske pljucnice pri bolnikih, ki so jih zdravili z radiacijo pred, med ali po njihovem zdravljenju s pemetreksedom, primerih radiacijskega izpušcaja pri bolnikih, ki so se v preteklosti zdravili z radioterapijo in o primerih periferne ishemije, ki je vcasih vodila v nekrozo okoncin. Imetnik dovoljenja za promet Eli Lilly Nederland B.V., Grootslag 1 5, NL 3991 RA, Houten, Nizozemska. Datum zadnje revizije besedila 21.09.2009. Nacin izdaje zdravila: H Podrobnejše informacije o zdravilu Alimta, so na voljo na lokalnem predstavništvu SIALM00005 Eli Lilly Farmacevtska družba, d.o.o. Brnciceva 41G, 1231 Ljubljana - Crnuce, Slovenija Telefon: +386 (0)1 5800 010 Faks: +386 (0)1 5691 705 Publisher Association of Radiology and Oncology Affiliated with Slovenian Medical Association – Slovenian Association of Radiology, Nuclear Medicine Society,Slovenian Society for Radiotherapy and Oncology, and Slovenian Cancer Society Croatian Medical Association – Croatian Society of RadiologySocietas Radiologorum HungarorumFriuli-Venezia Giulia regional groups of S.I.R.M. Italian Society of Medical Radiology Aims and scope Radiology and Oncology is a journal devoted to publication of original contributions in diagnostic and interventionalradiology, computerized tomography, ultrasound, magnetic resonance, nuclear medicine, radiotherapy, clinical andexperimental oncology, radiobiology, radiophysics and radiation protection. Editor-in-Chief Gregor SeršaLjubljana, Slovenia Executive Editor Viljem KovacLjubljana, Slovenia Deputy Editors Andrej Cör Izola, Slovenia Igor KocijancicLjubljana, Slovenia Editorial Board Advisory Committee Karl H. Bohuslavizki Hamburg, Germany Damir Miletic Rijeka, Croatia Marija Auersperg Ljubljana, Slovenia Maja Cemažar Ljubljana, Slovenia Maja Osmak Zagreb, Croatia Tomaž Benulic Ljubljana, Slovenia Christian Dittrich Vienna, Austria Branko Palcic Vancouver, Canada Jure Fettich Ljubljana, Slovenia Metka Filipic Ljubljana, Slovenia Dušan Pavcnik Portland, USA Valentin Fidler Ljubljana, Slovenia Tullio Giraldi Trieste, Italy Geoffrey J. Pilkington Portsmouth, UK Berta Jereb Ljubljana, Slovenia Maria Godény Budapest, Hungary Ervin B. Podgoršak Montreal, Canada Vladimir Jevtic Ljubljana, Slovenia Vassil Hadjidekov Sofia, Bulgaria Uroš Smrdel Ljubljana, Slovenia Stojan Plesnicar Ljubljana, Slovenia Marko Hocevar Ljubljana, Slovenia Maksimilijan Kadivec Ljubljana, Slovenia Miklós Kásler Budapest, Hungary Primož Strojan Ljubljana, SloveniaBorut Štabuc Ljubljana, SloveniaRanka Štern-Padovan Zagreb, Croatia Mirjana Rajer Ljubljana, SloveniaŽiva Zupancic Ljubljana, Slovenia Michael Kirschfink Heidelberg, Germany Justin Teissié Toulouse, France Janko Kos Ljubljana, Slovenia Sándor Tóth Orosháza, Hungary Tamara Lah Turnšek Ljubljana, Slovenia Gillian M. Tozer Sheffield, UK Damijan Miklavcic Ljubljana, Slovenia Andrea Veronesi Aviano, Italy Luka Milas Houston, USA Branko Zakotnik Ljubljana, Slovenia Radiol Oncol 2010; 44(2): A. Editorial office Radiology and Oncology Institute of Oncology Zaloška 2SI-1000 Ljubljana Slovenia Phone: +386 1 5879 369Phone/Fax: +386 1 5879 434E-mail: gsersa@onko-i.si Copyright © Radiology and Oncology. All rights reserved. Reader for English Vida Kološa Secretary Mira KlemencicZvezdana Vukmirovic Design Monika Fink-Serša, Samo Rovan, Ivana Ljubanovic Layout Matjaž Lužar Printed byTiskarna Ozimek,Slovenia Published quarterly in 600 copies Beneficiary name: DRUŠTVO RADIOLOGIJE IN ONKOLOGIJEZaloška cesta 2,1000 LjubljanaSlovenia Beneficiary bank account number: SI56 02010-0090006751IBAN: SI56 0201 0009 0006 751Our bank name: Nova Ljubljanska banka, d.d., Ljubljana, Trg republike 2,1520 Ljubljana; SloveniaSWIFT: LJBASI2X Subscription fee for institutions EUR 100, individuals EUR 50 The publication of this journal is subsidized by the Slovenian Research Agency. Indexed and abstracted by: Science Citation Index Expanded (SciSearch®)Journal Citation Reports/Science EditionScopusEMBASE/Excerpta MedicaOpen J-gateChemical AbstractsBiomedicina Slovenica This journal is printed on acid- free paper On the web: ISSN 1581-3207 http://versita.com/science/medicine/ro/http://www.onko-i.si/radioloncol/ Radiol Oncol 2010; 44(2): B. contents contents review 67 Radiotherapy in combination with vascular-targeted therapies Eva Ciric and Gregor Sersa 79 Hyperhomocysteinemia and the role of B vitamins in cancer Nadja Plazar and Mihaela Jurdana experimental radiology 86 Hemostatic efficacy of chitosan-based bandage for closure of percutaneous arterial access sites: An experimental study in heparinized sheep model Pawanrat Kranokpiraksa, Dusan Pavcnik, Hideaki Kakizawa, Barry T. Uchida, Miran Jeromel, Frederick S. Keller, Josef Rösch radiology 92 3T MRI in evaluation of asbestos-related thoracic diseases – preliminary results Janez Podobnik, Igor Kocijancic, Viljem Kovac, Igor Sersa 97 Reliability of diffusion weighted MR imaging in differentiating degenerative and infectious end plate changes Ozgur Oztekin, Cem Calli, Omer Kitis, Zehra Hilal Adibelli, Cemal Suat Eren, Melda Apaydin, Mehmet Zileli,Taskin Yurtseven 103 Post-traumatic high-flow priapism treated by endovascular embolization using N-butyl-cyanoacrylate Marko Rados, Vice Sunjara, Ivica Sjekavica, Ranka Stern Padovan Radiol Oncol 2010; 44(2): C. contents clinical oncology 107 Antigen expression on recurrent meningioma cells Andrej Vranic 113 Frequent MGMT (06-methylguanine-DNA methyltransferase) hypermethylation in long-term survivors of glioblastoma: a single institution experience Martina Baur, Matthias Preusser, Maria Piribauer, Katarzyna Elandt, Marco Hassler, Marcus Hudec, Christian Dittrich, Christine Marosi 121 A clinical case of the penile metastasis from the rectal carcinoma Mehmet Yildirim, Ali Coskun, Mete Pürten, Ozgur Oztekin, Enver Ilhan radiophysics 124 A fully electronic intensity-modulated radiation therapy quality assurance (IMRT QA) process implemented in a network comprised of independent treatment planning, record and verify, and delivery systems Daniel W Bailey, Lalith Kumaraswamy, Matthew B Podgorsak non-ionizing radiation 131 Effect of 905 MHz microwave radiation on colony growth of the yeast Saccharomyces cerevisiae strains FF18733, FF1481 and D7 Ivana Vrhovac, Reno Hrascan, Jasna Franekic I slovenian abstracts VIII notices Radiology and oncology is covered in Science Citation Index Expanded (SciSearch®),Journal Citation Reports/Science Edition, Scopus, EMBASE/Excerpta Medica, Open J-gate, Chemical Abstracts, Biomedicina Slovenica Radiol Oncol 2010; 44(2): D. review Radiotherapy in combination with vascular-targeted therapies Eva Ciric and Gregor Sersa Institute of Oncology Ljubljana, Ljubljana, Slovenia Received 11 December 2009 Accepted 20 April 2010 Correspondence to: Prof. Gregor Serša, Ph.D., Institute of Oncology Ljubljana, Department of Experimental Oncology, Zaloška 2, SI-1000 Ljubljana, Slovenia. E-mail: gsersa@onko-i.si Disclosure: No potential conflicts of interest were disclosed. Background. Given the critical role of tumor vasculature in tumor development, considerable efforts have been spent on developing therapeutic strategies targeting the tumor vascular network. A variety of agents have been developed, with two general approaches being pursued. Antiangiogenic agents (AAs) aim to interfere with the proc­ess of angiogenesis, preventing new tumor blood vessel formation. Vascular-disrupting agents (VDAs) target existing tumor vessels causing tumor ischemia and necrosis. Despite their great therapeutic potential, it has become clear that their greatest clinical utility may lie in combination with conventional anticancer therapies. Radiotherapy is a widely used treatment modality for cancer with its distinct therapeutic challenges. Thus, combining the two approaches seems reasonable. Conclusions. Strong biological rationale exist for combining vascular-targeted therapies with radiation. AAs and VDAs were shown to alter the tumor microenvironment in such a way as to enhance responses to radiation. The results of preclinical and early clinical studies have confirmed the therapeutic potential of this new treatment strategy in the clinical setting. However, concerns about increased normal tissue toxicity, have been raised. Key words: antiangiogenic agents; vascular-disrupting agents; radiotherapy Introduction Radiotherapy is an effective and widely used treat­ment modality for many tumors, with about half of all cancer patients undergoing radiation therapy as a part of their treatment.1Although widely used, tumor radioresistance remains a major problem and a need exists to improve the cure rate by radia­tion therapy alone. As the patient population treat­ed with radiotherapy is so enormous, enhancing the therapeutic outcome for even a relatively small proportion of these has the potential to translate to a highly significant clinical benefit. Combinations of cytotoxic chemotherapeutic agents with radia­tion have a synergistic effect on tumor response and are firmly established in clinical practice for a wide spectrum of tumors.2In recent years, there has been increasing interest in combining vascular-targeted therapies with radiation.3The enhanced antitumor efficacy of combined treatment may be explained by the alteration of the tumor microen­vironment by vascular-targeted agents resulting in increased radiosensitivity of the tumor. However, the mechanisms of interaction between the two treatment modalities are complex and involve in­teractions between tumor stroma, the vasculature and the tumor cells themselves, which are not cur­rently well understood. Therefore, the ideal way to use this potentially powerful combination for tu­mor cure has yet to be determined. Tumor angiogenesis Angiogenesis is a critical step in tumor progression, as tumors are unable to grow beyond 2 mm3with­out a vascular supply, due to lack of oxygen and nutrients.4Formation of new blood vessels occurs from pre-existing vessels and allows the tumor to grow and expand rapidly.5Tumors switch in their development to an angiogenic phenotype. The 68 Ciric E and Sersa G / Vascular-targeted therapies in radiotherapy transition from dormant to the angiogenic state of the tumor is termed the “angiogenic switch” and is caused by a shift in the balance of anti- and pro-an­giogenic factors.6It is regulated by environmental factors and by genetic alterations that act to either up-regulate pro-angiogenic factors, such as vas­cular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) and/or down-regu­late inhibitors of angiogenesis, such as angiostatin, endostatin, thrombospondin and interferons.7 The multistep process of tumor angiogenesis is characterized by degradation of the extracellular matrix, followed by proliferation and migration of the underlying endothelial cells into the tumor, with resultant vessel formation.5The initial step in the process is activation of quiescent endothelial cells by binding of tumor-produced or stromal-produced growth factors to endothelial receptors. VEGF is a potent and specific growth factor that plays a pivotal role in endothelial cell activation.8The main effects of VEGF are to increase vessel permeability and induce endothelial cell migration and proliferation, leading to the formation of en­dothelial sprouts, which then anastomose to form vascular loops and networks.9,10VEGF also acts as a survival factor for endothelial cells by inhib­iting apoptosis.11It is therefore a pivotal driver of tumor angiogenesis, allowing tumor progression from in situ lesions to widespread disease, and providing the tumor with a route via which cells can get into the circulation and form distant me­tastases.4,12VEGF is secreted by almost all solid tu­mors.13Proliferating endothelial cells found in and around tumors produce multiple growth factors that not only promote endothelial cell growth but also tumor cell growth, invasion, and survival.14,15Angiogenesis therefore provides both a perfusion effect and a paracrine effect for a growing tumor and tumor cells and endothelial cells can drive each other with resultant perpetuation and ampli­fication of the malignant phenotype.16 Newly formed tumor blood vessels are distinct from those of normal tissue. They are markedly dis­ordered, often dilated, tortuous and characterized by a relative lack of pericytes and other supporting cells, impaired blood flow and increased vascular permeability.17Extravasation of macromolecules and pertinent development of high interstitial fluid pressure often results in vascular collapse, which leads to acidic and hypoxic areas heterogeneously distributed within the tumor mass.18,19Hypoxia re­sulting from such functional vessel abnormalities is termed “acute” or “perfusion-limited”. The af­fected tumor cells are found much closer to blood vessels than would be expected from diffusion lim­itations and are exposed to oxygen concentrations that vary transiently between normal, anoxia and anywhere in-between. On the other hand, “chron­ic” or “diffusion-limited” hypoxia is found at an increased distance from blood vessels. In this type of hypoxia, individual perfused vessels are char­acterized by an oxygenation gradient surrounding them. Cells in this area exist at all possible oxygen concentrations ranging from anoxia at distant loca­tions to normal values next to the vessels.20 Hypoxia, angiogenesis and radioresistance Hypoxia is an important stimulus for angiogen­esis.21Hypoxia inducible factor-1 (HIF-1) is a ma­jor mediator of the response to hypoxia. It is a transcriptional factor that regulates a number of processes, including VEGF transcription, apop­tosis and cell cycle arrest.22,23HIF-1 is regulated mainly by hypoxia, but it can also be activated in response to radiation.24Both the hypoxic tumor mi-croenvironment and external stresses such as ion­izing radiation, lead to the up-regulation of many other pro-angiogenic factors, including VEGF, angiopoietin-2, nitric oxide synthase, platelet-de­rived growth factor (PDGF) and basic fibroblastic growth factors (bFGF).25,26It has been shown that radiotherapy alone can potentiate angiogenic proc­esses.27Increased VEGF production in response to irradiation has been observed in various cancer cell lines.28This is a part of the overall cellular response to stress and it is associated with the induction of a variety of transcription factors that can activate transcription of cytokines, growth factors, and cell cycle-related genes. Hypoxia in tumors is strongly associated with radiation resistance as oxygen is required to chem­ically modify free-radical damage to the target DNA. When radiation is absorbed by the tissue, it creates reactive oxygen species that react with and damage cellular DNA, thus triggering cell death by apoptosis and/or necrosis. Cells irradiated in the presence of air are about three times more sensitive than cells irradiated under conditions of severe hy­poxia.29Pre-treatment measurements of tumor ox­ygenation have been shown to predict the response to radiotherapy and the likelihood of tumor recur­rence, progression and metastatic disease in many human tumors.30A more moderate hypoxia than is needed for maximum resistance to radiation has al­ Ciric E and Sersa G / Vascular-targeted therapies in radiotherapy 69 so been shown to have a negative impact on tumor control. This may be due to the fact that hypoxia influences a number of biological responses that af­fect tumor properties important for the treatment outcome, including angiogenesis.20Different levels of hypoxia in a tumor thus provide the conditions for existence of viable cells that are not only radio-resistant but angiogenic as well.31,32 Vascular-targeted therapies The importance of targeting tumor vasculature development and function first became apparent in the 1970s through the seminal studies of Judah Folkman, who demonstrated that angiogenesis is crucial for the growth and survival of tumor cells. His findings suggest that both tumor cells and their supporting endothelial cells are potential tar­gets for cell killing and should be considered when planning cancer treatment.4Destroying the tumor vasculature deprives tumors of nutrients and oxy­gen necessary for their growth and should also inhibit metastatic spread, theoretically leading to tumor regression. As a therapeutic group, vascular-targeting agents are unique as they have highly specific tar­gets, while simultaneously having the potential to be effective against a broad range of tumor types. They are now divided into two classes; antiang­iogenic agents (AAs), which inhibit the formation of new blood vessels, and vascular-disrupting agents (VDAs), which act against existing tumor vasculature. AAs are considered to be cytostatic in nature in contrast to VDAs, which are thought to be cytotoxic. Although there are differences be­tween the two groups, including their administra­tion schedules, individual agents might show both antiangiogenic and vascular-disrupting effects.33 Antiangiogenic agents AAs aim to prevent the growth of new blood ves­sels in tumors. One of the most widely studied tar­gets for angiogenesis being explored clinically is VEGF and its receptors. VEGF is a ligand with a central role in signaling pathways controlling tu­mor blood vessel development and survival. The binding of VEGF ligands activates receptor tyro­sine kinases, designated VEGFR1, VEGFR2 and VEGFR3, which in turn activate a network of dis­tinct downstream signaling pathways. Although the effects of VEGF receptors (VEGFR) signaling were initially thought to be specific for the vascu­lature, VEGF can also play a role in many other processes.34VEGFR1 expression by colon cancer cells contributes to colon cancer cell motility and invasiveness but has little direct effect on prolif­eration of these cells. VEGFR2 expression by lung cancer cells may play a role in tumor cell survival after cytotoxic stress.35,36Many different strategies for inhibiting VEGF activity have been evaluated, including the neutralization of the ligand or recep­tor by antibodies, blocking VEGFR signaling with tyrosine kinase inhibitors and even antiangiogenic gene therapy based on modulating the expression of VEGF pathway-related genes.34,37 Bevacizumab is a humanized monoclonal anti­body that acts by binding and neutralizing VEGF. In a pivotal clinical trial conducted by Hurwtzet al., bevacizumab in combination with fluorouracil-based chemotherapy, significantly improved the overall survival for patients with metastatic color-ectal cancer over chemotherapy alone.38Improved overall survival with combination therapy was al­so shown for patients with NSCLC and improved progression-free survival for patients with meta­static breast cancer and renal cell cancer was ob­served.39-41 Small molecule tyrosine kinase inhibitors (TKIs) present another class of antiangiogenic agents. They act by preventing activation of growth factor receptors, thus inhibiting downstream signaling pathways. They offer the theoretical advantage of being simultaneously active against receptors for different growth factors. Sunitinib, for example, targets VEGFRs, platelet-derived growth factor re­ceptor (PDGFR) and c-kit and has shown signifi­cant efficacy in clinical trials for renal cancer.42 So far improvements in overall survival have only been seen in patients with colorectal and non-small cell lung cancers, when AAs were given in combination with chemotherapy. One possible reason why single-agent AAs ultimately fail is that there is up-regulation of other pro-angiogenic fac­tors leading to angiogenesis and tumor resistance, hence the rationale for these drugs to be combined with chemotherapy or radiotherapy.43 Vascular-disrupting agents VDAs cause a rapid shutdown of perfusion in the established tumor vasculature, leading to tumor cell ischemia and secondary tumor cell death. These agents have the potential to destroy existing 70 Ciric E and Sersa G / Vascular-targeted therapies in radiotherapy tumor masses and may be therefore particularly suitable for treating large tumors, which are typi­cally resistant to conventional therapies.33 Two major classes of VDAs that selectively tar­get tumor vessels are in clinical development; the ligand-directed VDAs and small molecule VDAs. Biological or ligand-directed VDAs work by using antibodies, peptides or growth factors which selec­tively bind to the endothelium. Coagulation and/or endothelial cell death is then achieved by coupling the vascular-targeting moiety with a toxin (e.g. ri­cin) or a pro-coagulant.44Small molecule VDAs are at a much more advanced stage of clinical devel­opment than ligand-based therapies. These agents work by inducing vascular collapse, leading to ex­tensive necrosis in tumors and include flavonoids and tubulin-depolymerizing/binding agents.33Flavone acetic acid and its derivatives, particularly 5,6-dimethyl-xanthenone-4-acetic acid (DMXAA), have a complex mechanism of action and are be­lieved to work by inducing the release of vasoac­tive agents and cytokines, such as tumor necrosis factor alpha (TNF-a), which leads to hemorrhagic necrosis.45The tubulin-binding agents (e.g. com-bretastatin A-4 disodium phosphate) are believed to work by selective disruption of the cytoskeleton in proliferating endothelial cells in tumors. The subsequent change in endothelial cell shape leads to vessel blockage, thrombus formation, rapid re­duction in tumor blood flow, and secondary tumor necrosis.46 Recently, electrochemotherapy has been recog­nized to have a vascular-disrupting effect besides a direct cytotoxic effect on tumor cells.47,48Due to non-selective permeabilization of cells in the tu­mors exposed to electric pulses, endothelial cells also undergo apoptosis by uptake of bleomycin or cisplatin.48,49This leads to permanent blood flow abrogation of the affected vessels leading to tumor hypoxia and necrosis, similar as in other vascular-disrupting agents.50It has been estimated that the vascular-disrupting effect contributes 20-30% to the overall antitumor effectiveness of electroche­motherapy.48,49 The result of selective vascular destruction com­mon to all of these strategies is extensive central tumor necrosis that leaves only a thin layer of vi­able cells at the tumor periphery. These cells are believed to obtain nutrients and oxygen from ves­sels of the surrounding normal tissue and their re­population may be the cause of treatment failure when VDAs are used in monotherapy, therefore combining VDAs with other standard treatment is an obvious option.33 Combined treatments As oxygen is crucial for maximal effectiveness of radiation, a logical concern when combining AAs and VDAs with radiation would be that compro­mising tumor vasculature by these agents would leave a tumor hypoxic and, thus, less radiosensi­tive. However, the mechanisms of interaction be­tween the two treatment modalities have proved to be more complex and involve changes in the tumor microenvironment that may in fact result in an im­proved treatment outcome.51 Radiotherapy and antiangiogenic agents The understanding that tumor micro environmen­tal factors, such as hypoxia, promote up-regulation of angiogenic and survival pathways leading to increased radioresistance, and that radiotherapy it­self has pro-angiogenic effects, has prompted stud­ies combining AAs with radiation. Teicher’s group was the first to show that AAs increase the tumor response when combined with single dose radiotherapy.52,53A number of preclini-cal studies have since indicated that AAs can en­hance the response to radiation (Table 1). The list of AAs evaluated in combination with radiation in­clude non-specific antiangiogenic agent angiosta-tin, agents targeting the VEGF signaling pathway (anti-VEGF, anti-VEGFR antibodies and tyrosine kinase inhibitors), COX-2 inhibitors and epidermal growth factor receptor (EGFR) inhibitors which also target tumor cells. The antiangiogenic and an-titumor effects have been reported to be additive as well as synergistic.71Lee et al.conducted important animal experiments using an anti-VEGF antibody in combination with radiotherapy, resulting in synergistic antitumor effects. The anti-VEGF an­tibody decreased tumor interstitial fluid pressure and increased tumor perfusion, probably due to an observed reduction of tumor vascular density with vessel reorganization.59In addition, AAs have been shown to increase oxygenation, thus increas­ing overall radiosensitivity. Jain tried to reconcile the paradoxical effects of AAs on oxygenation with the concept of “normalization” of the tumor vas­culature.72He postulated that rather than obliter­ating all tumor blood vessels, AAs destroyed only immature vessels, reduced vascular permeability and interstitial fluid pressure, and increased peri­cyte recruitment to stabilize intact vessels. Such normalization of tumor vasculature resulted in a more stable, organized vasculature, which could Ciric E and Sersa G / Vascular-targeted therapies in radiotherapy 71 TAble 1. Preclinical combination trials with antiangiogenic agents and radiotherapy TNP-470 Angiostatin Endostatin Anti-VEGF antibody VEGFR-2 blockade SU5416 DC101 VEGFR tyrosine kinase inhibitors PTK787/ZK222584 ZD6474 AZD2171 Multi-kinase inhibitors SU11248 (sunitinib) SU6668 Lewis lung carcinoma C3H mammary carcinoma U87 glioblastoma Lewis lung carcinoma D54 human glioblastoma SQ-20B squamous cell carcinoma Lewis lung carcinoma SQ-20B squamous cell carcinoma Seg-1 esophageal adenocarcinoma U87 glioblastoma LS1747 colon adenocarcinoma Seg-1 esophageal adenocarcinoma U87 glioblastoma GL261 murine glioblastoma 54A small cell lung cancer U87 glioblastoma MCa4 mammary carcinoma MCa35 mammary carcinoma SW480 human colon adenocarcinoma CaLu-6 non-small cell lung cancer HT49 colorectal carcinoma H460 non-small cell lung cancer CaLu-6 non-small cell lung cancer LoVo colorectal carcinoma Lewis lung carcinoma GL261 murine glioblastoma Lewis lung carcinoma GL261 murine carcinoma 52, 53 54 55 56 56 57 58 58 58 58, 59 59 60 59 61 62 62 63 63 64 65 66 67 68 69 69 69 70 70 deliver oxygen and nutrients to the tumor more efficiently via well-functioning vessels, thereby decreasing hypoxia and hence radioresistance (Figure 1). However, continued antiangiogenic ac­tivity could cause vessel regression and impaired delivery leading to exacerbation of hypoxic condi­tions and radioresistance. Benefits of such a combi­nation therapy may therefore be dependent upon a transient “normalization window” of opportunity when blood flow and tumor oxygenation are in­creased.73 Optimal timing for delivery of antiangiogenic therapy during the course of radiation to achieve the greatest enhancement of the radiation re­sponse, remains unknown and few studies have compared different sequences of radiation therapy and AAs.74Recently, ZD6474 (vandetanib), a small molecule inhibitor of VEGFR2 with additional ac­tivity against EGFR, was combined with radiation therapy in the treatment of tumor xenografts. Two combination schedules were examined with van-detanib administered before each dose of radiation (concurrent schedule) or 30 minutes after the last dose of radiotherapy (sequential schedule). The growth delay induced using the concurrent sched­ule was greater than that induced by vandetanib or radiation treatment alone but the sequential sched­ule maximally delayed tumor growth. The authors demonstrated that a less pronounced response in the concurrent schedule was due to reduced tu­mor vascular perfusion causedby administration of vandetanib, which impaired re-oxygenation between radiation fractions, thereby decreasing radiosensitivity. In addition, the enhanced effect of vandetanib and radiotherapy in the sequen­tial schedule could be explained by abrogation of VEGF-dependent survival signaling, which is sup­posed to have an important role in tumor recovery after irradiation.65 The enhancement of the effect of radiation ther­apy by antiangiogenic therapy may be also influ­enced by the tumor microenvironment. This was shown in a study by Lund et al.who treated mice with glioblastoma xenografts implanted into the 72 Ciric E and Sersa G / Vascular-targeted therapies in radiotherapy A C BD FIguRe 1. Theoretical model explaining the biological rationale for combining radiotherapy and AAs. A) Abnormal tumor vasculature largely composed of immature, disordered, often dilated and tortuous blood vessels is characterized by increased vascular permeability and impaired blood flow which leads to functional vessel abnormalities resulting in hypoxic areas in the tumor. B) After irradiation, oxygenated cells are destroyed, leaving behind the radioresistant hypoxic cells which release proangiogenic factors and further promote angiogen­esis. During the time between radiation fractions hypoxic cells partly reoxygenate and further stimulate tumor repopulation, ultimately resulting in a moderate response to fractionated radiation. C) Pretreatment with AA destroys immature, inefficient tumor vessels and cause vessel reorganization thus increasing tumor perfusion and oxygenation. D) With irradiation many radiosensitive oxygenated cells are killed. The few remaining hypoxic cells reoxy­genate, without angiogenesis being increased. The result is a less pronounced tumor repopulation and better overall response to fractionated radiation. Radiol Oncol 2010; 44(2): 66-78. Ciric E and Sersa G / Vascular-targeted therapies in radiotherapy 73 TAble 2. Preclinical combination trials with vascular-disrupting agents and radiotherapy Tumor necrosis factor Flavone acetic acid DMXAA Combretastatin A-4 disodium phosphate ZD6126 MN-029 MCA-K mammary carcinoma 76 MCA-K mammary carcinoma 77 C3H mammary carcinoma 78 RIF-1 fibrosarcoma 79 MDAH-MCa4 mammary carcinoma 79 C3H mammary carcinoma 80 KHT sarcoma 80 KHT sarcoma 81 Carcinoma NT 82 C3H mammary carcinoma 83 KHT sarcoma 83 Kaposi’s sarcoma 84 Rhabdomyosarcoma 85 C3H mammary carcinoma 86 A549 NSCLC 87 U87 glioblastoma 88 KHT sarcoma 89 KHT sarcoma 90 thigh or intracranially with TNP-470 and/or radia­tion therapy.55Significant enhancement of the tu­mor response to TNP-470 and radiation was seen in the thigh tumors, but no additive effect was observed in intracranial tumors. The authors pro­posed that differences in the capillary beds and mi-croenvironment of the brain and the subcutaneous tissues of the thigh may have contributed to the differences in response. Radiotherapy and vascular-disrupting agents The presence of a viable rim of tumor cells at the periphery after VDA treatment, as shown in pre-clinical studies, explains the modest tumor con­trol seen in the single-agent phase I studies.30It has been suggested that increased blood flow in the adjacent normal tissue, together with probable rapid up-regulation of angiogenic factors, such as VEGF, directly facilitates growth and expansion of the remaining rim of viable cells.75 These cells are believed to be well oxygenated and thus present an excellent target for conventional cytotoxic thera­pies. A logical rationale for combining VDAs with radiation would therefore be the interaction of the two treatments at the tumor microregional level; VDA reducing or eliminating the poorly oxygen­ated and hence radioresistant subpopulation of tu­mor cells and radiation killing the remaining well oxygenated peripheral cells (Figure 2). A number of pre-clinical studies performed on rodent tumor models over the past few years have reported en­hanced tumor killing when VDAs were given in combination with radiotherapy (Table 2). A study by Murata et al.showed the importance of scheduling.83In his study for the murine CH3 tumors no improvement in local control was seen when combretastatin A-4 disodium phosphate was given 60 minutes before radiation compared to improved results when given concurrently or after radiotherapy. A likely explanation for this finding is that the vascular shutdown induced by the VDAs may have rendered some tumor cells hy­poxic at the time of irradiation and that these cells later re-oxygenated and survived. It was suggested that blood flow needs to be re-established in the remaining viable tissue to obtain maximum radio-sensitization of the tumor. The greatest enhance­ment of the radiation response in fractionated dose regimens may be achieved when VDA is admin­istered within a few hours after radiation. Under such conditions, antitumor effects may be greater than additive.91An interesting animal study con­ducted by Siemann and Rojiani using the tubulin-binding agent N-acetyl-colchinol (ZD6126) and radiation showed that enhanced killing was more likely in larger tumors than in smaller ones.89This observation may be explained by the fact that larg­er tumors are less radiosensitive due to increased hypoxic regions, which can be compensated by 74 Ciric E and Sersa G / Vascular-targeted therapies in radiotherapy FIguRe 2. Schematic representation of the rationale for combining radiotherapy and VDAs. The result of VDA treatment is selective destruction of tumor vessels which causes extensive central tumor necrosis leaving only a thin layer of viable cells at the tumor periphery. These cells are believed to obtain nutrients and oxygen from vessels of the surrounding normal tissue and their repopulation may be the cause of treatment failure when VDAs are used in monotherapy. Combined treatment of VDA with radiotherapy may be more successful as radiation can destroy the viable tumor rim of well oxygenated and thus radiosensitive peripheral tumor cells remaining after the use of VDA. VDAs, whereas smaller tumors are more radiosen­sitive with fewer areas affected by VDAs. Studies combining electrochemotherapy with tumor irradiation were also performed. The poten­tiation of the radiation response in experimental tumors was demonstrated with a single dose and fractionated radiation regime. A potentiating effect of 2.7 was observed with single dose irradiation and 4.6 with the fractionated regime.92-95 The effect of combined treatment was also demonstrated on tubal dedifferentiated papillary adenocarcinoma skin metastases.96 An enhanced radiation response with this treatment modality can be explained in part by radiosensitization of tumor cells that occurs in the process of electropermeabilization leading to increased uptake of radiosensitizing chemothera­peutic drugs, and in part by a vascular-disrupting effect, which is a result of electrochemotherapy as described in the previous section. A therapeutic approach combining VDAs and radiotherapy may therefore be particularly suit­able for treating larger tumors. The greatest an-titumor effect may be achieved by administering VDA after radiation fractions. However, in order to determine the optimal treatment schedule in the course of fractionated radiation, further investiga­tions are needed. Clinical trials on radiation and vascular-targeted therapies The agents most widely explored in clinical tri­als are AAs targeting VEGF and its receptors. Of many explored in clinical trials, three have been approved for clinical use; two small molecule TKIs in monotherapy (sorafenib, sunitinib) for meta- Radiol Oncol 2010; 44(2): 66-78. Ciric E and Sersa G / Vascular-targeted therapies in radiotherapy 75 TAble 3. Clinical trials of vascular-targeted agents in combination with chemoradiation/radiation therapy Bevacizumab (B) I poor-prognosis head and neck cancer II glioblastoma multiforme after surgery II locally advanced rectal cancer I/II locally advanced inoperable colorectal cancer II locally advanced inoperable pancreatic cancer NSCLC Sunitinib (S) CA4P I oligometastatic cancer advanced NSCLC chemoradiotherapy + B 102 temozolomide + radiotherapy + B . 103 temozolomide + B standard preoperative chemoradiotherapy 104 + B chemoradiotherapy + B 105 chemoradiotherapy + B . maintenance 106 chemotherapy + B chemoradiotherapy + B 107 IGRT + S . maintenance S 108 palliative radiotherapy + CA4P 109 NSCLC = non-small cell lung cancer, IGRT = image-guided radiotherapy, CA4P = combretastatin A-4 disodium phosphate static renal and hepatocellular carcinoma and an anti-VEGF monoclonal antibody (bevacizumab) in combination with chemotherapy for metastatic colorectal cancer, NSCLC and breast cancer.97-101Today none of these agents is approved in combi­nation with radiation therapy. However, several phase I and II clinical trials have been concluded and numerous are ongoing (Table 3). Many of the trials have showed a promising antitumor re­sponse. However, increased toxicity, such as fistula formation, wound healing problems and throm­bosis, have been observed in some studies, espe­cially when the VEGF inhibitor was combined with chemoradiotherapy protocols.102,107 VDAs are in a less advanced stage of clinical development, with only a few early trials con­cluded, mainly evaluating VDAs in monotherapy or chemotherapy combinations.109-111Currently, the most widely explored VDA in clinical trials is combretastatin A-4 disodium phosphate, which has already been evaluated in several Phase I trials evaluating dosage schedules and toxicity, and has recently entered Phase II trials in combination with chemotherapy, radiation and radioisotopes.51 Conclusion Advances in the understanding of tumor biology have led to development of novel antitumor agents targeting tumor vasculature. Initial clinical trials testing these agents in monotherapy were some­what disappointing and it has now become clear, that in most advanced malignancies, vasculature-targeting strategies will be most effective when used in combination with conventional anticancer therapies. Preclinical experiments on animal tumor models using different AAs and VDAs revealed possible mechanisms responsible for the syner­gistic antitumor effects of radiation and vascular-targeting strategies, based on AAs/VDAs altering the tumor microenvironment in such a way as to enhance responses to radiation therapy. The im­portance of treatment sequencing has been demon­strated in these preclinical studies. Several early clinical trials combining AAs with radiation have showed the potential benefits of this treatment strategy in the clinical setting, warrant­ing further investigations. However, the potential for higher rates of normal tissue toxicity has been documented, particularly in trials where AAs were combined with chemoradiotherapy. This indicates the need for careful design of future clinical trials with optimal radiotherapy planning and delivery in order to minimize damage to normal tissues. It might be prudent to first evaluate in early trials the combination of AAs/VDAs with radiotherapy alone. Further attention should be placed on the doses of AAs/VDAs, as currently there is little data suggesting that higher doses are necessarily better at enhancing the radiation response. Conventional strategies for monitoring anticancer therapies may not apply for vascular-targeted agents and clinical trials need to be designed not only to determine if the agents are safe and have evidence of efficacy, 76 Ciric E and Sersa G / Vascular-targeted therapies in radiotherapy but also to validate both invasive and noninvasive surrogates of response. 