ADIOLOGY .,.11 NCOLOGY September 2006 Vol. 40 No. 3 Ljubljana ISSN 1318-2099 BIS1VENE INRIRMACIE IZ POVZE1'KA GlAIINIH ZNACaNosn ZDRAVIU AROMASIN" -.1n cMal zdmla: oblofena tabletavselqe 251111. ........ . napredovalega raka dojke pri :lenskah z ll8IIMlO ali ooietno povzroceno menopavzo, pri kaler1hje bolezen napadovala po 11ot1es111111enskl tmap!II. IJtllnkc,;ftost še ni bila dokazana pri bolnicah, pri katerih l1molSke ceOce nimajo estnJ(lensklll receplorjev. . In naSn upooabe: 25 1111 enkrat na dan,najbofe po jed. Zdravlerie naj traja, dokler Je opazna rast llm>lja. 0/Mn]etsr In IIHMc: odmerka ni potrebno priJllgodlll. 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IWln In rellm ......., zxlra'lllo se . le na recept. uporablja pa se po navodilu In pod posebnim nadzorom zdravnika specfallsla ali od njega pooblašc!enega zdravnika. -.lc .......... za promet Pflzer wxernbotq SARL, 283, route d'Arton, L-8011 Strassen, l..uksembl.Wg. _,, _.. ...... le beleclla: 5.10.2001 Prad predplsowqem se seznanlte s celolnlm povzetkom glavnih znacilnosti zllravlla. Podrobnejše Informacije o zdrevllu so na voljo pri: Pfizar, podružnica za svetovanje s podrocja farmacevtske dejavnosti, LJubljana, Letall§ka cesta 3c, 1000 LJubljana Editorial office Radiologij and Oncology Institute of Oncology Zaloška 2 SI-1000 Ljubljana Slovenia Phone: +386J 5879 369 Phone/Fax: +386 1 5879 434 E-lllail: gsersa@onko-i.si Aims and scope Septelllber 2006 Vol. 40 No. 3 Pages 143-202 ISSN 1318-2099 UDC 616-006 CODEN: RONCEM Radiologi; and OncologiJ is a joumal devoted to publication oj original contributions in diagnostic and interventional radiology, colllputerized tomography, ultrasound, magnetic resonance, nuclear medicine, radiotherapy, clinical and Editor-in-Chief Gregor Serša Ljubljana, Slovenia Executive Editor Viljem Kovac Ljubljana, Slovenia Editorial board Marija Auersperg Ljubljana, Slovenia Nada Bešenski Zagreb, Croatia Karl H. Bohuslavizki Hamburg, Germany Haris Boko Zagreb, Croatia Nataša V. Budihna Ljubljana, Slovenia Marjan Budihna Ljubljana, Slovenia Malte Clausen Hamburg, Gennany Christoph Clemm M11nchen, Germany Mario Corsi Udine, Italy Ljubomir Diankov Sofia, Bulgaria Christian Dittric/1 Vienna, Austria Ivan Drinkovic Zagreb, Croatia Gillian Duchesne Melbourne, Australia Editor-in-Chief Emeritus Tomaž Benulic Ljubljana, Slovenia Editor Uroš Smrdel Ljubljana, Slovenia Valentin Fidler Ljubljana, S/ovenia Be1aFornet Budapest, Hungary Titllio Giraldi Trieste, Italy Andrija Hebrang · Zagreb, Croatia Laszl6 Horvath Pecs, Hungary Berta Jereb Ljubljana, Slovenia Vladimir Jevtic Ljubljana, Slovenia H. Dieter Kogelnik Salzburg, Austria Jurij Lindtner Ljubljana, Slovenia Ivan Lovasic Rijeka, Croatia Marijan Lovrencic Zagreb, Croatia LukaMilas Houston, USA Metka Milcinski Ljubljana, Slovenia .agreb, Croatia .. Branko Palcic Vancouver, Canada Jurica Papa Zagreb, Croatia Dušan Pavcnik Portland, USA Stojan Plesnicar Ljubljana, Slovenia Ervin B. Podgoršak Montreal, Canada Jan C. Roos Amsterdam, Netherlands Slavko Šimunic Zagreb, Croatia Lojze Šmid Ljubljana,Slovenia Borut Štabuc Ljubljana, Slovenia Andrea Veronesi Aviano, Italy iiva Zupancic Ljubljana, Slovenia Publisher Association of Radiology and Oncology Affiliated with Slovenian Medica/ Association -Slovenian Association of Radiology, Nuclear Medicine Society, Slovenian Society for Radiotherapy and Oncology, and Slovenian Cancer Society Croatian Medica/ Association -Croatian Society of Radiology Societas Radiologorum Hungarorum Friuli-Venezia Giulia regional groups of S.I.R.M. (Italian Society of Medica/ Radiologi;) Copyright © Radiology and Oncologi;. Ali rights reserved. Reader for English Vida Kološa Key words Eva Klemencic Secretary Mira Klemencic Design Monika Fink-Serša Printed by Imprint d.o.o., Ljubljana, Slovenia Published quarterly in 700 copies Beneficiary name: DRUŠTVO RADIOLOGIJE IN ONKOLOGIJE Zaloška cesta 2, 1000 Ljubljana Slovenia Beneficiary bank account number: 5156 02010-0090006751 !BAN: 5!56020100090006751 Our bank name: Nova Ljubljanska banka, d.d., Ljubljana, Trg republike 2, 1520 Ljubljana; Slovenia SWIFT: LJBASI2X Subscription fee for instih1tions EUR 100 (20.000 SIT), individuals EUR 50 (5000 SIT) The publication of this journal is subsidized by the Slovenian Research Agency. Indexed and abstracted by: BIOMEDICINA SLOVENICA CHEMICAL ABSTRACTS EMBASE / Excerpta Medica Sci Base Scopus This journal is printed on acid- free paper Radiologi; and Oncologi; is available on the internet at: http:/ /www.onko-i.si/radioloncol ISSN 1581-3207 Ljubljana, Slovenia ISSN 1318-2099 September 2006 UDC 616-006 Vol. 40 No. 3 CODEN: RONCEM CONTENTS RADIOLOGY Partial subclavian steal syndrome in a congenitally anomalous subclavian artery Krnic A, Sucic Z, Vucic N, Krolo I IMAGING IN CLINICAL MEDICINE Duplication cyst of the esophagus Šubic T, Jamar B, Dolenšek M ONCOLOGY The cardiotoxicity of chemotherapy: New prospects for an old problem 149 Miolo GM, La Mura N, Nigri P, Murrone A, Da Ronch L, Viel E, Veronesi A, Lestuzzi C Electrochemotherapy of tumours 163 Serša G, Cemažar M, Miklavcic D, Rudolf Z Fertility preservation methods for female neoplastic patients 175 Del Pup L, Campagnutta E, Giorda G, De Piero G, Sopracordevole F, Sisto R Different localisation of cystatin C in immature and mature dendritic cells 183 Zavašnik-Bergant T, Bergant M, Jeras M, Griffiths G RADIATION PROTECTION Diagnostic reference levels for X-ray examinations in Slovenia 189 Škrk D, Zdešar U, Žontar D SLOVENIAN ABSTRACTS 197 NOTICES 201 Radiology and Oncology is covered in Biomedicina Slovenica, Chemical Abstracts, EMBASE / Excerpta Medica, Sci Base and Scopus Partial subclavian steal syndrome in a congenitally anomalous subclavian artery Anton Krnic,1 Zvonimir Sucic,1 Nikša Vucic,2 Ivan Krolo,3 1Department of Radiology, 2Internal Medicine Department, »Holy Ghost« General Hospital, Zagreb, Croatia, 3Department of Radiology, »Sisters of Mercy« Clinical Hospital, Zagreb, Croatia Background. A subclavian steal syndrome results from the abnormal flow of blood due to the occlusion in the subclavian artery proximal to the origin of the vertebral artery. A case of a male patient with a partial subclavian steal syndrome is presented. Case report. The syndrome was caused by a stenotic lesion of an aberrant right sublcavian artery (the so called »lusorian artery«). The partial subclavian steal was recognized using the duplex ultrasound which showed the »to and fro« pattern in the right vertebral artery. Angiography of the aortic arch revealed the arterial anomaly. In our case, duplex ultrasound was a crucial method in diagnosing the partial subclavian steal syndrome. However, in order to show the arterial anomaly, the final evaluation had to be performed using arteriography. Conclusions. The early recognized partial subclavian steal syndrome provides good understanding of patient’s symptoms, successful follow up, and a variety of treatment options. Key words: subclavian artery – abnormalities – radiography – ultrasonography; subclavian steal syn­drome; angiograph y; Doppler duplex; vertebral artery Introduction The most frequent congenital malformation of the aortic arch branches is the aberrant right subclavian artery.1 It is found in 0.5-1% of the population.1 Although the compres­sion of the oesophagus may occur, most Received 23 April 2006 Accepted 14 May 2006 patients are asymptomatic.1 The syndrome of subclavian steal caused by an occlusive lesion of the aberrant subclavian artery is a rare clinical finding.1-4 This syndrome re­sults from the abnormal flow of blood due to the occlusion in the subclavian artery proximal to the origin of the vertebral ar­tery. Blood flow through the vertebral artery is consequently reversed and the subclavian one thus »steals« cerebral blood. The syndrome of partial subclavian Correspondence to: Anton Krnic, MD, Department of steal, caused by the stenotic lesion of the Radiology, »Sveti Duh« General Hospital, Sveti Duh aberrant artery, has not been described in 64, HR-10000 Zagreb, Croatia; Fax: +385 1 37 72 136; E-mail: anton.krnic@zg.t-com.hr literature yet. Case report A 49 year-old man was seen because of dizziness and intermittent paresthesia of the right arm. When he was checked up, there was a blood pressure difference between both arms (right brachial pres­sure 90/60 mmHg, left 120/80 mmHg). Electronistagmography confirmed no ves­tibular lesion. Duplex scanning showed miscellane­ous plaque of the right internal carotid artery (30% stenosis), with moderately in­creased peak systolic velocity. The right vertebral artery was hypoplastic, with a spectral alteration characteristic for the initial subclavian steal syndrome (the ‘to and fro’ pattern, Figure 1a).5 There were increased velocities and turbulent flow in the proximal segment of the right subcla­vian artery which suggested significant proximal stenosis of the artery (Figure 1b). The transcranial Doppler also showed a flow asymmetry between right and left vertebral artery. Unlike the left vertebral ar­tery, which showed normal, towards brain directed flow (Figure 2a), the right vertebral artery showed bidirectional flow with de­creased peak systolic velocity (Figure 2b). The clinical and duplex findings indi­cated the presence of a partial subclavian steal caused by moderate stenosis of the right subclavian artery. Angiography ad­ditionally showed the abnormal origin and course of the right subclavian artery, well known as the »lusorian artery« (Figure 3a, 3b).1-4 It also showed the mild to moderate grade stenosis of the artery in the middle Radiol Oncol 2006; 40(3): 143-6. Figure 3a. Digital subtraction angiography of the aor­tic arch: there is an aberrant right subclavian artery, which arises as the most distal vessel from the aortic arch and crosses the middle line. The image provides evidence of a mild to moderate grade stenosis of the aberrant artery in middle line, but there is no clear evidence of steal syndrome. line (Figure 3a, 3b). It, however, did not provide clear evidence for the presence of the subclavian steal. Since the patient did not complain of dysphagia, no further evaluation (oesoph­agogram or CT) was done. The patient was finally released form the hospital and was referred for internist and neurological follow-up. Discussion The stenosis of the subclavian artery in our patient was likely the result of progression of an atherosclerotic lesion in the segment of the artery which was in contact with the esophagus.1-4 It resulted in the partial re­versal of blood flow in the vertebral artery. The partial reversal of blood flow in our patient could clearly be confirmed only by duplex ultrasound (Figure 1a). It resulted in clinical symptoms known as the partial subclavian steal syndrome.5 Figure 3b. Selective digital subtraction angiography of the aberrant right subclavian artery (lusorian artery). Unlike us, De Vleeschauwer et al.1, as well as other authors reported patients that had severe stenosis or occlusion of the aberrant subclavian artery and, thus, suffered steal syndrome in its advanced phase, known as the total subclavian steal syndrome. In those patients, the symptoms were much more pronounced and the syn­drome was easier to diagnose using duplex ultrasound or other imaging modalities as well.1-4 The anomalous origin of the right sub-clavian artery was first reported about 200 years ago by Bayford.6 The anomalous right subclavian artery (»arteria lusoria«) passes behind the oesophagus in about 80% of the cases and in these cases a posterior notch can be seen in oesophagogam and during endoscopy.1,7 The most common symptom is dysphagia, the so-called »dysphagia luso­ria« - dysphagia secondary to a freak of na­ture.8 CT, MR and endoscopic ultrasound help in differential diagnosis.7-9 Treatment options, if indicated, include conservative treatment, surgical treatment, and endovascular treatment, which is re­cently also considered in cases of subcla­ vian steal. 1,3,4,7-11 Radiol Oncol 2006; 40(3): 143-6. In conclusion, patients with the luso­rian artery can develop a subclavian steal syndrome, caused by a stentoic lesion of the retroesophageal segment of the aber­rant artery. The subclavian steal can be recognized in its early, partial phase. At that stage duplex ultrasound is the major and usually the only enough sensitive diag­nostic tool. In order to diagnose it, the ul­trasononographer should be familiar with the ‘to and fro’ flow pattern in the vertebral artery.5 Early ultrasonographic recognition of the condition could provide a better, on-time, understanding of patient’s symp­toms. In that way, it is possible to plan treatment and follow-up options in a more efficient way. References 1. De Vleeschauwer P, Horsch S. Subclavian steal syndrome in a congenitaly anomalous subclavian artery: a case report. Ann Vasc Surg 1986; 1: 389­91. 2. Rowe DM, Becker GJ, Scott JA, Conces DJ Jr. Right subclavian steal associated with aberrant right subclavian artery. AJNR Am J Neuroradiol 1988; 9: 604-6. 3. Azakie A, McElhinney DB, Dowd CF, Stoney RJ. Percutaneous stenting for symptomatic stenosis of aberrant right subclavian artery. J Vasc Surg 1998; 27: 756-8. 4. Basile A, Lomoschitz F, Lammer J. Transbrachial stenting of a critical ostial arteria lusoria stenosis. J Endovasc Ther 2003; 10: 829-32. 5. Paivansalo M, Heikkila O, Tikkakoski T, Leinonen S, Merikanto J, Suramo I. Duplex ultrasound in the subclavian steal syndrome. Acta Radiol 1998; 39: 183-8. 6. Bayford D. Account of singular case of obstructive deglutition. Mem Med Soc London 1794; 2: 271-82. 7. Janssen M, Baggen MG, Veen HF, Smout AJ, Bekkers JA, Jonkman JG, et al. Dysphagia lusoria: clinical aspects, manometric findings, diagnosis, and therapy. Am J Gastroenterol 2000; 95: 1411-6. 8. Brown DL, Chapman WC, Edwards WH, Coltharp WH, Stoney WS. Dysphagia lusoria: aberrant right subclavian artery with a Kommerell’s diverticu­lum. Am Surg 1993; 59: 582-6. 9. Maeder M, Binek J. Impact of endoscopic ultra-sonography in the diagnosis of aberrant rightsub­clavian artery: a case report. [German] Ultraschall Med 2004; 25: 296- 8. 10. Pome G, Vitali E, Mantovani A, Panzeri E. Surgical treatment of the aberrant retroesophageal right subclavian artery in adults (dysphagia lusoria). Report of two new cases and review of the litera­ture. J Cardiovasc Surg (Torino). 1987; 28: 405-12. 11. Vos AW, Wisselink W, Rijbroek A, Avontuur JA, Manoliu RA, Rauwerda JA. Endovascular repair of a type B aortic dissection with transposition of a coexistent aberrant subclavian (lusorian) artery. J Endovasc Ther 2002; 9: 549-53. Radiol Oncol 2006; 40(3): 143-6. Images in clinical medicine Duplication cyst of the esophagus Tanja Šubic,1 Breda Jamar,2 Marija Dolenšek2 1General Hospital of Novo mesto, Novo mesto, Slovenia 2Clinical Institute of Radiology, Clinical Centre, Ljubljana, Slovenia A 24 years old female, previously healthy, complained for fever and severe retrosternal pain. Chest X-ray showed a homogenous, well delineated, half-circular bulge of right mediastinum at the level of the left atrium (Figure 1), and antibiotic treatment alleviat­ed the symptoms. Afterwards barium swal­low, and CT of chest were performed. On Received 13 June 2006 Accepted 20 June 2006 Correspondence to: Tanja Šubic, MD, General Hospital of Novo mesto, Šmihelska cesta 1, 8000 Novo mesto, Slovenia. Figure 2. On barium swallow a semi-circular impres­sion on the right side of the middle third of esophagus is seen. barium swallow (Figure 2) a semi-circular impression on the right side of the middle third of esophagus is seen, the wall in this segment is rigid and peristalsis weak. Thin band of contrast medium is leaking out of the lumen. MDCT scans (Figures 3 and 4) showed an air-filled oval formation within the right posterior mediastnum. A com­munication between the esophagus and the formation cannot be demonstrated. Clinical features and radiological exami­nations suggest this formation most likely to be the duplication cyst of the esophagus. Other possible diagnoses are bronchogenic cyst, mediastinal lymphadenopathy, es­ophageal diverticuluma and neurinoma. This rare congenital anomaly is mostly asymptomatic and represents an inciden­tal finding on chest X- ray. Our patient had clinical symptoms because of cyst inflammation. Surgery was performed and a fibrotic, purulent structure, adherent to surrounding structures, was removed. Histology confirmed the radiological diag­nosis. Figure 4. MDCT coronal reconstructions showed an air-filled oval formation within the right posterior mediast­num. Radiol Oncol 2006; 40(3): 147-8. rewiev The cardiotoxicity of chemotherapy: New prospects for an old problem GianMaria Miolo1, Nicoletta La Mura1, Paola Nigri1, Antonio Murrone1, Laura Da Ronch1, Elda Viel2, Andrea Veronesi1, Chiara Lestuzzi2 1Division of Medical Oncology C, 2Cardiology Service, Centro di Riferimento Oncologico, Aviano, Italy Background. Cardiotoxicity caused by chemotherapy, with its diverse early and late presentations, can hamper potentially curative or palliative treatments. The drugs most often linked to cardiotoxicity include anthracyclines, trastuzumab, 5-fluorouracil and taxanes, but some forms of cardiotoxicity have been de­scribed, more or less sporadically, for most antitumour agents. It is likely that the widespread use of the new biological target therapies will lead to the identification of other less known toxic effects. The available data on its incidence and clinical presentation, the pathogenetic mechanisms involved, the diagnosis, prevention and management of cardiac toxicity from chemotherapy are briefly reviewed. Conclusions. The identification of novel molecular targets will increase the number of drugs available for the treatment of neoplastic disease. It will be important to evaluate the side effects related to treatment, particularly in organs with a limited regenerative capability such as the heart. Further studies will therefore be necessary. Key words: antineoplastic agents – adverse effects – toxicity; heart – drug effects Introduction The available data on the incidence, risk factors, pathogenetic mechanisms in­volved, clinical presentation, diagnosis and management of cardiac toxicity from anti- Received 14 July 2006 Accepted 22 August 2006 Correspondence to: GianMaria Miolo, MD, Division of Medical Oncology C, Centro di Riferimento Oncologico, Via Pedemontana 12, 33081 Aviano, Italy; Phone: + 39 0434-659653; Fax: + 39 0434-659453; E­mail: gmiolo@libero.it cancer drugs are briefly reviewed. The pos­sibilities to intervene through alterations of the chemotherapy schedule or the use of cardioprotective agents are discussed. Not mentioned are the possible cardiac com­plications of radiotherapy and hormonal therapy. Risk factors Risk factors for the development of hy­pokinetic cardiopathy from anthracyclines include: elevated serum anthracycline con­centrations following high doses and/or short infusions, previous or concomitant radiotherapy to the mediastinum or to the left hemithorax, age less than 15 years or over 65 years, cardiovascular risk factors such as arterial hypertension and diabetes, pre-existing hypokinetic cardiopathy and the female sex, in which morbidity and cardiac mortality are twice as frequent as in males in the presence of another cardio­vascular risk factor.1-6 The only identified risk factors for tras­tuzumab are the type of chemotherapy pretreatment and the age of the patient that appear to be particularly influential in the group also treated with anthracyclines.7 Regarding 5-fluorouracil, there is no con­sensus on age as a risk factor. Some authors report a more elevated risk in patients older than 50, while others do not.8-10 Anthracyclines The most frequently observed cardiotoxic effect after the use of anthracyclines (adri­amycin or doxorubicin, daunorubicin, epi­rubicin and mitoxantrone) is the depression of myocardial contractility. About 10-15% of neoplastic patients treated with anthracy­clines develop a hypokinetic cardiopathy11, and this depends mainly on the cumulative dose of the drug. The risk related to an adriamycin cumulative dose equal to 550 mg/m2 is 7%; this grows linearly with high­er dosages, reaching 50% for a total dose of 1000 mg/m2.1,12 However, cardiotoxicity can also occur at a cumulative dose of less than 400 mg/m2, especially if the anthracycline is combined with other cytotoxic drugs, for example, cyclophosphamide.13 Various hy­potheses on the pathogenetic mechanisms of cardiopathy from anthracyclines have been formulated: the production of free radicals14 and/or a reduction of free radi­cal scavengers; the inhibition of the ionic pumps through toxic metabolites15; altera­tion of the energetic mitochondrial metabo­lism16; the formation of a complex binding trivalent iron ions that damage cellular membranes and DNA3; the release of in­flammatory cytokines11; and the induction of an adrenergic dysfunction.17 Histological damage is characterized by the expansion of the sarcomere tubules and by the loss of actin and myosin myofilaments. Cardiotoxicity can occur early or in a later phase. Acute toxicity, that is during chemotherapy infusion, occurs in 0.4 - 41% of cases11, especially in the presence of electrolytic alterations15, presenting mainly as sinus tachycardia, supraventricular or ventricular non-repetitive arrhythmia, as-pecific alterations of repolarisation or an extension of the QT tract. In this phase, the patient is generally asymptomatic or only mildly so, and these alterations sub­side some hours after the discontinuation of the drug.1 This type of cardiotoxicity is not a contraindication for further treatment with anthracyclines, but only after an ade­quate correction of the plasma electrolytes. However, cases of sudden death correlated to arrhythmia have been reported.2 Chronic toxicity appears weeks to months to years after the end of chemo­therapy; it can also occur during treatment when the cumulative dose is elevated, or years after a first course of chemotherapy if the patient receives a second course. The in­cidence of chronic toxicity in anthracycline treated patients varies from 0.4 - 23%.18,19 The appearance of signs and/or symptoms of congestive heart failure such as pulmo­nary edema or cardiogenic shock, which may occur progressively or unexpectedly, is frequent.12,13 The ECG may be normal or may present ventricular conduction delays of different grades. Chest radiography may show possible signs of pulmonary conges­tion5 with cardiac dimensions in the up­per normal range. Echocardiography may Radiol Oncol 2006; 40(3): 149-61. show hypokinesia, often more evident at the septal level, with a reduced function of the left ventricular pump which may or may not be associated with the involvement of the right ventricle, depending on the duration of the heart failure. The cardiac dimensions are frequently in the normal range or only slightly increased, but in the more serious cases or in the case of a late diagnosis, these may be greatly increased with evident remodeling. Parietal thickness is generally preserved. The diastolic mi-tralic flow may be normal or altered when seen on echodoppler. An endomyocardial biopsy may highlight vacuolisation of the cytoplasm and mitochondrial degeneration as well as the loss of myofibrils and perivas­cular fibrosis.20 The use of less cardiotoxic second gen­eration anthracyclines such as idarubicin, epirubicin or mitoxantrone has been sug­gested and utilized in clinical practice. However, second generation anthracyclines are not devoid of cardiotoxicity, but gener­ally at higher doses.18,21,.22 Moreover, the damage is cumulative even among different anthracyclines. One strategy to limit doxorubicin car-diotoxicity is its encapsulation in lipo­somes which alters the tissue distribution and the pharmacokinetics of the drug, limiting its toxic effect on healthy tissue. The available data suggests a better cardio-logical safety profile of these formulations, both in monochemotherapy and in combi­nation with trastuzumab or other cytotoxic agents.23,24 Attention to the cumulative dose and a careful cardiological follow-up still have the greatest role in the prevention of an-thracycline cardiotoxicity. Measurement of the left ventricular pump function should be performed at baseline either by echo-cardiogram or by radionuclide angiography when a cumulative dose of 250-300 mg/m2 of adriamycin or 500 mg/m2 of epirubicin or 25-30 mg/m2 of mitoxantrone has been reached and repeated 10-30 days after the end of chemotherapy.14,25 Late cardiotoxic­ity can be detected by extending the cardio-logical follow-up annually for at least 4-5 years after chemotherapy termination. In the past, the prognosis of cardio­myopathy from anthracyclines was rather unfavourable. The two-year mortality rate of patients in the functional NYHA III-IV class, in treatment only with digitalis and diuretics, was 50%.26 The prognosis in children appears to be better than that of adults1, with a two-year mortality rate of 20%. After the introduction of the ACE-inhib­itors for the treatment of patients with left ventricular dysfunction irrespective of the aetiology, the prognosis of patients with anthracycline cardiopathy clearly improved in terms of both morbidity and mortality. Moreover, the possible reversibility of left ventricular dysfunction has also been dem- onstrated.11,27,28,29 The most used drugs are captopril and enalapril30 at the same recommended dose as in other types of hypokinetic cardiopathy. Currently, studies on the effectiveness of beta-blockers in this specific type of car-diopathy are less numerous.31,32 However, small studies have shown a significant improvement of the left systolic ventricular function33,34 and of the NYHA functional class35 in patients with anthracycline car-diopathy receiving beta-blocker treatment. The principal drugs used in these patients are metoprolol33,35 and carvedilol36 at the doses recommended by the international guidelines for cardiac failure. Trastuzumab Trastuzumab is a recombinant monoclonal antibody that targets the HER-2 receptor. It is effective in metastatic breast cancer Radiol Oncol 2006; 40(3): 149-61. with a high HER-2 expression both as a single agent or in combination therapy and as first-line treatment or in pretreated pa­tients. Being a very specific monoclonal an­tibody, it has a favourable toxicity/efficacy ratio. However, it can cause cardiotoxicity in the form of ventricular dysfunction and heart failure, especially if administered af­ter or together with anthracyclines.7,37,38,39 The mechanism of the toxicity is still not known, but some hypotheses have been formulated. Direct toxicity While HER-2 and HER-4 receptors are in­volved in cell growth, reparation and sur­vival during the embryonic development of the heart, the normal adult muscle has few HER-2 receptors. Both HER-2 and HER-4 receptors have been detected in the myocar­dium of some cases showing clinical toxicity from trastuzumab.39,40 In a small series of patients, the injection of radiomarked tras­tuzumab was followed by cardiac captation in about one third of the cases, and most of these presented subsequent cardiotoxicity.41 Therefore, trastuzumab could have a direct cardiotoxic effect, at least in some subjects. Probably, there is an interpersonal variabil­ity in the expression of HER-2 and HER-4 receptors in the human myocardium as well as in the tumor itself. Indirect toxicity Cardiotoxicity is greater when trastuzu­mab is administered together with anthra­cyclines and, if this occurs under stress conditions, the HER-2 receptor is possibly involved in the reparative processes of damaged myocytes. This could suggest that the HER-2 receptor blockade prevents the myocardium from repairing the damage caused by anthracyclines, increasing their eventual toxic effect.39,42 Trastuzumab has an antitumoral speci­ficity and a favourable toxicity/activity ra­tio that will probably make it a more widely used agent in the next years. The recently available results on the impact of trastu­zumab in the adjuvant therapy of breast carcinoma could be predictive of a more generalized use in patients with HER-2 hyperexpressing tumors after surgery. The evaluation of the cardiotoxicity will be of special importance in this context. 5-Fluorouracil In the literature, the frequency of fluorou-racil-related cardiotoxicity such as angina, myocardial infarction, arrhythmia, heart failure and sudden death ranges from 1.6% -18%, with a mortality rate ranging from 2.2% to 13%. This variability is partly due to patient selection (inclusion or exclusion of those with pre-existing cardiopathy, especially of the ischemic type), to the study methods (retrospective or prospec­tive; with provocative tests and/or close cardiological follow-up or limited to clini­cal observation only), to the method of drug administration (high or low doses; bolus or continuous infusion) and to the possible interference of other potentially cardiotoxic drugs in polychemotherapeu-tic regimens.8,9,43-47 From a metanalysis of the literature, cardiotoxicity generally con­cerns 5-10% of unselected subjects receiv­ing high doses (>800 mg/day), particularly in continuous infusion. In single bolus doses at 1-4 week intervals, as, for exam­ple, in the CMF regimen, it is generally well tolerated even in patients pre-treated with known cardiotoxic drugs such as anthracyclines.9,43-46,48 This could be ex­plained by the peculiar pharmacokinetics of fluorouracil, that has a brief plasmatic half-life of a few minutes only, but enters various tissues, including the myocardium, from which it is subsequently released in a variable number of days. In continuous Radiol Oncol 2006; 40(3): 149-61. infusions, circadian variations of the hae­matic levels of the drug occur, partly due to fluctuations of the dihydropirimidine­dehydrogenasis (DPD) activity.49 In most cases, the signs of cardiotoxicity appear 2-3 days after the beginning of treatment and they can also persist after the end of the infusion; therefore, only with very el­evated doses of fluorouracil or in the case of continuous infusions can a high con­centration of the drug in the cardiac tissue be sufficient to cause a toxic reaction. The presence of ischemic cardiopathy or ECG alterations of an ischemic type may identify a group of patients with a more elevated risk of cardiotoxicity, but not all authors agree with this.8,9,10 Hereditary congenital DPD defects can also cause significant fluorouracil toxicity, which is, however, mostly haematological.51 Generally, fluorouracil cardiotoxicity causes angina, with or without ECG signs of an acute heart attack or with ECG signs of ischemia in the absence of typical pain. Sudden death and arrhythmia, especially ventricular arrhythmia, follow in order of frequency. In several cases, ventricular ar­rhythmia and death have been described after the appearance of acute myocardial ischemia, while in other cases, sudden death occurred in the absence of cardio-logical follow-up and might have been due to a myocardial infarction. In conclusion, it can be hypothesized that in at least 90% of cases of toxicity, the primary cause is due to ischemia.28,47,51, 52 More rarely, toxicity appears in the form of cardiogenic shock, heart failure and a myocarditis type syndrome.47,51 The major­ity of events appear during the first cycle of chemotherapy, and in more than half of the cases, within 72 hours from the beginning of the infusion, but in studies with a direct follow-up of the patients, the events oc­curred on the third or fourth day of a high dose continuous infusion.28,51,52 The ECG alterations consist either in the elevation or in the depression of the ST tract, diffuse or localized.33,34,40,41 An onset with ST elevation leads to the sup­position that vasospasm is the cause of the ischemia, as confirmed in some cases by angiographic documentation and experi­mental studies.53-56 An interesting finding is the sporadically reported induction of vasospastic effort angina during the fluo­rouracil infusion. This event is probably underdiagnosed because of the generally scanty follow-up of outpatients.57 The mechanism of fluorouracil cardio-toxicity has not yet been entirely clarified, and could also be linked to different events which are more or less predominant in different patients. For example, the mecha­nisms underlying the induction of vaso-spasm are little known. Endothelin, that has been found to be particularly elevated in patients with tumors and fluorouracil toxicity, could possibly be a mediator, but it is also possible that other still unknown va­soactive compounds could be involved.57,58 In some cases of vasospastic effort angi­na, ischemia does not appear during stress but on recovery; therefore hyperventilation could play some role.52,59 Other factors that could explain this phenomenon include the formation of thrombi, an increase in the oxygen requirement of myocytes from the inotropic and positive chronotropic effect, an interference with cellular me­tabolism, ATP depletion, inhibition of the tricarboxylic acid cycle, delayed immune reactions with lymphokine activation and cellular toxicity like that induced by anthra­ cyclines.44,45,47,60 A specific therapy does not exist. The use of nitroderivatives, calcium channel blockers or a combination of these could be effective in the management or pre­vention of a recurrence of the toxicity in some patients; in others these measures could be completely or partially ineffective. Radiol Oncol 2006; 40(3): 149-61. Arrhythmia and cardiogenic shock could be resistant to all conventional therapies or regress with a simple type of support­ive therapy. As a rule, the discontinuation of fluorouracil leads to the regression of symptoms within 48-72 hours.9,28,43,51,52,5 4,61 Once a patient has manifested cardio-toxicity, each further fluorouracil admin­istration, even with the protection of anti-anginal drugs, carries an elevated risk of a repetition of the cardiotoxicity; therefore, a modification of the dose or of the method of administration can sometimes be effec­ tive.28,44,51 Taxanes Hypokinetic arrhythmia, particularly oligo- or asymptomatic sinus bradycardia but occasionally also transient atrioventricular II or III grade blocks, supraventricular and ventricular arrhythmia, depression of the systolic function and myocardial infarction have occurred in some studies involving paclitaxel, but in others no significant car-diotoxicity was reported.62-67 Some toxic effects, mainly arrhythmia, are analogous to those observed after ac­cidental poisoning with parts of the yew plant and are probably due to paclitaxel itself. Others, particularly myocardial isch­emia, ventricular arrhythmia and hypoten­sion, may be due to the action of cremo­phore, the paclitaxel carrier, through the induction of histamine release, or to other substances used in the premedication.64 The combination of paclitaxel with doxorubicin causes an increased incidence of cardiotoxicity. It has been suggested that the interaction between the two agents, with delayed elimination of doxorubicin when administrated after paclitaxel, is re­sponsible for this effect.68 Docetaxel, in­stead, does not seem to have cardiotoxic effects. The mechanism of paclitaxel cardiotox­icity is strictly related to its antitumoral activity. In fact, the antimitotic effect is due to its ability to stabilize the microtubules. At much higher doses than those effective in neoplastic cells, the same action occurs in the cardiac cells, affecting its elastic and electromechanical properties as seen in hypertrophy from fluid overload.67,69 The effect would be a decrease of the spontane­ous contraction frequency and a greater susceptibility to arrhythmia, without sig­nificant modifications of the contractil­ity or muscle compliance; however, if the dose and time of exposure to paclitaxel is increased, a depression of contractility has been observed.67,69 It has also been suggested that the car-diotoxicity of taxanes is due to coronary vasoconstriction through a mechanism that is independent from the action on microtu­bules. This effect is observed only with pa-clitaxel and not with other analogues such as docetaxel, which has the same antimi­totic mechanism. The partial discrepancy of the data can perhaps be explained by the different methods of drug administra­tion such as the dose, duration of the infu­sion and its association with other drugs. However, a general consensus seems to exist on the fact that the cardiotoxic effects of taxanes are transient and that the cel­lular dysfunction does not lead to necrosis as happens with anthracyclines; as a result, toxicity is not cumulative. Some toxic ef­fects such as vasomotor phenomena, dys­pnea and respiratory distress syndrome have an allergic origin, and are probably due to the drug carrier (Cremophor) or to the excipients rather than to the molecule itself; these symptoms generally respond to steroid treatment. It is, therefore, not necessary to submit all patients to a car-diological follow-up, but only those with bradyarrhythmia or conduction anomalies. The exclusion from taxane treatment of Radiol Oncol 2006; 40(3): 149-61. all patients with a cardiological history is probably not justified. Other drugs Effects like bradycardia, ventricular ar­rhythmia, heart failure, hypertension and myocardic ischemia have been ascribed to cisplatin and carboplatin. Some of these side effects have been described after poly-chemotherapies, generally in association with cyclophosphamide, vinca alkaloids, etoposide or fluorouracil, in which the role of platinum compounds is debatable. However, cisplatin can give haemodynam­ic and arrhythmic problems. The drug is nephrotoxic and therefore substantial amounts of hydration are given before and after its administration. In subjects with unstable cardiac compensation, hyperten­sion or who have suboptimal compliance of the left ventricle, this could favour an acute pulmonary edema and/or a hypertensive crisis. The toxic effect on the kidney could explain other events such as ventricular arrhythmia, caused perhaps by hypomag­nesemia and hypokalemia, a particularly frequent side effect, and hypertension.70-73 In the more recent prospective studies in which the prevention of dysionia was pre­scribed, cardiac toxicity was not a problem. Moreover, carboplatin appears to be gener­ally less toxic than cisplatin.74-76 Following gemcitabine therapy, arrhyth­mia, particularly atrial fibrillation, hyper­tension and heart failure have been de­scribed, but the role of gemcitabine as a cause of these phenomena is rather specu­lative.77,78 Cyclophosphamide can cause cardiotox­icity by damaging the capillary endotheli-um and also causing microthrombosis with subsequent hemorrhagic and ischaemic myocarditis, but this mainly concerns treat­ment with high doses of the drug.79 Important cardiotoxicity with arrhyth­mia and acute ischemic cardiopathy have been observed following cytokine treat­ment, in particular with intravenously ad­ministered interleukin-2 (IL-2) and a-in­terferon. The cause seems to be mainly capillary leak syndrome in the case of IL-2 and a direct toxicity or vasculitis for inter­feron. Currently, following the subcutane­ous administration of these drugs, their cardiotoxicity appears to be rare.80 Following the administration of vinorel­bine, cases of myocardial infarction have been reported, and, when associated with trastuzumab, cases of depression of the pump function and of heart failure have been reported.81 A more frequent problem is acute dyspnea, often associated with a hypertensive crisis and sometimes with thoracic pain generally related to broncho-spasm or respiratory distress that disap­pears with the administration of oxygen, bronchodilators and cortisone. The ECG is usually normal. Bleomycin does not have any particular cardiotoxicity, but is important with regard to the problems of differential diagnosis. This drug is an integral part of therapeutic regimens used in Hodgkin’s lymphoma, head and neck cancer and testicular cancer. In the most favourable forms of Hodgkin’s disease and testicular cancer, the percent­age of cure is higher than 90%. The latter tumors are particularly frequent in young people with a long life expectancy in whom late toxicities are of paramount importance. The main dose-limiting effect of bleomycin is pulmonary toxicity (25-30% acute and 7-10% chronic).82-87 Often, even long after