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Preclinical evaluations of therapies combin­ing the vascular targeting agent combretastatin A-4 disodium phosphate and conventional anticancer therapies in the treatment of Kaposi’s sar­coma. Acta Oncol 2002; 41: 91-7. 85. Ahmed B, Landuyt W, Griffioen AW, van Oosterom A, van den Bogaert W, Lambin P. In vivo antitumor effect of combretastatin A-4 phosphate added to fractionated radiation. Anticancer Res 2006; 26: 307-10. 86. Horsman MR, Murata R. Vascular targeting effects of ZD6126 in a C3H mouse mammary carcinoma and the enhancement of radiation response. Int J Radiat Oncol Biol Phys 2003; 57: 1047-55. 87. Raben D, Bianco C, Damiano V, Bianco R, Melisi D, Mignogna C, et al. Antitumor activity of ZD6126, a novel vascular-targeting agent, is enhanced when combined with ZD1839, an epidermal growth factor receptor tyrosine kinase inhibitor, and potentiates the effects of radiation in a hu­man nonsmall cell lung cancer xenograft model. Mol Cancer Ther 2004; 3: 977-83. 88. 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Electroporation of LPB sarcoma cells in vitro and tumors in vivo increases radiosensitizing effect of cisplatin. Anticancer Res 2003; 23: 275-82. 94. Kranjc S, Grosel A, Cemazar M, Sentjurc M, Sersa G. Improvement of com­bined modality therapy with bleomycin and radiation using electroporation of LPB sarcoma cells and tumors in mice. BMC Cancer 2005; 5: 115. 95. Kranjc S, Tevz G, Kamensek U, Vidic S, Cemazar M, Sersa G. Radiosensitizing effect of electrochemotherapy in a fractionated radiation regime in ra­diosensitive murine sarcoma and radioresistant adenocarcinoma tumor model. Radiat Res 2009; 172: 677-85. 96. Sersa G, Cemazar M, Rudolf Z, Fras AP. Adenocarcinoma skin metastases treated by electrochemotherapy with cisplatin combined with radiation. Radiol Oncol 1999; 33: 291-6. 97. Tamaskar I, Pili R. Update on novel agents in renal cell carcinoma. Expert Rev Anticancer Ther 2009; 9: 1817-27. 98. Zhu AX, Duda DG, Sahani DV, Jain RK. Development of sunitinib in hepa­tocellular carcinoma: rationale, early clinical experience, and correlative studies. Cancer J 2009; 15: 263-8. 99. Ocvirk J. Advances in the treatment of metastatic colorectal carcinoma. Radiol Oncol 2009; 43: 1-8. 100.Pallis AG, Serfass L, Dziadziusko R, van Meerbeeck JP, Fennell D, Lacombe D, et al. Targeted therapies in the treatment of advanced/metastatic NSCLC. Eur J Cancer 2009; 45: 2473-87. 101.Yang SX. Bevacizumab and breast cancer: current therapeutic progress and future perspectives. Expert Rev Anticancer Ther 2009; 9: 1715-25. 102.Seiwert TY, Haraf DJ, Cohen EE, Stenson K, Witt ME, Dekker A, et al. Phase I study of bevacizumab added to fluorouracil-and hydroxyurea-based concomitant chemoradiotherapy for poor-prognosis head and neck cancer. J Clin Oncol 2008; 26: 1732-41. 103.Lai A, Filka E, McGibbon B, Nghiemphu PL, Graham C, Yong WH, et al. Phase II pilot study of bevacizumab in combination with temozolomide and regional radiation therapy for up-front treatment of patients with newly diagnosed glioblastoma multiforme: interim analysis of safety and toler­ability. Int J Radiat Oncol Biol Phys 2008; 71: 1372-80. 104.Willett CG, Duda DG, di Tomaso E, Boucher Y, Ancukiewicz M, Sahani DV, et al. Efficacy, safety, and biomarkers of neoadjuvant bevacizumab, radiation therapy, and fluorouracil in rectal cancer: a multidisciplinary phase II study. J Clin Oncol 2009; 27: 3020-6. 105.Koukourakis MI, Giatromanolaki A, Sheldon H, Buffa FM, Kouklakis G, Ragoussis I, et al. Tumor and Angiogenesis Research Group. Phase I/II trial of bevacizumab and radiotherapy for locally advanced inoperable colorec­tal cancer: vasculature-independent radiosensitizing effect of bevacizumab. Clin Cancer Res 2009; 15: 7069-76. 106.Crane CH, Winter K, Regine WF, Safran H, Rich TA, Curran W, et al. Phase II study of bevacizumab with concurrent capecitabine and radiation fol­lowed by maintenance gemcitabine and bevacizumab for locally advanced pancreatic cancer: Radiation Therapy Oncology Group RTOG 0411. J Clin Oncol 2009; 27: 4096-102. 107.Spigel DR, Hainsworth JD, Yardley DA, Raefsky E, Patton J, Peacock N, et al. Tracheoesophageal fistula formation in patients with lung cancer treated with chemoradiation and bevacizumab. J Clin Oncol 2010; 28: 43-8. 108.Kao J, Packer S, Vu HL, Schwartz ME, Sung MW, Stock RG, et al. Phase 1 study of concurrent sunitinib and image-guided radiotherapy followed by maintenance sunitinib for patients with oligometastases: acute toxicity and preliminary response. Cancer 2009; 115: 3571-80. 109.Ng QS, Goh V, Carnell D, Meer K, Padhani AR, Saunders MI, et al. Tumor antivascular effects of radiotherapy combined with combretastatin a4 phosphate in human non-small-cell lung cancer. Int J Radiat Oncol Biol Phys 2007; 67: 1375-80. 110.Gridelli C, Rossi A, Maione P, Rossi E, Castaldo V, Sacco PC, et al. Vascular disrupting agents: a novel mechanism of action in the battle against non-small cell lung cancer. Oncologist 2009; 14: 612-20. 111.Heath VL, Bicknell R. Anticancer strategies involving the vasculature. Nat Rev Clin Oncol 2009; 6: 395-404. review Hyperhomocysteinemia and the role of B vitamins in cancer Nadja Plazar and Mihaela Jurdana College of Health Care Izola, University of Primorska, Izola, Slovenia Received 28 October 2009 Accepted 16 March 2010 Correspondence to: Nadja Plazar, PhD, University of Primorska, College of Health Care Izola, Polje 42, 6310 Izola, Slovenia. Phone: +386 5 662 64 60; Fax: +386 5 662 64 80; E-mail: nadja.plazar@vszi.upr.si Disclosure: No potential conflicts of interest were disclosed. Background. Patients suffering from malignancies have increased complications due to corresponding cardiovas­cular diseases and risk factor for the development of venous thromboembolism. Epidemiological studies have shown that increased homocysteine plasma concentration (hyperhomocysteinemia) is related to a higher risk of coronary heart disease, stroke, peripheral vascular disease and malignancies. Homocysteine (tHcy) is an intermediate sulfur-containing amino acid produced from methionine during processing of dietary proteins. The plasma homocysteine levels are strongly influenced by diet, as well as by genetic factors. Folic acid, vitamins B6 and B12 are dietary compo­nents which influence the plasma homocysteine levels the most. Several studies have found that high blood levels of B vitamins are related to the integrity and function of DNA, and, are at least related to lower concentration of homo-cysteine. Folate depletion has been found to change DNA methylation and DNA synthesis in both animal and human studies. Because of this critical role of folate, most studies including homocysteine have focused on these two actions. Conclusions. Hyperhomocysteinemia proves to be the most common condition highly associated with both venous and arterial thrombosis in many cancer patients, while the associated pathophysiology has not been precisely es­tablished yet. Therefore, of current interest is the possible role of folate metabolism developing into a cancer initiating hyperhomocysteinemia. This review will discuss this possibility. Key words: homocysteine; hyperhomocysteinemia; B vitamins; cancer Introduction It has been long postulated that the plasma ho-mocysteine concentration is inversely related to the occurrence of cardiovascular and cerebrovas­cular diseases.1More recently, increased plasma homocysteine concentration has been postulated as a risk factor for cancer and even as a novel tu­mour marker.2This increased risk can be attrib­uted to the high prevalence of classical factors in these patients, such as hypertension, diabetes, and dyslipidemia, but most certainly (also) to factors resulting from the malignant disease and the ap­plied selected therapy. For example, back in 1865 Trousseau described hypercoagulability and in­creasing risk of »spontaneous coagulation« in pa­tients with cancer.3Nowadays, it is established that breast, pancreas, and gastrointestinal cancers are associated with a higher incidence of thrombo­sis. With more advanced stages of cancer there is a lower overall survival rate4, but, also agreater risk of venous thromboembolism5, what can addition­ally influences on the survival of patients. A raised plasma homocysteine level is associated with se­rum B vitamins concentration, especially folate levels, since these are required in homocysteine metabolism. Adequate B vitamins intake is essen­tial for nucleotide biosynthesis, DNA replication and methyl group supply and thus for cell growth and repair.6Evidence suggests that folate deple­tion fosters the development of cancer, particularly colorectal cance6-10, whereas high doses of folic acid may enhance growth of cancer cells.7However, the complexity of the folate metabolic pathway may suggest that different metabolites of folate might induce multiple effects in normal, preneoplastic and malignantcells. 80 Plazar N and Jurdana M / Hyperhomocysteinemia and the role of B vitamins in cancer COOH COOH COOH homocystine homocysteine cysteine-homocysteine FIGURE 1. Structural formulae: Homocysteine, homocystine and mixed disulfide (cysteine-homocysteine). co-factor. Another pathway of homocysteine me- Homocysteine, metabolism and tabolism is the re-methylation pathway, which cardiovascular complications is connected with the folate metabolic pathway. It involves the transfer of a methyl group from Proliferating cells secrete more homocysteine com-5-methyl-tetrahydrofolate to homocysteine to form pared to non-proliferating cells. Homecysteine is methionine, and eventually S-adenosylmethionine. a sulphur-containing intermediate in the normal The methyl transfer from 5-methyl-tetrahydro-metabolism of the essential amino acid methionine folate to homocysteine is catalyzed by methio-present in almost all body cells and mostly 5 to nine-synthase, and requires vitamin B12 as a 10% of daily synthesized homocysteine (1.2 mmol/cofactor (Figure 2). Important to notice is that day)11is transferred into the blood through hepa-S-adenosylmethionine is the universal methyl tocytes. The thyol group of homocysteine makes donor for methylation reactions. The resulting it readily available to be oxidized in the blood at tetrahydrofolate transfers into the 5.10 methyltet-physiological pH upon which it forms disulfide rahydrofolate with the enzyme 5.10 methyltetrahy-bonds with other thyols (Figure 1).12drofolate reductase (MTHFR) and then into the 5 In a healthy population the frequency of mod-methyltetrahydrofolate (5-MTHF).18Cellular avail-erate hyperhomocysteinemia (12-30 µmol/L) is ability of 5-MTHF may be of great importance in 5 to 7%, with higher values for men being attrib-regulating cellular effects of homocysteine related uted to gender differences like estrogen presence to cell growth. Therefore, deficiencies of folate and in women. This is confirmed by the fact that after vitamin B12 and reduced activity of the involved menopause the blood levels of homocysteine of metabolic enzymes will inhibit the breakdown of woman approximate those in men.13Another cause homocysteine, which will lead to an increase of the for moderate hyperhomocysteinemia is an unbal-intracellular homocysteine concentration.19anced diet with suboptimal intake of vitamins (B6, Moreover, hereditary causes of increased homo­B12 and folates), acting as coenzymes in the me-cysteine blood concentrations exist (hyperhomo­tabolism of homocysteine.14,15In the elderly, such cysteinemia). Most studies refer to changes in the a moderate hyperhomocysteinemia due to lack of genes for those enzymes that lead to severe hyper-vitamins B and folates is very common. A survey, homocysteinemia, such as the CBS gene (cistation-ß carried out by Herrmann et al., showed that 32% of syntases) or in the GCT gene (. cistationase), both healthy elderly people aged 65 to 75, and 58% of coding the trans-sulfuration pathway. Further, those over 85 years of age suffer from hyperhomo-mutations do occur in genes coding for enzymes cysteinemia, indicating that hyperhomocysteine-involved in the remethylation pathway and the re-mia significantly increases with age16, therefore it lated folate metabolic pathway. For a homozygous decreases in younger people as the incidence of person with a mutation MTHFR 677C => T the en­malignancies.17zyme activity is reduced to 35% of the normal.20A Via the trans-sulfuration pathway homocysteine typical mutation in Europe occurs within the gene is converted into cystathionine to form cysteine by for MTHFR 677C => T, with different incidences be-cystathionine-ß-synthase, with vitamin B6 as a tween German (24.5%) and Italian (43.8%) popula­ Plazar N and Jurdana M / Hyperhomocysteinemia and the role of B vitamins in cancer Acceptor Nutrition protein dUMP Serine B6 Cystathionine B6 Cysteine Transsulfuration a-Ketobutyrate FIGURE 2. Metaboliolism of homocysteine. dUMP - desoxyuridine monophosphate, dTMP - desoxytimidine monophosphste, THF - tetrahydrofolate, DHF - dihydrofolate, 5-MTHF - 5-methyltetrahydrofolate, 5,10-MTHF - 5,10-methyltetrahydrofolate, 5,10 MTHFR - 5,10- methyltetrahydrofolate reductase, MS - metionin synthase, MSR - metionin synthase reductase, B12 - vitamin B12, SAM - S-adenosylmethionine, SAH - S-adenosylhomocysteine, CBS - cystath­ ionine ß-synthase, GCT - .-cystathionase, B6 - vitamin B6. tions.20Moreover, it seems that this mutation and the reduced activity of the enzyme MTHFR are not connected with hyperhomocysteinemia if persons have balanced diet with optimal intake of vitamins (B6, B12 and folates).19 Hyperhomocysteinemia is frequently associated with folate deficiency and it has been long postu­lated that the plasma homocysteine concentration is inversely related to the occurrence of cardiovas­cular disease and venous thrombosis.21-24 Disturbed homocysteine metabolism, hyperhomo­cysteinemia and cancer Hyperhomocysteinemiaiscommonlyoccurring inawiderangeofunrelateddiseases.Forexam­ple,inpatientswithrenalfailureastrong,positive correlationwasobservedbetweenhomocysteine (tHcy)levels,serumcreatinine,andtherenal 82 Plazar N and Jurdana M / Hyperhomocysteinemia and the role of B vitamins in cancer glomerularfiltrationrate.Rheumatoidarthritis impairedgastricandotherdisturbancesresults ofteninelevatedbloodtHcy.25Thediseaseamong whichelevatedtHcyareobservedare:Systemic lupuserythematosus,non-insulin-dependentand insulin-dependentdiabetesmellitus,hypothy­roidism,cognitiveimpairmentandneuropsychi­atricdisorders(dementia,depression,schizophre­nia),fibromyalgiaandchronicfatiguesyndrome, Parkinson’sdisease,cerebrovasculardisorders, andasepticmeningitis.26 Increased tHcy levels are often found in patients with neoplastic diseases.27In vitroit was shown that some cancer cell lines are incapable of remeth­ylating tHcy and it was recently shown that ovar­ian cancer cells from patients with elevated tHcy have impaired capacity to remethylate tHcy.28,29Tempting to conclude that hyperhomocysteinemia in cancer patients could be secondary to the can­cer. However, impaired methylation of DNA and polyamines has often been proposed to be involved in carcinogenesis30, so the combination of increased tHcy levels and impaired methylation capacity in patients has been proposed as being carcinogenic.31In lymphocytes, positive correlation between cellu­lar tHcy levels and increased chromosome damage was shown.31 Patient with malignancies often have an in­creased risk of venous thromboembolic (VTE) dis­ease5and as such being the second most common cause of death in cancer patients, second to the pri­mary disease itself. The pathophysiology of this as­sociation has not been precisely defined. However, it has been reported that in cancer patients several pro-coagulant factors are increased.32Other estab­lished contributors to the VTE increased risk is oncological therapy as chemotherapy, hormonal adjuvant therapy, surgery, central venous cath­eters, immobility and inherited thrombophilia.5,33,34 However, this oncological therapy canalso influ­ence the immunological response of treatment patients.35 In women with advanced breast cancer hyperhomocysteinemia is common.36This ob­servation could explain the high rate of venous thrombosis in women with metastatic breast ma­lignancy.6Furthermore, the association between MTHFR C677T polymorfism and breast cancer has been reported. However a positive correlation has not been confirmed by all studies.37MTHFR C677T polymorphism is associated with changes in intra­cellular folate cofactors, affecting DNA methyla­tion and synthesis via altered one-carbon transfer reactions. Of further notice on this association are potential ethnic differences.38 Al-Awadi et al.39demonstrated with nude mice implanting human breast, prostate and pancreas tumour cells leads to decreased plasma cysteine, homocysteine and methionine levels over a two-month period, which was a direct result from the progressing implanted tumour cells. In the case of methionine, the decrease was significant only due to progression of the breast tumours over a long time period. The results suggest that the sulphur amino acids cysteine, homocysteine and methio­nine can be potentially used as plasma or serum biomarkers for cancer progression. Many other studies showed that the raised tHcy is related to the cancer itself and to the extent of the disease.21,40 After remissionof the cancer in chil­dren with acute lymphoblastic leukemia the tHcy levels returned to normal.2 Both plasma concentration of homocysteine and neopterin, a catabolic product of guanosine triphos­phate-GTP and as such an immune system activa­tion marker, are closely associated and elevated in patients with various types of disease.3From in vit­rostudies it has been shown that tumour cells and other proliferating cells release homocysteine.,41This in vitronotion might be extrapolated to the in vivosituation and could explain why hyperhomo­cysteinemia is observed in patients with various kinds of cancers. Within cellular immune activa­tion, T cells release large amounts of the cytokine IFN-., which stimulates human monocyte-derived macrophages and dendritic cells to produce neop­terin.42For example, in cancer patients, increased urine and plasma neopterin concentrations have been reported, suggesting enhanced cellular im­mune activation.43,44Therefore, immune activation cascades might also be an important triggers for the accumulation of plasma homocysteine in vari­ous diseases, including malignancies. When different tumours are compared, the fre­quency of increased neopterin concentration is much lower in patients with breast cancer that it is in patients with other types of cancers.3Although no association between neopterin and tumour size or lymph node status has been shown in women with breast cancer, follow-up examinations re­veal that at diagnosis high urine neopterin con­centrations are associated with shorter survival.4However, these authors conclude that plasma ho-mocysteine and neopterin concentrations are only rarely elevated in breast cancer patients and they note that the activation and proliferation of immu­nocompetent cells rather than tumour cells prolif­eration is responsible for hyperhomocysteinemia in these breast cancer patients. Plazar N and Jurdana M / Hyperhomocysteinemia and the role of B vitamins in cancer 83 B vitamins for cancer prevention Folate, vitamin B12, and vitamin B6, have a number of biologic roles that make them potentially impor­tant in cancer. Within DNA synthesis they func­tion as coenzymes in the synthesis of purines and thymidylate. Diminished levels of these vitamins may result in misincorporation of uracil into DNA, leading to chromosome breaks and disruption of DNA repair.45As explained earlier both folate and vitamin B12 are involved in DNA methylation. Deficient folate and vitamin B12 levels can reduce the availability of S-adenosylmethionine, the uni­versal methyl donor, for DNA methylation and may thereby influence gene expression. Inadequate body levels of biologically active folate, vitamin B6, and vitamin B12 are primary determinants of high blood homocysteine levels.46Folic acid is a component of food which has been associated with lower cancer risk in epidemiologic studies.9,10,47,48 Wide geographical variation and migrant studies in cancer incidence and mortal­ity suggest that diet and other lifestyle factors as physical activity influence cancer risk.49,50Data on cancer incidence and mortality are available from 37 countries and analysis showed that incidence of colorectal cancer was inversely correlated with more cereals (grains) in the diet.51Folic acid present in a wide variety of plant foods, such the legumes, vegetables, fruits and whole grains is thought to be protective against colorectal cancer.52,53The lack of folic acid in animal cells studies resulted in DNA defects that resemble effects found in cancer cells. It has been hypothesized in many epidemiologic studies38,54,55that cancer can be initiated by DNA damage (increasing DNA methylation, and by re­pairing and reducing formation of DNA strand breaks of p53 and Apc genes) caused by folic ac­id deficiency.56,57A deficiency of folic acid leads to a low level of thymidilic acid and alterations in the pool of nucleotides available for DNA and RNA synthesis. It is even suggested that adequate folate intake may be important in the prevention of breast cancer, particularly among women who consume alcohol.58-60 Alcohol is a known folate an­tagonist and thus could increase an individual’s re­quirement for folate intake. For vitamin B12, unlike folate, variation in amount absorbed rather than intake is the main determinant of plasma levels in Western populations.61In a prospective study anal­ysis of collected blood from 195 case–control pairs, low plasma levels of vitamin B12 were associated with increased risk of breast cancer among post-menopausal women; however, low plasma levels of folate, and homocysteine were not associated with breast cancer risk.62Hypofolatemia and meta­bolic alteration in homocysteine, vitamin B12 could be associated with laryngeal cancer.63Therefore, a great effort was made to proof this association as it was made to find association between cysteine cathepsins as well as stefins and promoting and invasion of head and neck tumours.63,64Increased plasma vitamin B12 concentration may reduce the risk of rectal cancer.65 ArecentanimalstudydemonstratedthataB12­deficientdiet,whichwasofinsufficientseverityto causeanemiaorillness,disturbednormalhomeosta­sisofone-carbonmetabolisminthecolonicmucosa andresultedindiminishedgenomicDNAmethyla­tionandincreaseduracilmisincorporationinDNA, bothofwhicharepurportedmechanismsforone-carbonmetabolism-relatedcoloniccarcinogenesis.66 In a large prospective study on health care pro­fessionals, high intake of folic acid was found to be significantly correlated with low incidence of colorectal adenomas (polyps).67 Therefore the diet regime and life style factors should be consider as primary prevention beside also important second­ary prevention.49,50,68Case control studies have as well as found high folic acid intake to be correlated with low risk for either pancreatic cancer or breast cancer.69,70In Greece and in Argentina studies cor­relating breast cancer and diet found risk reduc­tions from six to ten fold in subjects eating mostly vegetables rich of B vitamins.71 The high risk of de­veloping cancer in a lifetime in the North American and Western European societies might be related to the low intake of vegetables and particularly folic acid might be lacking in diets. Folic acid deficien­cy could be the permissive condition that enables DNA damage to occur and accumulate. This can lead to DNA damage and cancer. Is synthetic folate fortification always good for us? Many countries have implemented mandatory folic acid fortification of flour and grain products to re­duce the risk of various diseases, especially neural-tube birth effects. Experimental evidence suggests that high doses of folic acid may enhance growth of cancer cells.7,53,72,73 These effects have resulted in substantial increase in circulating folate and un-metabolized folic acid concentration.72,74Described 84 Plazar N and Jurdana M / Hyperhomocysteinemia and the role of B vitamins in cancer the beneficial effects of folate in preventing cancer, it is also well known that high intake of synthetic folic acid might mask vitamin B12 deficiency.74Experimental studies suggest that excessing folic acid may promote the progression of already ex­isting preneoplasms.7Responsible mechanisms of high folates concentration causing cancer promot­ing effects include folates providing nucleotide precursors for the preneoplastic cells improving their replication and proliferation. Folates, methyl donors might lead to a de novo methylation and subsequent inactivation of tumour-suppressor genes, resulting in accelerated tumour progres­sion.7The safe upper limit for folate intake as well as the safe upper folate concentration in blood are not known.6The mandatory fortification of food with folic acid, it dose and the time of intervention depends by the country’s decision.72 Because the safety of folate might depend on its chemical structure (natural folate or synthetic folic acid), there is the question of potential adverse ef­fects of circulating unmetabolized folic acid.75 Conclusions Manydatasupportarelationshipbetweenhyper-homocysteinemia,lowBvitaminsconcentration andriskforvarioustypesofcancer.Defectiveme­tabolismoftHcyincarcinogenesisiswelldocu­mented,butthepathophysiologyofthisassociation isnotfullyunderstood.Manyauthorssuggestthat factorscontributingtofolatestatusarenotprotec­tiveagainstcertaintypeofcancer,sofurtherstudies areneededtoexplorefolatestudiesinhuman. Acknowledgement The authors thank Dr. Cčcil J.W.Meulenberg and MSc. Majda Šavle from College of Health Care, University of Primorska, Slovenia, for manuscript reading and their useful comments. References 1. FassbenderK,OrellM,HennericiM.PlasmaHomocyst(e)ineConcentrations in Cerebrovascular Disease. Stroke 1999; 30: 2224-38. 2. Sun CF, Haven TR, Wu TL, Tsao KC, Wu JT. Serum total homocysteine in­creases with the rapid proliferation rate of tumor cells and decline upon cell death: a potential new tumor marker. Clin Chim Acta 2002; 321: 55-62. 3. Trosseau A. Phlegmasia alba dolens. Clin de l’Hotel-Dieu de Paris 1865; 3: 654-712. 4. DebevecL, JericT, KovacV, SokM, Bitenc M. The progress in routine man­agement of lung cancer patients. A comparative analysis of an institution in 1996 and 2006. Radiol Oncol 2009; 43: 47-53. 5. Rickles FR, Levine M, Edwards RL. Hemostatic alteration in cancer patients. Cancer Metastasis Rev 1992; 11: 237-48. 6. Smith AD, Kim YI, Refsum H. 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Lancet 2009; 19: 959-61. 73. Ulrich CM, Potter JD. Folate and cancer: timing is everything. JAMA 2007; 297: 2408-9. 74. Shane B. Folate fortification: enough already? Am J Clin Nutr 2003; 77: 8-9. 75. Kalmbach RD, Choumenkovitch SF, Troen AM, D’Agostino R, Jacques PF, Selhub J. Circulating folic acid in plasma: relation to folic acid fortification. Am J Clin Nutr 2008; 88: 763-8. research article Hemostatic efficacy of chitosan-based bandage for closure of percutaneous arterial access sites: An experimental study in heparinized sheep model Pawanrat Kranokpiraksa, Dusan Pavcnik, Hideaki Kakizawa, Barry T. Uchida, Miran Jeromel, Frederick S. Keller, Josef Rösch Dotter Interventional Institute, Oregon Health Sciences University, Portland, OR, USA Received 4 March 2010 Accepted 22 March 2010 Correspondence to: Dusan Pavcnik, MD, PhD, Dotter Interventional Institute, Oregon Health & Science University, 3181 SW Sam Jackson Park Road, L342, Portland, Oregon 97239, USA. E-mail: pavcnikd@ohsu.edu Disclosure: No potential conflicts of interest were disclosed. Background. Most of the presently used percutaneous arterial closure devices (PACD) for hemostasis after inter-ventional vascular procedures are effective, but carry risk of complications by deposition of a foreign body. A new promising externally applied PACD – chitosan-based HemCon Bandage (HCB) was explored in sheep. The HCB hemostatic efficacy and complications occurring with its use were compared to those with the standard manual compression (SMC). Material and methods. Both superficial femoral arteries (SFA) of 9 heparinized sheep were catheterized with an 8F sheath for 5 minutes. After the sheath withdrawal, hemostasis with the HCB was compared with hemostasis achieved with SMC in the contralateral SFA. Iliac angiograms performed by carotid artery approach determined the hemostasis time. Results. The HCB use shortened time to hemostasis with a mean time of 6.9 ± 3.9 minutes versus 10.8 ± 2.8 minutes for the SMC (P-value 0.019). Seven SFAs in the HCB group and only 1 SFA in the SMC group exhibited hemostasis in 5 minutes. All nine SFAs using the HCB showed femoral artery patency and demonstrated less hematoma (2/9) than in the SMC group (8/9). No complications developed in the HCB group, one SFA occlusion was seen in the SMC group. Conclusions. The externally applied HCB in heparinized sheep was safe and effective. It significantly shortened time to hemostasis at the SFA access sites following 8F sheath removal. Proper application of the HCB was necessary to shorten hemostasis and prevent hematoma formation. The HCB should be tested in a clinically controlled study to evaluate its efficacy in humans. Key words: arterial catheterization; hemostasis; closure devices; hemostatic pads; chitosan-based pad Introduction Since the introduction of percutaneous catheteri­zation in 19531, manual compression over the puncture site has been the standard technique for achieving hemostasis in interventional radiology.2With diagnostic angiography using 5F to 6F cath­eters, manual compression followed by bed rest has been very efficient for achieving hemostasis and has led to less than 1% puncture site complica­tions.3Introduction of therapeutic vascular proce­dures with the need for 8F and larger introductory sheaths, adjuvant anticoagulation and antiplatelets or thrombolytic therapy has led to an increase in complications. Arterial access site complications as high as 17% have been reported with intervention-al procedures, some of them requiring corrective surgical treatment.4,5 Since the early nineties, several types of percu­taneous arterial closure devices (PACD) have been introduced to enhance hemostasis after interven­tional procedures and to decrease the rate of com­plications. These devices either replace or shorten the time of manual compression at the puncture Kranokpiraksa P et al./ Hemostatic efficacy of chitosan bandage for arterial closure 87 site.5,6Externally applied hemostatic patches and pads that accelerate the hemocoagulation process at the puncture site are one of the newest PACD types. Acceleration of hemostasis is caused by active ingredients of the patches and pads that contain procoagulants that potentiate clot forma­tion.7-9All procoagulants require compression for hemostatis, but substantially reduce compres­sion times. Most procoagulants require contact with blood for activation. There have been several clinical studies on the hemostatic efficacy of these patches and pads.7-15However, we have found only two experimental reports documenting the hemostatic efficacy of procoagulants in animals. One paper reported exploration of the efficacy of microfibrillar collagen and thrombin applied into the arterial puncture tract in dogs with the help of a balloon catheter.16The other report described a procoagulant (chitosan) installed into the arte­rial puncture tract in dogs.17Chitosan is a linear polysaccharide derived from chitin commercially extracted from marine arthropod shells. It is com­posed of positively charged molecules that attract red blood cells and platelets, thereby, promoting hemostasis. We report an exploration of the hemo-static efficacy of chitosan-based HemCon Bandage (HCB) (HemCon Medical Technologies, Portland, OR) and a comparison of these bandages with standard manual compressionin a heparinized sheep model. We used sheep for testing since their arteries are similar in size to humans.18,19In addi­tion, their coagulation and fibrinolytic systems are closer to those of humans when compared to ca­nine and swine.20 Material and methods The study protocol was approved by the Institutional Animal Care and Use Committee. Nine female sheep weighing from 56 to 70 kg were used in this study. A cardiac mobile system (GE/OEC 9800; GE Medical Systems, OEC, Salt Lake City, UT, USA) with digital imaging was used for fluoroscopy and angiography. Digital subtraction angiographies were performed with an injector (Medrad mark Plus, MEDRAD, Inc., Warrendale, PA, USA). Preparation of animals and their anesthesia were described in previous paper.18After induc­tion of general endotracheal anesthesia, the sheep were placed and secured with their backs on the ra­diographic table and their hind limbs in moderate abduction. The neck and both groins were shaved and prepped. The right common carotid artery (CCA) was exposed and a 9F, 50 cm long introduc­er sheath (Cook Medical, Bloomington, IN, USA) was retrogradely inserted into abdominal aorta. A standard dose of heparin (100 IU/kg) was then ad­ministered intra-arterially. Activated clotting times (ACT) were recorded at baseline prior to heparin administration, prior to arterial sheath removal and at the end of the procedures. A 5F multiside-hole catheter (Cook Medical) was then introduced through the 9F sheath for selective angiography of the external iliac arteries using an injection of 16 ml of Omnipaque (IOHEXOL 300 mg 1/mil, GE Healthcare, Princeton, NJ, USA) in 2 seconds. The access sites in each animal were the superfi­cial femoral arteries (SFA). One SFA received treat­ment with the HCB applied with manual compres­sion and the contralateral SFA served as a control with the use of SMC. The sequences of the punc­ture sites and treatment modes were randomized. With selective iliac angiography, a road map im­age was created and SFA diameter was measured. Single wall access of the SFA was done under road map guidance with the 21 gauge needle of the micropuncture set (Cook Medical). An 8F sheath was then placed into the artery and left there for 5 minutes. During the sheath removal and prior to the use of the HCB 2” X 2” in size, mild nonoc­clusive pressure was first applied above the skin puncture site. After the sheath was completely re­moved, a small amount of blood was first allowed to seep on the skin access site to contact the band­age and initiate hemostasis. The bandage was then applied with digital nonocclusive pressure. In the control SFA, significant digital pressure was ap­plied during the sheath removal to prevent blood penetration through the puncture tract, as used in clinical practice. Manual pressure was held in both the treatment and control groups for 5 minutes. Angiography was done immediately after pres­sure relief to confirm hemostasis. If angiography showed extravasation, compression was continued for a further 2.5 minutes. Angiography was then repeated every 2-1/2 minutes until no evidence of bleeding was seen. Compression was reapplied in the interval between repeat angiographies. Lack of extravasation was the endpoint. The study then proceeded on the contralateral SFA. Finally, angi­ography of each side was performed at about 30 minutes after cessation of bleeding to check the patency and status of the SFA. The access sites were then checked for hematomas defined as loss of definition of the fossa subinguinalis and raised appearance of the skin. The degree of hematoma at 88 Kranokpiraksa P et al./ Hemostatic efficacy of chitosan bandage for arterial closure TABLE 1. Comparison of angiographic findings, ACT values, hemostasis times and post procedure hematomas in 18 punctured superficial femoral arteries, 9 in the HCB and 9 in the control group SFA diameter (mm) 5.4 - 6.1 5.4 – 6.1 Baseline ACT 106 – 161 (136 +/- 19.8) 205-1061 267- 480 0.737 ACT prior sheath removal (sec) (404.4 +/- 262) (371.8+/-73.1) Hemostasis time (min) 5-15 (6.9 +/- 3.9) 5-12.5 (10.8 +/- 2.8) 0.019 Artery patency at 30 min 9 8 Subcutaneous Hematoma 2 8 (2 significant) TABLE 2. ACT values prior to sheath removal, hemostasis times and hematoma presence at puncture site in individual animals 1 348 5 0 344 12.5+ 2 2685 0 4127.5 + 3 447 5 0 257 12.5+ 4 800 12.5 + 1061 12.5 ++ 5 361 15 + 244 12.5 ++ 6 4805 0 20510 + 7 262 5 0 260 12.5+ 8 2725 0 4695 0 9 