treatment discontinuation, the patient may have to be referred to a cardiologist because of dyspnea. In these cases, the dose of anthracyclines and/or mediastinal radio­therapy need to be assessed, and a cardio-logical evaluation and respiratory function tests should be performed. The main risk Radiol Oncol 2006; 40(3): 149-61. factors for bleomycin pneumopathy are age > 40 years, cumulative dose > 300 mg and concomitant radiotherapy to the lungs or platinum chemotherapy. Cardioprotective agents A number of methods to reduce the risk of anthracycline cardiopathy have been sug­gested. The most widely used is the limita­tion of the cumulative dose of various drugs: 550 mg/m2 for adriamycin, 600 mg/m2 for daunorubicin, 1000 mg/m2 for epirubicin, 1900 mg/m2 for zorubicin and 160 mg/m2 for mitoxantrone.3 When an association with other antineoplastic cardiotoxic drugs or with radiotherapy to the mediastinum is scheduled, lesser doses should be used. The administration of anthracyclines in smaller, more frequent doses and/or the prolongation of the time of infusion to 48-96 hours can reduce the cardiotoxic risk linked to elevated plasma concentrations of the drug. The use of less cardiotoxic deriva­tives of first-generation anthracyclines such as idarubicin, epirubicin or mitoxantrone has been suggested and performed in the clinical setting. However, second-genera­tion anthracyclines can also cause cardio-toxicity, though at higher doses.18,21,22 The damage is cumulative even between differ­ent anthracyclines. Liposomes, used as adriamycin carri­ers23 have been elaborated to lessen their toxic effect on healthy tissues; however, in vivo studies have not shown the same degree of cardiotoxicity prevention noted in in vitro studies. Besides, the elevated cost of the liposomal formulations limit their generalized use. Many clinical studies have identified substances which are able to protect the myocardium from anthracycline toxicity without reducing their antineoplastic ac­tivity.88,89 Cytoprotector agents such as ICRF187 (dexrazoxane) at doses of 20:1 or 10:1 of the dose of adriamycin can reduce cardiac events to 30-50% and anthracy­cline cardiopathy to 10-15% of the original risk.90,91 This intracellular iron chelator re­duces the formation of free radicals92 and promotes an immunomodulating effect on the myocardial inflammation caused by anthracyclines.93 Although clinical stud­ies have shown the effectiveness of this substance in preventing the initial car-diotoxicity from anthracyclines, there are no current randomized studies that have determined its role in the prophylaxis of myocardial damage in the long term.93 Other studies have shown that dexrazox­ane at high doses (>900 mg/m2) could have a counter effect on the antineoplastic activity of adriamycin and epirubicin94, in­cluding increasing the systemic clearance of the drug95,96; in addition, it can cause bone marrow toxicity and phlebitis at the administration site.93,97 For these reasons, dexrazoxane is not used extensively in clinical practice, but mostly in patients with previous cardiac damage who have to receive further doses of anthracyclines. Other substances such as vitamin E, pro-bucol, ascorbic acid, melatonin and other antioxidants have not yet shown an ad­equate in vivo cardioprotective effect. Presently, the prevention of anthracy­cline cardiopathy is based on the cardiologi-cal follow-up of the patient. A knowledge of the possible cardiovascular risk factors, a sound clinical evaluation and the measure­ment of the left ventricular function before, during and after chemotherapy may lead to an early diagnosis of cardiotoxicity. In this case, the reduction or discontinuation of the drug and/or the beginning of an ade­quate cardioprotective therapy may improve and normalize the cardiac function. In this phase, the collaboration with the oncologist is of particular importance to clarify the risk/benefit ratio and to reduce, discontinue Radiol Oncol 2006; 40(3): 149-61. or modify the cardiotoxic regimen. Regarding trastuzumab, an in vitro study on human cardiomyocytes has shown a partial reversibility of the toxicity of the drug with the addition of recombinant neuregulines.41 Periodic clinical and in­strumental controls are advisable after 1-2 months and then every 6 months if there are no problems, more often if signs of tox­icity are present. In the case of a left ven­tricular dysfunction, chemotherapy should immediately be discontinued and the usual medical therapy for heart failure begun, that is, diuretics, ACE-inhibitors, digitalis and beta-blockers when necessary, with a follow-up planned after a few weeks. The discontinuation of trastuzumab may result in a complete recovery of the cardiac func­tion and the suspension of the cardiological therapy. In the event of trastuzumab car-diotoxicity, it should be permanently dis­continued. Antidotes for cardiotoxicity are not yet available. It has also been suggested that the exposure to trastuzumab for a long time may induce apoptosis in neoplastic cells, but this mechanism has not yet been shown in the myocardium.98 Patients scheduled for therapy with flu-orouracil should undergo an initial cardio-logical evaluation to rule out an ischemic cardiopathy. Patients with a higher risk should have a cardiological check-up 2-3 days after the start of the infusion if high doses are used (>800 mg/m2) or after two weeks in the event of chronic low doses, and, in the case of angina, a stress test should be performed. For patients with pre-existing ischaemic cardiopathy or with signs of cardiotoxicity from fluorouracil, the use of alternative drugs should be en­couraged. When fluorouracil is essential, it should be administered in a protected envi­ronment with continuous ECG monitoring and the association of nitrates and calcium antagonists. A modification of the thera­peutic scheme such as administering low doses in weekly boluses rather than in con­tinuous infusion could also be helpful.58,62 When, despite these precautions, serious toxicity appears, fluorouracil should be no longer be administered. New agents The cardiotoxic effects of some new anti-neoplastic agents are currently little known. As a matter of fact, the recent introduction of these drugs in clinical practice does not permit an adequate estimation of their car-diotoxic effects and further evaluation and investigation will be necessary. Examples of these drugs include monoclonal antibodies such as rituximab, a chimeric murine/human monoclonal antibody against the CD20 anti­gen on normal and malignant B-lymphocytes, bevacizumab, a monoclonal antibody that blocks vascular endothelial growth factor (VEGF) receptors, cetuximab, a chimeric anti­body targeting the human epidermal growth factor (EGFR) receptor, and tyrosine kinase inhibitors such as imatinib, gefitinib and er-lotinib and other classes of agents. The identification of novel molecular targets will increase the number of drugs available for the treatment of neoplastic disease. It will be important to evaluate the side effects related to treatment, particu­larly in organs with a limited regenerative capability such as the heart. Further stud­ies will therefore be necessary. Acknowledgments The authors thank Anna Maria Colussi for assisting with the preparation of the manuscript. References 1. Allen A. The cardiotoxicity of chemotherapeutic drugs. 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Randomized com­parison of cisplatin plus fluorouracil and carbo-platinum plus fluorouracil versus methotrexate in advanced squamous-cell carcinoma of the head and neck: a Southwest Oncology Group Study. J Clin Oncol 1992; 10: 1245-51. 74. Spaulding MB, Fisher SG, Wolf GT. Tumor re­sponse, toxicity, and survival after neoadjuvant organ-preserving chemotherapy for advanced la­ryngeal carcinoma. J Clin Oncol 1994; 12: 1592-9. 75. Birkenhake S, Leykamm S, Martus P, Sauer R. Concomitant radiochemotherapy with 5-FU and cisplatin for invasive bladder cancer. Acute toxic­ity and first results. Strahlentherapie und Onkologie 1999; 175: 97-101. Radiol Oncol 2006; 40(3): 149-61. 76. Sauer-Heilborn A, Kath R, Schneider CP, Hoffken K. Severe non-haematological toxicity after treat­ment with gemcitabine. J Cancer Res Clin 1999; 125: 637-40. 77. Bengala C, Danesi R, Guarneri V, Pazzagli I, Donati S, Favre C, et al. High-dose consolida­tion chemotherapy with idarubicin and alkylating agents following induction with gemcitabine-epi­rubicin-paclitaxel in metastatic breast cancer: a dose finding study. Bone Marrow Transpl 2003; 31: 275-80. 78. Tolba KA, Deliargyris EN. Cardiotoxicity of cancer therapy. Cancer Invest 1999; 17: 408-22. 79. Taniguchi I. Clinical significance of cyclophospha­mide-induced cardiotoxicity. Inter Med 2005; 44: 89-90. 80. Eskander ED, Harvey HA, Givani E, Lipton A. Phase I study combining tumor necrosis factor with interferon-alpha- and interleukin-2. Am J Clin Oncol 1997; 20: 511-4. 81. Burstein HJ, Harris LN, Marcom PK, Lambert-Falls R, Havlin K, Overmoyer B, et al. Trastuzumab and vinorelbine as first-line therapy for HER2-over-expressing metastatic breast cancer: multicenter phase II trial with clinical outcomes, analysis of serum tumor markers as predictive factors, and cardiac surveillance algorithm. J Clin Oncol 2003; 21: 2889-95. 82. Gobbi PG, Broglia C, Merli F, Dell’Olio M, Stentano C, Iannitto E, et al. Vinblastine, bleomicyn, and methotrexate chemotherapy plus irradiation for patients with early-stage, favorable Hodgkin lym­phoma. Cancer 2003; 98: 2393-401. 83. Chaudhary UB, Haldas JR. Long-term complica­tions of chemotherapy for germ cell tumours. Drugs 2003; 63: 1565-77. 84. Sleijfer S. Bleomycin-induced pneumonitis. Chest 2001; 120: 617-24. 85. O’Sullivan JM, Huddart RA, Norman AR, Nicholls J, Dearnaley DP, Horwich A. Predicting the risk of bleomycin lung toxicity in patients with germ-cell tumours. Ann Oncol 2003; 14: 91-6. 86. Diehl V, Franklin J, Pfeundschuch M, Lathan B, Paulus U, Hasenclever D, et al. Standard and in-creased-dose BEACOPP chemotherapy compared with COPP-ABVD for advanced Hodgkin’s dis­ease. New Engl J Med 2005; 18: 353:744. 87. Aleman BMP, Raemaekers JMM, Tirelli U, Bortolus R, van’t Veer MB, Lybeert ML, et al. Involved-field radiotherapy for advanced Hodgkin’s lymphoma. 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The interaction of the cardioprotec­tive agent ICRF-187 ((+)-1,2-bis (3,5-dioxopipera­zinyl-1-yl) propane); its hydrolysis product (ICRF­198); and other chelating agents with the Fe (III) and Cu (II) complexes of adriamycin. Agents Action 1990; 29: 374-81. 93. Lipshultz S. Dexrazoxane for protection against cardiotoxic effects of anthracyclines in children. Editorial. J Clin Oncol 1996; 14: 332-3. 94. Sehested M, Jensen PB, Sorensen BS, Holm B, Friche E, Demant EJ. Antagonistic effect of the cardioprotector (+)-1,2-bis (3,5-dioxopiperazinyl-1-yl) propane (ICRF-187) on DNA breaks and cytotoxicity induced by the topoisomerase II di­rected drugs daunorubicin and etoposide (VP-16). Biochem Pharmacol 1993; 46: 389-93. 95. Herman EH, Zhang J, Chadwick DP, Ferrans VJ. Comparison of the protective effects of amifostine and dexrazoxane against the toxicity of doxoru­bicin in spontaneously hypertensive rats. Cancer Chemother Pharmacol 2001; 45: 329-34. 96. Basser RL, Sobol MM, Duggan G, Cebon J, Rosenthal MA, Mihaly G, et al. Comparative study of the pharmacokinetics and toxicity of high-dose epirubicin with or without dexrazoxane in patients with advanced malignancy. J Clin Oncol 1994; 12: 1659-66. 97. Pedersen-Bjergaad J. The dioxopiperazine deri­vates, their leukemogenic potential and other biological effects. Leukemia Res 1992; 16: 1057-9. 98. Henson ES, Hu X, Gibson SB. Herceptin sensitizes ErbB2-overexpressing cells to apoptosis by reduc­ing antiapoptotic Mcl-1 expression. Clin Cancer Res 2006; 12: 845-53. Radiol Oncol 2006; 40(3): 149-61. review Electrochemotherapy of tumours Gregor Serša1, Maja Cemažar1, Damijan Miklavcic2 and Zvonimir Rudolf1 1Institute of Oncology Ljubljana, 2University of Ljubljana, Faculty of Electrical Engineering, Ljubljana, Slovenia Electrochemotherapy consists of chemotherapy followed by local application of electric pulses to the tumour to increase drug delivery into cells. Drug uptake can be increased by electroporation for only those drugs whose transport through the plasma membrane is impeded. Among many drugs that have been tested so far, only bleomycin and cisplatin found their way from preclinical testing to clinical trials. In vitro studies demonstrated several fold increase of their cytotoxicity after electroporation of cells. In vivo, electroporation of tumours after local or systemic administration of either of the drugs, i.e. electrochemotherapy, proved to be an effective antitumour treatment. In preclinical studies on several tumour models, electrochemotherapy either with bleomycin or cisplatin was elaborated and parameters for effective local tumour control were determined. In veterinary medicine, electrochemotherapy also proved to be effective in the treatment of primary tumours in cats, dogs and horses. In human clinical studies, electrochemotherapy was performed on the patients with progressive disease and accessible tumour nodules of different malignancies. All clinical studies demonstrated that electrochemotherapy is an effective treatment for local tumour control in cancer patients. Key words: neoplasms – drug therapy; electroporation; electrochemotherapy, drug delivery systems; bleomycin; cisplatin, Introduction Treatments for cancer may be divided into different categories based on their goals and mode of action. Very often the differ­ent types of treatment are used in combina- Received 28 March 2006 Accepted 15 June, 2006 Correspondence to: Prof. Gregor Serša, Institute of Oncology Ljubljana, Department of Experimental Oncology, Zaloška 2, SI-1000 Ljubljana, Slovenia. Phone/ Fax: +386 1 5879 434; E-mail address: gsersa@onko-i.si tion, either simultaneously or sequentially. In general, cancer treatment includes three main treatment modalities, surgery and ra­diation, which are local treatment modali­ties and chemotherapy which is a systemic treatment modality. Chemotherapy, a systemic treatment modality for cancer is effective if the drugs that have intracellular targets readily pass the plasma membrane. However, among highly cytotoxic chemotherapeutic drugs there are some whose transport through the plasma membrane is hampered. These drugs are good candidates for electroche­motherapy. Electrochemotherapy is a local treatment combining chemotherapy and application of electric pulses to the tumour, thus increasing plasma membrane perme­ability. In electrochemotherapy, the optimal antitumour effectiveness is achieved when electric pulses are given at the time of the highest extracellular concentration of hydrophilic chemotherapeutic drug in the tumour, thereby increasing their transport through the plasma membrane towards their intracellular targets.1-4 Preclinical data In vitro studies Electroporation proved to be effective in facilitating the transport of different molecules across the plasma membrane. Different biochemical and pharmacological studies on chemotherapeutic drug trans­port facilitated by means of electropora­tion, report that the increased intracellular drug accumulation improves the cytotoxici­ty of the drug. Since electroporation can fa­cilitate the drug transport through the cell membrane only for poorly or non-permeant molecules, suitable candidates for electro-chemotherapy are limited to those drugs that are hydrophilic and lack transport sys­tem in the membrane. Several chemothera­peutic drugs were tested in vitro on cells for potential application in combination with electroporation; some of them are dauno­rubicin, doxorubicin, etoposide, paclitaxel, actinomycin D, adriamycin, mitomycin C, 5-fluorouracil, vinblastine, vincristine, gem-citabine, cyclophosphamide, carboplatin, cisplatin and bleomycin. Electroporation of cells increases the cytotoxicity of some of these drugs ranging from 1.1 to up to several thousand fold. However, only two of these drugs have been identified as potential candidates for electrochemo-therapy of cancer patients.1,2,4 The first is bleomycin; it is hydrophilic and has very restricted transport capacity through the cell membrane, thus its cytotoxicity can be potentiated up to several 1000 fold by electroporation of cells. Few hundred internalized molecules of bleomycin are sufficient to kill the cell. 1,2,5 The second is cisplatin whose transport through the cell membrane is also hampered. Only 50% of cisplatin is transported through the plasma membrane by passive diffusion, the rest is transported by carrier molecules. The overall flux across the plasma membrane is thus limited. Electroporation of the plasma membrane enables greater flux and accu­mulation of the drug in the cells which re­sults in increase of cisplatin cytotoxicity by up to 80-fold.3-6 These promising preclinical data obtained in vitro on a number of differ­ent cell lines have paved the way for testing these two drugs in electrochemotherapy in vivo on different tumour models. In vivo studies Bleomycin and cisplatin were tested in elec­trochemotherapy protocol on a number of animal models in vivo (Figure 1). Extensive studies on different animal models with different tumours, either transplantable or spontaneous were performed. Antitumour effectiveness of electrochemotherapy was demonstrated on tumours in mice, rats, hamsters, cats and rabbits. Tumours treated by electrochemotherapy were either subcu­taneous, grew in the muscle, brain or in the liver, and were of different types, e.g. sarco­mas, carcinomas, glioma or melanoma.1,2,4 In these studies, different factors con­trolling antitumour effectiveness were de­termined: The drugs can be given by different routes of administration, they can be injected either intravenously or intra- tumourally. The prerequisite is that, at the time of the application of elec­ tric pulses to the tumour, a sufficient Radiol Oncol 2006; 40(3): 163-74. amount of drug is present in the tu­mour. Therefore, after intravenous drug administration into small laboratory animals (4 mg/kg of cisplatin or 0.5 mg/kg bleomycin), only a few minutes’ interval is needed to reach the maximal drug concentration in the tumours. After intratumoural administration, this interval is even shorter and the ap­plication of electric pulses has to follow the administration of the drug as soon as possible (within a minute). 