433 5 0 388 12.5+ Mean+/-372±73.1 6.94±3.9 404±262 10.8±2.8 ACT values in seconds, hemostasis time in minutes; Hematoma grades 0 = none, + = minor, ++ = significant the groin was graded: 0 = no hematoma, 1 = slight and 2 = significant. Groin area fullness with promi­nence <1 cm was considered slight hematoma. At the end of the study, the animals were euthanized. Statistical analysis Data were recorded into a worksheet (Excel 2007, Microsoft, Redmonds, WA, USA) and summary statistics (mean and standard deviation) were cal­culated. Student’s t-test was used to determine if there was a statistically significant difference be­tween the control and treatment with regard to achieving hemostasis. A value of P<0.05 was con­sidered significant. Results SFA diameters ranged from 5.4 to 6.1 mm with all pairs being matched in size. The mean ACT prior to the sheath removal in the HCB group was 404.4 ±262 seconds and in the control group 371.87 ± 73.1 (p=0.737). The mean time to achieve hemosta-sis in the HCB group was 6.9 ± 3.9 minutes, while the mean time of the control group was 10.8 ± 2.8 minutes (p=0.019). The results are summarized in Table 1 and 2. In the HCB group, hemostasis at 5 minutes post sheath removal was achieved in 7 of 9 SFAs (77.8%) (Figure 1). The other 2 SFAs exhibited hemostasis at 12.5 and 15 minutes, respectively. One of those two delayed times to hemostasis was equivalent to the control side and the other needed Kranokpiraksa P et al./ Hemostatic efficacy of chitosan bandage for arterial closure 89 FIguRE 1. Hemostatic control with the chitosan-based bandage. A - Baseline angiogram prior to the superficial femoral artery (SFA) puncture. B - Angiogram obtained after 8F sheath withdrawal and 5 minutes chitosan-based bandage compression shows complete hemostasis. C - Angiogram obtained 30 minutes after sheath withdrawal shows patent SFA. longer time to achieve hemostasis than the control SFA. Hemostasis was obtained at 5 minutes after sheath removal in one of 9 (11.1%) of the control group (Figure 1). Angiography at about 30 minutes after intervention showed no extravasation in ei­ther HCB or control group. All SFAs in the HCB group were patent without demonstrable arterial spasm. Eight of 9 (88.9%) SFAs in the control group were patent and spasm was found in 3 (33%) arter­ies. Two of these SFAs exhibited small defects, pre­sumably thrombi at the access sites. One SFA was occluded. In the HCB group, there were two grade 1 hematomas, while in the SMC group there were eight, two of which were grade 2. Discussion Numerous PACDs are now available for achieving rapid hemostasis at percutaneous arterial access sites after diagnostic or interventional procedures. Madigan et al. in 2007 reviewed 14 PACDs.6Other new PACDs are being developed and/or tested.7Based on their principle mechanism of hemostasis, PACDs are categorized into four groups. The first three groups include biodegradable sealing plugs, suture-mediated devices and staple-mediated de­vices. They are very effective and have been the most frequently used PACDs. However, they have not been used without complications. Their use has been associated with infections due to deposition of a foreign body, bleeding, pseudo aneurysm, ar­teriovenous fistula and arterial occlusion.5,6,18,21The fourth group of PACDs – patches and pads have recently received close attention. They are topically applied and their procoagulant ingredients accel­erate hemocoagulation at the access site without leaving any foreign material behind. Because of their action, they are called “noninvasive” PACDs.7The procoagulant components of the noninvasive PACDs include, among others, bovine thrombin (D-Stat-Dry-Vascular Solutions), poly-N-acetyl glucosamine derived from marine diatoms (Syvek Patch-Marine Polymer Technologies), polyprolate acetate (Clo-Sur Pad – Scion Cardiovascular) and chitosan obtained from exoskeleton of crustaceans (Chito-Seal, Abbot Vascular, HemCon®Bandage). Clinical studies of D-Stat-Dry, Syvek Patch, Clor-Sur Pad and Chito-Seal showed that these PACDs applied with compression reduce time to hemos­tasis after femoral artery catheterization compared with SMC and do not increase the complication rate when using 4-6 F sheaths.8,10,12-14 Literatureisavailableonthehemostaticefficacy oftheHCBintraumaticanimalmodels,andon HCBuseintraumapatientsinthemilitary,inemer­gencydepartmentsandduringsurgery.22However, itsefficacyforhemostasisafterfemoralarterycath­eterizationhasnotbeendocumentedbyeitherex­perimentalorclinicalstudies.Ourexperimental studydemonstratesthattheHCBcanbeeffectivein thissetting.Inourexperimentalheparinizedsheep 90 Kranokpiraksa P et al./ Hemostatic efficacy of chitosan bandage for arterial closure FIguRE 2. Hemostatic control with the standard manual compression. A - Baseline arteriogram prior SFA puncture. B - Angiogram obtained after 8F sheath withdrawal and 5 minutes standard manual compression shows extensive extravasation from the puncture site. C - After additional 2.5 minutes manual compression (total 7.5 minutes), there is decreased extravasation. D - After additional 2.5 minutes of manual compression (10 minutes total), complete hemostasis is achieved. modelusingan8Fsheath,theHCBshortenedthe timetohemostasistoalmosthalfofSMCalone. HemostasiswasachievedwithHCBin7of9punc­turesiteswithin5minutes,incomparisontoonly onesiteinthecontrolgroupthatwaswithin5min­utes.Thetwoinstanceswherehemostasiswiththe HCBtookthesameamountoftimeorslightlylong­ertimethaninthecontrolSFAwasduetoapplica­tionofthebandageinamannersimilartothatof standardcompressionwheretheneededbloodto activatetheHCBwasnotallowedtoseepthrough theaccesstract.Thisfindingstronglyreinforces theneedforthepresenceofbloodattheaccesssite toinitiatethehemocoagulativeactionofchitosan. TheHCBgroupalsodemonstratedlesshematoma formationthanthecontrolgroup.Thehigherinci­denceofhematomas,however,wasundoubtedly alsorelatedtoreleaseofpressureattheaccesssite forperformanceofangiographybeforehemostasis couldbeestablished.Nobleedingwasfoundin bothgroups30minutesaftercompressions,butin twoSFAssmalldefectssuspiciousofthrombiwere foundattheaccesssitesafterSMC. The study limitations include the small sample size and the impossibility of performing a blind study due to the distinctive appearance of the HCB. Another limitation is the absence of data on time to ambulation and long-term efficacy of the closure. Important information about time to am­bulation, thus, could not be evaluated. Another study should address these limitations and should also include histopathologic evaluation of the SFAs access site. Conclusions The chitosan-based HCB was effective and short­ened the time to hemostasis at SFA access sites following removal of an 8F sheath in heparinized sheep. Proper application of the bandage, however, was necessary to shorten hemostasis and decrease hematoma formation. A controlled clinical study needs to be done for evaluation of hemostatic effi­cacy of HCB following endovascular interventions in humans. Acknowledgement Supported by a research grant from HemCon Medical Technologies, Portland, OR, USA Kranokpiraksa P et al./ Hemostatic efficacy of chitosan bandage for arterial closure 91 References 1. Seldinger SI. Catheter replacement of the needle in percutaneous arteriog­raphy; a new technique. Acta Radiol 1953; 39: 368-76. 2. Gjikolli B, Hadzihasanovic B, Jaganjac S, Herceglija E, Niksic M, HadzimehmedagicA,etal.Treatmentofcomplicatedcasewithsubclaviasteal syndrome and stenosis of common iliac artery. Radiol Oncol 2008; 42: 1-12. 3. Singh H, Cardella JF, Cole PE, Grassi CJ, McCowan TC, Swan TL, et al. Quality improvement guidelines for diagnostic arteriography. J Vasc Interv Radiol 2002; 13: 1-6. 4. Popma JJ, Satler LF, Pichard AD, Kent KM, Campbell A, Chuang YC, et al. Vascular complications after balloon and new devices angioplasty. Circulation 1993; 88 (part 1): 1569-78. 5. Hoffer EK, Bloch RD. Percutaneous arterial closure devices. J Vasc Interv Radiol 2003; 14: 865-85. 6. Madigan JB, Ratnam LA, Belli AM. Arterial closure devices. J Cardiovasc Surg 2007; 48: 607-24. 7. Hirsch JA, Reddy SA, Capasso WE, Linfante I. Non-invasive haemostatic closure devices. Tech Vasc Interv Radiol 2003; 6: 92-5. 8. Nguyen N, Hasan S, Caufield L, Ling FS, Marins CR. Radnomized control trial of topical hemostasis pad use for achieving vascular hemostasis following percutaneous coronary intervention. Catheter Cardiovasc Interv 2007; 69: 801-7. 9. Wang DS, Chu LF, Olson SE, Miller FJ, Valji K, Wong WH, et al. Comparative evaluation of noninvasive compression adjuncts for hemostasis in percuta­neous arterial, venous and arteriovenous dialysis access procedures. J Vasc Interv Radiol 2008; 19: 72-9. 10. Mlekusch W, Dick P, Haumer M, Sabeti S, Minar E, Schillinger M. Arterial puncture site management after percutaneous transluminal procedures using a hemostatic wound dressing (Clo-Sur PAD) versus conventional manual compression: A randomized controlled trial. J Endovasc Ther 2006; 13: 23-31. 11. Hallak OK, Cubedder RJ; Griffith RA, Reyes BJ. The use of the D-STAT dry bandage for the control of vascular access site bleeding: A multicenter experience in 376 patients. Cardiovasc Intervent Radiol 2007; 30: 593-600. 12. Balzer JO, Schwarz W, Thalhammer A, Eichler K, Schmitz-Rixen T, Vogl TJ. Post interventional percutaneous closure of femoral artery access sites using the Clo-Sur PAD device: Initial findings. Eur Radiol 2007; 17: 693-700. 13. Rastan A, Sixt S, Swarzwalder U, Schwarz T, Frank U, Burgelin K, et al. VIPER-2: A prospective, randomized single-center comparison of 2 different closure devices with a hemostatic wound dressing for closure of femoral artery access sites. J Endovasc Ther 2008; 15: 83-90. 14. Applegate RJ, Sacrinty MT, Kutcher MA, Baki TT, Gandhi SK, Santos RM, et al. Propensity score analysis of vascular complications after diagnostic cardiac catheterization and percutaneous coronary intervention using thrombin hemostatic patch-facilitated manual compression. J Invasive Cardiol 2007; 19: 164-70. 15. Mlekusch W, Minar E, Dick P, Sabeti S, Bartok A, Haumer M, et al. Access site management after peripheral percutaneous transluminal procedures: Neptun Pad compared with conventional manual compression. Radiology 2008; 249: 1058-63. 16. Gershony G, Brock JM, Powell JS. Novel vascular sealing device for closure of percutaneous vascular access sites. Cathet Cardiovasc Diag 1998; 45: 82-8. 17. Hoekstra A, Struszczyk H, Kivekas O. Percutaneous microcrystalline chitosan application for sealing arterial puncture sites. Biomaterials 1998 19: 1467­71. 18. Ni RF, Kranokpiraksa P, Pavcnik D, Kakizawa H, Uchida BT, Keller FS, et al. Testing percutaneous arterial closure devices: An animal model. Cardiovasc Intervent Radiol 2009; 32: 313-6. 19. Wei Lu, Pavcnik D, Uchida B, Park WK, Liu L, Timmermans HA et al. The Ovine jugular vein as a model for interventional radiology procedures. Radiol Oncol 2008; 42: 59-65. 20. Narayanaswamy M, Wright KC, Kandarpa K. Animal models for athero­sclerosis, restenosis and endovascular graft research. J Vasc Interv Radiol 2000; 11: 5-17. 21. Wagner SC, Gonsalves CF, Eschelman DJ, Sullivan KL, Bonn J. Complications of a percutaneous suture-medicated closure device versus manual com­pression for arteriotomy closure: A case-controlled study. J Vasc Interv Radiol 2003; 14: 735-41. 22. Gustafson SB, Fulkerson P, Bildfell R, Aguilera L, Hazard TM. Chitosan dress­ing provides hemostasis in swine femoral arterial injury model. Prehosp Emerg Care 2007; 11: 172-8. research article 3T MRI in evaluation of asbestos-related thoracic diseases – preliminary results Janez Podobnik1, Igor Kocijancic1, Viljem Kovac2, Igor Sersa3 1 Institute of Radiology, University Clinical Centre Ljubljana, Ljubljana, Slovenia 2 Department of Radiation Oncology, Institute of Oncology Ljubljana, Ljubljana, Slovenia 3 Jožef Stefan Institute, Ljubljana, Slovenia Received 4 January 2010 Accepted 11 February 2010 Correspondence to: Janez Podobnik, Institute of Radiology, University Clinical Centre Ljubljana, Zaloška 2, Ljubljana, Slovenia. Phone: + 386 1 522 30 62; Fax: + 386 1 522 24 97; E-mail: janez.podobnik@kclj.si Disclosure: No potential conflicts of interest were disclosed. Background. 3T high-field magnetic resonance imaging (MRI) scanners have recently become available for the clinical use and are being increasingly applied in the field of whole-body imaging and chest imaging as well. The aim of this study was to evaluate the diagnostic potential of 3 T MRI as a complementary imaging modality to CT in detecting the pathological changes of asbestos-related thoracic diseases. Patients and methods. Fifteen patients with the asbestos-related thoracic disease were scheduled for 3T MRI. Five had a benign form of the disease and 10 had malignant pleural mesothelioma (MPM). From the patients with a be­nign form of the disease their last CT examination in digital form was acquired and patients with MPM were scheduled for CT examination with contrast media. The protocol of MR imaging consists of T2-weighted cardiac-gated breath-hold turbo spin echo (TSE) sequences in coronal, sagittal and axial plane and T1-weighted cardiac-gated breath-hold TSE black blood in axial plane. In T2-weighted sequences in axial plane, fat saturation was also used. CT examinations were obtained with the administration of the contrast medium from lung apices to the lower end of the liver. Images of 5 mm (mediastinum window) and 3 mm (lung window) in axial plan were reconstructed. MRI signal intensity of le­sions and adjacent muscles on Syngo MultiModality Work Place were measured. Results. Compared to muscles pleural plaques appeared hypo-intense to iso-intense on T1 weighted images (in 100%) and also hypo-intense on T2 fs-weighted images (in 100%). MPM appeared inhomogeneous hypo-intense to iso-intense on T1-weighted and hyperintense on T2 fs-weighted images in all patients (100%). Conclusions. These preliminary results pointed out that MRI was equal or even better compared with CT examination for detecting possible malignant potential of pleural changes in the asbestos-related pleural disease, using signal inten­sity measurements of T2 fs-weighted images. The 3T MRI enabled the accurate determination of chest pathology and it could be used for imaging of patients with the asbestos-related thoracic disease. MRI is particularly valuable because a patient is not exposed to the harmful radiation which is important if imaging methods are used repeatedly, like in screening programs or in monitoring of treatment results. This finding turned us to propose 3T MRI imaging technique as a non-ionizing imaging method for the follow-up of patients with the isolated pleural form of the asbestos-related disease. Key words: 3T, magnetic resonance imaging; asbestos-related thoracic disease; malignant pleural mesothelioma Introduction Asbestos is a generic term applied to a variety of naturally formed hydrated silicates, which were used because of their heat resistance properties. Asbestos-related thoracic diseases are benign pleural effusions, pleural plaques, diffuse pleural thickening, rounded atelectasis, asbestosis, mes-othelioma, and lung cancer. Asbestos is a mainly cause for malignant mesothelioma, much more im­portant than the potential other causes as Simian virus.1The incidence of malignant mesothelioma is expected to peak between 2010 and 2030 in indus­trialized countries despite the regulatory restric­tion during the 1980s and 1990s.2 CT is a gold standard tool for the detection of the asbestos-related thoracic disease although re­cent studies revealed that MRI was superior in Podobnik J et al. / MRI and asbestos-related thoracic diseases 93 the detection of the invasive growth of malignant pleural mesothelioma (MPM) in diaphragm and endothoracic fascia or the detection of single chest wall focus.3A high radiation dose in repeated CT examination and the use of iodine contrast media in patients with renal disease, diabetes and known allergy must be considered. 3T high-field MRI scanners have recently be­come available for the clinical use and are now in­creasingly being applied in the field of the whole body imaging and the thoracic imaging as well. Due to the fact that the signal-to-noise ratio is di­rectly related to the static magnetic field strength the spatial resolution can be increased and the ex­amination time can be shortened.4 MRI of the lungs is limited because of physical and physiological factors such as low proton den­sity, susceptibility effect, and respiratory move­ments as well as cardiac and vascular pulsation. Susceptibility artefacts, magnetic field distortion and motion artefacts are increased in high-field MRI. However, the signal loss from normal lung parenchyma due to the susceptibility effect and the theoretically increased signal from solid changes may result in a higher contrast between the nor­mal lungs and pathological changes in high-field MRI.5,6 To reduce susceptibility artefacts the turbo spin-echo (TSE) with a short echo spacing sequence has been recommended.7To avoid respiratory movements the breath-hold technique in inspira­tion with the examination time under 20 seconds should be used6and to prevent cardiac and vascu­lar pulsation artefacts ECG triggering must be ap­plied. To avoid additional movements phase array coil should be properly attached with belt.8 The aim of our study was to evaluate the diag­nostic performance of 3T MRI for detection and characterization of asbestos-related thoracic dis­eases in comparison to CT. Patients and methods Fifteen patients with the asbestos-related thoracic disease (ARTD) were scheduled for 3T MRI. Five had a benign form of the disease and 10 had MPM. From the patients with a benign form of ARTD their last CT examination in digital form, which was not older than one month, was acquired and patients with MPM were scheduled for CT exami­nation with contrast media. All patients with the benign form of ARTD were males and occupationally exposed to asbestos. The median age was 66 years in range from 53 to 76 years. In patients with MPM were 6 males and 4 females. The median age of this group was 62 years in range from 50 to 75 years. Only 3 males were oc­cupationally exposed to asbestos, in others the his­tory of the environmental exposure was described. The study was approved by the national medi­cal ethic committee of the Republic of Slovenia. A written consent was obtained from all patients. MRI studies MR studies were performed with the Trio team system (Siemens, Erlangen, Germany) equipped with the gradient system with a maximum gradi­ent amplitude 40 mT/m and slew rate of 200 mT/m/ms. A matrix body coil with 6 elements in combina­tion with a spine coil was used. The protocol of MR imaging consists of T2­weighted cardiac-gated breath-hold TSE se­quences in coronal, sagittal and axial plane and T1-weighted cardiac-gated breath-hold TSE black blood in axial plane. In T2-weighted sequences in axial plane Spectral pulse for saturation the signal from fat (SPIR - fs) was also used. T2-weighted TSE with following parameters were performed: Long TR (repetition time) – depends on heart rate (two cardiac cycles were used) • TE (echo time) 100 ms • Slice thickness 5 mm with 1 mm gap • Turbo factor 29 (Echo Train Length 5) • Field of view between 350 to 400 mm (depends on the patient size) • Matrixsize208x320withinterpolation • Parallel imaging factor (iPAT) 2 • Acquisition time <20 seconds. T1-weighted TSE with following parameters were performed: • Short TR – depends on heart rate (one cardiac cycles were used) • TE (echo time) 28 ms • Slicethickness5mmwith2.5mmgap • Turbo factor 9 (Echo Train Length 11) • Field of view between 350 to 400 mm (depends on the patient size) • Matrixsize106x256withinterpolation • Parallel imaging factor (iPAT) 2 • Acquisition time <14 seconds. The examination time takes from 25 to 30 min­utes depending on the patient’s cooperation. Podobnik J et al. / MRI and asbestos-related thoracic diseases CT examinations CT examinations were obtained with Somatom 16 or Definition scanners (Siemens, Erlangen, Germany) with the administration of the contrast medium using a power injector with 2 ml/s flow. CT scans with 120 kV and 100 mAs were performed from lung apices to the end of the liver. Kernel B31f medium smooth for mediastinal window (W: 350, C: 35) and B80f ultra sharp for the lung window (W: 1600, C: -600) were used. Images of 5 mm (me-diastinal window) and 3 mm (lung window) in axial plan were reconstructed. Measurement of MRI signal intensity MRI signal intensity of lesions on Syngo MultiModality Work Place were measured to es­tablish their hyper intensity or hypo intensity com­pared to the signal intensity of muscles. On T1and T2fs-weighted images in axial plane the circular region of interests (ROI) with area ex­panse from 0.3 to 0.4 cm2 were drawn. The region with the pronounced artefact was avoided. Data were collected and arranged regarding to the char­acteristics of the lesion. MRI and CT examinations were assessed by two radiologists experienced in chest imaging. Results Compared to muscles benign pleural plaques ap­peared hypo-intense on T2fs-weighted images and on T1weighted images. In some plaques a hyper-intense rim between the hypo-intense plaque centre and lung parenchyma on T2fs- weighted images was found (Figure 1). A diffuse pleural thickening was more distinctive on MRI than on CT images. MPM appeared inhomogeneous hypo-intense to iso-intense on T1-weighted and hyperintense on T2fs-weighted images. SPIR was used to saturate the signal from the fat and consequently a high signal compared to the muscle was only found in malig­nant lesions. In our study 3T MRI shows the extent of the tumour and accompanying pleural solid and fluid components with the greater accuracy com­pared to CT examination (Figure 2). Results of signal intensity measurements on T1-weighted images (Figure 3) have shown that the MR signal from benign pleural plaques com­pared to muscles was hypo-intense to iso-intense in all patients (100%). In eight patients with MPM Radiol Oncol 2010; 44(2): 92-96. Podobnik J et al. / MRI and asbestos-related thoracic diseases 95 (80%) the measured MR signal was hypo-intense, iso-intense in one patient (10%) and hyper-intense in one patient (10%). Results of measurements on T2fs-weighted im­ages (Figure 4) have shown that the MR signal from benign plaques compared to muscles was hypo-intense in all patients (100%) and hyper-intense in patients with MPM (100%). Discussion The benefit of 3T high field MR scanners which have recently become available for the clinical use is higher signal-to-noise ratio. For this reason the examination time can be shortened and the spatial resolution can be improved.4These facts are the reason that MRI is expanding on the field of chest imaging and becoming comparable imaging mo­dality to CT and proposed as a valuable alternative to certain patient groups.9 The most significant advantage of MRI of MPM is its excellent contrast resolution of soft tissues. Recent studies have demonstrated that MRI is su­perior to CT in evaluating of MPM invasive growth in diaphragm and abdominal cavity, invasion of endothoracic fascia and mediastinal structures.3,10Falaschi et al.11analyzed the potential usefulness of MR signal intensity in differentiating the malig­nant from the benign pleural disease and conclud­ed that the hypo-intense signal in pulse sequences with long TR is a reliable predictive sign of the be­nign disease. In our study the intensity of the sig­nal measured in benign pleural plaques compared to muscles on T1-weighted images were hypo-in­tense to iso-intense. On T2fs-weighted images the signal intensity from MPM compared to muscles was hyper-intense in all patients. CT using high resolution protocols is superior to MRI in imaging of parenchymal and early inter-and nature (calcified and non-calcified) of pleural stitial involvement in ARTD, but MRI achieved a plaques, but statistically significant conclusions on comparable interobserver agreement in detecting this statement needs to be confirmed on the larger pleural plaques compared to CT and a higher in-number of patients. terobserver agreement in revealing other pleural The incidence of ARTD is rising but the pattern pathologies.2,7In the same article it is also stated of ARTD has been changing due to the intensity of that a hyper-intense rim of benign asbestos plaques exposures. The incidence of MPM will probably next to lung parenchyma on T2weighted images increase at least twenty years after the interdic-possibly correlate to reduced plaque collagen fi-tion of asbestos.13This fact requires the evaluation brils and that diffuse pleural thickenings are more of screening programs for the detection of early pronounced on MRI than on CT. In our small stage malignant changes in a high risk group. MRI group of patients with asbestosis (altogether 5 pa-is a very suitable imaging method in repeatedly tients) we found that imaging of the asbestos pleu-screening programs because the patient is not ex-ral disease with 3T MR protocols was comparable posed to a harmful radiation. MRI can be also used with CT imaging showing almost the same extent in monitoring patients treated with chemotherapy. 96 Podobnik J et al. / MRI and asbestos-related thoracic diseases 7. Yi CA, Jeon TY, Lee KS, Lee JH, Seo JB, Kim YK, et al. 3-T MRI: usefulness for A3 A4 A6 A7 A9 A10 A15 A16 A19 A17 A18 A20 A12 A13 A14 PI JI KJ ŠJ VH ŽF SE PM JF MJ BR H CŽ BV KJ evaluating primary lung cancer and small nodules in lobes not containing primary tumors. AJR Am J Roentgenol 2007; 189: 386-92. FIGURE 3. Signal intensity measurement on T1 fs-weighted images. 8. Sofic A, Šehovic N, Bešlic Š, Prnjavorac B, Bilalovic N, Caluk J, et al. MR MPM = malignant pleural mesothelioma rectum imaging with ultra sound gel as instrumental contrast media in tubulovillous adenoma. Radiol Oncol 2008; 42: 136-42. 9. Fink C, Puderbach M, Biederer J, Fabel M, Dietrich O, Kauczor HU, et al. Lung MRI at 1.5 and 3 Tesla: observer preference study and lesion contrast using five different pulse sequences. Invest Radiol 2007; 42: 377-83. SIGNAL INTENSITY IN LESION 350 10. Bekkelund SI, Aasebo U, Pierre-Jerome C, Holmboe J. Magnetic resonance imaging of the thorax in the evaluation of asbestosis. Eur Respir J 1998; SIGNAL INTENSITY IN MUSCLE 300 11: 194-7. 11. Falaschi F, Battolla L, Mascalchi M, Cioni R, Zampa V, Lencioni R, et al. 250 Usefulness of MR signal intensity in distinguishing benign from malignant pleural disease. AJR Am J Roentgenol 1996; 166: 963-8. 200 12. Knuuttila A, Kivisaari L, Kivisaari A, Palomäki M, Tervahartiala P, Mattson K. Evaluation of pleural disease using MR and CT. With special reference to 150 malignant pleural mesothelioma. Acta Radiol 2001; 42: 502-7. 13. Zwitter M, Tumorji torakalnih organov. In: Novakovic S, Hocevar M, 100 Novakovic BJ, Strojan P, Žgajnar J, editors. Onkologija. Raziskovanje, diag­nostika in zdravljenje raka. 1. edition. Ljubljana: Mladinska knjiga; 2009. p. 295-6. 50 0 SIGNAL INTENSITY IN LESION 500 SIGNAL INTENSITY IN MUSCLE 150 400 350 300 250 200 150 100 50 0 References 1. Hmeljak J, Cör A. Presence and role of Simian Virus 40 (SV40) in malignant pleural mesothelioma. Radiol Oncol 2009; 54: 9-16. 2. Weber MA, Bock M, Plathow C, Wasser K, Fink C, Zuna I, et al. Asbestos-related pleural disease: value of dedicated magnetic resonance imaging techniques. Invest Radiol 2004; 39: 554-64. 3. Wang ZJ, Reddy GP, Gotway MB, Higgins CB, Jablons DM, Ramaswamy M, et al. Malignant pleural mesothelioma: evaluation with CT, MR imaging, and PET. Radiographics 2004; 24: 105-19. 4. Kim HY, Yi CA, Lee KS, Chung MJ, Kim YK, Choi BK, et al. Nodal metastasis in non-small cell lung cancer: accuracy of 3.0-T MR imaging. Radiology 2008; 246: 596-604. 5. Lutterbey G, Gieseke J, von Falkenhausen M, Morakkabati N, Schild H. Lung MRI at 3.0 T: a comparison of helical CT and high-field MRI in the detection MPM MPMMPMMPMMPMMPMMPMMPMMPMMPMPLAQUEPLAQUEPLAQUEPLAQUEPLAQUE of diffuse lung disease. Eur Radiol 2005; 15: 324-8. 6. Kauczor HU, Kreitner KF. Contrast-enhanced MRI of the lung. Eur J Radiol 2000; 34: 196-207. FIGURE 4. Signal intensity measurement on T2 fs-weighted images. MPM = malignant pleural mesothelioma The results of our preliminary study show that CT is still a gold standard in imaging patients with thoracic diseases, also because the large number of CT scanners is available. 3T MRI is a promising method which can be nowadays used as a comple­mentary method. research article Reliability of diffusion weighted MR imaging in differentiating degenerative and infectious end plate changes Ozgur Oztekin1, Cem Calli2, Omer Kitis2, Zehra Hilal Adibelli1, Cemal Suat Eren3, Melda Apaydin4, Mehmet Zileli5,Taskin Yurtseven5 1 Radiology Department, Izmir Education and Research Hospital, Izmir, Turkey 2 Department of Radiology, Ege University Medical School, Izmir, Turkey 3 Radiology Department, Tepecik Education and Research Hospital, Izmir, Turkey 4 Radiology Department, Ataturk Education and Research Hospital, Izmir, Turkey 5 Department of Neurosurgery, Ege University Medical School, Izmir, Turkey Received 12 June 2009 Accepted 15 September 2009 Correspondence to: Dr. Ozgur Oztekin, Albayrak mavisehir evleri, Yali mahallesi, 6525sok. No: 35 daire No: 31 Karsiyaka, Izmir, Turkey. Phone: +9053 2333 0201, +9050 5376 7387; Fax: +9023 2250 5050; E-mail: oztekinozgur@gmail.com Disclosure: No potential conflicts of interest were disclosed. Part of this study was presented as an poster presentation in 17th Symposium Neuroradiologicum of European Society of Neuroradiology (ESNR) Paris, France, 2002. Background. The aim of the study was to investigate the value of diffusion weighted MR imaging in the diagnosis of Modic type 1 change, which may be confused with the acute infectious spondylodiscitis on conventional MR imaging. Patients and methods. Twenty-seven patients with erosive intervertebral osteochondrosis, Modic type 1 and 18 patients with spondylodiscitis were included in this retrospective study. All images were acquired using on 1.5 Tesla MR units. Lumbar spinal MR imaging of 45 patients were retrieved from a digital database of a radiology record system and evaluated by one experienced radiologist. Patients with Modic type 1 change had CT slices obtained from the diseased disc space and the affected vertebrae. Results. Bone marrow adjacent to the vertebral end plate in both Modic type 1 change and acute spondylodis­citis were hypointense on T1-weighted images. On T2-weighted images corresponding levels of vertebral end-plates showed hyperintense signal intensity in both group. All the patients with spondylodiscitis and Modic type 1 change were hyperintense and hypointense on diffusion-weighted MR images, respectively. Conclusions. Our findings suggest that diffusion weighted MR imaging is an useful method in differentiating Modic type 1 changes from acute spondylodiscitis, both of which may mimic each other, either on clinical or conventional MRI findings. Key words: Modic type 1 change; spondylodiscitis; magnetic resonance imaging; diffusion-weighted imaging; verte­bral end-plate. Introduction Vertebral end-plate abnormalities of the lumbar spine are commonly seen on MR images.1Of these abnormalities most of them are frequently associ­ated with degeneration. Degenerative vertebral end-plate changes were first described independently by Roos et al.2and Modic et al.3as being a feature associated with the degenerative disk disease. These changes, also called as erosive intervertebral osteochondrosis (EIVO), were classified into three groups by Modic et al.3Type 1 change is vascular granulation tis­sue, demonstrated as low signal intensity on T1­weighted images (T1WI) and high signal intensity on T2-weighted images (T2WI). They are associ­ated with fissuring of cartilaginous end-plate and increased vascularity within the subchondral bone marrow on the histological examination.4Type 2 change is fatty infiltration of the end-plates, dem­ 98 Oztekin O et al. / MR in differentiating degenerative and infectious end plate changes FIguRe 1. Sagittal MR images from 53 years old male patient with low back pain. A) T1-weighted MR image demonstrating low-signal intensity changes adjacent to the L4-5 disk, and B) T2-weighted MR image of the same level demonstrating high-signal intensity changes. C) On post-contrast T1-weighted images end-plates disclose signal intensity increase, which can also be seen in spondylodiscitis. D) On DWI end-plates showing low signal intensity changes consistent with Modic type 1 change at L4 through L5. E) and F) Axial CT slices obtained from the L4-5 disk level showing discal vacuum phe­nomenon and sclerosis, supporting the diagnosis of degenerative disc disease. onstrated as hyper intense signal intensity on T1WI and hyper intense or isointense signal intensity on T2WI. In such cases biopsy reveals the fatty re­placement of the marrow, which is thought to be the result of marrow ischemia.4Type III changes consist of reduced signal intensity on both T1- and T2-weighted images representing bone sclerosis.3 Intervertebral disk space infections typically give rise to vertebral marrow oedema, manifesting as areas of low signal intensity on T1WI and high signal intensity on T2WI.5 Thereby, type 1 Modic changes may cause a di­agnostic dilemma in patients with low back pain since sometimes it resembles the MRI features of spondylodiscitis.6 Thediffusionweightedimaging(DWI)hasre­centlybeenusedinthespinebymanyauthors, mainlyforthedifferentiationofbenignandmalign oedemaofthevertebralbody.7-9DWHmightbe alsousefulinthedifferentialdiagnosisofbenign frommalignantlesionsinotherorgans.10Ithasbeen reportedthatbenignfractureoedemadepictshypo-isointensityonDWIwhereasmaligninfiltrationof thevertebraediscloseshyperintensity.Todistin­guishthebenignfromthemalignantdifferencesis crucialtochoosetherighttreatment.11DWIhasalso beenusedinspondylodiscitis.Ithasbeenshown thatDWIrevealshyperintensityintheaffectedver­tebraeandtheparavertebralinfectioussofttissue inacutespondylodiscitis.12Thepurposeofthisin­vestigationwastoevaluatetheusefulnessofdiffu­sionweightedMRimagingforthedifferentiationof Modictype1changesfromacutespondylodiscitis, bothofwhichmaymimickeachother,eitheronthe clinicalevaluationorconventionalMRIfindings. Patients and methods Patients Forty-five patients (18 patients with acute spond­ylodiscitis and 27 patients with Modic type 1 change) who underwent lumbar MRI examinations between January 2001 and December 2008 were in­cluded after a review of a digital database of a ra­diology record system. They were identified from a total of 1400 MR imaging examinations of the lumbar spine with a low back pain performed dur­ing this time at our institution. Patients with signal abnormalities limited to having previous surgery, recent vertebral fracture, metastatic disease, and pregnancy were excluded from the study. SpondylodiscitiswereprovenwithCT-guided biopsyin12patients.In6patientsthediagnosisof spondylodiscitiswerebasedonlaboratoryfindings. The diagnosis of Modic type 1 change were prov­en either by clinical or laboratory findings. In order to confirm the diagnosis radiologically two year-follow-up MRI was assessed. On the follow-up Modic type 1 in 8 of 27 patients partially converted Modic type 2 and 14 of 27 patients fully converted Modic type 2. Five of 27 patients were stable, and showed no change. CT slices were obtained from the diseased disc space and the affected vertebrae in all the patients to support the diagnosis. Spondylodiscitiswasdiagnosedinthepresence ofparavertebralorepiduralsignalabnormalities withorwithoutabscessformation.Ifsuchfindings wereabsent,threeofthefollowingfourcriteriahad tobefulfilledforthedisk-spaceinfection:signal abnormalityofthebonemarrowadjacenttothein­ Oztekin O et al. / MR in differentiating degenerative and infectious end plate changes 99 FIguRe 2. Sagittal MR images from 48 years old male patient with low back pain. A) Sagittal T1-weighted and B) Sagittal T2-weighted MR images showing low and high signal respectively at both end-plates of L4-5. C) On post-contrast T1-weighted images same level demonstrating heterogenic contrast enhancement. D) On DWI end-plates demonstrating low signal intensity changes consistent with Modic type 1 change at L4-5. E) Axial CT slices obtained from the L4-5 disk level and F) Coronal and G) Sagittal reconstructed CT image showing sclerosis on both end-plates of L4-5. tervertebraldisk(hypointenseonT1-weightedim­agesandhyperintenseonT2-weightedimages,sig­nalnotwelldemarcated);lossofthelow-intensity vertebralendplateonT1-weightedimages;hyperin-tensityofthediskonT2-weightedimages;anddisk enhancementaftertheinjectionofgadopentetate. All patients gave a written informed consent to use their clinical data for the study purposes. The study protocol was approved at the research ethics review committee of the hospital. Imaging technique A standard lumbar MRI protocol was employed to all patients. All images were acquired using on 1.5 Tesla MR units. All MR studies were performed on a 1.5 T unit (Magnetom Vision, Siemens, Erlangen, Germany) with gradient echo-planar capabilities and a standard phased array surface receiver coil for imaging the spine. The imaging protocol in­cluded axial and sagittal T1-weighted spin-echo sequences (552/ 12[TR/TE]), axial and sagittal T2­weighted turbo spin-echo (4000/120[TR/TE]) se­quences, sagittal STIR(3600/60[TR/TE]) sequences, and sagittal diffusion-weighted sequences. Sagittal spinal DW images (b = 150 s/mm2) were acquired in the same plane and orientation as used in the routine sequences by using a reversed fast imag­ing with steady-state precession (PSIF) sequence (TR/TE 1.400 ms/100 ms; field of view 320×80 mm; section thickness 5 mm; intersection gap 0.5 mm; sections 6; matrix 128×256; echo train length 69 and one excitation) with spectral presaturation and inversion recovery (SPIR). In addition, axial and sagittal fat-suppressed T1-weighted images were obtained after IV infusion of 0.1 mmol/kg of gado­pentate dimeglumine. Image assessment All MR images were reviewed and evaluated by one radiologist specializing in MR imaging of the spinal system. The abnormal levels were classified as either infected or degenerative. Signal intensity changes of the related disc and vertebral body marrow adjacent to the end plates of the degenerative spine on the conventional spin-echo sequence MR and the diffusion weighted MR were compared with those of spondylodiscitis. We categorized the signal intensity of the abnor­mal vertebra on T1-Weighted images as hypoin-tense relative to the presumed normal marrow. The signal of the abnormal vertebra on T2- weighted images was categorized as hypointense, isointense or hyperintense relative to the areas of the pre­sumed normal marrow. On the diffusion-weighted images, the areas of the abnormal signal intensity were categorized as hypointense, isointense and hyperintense with respect to the normal marrow. Statistical analyses The statistical analysis was carried out by using Statistical Package of Social Science (SPSS), version 13.0. Results The mean ages of patients with Modic type 1 change (Figures 1, 2) and spondylodiscitis (Figure3) were 52.2 years (range, 24-77 years) and 55.8 years (range, 18-85 years), respectively. These 27 patients had a total of 62 Modic chang­es and 18 patients with acute spondylodiscitis had 100 Oztekin O et al. / MR in differentiating degenerative and infectious end plate changes FIguRe 3. Sagittal MR images from 38 years old male patient with acute spondy­lodiscitis. A) T1-weighted image showing complete homogenous hypointensity at the L1 and L2 vertebrae corpus and adjacent intervertebral disc. There are also osteophytic changes on end-plate and loss of L1-2 intervertebral disc space. B) T2-weighted images showing hyperintense signal intensity corresponding to the same level. C) Postcontrast image showing homogenous enhancement of disc and adjacent vertebrae corpus. D) DWI revealing high signal intensity relative to neigh-bouring normal vertebrae. 46 vertebral involvement. Four patients with type 1 Modic changes and 5 patients with spondylodisci-tis had more than two vertebral involvements. On CT slices of affected vertebrae in patients with Modic type 1 change, 5 patients (18.5%) had discal vacuum phenomenon, 22 patients (81%) had well-defined sclerosis, and 4 patients (14.8%) had erosions of vertebral endplates without bone de­struction. Bone marrow adjacent to the vertebral end plate in both Modic type 1 change and and acute spondylodiscitis were hypointense on T1-weighted images. The hypointense areas in the vertebral end plates on T1-weighted images in patients with Modic type 1 change enhanced either moderately or strongly on postcontrast images, a finding which could not be differentiated from a disc space infec­tion. On T2-weighted images corresponding levels of vertebral end-plates showed hyperintense signal intensity in both groups. On diffusion-weighted MR images with relatively low b values, all verte­bral body marrow and end-plates with Modic type 1 change showed hypointense to normal signal intensities. Conversely, all vertebral body marrow and end-plates with acute infectious spondylo­discitis showed increased signal intensities when compared to presumed normal vertebrae. Discussion MR imaging (MRI) is commonly used in the di­agnosis of patients with low back pain (LBP) and sciatica.1In the search for causes of LBP, vertebral end-plate signal changes have come into focus. Vertebral end-plate changes are bone marrow and end-plate lesions visible in MRI. Different pathological processes can involve vertebral bone marrow adjacent to the end-plates, including degenerative disc disease, infection and tumours, and these may present with a variety of signal intensity as shown by MRI. After the initial description of Modic changes some studies have attempted to identify the cause of such changes. Modic type 1 changes were found to be associated with fissuring of the cartilaginous end-plate and increased vascularity within the subchondral bone marrow on the histological ex­amination.4Vertebral end-plate changes consistent with bone marrow oedema may also be seen in in­fective discitis, following intraosseous disc hernia­tion (Schmorl’s nodules) and within 3 months of chemonucleolysis.13-15 Spondylodiscitis is an infection of the interverte­bral disk and the adjacent vertebrae, with or with­out associated epidural or psoas abscesses. It is a serious diseaseboth due to its long-term course and the possible outcomes.16 Type 1 Modic change and spondylodiscitis may both reveal similar symptoms, mainly LBP. Clinical and laboratory findings such as white blood cell count, erythrocyte sedimentation rate and elevated body temperature are, supportive but not confirm­atory evidence in infectious spondylodiscitis.17 Stirling et al.18suggested a theory that bacteria might play a causative role in LBP in Modic type 1 changes and that patients might benefit from the antibiotic treatment. Stirling et al.18found bacteria in disc material from 19 of 36 patients with severe sciatica and Albert et al.19showed that 17 out of 32 patients with Modic type 1 changes and persist­ent LBP achieved long-lasting pain relief after the long-term antibiotic treatment. But Wedderkopp et al.20showed no evidence of bacteria in vertebrae in Modic type 1 changes in their recent study, al­though in this study possible presence of bacteria in the disc adjacent to the Modic type 1 changes in the vertebrae cannot be ruled out. Fayad et al.21found that patients with chronic LBP and predominantly type 1 Modic changes had a better short-term relief of symptoms following in-tradiskal steroid injection than those with predom­inantly type 2 changes, which further supports the inflammatory nature of Modic type 1 changes and the role of inflammation in the generation of LBP. Although the aetiology of the inflammatory process in the vertebrae is still unknown in Modic type 1 change, there are two accepted possible Oztekin O et al. / MR in differentiating degenerative and infectious end plate changes 101 theories.20The most well known theory is that the process is a part of the ‘‘normal’’ degenerative process of the spine, where dehydration of the nu­cleus and loss of disc height biomechanically leads to unphysiological load and shear forces causing microfractures followed by inflammation in the vertebral end-plate and adjacent bone marrow.20A second theory is that anaerobic bacteria with low virulence enter the vertebra from the bloodstream, resulting in infection, which is represented on MRI as Modic type 1 changes.20 On MRI, type 1 change reveals hypointensity on T1-weighted and hyperintensity on T2-weighted images. Affected bone marrow may show mild or strong contrast enhancement after the intravenous contrast administration. Differentiation of these findings from those of spondylodiscitis may not be always possible based on the conventional MRI findings alone.6,21,22CT may be helpful in the dif­ferentiation showing discal vacuum phenomenon or well-defined sclerosis and erosions of verte­bral endplates without bone destruction, findings that support the diagnosis of EIVO. In our patient group, CT revealed these changes compatible with EIVO at the end-plates adjacent to the degenerated disc. DWI imaging has been successful to some extent in the differentiation of benign versus pathologic compression fractures of the vertebral body.7-9,12It has been shown that benign bone marrow oedema has revealed hypointensity on DWI and malign vertebral compressions have disclosed hyperinten­sity in the vertebral bone marrow.7,9Recently it has been reported that infective discitis has also shown hyperintensity on DWI as well as malignancies of the affected vertebral bone marrow.12However, in this study we have found hypointensity on DWI at the Modic type 1 disorders. In general, histopathologic findings of the type 1 bone marrow are fibrovascular tissue totally re­placing normal marrow elements.5A possible ex­planation for our results on diffusion-weighted MR imaging is that in the fibrovascular degenera­tive change, the increased free water of bone mar­row caused by depletion of normal marrow ele­ments leads to an increase in the extracellular vol­ume fraction which produces low signal intensity in diffusion-weighted MR imaging. In contrast, in spondylodiscitis the reduction of the extracellular volume due to densly infiltrated inflammatory cells might lead to the increase in the signal inten­sity on diffusion-weighted MR imaging. Although there are some clues for the diagno­sis of type 1 changes on routine MRI, it sometimes may be very difficult to differentiate it from early onset spondylodiscitis. Intervertebral disk space infections typically give rise to vertebral marrow edema, manifesting as areas of low signal intensity on T1WI and high signal intensity on T2WI, there­by mimicking type 1 Modic changes. The contrast enhancement in the disk and endplates may occur in both conditions. Moreover, the enhancement of the intervertebral disc itself is not a definitive rule for spondylodiscitis, since sometimes this cannot be detected. Because of desiccation and dehydra­tion, the disk often appears normal or hypointense on T2WI in degenerative disc disease, whereas its T2WI signal intensity is typically increased in spondylodiskitis.21If there is enhancing paraverte­bral soft tissue mass adjacent to the intervertebral disc space, the MRI findings should orient the di­agnosis toward an infectious process.6The verte­bral endplates are usually preserved in degenera­tive disc disease rather than destroyed or eroded with the bone destruction and the presence of these findings should also remind the possibility of disk space infection.23 However, in the absence of these findings, the decision should be made between EIVO type I and spondylodiscitis. We believe that DWI can make this differentiation easily and elimi­nate the necessity of CT in these groups of patients. There are some limitations to the study pre­sented here. The interpretations of the MR images were performed by only one experienced radiolo­gist. Thus, the interobsever variability and accu­racy associated with a less experienced radiologist was not assessed. Another limitation was the small number of patients who participated in the study. The diagnosis of Modic type 1 was relied on the clinical and radiological follow-up and we had no histopathologic correlation in our study. Further studies comparing histopathologic results and DWI findings are needed. In clinical practice, most radiologic and clini­cal findings are sufficiently supportive for the differential diagnosis of pathological processes involving vertebral body marrow adjacent to the end-plates. But, sometimes Modic type 1 change causes a diagnostic dilemma in patients with low back pain since it has almost the same conven­tional MR imaging and clinical findings with the acute spondylodiscitis. Moreover postcontrast MR imaging will not lead to the differential diagnosis since the Modictype 1 change will also enhance as well as spondylodiscitis. However, DWI appears to be useful in the differentiation of EIVO from the acute spondylodiscitis with distinct documented features in this study. 102 Oztekin O et al. / MR in differentiating degenerative and infectious end plate changes For this reason, when results of clinical and con­ventional MR findings are equivocal, diffusion-weighted MR imaging may provide an excellent differential diagnosis between degenerative fibrov­ascular change of the spine and pyogenic spond­ylitis. Acknowledgement I would like to express my special appreciation to Ege University Medical School, Department of Radiology in the name of Professor Cem Calli for his permition of usage of their archive. References 1. Zhang YH, Zhao CQ, Jiang LS, Chen XD, Dai LY. Modic changes: a system­atic review of the literature. Eur Spine J 2008; 17: 1289-99. 2. de Roos A, Kressel H, Spritzer C, Dalinka M. MR imaging of marrow changes adjacent to end plates in degenerative lumbar disk disease. AJR Am J Roentgenol 1987; 149: 531-4. 3. Modic MT, Steinberg PM, Ross JS, Masaryk TJ, Carter JR. Degenerative disk disease: assessment of changes in vertebral body marrow with MR imaging. Radiology 1988; 166: 193-9. 4. Braithwaite I, White J, Saifuddin A, Renton P, Taylor BA. Vertebral end-plate (Modic) changes on lumbar spine MRI: correlation with pain repro­duction at lumbar discography. Eur Spine J 1998; 7: 363-8. 5. Rahme R, Moussa R. The Modic vertebral endplate and marrow changes: pathologic significance and relation to low back pain and segmental instability of the lumbar spine. AJNR Am J Neuroradiol 2008; 29: 838-42. 6. Stabler A, Reiser MF. Imaging of spinal infection. Radiol Clin North Am 2001; 39: 115-35. 7. Baur A, Reiser MF. Diffusion-weighted imaging of the musculoskeletal system in humans. Skeletal Radiol 2000; 29: 555-62. 8. Baur A, Huber A, Ertl-Wagner B, Dürr R, Zysk S, Arbogast S, et al. Diagnostic value of increased diffusion weighting of a steady-state free precession sequence for differentiating acute benign osteoporotic fractures from pathologic vertebral compression fractures. AJNR Am J Neuroradiol 2001; 22: 366-72. 9. Oztekin O, Ozan E, Hilal Adibelli Z, Unal G, Abali Y. SSH-EPI diffusion-weighted MR imaging of the spine with low b values: is it useful in dif­ferentiating malignant metastatic tumor infiltration from benign fracture edema? Skeletal Radiol 2009; 38: 651-8. 10. Inan N, Kilinc F, Sarisoy T, Gumustas S, Akansel G, Demirci A. Diffusion weighted MR imaging in the differential diagnosis of haemangiomas and metastases of the liver. Radiol Oncol 2010; 44: 24-9. 11. Rajer M, Kovac V. Malignant spinal cord compression. Radiol Oncol 2008; 42: 23-31. 12. Balliu E, Vilanova JC, Pelaez I, Puig J, Remollo S, Barceló C, et al. Diagnostic value of apparent diffusion coefficients to differentiate benign from malignant vertebral bone marrow lesions. Eur J Radiol 2009; 69: 560-6. 13. Kato F, Ando T, Kawakami N, Mimatsu K, Iwata H. The increased signal intensity at the vertebral body endplates after chemonucleolysis demon­strated by magnetic resonance imaging. Spine 1993; 18: 2276-81. 14. Dagirmanjian A, Schils J, McHenry M, Modic MT. MR imaging of vertebral osteomyelitis revisited. AJR Am J Roentgenol 1996; 167: 1539-43. 15. Stabler A, Bellan M, Weiss M, Gartner C, Brossmann J, Reiser MF. MR imaging of enhancing intraosseous disk herniation (Schmorl’s nodes). AJR Am J Roentgenol 1997; 168: 933-8. 16. StumpeKD,ZanettiM,WeishauptD,HodlerJ,BoosN,VonSchulthessGK. FDG positron emission tomography for differentiation of degenerative and infectious endplate abnormalities in the lumbar spine detected on MR imaging. AJR Am J Roentgenol 2002; 179: 1151-7. 17. Fouquet B, Goupille P, Jattiot F, et al. Discitis after lumbar disc surgery. Features of “aseptic” and “septic” forms. Spine 1992; 17: 356-8. 18. Stirling A, Worthington T, Rafiq M, Lambert PA, Elliott TS. Association be­tween sciatica and propionibacterium acnes. Lancet 2001; 357: 2024-5. 19. Albert HB, Manniche C, Sorensen JS, Deleuran BW. Antibiotic treatment in patients with low-back pain associated with Modic changes Type 1 (bone oedema): a pilot study. Br J Sports Med 2008; 42: 969-73. 20. Wedderkopp N, Thomsen K, Manniche C, Kolmos HJ, Secher Jensen T, Leboeuf Yde C. No evidence for presence of bacteria in Modic type I changes. Acta Radiol 2009; 50: 65-70. 21. Fayad F, Lefevre-Colau MM, Rannou F, Quintero N, Nys A, Macé Y, et al. Relation of inflammatory Modic changes to intradiscal steroid injection outcome in chronic low back pain. Eur Spine J 2007; 16: 925-31. 22. OztekinO,CalliC,AdibelliZ,KitisO,ErenC,AltinokT.Brucellarspondylodisci-tis: magnetic resonance imaging features with conventional sequences and diffusion-weighted imaging. Radiol Med DOI10.1007/s11547-010-0530-3 23. CalliC,YuntenN,KitisOetal.Diffusion-weightedMRimaginginspondylo­discitis and vertebral malignancies: a comparative study. Neuroradiology 2001; 42 (suppl 1):S55 103 case report Post-traumatic high-flow priapism treated by endovascular embolization using N-butyl-cyanoacrylate Marko Rados, Vice Sunjara, Ivica Sjekavica, Ranka Stern Padovan University Department of Diagnostic and Interventional radiology, University Hospital Center Zagreb, Zagreb, Croatia Received 5 October 2009 Accepted 25 November 2010 Correspondence to: Vice Šunjara, MD, Klinicki zavod za dijagnosticku i intervencijsku radiologiju, KBC Zagreb – Rebro, Kišpaticeva 12, 10000 Zagreb, Croatia. Phone: +385 98 1642 482, Fax: +385 1 2388 250; E-mail: vsunjara@yahoo.com Disclosure: No potential conflicts of interest were disclosed. Background. Priapism, persistent erection without arousal, can be classified into low-flow (venous or ischemic) and high-flow (arterial or non-ischemic). The diagnosis of high-flow priapism can be confirmed by colour Doppler and arteriography and it is usually treated by the endovascular embolization. Case report. We present a case of a 20-year-old man with a post-traumatic high-flow priapism as a result of the previous perineal trauma. After a period of watchful waiting and an unsuccessful attempt at endovascular emboliza­tion using the resorptive gelatinous foam he was successfully treated by the endovascular embolization using N-butyl­cyanoacrylate. Conclusions. High-flow priapism can be successfully treated by the endovascular embolization, but the optimal choice of the embolization agent and a careful technique is essential. Key words: priapism; endovascular embolization; angiography; Doppler duplex ultrasonography; MRI angiography Introduction Priapism is a relatively rare condition character­ized by the persistent erection in the absence of sexual arousal. There are two main subtypes: the more common ischemic, or low-flow, character­ized by the impaired outflow from the corpora cav­ernosa, and non-ischemic, or high-flow, most often caused by trauma, characterized by the formation of arteriocavernous fistulas and increased inflow of blood to the corpora cavernosa. While the painful low-flow priapism and the associated decreased oxygenation of cavernous tissue can quickly lead to a cavernous fibrosis and permanent damage to penile tissues and is, therefore, an urological emergency, high-flow priapism is often painless and can persist for months or years, in most cases without a permanent damage of penile tissues, but sometimes with the reduced potency.1 The diagnosis of high-flow priapism can be con­firmed by colour Doppler2, which can also be used to characterize the number and location of arterio-cavernous fistulas and concomitant arterial pathol­ogy such as pseudoaneurysms. Colour Doppler is also useful in the follow-up, avoiding the repeated angiography with its risks and the ionizing radia­tion dose, although the MR angiography is usually necessarily to evaluate the effect of the radiological invasive interventional procedures.3 There are many treatment options in high-flow priapism: those mentioned most often are watchful waiting4, Doppler-guided compression5, endovas­cular highly selective embolization and surgery. Because the more aggressive treatment methods are associated with a small but significant rate of the permanent erectile dysfunction, an initial watchful waiting period is commonly indicated. The surgery in high-flow priapism usually consists of the ligation of a cavernous artery or its branch and is reported to have the highest permanent erectile dysfunction rate, thus it is usually the last treatment option. 104 Rados M et al. / Embolization of post-traumatic priapism Doppler sonogram of cavernous artery which fills the pseudoaneurysm. D. Pulsed Doppler analysis with aliasing phenomena due to turbulent high-velocity flow in the cavernous artery. E. Venous drainage in the corpora cavenosa on Doppler sonogram. Case report fro signals into suspected cavernoma and a high velocity flow in the cavernous artery which fills A 20-year old patient presented with priapism the pseudoaneurysm (Figure 1C and 1D). Venous caused by previous perineal trauma. Gray-scale ul-drainage in corpora cavenosa was also found on trasound depicted anehoic region, 14.7 x 12.7 mm in Doppler examination (Figure 1E) size, within corpus cavernosum (Figure 1A) Colour The initial arteriography confirmed a pseudoan- Doppler ultrasound showed multiple colour sig-eurysm of the right cavernous artery with an arteri­ nals due to the extravasation of blood (Figure 1B). ocavernous fistula (Figure 2). A smaller arteriocav- The pulsed Doppler analysis confirmed typical to-ernous fistula was also present on the left cavern­ Rados M et al. / Embolization of post-traumatic priapism 105 FIGURE 2. Selective angiography before embolization shows the arteriocavernous fistula (marked with an arrow). ous artery. The communication of the left and right internal pudendal artery was noted, with the blood from the left pudendal artery flowing to the right and contributing to the filling of the pseudoaneu­rysm on the right. After a six-month period of watchful wait­ing priapism did not resolve spontaneously and a more aggressive approach was decided upon with an attempt at highly selective embolization of the fistulas of both cavernous arteries using the resorptive gelatinous foam. One-month follow-up showed a recurrence of the right-sided fistula ne­cessitating another embolization procedure dur­ing which the superselective catheterization was performed and a microcatheter was inserted into the pseudoaneurysm on the right cavernous ar­tery. The embolization agent used was N-butyl­cyanoacrylate (Glubran II, GEM S.r.l.,Viareggio, Italy). Two-month follow-up showed the closure of arteriocavernous fistulas with the persistence of pseudoaneurym on the right that had morphed in­to a small cavernoma, which was embolized using additional N-butyl-cyanoacrylate. The end-result was a complete occlusion of the fistula (Figure 3). Priapism was successfully resolved and the patient remained symptom-free and regained the erectile function. Contrast-enhanced MR angiography fol­low-up at 6 months showed no recurrence of the fistula (Figure 4). FIGURE 3. Selective angiography after the second emboliza­tion shows the occlusion of arteriocavernous fistula (filling artery marked with an arrow). 106 Rados M et al. / Embolization of post-traumatic priapism Discussion The endovascular selective embolization of the pathological arteriocavernous communication is firmly established as the invasive treatment of choice in high-flow priapism.6,7It is commonly performed using microcatheters and a range of embolization materials: autologous clots, ge­latinous foam, endovascular coils8or N-butyl­cyanoacrylate.9,10Autologous clots and gelatinous foam are often preferred because of their spontane­ous degradation and a reportedly lower risk of the permanent erectile dysfunction, but could have a greater recurrence rate. After our first unsuccessful attempt at emboli­zation using resorptible embolization materials we switched to N-butyl-cyanoacrylate (Glubran II) which could provide faster and more efficient occlusion of the fistula. The cyanoacrylate em-bolization is permanent and carries a higher risk of ischemia of the vessel in question, and it con­sequently requires a better embolization technique and more experienced interventionists capable of introducing the catheter and the embolization ma­terial directly into the site of the fistula. In our case the treatment was facilitated by the fistula being positioned on a pseudoaneurysm of the cavern­ous artery. Even though the fistula was occluded after the first embolization session with N-butyl­cyanoacrylate we elected to perform an additional session to obliterate the residual cavernoma in or­der to prevent a possible recanalization of the fis­tula and recurrence. Our case showed that embolization using N-butyl-cyanoacrylate (Glubran II) could be used as a second-line treatment in patients with recur­rence after the first embolization attempt with re-sorptible materials. References 1. Hakim LS, Kulaksizoglu H, Mulligan R, Greenfield A, Goldstein I. Evolving concepts in the diagnosis and treatment of arterial high flow priapism. J Urol 1996; 155: 541-8. 2. Feldstein VA. Posttraumatic “high-flow” priapism evaluation with color flow Doppler sonography. J Ultrasound Med 1993; 12: 589-93. 3. Gjikolli1 B, Hadžihasanovic B, Jaganjac S, Herceglija E, Nikšic M, Hadzimehmedagic A, et al. Treatment of complicated case with subclavia steal syndrome and stenosis of common iliac artery. Radiol Oncol 2008; 42: 1-12. 4. Hatzichristou D, Salpiggidis G, Hatzimouratidis K, Apostolidis A, Tzortzis V, Bekos A, et al. Management strategy for arterial priapism: therapeutic dilemmas. J Urol 2002; 168: 2074-7 5. Mabjeesh NJ, Shemesh D, Abramowitz HB. Posttraumatic high flow priapism: successful management using duplex guided compression. J Urol 1999; 161: 215-6. 6. SavocaG,PietropaoloF,ScieriF,BertolottoM,MucelliFP,BelgranoE.Sexual function after highly selective embolization of cavernous artery in patients with high flow priapism: long-term followup. J Urol 2004; 172: 644-7. 7. O’Sullivan P, Browne R, Mceniff N, Lee MJ. Treatment of “high-flow” pria­pism with superselective transcatheter embolization: a useful alternative to surgery. Cardiovasc Intervent Radiol 2006; 29: 198-201. 8. Abujudeh H, Mirsky D. Traumatic high-flow priapism: treatment with super-selective micro-coil embolization. Emerg Radiol 2005; 11: 372-4. 9. Fratezi AC, Martins VM, Pereira Porta RM, Prado MA, Prota R, Caldas JG, et al. Endovascular Therapy for Priapism Secondary to Perineal Trauma. J Trauma 2001; 50: 581-4. 10. Kutlu R, Soylu A. Deep dorsal vein embolization with N-butyl-2­cyanoacrylate and lipiodol mixture in venogenic erectile dysfunction: earlyand late results. Radiol Oncol 2009; 43: 17-25. 107 research article Antigen expression on recurrent meningioma cells Andrej Vranic Department of Neurosurgery, University Medical Centre Ljubljana, Ljubljana, Slovenia Received 1 February 2010 Accepted 1 March 2010 Correspondence to: Assist. Andrej Vranic MD, MSc, Department of Neurosurgery, University Medical Centre, Zaloška 7, 1525 Ljubljana. E-mail: andrej.vranic@kclj.si Disclosure: No potential conflicts of interest were disclosed. Introduction. Meningiomas are intracranial brain tumours that frequently recur. Recurrence rates up to 20% in 20 years for benign meningiomas, up to 80% for atypical meningiomas and up to 100% for malignant meningiomas, have been reported. The most important prognostic factors for meningioma recurrence are meningioma grade, meningi­oma invasiveness and radicality of neurosurgical resection. The aim of our study was to evaluate the differences in antigenic expression on the surface of meningioma cells between recurrent and non-recurrent meningiomas. Methods. 19 recurrent meningiomas and 35 non-recurrent meningiomas were compared regarding the expression of MIB-1 antigen, progesterone receptors, cathepsin B and cathepsin L, using immunohistochemistry. Results. MIB-1 antigen expression was higher in the recurrent meningioma group (p=0.001). No difference in pro­gesterone receptor status between recurrent and non-recurrent meningiomas was confirmed. Immunohistochemical intensity scores for cathepsin B (p= 0.007) and cathepsin L (p<0.001) were both higher in the recurrent than in the non-recurrent meningioma group. Conslusions. MIB-1 antigen expression is higher in recurrent compared to non-recurrent meningiomas. There is no difference in expression of progesterone receptors between recurrent and non-recurrent meningiomas. Cathepsins B and L are expressed more in recurrent meningiomas. Key words: meningioma; recurrence; tumour markers; proliferation index, MIB-1 antigen; cathepsin B; cathepsin L Introduction Meningiomas represent 10-20% of primary intrac­ranial tumours and show a wide range of histo-morphological subtypes. According to signs of ma­lignancy in their histological picture, they are clas­sified by the WHO classification as benign men-ingiomas (BM - grade I), atypical meningiomas (AM - grade II) and malignant meningiomas (MM - grade III). Although in general meningiomas are considered as slow-growing benign tumours, re­currence rates are quite high. Over a 20 year peri­od, the recurrence rate for BM is reported to be 10­26%, for AM 50-80% and for MM 78-100%.1-4The average recurrence-free interval ranges between 4 and 6.5 years. The most important risk factors for the meningioma recurrence are the meningioma grade, meningioma invasiveness and thorough­ness of neurosurgical resection, which is a prin­cipal approach for any surgical treatment in the brain tumour patient.5,6Meningioma invasiveness has been observed in all meningioma grades.1 Biological markers with the prognostic value for the meningioma recurrence have been sought in the last decades. The proliferation index Ki67 (Ki67 index), assessed as the percentage of MIB-1 positive cells in the area of greatest proliferation, is the most frequently used proliferation index for meningiomas. The Ki67 index was shown to cor­relate with higher malignancy grades of meningi­omas and was observed to be significantly more expressed in recurrent meningiomas compared to non-recurrent ones.7However, the Ki67 index has not been confirmed as a statistically significant predictor of recurrence in gross-totally removed benign meningiomas.8,9 The loss of progesterone receptors (PR), which are normally expressed in two thirds of BM, seems 108 Vranic A / Antigen expression on recurrent meningioma to correlate with a higher meningioma grade and the decreased recurrence-free survival time of pa­tients with meningiomas.9-12Other biochemical fac­tors, like the epithelial membrane antigen, S100, ac­tivated caspase 3 and its inhibitor survivin, HER2, metalloproteases MMP-2 and MMP-9 and others have all been proposed as tumour markers signifi­cant for the meningioma recurrence.13-19 Cathepsins are intracellular cysteine proteases normally present in most tissues. A high expres­sion of cathepsins B and L was found in several malignant tumours.20-24A high expression of cathe­psin D was observed in benign meningiomas.25A higher expression of cathepsins B in L was found in AM and MM2 3,27, compared to BM. Cathepsins B and L were found to be expressed more in recur­rent meningiomas.26,28However, only three recur­rent meningiomas were studied and the need for the evaluation of cathepsins B and L expression on a larger series of recurrent meningiomas, was em­phasized.28 In the present study, we focused on expression of the MIB-1 antigen, PR, cathepsin B and cathep-sin L on meningioma cells. The aim of the study was to compare recurrent and non-recurrent men-ingiomas. Differences between three meningioma grades were also sought. Patients and methods In our retrospective study, 54 patients (32 fe­male and 22 male), aged from 19 to 78 (mean 55.5 years), operated for meningioma at the University Medical Centre Ljubljana in the years 1996 - 2001, were selected randomly for the study. Thirty-three meningiomas were diagnosed as BM, 11 as AM and 10 as MM. Meningiomas were operated on by different surgeons and they were described by sur­geons as gross-totally removed, graded as Simpson I or Simpson II. All meningiomas were intracranial: 24 cranial base, 16 hemispheric and 14 parasagit­tal meningiomas were studied. Nineteen patients were in the recurrent meningioma group, and 35 patients were in the non-recurrent meningioma group. In the recurrent meningioma group, the mean time to the recurrence from the first opera­tion was 4.5 years (1-14 years). In the non-recurrent meningioma group, tumours have not recurred for at least 5-10 years (mean 6.5 years). Tissue samples were fixed in formaldehyde and embedded in paraffin wax. 5µm thick sections stained with haematoxylin and eosin (H&E) were assessed by the pathologist with regard to grade of FIGURE 1. Expression of MIB-1 antigen in different meningioma grades comparing non-recurrent and recurrent meningiomas. malignancy according to the WHO classification. In the recurrent meningioma group, 6 tumours (34%) were benign, 5 (23%) were atypical and 8 (43%) were malignant. In the non-recurrent group, 27 (75%) were benign, 6 (18%) were atypical and 2 (7%) were malignant. Immunohistochemistry (IH) was performed on tissue sections cut from the most representative paraffin block of each tumour using the standard procedures. IH was performed according to the routine protocols used in everyday practice at the Institute of Pathology, Ljubljana. Primary mouse monoclonal antibodies against human cathepsin B and human cathepsin L (Krka, Novo mesto), primary mouse anti-human Ki67 monoclonal an­tibodies (clone MIB-1, No 7240, DAKO, Denmark) and primary mouse anti-human progesterone re­ceptor antibodies (clone PgR 636, No M3569 dilut­ed, DAKO, Denmark) were used. Antibodies were incubated with slides for 26 min at 40°C. The Ki67 index was calculated with the help of the Leica Q Prodit computer program (Leica, Germany), count­ing the percentage of MIB-1 labelled nuclei in the most affected region. Positive or negative proges­terone receptor status was determined. Tumours were consideredpositive even if there were only a few cells with a positive IH reaction. The intensity of the IH reaction between cathepsin B and L anti­ Vranic A / Antigen expression on recurrent meningioma 109 bodies and the tumour cells was scored from 0 to 5 by two independent observers. Cases with a differ-1 ent score at the beginning were discussed and the agreement was reached. Intensity and frequency of immunostaining for cathepsins B and L was scored with: 0 no staining; 1 very mild; 2 mild, 3 moderate, 0,8 4 strong and 5 very strong staining observed. Statistical analysis was performed, using SPSS 16.0 for Windows (SPSS Inc., USA). The recurrent meningioma group was compared with the non-recurrent one. Variables used in the analysis in­cluded meningioma grade, absence or presence of recurrence, proliferation index, PR status, and IH intensity scoresfor cathepsins B and L. Differences PR expression 0,6 0,4 in expression of biological markers were analyzed using independent-samples T-test. Significance of differences between recurrent and non-recurrent meningiomas, as well as between different histo­logical subgroups, was given as the p value; p<0.05 was considered significant. Results Differences between recurrent and non-recurrent meningiomas Proliferation index The proliferation index was higher in the recurrent meningioma group (p=0.001), regardless of the tu­mour grade. Progesterone receptor status No differences in PR status between recurrent and non-recurrent meningiomas were confirmed. Ten out of 19 recurrent and 18 out of 35 non-recurrent meningiomas had a positive PR status. PR status did not correlate with other biological markers. Protein expression of cathepsins B and L IH intensity scores for cathepsin B (p=0.007) and cathepsin L (p<0.001) were both higher in the recur­rent than in the non-recurrent meningioma group. Differences between meningioma grades Theproliferationindexincreasedwiththemenin­giomagrade(Figure1).Ahigherproliferationin­dex(p=0.006)andasignificantlossofPR(p=0.002) wereobservedinMMcomparedtoBM(Figure2). HigherIHintensityscoresforcathepsinB(p=0.007), andforcathepsinL(p=0.006),wereobservedinMM comparedtoBM(Figures3and4). 