1,2,4 Good antitumour effectiveness may be achieved by good tissue electropo-ration. The plasma membrane elec­troporation is obtained if the cell is exposed to a sufficiently high electric field. This depends on the electric field distribution in the tissue which is control­led by the geometry of electrodes and tissue. The electric field distribution in the tissue and cell electroporation can be improved by rotating electric field. Surface tumours can be effective­ly treated by plate electrodes, whereas appropriate electric field distribution in deeper parts of the tumour is assured by using needle electrodes.7-11 The antitumour effectiveness is de­pendent on the amplitude, number and duration of the electric pulses applied. Several studies in which parallel plate electrodes were used for surface tu­mours showed that an amplitude over distance ratio above 1000 V/cm is need­ed for tumour electroporation, and that above 1500 V/cm, irreversible changes in the normal tissues adjacent to the tumour occur; so, the window for ef­fective and safe electrochemotherapy is between 1000 -1500 V/cm. in most studies the amplitude over distance ratio of 1300 V/cm induced good anti-tumour effectiveness without sub-op­timal electroporation of the tissue or damage to the tissue due to irreversible cell permeabilization.7 For other types of electrodes, the electric field distribu­tion and thus also the necessary ampli­tude of electric pulses need to be de­termined by numerical calculations.10 Repetition frequencies of the pulses for electrochemotherapy are either 1 Hz or 5 kHz. The minimal number of the pulses used is 4; most studies use 8 electric pulses of 100 µs.11,12 Figure 1. Protocol of electrochemotherapy of ex­perimental tumours presented schematically (A). The drug is injected either intravenously or intratumorally, at the doses that do not exert antitumour effect. After the interval that allows sufficient drug accumulation in the tumours, electric pulses are applied to the tu­mour either by plate or needle electrodes (1300 V/cm, 100 µs, 1 Hz or 5 kHz, 8 pulses). The plate electrodes are placed in that way that the whole tumour is en­compassed between the electrodes, providing good electric field distribution in the tumours for an opti­mal electroporation of the cells in the tumours (B). Radiol Oncol 2006; 40(3): 163-74. All the experiments conducted in vivo on animals provided sufficient data to dem­onstrate that electrochemotherapy with ei­ther bleomycin or cisplatin is effective in the treatment of solid tumours, using drug concentrations which without application of electric pulses have no or minimal anti-tumour effect. Already a one-time treatment by electrochemotherapy induces partial or complete regression of tumours, whereas the treatment with bleomycin or cisplatin alone or application of electric pulses alone has no or minimal antitumour effect (Figure 2). Mechanisms of action The principal mechanism of electrochemo-therapy is electroporation of the cells in the tumours, which increases the drug effec­tiveness by enabling the drugs to reach its intracellular targets. This was demonstra­ted in the studies that measured the intratu­moural drug accumulation and the amount of the drug bound to DNA. Basically, the amounts of bleomycin and cisplatin in the electroporated tumours were up to 2-4 fold higher than in those without application of electric pulses.13,14,15 Figure 2. Example of good antitumour effectiveness of electrochemotherapy with cisplatin on SA-1 tumours. Cisplatin was given intravenously (4 mg/kg), 3 min thereafter 8 electric pulses were applied to the tumour with plate electrodes. Electric pulses were applied in two directions; 4 pulses in one and the other 4 in the perpendicular direction. Eight days after the treatment, good antitumour effectiveness of electrochemotherapy with cisplatin is evident, compared to the treatments with cisplatin or electric pulses alone. Radiol Oncol 2006; 40(3): 163-74. Besides membrane electroporation, which facilitates drug transport and its accumu­lation in the cell, other mechanisms that are involved in antitumour effectiveness of electrochemotherapy were described. The application of electric pulses to the tissues induces a transient, and reversible reduction of blood flow.16 The restoration of the blood flow in normal tissue is much faster than of that in tumours.17 The decrease in tumour blood flow induces drug entrapment in the tis­sue, providing more time for the drug to act. Besides, this phenomenon prevents bleeding from the tissue, which is important in clini­cal situations of haemorrhagic tumours.17 The cytotoxic effect of electrochemo-therapy is not limited only to tumour cells in the tumours. Electrochemotherapy acts also on stromal cells, including endothelial cells in the lining of tumour blood vessels.5 This represents yet another mechanism involved in the antitumour effectiveness of electrochemotherapy, i.e. vascular disrupting effect.18 The difference in antitumour effective­ness of electrochemotherapy was observed between immunocompetent and immuno­deficient experimental animals, indicating to the involvement of immune response in antitumour effectiveness.19,20 Due to the massive tumour antigen shedding in the organisms after electrochemotherapy, sys­temic immunity can be induced, and up-regulated by additional treatment with bio­logical response modifiers like IL-2, GM­ 21-23 CSF and TNF-a. To sum up, the electrochemotherapy pro­tocol was optimized in preclinical studies in vitro and in vivo, and basic mechanisms were elucidated. In addition to the electropora­tion of cells, the tumour drug entrapment, vascular disrupting effect and involvement of immune response were also demonstrat­ed. Based on all these data, electrochemo-therapy with bleomycin and cisplatin was promptly evaluated in clinical trials. Other biomedical applications of electroporation and electrochemotherapy in cancer treatment Knowledge about the mechanisms involved in the antitumour effectiveness of electro-chemotherapy opened new possibilities for the application of electric pulses or electro-chemotherapy in the treatment of cancer. The chemotherapeutic drugs that in­crease effectiveness of radiation therapy are radiosensitizing drugs. Among them are also bleomycin and cisplatin. Since drug delivery induced by electroporation is site-specific, it could be used for tumour-specific delivery of radiosensitizing drugs. By the increased radiosensitizing drug de­livery into the tumours and not in the surrounding normal tissue the therapeutic index of tumour irradiation is increased. In our recent studies, we combined electro-chemotherapy either with bleomycin or cis­platin with radiotherapy and demonstrated a good potentiation of tumour radiation response: 1.9 fold for electrochemotherapy with bleomycin and 1.6 fold for electroche­motherapy with cisplatin.24-26 The application of electric pulses was shown to modulate the tumour blood flow. Both, reduced blood flow and lowered par­tial oxygen pressure (pO2) in the tumours are consequences of the applied electric pulses.18 The reduced pO2 can activate biore­ductive drugs to exhibit cytotoxic effect on hypoxic cells.27 In well oxygenated cells, the drug remains inactive. On the other hand, tumour hypoxia induced by application of electric pulses can improve therapeutic con­ditions for the use of hyperthermia since tumour cells are more sensitive to heat in sub-optimal physiological conditions.28 Electrochemotherapy with cisplatin or bleomycin was successfully used also in the veterinary medicine. It was used to treat different tumours, such as mammary adenocarcinoma, fibrosarcoma, cutaneous Radiol Oncol 2006; 40(3): 163-74. mast cell tumour, hemangioma, hemangiosa­rcoma, perianal tumours, neurofibroma and sarcoids in dogs, cats, hamsters, rabbits and horses.29-33 Recent reports demonstrated a successful treatment of different neoplasms in companion animals and sarcoids in hor­ses.30-33 Hopefully, electrochemotherapy will be broadly used in veterinary medicine for the treatment of different malignancies, both primary and metastatic disease. Electrochemotherapy is an effective cy­toreductive treatment; however, its curative effect is dependent on the permeabilisation of possibly all cells in the tumours. Since permeabilisation of every single cell in the tumour is virtually impossible, electroche­motherapy could be combined with other cy­toreductive treatments. Another approach is a combination of electrochemotherapy with electrogene therapy. The first promising re­ports and data are already available, support­ing the effectiveness of this concept. 23,34 In conclusion, the electroporation in electrochemotherapy has already been very well exploited; however, there are new bio­medical applications of electroporation in cancer treatment that still need testing and development. Clinical studies on electrochemotherapy The first clinical study on electrochemo-therapy was published in 1991, reporting good treatment effectiveness of electro-chemotherapy on cutaneous tumour nod­ules of head and neck tumours.35 The results of this study by the group from the Institute Gustave Roussy, have stimulated other groups to initiate their own clinical studies. The first clinical centres which performed electrochemotherapy were Villejuif and Toulouse in France, the group in Tampa in USA, and our group at the Institute of Oncology Ljubljana in Slovenia. Recently, also new centres reported clinical experience on electrochemotherapy, e.g. Copenhagen in Denmark, Mexico City in Mexico, Chicago in USA, Vienna in Austria, Matsumoto and Jamagata in Japan, Sydney in Australia and Cork in Ireland.35-63 In all clinical studies, 247 patients were included; 202 patients with 655 tumour nodules were treated by electrochemother­apy with bleomycin and 45 patients with 354 tumour nodules were treated by elec­trochemotherapy with cisplatin. The ma­jority were malignant melanoma patients, and also the patients with metastases in head and neck region, mammary carci­noma, skin cancer, ovarian cancer, Kaposi sarcoma and chondrosarcoma were treated by electrochemotherapy. The results of the studies can be summarized as supporting the assumption that electrochemotherapy has good antitumour effectiveness either using bleomycin or cisplatin, resulting in ~80% objective responses of the treated tumour nodules.3,60 Based on these results, the European project that was aimed at developing and producing electric pulses generator was launched. In the CLINIPORATOR project, this electric pulses generator was developed and is now commercially available for those who would like to perform electrochemother­apy. This generator under the same name as the project - CLINIPORATOR™ (IGEA S.r.l., Carpi, Italy) is certified as a medical devices and is therefore appropriate for clinical use. Along with the development of the electric pulse generator, also plate and needle elec­trodes were developed (Figure 3). The next step was to gather clinical ex­perience of four cancer centres in Villejuif, Copenhagen, Cork and Ljubljana and pre­pare Standard Operating Procedures (SOP) of electrochemotherapy. This was a prereq­uisite step to bring electrochemotherapy into standard clinical practice. SOP is now completed and the drug licensing for elec­trochemotherapy in process, so electroche- Radiol Oncol 2006; 40(3): 163-74. CLINIPORATOR™ Plate electrodes Figure 3. CLINIPORATOR™, an electric pulse gen­erator for clinical use in electrochemotherapy. For ap­plication of electric pulses plate and needle electrodes were developed. motherapy can be used as standard proce­dure for local tumour treatment. Treatment procedures for electrochemotherapy Treatment advantages and clinical uses Electrochemotherapy is used for the treat­ment of cutaneous and subcutaneous tu­mour nodules of different malignancies. The treatment advantages and clinical uses for electrochemotherapy can be summarized: easy and effective treatment of single or multiple tumour nodules of any histol­ogy in the cutaneous and subcutaneous tissue,3,60 treatment that improves quality of life of patients with progressive disease,3,60 treatment of choice for tumours refrac­tory to conventional treatments,3,60 neoadjuvant treatment in form of cy­toreductive therapy before conventional treatment,62 organ sparing and function saving treat­ment,62,63 treatment of hemorrhagic or pain­ful nodules, since it reduces bleed­ing and in some cases pain level.50,62 Treatment procedure The treatment procedure is as follows: based on SOP, tumour nodules can be treated by electrochemotherapy with in­jection of bleomycin intravenously or in-tratumourally and by electrochemother­apy with cisplatin given intratumourally. The choice of the chemotherapeutic drug in not based on tumour histology, but depends on the number and size of the nodules. After drug injection the tumour nodules are exposed to electric pulses. The interval between the intravenous drug injection and application of electric pulses is 8-28 min, and after the intratumoural in­jection, as soon as possible. Different sets of electrodes are available for application; plate electrodes for smaller tumour nod­ules and needle electrodes for the treat­ment of larger (3 cm) and thicker tumour nodules. The treatment can be performed in one-session or can be repeated in case of new emerging nodules or on those nod­ules that relapsed in some regions not well treated in the first treatment. Electrochemotherapy does not induce side effects due to chemotherapeutic drugs since the drug dosage is very low. However, the application of electric pulses to the tumours induces contraction of the under­lying muscles. For electroporation, square wave electric pulses of the amplitude over distance ration of 1000-1300 V/cm, dura­tion of 100 µs, frequency 1 Hz or 5 kHz are used. These muscle contractions are painful, but the pain dissipates imme­diately after electric pulses application. Nevertheless, in SOP, the procedures for alleviating the pain by local anaesthesia Radiol Oncol 2006; 40(3): 163-74. Figure 4. Antitumour effectiveness of electrochemotherapy with intratumoural injection of cisplatin in a subcuta­neous metastasis of the patient with malignant melanoma. Twelve weeks after the treatment the tumour nodule was in complete response (CR), with pigmentation and good cosmetic effect. or by general anaesthesia in case of treat­ing multiple nodules are also described. Treatment effectiveness The treatment after single electrochemo-therapy session results in most cases in complete tumour eradication. When ne­cessary, treatment can be repeated at 4­8 weeks intervals with equal antitumour effectiveness. The treatment has a good cosmetic effect without scaring the treated tissue (Figure 4,5). Conclusion Electrochemotherapy is now on the verge being standard treatment in palliative treatment of cutaneous and subcutane­ous tumour nodules of different malig­nancies. However, further progress of electrochemotherapy will continue by de­veloping new electrodes that will enable the treatment of larger tumours and tu­mours in internal organs. Consequently, the indications for electrochemotherapy may be extended. Acknowledgement The authors acknowledge the financial support of the state budget by the Slovenian Research Agency (programme No. P3-0003; project No. J3-7044) and EU funded project ESOPE (QLK-2002-02003). Radiol Oncol 2006; 40(3): 163-74. Figure 5. Antitumour effectiveness of electrochemotherapy with intratumour­al injection of cisplatin of malignant melanoma tumour nodules after repeated treatments. Nodule No.1 was treated only once, and was in complete remission one year after the treatment. Nodule No. 2 was treated three times at two-month intervals. After each treatment session, good antitumour effect was observed, but the tumour relapsed. After the third treatment, no recurrence was observed in one-year observation period. References 1. Mir LM. Therapeutic perspectives of in vivo cell electropermeabilization. Bioelectrochem 2001; 53: 1-10. 2. Gehl J. Electroporation: theory and methods, perspectives for drug delivery, gene therapy and research. Acta Physiol Scan 2003; 177: 437-47. 3. Sersa G, Cemazar M, Rudolf Z. Electrochemotherapy: advantages and drawbacks in the treatment of patients. Cancer Therapy 2003; 1: 133-42. www.cancer-therapy.org 4. Sersa G. Electrochemotherapy: animal work re­view. In. 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Biochim Biophys Acta 2000: 1519: 73-83. 9. Pavselj N, Bregar Z, Cukjati D, Batiuskaite D, Mir LM, Miklavcic D. The course of tissue permea­bilization studied on a mathematical model of a subcutaneous tumor in small animals. IEEE Trans Biomed Eng 2005: 52: 1373-81. 10. Sel D, Cukjati D, Batiuskaite D, Slivnik T, Mir LM, Miklavcic D. Sequential finite element model of tissue electropermeabilization. IEEE Trans Biomed Eng 2005: 52: 816-27. Radiol Oncol 2006; 40(3): 163-74. 11. Miklavcic D, Pucihar G, Pavlovec M, Ribaric S, Mali M, Macek-Lebar A, Petkovsek M, Nastran J, Kranjc S, Cemazar M, Sersa G. The effect of high frequency electric pulses on muscle contractions and antitumor efficiency in vivo for a potential use in clinical electrochemotherapy. Bioelectrochemistry 2005; 65: 121-8. 12. Sersa G, Cemazar M, Semrov D, Miklavcic D. Changing electrode orientation improves the ef­ficacy of electrochemotherapy of solid tumors in mice. Bioelectroch Bioeneg 1996; 39: 61-6. 13. Belehradek JJr, Orlowski S, Ramirez LH, Pron G, poddevin B, Mir LM. Electropermeabilization of cells and tissues assessed by the quantitative and qualitative electroloading of bleomycin. Biocim Biophys Acta 1994; 1190: 155-63. 14. Cemazar M, Milacic R, Miklavcic D, Dolzan V, Sersa G. Intratumoral cisplatin administration in electrochemotherapy: antitumor effectiveness, sequence dependence and platinum content. Anticancer Drugs 1998; 9: 525-30. 15. Cemazar M, Miklavcic D, Scancar J, Dolzan V, Golouh R, Sersa G. Increased platinum accumula­tion in SA-1 tumour cells after in vivo electroche­motherapy with cisplatin. Brit J Cancer 1999; 79: 1386-1391. 16. Sersa G, Cemazar M, Parkins CS, Chaplin DJ. Tumour blood flow changes induced by application of electric pulses. Eur J Cancer 1999; 35: 672-7. 17. Gehl J, Skovsgaard T and Mir LM. Vascular reac­tions to in vivo electroporation: characterization and consequences for drug and gene delivery. Biochim Biophys Acta 2002; 1569: 51-8 18. Sersa G, Krzic M, Sentjurc M, Ivanusa T, Beravs K, Kotnik V, Coer A, Swartz HM, Cemazar M. Reduced blood flow and oxygenation in SA-1 tumours after electrochemotherapy with cisplatin. Br J Cancer 2002; 87:1047-54. 19. Sersa G, Miklavcic D, Cemazar M, Belehradek JJr, Jarm T, Mir LM. Electrochemotherapy with CDDP on LPB sarcoma: comparison of the anti-tumor ef­fectiveness in immunocompetent and immunodefi­cient mice. Bioelectroch Bioener 1997; 43: 279-283. 20. Sersa G, Kotnik V, Cemazar M, Miklavcic D, Kotnik A. Electrochemotherapy with bleomycin in SA-1 tumor-bearing mice - natural resistance and immune responsiveness. Anti-Cancer Drugs 1996; 7: 785-791. 21. Sersa G, Cemazar M, Menart V, Gaberc-Porekar V, Miklavcic D. Antitumor effectiveness of elec­trochemotherapy is increased by TNF-a on SA-1 tumors in mice. Cancer Letters 1997; 116: 85-92. 22. Mir LM, Rath C, Orlowski S, Quintin-Colona F, Fradelizi D, Belahradek J, Kourilsky P. Systemic antitumor effects of electrochemotherapy com­bined with histoincompatible cells secreting inter­leukin 2. J Immunother 1995; 17: 30-8. 23. Heller L, Pottinger C, Jaroszeski MJ, Gilbert R, Heller R. In vivo electroporation of plasmids encoding GM-CSF or interleukin-2 into existing B16 melanoma combined with electrochemother­apy inducing long-term antitumour immunity. Melanoma Res 2000; 10: 577-83. 24. Sersa G, Kranjc S, Cemazar M. Improvement of combined modality therapy with cisplatin and radiation using electroporation of tumors. Int J Radiat Oncol Biol Phys 2000; 46: 1037-41. 25. Kranjc S, Cemazar M, Grosel A, Scancar J, Sersa G. Electroporation of LPB sarcoma cells in vitro and tumors in vivo increases radiosensitizing ef­fect of cisplatin. Anticancer Res 2003; 23: 275-82. 26. 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. 27. Cemazar M, Parkins CS, Holder AL, Kranjc S, Chaplin DJ, Sersa G. Cytotoxicity of bioreductive drug tirapazamine is increased by application of electric pulses in SA-1 tumours in mice. Anticancer Res 2001; 21: 1151-6. 28. Karner KB, Lesnicar H, Cemazar M, Sersa G. Antitumour effectiveness of hyperthermia is poten­tiated by local application of electric pulses to LPB tumours in mice. Anticancer Res 2004; 24: 2343-8. 29. Mir LM, Devauchelle P, Quintin-Colonna F, Delisle F, Dolinger S, Fradelizi D, Belehradek JrJ, Orlowski S. First clinical trial of cat soft-tissue sarcomas treatment by electrochemotherapy. Br J Cancer 1997; 76: 1617-22. 30. Tozon N, Sersa G, Cemazar M. Electrochemotherapy: Potentiation of local antitumour effectiveness of cisplatin in dogs and cats. Anticancer Res 2001; 21: 2483-6. 31. Rols MP, Tamzali Y, Teissie J. Electrochemotherapy of horses. A preliminary clinical report. Bioelectrochem 2002; 55: 101-5. 32. Spugnini EP, Porrello A. Potentiation of chemo­therapy in companion animals with spontaneous large neoplasms by application of biphasic electric pulses. J Exp Clin Cancer Res 2003; 22: 571-80. Radiol Oncol 2006; 40(3): 163-74. 33. Tozon N, Kodre V, Sersa G, Cemazar M. Effective treatment of perianal tumors in dogs with electro-chemotherapy. Anticancer Res 2005; 25: 839-46. 34. Matsubara H, Maeda T, Gunji Y, Koide Y, Asano T, Ochiai T, Sakiyama S, Tagawa M. Combinatory anti-tumor effects of electroporation-mediated chemotherapy and wild-type p53 gene transfer to human esophageal cancer cells. Int J Oncol 2001; 18: 825-9. 35. Mir LM, Belehradek M, Domenge C, Orlowski S, Poddevin B, Belehradek J Jr, Schwaab G, Luboinski B, Paoletti C. Electrochemotherapy, a new antitu­mor treatment: first clinical trial. C R Acad Sci III 1991; 313: 613-8. 36. Belehradek M, Domenge C, Luboinski B, Orlowski S, Belehradek J Jr, Mir LM. Electrochemotherapy, a new antitumour treatment. First clinical phase I-II trial. Cancer 1993; 72: 3694-700. 37. Rudolf Z, Stabuc B, Cemazar M, Miklavcic D, Vodovnik L, Sersa G. Electrochemotherapy with bleomycin: The first clinical experience in malig­nant melanoma patients. Radiol Oncol 1995; 29: 229-35. 38. Glass LF, Fenske NA, Jaroszeski M, Perrott R, Harvey DT, Reintgen DS, Heller R. Bleomycin-mediated electrochemotherapy of basal cell carci­noma. J Am Acad Dermatol 1996; 34: 82-6. 39. Glass LF, Pepine ML, Fenske NA, Jaroszeski M, Reintgen DS, Heller R. Bleomycin-mediated elec­trochemotherapy of metastatic melanoma. Arch Dermatol 1996; 132: 1353-7. 40. Heller R, Jaroszeski MJ, Glass LF, Messina JL, Rapaport DP, DeConti RC, Fenske NA, Gilbert RA, Mir LM, Reintgen DS. Phase I/II trail for the treatment of cutaneous and subcutaneous tumors using electrochemotherapy. Cancer 1996; 77: 964­71. 41. Reintgen DS, Jaroszeski MJ, Heller R. Electrochemotherapy, a novel approach to cancer. The Skin Cancer Foundation Journal 1996; 14: 17-9. 42. Domenge C, Orlowski S, Luboinski B, De Baere T, Belehradek J-Jr, Mir LM. Antitumor electroche­motherapy. New advances in the clinical protocol. Cancer 1996; 77: 956-63. 43. Glass LF, Jaroszeski M, Gilbert R, Reintgen DS, Heller R. Intralesional bleomycin-mediated elec­trochemotherapy in 20 patients with basal cell carcinoma. J Am Acad Dermatol 1997; 37: 596-9. 44. Mir LM, Glass LF, Sersa G, Teissie J, Domenge C, Miklavcic D, Jaroszeski M-J, Orlowski S, Reintgen DS, Rudolf Z, Belehradek M, Gilbert R, Rols M-P, Belehradek JJr, Bachaud JM, DeConti R, Stabuc B, Cemazar M, Coninx P, Heller R. Effective treat­ment of cutaneous and subcutaneous malignant tumours by electrochemotherapy. Brit J Cancer 1998; 77: 2336-42. 45. Heller R, Jaroszeski MJ, Reintgen DS, Puleo CA, DeConti RC, Gilbert RA, Glass LF. Treatment of cutaneous and subcutaneous tumors with elec­trochemotherapy using intralesional bleomycin. Cancer 1998; 83: 148-57. 46. Panje WR, Hier MP, Garman GR, Harrel E, Goldman A, Bloch I. Electroporation therapy of head and neck cancer. Am Otol Rhinol Laryngol 1998; 107: 779-85. 47. Kubota Y, Mir LM, Nakada T, Sasagawa I, Suzuki H, Aoyama N. Successful treatment of metastatic skin lesions with electrochemotherapy. J Urol 1998; 160: 1426. 48. Sersa G, Stabuc B, Cemazar M, Jancar B, Miklavcic D, Rudolf Z. Electrochemotherapy with cisplatin: potentiation of local cisplatin antitumour effec­tiveness by application of electric pulses in cancer patients. Eur J Cancer 1998; 34: 1213-8. 49. 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. 50. Gehl J, Geertsen P. Efficient palliation of hemor­rhaging malignant melanoma skin metastases by electrochemotherapy. Melanoma Res 2000; 10: 585-9. 51. Rebersek M, Cufer T, Rudolf Z, Sersa G. Electrochemotherapy with cisplatin of breast can­cer tumor nodules in a male patient. Radiol Oncol 2000; 34: 357-61. 52. Sersa G, Cufer T, Cemazar M, Rebersek M, Zvonimir R. Electrochemotherapy with bleomycin in the treatment of hypernephroma metastasis: case report and literature review. Tumori 2000; 86:163-5. 53. Rols MP, Bachaud JM, Giraud P, Chevreau C, Roche H, Teissie J. Electrochemotherapy of cutane­ous metastases in malignant melanoma. Melanoma Res 2000; 10: 468-74. 54. Sersa G, Stabuc B, Cemazar M, Miklavcic D, Rudolf Z. Electrochemotherapy with cisplatin: the systemic antitumour effectiveness of cisplatin can be potentiated locally by application of electric pulses in the treatment of malignant melanoma skin metastases. Melanoma Res 2000; 10: 381-5. Radiol Oncol 2006; 40(3): 163-74. 55. Sersa G, Stabuc B, Cemazar M, Miklavcic D, Rudolf Z. Electrochemotherapy with cisplatin: clinical experience in malignant melanoma pa­tients. Clin Cancer Res 2000; 6: 863-7. 56. Rodriguez-Cuevas S, Barroso- Bravo S, Almanza- Estrada J, Cristobal- Martinez L, Gonzales-Rodriguez E. Electrochemotherapy in primary and metastatic skin tumors: Phase II trial using intralesional bleomycin. Arch Med Res 2001; 32: 273-6. 57. Allegretti JP, Panje WR. Electroporation therapy for head and neck cancer including carotid artery involvement. Laryngoscope 2001; 111: 52-6. 58. Burian M, Formanek M, Regele H. Electroporation therapy in head and neck cancer. Acta Otolaryngol 2003; 123: 264-8. 59. Shimizu T, Nikaido T, Gomyo H, Yoshimura Y, Horiuchi A, Isobe K, Ebara S, Takaoka K. Electrochemotherapy of digital chondrosarcoma. J Ortop Sci 2003; 8: 248-51. 60. Gothelf A, Mir LM, Gehl J. Electrochemotherapy: results of cancer treatment using enhanced deliv­ery of bleomycin by electroporation. Cancer Treat Rew 2003; 29: 371–87. 61. Byrne CM, Thompson JF, Johnston H, Hersey P, Quinn MJ, Hughes M, McCarthy WH. Treatment of metastatic melanoma using electroporation therapy with bleomycin (electrochemotherapy). Melanoma Res 2004; 15: 45-51. 62. Snoj M, Rudolf Z, Cemazar M, Jancar B, Sersa G. Successful sphincter-saving treatment of anorectal malignant melanoma with electrochemotherapy, local excision and adjuvant brachytherapy. Anti-Cancer Drugs 2005; 16: 345–8. 63. Kubota Y, Tomita Y, Tsukigi M, Kurachi H, Motoyama T, Mir LM. A case of perineal malignant melanoma successfully treated with electrochemo-therapy. Melanoma Res 2005; 15: 133-4. Radiol Oncol 2006; 40(3): 163-74. review Fertility preservation methods for female neoplastic patients L. Del Pup, E. Campagnutta, G. Giorda, G. De Piero, F.Sopracordevole, R. Sisto Gynecological Oncology Department, National Institute of Cancer, Aviano, Pordenone, Italy Background. The ability to have biological children is of great importance to cancer patients and fertility preservation before the oncological treatment is nowadays not rare. Oncologists have the responsibility to inform patients about the risks that their cancer treatment will permanently impair fertility and about the ways to limit this iatrogenic damage. Methods of fertility preservation are evolving quickly, yet the medical oncology literature is still poor regarding this topic. Indications and contraindications, limits and controver­sies of the fertility sparing techniques are reviewed in order to help the oncologist to counsel patients. Conclusions. Any oncologist seeing reproductive-aged patients should discuss the fertility implications of the oncological treatment and the fertility preservation options. A referral to appropriate fertility specialists as early as possible is recommended. People attempting fertility preservation in the context of cancer treat­ment are encouraged to enrol in clinical trials. Key words: neoplasms; fertility; ovary; oocytes; embryo; cryopreservation; fertilization in vitro Introduction Nearly 25% of cancers affect women who have not had a child or who have delayed childbearing. The number of women sur­viving cancer is increasing, but at the same time the long-term fertility adverse effects of the treatment that they received are growing. This iatrogenic damage sub­stantially impairs the quality of life of the cancer survivor, leading to premature ovar- Received 1 September 2006 Accepted 14 September 2006 Correspondence to: Lino Del Pup, Gynecological Oncology Dept., National Institute of Cancer, Aviano, Pordenone, Italy. Phone: +39434 659051; Fax: +39434 659439; E mail address: ldelpup@cro.it ian failure and infertility in the majority of these patients. These women can experience persistent sexual problems, fertility concerns, and related adverse psychosocial sequel even many years after their cancer treatment. Reproductive concerns are significant mostly for those who »very much« desire children prior to cancer, had none prior, and are unable to reproduce subsequently. Cancer diagnosis reduces the desire to have children in 6-13% of patients, but it increases such desire in 19-24% of them. Among cancer survivors, 76% of those without children and 31% of those who are already parents, desire to have children in the future.1 This suggests that there may be a need for more formalized intensive counselling both prior to and after cancer treatment to aid patients in resolving or managing psychosocial sequel resulting from the un­planned infertility (Table 1).2,3 Table 1. Percentages of cancer survivors having specific reproductive concerns3 Reproductive Concerns Scale % Loss of control over reproductive future 30 Discontent with number of children 27 Inability to talk openly about fertility 18 Illness affected ability to have children 15 Sad about inability to have children 13 Frustrated ability to have children affected 11 Angry ability to have children affected 11 Mourned loss of ability to have children 11 Concerns of having children 8 Guilt about reproductive problems 8 Less satisfied with life because of problem 6 Less of a woman 6 Blame self for reproductive problems 4 Others are to blame for reproductive problems 40 Discussing fertility issues at the time of diagnosis provides the patient and her fam­ily with the reassurance that the oncology team believes in a future of survival and even of acceptable quality of life. While none of the fertility preservation options currently available provide total reassur­ance regarding the future fertility, for many young women, both the counselling in­volved in discussions of fertility preserva­tion and the potential for optimisation of the chances of fertility in the future, together have a positive psychological im­pact during a very traumatic time in their lives.4,5 Oncologists have traditionally focused more on providing the most effective treat­ments available, and less on the patient’s post treatment quality of life. Physicians treating younger patients for cancer should now be aware of the adverse effects of treat­ment on fertility and of ways to minimize those effects. If gonadal toxicity is unavoid­able, they should be knowledgeable about options for fertility preservation and must discuss with patients the following op­tions.2,6 Gonadotropin-releasing hormone agonist (GnRHa) co-treatment with chemotherapy The results of gonadoprotective hormo­nal therapy are considered contradictory and the controversy will only be resolved by prospective randomized clinical trials. Following encouraging findings in animal models, nonrandomized studies with a short-term follow-up suggested a protective role for GnRHa co-treatment, 7-11 but these studies were criticized for their lack of ran­domization, different follow-up periods for treatment and control groups, and the use of ovarian failure as the endpoint, which may not reflect the decrease in primordial follicle count in response to chemotherapy in young women. 12 The mechanism by which GnRHa co-treatment may protect against chemother­apy-induced gonadal damage is still debat­ed, as is the presence of follicle-stimulating hormone (FSH) receptors in primordial follicles.7,12 Mechanisms by which GnRH-a could minimize chemotherapy-associated gonadotoxicity:7 1. the hypogonadotropic state generated by the GnRH-a creates a prepubertal hormonal milieu that decreases the ac­tivity and so the rate of follicular apop­tosis and degeneration. 2. the hypoestrogenic state may decrease utero-ovarian perfusion, resulting in a de­creased total cumulative exposure of the ovaries to the chemotherapeutic insult. 3. gonads contain GnRH-I and GnRH-II receptors the activation of which could decreases apoptosis. Radiol Oncol 2006; 40(3): 175-81. 4. GnRH-a may up-regulate an intrago­nadal antiapoptotic molecules such as sphingosine-1 phosphate (S-1-P). 5. GnRH-a may protect the undifferentiat­ed germline stem cells, which ultimately generate de novo primordial follicles. Criticism to Gn-RH agonist use is based on these considerations. Primordial follicles initiate follicle growth through an unknown mechanism, which is not gonadotropin de­pendent. There is some controversy regard­ing the existence of GnRH receptors on the human ovary, whereas GnRH receptors have clearly been detected in the rat ovary. The response may thus not be similar across species. If the sole mechanism of gonad pro­tection with GnRH agonists were through the suppression of gonadotropins, especially FSH, then the treatment would not be ex­pected to protect the primordial follicle pop­ulation that represents the ovarian reserve. Some prepubertal children receiving gonadotoxic chemotherapy may eventu­ally have POF. As younger patients have a larger ovarian reserve, a decreased fre­quency of immediate amenorrhea does not mean that the gonads are unaffected by the chemotherapy, but simply that they have a sufficient number of oocytes not to demon­strate immediate ovarian failure.13 The hypoestrogenic state induced by Gn RH agonists may have negative effects in breast cancer patients by arresting tumours cells in G0 phase and making them less responsive to chemotherapy. At present, despite encouraging reports, the benefits and long-term effects of GnRHa co-treatment are unclear, and a consensus regarding the effectiveness of ovarian sup­pression is lacking. Therefore, GnRHa co-treatment for prevention of chemotherapy-induced gonadotoxicity should be offered to patients only with appropriate informed consent in an institutional review board ap­proved investigational protocol. Gonadotropin-releasing hormone agonist co-treatment with chemotherapy can be used: 1. as the only strategy, if no other option is available and the patient is informed about its limits 2. combined with other options. Ovarian tissue cryopreservation Ovarian tissue consisting of germ cells can be removed and stored before the gonado-toxic treatment. After patients are cured, this tissue might either be returned to pa­tients via autotransplantation or matured in vitro to produce offspring by in vitro fertilisation. Ovarian tissue can be removed by the use of multiple biopsy samples from the ovary or by oophorectomy. The removal of ovarian cortical strips that can be done laparoscopically is better and it produces tissue that is rich in primordial follicles. Cortical strips and biopsies are ideal be­cause the tissue survives cryopreservation and undergoes revascularisation on return, although most primordial follicles are lost. The autologous transplantation of this tissue aims to restore natural fertility and also maintain sex-steroid production. The feasibility of this process has been shown in sheep and other mammals, with both the return of ovarian hormonal activity and the subsequent production of offspring. After such success in animals, the evidence of ovulation after orthotopic transplantation in a woman was reported.14 The reports by Oktay and colleagues15 and Donnez and co-workers16 are important in showing that the ovarian function could realistically be preserved after the sterilising treatment, although the continuing intermittent ovula­tion in the Donnez study raises questions as to whether pregnancy clearly resulted from the grafted tissue. This technique of fertility preservation remains experimental and several issues Radiol Oncol 2006; 40(3): 175-81. remain to be clarified, but perhaps the greatest concern is the potential to return malignant cells back to patients after they are cured. This factor is of particular im­portance in patients with haematological malignant disease. Oocyte maturation in vitro, followed by assisted reproduction, would eliminate this risk. Techniques to mature oocytes artificially, even from early stages of development, have yielded some success in mice. At present, little is known about the support needed for this process to take place in human tissue, and the clini­cal potential of this technique will need to be established. Candidates to ovarian tissue cryopreservation are cancer patients who: 1. wish to be pregnant in the future or who don’t exclude such possibility 2. have a realistc chance of long-term sur­vival 3. still have at least a certain amount of follicles, possibly not damaged by previ­ous treatments 4. accept, must be performed and don’t have surgical contraindications to lapar­oscopy 5. have a low risk of primary tumour reim-plantation or ovarian cancer 6. can’t use ovarian hyperstimulation, be­cause of neoplatic and/or thrombotic risk 7. need to start chemo/radiotherapy as soon as possible and who have not enough time to wait for in vitro fertiliza­tion (IVF) cycles 8. don’t yet have a partner or have him but can’t do IVF 9. are well informed about all the options and their risks 10. choose ovarian cryopreservation con­scious that it is still experimental 11. have ethical concerns regarding ovula­tion induction and oocyte retrieval or other options. Cryopreservation of unfertilized human oocytes Fertility might be preserved by obtain­ing mature oocytes before the gonadotoxic treatment for IVF and subsequent embryo cryopreservation. This is the most effective method, but it is only applicable to sexu­ally mature women, and needs a partner or donor sperm for fertilisation. For women without a partner, cryop-reservation of mature oocytes is an option, but subsequent pregnancy rates are sub­stantially lowered because these cells sus­tain more damage during the freeze–thaw process than do embryos.17 These tech­niques are not suitable for most patients with cancer, because they need a period of ovarian stimulation that will delay treat­ment. The technique is also inappropriate for prepubertal patients, in whom all fertil­ity preservation strategies remain experi­mental. Cryopreservation of human oocytes can be performed if: 1. the laboratory is specifically highly com­petent on oocyte cryopreservation 2. a partner is not available 3. ovarian stimulation is possible 4. other options are discussed and dis­carded 5. the patient is properly conscious of the actual limited results of this technique. Embryo cryopreservation Embryo cryopreservation is still the most efficient method to preserve future fertility because of reasonable post-thaw survival, implantation, and delivery rates. Because the efficacy of IVF is dramatically reduced after even one round of chemotherapy, IVF should be performed before chemotherapy. Obviously, embryo freezing is predomi- Radiol Oncol 2006; 40(3): 175-81. nantly suitable for women with a partner with whom they wish to procreate and it has legal limitations. Embryo cryopreservation is an established technique that is available for fertility preservation if: 1. a small delay in the initiation of chemo­therapy or radiotherapy is acceptable 2. a partner sperm is available (or a donor outside Italy) 3. ovarian hyperstimulation can be safely performed 4. this technique is chosen knowing its ef­ficiency and the alternatives 5. there are non ethical or legal limita­tions. Fertility-sparing surgery Preservation of at least a part of an ovary and/or of the uterus can be done in cer­tain neoplastic situations. Optimal cancer therapy should always supersede fertility preservation as a primary objective. Ovarian neoplasms candidates for fertil­ity-sparing surgery are ovarian tumours of low malignant potential, malignant ovarian germ cell tumours and ovarian sex cord-stro­mal tumours. Fertility-sparing surgery may be an option for invasive epithelial ovarian cancer which have early-stage disease, if the patient is well informed about risks, but this is highly controversial. Surgical procedures that would constitute fertility-sparing surgery for an ovarian malignancy include ovarian cystectomy, unilateral salp­ingo-oophorectomy, unilateral salpingo­oophorectomy plus hysterectomy, with the preservation of the contralateral ovary, and bilateral salpingo-oophorectomy, with the preservation of the uterus. Of course, af­ter the latter two procedures, the assisted reproductive technology (ART) would are necessary to achieve a pregnancy. The fertility sparing options for invasive cervical cancer are conization alone for stage IA1 or IA2 disease or radical trache­lectomy for stage IA2 or IB1 disease. In addition, IVF techniques may be employed prior to definitive therapy if time delays are not significant. The optimal candidate for medical treat­ment of endometrial cancer is a woman of childbearing age who has a stage IA, grade 1, adenocarcinoma. If such treatment is contemplated, it is recommended that a thorough hysteroscopy and curettage be performed to rule out a worse lesion prior to initiation.2 Candidates for fertility-sparing surgery or therapies are well informed patients with: 1. ovarian tumours of low malignant po­tential, malignant ovarian germ cell tumours, ovarian sex cord-stromal tu­mours and selected cases of epithe­lial malignant ovarian cancers stage Ia where one ovary could be saved. 