0,2 0 FIGURE 2. Expression of PR in different meningioma grades comparing non-recurrent and recurrent meningiomas. Differences between recurrent and non-recurrent meningiomas in the BM subgroup In the subgroup of 33 BM, recurrent BM expressed more cathepsin L (p=0.035) than non-recurrent BM. Discussion In our series of 54 meningiomas, treated in a single institution, we showed that MIB-1 antigen, cathep-sin B and cathepsin L were expressed more in re­current compared to non-recurrent meningiomas. No difference in PR expression between recurrent and non-recurrent meningiomas was noticed. Comparing different meningioma grades, we showed that higher meningioma grades express more MIB-1 antigen, less PR and more cathep-sins B and L. Differences between BM and MM were statistically important in all four parameters. Differences between BM and AM as well as differ­ences between AM and MM were statistically in­significant. Although often studied and even used in eve­ryday diagnostics of meningiomas, the prognostic significance of the Ki67 index remains poorly de­ 110 Vranic A / Antigen expression on recurrent meningioma 4,0 mean cathepsin B IH intensity score 3,0 2,0 1,0 0,0 FIGURE 3. IH intensity score of cathepsin B expression in differ­ent meningioma grades comparing non-recurrent and recur­rent meningiomas. fined.9,29,30 The Ki67 index significantly increases from BM through AM to MM, but there is a consid­erable overlap between different grades.9,29,31In a series of primary meningiomas of all three grades, Perry et al.reported a Ki67 index = 4.2% associated with the decreased recurrence-free survival in uni­variate but not in multivariate analysis.29 No correlation between the outcome and the Ki-67 index was found in 600 cases of benign men­ingiomas.9The principal limitation of the Ki67 in­dex seems to be the lack of standardization of the technique and difficulties in defining cut-off val­ues.9,29,30Our study confirms an important role of the Ki67 index in meningioma grading, suggesting different cell proliferation rates in different menin­gioma grades (Figure 1). No differences in PR expression between recur­rent and non-recurrent meningiomas were found in our study. PR were expressed more frequently in female patients` meningiomas. A significant loss of PR was observed in MM (Figure 2). The loss of PR could be responsible for malignant progres­sion of meningiomas with PR acting protectively. According to our observations, meningioma re­currences are most frequent for female patients at the onset of menopause. More research work is re­quired to confirm this observation. FIGURE 4. IH intensity score of cathepsin L expression in different meningioma grades comparing non-recurrent and recurrent meningiomas. Suggesting the invasive nature of recurrent men-ingiomas, most research work has been focused on proteases. Lysosomal proteases cathepsin B and L have been associated with tumour invasiveness.20-22They were considered as factors contributing to in­vasiveness of meningiomas.26-28Higher expressions of cathepsin B, metalloprotease-2 and metallopro­tease-9 were also detected in meningiomas, histo-logically described as invasive.18,32 Inthepresentstudy,wefoundthatcathepsinsB andLwereexpressedmoreinrecurrentmeningi­omascomparedtonon-recurrentones.Thisfinding isinaccordancewithpreviousreportsaboutcathe­psinexpressioninrecurrentmeningiomaswhich sofarhavenotbeenconfirmedonalargerseries.28Ourstudyshowsobviousdifferencesbetweenthe twogroups,suggestingrecurrentmeningiomas werebiologicallydifferent–i.e.moreinvasivethan non-recurrentmeningiomas.Ourfindingssupport theideaofcathepsinsasindicatorsofinvasive­nessofmeningiomas.Theysuggestthatmeningi­omasrecurnotonlyduetoahighercellprolifera­tionmarkedbytheKi67indexbutalsoduetoa morepowerfulinvasivenessofmeningiomacells, markedbyhighercathepsinBandLexpression. Our study also confirms that higher expressions of cathepsins B and L correlate with a higher men­ Vranic A / Antigen expression on recurrent meningioma 111 ingioma grade, as already shown in previous stud­ies.26-28However, since invasiveness is also found in BM, the meningioma grade is probably not di­rectly correlated to expression of cathepsins B and L. Several cellular mechanisms of cellular prolif­eration, invasiveness and others are responsible for the malignant transformation of meningiomas. The expression of antigenic markers,the Ki67 in­dex and cathepsins B and L seem to correlate with a tendency of meningioma to recur. Measuring the expression of these three antigens on meningioma cells could have prognostic value already at the first appearance of a meningioma. These antigenic markers could be proposed as prognostic indica­tors of the recurrence-free survival of patients with meningiomas. A larger multivariate study on a larger population is needed to confirm the prog­nostic value of MIB-1 antigen, PR, cathepsin B and cathepsin L on meningioma cells. A higher probability of an individual meningi­oma to recur would mean an alarm to start a more aggressive therapeutic approach for the patient. This would include more frequent control check­ups, more frequent control MRI scans and possi­bly immediate postoperative irradiation with the proper radiation delivery.33A more frequent fol­low-up is particularly important since the median time to recurrence in meningiomas is rather long (4.5 years), which gives us enough time for thera­peutic intervention. Conclusions The recurrence rate of meningiomas, especially of AM and MM is quite high. So far, the most impor­tant known prognostic factors for the meningioma recurrence have been meningioma grade, meningi­oma invasiveness and completeness of neurosurgi-cal resection. Since the WHO grading system alone does not always correctly predict the biological be-haviour of meningiomas, antigenic markers with prognostic significance are being sought. The MIB­1 antigen, cathepsin B and cathepsin L are shown to be expressed more on cells of recurrent meningi­omas compared to non-recurrent ones. Expression of these antigens could possibly help us to assess the risk of meningioma recurrence with each indi­vidual meningioma patient. References 1. Ayerbe J, Lobato RD, Cruz J, Cabrera A. Risk factors predicting recurrence in patients operated on for intracranial meningioma. A multivariate analysis. Acta Neurochir 1999; 141: 921-32. 2. Black P. 643 meningiomas: current perspective. Neurosurgery 2003; 32: 643-57. 3. De Monte F, Marmor E, Al-Mefty O. Meningiomas. In: Kaye AH, editor. Brain tumours. London: Harcourt Publishers; 2001. p. 719-50. 4. Jäaskelainen J. 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Survivin in brain tumours: an attractive target for immunotherapy. J Neurooncol 2003; 64: 71-6. 16. LoussouarnD,BrunonJ,Avet-LoiseauH,CamponeM,MosnierJF.Prognostic value of HER2 expression in meningiomas: an immunohistochemical and fluorescence in situ hybridization study. Hum Pathol 2006; 37: 415-21. 17. Puduvalli VK, Li JT, Chen L, McCutcheon IE. Induction of apoptosis in primary meningioma cultures by fenretinide. Cancer Res 2005; 65: 1547-53. 18. Okada M, Miyake K, Matsumoto Y, Kawai N, Kunishio K, Nagao S. Matrix metalloproteinase-2 and matrix metalloproteinase-9 expressions correlate with the recurrence of intracranial meningiomas. J Neurooncol 2004; 66: 29-37. 19. Perret AG, Duthel R, Fotso MJ, Brunon J. Stromelysin-3 is expressed by ag­gressive meningiomas. Cancer 2002; 94: 765-72. 20. Mikkelsen T, Yan PS, Ho KL, Sameni M. Immunolocalization of cathepsin B in human glioma: implications for tumor invasiveness and angiogenesis. J Neurosurg 1995; 83: 285-90. 21. Sivaparvathi M, Sawaya R, Wang SW, Rayford A, Rao JS. Overexpression and localization of cathepsin B during the progression of human gliomas. Clin Expl Metastasis 1995; 13: 49-56. 22. Sivaparvathi M, Yamamoto M, Nicolson GL, Rao JS. Expression and immu­nohistochemical localization of cathepsin Lduring the progression of human gliomas. Clin Exp Metastasis 1996; 14: 27-34. 112 Vranic A / Antigen expression on recurrent meningioma 23. Strojnik T, Kos J, Židanik B, Lah TT. Cathepsin B immunohistochemical stain­ing in tumor and endothelial cells is new prognostic factor in patients with brain tumors. Clin Cancer Res 1999; 5: 559-67. 24. Strojan P. Cysteine cathepsins and stefins in head and neck cancer: an upda­te of clinical studies. Radiol Oncol 2008; 42: 69-81. 25. Castilla EA, Prayson RA, Abramovich CM, Cohen ML. Immunohistochemical expression of cathepsin D in meningiomas. Am J Clinl Pathol 2003; 119: 123-8. 26. Trinkaus M, Vranic A, Dolenc VV, Lah TT. Cathepsins B and L and their inhibi­tors stefin B and cystatin C as markers for malignant progression of benign meningiomas. Int J Biol Markers 2005; 20: 50-9. 27. Trinkaus M, Vranic A, Dolenc VV, Lah TT. Cathepsin L in human meningio-mas. Radiol Oncol 2003; 37: 89-99. 28. Strojnik T, Zidanik B, Kos J, Lah TT. Cathepsins B and L are markers for clini­cally invasive types of meningiomas. Neurosurgery 2001; 48: 598-605. 29. Perry A, Scheithauer BW, Stafford SL, Lohse CM, Wollan PC. “Malignancy” in meningiomas: a clinicopathologic study of 116 patients, with grading implications. Cancer 1999; 85: 2046-56. 30. Modha A, Gutin PH: Diagnosis and treatment of atypical and malignant meningiomas: a review. Neurosurgery 2005; 57: 538-50. 31. Yang SY, Park CK, Park SH, Kim DG, Chung YS, Jung HW. Atypical and anaplas-tic meningiomas: prognostic implications of clinicopathological features. J Neurol Neurosurg Psychiatry 2008; 79: 574-80. 32. Nordqvist ACS, Smurawa H, Mathiesen T. 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Radiol Oncol 2008; 42: 170-2. 113 research article Frequent MGMT (06-methylguanine-DNA methyltransferase) hypermethylation in long-term survivors of glioblastoma: a single institution experience Martina Baur1,2, Matthias Preusser3,4, Maria Piribauer3, Katarzyna Elandt3, Marco Hassler3, Marcus Hudec5, Christian Dittrich1,2, Christine Marosi3 1 Applied Cancer Research-Institution for Translational Research Vienna (ACR-ITR VIEnna/CEADDP), Vienna, Austria 2 Ludwig Boltzmann Institute for Applied Cancer Research (LBI-ACR VIEnna)-Ludwig Boltzmann Cluster Translational Oncology, Kaiser Franz Josef-Spital, Vienna, Austria 3 Department of Internal Medicine I, Medical University Vienna, Vienna, Austria 4 Institute of Neurology, Medical University Vienna, Vienna, Austria 5 Department of Scientific Computing, University of Vienna, Vienna, Austria Received 18 March 2010 Accepted 6 April 2010 Correspondence to: Martina Baur, M.D., Assoc. Prof. of Medicine, Ludwig Boltzmann-Institute for Applied Cancer Research (LBI-ACR Vienna)­Ludwig Boltzmann Cluster Translational Oncology, 3rd Medical Department-Centre for Oncology and Haematology, Kaiser Franz Josef-Spital, Vienna, Austria. Phone: +43 1 60 1 91 2308; Fax: +43 1 60 1 91 2309; E-mail: martina.baur@gmx.net Disclosure: No potential conflicts of interest were disclosed. Background. The aim of this retrospective study was to analyse the MGMT (06-methylguanine-DNA methyltrans­ferase) promoter methylation status in long-term surviving (= 3 years) patients with glioblastoma multiforme (GBM). Methods. The methylation status of the MGMT promoter was determined by bisulfite modification of the DNA and subsequent methylation-specific polymerase-chain-reaction (MSP). DNA was extracted from routinely formalin-fixed and paraffin-embedded tumour tissue samples. Results. MSP yielded interpretable results in only 14 of 33 (42%) long-term surviving patients with GBM. A methylated band was seen in 3 of 14, methylated as well as unmethylated bands in 8 of 14 and an only unmethylated band in 3 of 14 patients, thus, yielding MGMT promoter methylation in 11 of 14 patients. The two groups of patients with methyl­ated and unmethylated MGMT promoter status were too small to draw any firm statistical conclusions. Conclusions. Long-term surviving patients with GBM have very frequently intratumoural MGMT promoter methyla­tion. This phenomenon discriminates long-term survivors from a non-selected group of patients with GBM. The stand­ardization of the MSP for the determination of the MGMT promoter methylation status seems to be necessary in order to make this methodology a more reliable one. Key words: glioblastoma multiforme (GBM); high grade glioma; MGMT promoter methylation; hypermethylation; long-term survival Introduction Glioblastomamultiforme(GBM)isthemostcom­monprimarybraintumourinadults.Itrepresents themostfrequentlyencounteredtypeofglialtu­moursandcanalsooccurinchildren.1,2Median survivalisgenerallyonlyslightlylongerthanone yearbasedonmultimodalapproachesconsisting ofmaximalfeasibleresection,radiotherapyand chemotherapy.Asubstantialstepforwardinthe treatmentofGBMwasreachedbytherandomized phaseIIItrialbyStuppetal.,demonstratingasig­nificantlylongersurvivalinpatientstreatedwith temozolomideinadditiontoradiotherapyfollowed byadjuvanttemozolomidewithamediansurvival of15monthsandafive-yearsurvivalrateof9.8%.3 Distinct from unselected GBM patients, who survive about one year, there is a small subgroup 114 Baur M et al. / MGMT Promoter methylation in glioblastoma of 1% - 5% of patients with GBM that survive at least 3 years after the diagnosis of GBM.4-12They are designated as long-term glioblastoma survi­vors. This period of 36 months survival was also adopted in our study as the lower limit for long­term surviving GBM patients; yet, there is no gen­erally accepted definition. All histologic diagnoses of the putative long-term surviving GBM patients have to be reviewed because in about one half of the cases the histologic diagnosis of GBM is re­classified to represent a less malignant tumour, namely oligodendroglioma, malignant mixed oli­godendroglioma-astrocytoma or anaplastic astro­cytoma.6-10,13Although the histologic aspect of the tumours from long-term survivors does not differ from that of classical survivors, it is postulated that the long-term surviving patients are a subgroup of GBM patients with a different biological behav­iour, a different therapeutic responsiveness and a distinct genetic characterization. Clinicalparameterssuchasyoungage,high Karnofskyperformancestatusandtheextentof radicalityofsurgeryareassociatedwithabetter prognosisdespitethehistologyofaGBM.4,6,8,10,14,15Scottetal.foundadditionalfactorsastheneuro-logicfunctionandthedoseofradiotherapyap­pliedintheirrecursivepartitioninganalysistobe importantprognosticvariables.16Theperiodof symptomsbeforethediagnosisinlong-termGBM survivorsincontrasttoaverageGBMpatientsis significantlylonger.10AsignificantlylowerKi­67-labelingindexcomparedtocontrolshasbeen describedintumoursfromlong-termsurvivors.10Suchpatientsexhibitfewergeneticaberrations thantypicalGBMpatients.7Likeinoligodenrog­liomapatientsthelossof19qisexclusivetothe long-termsurvivors.1Usually6qloss,10qloss and19qgainareassociatedwiththeshort-term survival7whereasmdm2overexpressionisless likelyexhibitedbythelong-termGBMsurvivors.5Molecularparameters,whichcandeterminatethe stepoftumourmalignancy17,arealsoimportantin GBMpatients.5Theoverexpressionoftheprotein p53andthenuclearp53expressionaresignificant­lymorefrequentlyfoundinlong-termsurviving patients.5Abettermolecularcharacterizationof long-term GBM patients is achieved by examining ofmultiplemarkerssuggestingthatdifferingpat­ternsofgeneticlesionsmaydiscriminatebetween thelongandtheshort-termsurvivalofGBMpa­tients.7 Ithasbecomeclearthatcancersingeneral arisefrombothgeneticandepigeneticchanges. Epigeneticchanges,suchashypermethylation, mayinactivategeneswithoutchangingthebase sequence.Analysingadifferentpromotermeth-ylationstatusofkeyregulatorgenesimplicated inapoptosisandinflammationhypermethylation ofTMS1/ASCwassignificantlymorefrequentin long-termsurvivingGBMpatientsandDAPK promoterhypermethylationwasonlyfoundinthe long-termsubsetcomparedtounselectedGBMpa­tients.4Martinezetal.18foundasignificantlyhigh­ermethylationrateofMGMTinlong-termGBM patientscomparedtounselectedGBMpatients. TheMGMTgeneislocatedonchromosome10q26. Methylationofthegenepromoterisassociated withthelossofMGMTexpressionwhichresults indiminishedDNA-repairactivity.Tumourcells lackingMGMTarepronetocelldeathinducedby alkylatingsubstancessuchastemozolomide.In thisprocessthealkyl-groupistransferredtothe activesiteoftheMGMTproteinthattherebybe­comesirreversiblyinactivatedandsubsequently degraded,requiringresynthesis.AlthoughO6­methylguanineaccountsforlessthan10%ofthe lesionsinducedbyalkylatingagents,itplaysama­jorroleasatriggerforcytotoxicityandapoptosis. Ifleftunrepaired,e.g.duetoepigeneticsilencing oftheMGMTgeneordepletiontheMGMTpro­teinbysaturationoftheprocess,O6-methylgua­ninepersistsintheDNA.19Recently,Hegietal.20, Glasetal.21andSonodaetal.22describedpromoter methylationofMGMTasanindependentfavour-ableprognosticfactor.PatientswithGBMcontain­ingamethylatedMGMTpromoterbenefitedfrom temozolomide,whereasthosewhodidnothavea methylatedMGMTpromoterdidnothavesucha benefit.20 To further characterize long-term glioblastoma patients genetically we investigate retrospectively the MGMT promoter methylation status by the bi-sulfite modification of the DNA and subsequent methylation-specific polymerase-chain-reaction (MSP) in formalin-fixed and paraffin-embedded tumour tissue samples of 33 long-term survivors with GBM from a single centre. Patients and methods Patient recruitment Primary and secondary GBM patients surviving longer than 36 months after the diagnosis were retrospectively identified in a single centre, the Department of Internal Medicine I, University of Vienna, Vienna, Austria starting from the year 1995 Baur M et al. / MGMT Promoter methylation in glioblastoma 115 up to 2003. The histologic diagnosis of GBM ac­cording to the World Health Organization (WHO) classification of the brain tumours was confirmed by the pathology review by M.P. All patients have been treated with alkylating agents. The clinical data were evaluated by checking patients` records, the presence and the extent of oedema by review­ing the radiologic films. A cognitive impairment was assessed by analysing the dialogues between the treating physicians and the patients; addition­ally, the functional capacities regarding ADL (ac­tivities of daily living) and IADL (instrumental ac­tivities of daily living) documented as reported by the relatives were scored. This study has been ap­proved by the local ethics committee and has been performed in accordance with the ethical standards laid down in the 1964 Declaration of Helsinki. The informed consent for samples and the data analysis from each patient had been obtained. MGMT promoter methylation analysis A MGMT promoter methylation status was ana­lysed using methylation-specific PCR (MSP) as de­scribed by Hegi et al.20In brief, genomic DNA was isolated from paraffin sections of GBM tissue us­ing Ex-Wax DNA Extraction (Chemicon, Temecula, California, USA). The DNA was subjected to bi-sulfite treatment at 56°C for 16-20h. Then the DNA was purified using Wizard DNA Clean-Up System A7280 (Promega, Madison, Wisconsin, USA). MSP was performed in a two-step “nested” approach using previously defined primer sets.20The PCR products were separated on two percent agarose gels. A glioblastoma case with a known methyl­ated MGMT promoter was used as the positive control and water was used as the negative control for MSP analysis. MIB-1 proliferation index immunohistochemistry Tumour sections (3-5 micrometers thick) were im­munostained with a monoclonal mouse anti-Ki-67 antibody (clone MIB-1, Dako, Glostrup, Denmark) at a dilution of 1:50 for 25 minutes. For the deter­mination of the MIB-1 proliferation index, the frac­tion of labelled nuclei per 500 tumour cell nuclei was manually counted using an eye grid and was expressed as percentage. Statistical analysis Time to progression reached from the date of the first neurosurgical procedure or diagnosis of gliob­lastoma to the time of the first objective evidence of tumour progression or the time of censoring. Survival time was defined as the time lapse from the initial surgery or diagnosis to the patient’s death or the time of censoring. Time to progression and survival were estimated using the Kaplan-Meier method. The influence on time to progres­sion and overall survival by sex, age, presence of primary or secondary glioblastoma, side and re­gion of the brain of the primary tumour, presence of cognitive impairment, presence of oedema and Karnofsky performance status was calculated by the log-rank test. For the analysis of the influence of the age of the formalin-fixed or paraffin embed­ded tumour tissue on the feasibility of determina­tion of the MGMT promoter methylation status a .˛-test was used. All statistical analyses were per­formed using the SPSS software 15.0. Results In this retrospective analysis 35 long-term surviv­ing patients with GBM were identified from one centre. 33 of them were confirmed GBM patients after the histologic review indicating a percentage of 6% of revised histologies. The two diagnoses of the reclassified histologies were anaplastic oli­godendroglioma and anaplastic astrocytoma. As 40 patients per year with primary GBM are treated in the institution this results in about four patients per year becoming long-term surviving GBM pa­tients. This corresponds to an estimated percentage of 10% of long-term survivors in the institution. The median follow-up was 54.2 ±SD 26.1 months. The patient’s characteristics are shown in Table 1. Karnofsky performance score The postoperative Karnofsky performance score of the long-term surviving patients was at least 80%. At the time of writing two women and three men were professionally still active, the two women as computer clerks with full time employment, one of the men as a teacher for mathematics in a vo­cational school, one as a farmer and the third as a pizza cook. 116 Baur M et al. / MGMT Promoter methylation in glioblastoma TABLE 1. Patient characteristics Number of patients Age (years) median (range) Sex Male Female Performance status acc. to Karnofsky (%) 60 70 80 90 100 Oedema = 1 cm > 1 cm n.e. History of glioblastoma Primary glioblastoma Secondary glioblastoma Localisation of the tumour Frontal Parietal Trigonal Temporal Occipital Frontoparietal Insula Parietooccipital Thalamus Side of tumour localisation Right Left Bilateral Extent of resection Biopsy Subtotal Total Cognitive impairment Yes No Stroke Yes No Initial chemotherapy Temozolomide CCNU Fotemustine/Dacarbacine N = numbers, n.e. = not evaluable 33 38 (22-66) 22 (66.7) 11 (33.3) 1 (3.1) 3 (9.1) 11 (33.3) 14 (42.4) 4 (12.1) 11 (33.3) 19 (57.6) 3 (9.1) 31 (93.9) 2 (6.1) 12 (36.4) 4 (12.1) 2 (6.1) 8 (24.2) 1 (3.0) 1 (3.0) 1 (3.0) 2 (6.1) 2 (6.1) 10 (30.3) 22 (66.7) 1 (3) 4 (12.1) 10 (30.3) 19 (57.6) 11 (33.3) 22 (66.7) 2 (6.1) 31 (93.9) 6/33 (18.2%) 15/33 (45.5%) 12/33 (36.4%) Local relapses 20 of 33(61%) patients suffered a local relapse, elev­en of them after gross total resection of the primary tumour. Median time to progression Themediantimetoprogression(TTP)was39 months[95%CI:0;105.6](Figure1).Patientswith abiopsyatinitialdiagnosishadamedianTTPof 5.3months[95%CI:0;17.8months](n=4),patients withasubtotalresectionhadaTTPof39.3months [95%CI:0-92months](n=10)andpatientswitha totalresectiononeof66.9months[95%CI:0;145.3 months](n=19),respectively.Theclinicalparam­etersage,sex,oedema,sideandregionofthebrain oftheprimarytumourandKarnofskyperformance statusdidnotimpactontheTTP.Patientswiththe unmethylatedMGMTpromoterhadatimetopro­gressionof7,39and79+monthswhereaspatients withmethylatedMGMTpromoterhadaTTPof5 to56+months. Survival At the time of evaluation, 15 patients were alive, seven of them without tumour recurrence for up to 151+ months. The median survival was 83 months [95% CI; 43.8-122.3] (Figure 2). Patients with a sub­total resection survived 47.2 months [95% CI: 28.8­ 65.6 months] and patients with a total resection 83.0 Baur M et al. / MGMT Promoter methylation in glioblastoma 117 TABLE 2. MGMT promoter determination months [95% CI: 43.2; 122.9 months], respectively. The clinical parameters age, sex, oedema, side and region of the brain of the primary tumour and Karnofsky performance status did not impact on the survival. Of note, nine of the patients with a lo­cal relapse survived longer than five years. Patients with the unmethylated MGMT promoter had a survival of 43, 79+ and 97+ months, respectively. Patients with the methylated MGMT promoter had a median survival of 48 ± SD 0.97 months. MIB-1 scoring Insevenpatientstheimmunohistochemicalstaining ofMIB-1wasdetermined.ThemeanMIB-1score was29.1%,themedian30.3%(range12.1-49%). MGMT promoter methylation status (Table 2) Only in 14 of 33 (42%) patients the determination of the MGMT promoter methylation status by MSP yielded interpretable results. There was no linear correlation of the success rate to the age of the par­affin block (p=0.5). Of 14 patients with interpretable MSP results, three of 14 patients had a methylated MGMT promoter, three of 14 patients an unmeth­ylated MGMT promoter and 8 of 14 patients partly a methylated and partly an unmethylated MGMT promoter. Thus, the MGMT promoter methyla­tion was found in 11 of the 14 patients. These two groups of patients with the methylated and the un-methylated MGMT promoter status, respectively were too small to draw reliable conclusions based on statistical testing. Feasible 14 (42) Methylated 3 (21.4) Unmethylated 3 (21.4) Methylated and unmethylated 8 (57.2) Not feasible 19 (58) Discussion Reports on the MGMT promoter methylation sta­tus in long-term surviving patients with glioblas­toma multiforme are scarce. 78.5% of long-term survivors presented with MGMT promoter hyper-methylation. This is in the same range as reported by Martinezet al.,Sonoda et al.and Krex et al.18,22.23This high proportion of patients with the MGMT promoter methylation is in clear contrast the 44% (range 25- 68%) determined from 13 different studies of unselected patients with GBM.18,20,24,34However, the high rate of methylated tumours in the long-term surviving patients let suggest that MGMT promoter methylation is of paramount im­portance for response to the actual standard ther­apy with alkylating agents in GBM.3The proof of this principle is eagerly awaited in form of the re­sults of the prospective Intergroup trial RTOG0525/EORTC26052 which will not yet be presented for the first time at ASCO 2010 testing dose-intense temozolomide in comparison to standard-dose te­mozolomide dependent on the MGMT promoter methylation status in GBM patients. Although 33 patients were initially included in our analysis of long-term surviving patients with GBM, the paraffin embedded tissue blocks of only 14 out of 33 (42%) patients were suitable for the MGMT hypermethylation test by MSP. Because of the small number of patients it was impossible to determine whether the MGMT methylation status was of prognostic impact in our patient cohort. The MGMT promoter methylation status as a prog­nostic factor in long-term surviving GBM patients should be further evaluated in prospective studies. The statistical analysis did not show a signifi­cant difference between older paraffin embed­ded tissue in contrast to younger patient samples 118 Baur M et al. / MGMT Promoter methylation in glioblastoma (p=0.5); however, only higher patient numbers in the subgroups could provide reliable significant results. The success rate of the methylation specific PCR determination on paraffin-embedded tumour samples is highly variable and centre dependent.20Hegi et al. reported on a median success rate of 75% (range 0-100%)20, Brandes et al.of 66%.29In contrast to these results Aldape et al.found prospectively a success rate of 91% in 995 patients with GBM.35 Intheliteratureseveralreasonsforthelowsuc­cessrateofMSPtestinginparaffin-embeddedtu­mourtissueofpatientswithGBMcomparedto freshfrozentissuearediscussed.Frequentlyonly averysmallamountofpartiallydegradedDNAis recoveredduetoextensivenecrosisandscarcityof malignantcells.Herrlingeretal.observedthat17% ofthetumourspecimensdidnotcontainenough DNA.33Especiallyintumourbiopsiestumourcells arenoteasilyfound.24Hauetal.recommendeda goodqualityparaffinembeddedtissuethatisnot overfixed.19Theaccumulationofnormalcellsin thetumour,includinginfiltratinglymphocytes, maycomplicateaccurateassessmentofMGMT.28Thebestresultswithmethylation-specificPCRare obtainedwithcryopreservedtumourspecimens, thusavoidingthefixation-relateddeteriorationof thequalityofDNA.368of14ourpatientsexhibited boththemethylatedandtheunmethylatedMGMT promoter.Ourobservationisinthesamerange asreportsbyMartinezetal.,Blancetal.,Criniere etal.,Cankovicetal.andGonzalez-Gomezetal.whofoundthatthemajorityofthemethylatedtu­moursalsoexhibitedanunmethylatedband,which mayarisefromeithernormalcellswithinthetu­moursampleorfromatumourcellsidepopula­tion.18,26,31,34,37OurexperienceshowsthatMGMT promotermethylationtestingmaybetechnically challenging.Severalmethodsincludingmultiplex ligationprobeamplificationMLPA,realtimequan­titativepolymerasechainreaction(quantitative rt-PCR),havebeenproposedaspotentialalterna­tivestoconventionalMSP.Thesemethodsneedto becriticallyevaluatedinfuturestudiesandreliable cut-offvaluesfortheprognosticationandthepre­dictionhavetobeprospectivelyvalidated.38,39 Two of 33 (6%) patients of our study suffered from secondary GBM. This percentage was mark­edly lower than the incidence of 20% reported by Steinbach et al..6In these two patients with second­ary GBM in our study the MGMT promoter meth-ylation determination was not feasible due to tech­nical reasons. The determination of the proliferative activity in form of MIB-1 evolved a low median score of 30.3 (range 12.1-49). This observation correlated well with the results reported by Ho et al., demonstrat­ing a cut-off value of = 35 being related to worse outcome in unselected GBM patients.40However, due to sampling differences, there has no clear prognostic impact of Ki-67 on the survival of GBM patients been detected.10,22,41 In addition to the MGMT methylation status, we compared clinical parameters of our long-term GBM patients like age, Karnofsky-performance score, ratio male/female, localisation of the primary tumour, extent of surgery, laterality of the primary tumour, incidence of cognitive impairment and of ischemic events, incidence of relapses, median survival, to those of other reports of long-term sur­vivors with GBM. Most authors included patients surviving = 3 years, Vertosicket al. those > 4 years, McLendonet al.,Steinbach et al. and Salvatiet al. patients surviving = 5 years and Morita et al. those = 7 years.6,8,11-13 The median age at diagnosis of the patients of this series was 38 years, which is only slightly younger than the median of 41 years (range 37-51 years) observed in 10 different studies5,6,8,10-13,18,23,42and 12 years younger than the average age at diag­nosis of unselected GBM patients (median 53 years ± 0.55 years).10 One of the most important prognostic factors in cancer patients43, the Karnofsky performance score at the beginning of radiochemotherapy in our 33 patients reached median 90%. This equals the median of 90% (range 80-90%) observed in nine other studies of long-term surviving GBM pa­tients5-7,10-12,18,23,42and is clearly higher than the me­dian of 76.1% of unselected GBM.10The sex ratio male:female in our patient cohort was 67:33, quite similar to other studies. There was no predilection in laterality or in localization in a given cerebral lobe, as in the other series of long-term surviving GBM patients.6,13,23 The radiological parameter “extent of oedema > 1 cm” at diagnosis of GBM was present in 58% of our patients and did not impact on time to pro­gression or survival in this series. In other studies of long-term surviving patients with GBM oede-ma was not investigated as a prognostic factor. However, in average patients with GBM, oedema larger than 1 cm has been reported to influence the survival, negatively.44,45 Of note, a gross total resection was achieved in 58% of the patients of this series. This has been re­ported accordingly in the series by Scott et al. with 40%, Salvati et al. with 46% and Hottinger et al. with 48% but not by Mc Lendon et al. with 27%.8,10,12,42 Baur M et al. / MGMT Promoter methylation in glioblastoma 119 Obtaining a total gross resection appears to be of paramount importance for achieving a long-term survival in GBM patients. A considerably lower percentage of gross total resections of about 40% were recorded in studies of unselected patients.3However, in this series four patients (12%) under­went only a biopsy of the primary tumour. In three of these four patients the MGMT status was not evaluable and in the remaining patient the MGMT promoter was unmethylated. This raises hope that even patients without tumour debulking and with unmethylated MGMT promoter status can eventu­ally achieve a long-term survival. 61% of our patients relapsed locally. This was in line with three other reports of long-term surviv­ing GBM patients specifying a percentage ranging from 45-73%.10,12,42Of note, we did not observe dis­tant relapses in the cohort of long-term surviving patients. In 33% of our patients a cognitive impairment was recorded. A similar rate of 28% has been re­ported by Hottinger et al.42 Two of 33 (6%) of our patients suffered from an ischemic event, this was nearly identical to the 10% observed by Steinbach et al. but clearly lower than the 23% found by Hottinger et al..6,42Further tri­als will have to evaluate the incidence of ischemic events in long-term surviving patients with GBM, to identify risk factors and establish preventive strategies. In summary, this series of patients achieving a long-term survival after the diagnosis of GBM il­lustrates the validity of the prognostic factors de­veloped in the nomogram by Gorlia et al.15on the patients of EORTC and NCIC trials as well as of other series with long-term surviving patients with GBM: young age, extensive tumour resection, fa-vourable performance status and treatment ac­cording to the standard of care, as well as a high percentage of glioblastomas with MGMT promoter methylation. The definitive role of MGMT promot­er methylation in directing tailored chemotherapy in GBM patients will be elucidated in the large ran­domized international intergroup trial RTOG0525/EORTC26052 stratifying GBM patients by MGMT methylation status and randomizing for standard temozolomide in contrast to dose-dense temozolo­mide therapy. MGMT promoter methylation test­ing represents a substantial step forward in the treatment of patients with glioblastoma multiforme and enables us to better understand the mode of action of alkylating therapies and the course of the disease. Further, new treatment options exploiting the MGMT promoter methylation mechanism may add to the improvements achieved in this disease. Acknowledgment We would like to acknowledge the cooperation with Johannes Hainfellner, M.D., Institute of Neurology, Medical University of Vienna, Vienna, Austria in performing this study. References 1. Velnar T, Smrdel U, Popovic M, Bunc G. Genetic markers in oligodendroglial tumours. Radiol Oncol 2010; 44: 13-18. 2. Kachanov DY, 2, Dobrenkov KV, Shamanskaya TV, 2, Abdullaev RT, 2, Inushkina EV, Savkova RF, et al. Solid tumors in young children in Moscow Region of Russian Federation. Radiol Oncol 2008; 42: 39-44. 3. Stupp R, Hegi ME, Mason WP, van den Bent MJ, Taphoorn MJB, Janzer RC, et al. 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Sonoda Y, Kumabe T, Watanabe M, Nakazato Y, Inoue T, Kanamori M, et al. Long-term survivors of glioblastoma: clinical features and molecular analy­sis. Acta Neurochir (Wien) 2009; 151: 1349-58. 23. Krex D, Klink B, Hartmann C, von Deimling A, Pietsch T, Simon M, et al. Long­term survival with glioblastoma multiforme. Brain 2007; 130: 2596-606. 24. Balana C, Ramirez JL, Taron M, Roussos Y, Ariza A, Ballester R, et al. O6­methyl-guanine-DNA methyltransferase methylation in serum and tumor DNA predicts response to 1,3-bis(2-chloroethyl)-1-nitrosourea but not to temozolomide plus cisplatin in glioblastoma multiforme. Clin Cancer Res 2003; 9: 1461-8. 25. Kamiryo T, Tada K, Shiraishi S, Shinojima N, Kochi M, Ushio Y. Correlation between promoter hypermethylation of the O6-methylguanine-deoxyribonucleic acid methyltransferase gene and prognosis in patients with high-grade astrocytic tumors treated with surgery, radiotherapy, and 1-(4-amino-2-methyl-5-pyrimidinyl) methyl-3-(2-chloroethyl)-3-nitrosou­rea-based chemotherapy. Neurosurgery 2004; 54: 349-57. 26. Blanc JL, Wager M, Guilhot J, Kusy S, Bataille B, Chantereau T, et al. Correlation of clinical features and methylation status of MGMT gene pro­moter in glioblastomas. J Neurooncol 2004; 68: 275-83. 27. Weaver KD, Grossman SA, Herman JG. Methylated tumor-specific DNA as a plasma biomarker in patients with glioma. Cancer Invest 2006; 24: 35-40. 28. EstellerM,Garcia-FoncillasJ,AndionE,GoodmanSN,HidalgoOF,Vanaclocha V, et al. Inactivation of the DNA-repair gene MGMT and the clinical response of gliomas to alkylating agents. N Engl J Med 2000; 343: 1350-4. 29. Brandes AA, Tosoni A, Cavallo G, Bertorelle R, Gioia V, Franceschi E, et al. 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Phase II trial of lomustine plus temozolomide chemotherapy in addition to radiotherapy in newly diagnosed glioblastoma: UKT-03. J Clin Oncol 2006; 24: 4412-7. 34. Cankovic M, Mikkelsen T, Rosenblum ML, Zarbo RJ. A simplified laboratory validated assay for MGMT promoter hypermethylation analysis of glioma specimens from formalin-fixed paraffin-embedded tissue. Labor Invest 2007; 87: 392-7. 35. Aldape KD, Jones G, Wang M, Hegi M, Janzer RC, Stupp R, et al. MGMT methylation testing in RTOG 0525: A phase III trial of newly diagnosed gliob­lastoma [abstract]. J Clin Oncol 2009; 27(Suppl), 15S. No. 2051. 36. Dehdashti AR, Hegi ME, Regli L, Pica A, Stupp R. New trends in the medi­cal management of glioblastoma multiforme: the role of temozolomide chemotherapy. Neurosurg Focus 2006; 20: 1-6. 37. Gonzalez-Gomez P, Bello MJ, Arjona D, Lomas J, Alonso ME, De Campos JM, et al. Promoter hypermethylation of multiple genes in astrocytic gliomas. Int J Oncol 2003; 22: 601-8. 38. 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GojkovicHorvatA,KovacV,StrojanP.Radiotherapyinpalliativetreatmentof painful bone metastases. Radiol Oncol 2009; 43: 213-24. 44. Pope WB, Sayre J, Perlina A, Villablanca JP, Mischel PS, Cloughesy TF. MR imaging correlates of survival in patients with high-grade gliomas. Am J Neuroradiol 2005; 26: 2466-74. 45. Hammoud MA, Sawaya R, Shi W, Thall PF, Leeds NE. Prognostic significance of preoperative MRI scans in glioblastoma multiforme. J Neurooncol 1996; 27: 65-73. 