2. stage IA1 or IA2 cervical cancer treated with conization alone or stage IA2 or IB1 where radical trachelectomy is per­formed 3. selected cases of stage IA, low-grade, endometrial cancer treated with pro­gestins. Transposition of the ovaries Patients who receive pelvic irradiation might have their ovaries shielded or re­moved from the radiation field, a proce­dure known as oophoropexy which can be undertaken laparoscopically. Although the ovarian function can be preserved in 50% of cases, ischemia and radiation induced uterine and ovarian damage will reduce the chances of a successful pregnancy. Radiol Oncol 2006; 40(3): 175-81. Transposition of the ovaries should be considered in case of: 1. planned pelvic or whole body irradia­tion 2. chemotherapy is not necessary 3. ovarian cancer involvement is unlikely 4. ovarian hypestimulation can be per­formed 5. can be combined with ovarian tissue cryopreservation. Egg or uterus donation Premature ovarian failure affects expecially young female cancer patients who can only rely on egg donation. This technique has the highest effectiveness among fertility preservation options even for women can­didates to other fertility preventive options: cumulative pregnancy rates are over 60%, if embrios are of good quality. Uterus donation is still anedoctal and it is a possibility for women who did hyster­ectomy or pelvic radiotherapy. Strong ethi­cal and legal concerns are the main limits. Candidates to egg or uterus donation are women who: 1. are affected by premature ovarian failure 2. did hysterectomy or pelvic radiotherapy 3. have no ethical concerns nor legal limits to this. Conclusions Fertility preservation is often possible and should always be proposed and discussed to women undergoing treatment for can­cer. Oncologists should be prepared on this subject or they must refer patients to proper reproductive specialists. To preserve the full range of options, fertility preser­vation approaches should be considered as early as possible during the treatment planning. These methods have psychologi­cal, ethical and legal aspects that should be fully discussed before choosing the most appropriate for each case.18-20 The fertility preservation techniques should be consid­ered investigational and must be performed in centres with the necessary expertise. References 1. Schover LR. Motivation of parenthood after can­cer: a review. J Natl Cancer Inst Monogr 2005; 34: 2-5. 2. Lee SJ, Schover LR, Partridge AH, Patrizio P, Wallace WH, Hagert K, et al. American Society of Clinical Oncology recommendations on fertility preservation in cancer patients. J Clin Onc 2006; 24: 1-15. 3. Wenzel L, Dogan-Ates A, Habbal R, Berkowitz R, Goldstein DP, Bernstein M, et al. Defining and measuring reproductive concerns of female cancer survivors. J Natl Cancer Inst Monogr 2005; 34: 94-8. 4. Schover LR, Rybicki LA, Martin BA, Bringelsen KA. Having children after cancer: A pilot survey of survivors’ attitudes and experiences. Cancer 1999; 86: 697-709. 5. 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Blumenfeld Z, Shapiro D, Shteinberg M, Avivi I, Nahir M. Preservation of fertility and ovarian function and minimizing gonadotoxicity in young women with systemic lupus erythematosus treated by chemotherapy. Lupus 2000; 9: 401-5. 11. Blumenfeld Z. Ovarian rescue/protection from chemotherapeutic agents. J Soc Gynecol Investig 2001; 8(1 Suppl Proceedings): S60-4. 12. Sonmezer M, Oktay K. Fertility preservation in female patients. Hum Reprod Update 2004; 10: 251–66. 13. Falcone T, Bedaiwy MA. Fertility preservation and pregnancy outcome after malignancy. Curr Opin Obstet Gynecol 2005; 17: 21-6. 14. Oktay K, Karlikaya G. Ovarian function after transplantation of frozen, banked autologous ovar­ian tissue. N Engl J Med 2000; 342: 1919. 15. Oktay K, Buyuk E, Veeck L, Zaninovic N, Xu K, Takeuchi T, et al. Embryo development after het­erotopic transplantation of cryopreserved ovarian tissue. Lancet 2004; 363: 837-40. 16. Donnez J, Dolmans MM, Demylle D, Jadoul P, Pirard C, Squifflet J, et al. Livebirth after ortho-topic transplantation of cryopreserved ovarian tissue. Lancet 2004; 364: 1405-10. 17. Porcu E, Fabbri R, Damiano G. Oocyte cryopreser­vation in oncological patients. Eur J Obstet Gynecol Reprod Biol 2004; 113(Suppl 1): S14–6. 18. Patrizio P, Butts S, Caplan A. Ovarian tissue pres­ervation and future fertility: emerging technolo­gies and ethical considerations. J Natl Cancer Inst Monogr 2005; 34: 107-10. 19. Wallace WH, Anderson RA, Irvine DS. Fertility preservation for young patients with cancer: who is at risk and what can be offered? Lancet Oncol 2005; 6(4): 209-18. 20. Stern CJ, Toledo MG, Gook DA, Seymour JF. Fertility preservation in female oncology patients. Aust N Z J Obstet Gynaecol 2006; 46: 15–23. Radiol Oncol 2006; 40(3): 175-81. Different localisation of cystatin C in immature and mature dendritic cells Tina Zavašnik-Bergant1, Martina Bergant2, Matjaž Jeras2, Gareth Griffiths3 1Jožef Stefan Institute, Dept. of Biochemistry and Molecular Biology, Ljubljana, Slovenia; 2Blood Transfusion Centre of Slovenia, Tissue Typing Centre, Ljubljana, Slovenia; 3European Molecular Biology Laboratory (EMBL), Cell Biology and Biophysics Unit, Heidelberg, Germany Background. Limited antigen degradation by proteolytic enzymes and their control by protease inhibitors represent a crucial step in generating antigenic peptides inside the endocytic pathway of antigen-presenting cells such as dendritic cells. Methods. Human dendritic cells were used as a cell model in which quantitative immunogold electron microscopy was applied in order to study endogenous protease inhibitor cystatin C. Ultrathin cryosec­tions were prepared from immature and mature dendritic cells and labelled with specific antibody. Under the transmission electron microscope gold particles, bound to specific probe (antibody), pointed the exact localization of labelled inhibitor. Results. Quantification of immunogold labelling and further statistical analysis by chi-squared test empha­sized the differences in cystatin C content in different cell compartments. Conclusions. Statistically significant differences in intracellular distribution of cystatin C have been deter­mined between immature and mature dendritic cell population. Key words: dendritic cells; cryoultramicrotomy; microscopy, electron; immunohistochemistry Received 1 June 2006 Accepted 21 July 2006 Correspondence to: Tina Zavašnik-Bergant, Department of Biochemistry and Molecular Biology, Jožef Stefan Institute, Jamova 39, SI-1000 Ljubljana, Slovenia. Tel.: +386 1 477 3474; Fax: +386 1 477 3984; E-mail: tina.zavasnik@ijs.si The paper was presented at the 4th Conference on Experimental and Translational Oncology (Kranjska gora, Slovenia, March 22-26, 2006) as invited lecture. Abbreviations: Ag – antigen, APC – antigen-pre­senting cells, DC – dendritic cells, MHC II – major histocompatibility complex class II molecules, TNF-a• – tumour necrosis factor-alpha, TEM – transmission electron microscopy Introduction Following the processes of internaliza­tion, different particles, parts of apoptotic and necrotic cells, pathogens (bacteria, viruses) and other foreign proteins are taken up into the endocytic pathway of immune cells such as macrophages and dendritic cells (DC).1 As antigen-present­ing cells (APC) these cells play a pivotal role in the adaptive immunity by initia­tion of efficient T-cell mediated immune response.2 DC circulate through tissues like veiled scavengers. Their maturation occurs as they migrate from peripheral tis­sues, where they search for antigen (Ag), to lymphoid organs, where they present captured and processed Ag to specific T cell receptors. Limited Ag degradation performed by proteolytic enzymes (cathe­psins) and their control by intracellular protease inhibitors represent a crucial step in generating antigenic peptides.3,4 Due to the increased research interest in events taking place inside the endocytic pathway5 of immune cells, a high-resolu­tion technique has been applied to observe cell organelles and search for specific intra­cellular proteins. Differentiation of mono-cytes to immature DC and their further maturation with TNF-a to mature DC pop­ulation6,7 was applied as a model system for studying transport pathway of endog­enous protease inhibitor cystatin C in im­mune cells. Described method consists of preparation of ultrathin cryosections from frozen cells,8 their immunogold labelling,9 quantification of attached gold particles by transmission electron microscopy (TEM) and statistical analysis of obtained data.10 Material and methods Immature DC were generated from isolated blood monocytes and further stimulated with TNF-a as described.6,7 Cells were fixed with 4% paraformaldehyde (Sigma –Aldrich). They were embedded into 10% gelatin (Merck), infused with 2.3 M sucrose (Merck) and cut to ultrathin cryosections (70 – 80 nm) at minus 120 °C8,9 using Leica EM FC6 ultramicrotome for cryosectioning. Sections were thawed in a droplet of 1:1 mixture of 2.3 M sucrose (Merck) and 2% methyl cellulose (Sigma-Aldrich). Retrieved ultrathin sections were put on carbon-coated formvar films (TAAB Lab.) and spread over hexagonal Cu/Pd grids (Agar). Sections were labelled with rabbit anti-hu­man cystatin C antibody11 or rabbit anti-hu­man cathepsin S antibody.12 Their specifici­ty and cross-reactivity against recombinant human antigens 13,14 was checked as report­ed 7,11,12,15 prior to described immunogold labelling experiments. After the removal of an excess of primary antibody thin sections of DC were incubated with Protein A–gold conjugate with 10-nm gold particles (Cell Microscopy Centre, University Medical Centre Utrecth). Sections were contrasted with uranyl acetate (Aurion) and viewed with Philips CM120 BioTWIN transmis­sion electron microscope. KeenView digital camera and iTEM software were used for taking pictures. Quantification of immunogold labelling patterns in the same sets of compartments (or organelles) in different groups of cells (immature and mature DC) was performed for protease inhibitor cystatin C. Random sampling of specimen was followed by counting of approximately 200 gold par­ticles on two labelled grids. The distribu­tions of gold particles in different compart­ments was compared by contingency table analysis with statistical degrees of freedom for chi-squared values being determined by the number of compartments and the number of experimental groups of cells. Compartmental chi-squared values making substantial contributions to the total chi-squared values identified where the main differences between groups resided.10 Results Strong immunogold labelling of Golgi was observed in immature DC (Figure 1) where­as in mature DC Golgi remained practically unlabelled after incubation with specific antibody (Figure 2). Gold counts and re­sults of calculations for Golgi and other compartments (organelles) are provided in Figure 3 and Table 1. High number of gold Radiol Oncol 2006; 40(3): 183-8. particles associated with Golgi (Figure 1, Figure 3A), endoplasmic reticulum (Figure 3A) and nuclear membrane (Figure 3A) indicated high expression of cystatin C in immature DC. In mature DC number of gold particles increased in multivesicular bodies and small vesicles near the plasma membrane (Figure 3B). Of 204 gold particles counted in im­mature DC 59 were associated with Golgi (Figure 3A, Table 1). In contrast, in mature DC 203 gold particles were distributed so that only 2 were observed on Golgi (Figure 3B, Table 1). By contingency table analysis, the predicted number of gold particles on a given compartment in a given cell group is given by [(column total × row total)/grand total] where the grand total is the sum of the column (or row) totals.10 Consequently, the predicted number of Golgi gold parti­cles in immature DC, for example, is calcu­lated to be [(61x204)/407] or roughly 30.6 (Table 1). For each compartment and cell type, the partial chi-squared value is given Figure 3. Comparison of immunogold labelling of inhibitor cystatin C in immature (A) and mature (B) den-dritic cells. Gold particles were counted on two grids. Distributions of app. 200 gold particles (+ SD) are shown. Compartments: (a) nucleus, (b) nuclear membrane, (c) rough endoplasmic reticulum, (d) Golgi apparatus, (e) small vesicles, (f) multivesicular bodies, (g) multilamelar bodies, (h) vesicles near plasma membrane, (i) plasma mem­brane, (j) cytoplasm, (k) mitochondria, (l) unknown structures, other. Radiol Oncol 2006; 40(3): 183-8. Table 1. Labelling distributions of inhibitor cystatin C and chi-squared test. Values represent observed and ex­pected (in brackets) numbers of 10-nm gold particles in immature and mature dendritic cells. For a total chi-squared of 120.02 and 11 degrees of freedom, P < 0.001. These two labelling distributions are significantly different. by [(observed golds - expected golds)2 / 10 expected golds]. Therefore, the partial chi-squared value for Golgi in immature DC is equivalent to [(59 - 30.6)2/30.6] or 26.4. Table 1 shows that the total chi-squared value is 120.02. For 11 degrees of freedom [(2 - 1 columns) by (12 - 1 rows)] this indicates that the null hypothesis of no difference between groups must be rejected (P < 0.001). Therefore, the distribution of cystatin C is significantly different in immature and mature DC and the gold counts are consist­ent with a shift towards greater-than-pre­dicted labelling of Golgi in immature DC (Table 1). Furthermore, the partial (or compart­mental) chi-squared values indicate that immature DC have more-than-expected gold particles on Golgi, endoplasmic re­ticulum and nuclear membrane (Table 1). On the contrary, partial chi-squared val­ues indicate that mature DC have fewer-than-expected gold particles on Golgi but more-than-expected gold particles on the vesicles near the plasma membrane (within a distance of 200 nm), small vesicles and multivesicular bodies (Table 1). Population of multivesicular bodies (in addition to some other organelles) has also been labelled with anti-cathepsin S antibody and gold in both, immature and mature, DC population (data not shown). Discussion Antigen processing by MHC class II mol­ecules (MHC II) is tightly linked to the ac­tivity and stability of proteolytic enzymes present inside the endocytic pathway of DC.16 Lysosomal cysteine proteases (cathe­psins), which constitute a major portion of this proteolytic system, have an essential role in both Ag processing and matura­tion of MHC II.17 Among the regulatory molecules of cathepsins a low molecular weight (MW 13.000) type II inhibitor cysta-tin C was suggested to have a role in con­trolling the proteolytic activity of cathepsin S inside the endocytic route of mouse DC.18,19 By binding to cathepsin S during Radiol Oncol 2006; 40(3): 183-8. DC maturation cystatin C was suggested to compromise the step-wise degradation of MHC II-associated chaperone molecule (invariant chain). We have confirmed that in human imma­ture DC cystatin C content was highly el­evated compared to mature DC population as well as to their precursors – monocytes.7 But with this study, cystatin C extremely high content in Golgi apparatus as well as its presence in endoplasmatic reticulum and nuclear membrane, continuous with the endoplasmic reticulum, was quanti­fied for the first time in human immature DC. Furthermore, the transport of cystatin C was shown from Golgi towards the cell membrane, supported by the decrease of cystatin C in Golgi and endoplasmic reticu­lum and increase in labelling of different populations of transport vesicles (small vesicles, vesicles near plasma membrane) and multivesicular bodies. MHC II–loading compartments (multive­sicular bodies) were positive for cathepsin S 13,14 (proposed target enzyme of cystatin C, involved in maturation of MHC II) in both, immature and mature DC (Zavašnik-Bergant, unpublished data). Labelling of multivesicular bodies for cystatin C was higher-than-expected in mature but not immature DC indicating higher content of cystatin C inside Ag-loading compart­ments in mature DC compared to imma­ture population. Differentiation and matu­ration dependence of endogenous cystatin C supports its intracellular regulatory po­tential in human DC7, as well as it further suggests its new role in Golgi apparatus of immature DC. In conclusion, preparation of cryosections of human DC, their immunogold label-ling with specific anti-cystatin C antibody and transmission electron microscopy were successfully applied to quantify protease inhibitor cystatin C in different organelles of immature and mature DC. Acknowledgements This work has been performed at EMBL Heidelberg and granted by EMBO fellow­ship (ASTF 90.00-05) to Tina Zavašnik-Bergant. An introduction to the described technique by Uta Haselmann and a support given by Claude Antony (EMBL Electron Microscopy Core Facility) is acknowl­edged. References 1. Griffiths G. On vesicles and membrane compart­ments. Protoplasma 1996; 195: 37-58. 2. Honey K, Rudensky AY. Lysosomal cysteine pro­teases regulate antigen presentation. Nat Rev Immunol 2003; 3: 472-82. 3. Villadangos JA, Schnorrer P, Wilson NS. Control of MHC class II antigen presentation in dendritic cells: a balance between creative and destructive forces. Immunol Rev 2005; 207: 191-205. 4. Chapman HA. Endosomal proteases in antigen pre­sentation. Curr Opin Immunol 2006; 18: 78-84. 5. Pillay, CS, Elliott E, Dennison C. Endosomal pro­teolysis and its regulation. Biochem J 2002; 263: 417-29. 6. Repnik U, Kneževic M, Jeras M. Simple and cost-effective isolation of monocytes from buffy coats. J Immunol Methods 2003; 278: 283-92. 7. Zavašnik-Bergant T, Repnik U, Schweiger A, Romih R, Jeras M, Turk V, et al. Differentiation- and maturation-dependent content, localization and secretion of cystatin C in human dendritic cells. J Leukoc Biol 2005; 78: 122-34. 8. Tokuyasu KT. A study of positive staining of ul­trathin frozen sections. J Ultrastruct Res 1978; 63: 287-307. 9. Tokuyasu KT. Immunocytochemistry on ultrathin cryosections. In: Specer DL, Goodman RD, Leinwand LA, editors. Cells, a laboratory manual. Vol. 3. Subcellular localization of genes and their products. Cold Spring Harbour: Laboratory Press; 1997. p. 131.1-131.27. 10. Mayhew TM, Griffiths G, Lucocq JM. Applications of an efficient method for comparing immunogold labelling patterns in the same sets of compart­ments in different groups of cells. Histochem Cell Biol 2004; 122: 171-7. Radiol Oncol 2006; 40(3): 183-8. 11. Kos J, Štabuc B, Schweiger A, Krašovec M, Kopitar-Jerala M, Vrhovec I. Cathepsins B, H and L and their inhibitors stefin A and cystatin C in sera of melanoma patients. Clin Cancer Res 1997; 3: 1815-22. 12. Kos J, Šmit A, Krašovec M, Svetic B, Lenarcic B, Vrhovec I, et al. Lysosomal proteases cathepsins D, B, H, L and their inhibitors stefins A and B in head and neck cancer. Biol Chem 1995; 376: 401-5. 13. Cimerman N, Trstenjak-Prebanda M, Turk B, Popovic T, Dolenc I, Turk V. Interaction of cystatin C variants with papain and human cathepsins B, H and L. J Enzym Inhib 1999; 14: 167-74. 14. Kopitar G, Dolinar M, Štrukelj B, Pungercar J, Turk V.Folding and activation of human procathepsin S from inclusion bodies produced in Escherichia coli. Eur J Biochem 1996; 236: 558-62. 15. Zavašnik-Bergant V, Sekirnik A, Golouh R, Turk V. Immunochemical localization of cathepsin S, cathepsin L and MHC II-associated p41 isoform of invariant chain in human lymph node tissue. Biol Chem 2001; 382: 799-804. 16. Lennon-Duménil AM, Bakker AH, Maehr R, Fiebiger E, Overkleeft HS, Rosemblatt M, et al. Analysis of protease activity in live antigen-pre­senting cells shows regulation of the phagosomal proteolytic content during dendritic cell activa­tion. J Exp Med 2002; 196: 529-40. 