121 case report A clinical case of the penile metastasis from the rectal carcinoma Mehmet Yildirim, Ali Coskun, Mete Pürten, Ozgur Oztekin, Enver Ilhan Department of Surgery, Department of Radiology, Izmir Bozyaka Teaching and Research Hospital, Izmir, Turkey Received 23 November 2009 Accepted 10 December 2009 Correspondence to: Mehmet Yildirim, MD, Atakent Mah. Bergama 2 Apt. 32/1, Bostanli, Izmir, Turkey. Phone: + 90 232 3625692; Fax: + 90 232 2614444; E-mail: mehmetyildi@gmail.com Disclosure: No potential conflicts of interest were disclosed. Background. Penile metastases are rare and usually secondary to genitourinary and colorectal cancer. Case report. We present a case of a 77-year-old man with penile metastasis who was operated for rectal carcino­ma. He was referred to our clinic for penile ulcerous lesion, semierectile penis and voiding dysfunction. Imaging studies showed nodular lesion at glans penis and multiple bone metastases. He did not respond to chemoradiotherapy and he had bad prognosis. Conclusions. Imaging methods and biopsy may help to clarify the diagnosis but the treatment modalities are insuf­ficient in these patients. Key words: penis; metastasis; rectum; carcinoma Introduction The metastatic involvement of the penis is ex­tremely rare, despite rich vascularisation between the penis and the neighbouring organs. The first re­port of secondary penile malignancy from an ade­nocarcinoma of the rectum was defined by Eberth.1 The commonest sites of primary malignancy are genito-urinary organs, rectum and recto-sigmoid areas.2In literature less than 300 penile metastas­es were reported; 50 of them are originated from colorectal carcinoma.3Penile metastases lead to semi-erectile penis/priapism and skin lesions.2 Like in colorectal lesions MRI is the most useful method for the accurate diagnosis in suspected penile me­tastases.4,5An open biopsy is definitive for a the accurate diagnosis. A high rate of suspicion is re­quired to detect them. In this article, we report a case of a penile me­tastasis secondary to a rectal carcinoma three years after the surgery. Case report A 78-year-old male presented in March 2002 with a 2 months history of change in bowel habit and rec­tal bleeding. The examination revealed a tumour of the lower rectum. An abdomino-perineal excision was performed and the pathological examination revealed a Dukes C undifferentiated adenocarcino-ma with the lymph nodes involvement (15/22). He had received adjuvant chemo-radiotherapy post­operatively. He remained well until 2 years later, when he presented with urogenital complaints: pe­nile ulcerous lesion, semierectile penis and voiding dysfunction. 122 Yildirim M et al. / Penile metastasis from the rectal carcinoma A follow-up examination was performed in April 2005 following the development of indura­tions, hyperaemia, and oedema of the penis and demonstrated 0.5 cm and 0.3 cm ulcerative lesions on glans penis (Figure 1). The examination of the superficial inguinal lymph nodes and the abdomi­nal examination showed no remarkable signs. There was also no history of trauma or infectious disease. A complete blood cell count showed an in­creased white blood cell count of 16 x 103 / µL (nor­mal range: 5-10.0 x 103/µL), anaemia (with a hae­moglobin of 10.2 g/dL), normal level of PSA but increased levels of CEA (66.6 ng/ml, normal value <3 ng/ml) and CA19.9 (67.5 U/ml, normal value <37 U/ml). Penile US was normal for corpus spongi­osumand cavernosus. CT demonstrated metastases of several thoracic vertebras and sacroiliac joint. Furthermore, MRI of the penis showed multiple pathological focuses in T2 signal and a nodular le­sion that measured 15 mm in size, occupying the glans penis (Figures 2,3).The patient underwent excisional biopsy of the glans penis and inguinal lymph node, which revealed metastatic adenocar­cinoma consistent with his rectal carcinoma. The patient received chemotherapy for metastatic dis­ease until disease progression and unacceptable toxicity. The order of drug administration each week was irinotecan (80 mg/m2i.v. day 1) followed by leucovorin (500 mg/m2i.v day 1) and 5-fluor­ouracil (2300 mg/m2continuous infusion day 1). Two days after chemotherapy administration of the third cycle the patient was admitted to our de­partment with fatigue, weakness, dysuria and hy­potension. Increased level of BUN, creatinin and decreased level of blood count were registered. Despite the whole blood transfusion, the values of blood count decreased (down to 5.4 g/L ) during hospitalization. He was admitted for further medi­cal therapy but during this admission his general condition deteriorated and he died with progres­sive disease. Overall it appeared that this patient had a dis­seminated metastatic rectal carcinoma, with an unusual settled for the penis. Discussion The penile metastasis in the course of rectal cancer is an unexpected complication, despite its proxim­ity to the rectum and its rich vascularity. Recently published studies showed that the penile metas­tases are associated with prostate, urinary bladder cancer, and infrequently with rectal carcinoma.6There have also been some reports regarding dis­seminated metastases from the oeasophagus, pancreas and stomach carcinoma. There are a few pathways of metastasis to penis; frequently, retro­grade venous route, embolism to arterial system, retrograde lymphatic spread into the penile lym­phatic channels, direct extension, and operative manuplations. The retrograde venous transporta­tion suggested the main pathways to penis metas­tasis.1 The most common symptoms and signs are dys­uria, voiding dysfunction, perineal pain, priapism, penile nodules and mass.6In our case we found voiding dysfunction, semi-erectile penis and ul­ Yildirim M et al. / Penile metastasis from the rectal carcinoma 123 cerative lesions of the glans penis. Perineal pain, which it was a feature in one-third of patients in the series5, was not found in our case. We found a three-year interval between the primary tumour and penile metastasis. However, there are reports of the metastatic involvement years after the rec­tum carcinoma, but, penile metastases were usu­ally reported on the average of 13 months after the primary carcinoma.1 The lung, vertebra and liver metastasis can be associated with the penile me­tastasis in patients with rectal carcinoma2 which, in our case, we found vertebral metastases at presen­tation. This might reflect the fact that penile metas­tases reflect the disseminated disease. We believe that it would be beneficial in the dif­ferential diagnosis to take into consideration the findings related to voiding dysfunction, priapism and ulcerous lesions of the penis. Peyronie’s dis­ease, traumatic and infectious skin and syphilitic lesions or primary carcinoma of penis, must also be in the differential diagnosis of the penile metas­tasis.2,7 Tissue tumour markers: AFP, CEA, HCG can be used for differential diagnosis. Penile US and CT is the first choice for imaging but its sensitivity is lim­ited. Cavernosogram was reported a preferable ap­plication in some patients.8MRI reported as a very useful method even at the beginning of the symp­toms.5MRI demonstrates multiple metastatic nod­ules, with a low signal intensity and is intense with the surrounding corpus cavernosumon T1-weighted images, and low signal intensity against the high background intensity of the cavernous bodies on T2-weighted imaging.9Biopsy of the penile lesions is a favoured diagnostic modality to confirm the diagnosis as outlined in our case report. Regardless of the length to metastasis and difference in the treatment of the metastatic focus, the metastasis of the penis reflects a widely disseminated disease and poor prognosis. Treatment options are palliative with local surgery, systemic chemotherapy and local radio­therapy.4,7Surgery (panectomy) remains the only treatment with a long survival but mostly a pa­tient dies in a year. Radiotherapy has an average survival of 8 months, whereas chemotherapy has not been studied for metastases from the rectum carcinoma. In our case, metastases did not respond to palliative therapy and showed progression of the local disease. Despite the studies showed that rectal primaries had longer surviving comparing to genitor-urinary primary, we did not find any forms of result.7,8,10 In conclusion, penile metastases are rarely seen as a form of metastasis among rectal cancer pa­tients. Imaging methods may help to clarify the diagnosis, but penile lesions should be biopsied to confirm the diagnosis. On the other hand, the treat­ment modalities are insufficient for a long survey and quality of life in these patients. references 1. Haddad FS, Manne RK. Involvement of the penis by rectocolic adenocar­cinoma. Report of a case and review of the literature. Dis Colon Rectum 1987; 30: 123-9. 2. Abeshouse BS, Abeshouse GA. Metastatic tumours of the penis: a review of the literature and a report of two cases. J Urol 1961; 86: 99-112. 3. Cherian J, Rajan S, Thwaini A, Elmasry Y, Shah T, Puri R. Secondary penile tumours revisited. [Abstract]. Int Semin Surg Oncol 2006; 3: 33. 4. Sofic A, Šehovic N, Bešlic Š, Prnjavorac B, Bilalovic N, Caluk J, et al. MR rectum imaging with ultra sound gel as instrumental contrast media in tubulovillous adenoma. Radiol Oncol 2009; 42: 136-42. 5. Andresen R, Wegner HE, Dieberg S. Penile metastasis of sigmoid carci­noma: comparative analysis of different imaging modalities. Br J Urol 1997; 79: 477-8. 6. Appu S, Lawrentschuk N, Russell JM, Bright NF. Metachronous metastasis to the penis from carcinoma of the rectum. Int J Urol 2006; 13: 659-61. 7. Kragelj B. Irradiation of regionally advanced carcinoma of the penis. Radiol Oncol 2009; 43: 41-6. 8. Escribano G, Allona A, Burgos FJ, Garcia R, Navio S, Escudero A. Cavernosography in diagnosis of metastatic tumours of penis: 5 new cases and a review of the literature. J Urol 1987; 138: 1174-7. 9. Lau TN, Wakeley CJ, Goddard P. Magnetic resonance imaging of penile metastais; a report on five cases. Australasian Radiology 1999; 43: 378-81. 10. Ketata S, Boulaire JL, Soulimane B, Bargain A. Metachronous metastasis to the penis from a rectal adenocarcinoma. Clin Colorectal Cancer 2007; 6: 657-9. 124 research article A fully electronic intensity-modulated radiation therapy quality assurance (IMRT QA) process implemented in a network comprised of independent treatment planning, record and verify, and delivery systems Daniel W Bailey1, Lalith Kumaraswamy2, Matthew B Podgorsak2 1 Department of Physics, State University of New York at Buffalo, Buffalo NY, USA 2 Department of Radiation Medicine, Roswell Park Cancer Institute, Buffalo NY, USA Received 28 January 2010 Accepted 16 February 2010 Correspondence to: Prof. Matthew B Podgorsak, Department of Radiation Medicine, Roswell Park Cancer Institute, Buffalo NY 14263, USA. E-mail: Matthew.Podgorsak@RoswellPark.org Disclosure: No potential conflicts of interest were disclosed. Background. The purpose of this study is to implement an electronic method to perform and analyze intensity-mod­ulated radiation therapy quality assurance (IMRT QA) using an aSi megavoltage electronic portal imaging device in a network comprised of independent treatment planning, record and verify (R&V), and delivery systems. Methods. A verification plan was generated in the treatment planning system using the actual treatment plan of a patient. After exporting the treatment fields to the R&V system, the fields were delivered in QA mode with the aSi imager deployed. The resulting dosimetric images are automatically stored in a DICOM-RT format in the delivery sys­tem treatment console computer. The relative dose density images are subsequently pushed to the R&V system. The absolute dose images are then transferred electronically from the treatment console computer to the treatment plan­ning system and imported into the verification plan in the dosimetry work space for further analysis. Screen shots of the gamma evaluation and isodose comparison are imported into the R&V system as an electronic file (e.g. PDF) to be reviewed prior to initiation of patient treatment. A relative dose image predicted by the treatment planning system can also be sent to the R&V system to be compared with the relative dose density image measured with the aSi imager. Results. Our department does not have integrated planning, R&V, and delivery systems. In spite of this, we are able to fully implement a paperless and filmless IMRT QA process, allowing subsequent analysis and approval to be more efficient, while the QA document is directly attached to its specific patient chart in the R&V system in electronic form. The calculated and measured relative dose images can be compared electronically within the R&V system to ana­lyze the density differences and ensure proper dose delivery to patients. Conclusions. In the absence of an integrated planning, verifying, and delivery system, we have shown that it is nevertheless possible to develop a completely electronic IMRT QA process. Key words: EPID; IMRT; QA; paperless; portal dosimetry; PACS: 87.53.Bn, 87.53.Kn Introduction Intensity-modulated radiation therapy (IMRT) involves complex treatment plans that are com­pletely patient specific in order to highly conform delivered dose to the treatment volume, thus im­proving normal tissue sparing as compared to more traditional radiotherapy techniques.1,2As a consequence, the complexity and uniqueness of these treatment plans demand patient-specific pretreatment quality assurance (QA) of all IMRT treatments. Standard methods of IMRT QA involve ionization chambers, diode arrays and radiograph­ic films, often used in some combination to pro­vide verification of absolute dose, field geometry, number of monitor units, etc. However, these tra­ Bailey DW et al. / Paperless IMRT QA 125 ditional QA methods carry some distinct disadvan­tages, especially in the clinic that delivers a large number of IMRT treatments. These methods can be exceedingly time and resource demanding, requir­ing calibration and constancy checks of ionization chambers, set-up and calibration of diode arrays, calibration of films and expensive processing (un­less self-developing dosimetry film is chosen as a more convenient yet still expensive alternative). Furthermore, for all of the QA methods listed above, the QA analysis report may not be read­ily available in electronic form, demanding direct attachment to the patient paper chart (or manual scanning into the patient electronic chart, a proc­ess that is still not paperless). The disadvantages of paper charts are well known and well docu­mented3- including illegible signatures, notes and prescriptions; inaccessibility to multiple reviewers at one time; difficulties in locating charts and in­ability to access them remotely; etc. Meanwhile, the benefits of implementing an entirely paperless electronic medical record process have also been expounded in the literature.3-7A study published by the National Institute of Health and the Journal of the American Medical Association concluded that “EMRs will eventually become the standard of care,” citing that electronic patient charts provide complete, legible, and organized patient informa­tion in a format that is accessible at any time, even to multiple viewers in multiple (even remote) lo­cations.3Given the current shift toward adopting electronic medical records over paper charts, it is all the more important that the pretreatment IMRT QA process be fully electronic: no films, no print­ing and no scanning of QA reports, treatment plans and other documents. Inrecentyears,ithasbeendemonstratedthatan electronicportalimagingdevice(EPID),previously employedtoreplaceradiographicportalimagesfor patientalignment,caneffectivelybeusedforab­solutedosemeasurementandpretreatmentIMRT verification.8-12InEPIDIMRTQA,portaldosimet­ricimagesarecomparedtorespectiveportaldose predictionscreatedbyatreatmentplanningsystem (TPS)usinggeometricanddosimetrictools(such asdoseprofilesandgammaevaluation).13-15Thus, withaproperlycalibratedandcommissionedEPID, allqualitativeandquantitativedatanecessaryfor verificationofanIMRTfluenceisacquiredinasin­gleexposure,andallinformationisreadilyavail­ableinelectronicformallowingforthepossibility ofanentirelypaperlessIMRTQAprocess. A significant roadblock to the paperless EPID IMRT QA process is the common situation in which the treatment planning, record and verify (R&V), and radiotherapy delivery systems are not manufactured by the same vendor and thus com­munication between these systems is not entirely integrated. The purpose of this study is to imple­ment a fully electronic method to perform and analyze patient-specific IMRT QA using an EPID in a network comprised of independent treatment planning, R&V, and delivery systems. The advan­tages of such a QA process over standard methods of IMRT QA include: 1. Excellent efficiency, acquiring complete qualita­tive and quantitative information in a single ex­posure for each field, with no processing and no other calibration than the absolute and relative dose calibrations of the EPID (at intervals sug­gested by the vendor). 2. Excellent resolution compared to ionization chambers and diode arrays, with arrays as high as 1024x768 pixels with 0.392 mm pixel pitch (Varian PortalVision aS1000, Varian Medical Systems, Palo Alto CA). 3. Possibility of weekly QA by quick acquisition of EPID relative dose density images and compari­son within the R&V system to TPS predictions of those dose densities. 4. IMRTQA report electronically attached to the patient chart within the R&V system in a pa­perless processwith no manual attachment or tracking of QA reports, thereby decreasing the probability of errors (e.g. misplacement of QA document, etc.). Methods Wehavecommissionedanelectronicportaldosim­etrysystemconsistingofanamorphoussilicon (aSi) EPID (Varian PortalVision aS1000), coupled to aVarianTrilogylinearacceleratorwiththeVarian MilliniumMulti-LeafCollimator(MLC,120leaves). ThePortalVisionaS1000isa40x30cm2flat-panel, indirectdetectionEPIDwithamatrixof1024x768 pixelswith0.392mmpixelpitch.Forthisstudy,all EPIDimageswereacquiredattheminimumSSDof 105cmwithgantryandcollimatoratzerodegrees (unlessthecollimatorneededalternatepositioning toavoidregionsofhighbackscatterintheEPID).16TheEPIDwasfullycalibratedusingtheproce­duressuppliedbythevendor17,usingthefollow­ingintervals:thedarkfieldbackgroundcorrection andfloodfieldrelativedosecalibrationwereboth performedweekly;whiletheabsolutedosecalibra­tionwasperformedeachdaythattheEPIDwas 126 Bailey DW et al. / Paperless IMRT QA inuseforIMRTQA(alsoemployingthediagonal doseprofilecorrectionsuggestedbyBaileyetal.16). Thebeamsymmetry,energyandoutputwereveri­fiedeachweek.TheTPSemployedforthisstudy isVarianEclipse(Version8.6,includingPortal DosimetryVersion8.2.24),andtheR&Vsystemis thevendor-independentImpacMosaiq(Version 1.6,ElektaOncologySystems,NorcrossGA). From TPS to the R&V system Our electronic QA process begins with a patient-specific radiotherapy treatment plan created in the TPS using inverse-planning IMRT techniques based upon the patient’s 3-D computed-tomogra­phy (CT) data and the dose criteria predefined by the radiation oncologist. Each specific treatment field within this plan contains 320 control points that dictate the dynamic motion of the MLC leaves. Firstly in this process, the TPS uses the input geo­metric and dosimetric criteria to calculate an ideal fluence matrix referred to as the optimal fluence. Secondly, the optimal fluence is sent to the Leaf Motion Calculator which incorporates various me­chanical and geometric aspects of the delivery sys­tem (e.g.MLC beam transmission, minimum leaf gap, maximum leaf speed, MLC position deviation tolerance, etc.) to calculate leaf trajectories for the fluence that the system can capably deliver, known as the actual fluence.18Routine IMRT QA is partly designed to check the accuracy of these beam mod­els and parameters. If the necessary LINAC colli­mator jaw settings are beyond a certain separation (approximately 15 cm), the TPS splits the treatment field into multiple overlapping carriages, maintain­ing maximum degrees of freedom in MLC position and motion. After the treatment plan is completed and approved, the plan is electronically exported to the R&V system as a DICOM-RT file which in­cludes all necessary patient information and deliv­ery information, such as number of monitor units (MU), dose rate, collimator settings, and dynamic MLC positions. From R&V system to LINAC delivery The R&V system communicates the delivery pa­rameters from the TPS to the delivery system (and allows for automatic field setup), and further pro­vides an electronic medical record (EMR) which tracks the fractions and doses that have been de­livered to the patient, the delivery system settings for each field and fraction delivered, portal images and IMRT QA dosimetric images acquired with the EPID (or scanned films), among other information. When the treatment plan is delivered, whether for pretreatment QA or actual treatment delivery, the R&V system communicates the field setup and de­livery information to the LINAC delivery system as an RTP file and stands by to record the subsequent delivered parameters and capture the acquired im­ages. The IMRT QA process also checks the accura­cy of communication and file transfer between the R&V and delivery systems for each delivered field. From image acquisition to the electronic medical record In order to acquire IMRT dosimetric images with the Varian delivery system and portal imager, the EPID is positioned with the center of the detecting surface aligned to the LINAC cross-hairs and at the desired SSD (minimum of 105 cm, maximum of 140 cm). Since the Varian TPS is programmed to predict non-transit EPID response, no phantom or other buildup is placed between the source and the EPID detecting surface. The delivery system is prompted by the user to acquire a portal dose im­age for each field, and the image must be acquired in “Integrated Acquisition” mode, meaning that the EPID continuously collects data throughout the duration of beam-on time (with maximum readout of 20-30 frames per second19) with no dependence on the timing of LINAC beam pulses, and sums all the collected data from one acquisition to form one image. The patient plan is delivered from the R&V system in QA Mode such that the delivery does not contribute to the tracking of patient dose deliv­ery, but the chart reviewer can see whether or not the fields have been delivered for QA. When the delivery of a single field is complete, the delivery system calculates two images simultaneously from one acquisition: (1) an integrated relative dose im­age of the fluence (Figure 1, right panel), and (2) an absolute dose image computed from the EPID response and the most recent calibration data for the appropriate energy and dose rate (Figure 2). The absolute dose image can be collected from the delivery system treatment console computer via portable drive or network (we have used both methods), but cannot be automatically exported to the R&V system since this system has no informa­tion about the dosimetric calibration of the imager. However, a filter can be set up within the R&V system to automatically collect the relative dose Bailey DW et al. / Paperless IMRT QA 127 FIguRe 1. 2D integrated relative dose images displayed in the R&V software: 1) acquired using the EPID in integrated acquisition mode (right); and 2) predicted by and exported from the TPS (left). These images are saved within the patient’s EMR, attached directly to the appropriate treatment field, and can be compared with various measuring tools within the R&V software (for exam­ple, the measuring tool illustrated in the figure). density image and attach it to the respective field within the patient’s EMR (in DICOM-RT format). Results and discussion Analysis of the acquired EPID images takes two paths, one for the relative dose density image and one for the absolute dose image. Qualitative analysis As mentioned in the previous section, the relative dose density image is collected by the R&V sys­tem and attached to the specific patient field. To check the field geometry and relative dose distri­bution, a respective planned relative dose density map must be exported from the TPS (for the same SSD at which the EPID image was acquired) and similarly attached to the specific field. In this man­ner, the planned fluence and the acquired fluence can be placed side by side in the R&V software for qualitative comparison (Figure 1). This process is analogous to comparing a TPS printout of the flu-ence at a certain SSD to a radiograph exposed to the same IMRTfield at the same SSD as the print­out. The R&V software contains a number of meas­uring tools which can be used to compare the field size, leaf position, qualitative dose distribution, etc. ThoughthistypeofQAdoesnotcontributesub­stantialamountsofinformationtotheabsolutedose QA(discussedbelow)whenperformedonlyonce, it does have one distinct advantage. Currently, dai­lyQAforIMRTtreatmentsisvirtuallynon-existent (thoughsomeinstitutionsarepursuinginvivoQA withEPIDs).20-22However,oneofthemainobjec­tivesofpatient-specificQAistoensurethatthe electronicfilescontainingtreatmentanddelivery systeminformationaccuratelyreflectwhatwas plannedandapprovedintheTPS.IftheEPIDwere usedtotakeaquicknon-transitimageofoneortwo fieldsintheradiotherapyplan(muchthesameway portalimagesarecurrentlyusedforpatientposi­tioning),theseEPIDimagescaneasilyandquickly becomparedtothefluencesalreadyexportedfrom theTPSandstoredinthepatient’sEMR.Inthis way,radiotherapyprofessionalscanquicklyverify 128 Bailey DW et al. / Paperless IMRT QA FIguRe 2. Absolute dose image computed from the EPID response and the most recent calibration data for the appropriate en­ergy and dose rate, as displayed by the delivery system computer upon acquisition. This image is exported from the delivery system treatment console computer to the TPS for comparison to the calculated portal dose prediction for the appropriate field and SSD. that,throughoutthecourseoftreatmentanddaily filetransfer,thecorrecttreatmentfieldsandDMLC positionsarebeingdeliveredaccurately.Usingthis technique,itmayalsobepossibletocatchmechani­calproblems(suchaserrorsinMLCleafandcol­limatorjawpositions)beforethepatientistreated, evenbetweentheextensivemonthlyLINACQA intervals.Thus,thisquick,qualitativeanalysiswith thegrayscaleEPIDimagecouldbeusedonaweek­lybasistoprovidefastandefficientsystemQA, muchasweeklyportfilms(withstaticMLC)are usedtoprovideclinicaltreatmentQA. Quantitative analysis To complete quantitative analysis on the absolute dose EPID image, this file must first be exported from the delivery system treatment console com­puter via portable drive or network connection, and imported to the computer with which the analysis will be completed. It is possible to perform this analysis via custom made software10,12,18, com­mercially available software modalities alterna­tive to the TPS in use23(see EPIDose, SunNuclear, Melbourne, FL), or the portal dose prediction and analysis capabilities of the TPS in use. We current­ly employ the Varian Portal Dosimetry algorithm (Dosimetric Portal Image Calculation, DPIC) with­in the Eclipse TPS to create portal dose predictions for the aS1000 PortalVision EPID at desired SSD. Commissioning of this algorithm requires captur­ing two vendor-specified EPID images (at two dif­ferent SSDs), the diagonal beam profile measured during LINAC commissioning (i.e.along the major diagonal of a 40 x 40 cm2field of desired energy at dmaxin water), and the EPID acquisition of field-size output factors for various field sizes specified by the vendor.24,25To perform IMRT QA with the PortalVision EPID, the Varian TPS has been pro­grammed to predict the response of the EPID to an IMRT field delivered with no buildup or phan­tom between the MLC and the EPID, following the methods pioneered by Van Esch et al. in 2004.11,20 With the EPID dose image imported into the TPS and the respective portal dose prediction calculat­ed, these two planar dose maps can be evaluated through dose difference analysis, gamma evalua­tion, dose profile line scans, isodose comparisons, various measuring tools, etc. (Figure 3). For gam­ma evaluation and dose evaluation, the region of interest can be selected to only include the area of Bailey DW et al. / Paperless IMRT QA 129 FIguRe 3. Portal dose prediction and acquired EPID absolute dose image as compared in the TPS via: 1) predicted vs. measured isodose lines (left panel); and 2) predicted vs. measured dose line profiles (right two panels). This analysis (in PDF or other desired format) is attached to patient’s EMR in the R&V software for approval prior to treatment. The bottom left panel shows the record status dialogue window within the R&V system, including reviewer options such as “pending,” “approved,” “voided,” etc. the detector within the collimator jaws, or a low-dose threshold can be specified by the user which effectively limits the analysis to the image within the collimator jaws. The resolution of the EPID im­age and subsequent analysis is far superior to ioni­zation chambers and 2D arrays, while the ease of calibration and image analysis is far more resource and time efficient than the use of films. To complete the IMRT QA report, the QA analy­sis can be easily and electronically transferred from the TPS to the patient’s EMR in the R&V system by copying the screen to any standard word process­ing or image editing software, or the screen can similarly be printed to PDF or postscript with the appropriate open-source software installed. A QA report can thus be created for each field within the radiotherapy plan and electronically attached to the patient’s chart, requiring no paper, no films, no scanning documents, and no searching for mis­placed QA reports. Furthermore, the R&V system can be set up such that this QA analysis must be approved before the fields are treated (see the sta­tus dialogue window in Figure 3). Conclusions Our radiotherapy department does not have inte­grated planning, R&V, and delivery systems - and yet we have shown that even in this hybrid envi­ronment it is nonetheless possible to develop a completely electronic IMRT QA process. Given the current demandfor paperless patient charts, devel­oping a paperless IMRT QA process is vital, even in systems that understandably include components made by diverse vendors. The process suggested in this study is paperless, filmless, time saving and reliable, enabling the pretreatment IMRT QA 130 Bailey DW et al. / Paperless IMRT QA process to be far more efficient. Furthermore, QA analysis documentation can be directly attached to its specific patient EMR within the R&V system, eliminating searching for documents and running around to obtain signatures, while greatly reduc­ing the risk of misplacing or losing the QA report. The calculated and measured relative dose density images can be viewed electronically side by side within the patient’s EMR to quickly and qualita­tively analyze the density differences, field sizes and MLC trajectories, ensuring proper dose deliv­ery to patients - even on a weekly basis. The ab­solute dose EPID images can be analyzed quickly and thoroughly with custom software or programs supplied by the TPS vendor or a secondary vendor, providing an absolute dose verification system that is of substantially higher resolution than arrays of diodes or ionization chambers, and substantially more efficient than exposing, processing, calibrat­ing, scanning, analyzing and storing films. Acknowledgement Presented in part as an oral presentation at the Annual Meeting of the American Association of Physicists in Medicine 2008, Houston, Texas, USA. References 1. Grabec D, Kragelj B. The sigmoid colon and bladder shielding in whole pel­vic irradiation at prostate cancer (forward planned IMRT from Institute of Oncology Ljubljana). Radiol Oncol 2009; 43: 56-64. 2. Atalar B, Ozyar E, Gunduz K, Gungor G. Intensity modulated radiotherapy (IMRT) in bilateral retinoblastoma. Radiol Oncol 2010; 44: in press. 3. Bates DW, Ebell M, Gotlieb E, Zapp J, Mullins, HC. A proposal for electronic medical records in US primary care. J Am Med Inform Assn 2003; 10: 1-10. 4. Angers C, Renaud J, MacPherson M, Clark B. Saving trees and improving work-flow. [Abstract]. Med Phys 2008; 35: 3413. 5. Hippisley-Cox J, Pringle M, Cater R, Wynn A, Hammersley V, Coupland C, et al. The electronic patient record in primary care: regression or progression? A cross sectional study. Brit Med J 2003; 326: 1439-43. 6. Siochi RAC, Pennington EC, Waldron TJ, Bayouth JE. Radiation therapy plan checks in a paperless clinic. J Appl Clin Med Phys 2009; 10: 43-62. 7. Safran C. Electronic medical records: a decade of experience. J Am Med Inform Assn 2001; 285: 1766. 8. Howell RM, Smith IP, Jarrio CS. Establishing action levels for EPID-based QA for IMRT. J Appl Clin Med Phys 2008; 9: 16-25. 9. Pasma KL, Dirkx MLP, Kroonwijk M, Visser AG, Heijmen BJM. Dosimetric verification of intensity modulated beams produced with dynamic multileaf collimation using an electronic portal imaging device. Med Phys 1999; 26: 2373-8. 10. Talamonti C, Casati M, Bucciolini M. Pretreatment verification of IMRT absolute dose distributions using a commercial a-Si EPID. Med Phys 2006; 33: 4367-78. 11. Van Esch A, Depuydt T, Huyskens DP. The use of an aSi-based EPID for rou­tine absolute dosimetric pre-treatment verification of dynamic IMRT fields. Radiother Oncol 2004; 71: 223-34. 12. van Zijtveld M, Dirkx MLP, de Boer HCJ, Heijmen BJM. Dosimetric pre­treatment verification of IMRT using an EPID: clinical experience. Radiother Oncol 2006; 81: 168-75. 13. Low DA, Harms WB, Mutic S, Purdy JA. A technique for the quantitative evaluation of dose distributions. Med Phys 1998; 25: 656-61. 14. Low DA, Dempsey JF. Evaluation of the gamma dose distribution compari­son method. Med Phys 2003; 30: 2455-64. 15. Depuydt T, Van Esch A, Huyskens DP. A quantitative evaluation of IMRT dose distributions: refinement and clinical assessment of the gamma evaluation. Radiother Oncol 2002; 62: 309-19. 16. Bailey DW, Kumaraswamy L, Podgorsak MB. An effective correction algo­rithm for off-axis portal dosimetry errors. Med Phys 2009; 36: 4089-94. 17. Image Acquisition System 3 reference guide. Palo Alto: Varian Medical Systems, Inc.; 2007. 18. Vial P, Greer PB, Hunt P, Oliver L, Baldock C. The impact of MLC transmitted radiation on EPID dosimetry for dynamic MLC beams. Med Phys 2008; 35: 1267-77. 19. Nicolini G, Fogliata A, Vanetti E, Clivio A, Vetterli D, Cozzi L. Testing the GLAaS algorithm for dose measurements on low-and high-energy photon beams using an amorphous silicon portal imager. Med Phys 2008; 35: 464-72. 20. van Elmpt W, McDermott L, Nijsten S, Wendling M, Lambin P, Mijnheer B. A literature review of electronic portal imaging for radiotherapy dosimetry. Radiother Oncol 2008; 88: 289-309. 21. Wendling M, Louwe RJW, McDermott LN, Sonke JJ, van Herk M, Mijnheer BJ. Accuratetwo-dimensional IMRT verification using a back-projection EPID dosimetry method. Med Phys 2006; 33: 259-73. 22. Wendling M, McDermott LN, Mans A, Sonke JJ, van Herk M, Mijnheer BJ. A simple backprojection algorithm for 3D in vivo EPID dosimetry of IMRT treatments. Med Phys 2009; 36: 3310-21. 23. Malhotra H, Bakhtiari M, Kumaraswamy L, Bailey D, Liu J, Tran T, et al. IMRT QA using a Hybrid Mapcheck/Electronic Portal Dosimetry Environment. [Abstract]. Med Phys 2009; 36: 2574. 24. Vision reference guide: portal vision and portal dosimetry. Palo Alto: Varian Medical Systems Inc.; 2008. 25. Configuration of dosimetric portal image calculation (DPIC). Palo Alto: Varian Medical Systems Inc.; 2004. 131 research article Effect of 905 MHz microwave radiation on colony growth of the yeast Saccharomyces cerevisiae strains FF18733, FF1481 and D7 Ivana Vrhovac, Reno Hrascan, Jasna Franekic Faculty of Food Technology and Biotechnology, University of Zagreb, Zagreb, Croatia Received 1 January 2010 Accepted 12 March 2010 Correspondence to: Reno Hrašcan, Department of Biochemical Engineering, Faculty of Food Technology and Biotechnology, University of Zagreb, Pierottijeva 6, 10000 Zagreb, Croatia. Phone: +385 1 48 36 013; Fax: 385 1 48 36 016; E-mail: rhrascan@pbf.hr Disclosure: No potential conflicts of interest were disclosed. Background. The aim of this study was to evaluate the effect of weak radiofrequency microwave (RF/MW) radiation emitted by mobile phones on colony growth of the yeast Saccharomyces cerevisiae. Materials and methods. S. cerevisiae strains FF18733 (wild-type), FF1481 (rad1 mutant) and D7 (commonly used to detect reciprocal and nonreciprocal mitotic recombinations) were exposed to a 905 MHz electromagnetic field that closely matched the Global System for Mobile Communication (GSM) pulse modulation signals for mobile phones at a specific absorption rate (SAR) of 0.12 W/kg. Results. Following 15-, 30-and 60-minutes exposure to RF/MW radiation, strain FF18733 did not show statistically significant changes in colony growth compared to the control sample. The irradiated strains FF1481 and D7 demon­strated statistically significant reduction of colony growth compared to non-irradiated strains after all exposure times. Furthermore, strain FF1481 was more sensitive to RF/MW radiation than strain D7. Conclusions. The findings indicate that pulsed RF/MW radiation at a low SAR level can affect the rate of colony growth of different S. cerevisiae strains. Key words: microwave radiation; Saccharomyces cerevisiae; colony growth Introduction Microwave radiation is a type of non-ionizing elec­tromagnetic radiation widely used in industry, commerce, medicine and for private purposes, es­pecially in mobile communication. In recent years, the use of mobile phones has accelerated, result­ing in increasing exposure of the environment to weak radiofrequency microwave (RF/MW) radia­tion generated by these devices. Although the aver­age exposure levels are low compared to exposure limits, public concern about the potential hazard on human health is growing.1Numerous experi­mental studies evaluating the biological effects caused by RF/MW radiation are controversial and no unanimous conclusion has been reached.2-4 It is well-documented that yeasts are representa­tive of eukaryotes, including human cells, in many aspects of fundamental cellular processes.5Many experiments, with the yeast Saccharomyces cerevi­siaeas a model organism, can be performed under biologically and technically well-controlled condi­tions after exposure to microwave radiation.6 Theobjectiveofthisstudywastoevaluatethepo­tentialeffectof905MHzRF/MWradiationsimilar tothatemittedbymobilephonesoncolonygrowth ofS.cerevisiaestrainsFF18733,FF1481andD7. Materials and methods Yeast strains This experiment was carried out using three S. cer­evisiaestrains. The FF18733 strain (MATa leu2-3,112 trp1-289 ura3-52 his7-2 lys1-1) is a wild-type, where­as the derived FF1481 strain (MATa leu2-3,112 trp1­289 ura3-52 his7-2 lys1-1 rad1::LEU2) is deficient in nucleotide excision repair.7The D7 strain (MATa/a ade2-40/ade2-119 trp5-12/trp5-27 ilv1-92/ilv1-92) is 132 Vrhovac I et al. / Effect of microwave radiation in Saccharomyces cerevisiae FIGURE 1. Colony growth of yeast S. cerevisiae strains FF18733, FF1481 and D7 after 15-, 30- and 60-minutes exposure to 905 MHz microwave radiation. Values represent means and standard deviations. relatively genetically unstable. Therefore, changes in mitotic crossing-over, mitotic gene conversion and reverse mutations may occur spontaneously.8 Experimental procedures A preculture of three strains of S. cerevisiaewas suspended in yeast extract liquid (YEL) and grown for 48 h at 28°C. Precultured cells (2 x 106cells/ml) were then suspended in YEL and grown for 18 h at 28°C. Half of each culture (2 x 108cells/ml), prepared in triplicate, were exposed to 905 MHz microwave radiation for 15, 30 and 60 minutes, whereas the other half served as a control. After radiation treatment, yeast cells were inoculated on solid complete growth medium and grown over­night at 28şC. Thereafter, the number of colonies of the three strains (irradiated and non-irradiated) was counted under a magnifier. Exposure conditions An electromagnetic field was generated using a Gigahertz Transversal Electromagnetic Mode Cell (GTEM-cell) model 5402 (ETSTMLindgren, USA) equipped with a signal generator (Antrisu MS27211B, Japan), signal amplifier (RF 3146 Power Amp Module, RF Micro Devices, Greensboro, USA) and a signal modulator (RF 2722 Polaris Chip, RF Micro Devices, Greensboro, USA). The signal am­plifier was used to amplify the RF/MW signal in­duced by the signal generator, whereas the signal modulator was used to modulate a continuous wave to pulse signal used in the Global System for Mobile Communication (GSM) mobile phones. Yeast suspensions were exposed to 905 MHz RF/MW with the GSM basic signal modulation for 15, 30 and 60 minutes. Inside the GTEM-cell, the electromagnetic field strength was 10 V/m, and the temperature was 28°C. The average specific ab­sorption rate (SAR) for a single cell was 0.12 W/kg. SAR was calculated by averaging the individual parameters of the cell components in accordance with their volume fraction in live cells.9 Statistical analysis Statistical analyses were carried out with descrip­tive statistics. Significant differences in colony growth were determined using the Student’s t-test. Values of Plower than 0.05 were considered statis­tically significant. Results Figure 1 shows the colony growth of three S. cer­evisiaestrains after 15-, 30- and 60-minutes expo­sure to 905 MHz RF/MW radiation similar to that emitted by mobile phones at SAR of 0.12 W/kg. The number of non-irradiated colonies of each strain was taken as 100% and the percent of irradiated colonies after different exposure times was calcu­lated with respect to this control sample. Following a 15-, 30- and 60-minutes exposure to RF/MW ra­diation, the wild-type strain FF18733 did not show statistically significant changes in colony growth compared to the control sample. Irradiated strains FF1481 and D7 demonstrated statistically signifi­cant reduction of colony growth compared to non-irradiated strains after all exposure times. The data indicate that RF/MW radiation decreased colony growth of strains FF18733, FF1481 and D7 result­ing in a 19.30±2.06%, 56.37±1.49% and 34.29±3.21% growth reduction after 60-minutes exposure, re­spectively. Discussion Users of mobile phones are exposed to weak mi­crowave radiation. In this context, the possible ef­fects of RF/MW radiation on genetic material are Vrhovac I et al. / Effect of microwave radiation in Saccharomyces cerevisiae 133 very important. Many studies on mammalian cells failed to find microwave-induced DNA damage and cell proliferation10-12; in contrast with expo­sure to ionizing radiation where the DNA damage is well known.13Other studies have reported that modulated RF/MW radiation is capable of causing DNA lesions and inhibition of cell proliferation.14,15 In this study, we estimated the effect of mobile phones radiofrequency of 905 MHz on the yeast S. cerevisiaestrains FF18733, FF1481 and D7. Strains FF1481 and D7 demonstrated a statistically signifi­cant difference in colony growth after 15-, 30- and 60-minutes exposure to pulsed RF/MW radiation at SAR 0.12 W/kg. Therefore, these strains showed increased sensitivity to RF/MW radiation and re­duction of colony growth was time-dependent. An earlier experiment with the yeast S. cerevisiaedemonstrated either an increased (up to 15%) or decreased (up to 38%) cell growth rate by certain frequencies of microwave radiation within a 41.6­41.8 GHz band.16-18 It is known that microwave radiation may occur directly by DNA lesion and/or indirectly by dam­age to DNA repair mechanisms. Strain FF1481 of S. cerevisiaeis deficient in nucleotide excision repa­ir due to an insertion of the functional LEU2gene at the RAD1locus and rad1becomes non-functio­nal. Rad1, in complex with Rad10, exhibits single-stranded DNA endonuclease activity and cleaves 3’-ended single-stranded DNA at its junction with the duplex DNA.19Since we observed a significant decrease of rad1mutant cell proliferation, it seems that pulsed RF/MW radiation at a low SAR level during short exposure times could induce DNA damage in S. cerevisiae cells. Mitotic recombination is necessary during mi­tosis for the repair of DNA single- and double-strand breaks, and mutagenic lesions generated by exposure to chemicals and radiation.20Strain D7 of S. cerevisiaeis commonly used to detect re­ciprocal (crossing-over) and nonreciprocal (gene conversion) mitotic recombinations and reverse mutations. Besides evaluation of the RF/MW effect on cell growth rate of strain D7, we estimated the induction of mitotic gene conversion and reverse mutations in strain D7 after 15-, 30- and 60-minutes exposure to 905 MHz RF/MW at SAR 0.12 W/kg. The frequency of gene conversion at the trplocus and reverse mutation at the ilvlocus showed only a slight tendency to increase compared to the con­trol sample (data not shown). Preliminary results indicate that modulated RF/MW radiation with a low SAR value did not affect either the rate of gene conversion nor reverse mutations in strain D7. Gos et al.21reported that mobile phone fields at 900 MHz with SAR of 0.13 and 1.3 W/kg did not exhibit any effect on mutations or recombinations in S. cerevisiaecells either in the absence or presence of genotoxic stress. In conclusion, our study showed that three S. cerevisiaestrains exhibit different patterns of colony growth after 15-, 30- and 60-minutes exposure to a mobile phones radiofrequency of 905 MHz at SAR 0.12 W/kg. Strains FF1481 (DNA repair mutant) and D7 (relatively genetically unstable) demon­strate an increased sensitivity to RF/MW radiation in comparison to strain FF18733 (wild-type). The data indicate that pulsed RF/MW radiation at a low SAR level could induce DNA damage in S. cerevi­siaecells. This points to the need for further studies of DNA repair mechanisms in yeast cells. Acknowledgments This work was supported by grant 058-0582261­2246 from the Ministry of Science and Technology, Republic of Croatia. The authors thank Ivancica Trošic, PhD (Radiation Dosimetry and Radiobiology Unit, Institute for Medical Research and Occupational Health) for the opportunity to use the GTEM-cell in this experiment. 134 Vrhovac I et al. / Effect of microwave radiation in Saccharomyces cerevisiae References 1. WorldHealthOrganization(WHO).Electromagneticfieldsandpublichealth: mobile telephones and their base stations, displayed 22 February 2010. Available at: http://www.who.int/mediacentre/factsheets/fs193/en. 2. Grundler W, Keiser F, Keilmann F, Walleczek J. Mechanisms of electromag­netic interaction with cellular systems. Naturwissenschaften 1992; 79:551-9. 3. Pacini S, Ruggiero M, Sardi I, Aterini S, Gulisano F, Gulisano M. Exposure to global system for mobile communication (GSM) cellular phone radiof­requency alters gene expression, proliferation, and morphology of human skin fibroblasts. Oncol Res 2002; 13: 19-24. 4. Zeni O, Romano M, Perrotta A, Lioi MB, Barbieri R, dAmbrosio G, et al. Evaluation of genotoxic effects in human peripheral blood leukocytes following an acute in vitro exposure to 900 MHz radiofrequency fields. Bioelectromagnetics 2005; 26: 258-65. 5. Botstein D, Fink GR. Yeast: an experimental organism for modern biology. Science 1988; 240: 1439-43. 6. Gos P, Eicher B, Kohli J, Heyer WD. Extremely high frequency electromagnet­ic fields at low power density do not affect the division of exponential phase Saccharomyces cerevisiae cells. Bioelectromagnetics 1997; 18: 142-55. 7. Štafa A, Svetec I-K, Zgaga Z. Inactivation of the SGS1 and EXO1 genes syn-ergistically stimulates plasmid integration in yeast. Food Technol Biotechnol 2005; 43: 103-8. 8. Zimmermann FK, Kern R, Rasenberger H. A yeast strain for simultaneous detection of induced mitotic crossing over, mitotic gene conversion and reverse mutation. Mutat Res 1975; 28: 381-8. 9. Steffensen KV, Raskmark P, Penersen GF. FTDT calculations of the EM-field distribution in a microtiter suspension well. In: Chiabrern A, Juutilainen J, editors. Proceedings of the COST 244 Workshop: Biomedical Effects of Electromagnetic Fields; 3-4 Sep 1995; Kuopio, Finland, p. 80-7. 10. Malyapa RS, Ahern EW, Straube WL, Moros EG, Pickard WF, Roti Roti JL. Measurement of DNA damage after exposure to electromagnetic radiation in the cellular phone communication frequency band (835.62 and 847.74). Radiat Res 1997; 148: 618-27. 11. Zeni O, Chiavoni AS, Sannino A, Antolini A, Forgio D, Bersani F. Lack of genotoxic effects (micronucleus induction) in human lymphocytes exposed in vitro to 900 MHz electromagnetic fields. Radiat Res 2003; 160: 152-8. 12. Gurisik E, Warton K, Martin DK, Valenzuela SM. An in vitro study of the ef­fects of exposure to a GSM signal in two human cell lines: monocytic U937 and neuroblastoma SK-N-SH. Cell Biol Int 2006; 30: 793-9. 13. Miklos M, Gajski G, Garaj-Vrhovac V. Usage of the standard and modified comet assay in assessment of DNA damage in human lymphocytes after exposure to ionizing radiation. Radiol Oncol 2009; 43: 97-107. 14. Phillips JL, Ivaschuk O, Ishida-Jones T, Jones RA, Campbell-Beachler M, Haggren W. DNA damage in Molt-4 T-lymphoblastoid cells exposed to cellu­lar radiofrequency field in vitro. Bioelectrochem Bioenerg 1998; 45: 103-10. 15. Diem E, Schwarz C, Adlkofer F, Jahn O, Rüdiger. Non-thermal DNA breakage by mobile-phone radiation (1800 MHz) in human fibroblasts and in trans­formed GFSH-R17 rat granulosa cells in vitro. Mutat Res 2005; 583: 178-83. 16. Grundler W, Keilmann F. Nonthermal effects of millimeter microwaves on yeast growth. Z Naturforsch C Biosci 1978; 33: 15-22. 17. Grundler W. Intensity-and frequency-dependent effects of microwaves on cell growth rates. Bioelectrochem Bioenerg 1992; 27: 361-5. 18. Grundler W, Kaiser F. Experimental evidence for cohorent excitations cor­related with cell growth. Nanobiology 1992; 1: 163-76. 19. Bardwell AJ, Bardwell L, Tomkinson AE, Friedberg EC. Specific cleavage of model recombination and repair intermediates by the yeast Rad1-Rad10 DNA endonuclease. Science 1994; 265: 2082-5. 20. Paques F, Haber JE. Multiple pathways of recombination induced by double-strand breaks in Saccharomyces cerevisiae. Microbiol Mol Biol Rev 1999; 63: 349-404. 21. Gos P, Eicher B, Kohli J, Heyer WD. No mutagenic or recombinogenic effects of mobile phone fields at 900 MHz detected in the yeast Saccharomyces cerevisiae. Bioelectromagnetics 2000; 21: 515-23. Radiol Oncol 2010; 44(2): 67-78. doi:10.2478/v10019-010-0025-9 Radioterapija v kombinaciji z zdravili usmerjenimi proti tumorskemu žilju Ciric E, Serša G Izhodišca. Glede na odlocilno vlogo tumorskega žilja pri razvoju malignih tumorjev je bilo veliko naporov vloženih v razvoj zdravil usmerjenih v tumorsko žilje. Razvitih je bilo veliko razlicnih zdravil, ki jih v splošnem uvršcamo v dve skupini. Antiangiogena zdravila se vpletajo v proces angiogeneze in v tumorjih zavirajo nastanek novih žil. Žilno razdiralna zdravila unicujejo obstojece tumorsko žilje in v tumorju povzrocajo ishemijo s posledicno nekrozo tumorja. Kljub velike-mu terapevtskemu potencialu obeh skupin zdravil postaja jasno, da bodo klinicno najbolj uporabna v kombinacijah z drugimi vrstami protirakavega zdravljenja. Radioterapija je zelo široko uporabljan nacin zdravljenja, ki je povezan z drugacnimi terapevtskimi izzivi in je zato kombinacija obeh pristopov smiselna. Zakljucki. Za ucinkovito uporabo zdravil usmerjenih proti tumorskemu žilju v kombinaciji z radioterapijo obstaja moc­na biološka podlaga. Izkazalo se je, da antiangiogena in žilno razdiralna zdravila vplivajo na tumorsko mikrookolje na takšen nacin, da povecajo odgovor tumorja na obsevanje. Rezultati predklinicnih in prvih klinicnih raziskav so potrdili terapevtski potencial te nove strategije zdravljenja, vendar so opozorili tudi na vecjo možnost stranskih ucinkov na zdrava tkiva. Radiol Oncol 2009; 44(2): 79-85. doi:10.2478/v10019-010-0022-z Hiperhomocisteinemija in vloga B vitaminov pri raku Plazar N, Jurdana M Izhodišca. Bolniki z rakom imajo pogoste srcno-žilne zaplete in predstavljajo tveganje za razvoj venske tromboze. Epidemiološke raziskave porocajo, da so bolezni srca in ožilja povezane z povišano koncentracijo homocisteina v krvi (hiperhomocisteinemija), ki se pojavlja tudi pri raku. Homocistein je aminokislina v krvi, ki nastaja pri presnovi metionina in ga v telo vnašamo s hrano. Zato na raven homocisteina v krvni plazmi mocno vpliva prehrana pa tudi genetski dejavniki. Prehrambene ucinkovine z najmocnejšim vplivom so B vitamini: folna kislina, vitamina B6 in B12. Vloga B vita-minov je pomembna za normalno vzdrževanje strukture DNK-ja. Raziskave opravljene na živalih in ljudeh so pokazale, da pomanjkanje folatov vpliva na proces metilacije in sintezo DNK-ja ter posledicno na visoko raven homocisteina v krvi. Zaradi pomembne vloge folatov mnogo raziskav že vrsto let proucuje omenjeno povezavo. Zakljucki. Hiperhomocisteinemija je eden glavnih skupnih znacilnosti povezanih z vensko in arterijsko trombozo pri mnogih rakavih obolenjih, patofiziologija slednje povezave pa še ni povsem pojasnjena. Ker ima metabolizem B vita-minov, predvsem folatov kljucni pomen pri razvoju hiperhomocisteinemije pri raku, smo v preglednem clanku navedli literaturo, ki opisuje tovrstno povezavo. Radiol Oncol 2010; 44(2): I-VII. Radiol Oncol 2010; 44(2): 86-91. doi:10.2478/v10019-010-0021-0 Ucinkovito zaustavljanje krvavitve s prevezo iz hitina po perkutani punkciji arterije. Eksperimentalna raziskava na hepariniziranih živalih Kranokpiraksa P, Pavcnik D, Kakizawa H, Uchida BT, Jeromel M, Keller FS, Rösch J Izhodišca. V raziskavi smo testirati ucinkovitost preveze iz hitina, da bi zaustavili krvavitev po perkutani punkciji povrh­nje femoralne arterije in po uporabi žilnega uvajala velikosti 8 F. Ucinkovitost preveze iz hitina smo primerjali z metodo standardne rocne kompresije. Material in metode. Pri devetih hepariniziranih ovcah smo perkutano punktirali povrhnjo femoralno arterijo obo­jestransko in vanjo za 5 minut uvedli žilno uvajalo velikosti 8 F. Po odstranitvi žilnega uvajala iz obeh punkcijskih mest smo randomizirano zaustavljali krvavitev s kompresijo s prevezo iz hitina ali pa s standardno rocno kompresijo. Obe kompresiji smo izvajali 5 minut. Nato smo uspešnost zaustavitve krvavitev preverjali angiografsko. Kadar smo angiograf­sko prikazali krvavitev iz punkcijskega mesta, smo nadaljevali z rocno kompresijo in ponavljali angiografske prikaze v razmaku 2,5 minut, dokler nismo popolno zaustavili krvavitve. Rezultati. Po 5-ih minutah kompresije s prevezo iz hitina smo uspešno zaustavili krvavitev pri sedmih živalih (77,8 %). Srednje trajanje kompresije, s katero smo s prevezo iz hitina zaustavili krvavitev (6,9 ± 3,9 minut), se je statisticno po­membno (p=0,019) razlikovalo od srednjega trajanja pri standardni rocni kompresiji (10,8 ± 2,8 minut). Po zaustavitvi krvavitve s kompresijo s prevezo iz hitina je bil pretok neoviran v vseh devetih arterijah (100%). V tej skupini smo odkrili, da je hematom manjši (2/9) v primerjavi s kontrolno skupino (8/9). Zakljucki. Uporaba kompresije s prevezo iz hitina po odstranitvi žilnega uvajala iz arterije v primerjavi s standardno rocno kompresijo statisticno pomembno skrajša cas, v katerem smo dosegli hemostazo. Pri tem ni bilo vec zaple­tov. Za skrajšanje casa za hemostazo in preprecitev hematoma je pomembna pravilna uporaba preveze iz hitina. Pricakujemo lahko, da bomo takšno, prevezo, ki jo sedaj uporabljamo za zaustavitev krvavitve pri poškodbah, v pri­hodnosti uporabljali tudi za zaustavitev krvavitve po diagnosticnih in terapevtskih endovaskularnih posegih. Radiol Oncol 2010; 44(2): I-VII. Radiol Oncol 2010; 44(2): 92-96. doi:10.2478/v10019-010-0027-7 Uporaba 3T MR slikanja pri odkrivanju bolezni, povezanimi z azbestom Podobnik J, Kocijancic I, Kovac V, Serša I Izhodišca. Namen raziskave je bil preveriti možnosti uporabe 3T MR slikanja pri odkrivanju in spremljanju bolezenskih sprememb, ki so posledica izpostavljenosti azbestnemu prašenju. Bolniki in metode. V raziskavo smo vkljucili 15 bolnikov, 5 z azbestno boleznijo in 10 z malignim plevralnim mezote­liomom (MPM). Bolniki z azbestno boleznijo so nam v digitalni obliki posredovali svoje prejšnje CT preiskave, ki niso bile starejše od enega meseca. Pri bolnikih z MPM pa smo zaradi primerjave sami opravili CT preiskavo, ki je bila potrebna pred pricetkom zdravljenja. Pri vseh smo naredili MR slikanje prsnega koša. Protokol je bil sestavljen iz T2 obteženih pulznih zaporedij v koronarni, sagitalni in transverzalni ravnini ter T1 obteženih pulznih zaporedij v transverzalni ravnini. Vse meritve smo naredili, ko je bolnik zadrževal dih in z uporabo EKG prožilca. Pri T2 obteženih pulznih zaporedjih smo uporabili tudi SPIR tehniko za zasicenje signala iz mašcevja. CT preiskave smo naredili z uporabo jodnega kontrastne­ga sredstva. Slikali smo podrocje od pljucnih apeksov do spodnjega roba jeter. Rekonstruirali smo slike v transverzalni ravnini debeline 5 mm v mediastinalnem oknu ter 3 mm v pljucnem oknu. Na delovni postaji Singo MultiModality Work Place smo izmerili intenzivnost MR signala v leziji in v mišici v isti rezini. Rezultati. Plevralni plaki imajo na T1 in T2 obteženih slikah v primerjavi z mišico hipointenziven signal. Na T1 obteženih MR slikah je MPM izrazito nehomogen in v primerjavi z mišico hipointenziven, na T2 obteženih slikah pa hiperintenziven. Zakljucki. Raziskava je pokazala, da je MR slikanje prsnega koša primerna diagnosticna slikovna metoda za prikaz sprememb, ki so posledica izpostavljenosti azbestu. T2 obtežene slike lahko celo nakazujejo maligno naravo bolezni. Pri MR slikanju bolniki niso izpostavljeni škodljivemu ionizirajocemu sevanju, zato lahko to slikovno metodo uporabljamo pri spremljanju rezultatov zdravljenja ali razvoja bolezni. Kontrastna sredstva, ki se uporabljajo pri MR slikanju so varnejša in manj nefrotoksicna v primerjavi z jodnimi kontrastnimi sredstvi, kar je zlasti pomembno pri bolnikih, ki jih zdravimo s kemoterapijo. Radiol Oncol 2010; 44(2): 97-102. doi:10.2478/v10019-010-0006-z Zanesljivost difuzijskega magnetnoresonancnega slikanja pri razlikovanju med degenerativnimi in vnetnimi spremembami terminalne plošce Oztekin O, Calli C, Kitis O, Adibelli ZH, Eren CS, Apaydin M Izhodišca. Namen raziskave je bil preveriti uporabnost difuzijskega magnetnoresonancnega (MR) slikanja pri razliko­vanju med degenerativno spremembo Modic tip 1 in akutnim infekcijskim spondilodiscitisom. S konvecionalnim MR slikanjem je razlikovanje med obema patološkima spremembama težavno in pogosto nezanesljivo. Boniki in metode. V retrospektivno raziskavo smo vkljucili 27 bolnikov z erozivno intervertebralno osteohondrozo Modic tip 1 in 18 bolnikov s spondilodiscitisom. Magnetnoresonancne preiskave smo naredili z napravo 1,5 Tesla. Izkušen radiolog je pregledal MR slike lumbalne hrbtenice 45 bolnikov. Bolnikom s spremembo Modic tip 1 smo naredili tudi CT slikanje prizatedega vretenca in medvretencne plošcice. Rezultati. Slika kostnega mozga je bila v bližini terminalne plošce tako pri bolnikih z Modic tip 1 kakor tudi pri bolnikih s spondilodiscitisom hipointenzivna na T1 poudarjenih slikah. Na T2 poudarjenih slikah pa je bila pri obeh vrstah bolnikov hiperintenzivna. Slika Modic tip 1 je bila pri difuzijski MR hipointenzivna, pri spondilodiscitisu pa je bila hiperintenzivna. Zakljucki. Naša raziskava je pokazala, da z difuzijsko MR preiskavo lahko zanesljivo locimo med degenerativno spremembo Modic tip 1 in akutnim spondilodiscitisom, s konvecionalnim MR slikanjem in tudi s klinicnim pregledom pa je locevanje nezanesljivo. Radiol Oncol 2010; 44(2): I-VII. Radiol Oncol 2009; 44(2): 103-106. doi:10.2478/v10019-010-0024-x Embolizacija z N-butyl-cyanoakrilatom pri znotrajžilnem zdravljenju priapizma z visokim pretokom po poškodbi Radoš M, Šunjara V, Sjekavica I, Štern Padovan R Izhodišca. Priapizme (trajna erekcija brez vzburjenja) delimo v nizko pretocne (venske ali ishemicne) in visoko pretoc­ne (arterijske ali neishemicne). Diagnozo visoko pretocnega priapizma ugotovimo z barvno Dopplersko ultrazvocno preiskavo in arteriografijo ter ga obicajno zdravimo z znotrajžilnim zapiranjem fistule. Prikaz primera. Opisujemo primer 20-letnega bolnika z visokopretocnim tipom priapizma, ki je nastal po poškodbi perineja. Bolnika smo najprej opazovali in nato neuspešno zdravili z embolizacijo fistule, pri kateri smo uporabili resorp­tivno gelatinozno peno. Ponoven endovaskularni poseg, ki smo naredili embolizacija z N-butyl-cyanoakrilatom, je bil uspešen. Zakljucki. Visokopretocni priapizem lahko uspešno zdravimo z znotrajžilnim posegom, pri cemer je pomembna op-timalna izbira embolizacijskega sredstva in natancna izvedba posega. Radiol Oncol 2009; 44(2): 107-112. doi:10.2478/v10019-010-0028-6 Izraženost antigenov pri ponavljajocih se meningiomih Vranic A Izhodišca. Meningiomi so intrakranialni tumorji, ki se pogosto ponovno pojavljajo. V literaturi opisujejo, da pride do ponovitve pri 20% benignih, do 80% atipicnih in do 100% malignih meningiomih. Najpogostejši napovedni dejavniki za ponovni vznik bolezni so gradus meningioma, invazivnost v možganovino in radikalnost nevrokirurške resekcije. Namen naše raziskave je bil oceniti razlike med ponavljajocimi se meningiomi in meningeomi, ki se ne ponavljajo, v izražanju nekaterih antigenov na površini njihovih celic. Metode. Primerjali smo 19 ponavljajocih se in 35 ne ponavljajocih se meningiomov. S pomocjo imunohistokemije smo ocenjevali razlike v izražanju antigena MIB-1, progesteronskih receptorjev, katepsina B in katepsina L. Rezultati. Izražanje MIB-1 antigena je vecje pri ponavljajocih se meningiomih (p=0,001). V izražanju progesteronskih receptorjev nismo opazili statisticno pomebnih razlik med skupinama. Imunohistokemicno ocenjeno izražanje katep­sina B (p= 0,007) in katepsina L (p<0,001) je bilo vecje v skupini ponavljajocih se meningiomov. Zakljucki. V raziskavi smo ugotovili, da je pri ponavljajocih se meningiomih veckrat izražen antigen MIB. V prihodnje bi to ugotovitev lahko uporabili kot dodaten napovedni dejavnik, ki bi nam poleg standardnih dejavnikov pomagal pri ocenjevanju tveganja za ponovitev bolezni. Radiol Oncol 2010; 44(2): I-VII. Radiol Oncol 2010; 44(2) 113-120. doi:10.2478/v10019-010-0023-y Pogostnost hipermetilacije MGMT (06-metilgvanin-DNK metiltransferaza) pri bolnikih z glioblastomom in dolgotrajnim preživetjem; izkušnje dunajske ustanove Baur M, Preusser M, Piribauer M, Elandt K, Hassler M, Hudecm M, Dittrich C, Marosi C Izhodišca. Namen retrospektivne raziskave je bila analiza metilacijskega statusa promotorja za MGMT (06-metilgva- nin-DNK metiltransferazo) pri bolnikih z glioblastomom (GBM), pri katerih smo ugotovili dolgotrajno preživetje (= 3 leta). Metode. Metilacijski status promotorja za MGMT smo ugotavljali z bisulfidno modifikacijo DNK in nato z verižno poli­merazno reakcijo specificno za metilacijo. DNK smo izlocili iz vzorcev tumorja, ki so bili fiksirani v formalinu in vkljuceni v parafinske bloke. Rezultati. Vzorce, primerne za interpretacijo smo z verižno polimerazno reakcijo dobili le pri 14 od 33 (42%) bolnikih z GBM in z dolgotrajnim preživetjem. Metilirane proge smo videli pri 3 od 14 bolnikih, metilirane in nemetilirane pri 8 od 14, samo nemetilirane pa pri 3 bolnikih. Metiliacijo promotorja za MGMT smo tako dokazali pri 11 od 14 bolnikih. Skupini bolnikov z metiliranim in nemetiliranim statusom promotorja za MGMT sta bili premajhni za kakršnekoli zanesljive statisticne zakljucke. Zakljucki. Bolniki z GBM, pri katerih smo ugotovili dolgotrajno preživetje, imajo pogosto prisotno intratumorsko me-tilacijo promotorja za MGMT. Ta metilacija locuje bolnike z dolgotrajnim preživetjem od neizbrane skupine bolnikov z GBM. Za povecanje zanesljivosti metodologije dolocanja statusa metilacije MGMT je potrebna standardizacija po­stopka verižne polimerazne reakcije specificne za metilacijo. Radiol Oncol 2010; 44(2): 121-123. 10.2478/v10019-010-0004-1 Klinicni primer zasevka raka danke v penis Yildirim M, Coskun A, Pürten M, Oztekin O, Ilhan E Izhodišca. Zasevki v penis so redki, obicajno pa jih povzrocajo genitourinarni in kolorektalni raki. Prikaz primera. Prikazujemo 77-letnega bolnika z zasevkoma raka danke v penis. Na kliniko je bil napoten zaradi razjede na penisu, semierektilnega penisa in motenj uriniranja. Slikovne preiskave so pokazale vozlicasto spremembo v glansu penisa in številne kostne zasevke. Na zdravljenje s kemoterapijo ni odgovoril in ocenili smo, da je napoved poteka bolezni slaba. Zakljucki. Slikovne preiskave in biopsija so razjasnili diagnozo bolezni, zdravljenje takšnih bolnikov pa ni dovolj uspe­ šno, kot kaže tudi primer prikazanega bolnika. Radiol Oncol 2010; 44(2): I-VII. Radiol Oncol 2010; 44(2): 124-130. doi: 10.2478/v10019-010-0017-9 Elektronska metoda za zagotavljanje kakovosti pri obsevanju IMRT s povezovanjem megavoltnih elektronskih slikovnih naprav z nacrtovalnim sistemom, sistemom zabeležbe in overitve ter obsevalnimi napravami Bailey DW, Kumaraswamy L, Podgorsak MB Izhodišca. Namen raziskave je bil uvajanje elektronske metode za zagotavljanje kakovosti pri obsevanju IMRT, to je obsevanju z modulirano intenziteto žarkovnega snopa. Uporabili smo megavoltne elektronske slikovne naprave iz amorfnega silicija in jih povezali z nacrtovalnim sistemom, sistemom zabeležbe in overitve (R&V) ter z obsevalnimi napravami neodvisnih proizvajalcev. Metode. Z nacrtovalnim sistemom smo na podlagi dejanskega obsevalnega nacrta ustvarili verifikacijski obsevalni nacrt. Obsevalna polja smo prenesli na sistem R&V, nato smo jih obsevali in pri tem kontolirali kakovost (nacin QA) z iztegnjeno elektronsko portalno slikovno napravo. Zajete dozimetricne slike smo avtomaticno shranili v formatu DICOM-RT na trdi disk racunalnika, ki je vodil obsevalno napravo. Slike relativne doze smo nato poslali na sistem R&V. Slike z informacijo o absolutni dozi smo prenesli z racunalnika, ki je vodil obsevalno napravo, na nacrtovalni sistem in uvedli v verifikacijski obsevalni nacrt za nadaljnjo analizo. Zajete slike z ekrana smo uvedli v sistem R&V v obliki elek­tronske datoteke (npr. PDF). Pri tem smo lahko na ekranu videli primerjavo uporabe analize gama in tudi primerjavo izodoznih crt. To smo naredili pred pricetkom obsevanja bolnika. Tudi sliko relativne doze, ki jo je izracunal nacrtovalni sistem, smo lahko poslali na sistem R&V in jo na ta nacin primerjali s sliko relativne doze, ki smo jo napravili z elektronsko portalno slikovno napravo. Rezultati. Na našem oddelku nacrtovalni sistem, sistem R&V ter obsevalne naprave niso integrirani. Kljub temu smo uspeli zagotoviti kakovost obsevanja IMRT brez beleženja na papir in na film. V sistemu R&V smo bolnikovim podatkom pripeli ustrezne dokumente v elektronski obliki. Na ta nacin smo omogocili bolj ucinkovito analizo in overovitev. Sliki, ki sta kazali izracunano in izmerjeno relativno dozno porazdelitev, smo lahko v elektronski obliki primerjali v sistemu R&V. Tako smo lahko analizirali razlike med njima in zagotovili, da bolnika obsevamo s pravilno dozo. Zakljucki. Pokazali smo, da je elektronsko zagotavljanje kakovosti obsevanja IMRT mogoce, tudi kadar planirni sis-tem ni integriran s sistemom R&V in obsevalnimi napravami. Radiol Oncol 2010; 44(2): I-VII. Radiol Oncol 2010; 44(2): 131-134. doi: 10.2478/v10019-010-0019-7 Ucinek 905 MHz mikrovalovnega sevanja na rast kolonij kvasovk Saccharomyces cerevisiae sevov FF18733, FF1481 in D7 Vrhovac I, Hrašcn R, Franekic J Izhodišca. Namen raziskave je bil prouciti ucinek nizkoradiofrekvencnega valovanja (RF/MW), ki ga sevajo mobilni telefoni, na rast kvasovk Saccharomyces cerevisiae. Materiali in metode. Sevi S. cerevisiae FF18733 (divji tip), FF1481 (mutacija rad1) in D7 (obicajno uporabljen za detekcijo reciprocne in nereciprocne mitoticne rekombinacije) so bili izpostavljeni 905 MHz elektromagnetnemu polju ki je zelo podoben signalu pri globalnem sistemu prenosnih komunikacij (Global System for Mobile Communication, GSM) uporabljenem v prenosnih telefonih pri specificni absorpciji (SAR) 0.12 W/kg. Rezultati. Ko smo izpostavili sev FF18733 S. Cerevisiae 15-, 30-in 60-minutnemu obsevanju RF/MW, nismo statisticno znacilno vplivali na rast kvasovk v primnerjavi s kontrolnimi neobsevanimi kvasovkami. Ko pa smo obsevali seve FF1481 in D7, smo statisticno znacilno vplivalo na rast kvasovk pri vseh dolžinah obsevanja. Pokazala se je razlika v obcutljivosti, sev FF1481 je bil bolj obcutljiv na obsevanje RF/MW kot sev D7. Zakljucki. Rezultati nakazujejo, da lahko pri nizkem SAR pulzno obsevanje RF/MW zavira rast razlicnih sevov S. ce­revisiae. Radiol Oncol 2010; 44(2): I-VII. Notices Notices submitted for publication should contain a mailing address, phone and/or fax number and/or e-mail of a Contact person or department. Radiol Oncol 2010; 44(2): VIII-IX. As a service to our readers, notices of meetings or courses will be inserted free of charge. Please send information to the Editorial office, Radiology and Oncology, Zaloška 2, SI-1000 Ljubljana, Slovenia. Radiol Oncol 2010; 44(2): VIII-IX. Fundacija "Docent dr. J. Cholewa" je neprofitno, neinstitucionalno in nestrankarsko zdrućenje posameznikov, ustanov in organizacij, ki ćelijo materialno spodbujati in poglabljati raziskovalno dejavnost v onkologiji. Dunajska 106 1000 Ljubljana ĆR: 02033-0017879431 Zadeli smo pravo tar;o Izredno u;inkovito zdravljenje prvega reda pri nedrobnoceli;nem plju;nem raku z mutacijo EGFR Iressa je prva in edina tar;na monoterapija, ki dokazano podaljša preživetje brez napredovanja bolezni v primerjavi z dvojno kemoterapijo kot zdravljenje prvega reda pri bolnikih z napredovalim nedrobnoceli;nim plju;nim rakom z mutacijo EGFR. 1 IRESSA® (GEFITINIB) 1. Povzetek glavih zna;ilnosti zdravila Iressa (gefitinib). Junij 2009. SKRAJŠAN POVZETEK GLAVNIH ZNA:ILNOSTI ZDRAVILA Sestava' Filmsko obložene tablete vsebujejo 250 mg gefitiniba. Indikacije' zdravljenje odraslih bolnikov z lokalno napredovalim ali metastatskim nedrobnoceli;nim plju;nim rakom z aktivacijskimi mutacijami EGFR-TK Odmerjanje in na;in uporabe' Zdravljenje z gefitinibom mora uvesti in nadzorovati zdravnik, ki ima izkušnje z uporabo zdravil proti raku. Priporo;eno odmerjanje zdravila IRESSA je ena 250-mg tableta enkrat na dan. Tableto je mogo;e vzeti s hrano ali brez nje, vsak dan ob približno istem ;asu. Kontraindikacije' preob;utljivost za zdravilno u;inkovino ali katerokoli pomožno snov, dojenje Opozorila in previdnostni ukrepi' Pri 1,3 % bolnikov, ki so dobivali gefitinib, so opažali intersticijsko bolezen plju; (IBP). Ta se lahko pojavi akutno in je bila v nekaterih primerih smrtna. :e se bolniku poslabšajo dihalni simptomi, npr. dispneja, kašelj in zvišana telesna temperatura, morate zdravljenje z zdravilom IRESSA prekiniti in bolnika takoj preiskati. ;e je potrjena IBP, morate terapijo z zdravilom IRESSA kon;ati in bolnika ustrezno zdraviti. ;eprav so bile nepravilnosti testov jetrnih funkcij pogoste, so jih redko zabeležili kot hepatitis. Zato so priporo;ljive redne kontrole delovanja jeter. V primeru blagih do zmernih sprememb v delovanju jeter je treba zdravilo IRESSA uporabljati previdno. :e so spremembe hude, pride v poštev prekinitev zdravljenja. Zdravilo IRESSA vsebuje laktozo. Bolniki z redko dedno intoleranco za galaktozo, laponsko obliko zmanjšane aktivnosti laktaze ali malabsorpcijo glukoze/galaktoze ne smejo jemati tega zdravila. Bolnikom naro;ite, da morajo takoj poiskati zdravniško pomo;, ;e se jim pojavijo kakršnikoli o;esni simptomi, huda ali dolgotrajna driska, navzea, bruhanje ali anoreksija, ker lahko vse te posredno povzro;ijo dehidracijo. Medsebojno delovanje zdravile' Induktorji CYP3A4 lahko pove;ajo presnovo gefitiniba in zmanjšajo njegovo koncentracijo v plazmi. Zato lahko so;asna uporaba induktorjev CYP3A4 (npr. fenitoina, karbamazepina, rifampicina, barbituratov ali zeliš;nih pripravkov, ki vsebujejo šentjanževko\Hypericum perforatum) zmanjša u;inkovitost zdravljenja in se ji je treba izogniti. Pri posameznih bolnikih, ki imajo genotip slabih metabolizatorjev s CYP2D6, lahko zdravljenje z mo;nim zaviralcem CYP3A4 pove;a koncentracijo gefitiniba v plazmi. Na za;etku zdravljenja z zaviralcem CYP3A4 je treba bolnike natan;no kontrolirati glede neželenih u;inkov gefitiniba. Pri nekaterih bolnikih, ki so jemali varfarin skupaj z gefitinibom, so se pojavili zvišanje internacionalnega normaliziranega razmerja (INR) in\ali krvavitve. Bolnike, ki so;asno jemljejo varfarin in gefitinib, morate redno kontrolirati glede sprememb protrombinskega ;asa (P:) ali INR. Zdravila, ki ob;utno in dolgotrajno zvišajo pH v želodcu npr. zaviralci protonske ;rpalke in antagonisti H2, lahko zmanjšajo biološko uporabnost gefitiniba in njegovo koncentracijo v plazmi in tako zmanjšajo u;inkovitost. Redno jemanje antacidov, uporabljenih blizu ;asa jemanja zdravila IRESSA, ima lahko podoben u;inek. Neželeni u;inki' V kumulativnem naboru podatkov klini;nih preskušanj III. faze so bili najpogosteje opisani neželeni u;inki, ki so se pojavili pri ve; kot 20 % bolnikov, driska in kožne reakcije (vklju;no z izpuš;ajem, aknami, suho kožo in srbenjem). Neželeni u;inki se ponavadi pojavijo prvi mesec zdravljenja in so praviloma reverzibilni. Ostali pogostejši neželeni u;inki so' anoreksija, konjunktivitis, blefaritis in suho oko, krvavitev, npr. epistaksa in hematurija, intersticijska bolezen plju; (1,3 %), navzea, bruhanje, stomatitis, dehidracija, suha usta, nepravilnosti testov jetrnih funkcij, bolezni nohtov, alopecija, asimptomati;no laboratorijsko zvišanje kreatinina v krvi, proteinurija, astenija, pireksija. Vrsta in vsebina ovojnine' škatla s 30 tabletami po 250 mg gefitiniba Na;in izdajanja zdravila' samo na recept Datum priprave besedila' junij 2009 Imetnik dovoljenja za promet' AstraZeneca AB, S-151 85, Sodertalje, Švedska Pred predpisovanjem, prosimo, preberite celoten povzetek glavnih zna;ilnosti zdravila. Dodatne informacije so na voljo pri' AstraZeneca UK Limited, Podružnica v Sloveniji, Verovškova 55, 1000 Ljubljana, telefon' 01\51 35 600. Erbitux 5 mg/ml raztopina za infundiranje (skrajšana navodila za uporabo) Cetuksimab je monoklonsko IgG1 protitelo, usmerjeno proti receptorju za epidermalni rastni faktor (EGFR). Terapevtske indikacije: Zdravilo Erbitux je indicirano za zdravljenje bolnikov z metastatskim kolorektalnim rakom in nemutiranim tipom KRAS; v kombinaciji s kemoterapijo in kot samostojno zdravilo pri bolnikih, pri katerih zdravljenje z oksaliplatinom in irinotekanom ni bilo uspešno. Zdravilo Erbitux je indicirano za zdravljenje bolnikov z rakom skvamoznih celic glave in vratu; v kombinaciji z radioterapijo za lokalno napredovalo bolezen in v kombinaciji s kemoterapijo na osnovi platine za ponavljajoco se in/ali metastatsko bolezen. Odmerjanje in nacin uporabe: Zdravilo Erbitux pri vseh indikacijah infundirajte enkrat na teden. Zacetni odmerek je 400 mg cetuksimaba na m2 telesne površine. Vsi naslednji tedenski odmerki so vsak po 250 mg/m2. Kontraindikacije: Zdravilo Erbitux je kontraindicirano pri bolnikih z znano hudo preobcutljivostno reakcijo (3. ali 4. stopnje) na cetuksimab. Posebna opozorila in previdnostni ukrepi: Ce pri bolniku nastopi blaga ali zmerna reakcija, povezana z infundiranjem, lahko zmanjšate hitrost infundiranja. Priporocljivo je, da ostane hitrost infundiranja na nižji vrednosti tudi pri vseh naslednjih infuzijah. Ce se pri bolniku pojavi huda kožna reakcija (= 3. stopnje po kriterijih US National Cancer Institute, Common Toxicity Criteria; NCI-CTC), morate prekiniti terapijo s cetuksimabom. Z zdravljenjem smete nadaljevati le, ce se je reakcija pomirila do 2. stopnje. Priporoca se dolocanje koncentracije elektrolitov v serumu pred zdravljenjem in periodicno med zdravljenjem s cetuksimabom. Po potrebi se priporoca nadomešcanje elektrolitov. Posebna previdnost je potrebna pri oslabljenih bolnikih in pri tistih z obstojeco srcno-pljucno boleznijo. Neželeni ucinki: Zelo pogosti (= 1/10): dispneja, blago do zmerno povecanje jetrnih encimov, kožne reakcije, blage ali zmerne reakcije povezane z infundiranjem, blag do zmeren mukozitis. Pogosti (= 1/100, < 1/10): konjunktivitis, hude reakcije povezane z infundiranjem. Pogostost ni znana: Opazili so progresivno zniževanje nivoja magnezija v serumu, ki pri nekaterih bolnikih povzroca hudo hipomagneziemijo. Glede na resnost so opazili tudi druge elektrolitske motnje, vecinoma hipokalciemijo ali hipokaliemijo. Posebna navodila za shranjevanje: Shranjujte v hladilniku (2 °C -8 °C). Ne zamrzujte. Vrsta ovojnine in vsebina: 1 viala po 20 ml ali 100 ml. Imetnik dovoljenja za promet: Merck KGaA, 64271 Darmstadt, Nemcija. Podrobne