17. Zavašnik-Bergant T, Turk B. Cysteine cathepsins in the immune response. Tissue Antigens 2006; 67: 349-55. 18. Pierre P, Turley SJ, Gatti E, Hull M, Meltzer J, Mirza A, et al. Developmental regulation of MHC II class II transport in mouse dendritic cells. Nature 1997; 388: 787-92. 19. Pierre P, Mellman I. Developmental regulation of invariant chain proteolysis controls MHC class II trafficking in mouse dendritic cells. Cell 1998; 93: 1135-45. Radiol Oncol 2006; 40(3): 183-8. Diagnostic reference levels for X-ray examinations in Slovenia Damijan Škrk 1, Urban Zdešar2, Dejan Žontar1 1Slovenian Radiation Protection Administration, 2Institute of Occupational Safety, Ljubljana, Slovenia Background. Medical applications of ionizing radiation are by far the largest man-made source of radiation exposure for the population in most developed countries. A good practice in diagnostic radiology should pro­duce an image containing all necessary information needed for accurate diagnosis and should result in the minimum dose to the patient. After introduction of diagnostic reference level by International Commission on Radiological Protection in 1996 the process of patient exposure optimization has been enhanced. Methods. Local performance in patient exposure for particular type of X-ray examination in radiological department can be assessed by comparison of mean patient dose to the diagnostic reference level derived from relevant regional or national data. Results. Results of extensive five-year national patient dose survey in Slovenia are reviewed. The proposed Slovenian diagnostic reference levels for fifteen different X-ray examinations are presented, commented and compared with international and national levels of other countries. Conclusions. The introduction of national diagnostic reference levels will increase the awareness of patient doses in Slovenia. Their proper use should promote good radiological practice by reducing doses where cur­rent practice is not optimised. Key words: radiography; radiation exposure; diagnostic reference level Received 18 July 2006 Accepted 18 September 2006 Correspondence to: Assist. Prof. Damijan Škrk, physicist, PhD, Slovenian Radiation Protection Administration, Langusova ulica 4, 1000 Ljubljana, Slovenia; Phone: +386 1 4788702; Fax: +386 1 4788715; E-mail: damijan.skrk@gov.si Introduction Numerous national and regional surveys have revealed large dose variations for patients undergoing the same type of diag­nostic X-ray examination.1,2,3,4 The findings have clearly indicated a need for improve­ments that would lead to patient dose re­duction without compromising diagnostic information. The concept of investigation levels for diagnostic medical exposures was first proposed by International Commission on Radiological Protection (ICRP) in its 1990 recommendations5 and further devel­oped into diagnostic reference level (DRL) in 1996 ICRP Publication 73.6 In line with the principle of keeping doses As Low As Reasonably Achievable (ALARA) the European Union member states regulated the optimisation of medical exposures by adoption and implementation of Medical Exposure Directive (MED).7 The directive defines DRLs as the dose levels in medi­cal radiodiagnostic practices for typical examinations for groups of standard-sized patients or standard phantoms for broadly defined types of equipment. DRLs are expected not to be exceeded in stand­ard procedures, when good and normal practice regarding diagnostic and technical performance is applied. With this func­tion in mind, the values at which they are set should be at the borderline between the acceptable and the unacceptable cur­rent national practice rather than at some optimum level based on the latest and best technology. Although it is known that many aspects of radiodiagnostic practice differ between European Union member states and that these differences may affect the patient dose distributions within each country, it is acceptable to use the levels published by the European Commission when national DRLs are not available. Nevertheless, the regulatory authorities in coordination with the professional bod­ies of medical radiology and radiological protection, are encouraged to develop guid­ance and establish DRLs based on their own national dose distributions as soon as feasible. DRLs essentially act as the initial standard in a local radiology audit process for identifying situations where patient doses are unusually high. Local reviews should be undertaken whenever relevant diagnostic reference levels are consistently exceeded and appropriate corrective ac­tions should be taken in order to improve practice and avoid unnecessary risk due to radiation health effects. Material and methods Diagnostic reference levels should be ex­pressed in terms of dose quantities that are well defined and that can be easily measured with sufficient precision and accuracy. A number of different dose quantities can be used, although in practice two are usually ap­plied for routine monitoring of patient doses in conventional radiology. For individual ra­diograph projections the recommended dose quantities are Entrance Surface Dose (ESD) and Dose-Area Product (DAP). The Slovenian Radiation Protection Administration and Institute of Occupational Safety started a national patient dose sur­vey in the year 2000.10 In line with the international recommendations concerning application of appropriate dose quantities, the measurement of the entrance surface dose (ESD) using thermoluminescence dosemeters (TLDs) was chosen. ESD can be directly measured by attaching TLDs to the skin at the point where the centre of the X-ray beam enters the patient. Due to the small size of the TLDs the imaging process is not affected by measurements and there is no loss of diagnostic information on X-ray images. Examples of ESD measurement for three types of X-ray examinations are presented in Figure 1. Another approach that was also used in the survey is the calculation of patient dose using X-ray examination technical settings and data from X-ray tube output measurements. The relevant parameters and their relation with the ESD are shown in Equation 1.12,14 As seen in the equation an appropriate backscatter factor should be applied and the measurement corrected for any differences between the position of the detector and the position of the entrance Radiol Oncol 2006; 40(3): 189-95. Figure 1. ESD measurement for a) chest examination in PA projection15, b) cervical spine examination in AP projec­tion and c) cervical spine examination in LAT projection16. Arrows indicate the TLDs positions. surface on the patient. Backscatter factors are significant usually ranging from 1.2 to 1.4. for the X-ray spectra and beam sizes used in diagnostic radiology. Equation 1. ESD calculation using X-ray tube output and exposure parameters. (D/It)0 is tube output at distance FDD from focus measured at the same tube potential as used for examination, It is product of tube current and exposure time used in the actual exami­nation, FDD is focus detector distance, FPD is focus patient distance and BSF is backscatter factor. The measurements were carried out with Li2B4O7 TLDs. Dosimeters were calibrated with 137Cs gamma rays and their energy responses were corrected to X-ray spectra. It was also found out that the response of TLDs varies less than 10% over radiation energies of X-rays produced at potentials between 50 kV to 120 kV. Since doses are critically dependent on the patient size, measure­ments were performed on a representative group of patients with weight close to 70 kg. Taking into consideration only examina­tions with acceptable image quality, patient related data were collected and radiographic techniques parameters (tube voltage, prod­uct of current and exposure time, focus-film distance, screen-film sensitivity and filtra­tion) for each tube and examination type were registered. The mean dose for groups of about 10 patients (for each type of exami­nation performed on a particular X-ray tube) provides a good indication of typical clinical practice in a given institution. Diagnostic ref­erence levels were derived from mean doses for each type of examination performed in different X-ray departments. Diagnostic reference levels should be set with clear understanding of their intended purpose. A pragmatic way of setting diagnostic level for particular examination is to use the rounded third quartile value of mean dose distribu­tion. If 75% of X-ray departments can oper­ate satisfactorily below this dose level, then the remaining 25% should be made aware of their non optimal performance. Radiol Oncol 2006; 40(3): 189-95. Results By the end of 2005 the Slovenian national patient dose database contained results of over 2000 measurements.10,11,12,13 The survey took place at 33 radiological departments, out of which 15 belong to general hospitals and 18 to primary health care centres. Based on frequencies of their use and contribution to the collective dose of the population, the following fifteen examinations were tak­en into consideration: skull (AP/PA), skull (LAT), cervical spine (AP,LAT), chest (PA, LAT, AP), thoracic spine (AP, LAT), lumbar spine (AP,LAT), lumbo sacral joint (LAT), pelvis (AP), hip (AP) and abdomen (AP). Not all of the listed examination types were assessed in each radiological department, leading to a ground total of 184 analysis performed. It was estimated that measure­ments were done at institutions performing approximately 25% of all listed diagnostic radiological procedures made in Slovenia. The minimum, mean, 3rd quartile and maxi­mum ESD values for each X-ray examination obtained from distribution of ESD mean values for participating X-ray departments are presented in Table 1. The proposed Slovenian national refer­ence dose values of ESD per X-ray exami­nation are summarized in Table 2, together with DRLs proposed by IAEA (1994)8, EC (1999)9, Germany (2002)2 and the United Kingdom (2002).4 Slovenian DRL values are in general lower than German and higher than United Kingdom values pro­posed in 2002. At the same time, they are well below the corresponding values proposed by IAEA in 1994 and EC in 1999. Apart from general conclusion one can note that the DRL of chest examination in PA projection and DRL of thoracic spine in LAT projection are slightly higher than the EC recommendations, indicating a non-optimised radiological practice. The DRL of chest examination is higher than the recommended one because in some departments tube voltage settings are low­er, while in the others screen-films have lower sensitivity than proposed by the European Guidelines on Quality Criteria for Diagnostic Radiographic Images. The DRL of thoracic spine is higher because tube voltage settings are not adjusted properly according to patient chest thick­ness. Table 1. ESD data for fifteen X-ray examination types based on national patient dose survey measurements Radiol Oncol 2006; 40(3): 189-95. More specific information about the na­tional patient dose survey is given in Table 3. In the first column the number of X-ray departments where a particular examina­tion was assessed is given, and in the sec­ond column the number of departments where international DRLs were exceeded is listed. The DRL values were in general compared to EU levels, with the exception of thoracic spine where EU9 levels were not available and the IAEA8 levels were used. For the chest in AP projection and the cervical spine in AP and LAT projections international levels were not available. A re­view of these data reveals that in about 12% out of 184 examinations analysed the in­ternational DRLs were exceeded. The ESD values for the same type of examination in the same X-ray department generally vary due to differences in patient size and in the radiographic technique used by differ­ent radiographers. Variations of the mean ESD values for individual X-ray examina­tion between different X-ray departments are also due to differences in radiographic equipment, film type sensitivity, process­ing chemicals and processing conditions. The range factor is defined as the ratio of maximum to minimum dose for the same type of examination. The range factors of the mean ESD values for X-ray examination types are in the interval from 3 to nearly 10, which is shown in the third column of Table 3. Considering individual measurements instead of department mean values the vari­ations are naturally higher, ranging from 4 to nearly 24 (Table 3 fourth column). Although the mean ESDs were found to be well below the corresponding European DRLs for most examinations, this does not mean that the dose levels could not be further reduced without loss of diagnostic value by improving the radiographic tech­nique. Considerable dose variations for the same type of X-ray examination strongly support the idea that further optimisation is possible, especially since written ex­amination protocols were found lacking in almost all institutions. The data obtained demonstrate the importance of radiograph­ic staff awareness of regular quality control testing of radiographic equipment. Radiol Oncol 2006; 40(3): 189-95. Table 3. The number of radiological departments where individual examinations were assessed and the interna­tional DRLs were exceeded are presented in the first and in the second column respectively. In the third and the fourth column the range factors of mean ESD values from each department, and the range factors of ESD values for individual measurements are summarized respectively Discussion The patient dose survey was performed in recent years with a goal to get informa­tion about the patient doses in common X-ray examinations performed in Slovenia. The data obtained provide a useful base line against which the mean values of patient doses at individual X-ray department may be compared. DRLs are not intended to be applied as investigation levels for individual patients but should be compared with meas­ured or assessed mean values for a repre­sentative sample of patients. If the typical dose for a specific type of diagnostic proce­dure is consistently exceeding the relevant DRL, appropriate corrective action should be taken to improve practice. This could involve changes in procedures or equipment to reduce doses without compromising the quality of the diagnostic information. The radiological department staff should be en­couraged to alter their imaging equipment or examination techniques to bring the pa­tient doses in line with the majority of de­partments. Periodic monitoring of patient doses and collecting results on a national level are planned to become widespread throughout the Slovenian radiological de­partments. According to United Kingdom’s experience4 we expect patient exposure for common conventional X-ray examinations to show a clear trend towards lower doses in a few years. While dose variations for the same type of X-ray examination seems to be more difficult to overcome, a continuing need for bringing radiological procedures in line with European Guidelines on Quality Criteria for Diagnostic Radiographic Images should be emphasized. Radiol Oncol 2006; 40(3): 189-95. References 1. Shrimpton PC, Wall BF, Jones DG, Fisher ES, Hillier MC, Kendall G M, et al. National survey of doses to patients undergoing a selection of routine X-ray examinations in english hospitals. NRPB-R200. London: HMSO; 1986. 2. Veit R, Bauer B. Establishment of diagnostic reference levels for diagnostic radiology in Germany European Commission Doc RTD/0034/20, pp. 285-291 (2000), Medizinische Physik, DGMP; 2002. 3. Hart D, Hillier MC, Wall BF, Shrimpton PC, Bungay D. Doses to patients from medical X-ray examinations in the UK- 1995 Review. National Radiological Protection Board NRPB-R289. London: HMSO; 1996. 4. Hart D, Hillier MC, Wall BF. Doses to patients from medical X-ray examinations in the UK- 2000 Review. National Radiological Protection Board, NRPB- W14, Chilton Didcot; 2002. 5. International Commission on Radiological Protection. 1990 Recommendations of the International Commission on Radiological Protection. ICRP Publication 60, Ann. ICRP 21. Oxford: Pergamon Press; 1991. 6. International Commission on Radiological Protection. Radiological protection and safety in medi­cine. ICRP Publication 73, Ann. ICRP 26, Oxford: Pergamon Press; 1996. 7. European Council Directive 97/43 Euratom on Health Protection of Individuals against the Dangers of Ionizing Radiation in Relation to Medical Exposure. Official Journal of the European Communities L180/22 (9 July 1997). 8. International Atomic Energy Agency. International Basic Safety Standards for Protection against Ionization Radiation and for the Safety of Radiation Sources. Safety Series No 115. Vienna: IAEA; 1996. 9. European Commission. European Guidelines on Quality Criteria for Diagnostic Radiographic Images EUR 16260 EN, Luxembourg; 1996. 10. Institute of Occupational Safety, Patient doses from X-ray examinations in General hospital of Slovenj Gradec. Ljubljana: SRPA report; 2000. 11. Institute of Occupational Safety, Patient doses from X-ray examinations in General hospital of Maribor, Ljubljana: SRPA report; 2001. 12. Institute of Occupational Safety. Patient doses from X-ray examinations in Slovenia, Ljubljana : SRPA report; 2004. 13. Institute of Occupational Safety, Diagnostic ref­erence levels for X-ray examinations in Slovenia, Ljubljana: SRPA report; 2005. 14. Šešek T. Calculation of Entrance Skin Dose out of Parameters of Exposition at X-ray examination of Lumbar Spine. Diploma thesis. Ljubljana: University of Ljubljana, University College of Health Care; 2005. 15. Koren M, Patient doses from X-ray chest examina­tions in General Hospital »Dr. Franca Derganca« of Nova Gorica. Diploma thesis. Ljubljana: University of Ljubljana, University College of Health Care; 2005. 16. Šmon P. Patient doses from X-ray chest examinations in General Hospital of Slovenj Gradec. Diploma the­sis. Ljubljana: University of Ljubljana, University College of Health Care; 2005. Radiol Oncol 2006; 40(3): 189-95. Radio/ Oncol 2006: 40(3): 143-6. Sindrom kraje krvi pri prirojeni anomaliji podkljucnicne arterije Krnic A, Sucic Z, Vucic N, Krolo I Izhodišca. Sindrom podkljucnicne kraje krvi nastane zaradi motenega pretoka podkljucnic­ne arterije, ki ga povzroca zapora proksimalno od ustja vratne arterije. V clanku opisujemo primer bolnika z delnim sindromom podkljucnicne kraje krvi. Prikaz primera. Sindrom je povzrocila zožitev, ki je nastala zaradi aberantne desne podkljucnicne arterije. Krajo krvi smo prepoznali z dvojnim Doppler ultrazvokom, ki je pokazal znacilen vzorec pretoka v vratni arteriji; angiografija aortnega loka pa je pokazala arterijsko anomalijo. Ceprav je bila preiskava z dvojnim Doppler ultrazvokom odlocilna, smo arterijsko anomalijo morali potrditi z angiografijo. Zakljucki. Zgodnje odkritje delnega sindroma podkljucnicne kraje krvi nam omogoca, da razumemo vzrok nastanka bolnikovih simptomov ter uspešno sledenje in zdravljenje bolezni. Radio/ Onco/ 2006; 40(1): 197-200. Radio/ Oncol 2006; 40(3): 149-62. Kardiotoksicnost kemoterapije. Nove rešitve starega problema Miolo GM, La Mura N, Nigri P, Murrone P, Da Ronch L, Viel E, Veronesi A, Lestuzzi C Izhodišca. Kardiotoksicnost, ki jo povzroca kemoterapija ima raznolike zgodnje in kasne oblike. Zmanjšuje možnost ucinkovitega zdravljenja z namenom ozdravitve pa tudi paliativ­nega zdravljenja. Onkološka zdravila, ki jih najpogosteje povezujemo s kadiotoksicnostjo so antraciklini, trastuzumab, 5-flurouracil in taksani. Nekatere oblike kardiotoksicnosti, ki jih lahko povzroca vecina protitumorskih zdravil, pa avtorji redko opisujejo in navajajo. Velika verjetnost je, da bo širša uporaba novih bioloških zdravil privedla do odkritja drugih manj poznanih stranskih pojavov. Zakljucki. Ker srce razvršcamo med organe z omejeno regeneracijsko sposobnostjo, je pomembno, da poznamo incidenco, klinicno sliko in patogene mehanizme, ki so povezani s stranskimi ucinki zdravil na srce. To nam lahko pomaga pri ugotavljanju, prevenciji in zdravljenju kardiotoksicnosti, ki jo povzroca kemoterapija. Ob novih nacinih zdravljenja so nujno potrebne še nadaljnje raziskave. Radio/ Oncol 2006; 40(3): 163-74. Elektrokemoterapija tumorjev Serša G, Cemažar M, Miklavcic D, Rudolf Z Elektrokemoterapija je nacin zdravljenja raka, ki združuje uporabo standardnih kemote­rapevtikov in aplikacijo elektricnih pulzov na obmocje tumorja. Z aplikacijo elektricnih pulzov na tumor povzrocimo destabilizacijo celicne membrane, s cimer omogocimo, da ci­tostatiki, ki imajo slabo prehajanje skozi membrano, lažje vstopajo v celico. Tako se veckrat poveca citotoksicnost citostatikov, kot sta cisplatin ali bleomicin, s tem pa se poveca tudi njihova protitumorska ucinkovitost, posebno na mestu aplikacije elektricnih pulzov. Zaradi selektivno povecanega vnosa samo na obmocju tumorja je terapevtski indeks elektrokemo­terapije zelo dober, dobra je namrec lokalna protitumorska ucinkovitost brez lokalnih ali sistemskih stranskih pojavov, zaradi kemoterapevtikov ali aplikacije elektricnih pulzov. Po številnih predklinicnih raziskavah je bila elektrokemoterapija preizkušena tudi v mnogih klinicnih raziskavah. V veterinarski onkologiji je bila uspešnost elektrokemoterapije doka­zana pri zdravljenju razlicnih primarnih tumorjev mack, psov in konjev.V humani onko­logiji se je elektrokemoterapija izkazala pri zdravljenju kožnih in podkožnih tumorjev pri bolnikih z napredovalo boleznijo razlicnih vrst rakov. Rezultati vseh teh študij dokazujejo uspešnost elektrokemoterapije v onkologiji za pri lokalnem nadzoru rasti kožnih in podko­žnih lezij razlicnih vrst raka. Radio/ Oncol 2006; 40(1): 197-200. Radio/ Oncol 2006: 40(3): 175-81. Zašcita plodnosti pri bolnicah z rakom Del Pup L, Campagnutta E, Giorda G, De Piero G, Sopracordevole F, Sisto R Izhodišca. Biološko starševstvo je pomembno tudi za bolnike z rakom, zato vedno pogosteje že pred pricetkom onkološkega zdravljenja zašcitimo njihovo plodno sposobnost. Naloga onkologov je bolnike seznaniti z možnostjo, da bo onkološko zdravljenje lahko trajno okvarilo njihovo plodnost in kakšne so možnosti, da bi to jatrogeno okvaro zmanjšali ali preprecili. Metode zašcite plodnosti se zelo hitro razvijajo, ceprav o njih v strokovni litera­turi ne porocajo velikokrat. Da bi onkologom pomagali pri svetovanju bolnicam z rakom, v clanku navajamo indikacije, kontraindikacije, omejitve in kontraverznosti razlicnih zašcit plodnosti. Zakljucki. Ko onkolog obravnava bolnice z rakom, ki so v rodnem obdobju, naj bolnice seznani z možnimi posledicami zdravljenja in z nacini, da bi bolnica ohranila plodno sposobnost. Priporocamo, da bolnico cim prej napoti na posvet k strokovnjaku, ki ima izkušnje z zdravljenjem plodnosti. Ce se bolnice odlocijo za eno od metod zašcite plodnosti, predlagamo vkljucitve v študijske raziskave, ki bodo lahko pokazale ucinkovitost posamicnih metod. Radio/ Oncol 2006; 40(3): 183-8. Razlicna lokalizacija cistatina C v nezrelih in zrelih dendriticnih celicah Zavašnik-Bergant T, Bergant M, Jeras M, Griffiths G Izhodišca. Razgradnja antigenov s proteolitskimi encimi (proteazami) v endocitozni poti antigena predstavitvenih celic (dendriticnih celic) ter njihova regulacija z inhibitorji proteaz predstavlja pomemben korak pri nastanku antigenskih peptidov. Metode. Cloveške dendriticne celice so bile uporabljene kot celicni model za študij pro­teaznega inhibitorja cistatina C. Pripravljene so bile tanke zamrznjene rezine nezrelih in zrelih dendriticnih celic ter oznacene s specificnimi protitelesi za kvantitativno elektronsko mikroskopijo. Pod transmisijskim elektronskim mikroskopom so zrna koloidnega zlata, vezana na specificne sonde (protitelesa), pokazala natancno lokalizacijo oznacenega inhi­bitorja. Rezultati. Ovrednotenje oznacenih celic s statisticnim testom Hi-kvadrat je potrdilo razlike v vsebnosti cistatina C v razlicnih celicnih organelih. Zakljucki. Potrjena je bila statisticno znacilna razlika v znotrajcelicni porazdelitvi cistatina C med populacijama nezrelih in zrelih dendriticnih celic. Radio/ Oncol 2006; 40(1): 197-200. Radio! Oncol 2006; 40(3): 189-95. Diagnosticne referencne ravni pri rentgenskih preiskavah v Sloveniji Škrk D, Zdešar U, Žontar D Izhodišca. Uporaba virov ionizirajocih sevanj v zdravstvu prispeva najvecji delež k izpo­stavljenosti prebivalstva zaradi uporabe umetnih virov ionizirajocih sevanj. Izvedba rent­genskih preiskav v skladu z dobro radiološko prakso vodi do radiograma, ki vsebuje vse potrebne podatke za postavitev prave diagnoze, ob najnižji pacientovi izpostavljenosti. Mednarodna komisija za varstvo pred sevanji je leta 1996 predstavila koncept diagnosticnih referencnih ravni in s tem spodbudila proces optimizacije radioloških posegov. Metode. Na posameznem radiološkem oddelku lahko raven izpostavljenosti pacientov pri izbrani preiskavi ocenimo s primerjavo med povprecno izpostavljenostjo in vrednostjo diagnosticne referencne ravni, pridobljene na podlagi ustreznih regionalnih ali lokalnih podatkov. Rezultati. Podani so rezultati obsežnega petletnega zbiranja podatkov o izpostavljenosti pacientov pri rentgenskih preiskavah v Sloveniji. Predlagane diagnosticne referencne ravni petnajstih rentgenskih preiskav so predstavljene, opisane in primerjane z mednarodnimi in nacionalnimi vrednostmi v drugih državah. Zakljucki. Vpeljava nacionalnih diagnosticnih referencnih ravni bo povecala zavedanje o izpostavljenosti pacientov pri rentgenskih preiskavah v Sloveniji. Njihova ustrezna uporaba bo vplivala na zmanjšanje izpostavljenosti v še ne optimiziranih postopkih in tako prispevala k dobri radiološki praksi. Radio! Oncol 2006; 40(1): 197-200. Notices Notices submitted for publication should contain a mailing address, phone and/ or fax number and/ or e-mail oj a Contact person or department. Oncology October 8-12, 2006 The ESTRO 25 / ECCO 14 Conference will take place in Leipzig, Germany. Contact FECS office, Av. E. Mounierlaan, 83/4, B-1200 Brussels, Belgium; or call +32 2 775 93 40; or fax +32 2 779 54 94; or e-mail info@estro.be; or see http://www. estroweb.org Radiation oncology October 22-27, 2006 The ESTRO teaching course »Evidence-Based Radiation Oncology: Methodological Basis and Clinical Application« will take place in Giardini Naxos, Italy. Contact ESTRO office, Avenue E. Mounierlaan, 83/12, B-1200 Brussels, Belgium; or call +32 2 775 93 40; or fax + 32 2 779 54 94; or e-mail info@estro.be; or see http://www.estro.be Lung and head & neck October 25-28, 2006 The »4th Lung & Head and Neck Conference« will be offered in Chicago, Illinois. Contact: Taryn Klocke; call +1 770-984-5113; or e-mail evokes@medicine.bsd.uchicago.edu Oncology October 26-29, 2006 The »2nd Congress of the Polish Oncology« will take place in Poznan, Poland. See http://www.kongresonkologii.pl Lung cancer November 8-12, 2006 »3rd The IASLC/ ASCO/ESMO International Conference on Targeted Therapies in Lung Cancer« will be held in Taormina, Sicily, Italy. Contact E-mail: fred.hirsch@UCHSC.edu Head &neck November 16-18, 2006 The »5th European Workshop on Basic Biology of Head & Neck Cancer« will take place in Poznan, Poland. See http://www.orl.amp.wdu.pl/5workshop Radiotherapy November 19-23, 2006 The ESTRO teaching course »IMRT and Other Conformal Techniques in Practice« will take place in Gliwice, Poland. Contact ESTRO office, Avenue E. Mounierlaan 83/12, B-1200 Brussels, Belgium; or call +32 2 775 93 40; or fax +32 2 779 54 94; or e-mail info@estro.be; or see http://www.estro.be Surgical oncology November 30 -December 2, 2006 The » 13th Congress of the European Society of Surgical Oncology ESSO 2006» will take place in Venice, Italy. Contact Conference Secretariat, ESSO 2006, Federation of European Cancer Societies, Avenue E. Mounier, 83, B-1200 Brussels, Belgium; or call +32 2 775 02 01; or fax +32 2 775 02 00; or e-mail ESS02006@fecs.be; or see http://www.fecs.be Radio/ Oncol 2006; 40(3): 201-2. Radiotherapy December 3-7, 2006 The ESTRO teaching course »Image-guided Radiotherapy (IGRT)« will take place in Brussels, Belgium. Contact ESTRO office, Avenue E. Mounierlaan 83/12, B-1200 Brussels, Belgium; or call +32 2 775 93 40; or fax +32 2 779 54 94; or e-mail info@estro.be; or see http://www.estro.be Toxicology July 15-19, 2007 The »11th International Congress of Toxicology« will be offered in Montreal, Canada. Contact Congress Secretariat, e-mail: ict2007@nrc­cnrc.gc.ca; or see http://www.ict2007.org Lung cancer September 2-6, 2007 The »12th World Conference on Lung Cancer« will be offered in Seoul, Korea. Contact Conference Secretariat; e-mail WCLC2007@ ncc.re.kr; or see http://www.iaslc.orgiumages/ 12worldconfannounce.pdf Oncology September 23-27, 2007 The »14th European Cancer Conference ECCO 14» will take place in Barcelona, Spain. Contact Conference Secretariat, ECCO 14, The European Cancer Conference, European Cancer Societies (FECS), Avenue E. Mounier, 83, B-1200 Brussels, Belgium; or call +32 2 775 02 01; or fax +32 2 775 02 00; or e-mail ECC014@fecs.be; or see http:// www.fecs.be Lungcancer August 21-24, 2009 The »13th World Conference on Lung Cancer« will be offered in San Francisco, USA. Contact Conference Secretariat; e-mail WCLC2007@ ncc.re.kr; or see http://www.iaslc.orgiumages/ 12worldconfannounce.pdf 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. Radio/ Onco/ 2006; 40(3): 201-2. C 2nd CENTRAL EUROPEAN COURSE (CEC 2006) METHODOLOGY OF CLINICAL TRIALS IN ONCOLOGY REPORT The second Central European course „Methodology of Clinical Trials in Oncology", held by the CESAR Central European Society for Anticancer Drug Research-EWIV (CESAR) in collaboration with the Ludwig Boltzmann-Institute for Applied Cancer Research (LBI-ACR VIEnna) and organised by the Applied Cancer Research -lnstitution for Translational Research Vienna (ACR-ITR VIEnna), took place in Vienna from August 24th -26th , 2006. This course has been established by Christian Dittrich during his presidency of the CESAR and in his function as president of the ACR-ITR VIEnna as well as director of the LBI-ACR VIEnna. The aim of the practice-orientated course was to provide the participants with an overview on the actual trial's methodology and to improve their knowledge in performing clinical trials focussed on oncology. The attendees were confronted with bio-statistical as well as pharmacological basic issues of the clinical trial's methodology. Special emphasis was put on the translational aspect of analysing and using preclinical data to be implemented in clinical research. The participants were introduced into the planning, performance and evaluation of clinical trials of the phases 1, II and III. Moreover, practical aspects of the separate phases such as aims, end points, criteria of evaluation and quality assurance were presented and discussed with the audience. Half a day was dedicated to the development of a phase II study protocol. In the frame of a workshop, the referee designed, together with the audience, a phase II protocol suitable for assessing a novel raf kinase inhibitor in patients with locally advanced and/or metastatic hormone refractory prostate cancer, pancreatic cancer and non-small celi lung cancer. Basic knowledge of the study design, organisation and administration comprising both medical and legal aspects of the clinical trial's methodology was presented and analysed in relation to the ICH-GCP guidelines and to multidisciplinary interactions between individual persons/organisations such as clinical investigator, monitor, sponsor, CRO, etc. Data management was discussed with regard to guidelines, recommendations, and warnings as well as actual experiences. The two-day course closed with a presentation summarising the problems and pitfalls of performing clinical trials. This second course was attended by 27 participants from 1 O nations (Austria, Belgium, Germany, Greece, Hungary, Mexico, Portugal, Slovenia, Switzerland, United Kingdom) with different professional background (physician/medical oncologist, pharmacist, biologist, statistician, technician) as well as with different relationship to the performance of clinical studies (investigator, monitor, sponsor, trial manager/coordinator, project manager, clinical research associate). There existed Glose interactions between participants and faculty; discussions and exchange of experiences took place during the sessions as well as the breaks and meals. The acceptance of the course is reflected by the evaluation forms, analysing the referees with regard to the content of their presentations, their presentation skills and the quality of their course materials provided as well as the course in general and the choice of the conference facilities, which were completed by 17 out of 27 participants. The final evaluation encourages us to organise a 3rd course in Vienna from August 23rd -25th , 2007. IIIIIDtlfllllilllJltt,11,1 .... WNlll,CM.Alt_,.\.INlf The CEC 2006 was supported by the Federal Ministry of Economic Affairs and Labour FONDACIJA "DOCENT DR. J. CHOLEWA" JE NEPROFITNO, NEINSTITUCIONALNO IN NESTRANKARSKO ZDRUŽENJE POSAMEZNIKOV, USTANOV IN ORGANIZACIJ, KI ŽELIJO MATERIALNO SPODBUJATI IN POGLABLJATI RAZISKOVALNO DEJAVNOST V ONKOLOGIJI. MESESNELOVA 9 1000 LJUBLJANA TEL 0 1 51 9 1 2 77 FAKS 01 251 81 13 ŽR: 501 00-620-1 33-05-1 0331 1 5-2 1 4 779 Activity of »Dr. J. Cholewa« Foundation for Cancer Research and Education -a report for the third quarter of 2006 The Dr. J. Cholewa Foundation for Cancer Research and Education is preparing plans to continue to support activities associated with cancer research and education in Slovenia in the coming months in late 2006 and early 2007. A number of requests for research grants and other forms of financial support were received from experts in various disciplines of cancer research and education in Slovenia. Needless to say, all the requests have been dealt with great attention and responsibly in respect to their contents and details by the Foundation members with clinical and research experience in cancer and by members with important experience in finance. It is important to note that a number research and study grants have simultaneously been bestowed to researchers and experts from various scientific fields associated with oncology already this year in Slovenia. Many of them were also given various forms of support to attend scientific meetings, conferences and symposia dealing with oncology worldwide. As a result of these activities, the information and knowledge of cancer in general and problems associated with cancer research have thus spread among interested professionals and others in Slovenia. For this reason, the Dr. J. Cholewa Foundation for Cancer Research and Education also continues to support the regular publication of »Radiology and Oncology« international scientific journal, which is edited, published and printed in Ljubljana, Slovenia. This support is considered to be one of its more important commitments and with this in mind, the Foundation will also continue to support the publication of the results from research it supported in any other respectable international scientific oncology journal and other form of dissemination of scientific information deal­ing with cancer research and education. Again, it is also important to emphasize that the Foundation remains active in promoting any form of cancer education in general popula­tion, among medica! and nursing students and among all the others with a particular inter­est in cancer research and education. The Dr. J. Cholewa Foundation for Cancer Research and Education thus has every reason to respectfully acknowledge the importance of various forms of support from various public companies and private individuals to its cause. Without their goodwill the Foundation's activities would have been to a large extent limited. 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Through 1ndustry­the d1agnost1c and treatment modal,t,es That's what we leading technology, increased product1v1ty measures for call Best Practice Oncology Care Siemens medica 1 Solutions that help Sanolabor •.•:• k o tt e r m a n n • Kottermann (Nemcija): laboratorijsko pohištvo, varnostne omare za kisline, luge, topila, pline in strupe, ventilacijska tehnika in digestorji Angelantoni scientifica (Italija): hladilna tehnika in aparati za laboratorije, transtuzijo, patologijo in sodno medicino CORNING Corning (Amerika): specialna laboratorijska plastika za aplikacijo v imunologiji, mikrobiologiji, virologiji, ipd., mehanske eno-in v. ck.na.n: pipete in nastavki • l t.: . . MICRONIC Micronic (Nizozemska): sistemi za shranjevanje vzorcev, pipete, nastavki za pipete Tmpl/anL"eeh· Therc's No Rtason to Operatc with Anyonc Elsc Implantech (Amerika): obrazni in glutealni vsadki Biomerica (Amerika): hitri testi za diagnostiko, EIA /RIA testi LABOR MED d.o.o. Bežigrajski dvor Ehret (Nemcija): Laminar flow tehnika, inkubatorji, sušilniki, suhi sterilizatorji in oprema za laboratorijsko vzrejo živali -kletke Dako (Danska): testi za aplikacijo v imunohistokemiji, patologiji, mikrobiologiji, virologiji, mono-in poliklonalna 'r§ Sakura finetek (Evropa): aparati za pripravo histoloških preparatov: mikro­inkriotomi, zalivalci, tkivni procesorji, barvalci, pokrivalci IBS INTEGRA · WWc11'1if1tl Integra Biosciens (Švica): laboratorijska oprema za mikrobiologijo, biologijo celic, molekularno biologijo in biotehnologijo SpectrumDesigns MEDICAL (Amerika): moški pektoralni vsadki Byron (Amerika): liposuktorji in kanile za liposukcijo Periceva 29, Ljubljana in f o@I ob orm ed. si Tel.: (0)1 436 49 01 Fax: (0)1 436 49 OS www.laborme Erbitux 2 mg/ml raztopina za infundiranje (skrajšana navodila za uporabo) Cetuksimab je monoklonsko lgGl protitelo, usmerjeno proti receptorju za epidermalni rastni faktor (EGFR). Terapevtske indikacije: Zdravilo Erbitux je v kombinirani terapiji z irinotekanom indici rano za zdravljenje bolnikov z metastatskim rakom debelega crevesa in danke, in sicer po neuspešni citotokslcni terapiji, ki je vkljucevala tudi irinotekan. Zdravilo Erbitux je v kombinaciji z radioterapijo indicirano za zdravljenje bolnikov z lokalno napredovalim rakom skvamoznih celic glave in vratu. Odmerjanje in nacin uporabe: Zdravilo Erbitux 2 mg/ml se daje z intravensko infuzijo prek linijskega filtra. Zdravilo Erbitux pri·vseh· indikacijah infundirajte enkrat na teden. Zacetni odmerek je 400 mg cetuksimaba/m' telesne površine, vsi naslednji tedenski odmerki so vsak po 250 mg/m'. Pred prvo infuzijo mora bolnik prejeti premedikacijo z antihistaminikom. Ta premedikacija je priporocljiva tudi pred vsemi naslednjimi infuzijami. Kontraindikacije: Zdravilo Erbitux je kontraindicirano pri bolnikih z znano hudo preobcutljivostno reakcijo (3. ali 4. stopnje) na cetuksimab. Pred zacetkom kombiniranega zdravljenja morate upoštevati kontraindikacije za irinotekan ali radioterapijo. Posebna opozorila in previdnostni ukrepi: ce pri bolniku nastopi blaga ali zmerna reak5ija, 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 (2c 3. stopnje po kriterijih NCI-CTC), morate prekiniti terapijo s cetuksimabom. Z zdravljenjem smete nadaljevati le, ce se je reakcija pomirila do 2. stopnje. 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 ravni jetrnih encimov, kožne reakcije, blage ali zmerne reakcije, povezane z infundiranjem, blag do zmeren mukozitis. Pogosti (2c 1/100, < 1/10): konjunktivitis, hude reakcije, povezane z infundiranjem. Pogostost ni znana: hipomagneziemija. Pakiranje: 1 viala po 50 ml. Imetnik dovoljenja za promet: Merek KGaA, 64271 Darmstadt, Nemcija. Podrobne informacije o zdravilu so objavljene na spletni strani Evropske agencije za zdravila (EMEA) http://www.emea.eu.int/ Dodatne informacije so vam na voljo pri: Merek, d.o.o„ Dunajska cesta 119, 1000 Ljubljana, tel.: 01 560 3810, faks: 01 560 3831, el. pošta: info@merck.si ' plošcatocelicni rak glave in vratu 2 1 v primerjavi z radioterapijo II 3 Bonner et al. Radiotherapy plus Cetuximab for Squamous -Celi Carcinoma 1 II opo n1 no zdravljenje* rimidex vodilni zaviralec aromataze anastrozol Kratka informacija o zdravilu Arimidex 1 mg ,stava: Filmsko obložena tableta vsebuje ng anastrozola. jikaclje: Adjuvantno zdravljenje žensk po mopavzi, ki imajo zgodnji invazivni rak jke s pozitivnimi estrogenskimi receptorji se ne morejo zdraviti s tamoksifenom radi povecanega tveganja za mboembolizem ali nenormalnosti dometrija. Zdravljenje napredovalega