informacije o zdravilu so objavljene na spletni strani Evropske agencije za zdravila PM-ONC-02/09/10.04.2009 (EMEA) http://www.emea.europa.eu. Dodatne informacije so vam na voljo pri: Merck d.o.o., Dunajska cesta 119, 1000 Ljubljana, tel.: 01 560 3810, faks: 01 560 3831, el. pošta: info@merck.si www.oncology.merck.de Povzetek glavnih znacilnosti zdravila Ime zdravila: Temodal 20 mg, 100 mg, 140mg, 180 mg, 250 mg, Temodal 2,5 mg/ml prašek za raztopino za infundiranje Kakovostna in kolicinska sestava: Vsaka kapsula zdravila Temodal vsebuje 20 mg, 100 mg, 140 mg, 180 mg ali 250 mg temozolomida. Ena viala vsebuje 100 mg temozolomida Po rekonstituciji 1 ml raztopine za infundiranje vsebuje 2,5 mg temozolomida. Pomožna snov: Ena viala vsebuje 2,4 mmol natrija. Terapevtske indikacije: Zdravilo Temodal 2,5 mg/ml je indicirano za zdravljenje: odraslih bolnikov z novo diagnosticiranim multiformnim glioblastomom, socasno z radioterapijo (RT) in pozneje kot monoterapija in otrok, starih 3 leta in vec, mladostnikov in odraslih bolnikov z malignimi gliomi, npr. multiformnimi glioblastomi ali anaplasticnimi astrocitomi, ki se po standardnem zdravljenju ponovijo ali napredujejo. Odmerjanje in nacin uporabe: Zdravilo Temodal 2,5 mg/ml smejo predpisati le zdravniki, ki imajo izkušnje z zdravljenjem možganskih tumorjev. Odrasli bolniki z novo diagnosticiranim multiformnim glioblastomom Zdravilo Temodal 2,5 mg/ml se uporablja v kombinaciji z žarišcno radioterapijo (faza socasne terapije), temu pa sledi do 6 ciklov monoterapije (monoterapijska faza) z temozolomidom (TMZ). Faza socasne terapije TMZ naj bolnik jemlje v odmerku 75 mg/m2 na dan 42 dni, socasno z žarišcno radioterapijo (60 Gy, danih v 30 delnih odmerkih). Zmanjševanje odmerka ni priporoceno, vendar se boste vsak teden odlocili o morebitni odložitvi jemanja TMZ ali njegovi ukinitvi na podlagi kriterijev hematološke in nehematološke toksicnosti. TMZ lahko bolnik jemlje ves cas 42-dnevnega obdobja socasne terapije (do 49 dni), ce so izpolnjeni vsi od naslednjih pogojev: • absolutno število nevtrofilcev (ANC – Absolute Neutrophil Count) = 1,5 x 109/l; • število trombocitov = 100 x 109/l; • skupna merila toksicnosti (SMT) za nehematološko toksicnost = 1. stopnje (z izjemo alopecije, navzee in bruhanja). Med zdravljenjem morate pri bolniku enkrat na teden pregledati celotno krvno sliko. Faza monoterapije Štiri tedne po zakljucku faze socasnega zdravljenja s TMZ in RT naj bolnik jemlje TMZ do 6 ciklov monoterapije. V 1. ciklu (monoterapije) je odmerek zdravila 150 mg/m2 enkrat na dan 5 dni, temu pa naj sledi 23 dni brez terapije. Na zacetku 2. cikla odmerek povecajte na 200 mg/m2, ce je SMT za nehematološko toksicnost za 1. cikel stopnje = 2 (z izjemo alopecije, slabosti in bruhanja), absolutno število nevtrofilcev (ANC) = 1,5 x 109/l in število trombocitov = 100 x 109/l. Ce odmerka niste povecali v 2. ciklu, ga v naslednjih ciklih ne smete povecevati. Ko pa odmerek enkrat povecate, naj ostane na ravni 200 mg/m2 na dan v prvih 5 dneh vsakega naslednjega cikla, razen ce nastopi toksicnost. Zmanjšanje odmerka in ukinitev zdravila med fazo monoterapije opravite, kot je opisano v preglednicah 2 in 3. Med zdravljenjem morate 22. dan pregledati celotno krvno sliko (21 dni po prvem odmerku TMZ). Odrasli in pediatricni bolniki, stari 3 leta ali vec, s ponavljajocim se ali napredujocim malignim gliomom:Posamezen cikel zdravljenja traja 28 dni. Bolniki, ki še niso bili zdravljeni s kemoterapijo, naj jemljejo TMZ v odmerku 200 mg/m2 enkrat na dan prvih 5 dni, temu pa naj sledi 23-dnevni premor (skupaj 28 dni). Pri bolnikih, ki so že bili zdravljeni s kemoterapijo, je zacetni odmerek 150 mg/m2 enkrat na dan, v drugem ciklu pa se poveca na 200 mg/m2 enkrat na dan 5 dni, ce ni bilo hematoloških toksicnih ucinkov. Kontraindikacije: Preobcutljivost za zdravilno ucinkovino ali katerokoli pomožno snov. Preobcutljivost za dakarbazin (DTIC). Posebna opozorila in previdnostni ukrepi: Pljucnica, ki jo povzroca Pneumocystis carinii Pilotno preskušanje podaljšane 42-dnevne sheme zdravljenja je pokazalo, da pri bolnikih, ki so socasno prejemali TMZ in RT, obstaja še posebej veliko tveganje za nastanek pljucnice zaradi okužbe s Pneumocystis carinii (PCP). Malignosti Zelo redko so porocali tudi o primerih mielodisplasticnega sindroma in sekundarnih malignostih, vkljucno z mieloidno levkemijo. Antiemeticno zdravljenje Navzea in bruhanje sta pogosto povezana z zdravljenjem s TMZ. Antiemeticno zdravljenje se lahko da pred uporabo TMZ ali po njej. Odrasli bolniki z novo diagnosticiranim multiformnim glioblastomom Antiemeticna profilaksa je priporocljiva pred zacetnim odmerkom socasne faze in je mocno priporocljiva med fazo monoterapije. Ponavljajoci se ali napredujoci maligni gliom Pri bolnikih, ki so mocno bruhali (stopnja 3 ali 4) v prejšnjih ciklih zdravljenja, je potrebno antiemeticno zdravljenje. Laboratorijske vrednosti Pred jemanjem zdravila morata biti izpolnjena naslednja pogoja za laboratorijske izvide: ANC = 1,5 x 109/l in število trombocitov = 100 x 109/l. Na 22. dan (21 dni po prvem odmerku) ali v roku 48 ur od navedenega dne, morate pregledati celotno krvno sliko in jo nato spremljati vsak teden, dokler ni ANC > 1,5 x 109/l in število trombocitov > 100 x 109/l. Ce med katerimkoli ciklom ANC pade na < 1,0 x 109/l ali število trombocitov na < 50 x 109/l, morate odmerek zdravila v naslednjem ciklu zmanjšati za eno stopnjo (glejte poglavje 4.2). Stopnje odmerka so 100 mg/m2, 150 mg/m2 in 200 mg/m2. Najmanjši priporoceni odmerek je 100 mg/m2. Pediatricna uporaba Klinicnih izkušenj z uporabo TMZ pri otrocih, mlajših od 3 let, ni. Izkušnje z uporabo tega zdravila pri starejših otrocih in mladostnikih so zelo omejene. Starejši bolniki (stari > 70 let) Videti je, da je pri starejših bolnikih tveganje za nevtropenijo ali trombocitopenijo vecje, kot pri mlajših. Zato je pri uporabi zdravila TMZ pri starejših bolnikih potrebna posebna previdnost. Moški bolniki Moškim, ki se zdravijo s TMZ je treba svetovati, naj ne zaplodijo otroka še šest mesecev po prejetem zadnjem odmerku in naj se pred zdravljenjem posvetujejo o možnostih za shranitev zmrznjene sperme. Natrij To zdravilo vsebuje 2,4 mmol natrija na vialo. To je treba upoštevati pri bolnikih na nadzorovani dieti z malo natrija. Medsebojno delovanje z drugimi zdravili in druge oblike interakcij: Študije medsebojnega delovanja so izvedli le pri odraslih. V loceni študiji 1. faze, socasna uporaba TMZ in ranitidina ni povzrocila spremembe obsega absorpcije temozolomida ali izpostavljenosti njegovem aktivnem presnovku monometiltriazenoimidazol karboksamidu (MTIK). Analiza populacijske farmakokinetike v preskušanjih 2. faze je pokazala, da socasna uporaba deksametazona, proklorperazina, fenitoina, karbamazepina, ondansetrona, antagonistov receptorjev H2 ali fenobarbitala ne spremeni ocistka TMZ. Socasno jemanje z valprojsko kislino je bilo povezano z majhnim, a statisticno pomembnim zmanjšanjem ocistka TMZ. Študij za dolocitev ucinka TMZ na presnovo ali izlocanje drugih zdravil niso izvedli. Ker pa se TMZ ne presnavlja v jetrih in se na beljakovine veže le v majhni meri, je malo verjetno, da bi vplival na farmakokinetiko drugih zdravil. Uporaba TMZ v kombinaciji z drugimi mielosupresivnimi ucinkovinami lahko poveca verjetnost mielosupresije. Neželeni ucinki: Pri bolnikih, ki se zdravijo s TMZ v kombinaciji z RT ali monoterapijo po RT zaradi novo diagnosticiranega multiformnega glioblastoma ali z monoterapijo pri bolnikih s ponavljajocim se ali napredujocim gliomom, so bili zelo pogosti neželeni ucinki podobni; slabost, bruhanje, zaprtje, neješcnost, glavobol in utrujenost. Pri bolnikih z novo diagnosticiranim glioblastomom multiforme na monoterapiji so zelo pogosto porocali o konvulzijah, medtem ko je bil izpušcaj opisan zelo pogosto pri bolnikih z novo diagnosticiranim multiformnim glioblastomom, ki so prejemali TMZ socasno z RT, ter pri tistih, ki so zdravilo prejemali v obliki monoterapije, pogosto pa pri tistih s ponavljajocim se gliomom. Pri obeh indikacijah so o vecini hematoloških neželenih reakcij porocali pogosto ali zelo pogosto. Imetnik dovoljenja za promet: Schering-Plough Europe, Rue de Stalle 73, Bruselj Belgija Nacin in režim izdaje zdravila: Zdravilo Temodal 20 mg, 100 mg, 140mg, 180 mg, 250 mg se izdaja na recept (Rp/Spec), Temodal 2,5 mg/ml prašek za raztopino za infundiranje pa je namenjeno uporabi samo v bolnišnicah (H). Datum priprave informacije: februar 2010 Literatura: 1 Povzetek temeljnih znacilnosti zdravila Temodal 2 Stupp R, et. al. Effects of radiotherapy with concomitant and adjuvant temozolomide versus radiotherapy alone on survival in glioblastoma in a 0233 Nova, intravenska oblika zdravila Temodal, za zdravljenje bolnikov z novo diagnosticiranim glioblastomom multiforme in bolnikov s ponavljajocim se ali napredujocim malignim gliomom.1 Resnicni napredek Pomembno izboljšanje preživetja potrjeno tudi ob daljšem spremljanju bolnikov.2 Dunajska 22, 1000 Ljubljana tel: 01 300 10 70 fax: 01 300 10 80 SKRAJ[AN POVZETEK GLAVNIH ZNA^ILNOSTI ZDRAVILA Samozastrokovnojavnost. Ime zdravila: Tarceva25mg/100mg/150mgfilmskoobložene Kakovostna in koli~inska sestava: Enafilmskoobloženatabletavsebuje25mg,100mgali150mgerlotiniba(voblikierlotinibijevegaklorida).Terapevtske indikacije:Nedrobnoceli~nirakplju~:ZdraviloTarcevajeindiciranozasamostojnovzdrževalnozdravljenjebolnikovzlokalnonapredovalimalimetastatskimnedrobnoceli~nimrakomplju~sstabilnoboleznijopo4ciklihstandardnekemoterapijenaosnoviplatinevprvilinijizdravljenja.ZdraviloTarcevajeindiciranotudizazdravljenjebolnikovzlokalnonapredovalimalimetastatskimnedrobnoceli~nimrakomplju~poneuspehuvsajenepredhodnekemoterapije.PripredpisovanjuzdravilaTarcevajetrebaupo{tevatidejavnike,povezanespodalj{animpreživetjem.Koristnegavplivanapodalj{anjepreživetjaalidrugihklini~nopomembnihu~inkovzdravljenjanisodokazalipribolnikihzEGFR-negativnimitumorji.Raktrebu{neslinavke:ZdraviloTarcevajevkombinacijizgemcitabinomindiciranozazdravljenjebolnikovzmetastatskimrakomtrebu{neslinavke.PripredpisovanjuzdravilaTarcevajetrebaupo{tevatidejavnike,povezanespodalj{animpreživetjem.Koristnegavplivanapodalj{anjepreživetjanisodokazalizabolnikezlokalnonapredovaloboleznijo.Odmerjanje in na~in uporabe: ZdravljenjezzdravilomTarcevamoranadzorovatizdravnikzizku{njamiprizdravljenjuraka.ZdraviloTarcevavzamemonajmanjenouropredzaužitjemhranealidveuripotem.Kadarjepotrebnoodmerekprilagoditi,gazmanj{ujemovkorakihpo50mg.Priso~asnemjemanjusubstratovinmodulatorjevCYP3A4bomordapotrebnaprilagoditevodmerka.PridajanjuzdravilaTarcevabolnikomzjetrnookvarojepotrebnaprevidnost.^esepojavijohudineželeniu~inki,pridevpo{tevzmanj{anjeodmerkaaliprekinitevzdravljenjazzdravilomTarceva.UporabazdravilaTarcevapribolnikihshudojetrnoaliledvi~nookvaroterpriotrocihnipriporo~ljiva.Bolnikomkadilcemjetrebasvetovati,najprenehajokaditi,sajsoplazemskekoncentracijeerlotinibaprikadilcihmanj{ekotprinekadilcih.Nedrobnoceli~nirakplju~:Priporo~enidnevniodmerekzdravilaTarcevaje150mg.Raktrebu{neslinavke:Priporo~enidnevniodmerekzdravilaTarcevaje100mg,vkombinacijizgemcitabinom.Pribolnikih,prikaterihsekožniizpu{~ajvprvih4do8tednihzdravljenjanepojavi,jetrebaponovnopretehtatinadaljnjezdravljenjezzdravilomTarceva.Kontraindikacije: Hudapreob~utljivostzaerlotinibalikaterokolipomožnosnov.Posebna opozorila in previdnostni ukrepi: Mo~niinduktorjiCYP3A4lahkozmanj{ajou~inkovitosterlotiniba,mo~nizaviralciCYP3A4palahkopove~ajotoksi~nost.So~asnemuzdravljenjustemizdravilisejetrebaizogibati.Bolnikom,kikadijo,jetrebasvetovati,najprenehajokaditi,sajsoplazemskekoncentracijeerlotinibaprikadilcihzmanj{anevprimerjavisplazemskimikoncentracijamiprinekadilcih.Verjetnoje,dajevelikostzmanj{anjaklini~nopomembna.Pribolnikih,prikaterihseakutnopojavijonoviin/aliposlab{ajonepojasnjeniplju~nisimptomi,kotsodispneja,ka{eljinvro~ina,jezdravljenjezzdravilomTarcevatrebaprekiniti,doklerniznanadiagnoza.Bolnike,kiseso~asnozdravijozerlotinibomingemcitabinom,jetrebaskrbnospremljatizaradimožnostipojavatoksi~nosti,podobniintersticijskiplju~nibolezni.^ejeugotovljenaintersticijskaplju~nabolezen,zdraviloTarcevaukinemoinuvedemoustreznozdravljenje.Pripribližnopolovicibolnikov,kisosezdravilizzdravilomTarceva,sejepojaviladriska.Zmernodohudodriskozdravimozloperamidom.Vnekaterihprimerihbomordapotrebnozmanj{anjeodmerka.Vprimeruhudealidolgotrajnedriske,navzeje,anoreksijealibruhanja,povezanihzdehidracijo,jezdravljenjezzdravilomTarcevatrebaprekinitiindehidracijoustreznozdraviti.Ohipokaliemijiinledvi~niodpovedisoporo~aliredko.Posebnopribolnikihzdejavnikitveganja(so~asnojemanjedrugihzdravil,simptomi,boleznialidrugidejavniki,vklju~nozvisokostarostjo)moramo,~ejedriskahudaalidolgotrajnaoziromavodivdehidracijo,zdravljenjezzdravilomTarcevaprekinitiinbolnikomzagotovitiintenzivnointravenskorehidracijo.Dodatnojetrebapribolnikihsprisotnimtveganjemzarazvojdehidracijespremljatiledvi~nodelovanjeinserumskeelektrolite,vklju~noskalijem.PriuporabizdravilaTarcevasoporo~alioredkihprimerihjetrneodpovedi.Knjenemunastankujelahkopripomoglapredhodnoobstoje~ajetrnabolezenaliso~asnojemanjehepatotoksi~nihzdravil.Pritehbolnikihjetrebazatopremislitiorednemspremljanjujetrnegadelovanja.DajanjezdravilaTarcevajetrebaprekiniti,~esospremembejetrnegadelovanjahude.Bolniki,kiprejemajozdraviloTarceva,imajove~jetveganjezarazvojperforacijvprebavilih,kisojihopaziliob~asno.Pribolnikih,kiso~asnoprejemajozdravila,kizavirajoangiogenezo,kortikosteroide,nesteroidnaprotivnetnazdravila(NSAID)in/alikemoterapijonaosnovitaksanov,alisovpreteklostiimelipepti~niulkusalidivertikularnobolezen,jetveganjeve~je.^epridedotega,jetrebazdravljenjezzdravilomTarcevadokon~noukiniti.Poro~alisooprimerihkožnihboleznizmehurjiinlu{~enjemkože,vklju~nozzeloredkimiprimeri,kisonakazovalinaStevens-Johnsonovsindrom/toksi~noepidermalnonekrolizoinsobilivnekaterihprimerihsmrtni.ZdravljenjezzdravilomTarcevajetrebaprekinitialiukiniti,~esepribolnikupojavijohudeoblikemehurjevalilu{~enjakože.Zeloredkosoporo~alioprimerihperforacijealiulceracijeroženice;opazilisotudidrugeo~esnebolezni. ZdravljenjezzdravilomTarcevajetrebaprekinitialiukiniti,~esepribolnikihpojavijoakutneo~esnebolezni,kotjebole~inavo~eh,alisele-teposlab{ajo.Tabletevsebujejolaktozoinjihnesmemodajatibolnikomzredkimidednimistanji:intolerancozagalaktozo,laponskooblikozmanj{aneaktivnostilaktazealimalabsorpcijoglukoze/galaktoze. Medsebojno delovanje z drugimi zdravili in druge oblike interakcij: Erlotinibsepriljudehpresnavljavjetrihzjetrnimicitokromi,primarnosCYP3A4invmanj{imerisCYP1A2.PresnovaerlotinibazunajjeterpotekasCYP3A4v~revesju,CYP1A1vplju~ihinCYP1B1vtumorskihtkivih.Zzdravilnimiu~inkovinami,kisepresnavljajostemiencimi,jihzavirajoalipasonjihoviinduktorji,lahkopridedointerakcij.Erlotinibjesrednjemo~anzaviralecCYP3A4inCYP2C8,kottudimo~anzaviralecglukuronidacijezUGT1A1in vitro.Prikombinacijiciprofloksacinaalimo~negazaviralca CYP1A2 (npr.fluvoksamina)zerlotinibomjepotrebnaprevidnost.Vprimerupojavaneželenihdogodkov,povezanihzerlotinibom,lahkoodmerekerlotinibazmanj{amo.Predhodnoaliso~asnozdravljenjezzdravilomTarcevanispremeniloo~istkaprototipovsubstratov CYP3A4,midazolamaineritromicina.Inhibicijaglukoronidacijelahkopovzro~iinterakcijezzdravili,kisosubstrati UGT1A1 inseizlo~ajosamopotejpoti.Mo~nizaviralci aktivnosti CYP3A4 zmanj{ajopresnovoerlotinibainzve~ajokoncentracijeerlotinibavplazmi.Priso~asnemjemanjuerlotinibainmo~nihzaviralcevCYP3A4jezatopotrebnaprevidnost.^ejetreba,odmerekerlotinibazmanj{amo,{eposebnopripojavutoksi~nosti.Mo~nispodbujevalci aktivnosti CYP3A4 zve~ajopresnovoerlotinibainpomembnozmanj{ajoplazemskekoncentracijeerlotiniba.So~asnemudajanjuzdravilaTarcevaininduktorjevCYP3A4sejetrebaizogibati.Pribolnikih,kipotrebujejoso~asnozdravljenjezzdravilomTarcevainmo~niminduktorjemCYP3A4,jetrebapremislitiopove~anjuodmerkado300mgobskrbnemspremljanjunjihovevarnosti.Zmanj{anaizpostavljenostselahkopojavitudizdrugimiinduktorji,kotsofenitoin,karbamazepin,barbituratiali{entjanževka.^etezdravilneu~inkovinekombiniramozerlotinibom,jepotrebnaprevidnost.Kadarjemogo~e,jetrebarazmislitiodrugihna~inihzdravljenja,kinevklju~ujejomo~negaspodbujanjaaktivnostiCYP3A4.Bolnikom,kijemljejovarfarinalidrugekumarinske antikoagulante,jetrebarednokontroliratiprotrombinski~asaliINR.So~asnauporabazaviralcev P-glikoproteina,kotstaciklosporininverapamil,lahkovodivspremenjenoporazdelitevin/alispremenjenoizlo~anjeerlotiniba.Zaerlotinibjezna~ilnozmanj{anjetopnostipripHnad5.Zdravila, ki spremenijo pH v zgornjem delu prebavil,lahkospremenijotopnosterlotinibainposledi~nonjegovobiolo{kouporabnost.U~inkaantacidovnaabsorpcijoerlotinibanisoprou~evali,vendarjetalahkozmanj{ana,karvodivnižjeplazemskekoncentracije.Kombinacijierlotinibainzaviralcaprotonske~rpalkesejetrebaizogibati.^emenimo,dajeuporabaantacidovmedzdravljenjemzzdravilomTarcevapotrebna,jihjetrebajematinajmanj4urepredali2uripodnevnemodmerkuzdravilaTarceva.^erazmi{ljamoouporabiranitidina,moramozdravilijematilo~eno:zdraviloTarcevajetrebavzetinajmanj2uripredali10urpoodmerkuranitidina.V{tudijifazeIbnibilopomembnihu~inkovgemcitabina nafarmakokinetikoerlotiniba,pravtakonibilopomembnihu~inkoverlotinibanafarmakokinetikogemcitabina.Erlotinibpove~akoncentracijoplatine.Pomembnihu~inkovkarboplatina alipaklitakselanafarmakokinetikoerlotinibanibilo.Kapecitabin lahkopove~akoncentracijoerlotiniba.Pomembnihu~inkoverlotinibanafarmakokinetikokapecitabinanibilo.Neželeni u~inki: Zelo pogosti neželeni u~inki sokožniizpu{~ajindriska,kottudiutrujenost,anoreksija,dispneja,ka{elj,okužba,navzeja,bruhanje,stomatitis,bole~inavtrebuhu,pruritus,suhakoža,suhikeratokonjunktivitis,konjunktivitis,zmanj{anjetelesnemase,depresija,glavobol,nevropatija,dispepsija,flatulenca,alopecija,okorelost,pireksija.Pogosti neželeni u~inki sogastrointestinalnekrvavitve,krvaviteviznosu,nenormalnostitestovjetrnefunkcije,keratitis,zanohtnica.Redko soporo~aliojetrniodpovedi.Ob~asno pa operforacijahvprebavilih,pora{~enostimo{kegatipapriženskah,spremembahobrvi,krhkihnohtih,odstopanjunohtovodkože,blagihreakcijahnakoži(npr.hiperpigmentacija),spremembahtrepalnic,resniintersticijskiplju~nibolezni,vklju~nossmrtnimiprimeri.Zelo redko soporo~alioprimerih,kisonakazovalinaStevens-Johnsonovsindrom/toksi~noepidermalnonekrolizoinsobilivnekaterihprimerihsmrtni,teroulceracijahinperforacijahroženice.Režim izdaje zdravila: H/Rp.Imetnik dovoljenja za promet: RocheRegistrationLimited,6FalconWay,ShirePark,WelwynGardenCity,AL71TW,VelikaBritanija.Verzija: 1.0/10.Informacija pripravljena:maj2010. DODATNEINFORMACIJESONAVOLJOPRI:Rochefarmacevtskadružbad.o.o.Vodovodnacesta109,1000Ljubljana.Povzetekglavnihzna~ilnostizdravilajedosegljivnawww.roche.si. 033-10-TAR ^AS ZA ŽIVLJENJE. DOKAZANO PODALJŠA PREŽIVETJE PRI BOLNIKIH: zlokalnonapredovalimalimetastatskim zmetastatskimrakomtrebu{neslinavke1 1 Povzetek glavnih znacilnosti zdravila TARCEVA, www.ema.europa.eu Lek farmacevtska družba d.d., Verovškova 57, 1526 Ljubljana, Slovenija, www.lek.si SKRAJŠAN POVZETEK GLAVNIH ZNACILNOSTI ZDRAVILA Epufen 12,5, 25, 50, 100 in 150 mikrogramov/uro transdermalni obliži SESTAVA: 1 transdermal­ni obliž vsebuje 2,89 mg, 5,78 mg 11,56 mg, 23,12 mg ali 34,65 mg fentanila. TERAPEVTSKE INDIKACIJE: Huda kronicna bolecina, ki se lahko ustrezno zdravi le z opioidnimi analgetiki. ODMERJANJE IN NACIN UPORABE: Odmerjanje je tre­ba individualno prilagoditi ter ga po vsaki uporabi redno oceniti. Izbira zacetnega odmerka: velikost odmerka fentanila je odvisna od predhodne uporabe opioidov, kjer se upošteva možnost pojava tolerance, socasnega zdravljenja, bolnikovega splošnega zdravstvenega stanja in stopnje resnosti obolenja. Pri bolnikih, ki pred tem niso dobivali mocnih opioidov, zacetni odmerek ne sme preseci 12,5-25 mi-krogramov na uro. Zamenjava opioidnega zdravljenja: pri zamenjavi peroralnih ali parenteralnih opioidov s fentanilom je treba zacetni odmerek izracunati na osnovi kolicine analgetika, ki je bila potrebna v zadnjih 24 urah, jo pretvoriti v odgovarjajoci odmerek morfina s pomocjo razpredelnice in nato preracunati ustrezen odmerek fentanila, spet s pomocjo razpredelnice (glejte SmPC). Prvih 12 ur po prehodu na transdermalni obliž Epufen bolnik še vedno dobiva predhodni analgetik v enakem odmerku kot prej; v naslednjih 12 urah se ta analgetik daje po potrebi. Titracija odmerka in vzdrževalno zdravljenje: obliž je treba zamenjati vsakih 72 ur. Odme­rek je treba titrirati individualno, dokler ni dosežen analgeticni ucinek. Odmerek 12,5 mikrogramov/uro je primeren za titriranje odmerka v manjšem odmernem ob-mocju. Ce analgezija na koncu zacetnega obdobja nošenja obliža ni zadostna, se lahko odmerek po 3 dneh zveca. Možno je, da bodo bolniki potrebovali obcasne dodatne odmerke kratko delujocih analgetikov (npr. morfina) za prekinitev bolecine. Sprememba ali prekinitev zdravljenja: vsaka zamenjava z drugim opioidom mora potekati postopoma, z majhnim zacetnim odmerkom in pocasnim zvecevanjem. Splošno veljavno pravilo je postopna ustavitev opioidne analgezije, da bi preprecili odtegnitvene simptome, kot so navzeja, bruhanje, diareja, anksioznost in mišicni tremor. Uporaba pri starejših bolnikih: starejše in oslabljene bolnike je treba skrb-no opazovati zaradi simptomov prevelikega odmerjanja ter odmerek po potrebi zmanjšati. Uporaba pri otrocih: transdermalni obliži Epufen se lahko uporabljajo le pri pediatricnih bolnikih (starih od 2 do 16 let), ki tolerirajo opioide in peroralno že dobivajo opioide v odmerku, enakovrednemu najmanj 30 mg morfina na dan. Bolnik mora prvih 12 ur po prehodu na Epufen še vedno dobivati predhodni anal-getik v enakem odmerku kot prej. V naslednjih 12 urah je treba ta analgetik dajati odvisno od klinicnih potreb. Titracija odmerka in vzdrževalno zdravljenje: ce je anal­geticni ucinek Epufena prešibak, je treba bolniku dodati morfin ali drugi opioid s kratkim delovanjem. Odvisno od dodatnih potreb po analgeziji in jakosti bolecine pri otroku se lahko uporabi vec obližev. Odmerek je treba prilagajati korakoma, po 12,5 mikrogramov/uro. Uporaba pri bolnikih z jetrno ali ledvicno okvaro: Zara-di možnosti pojava simptomov prevelikega odmerjanja je treba te bolnike skrbno spremljati in odmerek ustrezno zmanjšati. Uporaba pri bolnikih s povecano telesno temperaturo: Pri teh bolnikih bo morda treba prilagoditi odmerek. Nacin uporabe: transdermalni obliž Epufen je treba takoj po odprtju vrecke nalepiti na nerazdraženo, neobsevano kožo, na ravno površino prsnega koša, zgornjega dela hrbta ali nadlakti. Po odstranitvi zašcitne plasti je treba obliž trdno pritrditi na izbrano mesto in z dlanjo pritiskati približno 30 sekund, da se obliž popolnoma nalepi, še zlasti na robovih. Uporaba pri otrocih: pri mlajših otrocih je obliž priporocljivo nalepiti na zgornji del hrbta, ker je manjša verjetnost, da bi otrok odstranil obliž. Transdermalnega obliža se ne sme deliti, ker podatkov o tem ni na voljo. KONTRAINDIKACIJE: Preobcu­tljivost za zdravilno ucinkovino, hidrogenirano kolofonijo, sojo, arašide ali katerokoli pomožno snov. Akutna ali pooperativna bolecina, ko v kratkem casovnem obdobju ni možno titriranje odmerka in obstaja verjetnost za življenjsko ogrožajoco respirator-no depresijo. Huda okvara osrednjega živcnega sistema. POSEBNA OPOZORILA IN PREVIDNOSTNI UKREPI: Zaradi razpolovne dobe fentanila je treba bolnika v primeru pojava neželenega ucinka opazovati še 24 ur po odstranitvi obliža. Pri nekaterih bol­nikih, ki uporabljajo transdermalni obliž Epufen, se lahko pojavi respiratorna depre­sija. Epufen je treba previdno dajati: bolnikom s kronicno pljucno boleznijo, zvišanim intrakranialnim tlakom, možganskim tumorjem, boleznimi srca, jeter in ledvic, tistim z zvišano telesno temperaturo, pri starejših bolnikih in otrocih, bolnikih z miaste­nijo gravis. Odvisnost od zdravila: kot posledica ponavljajoce se uporabe se lahko razvijeta toleranca na ucinkovino ter psihicna in/ali fizicna odvisnost od nje. Ostali: lahko se pojavijo neepilepticne (mio)klonicne reakcije. MEDSEBOJNO DELOVANJE Z DRUGIMI ZDRAVILI IN DRUGE OBLIKE INTERAKCIJ: Derivati barbiturne kisline, opioidi, anksiolitiki in pomirjevala, hipnotiki, splošni anestetiki, fenotiazini, mišicni relaksanti, sedativni antihistaminiki in alkoholne pijace, zaviralci MAO, itrakonazol, ritonavir, ketokonazol, nekateri makrolidni antibiotiki, pentazocin, buprenorfin. VPLIV NA SPOSOBNOST VOŽNJE IN UPRAVLJANJA S STROJI: Zdravilo ima mocan vpliv na sposobnost vožnje in upravljanja s stroji. NEŽELENI UCINKI: Najbolj resen neželen ucinek fentanila je respiratorna depresija. Zelo pogosti (= 1/10): drema­vost, glavobol, navzeja, bruhanje, zaprtje, znojenje, srbenje, somnolenca. Pogosti (= 1/100 do < 1/10): kserostomija, dispepsija, reakcije na koži na mestu aplikacije, sedacija, zmedenost, depresija, tesnoba, živcna napetost, halucinacije, zmanjšan apetit. Obcasni (= 1/1000 do < 1/100): tahikardija, bradikardija, tremor, parestezija, motnje govora, dispneja, hipoventilacija, diareja, zastajanje urina, izpušcaj, rdecina, hipertenzija, hipotenzija, evforija, amnezija, nespecnost, vznemirljivost. Nekateri od naštetih neželenih ucinkov so lahko posledica osnovne bolezni ali drugih zdravljenj. Drugi neželeni ucinki: odpornost, fizicna in psihicna odvisnost se lahko razvijejo med dolgotrajno uporabo fentanila. Pri nekaterih bolnikih se lahko pojavijo odtegnitveni simptomi, ko zamenjajo prejšnje opiodne analgetike s transdermalnim obilžem s fentanilom ali po nenadni prekinitvi zdravljenja. NACIN IZDAJE: Samo na zdravni­ški recept. OPREMA: Škatle s 5 transdermalnimi obliži. IMETNIK DOVOLJENJA ZA PROMET: Lek farmacevtska družba, d.d., Verovškova 57, Ljubljana, Slovenija INFOR­MACIJA PRIPRAVLJENA: avgust 2009 Za podro~ja: • bioznanosti SYNGENE, INVITROGEN, BIOTEK diagnostike MINERVA, MEDAC, BIOTEK • gojenja celi~nih kultur INVITROGEN-GIBCO, TPP, SANYO • merjenja absorbance, fluorescence in luminiscence BIOTEK, SHIMADZU • pipetiranja BIOHIT in BIOTEK • laboratorijske opreme in instrumentov SANYO, SHIMADZU • ~iste vode za laboratorije ELGA LABWATER • HPLC in GC instrumentov, kolon, vial in filtrov PHENOMENEX, CHROMACOL/NATIONAL SCIENTIFIC, SHIMADZU Novartis Oncology prinaša spekter inovativnih zdravil, s katerimi poskuša spremeniti življenje bolnikov z rakavimi in hematološkimi obolenji. Ta vkljucuje zdravila kot so Glivec® (imatinib), Tasigna® (nilotinib), A. nitor® (everolimus), Zometa® (zoledronska kislina), Femara® (letrozol), Sandostatin® LAR® (oktreotid/i.m. injekcije) in Exjade® (deferasiroks). Novartis Oncology ima tudi obširen razvojni program, ki izkorišca najnovejša spoznanja molekularne genomike, razumskega nacrtovanja in tehnologij za odkrivnje novih ucinkovin. Novartis Pharma Services Inc. • Podružnica v Sloveniji • Tivolska cesta 30 • 1000 Ljubljana Samo za strokovno javnost. NV-JA-02/09-SI zdravnikom ali sfarmacevtom. Širok izbor jakosti zdravila EPREX®za zdravljenje anemije pri bolnikih z rakom: • EPREX® 10.000 i.e./1,0 ml • EPREX® 20.000 i.e./0,5 ml • EPREX® 30.000 i.e./0,75 ml • EPREX® 40.000 i.e./1,0 ml SKRAJŠANO NAVODILO ZA PREDPISOVANJE ZDRAVILA EPREX® EPREX® 1000 i.e./0,5 ml, EPREX® 2000 i.e./0,5 ml, EPREX® 3000 i.e./0,3 ml, EPREX® 4000 i.e./0,4 ml, EPREX® 5000 i.e./0,5 ml, EPREX® 6000 i.e./0,6 ml, EPREX® 8000 i.e./0,8 ml, EPREX® 10.000 i.e./1,0 ml, EPREX® 20.000 i.e./0,5 ml, EPREX® 30.000 i.e./0,75 ml, EPREX® 40.000 i.e./1,0 ml raztopina za injiciranje v napolnjenih injekcijskih brizgah in EPREX® 40.000 i.e./1,0 ml raztopina za injiciranje. Sestava: epoetin alfa, natrijev dihidrogenfosfat dihidrat, dinatrijev hidrogenfosfat dihidrat, natrijev klorid, polisorbat, glicin, voda za injekcije. Terapevtske indikacije: zdravljenje simptomatske anemije, ki je posledica kronicnega odpovedovanja ledvic pri odraslih in otrocih, zdravljenje anemije in zmanjšanje potreb po transfuziji pri odraslih bolnikih, pri katerih s kemoterapijo zdravimo solidne tumorje, maligni limfom ali multipli mielom, povecanje proizvodnje avtologne krvi pri bolnikih v programu samodarovanja krvi pred operacijo, zmanjšanje izpostavljenosti alogenim transfuzijam krvi pred vecjimi elektivnimi ortopedskimi kirurškimi posegi. Odmerjanje in nacin uporabe: Bolniki s kronicno ledvicno odpovedjo na hemodializi: Zdravilo injicirajte i.v. ali s.c., ciljna koncentracija Hb je 100-120 g/l pri odraslih in 95-110 g/l pri otrocih. Korekcijska faza: 50 i.e./kg 3 x tedensko. Odmerek prilagajamo postopno, z vsaj štiritedenskimi casovnimi presledki za 25 i.e./kg 3 x tedensko. Vzdrževalna faza: priporocen skupni tedenski odmerek je od 75 do 300 i.e./kg. Odrasli bolniki z zmanjšanim ledvicnim delovanjem, ki se še ne zdravijo z dializo: zacetni odmerek je 50 i.e./kg s.c. 3 x tedensko. Odmerek prilagajamo postopno, z vsaj štiritedenskimi casovnimi presledki za 25 i.e./kg 3 x tedensko. Vzdrževalni odmerek je od 17 do 33 i.e./kg 3 x tedensko, najvecji tedenski odmerek ne sme presegati 200 i.e./kg 3 x tedensko. Odrasli bolniki na peritonealni dializi: Korekcijska faza: 50 i.e./kg s.c. 2 x tedensko.Vzdrževalni odmerek je od 25 do 50 i.e./kg 2 x tedensko. Odrasli bolniki z rakom s simptomatsko anemijo, ki se zdravijo s kemoterapijo: Bolnike z anemijo zdravimo do ciljne koncentracije Hb 100-120 g/l, Hb pa ne sme preseci 120 g/l. Zacetni odmerek je 150 i.e./kg s.c. 3 x tedensko ali 450 i.e./kg s.c. 1 x tedensko. Odrasli kirurški bolniki, vkljuceni v program avtolognega zbiranja krvi za avtotransfuzijo: 600 i.e./kg i.v., 2-krat na teden v obdobju treh tednov pred kirurškim posegom. Odrasli kirurški bolniki, ki niso vkljuceni v program avtolognega zbiranja krvi za avtotransfuzijo: 600 i.e./kg, s.c., enkrat tedensko v obdobju treh tednov pred kirurškim posegom in na dan kirurškega posega. Kontraindikacije: cista aplazija rdecih krvnih celic (PRCA), nenadzorovana arterijska hipertenzija, kontraindikacije povezane s programom avtolognega zbiranja krvi, preobcutljivost za katerokoli sestavino zdravila, bolniki, pri katerih je predviden vecji elektiven kirurški poseg in niso vkljuceni v program avtolognega zbiranja krvi s hudo koronarno, cerebrovaskularno, karotidno ali periferno arterijsko bolezen ali so nedavno preboleli miokardni infarkt ali cerebrovaskularni dogodek, bolniki, ki ne morejo prejemati ustrezne antitromboticne pro.lakse. Posebna opozorila in previdnostni ukrepi: Med zdravljenjem moramo spremljati in nadzorovati krvni tlak, ce ga ne moremo urediti, moramo zdravljenje prekiniti. Potrebna je previdna uporaba zdravila pri bolnikih z epilepsijo in kronicno boleznijo jeter. Prvih osem tednov zdravljenja priporocamo redno spremljanje števila trombocitov. Za optimalen odgovor na zdravljenje, je treba zagotoviti ustrezne zaloge železa. Po vec mesecih ali letih zdravljenja s subkutano apliciranim zdravilom so redko porocali o PRCA, povzroceni s protitelesi. Ce sumimo PRCA moramo zdravljenje takoj prekiniti. Zaradi verjetnosti navzkrižne reakcije s protitelesi, bolniku ne smemo dati drugega epoetina in mu moramo zagotoviti ustrezno zdravljenje. Pri ocenjevanju ustreznosti odmerka pri bolnikih z rakom, ki prejemajo kemoterapijo, moramo upoštevati, da minejo 2-3 tedni od zacetka zdravljenja do pojava eritrocitov, nastalih pod njegovim vplivom v krvi. Kot pri vseh rastnih faktorjih obstaja verjetnost, da bi lahko spodbujali razvoj katere koli vrste rakave bolezni. Pri bolnikih, pri katerih je predviden vecji elektivni ortopedski kirurški poseg, je treba ugotoviti vzrok za anemijo in ga odpraviti pred zacetkom zdravljenja. Pri bolnikih s kronicnim ledvicnim odpovedovanjem je potrebna previdnost. Interakcije: Ni dokazov, da zdravljenje z epoetinom alfa vpliva na metabolizem drugih zdravil. Ker se ciklosporin veže na eritrocite, obstaja možnost interakcije med zdraviloma. Neželeni ucinki: trombocitemija, PRCA, ana.lakticna reakcija, hipersenzitivnost, krci, glavobol, cerebralna krvavitev, cerebrovaskularni dogodki, hipertenzivna encefalopatija, tranzitorna ishemicna ataka, hipertenzija, tromboze, pljucna embolija, navzea, diareja, bruhanje, izpušcaj, angionevroticni edem, urtikarija, artralgija, mialgija, por.rija, pireksija, gripi podobni simptomi, neucinkovitost zdravila, periferni edem, reakcija na mestu injiciranja, tromboza žilnega pristopa. Imetnik dovoljenja za promet: Johnson & Johnson d.o.o. Šmartinska 53, 1000 Ljubljana Režim izdajanja zdravila: H/Rp. Datum revizije: 11. 12. 2009. Instructions Instructions for authors The editorial policyof the journal Radiology and Oncologyis topublish original scientific papers, professional papers, review articles case reports and varia (editorials, short communications, professional information, book reviews, letters, etc.) perti­nent to diagnostic and interventional radiology, computerized tomography, magnetic resonance, ultrasound, nuclear medi­cine, radiotherapy, clinical and experimental oncology, radiobiology, radiophysics and radiation protection. The Editorial Board requires that the paper has not been published or submitted for publication elsewhere; the authors are responsible for all statements in their papers. Accepted articles become the property of the journal and therefore cannot be published else­where without written permission from the editorial board. Papers concerning the work on humans must comply with the principles of the declaration of Helsinki (1964). 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If corrections are not received by the stated deadline, proof-reading will be carried out by the editors. Reprints: The electronic version of the published papers will be available on www.versita.com free of charge. Radiol Oncol 2010; 44(2) BISTVENE INFORMACIJE IZ POVZETKA GLAVNIH ZNACILNOSTI ZDRAVILA SUTENT 12,5 mg, 25 mg, 37,5 mg, 50 mg trde kapsule Sestava in oblika zdravila: Vsaka trda kapsula vsebuje 12,5 mg, 25 mg, 37,5 mg ali 50 mg sunitiniba v obliki sunitinibijevega malata. Indikacije: Zdravljenje neizrezljivega in/ali metastatskega malignega gastrointestinalnega stromalnega tumorja (GIST), ce zdravljenje z imatinibijevim mesilatom zaradi odpornosti ali neprenašanja ni bilo uspešno. Zdravljenje napredovalega in/ali metastatskega karcinoma ledvicnih celic (MRCC). Odmerjanje in nacin uporabe: Terapijo mora uvesti zdravnik, ki ima izkušnje z zdravljenjem MRCC ali GIST. Priporoceni odmerek je 50 mg enkrat dnevno, peroralno vsak dan 4 tedne zapored; temu sledi 2-tedenski premor (Shema 4/2), tako da celotni ciklus traja 6 tednov. Odmerek je mogoce prilagajati v povecanjih po 12,5 mg, upoštevaje individualno varnost in prenašanje. Dnevni odmerek ne sme preseci 75 mg in ne sme biti manjši od 25 mg. Pri socasni uporabi z mocnimi zaviralci ali induktorji CYP3A4 je potrebno odmerek ustrezno prilagoditi. Uporaba pri otrocih in mladostnikih (< 18 let): Sutenta ne smemo uporabljati, dokler ne bo na voljo dodatnih podatkov. Uporaba pri starejših bolnikih (= 65 let): med starejšimi in mlajšimi bolniki niso opazili pomembnih razlik v varnosti in ucinkovitosti. Insuficienca jeter: pri bolnikih z jetrno okvaro razreda A in B po Child-Pughu prilagoditev odmerka ni potrebna; pri bolniki z okvaro razreda C Sutent ni bil preizkušen. Insuficienca ledvic: klinicnih študij niso izvedli. Sutent se uporablja peroralno, bolnik ga lahko vzame z ali brez hrane. Ce pozabi vzeti odmerek, ne sme dobiti dodatnega, temvec naj vzame obicajni predpisani odmerek naslednji dan. Kontraindikacije: Preobcutljivost za zdravilno ucinkovino ali katerokoli pomožno snov. Posebna opozorila in previdnostni ukrepi:Koža in tkiva. Krvavitve v prebavila, dihala, secila, v možganih ter krvavitve tumorja. Ucinki na prebavila: poleg navzee in driske tudi resni zapleti. Hipertenzija. Hematološke bolezni. Bolezni srca in ožilja: zmanjšanje LVEF in srcno popušcanje. Podaljšanje intervala QT. Venski trombembolicni dogodki. Dogodki na dihalih: dispneja, plevralni izliv, pljucna embolija ali pljucni edem. Moteno delovanje šcitnice. Pankreatitis. Delovanje jeter. Delovanje ledvic. Fistula. Preobcutljivost/angioedem. Motnje okušanja. Konvulzije. Pri krvavitvah, ucinkih na prebavila, hematoloških boleznih, dogodkih na dihalih, venskih trombembolicnih dogodkih, pankreatitisu in ucinkih na jetra so opisani tudi smrtni izidi. Medsebojno delovanje z drugimi zdravili: Zdravila, ki lahko zvišajo koncentracijo sunitiniba v plazmi (ketokonazol, ritonavir, itrakonazol, eritromicin, klaritromicin ali sok grenivke). Zdravila, ki lahko znižajo koncentracijo sunitiniba v plazmi (deksametazon, fenitoin, karbamazepin, rifampin, fenobarbital, Hypericum perforatum oz. šentjanževka). Antikoagulanti. Nosecnost in dojenje: Sutenta se ne sme uporabljati med nosecnostjo in tudi ne pri ženskah, ki ne uporabljajo ustrezne kontracepcije, razen ce možna korist odtehta možno tveganje za plod. Ženske v rodni dobi naj med zdravljenjem s Sutentom ne zanosijo. Ženske, ki jemljejo Sutent, ne smejo dojiti. Vpliv na sposobnost vožnje in upravljanja s stroji: Sutent lahko povzroci omotico. Neželeni ucinki: Najpogostejši neželeni ucinki: pljucna embolija, trombocitopenija, krvavitev tumorja, febrilna nevtropenija, hipertenzija, utrujenost, diareja, navzea, stomatitis, dispepsija, bruhanje, obarvanje kože, disgevzija, anoreksija, zvišanje ravni lipaze. Zelo pogosti: anemija, nevtropenija, hipotiroidizem, zmanjšanje teka, motnje okušanja, glavobol, bolecina v trebuhu / napihnjenost, flatulenca, bolecine v ustih, sindrom palmarno plantarne eritrodizestezije, spremembe barve las, astenija, vnetje sluznice, edemi. Nacin in režim izdajanja: Izdaja zdravila je le na recept, uporablja pa se samo v bolnišnicah. Izjemoma se lahko uporablja pri nadaljevanju zdravljenja na domu ob odpustu iz bolnišnice in nadaljnjem zdravljenju. Imetnik dovoljenja za promet: Pfizer Limited, Ramsgate Road, Sandwich, Kent, CT13 9NJ, Velika Britanija. Datum zadnje revizije besedila: 28.10.2009 Pred predpisovanjem se seznanite s celotnim povzetkom glavnih znacilnosti zdravila. SUT-01-10 Pfizer Luxembourg SARL, Grand Duchy of Luxembourg, 51, Avenue J.F. Kennedy, L-1855, PFIZER, Podružnica za svetovanje s podrocja farmacevtske dejavnosti, Ljubljana, Letališka 3c, 1000 Ljubljana, SLOVENIJA