ADIOLOGY AND NCOLOGY December 2007 Vol. 41 No. 4 Ljubljana ISSN 1318-2099 farmacevtska družba, d.o.o. 1 mm, treated with radical vulvectomy and inguinofemoral lymphadenectomy, is nearly 70%. The most significant prognostic factor is the number of metastatic inguinofemoral lymph nodes. Despite fairly high survival of patients, short- or long-term morbidities associated with the above mentioned radical surgical technique are worryingly high. These mor­bidities, which considerably prolong hospi­talization, are infections, lymphocysts, scar dehiscence and lymphatic edema of the lower limbs. A standard treatment modality applied in early stages of vulvar carcinoma (T1, tumor <2cm; T2, tumor >2cm), not accom­panied by palpable inguinal lymph nodes enlargement, is extensive local excision of the tumour with the safety margin of 1 cm and uni- or bilateral inguinofemoral lymphadenectomy, both performed in two separate incisions. Postoperative radiother­apy is indicated if more than one node is metastatic. In early stages of the disease, metastat­ic involvement of inguinofemoral lymph nodes is observed in 20-30% of patients. The rest of the patients (70%) do not ben­efit significantly from surgical intervention; however, a considerable increase in morbid­ity rates has been observed. Since the intro­duction of the technique with two separate incision sites, the number of complications decreased, though lymphoedema, lym­phocysts or scar dehiscence are still fairly frequent. No reliable method for determining the inguinofemoral nodes status has been so far developed. Palpation can detect just about 25% of all metastatic nodes. The re­sults of ultrasound (US) and positron emis­sion tomography (PET) examinations are unsatisfactory, whereas the potential of the computer tomography (CT) examination in the search of metastatic nodes has not been extensively described in the literature. The sensitivity of magnetic resonance imaging (MRI) was assessed to range between 40­50% and the specificity, between 90-100%. Among the most promising methods for detecting metastases in the regional nodes is US-guided fine-needle biopsy though, ad­mittedly, it requires highly skilled operators in order to yield success and efficiency.1 On account of so sparse non-invasive techniques for determining the inguinofem-oral nodes status, of non-involved nodes in the majority of patients with a low stage vulvar carcinoma and on account of fre­quent morbidity following lymphadenec­tomy, there was an urgent need to develop a minimally invasive surgical technique for diagnosing vulvar carcinoma, i.e. sentinel lymph node detection (SLND). Sentinel lymph node – what is it? The sentinel lymph node is the first node in the lymphatic basin into which the lymph from the primary tumour is drained; histological examination of the sentinel lymph node is hypothetically representa­tive for the rest of the nodes in the region. A histologically negative sentinel lymph node should be indicative for the absence of the metastases in all other non-sentinel nodes. 2 Sentinel lymph node biopsy – when and how? The sentinel lymph node biopsy should be performed in the patients in whom • vulvar carcinoma with the in-depth inva­sion of more than 1 mm was confirmed histologically, • injection of the necessary substances into the tumour surrounding is possible, and • no enlarged or fixed lymph nodes were de­tected in the inguinal region. Radiol Oncol 2007; 41(4): 167-73. Before the operation, the patient should be fully informed of the details of the ex­amination, to what purpose the results of the examination would serve, how the ex­amination would affect further treatment, and that the treatment is only experimen­tal. The treatment can be started only after obtaining the informed consent signed by the patient. The sentinel lymph node should always be marked in two ways, i.e. with 99mTc (Technetium) labeled nanocolloid and with blue dye; this is the most reliable marking method assuring that the node will abso­lutely be found later. According to the litera­ture, the injection of blue dye alone detects the sentinel node only in 56-88% of cases.3 Given that the injection of nanocolloid is a painful procedure, it is recommended that an anaesthetic gel is applied to vulva before the injection;4 at our Institute, we usually apply local anaesthetic cream EMLA. Colloid is then injected with a fine nee­dle at four separate injection sites along­side the peripheral tumour margin. Care should be taken that nanocolloid does not get spilled over the vulva or inguinal area because tiny radioactive particles detected by scintigraphy may compromise the image interpretation. The patient is immediately transported to the isotope treatment unit where first a dynamic and then a static lymphoscintig­raphies are performed by gamma camera. The first active and persistent spot found by this technique is sentinel node; its loca­tion is marked on the skin. Sometimes, two highly active sites may be found; in these cases, both are marked on the skin.1 The blue dye is applied immediately be­fore surgery. It is injected at four separate sites alongside the tumour margin. On the site marked on the skin incision, approxi­mately 3-4 cm long, is made; the tissue is then carefully pushed aside in order to avoid severe haemorrhage that would ob­struct the detection of the blue-dyed node. The node’s activity is checked by portable gamma-ray detector. The node is then cau­tiously excised. If the next node is also dyed blue, it should be removed, too despite lower activity. If the carcinoma is located in the centre, the sentinel node on the other side should be removed, too. After sentinel lymph nodectomy, the excision of tumour with a wide safety margin should be per­formed. Radical vulvectomy is recommend­ed only in cases of multifocal growth of the tumour. The nodes excised during surgery may be sent to cytology laboratory for intraop­erative evaluation of the samples by imprint cytology. The frozen section technique is not advised because it destroys the excised node completely so that no further exami­nations are possible. However, the surgeon may finish the operation a few days later af­ter having received the results of the imprint cytology. If the results confirm metastatic involvement in the node, inguinofemoral lymphadenectomy or postoperative irra­diation of the affected area should be per­formed; the decision between the two op­tions depends on health condition of each patient and on other of prognostic factors. Patients and methods In 2003, in view of the data from the literature reporting a 100% negative prog­nostic value3 we decided to apply a new method of treatment, i.e. removal of sen­tinel lymph nodes alone. The Department of Surgical Oncology at the Institute of Oncology Ljubljana is known to be very skilled in sentinel lymph nodes biopsies in the treatment of breast cancer and malig­nant melanoma [2005 - 537 biopsies (383 breast cancer, 147 melanoma malignum), 2006 – 529 biopsies (351 breast cancer, 172 melanoma malignum)]. Their experience was most welcome also in gynaecological oncology and facilitated the introduction of these techniques also in gynaecological oncology. From March 2003 to the end of 2006, 35 patients were treated for vulvar carcinoma with the technique of sentinel lymph node dissection. Mean age of the patient was 65.8 years (range 36-88 years). According to histology analysis results, 32 patients had squamous cell carcinoma, one malignant melanoma, one basal cell carcinoma, and one adeno-squamous cell carcinoma. Two hours before surgery, the patients were injected intradermally with 99mTc na­nocolloid in four quadrants lining the out­er margins of the tumour. Static and also dynamic lymphoscintigraphies (Figure 1) were then performed detecting active nodes and the locations were marked on the skin (Figure 2). Before surgery, methylene blue dye was injected intradermally, too. The first five operations were performed under the su­pervision of an IAEA instructor. During operation, sentinel nodes were detected by hand-held gamma probe. In all cases, we first excised the lymph nodes that were dyed blue and active and then the nodes that were not dyed, but still active. After the removal of the nodes, the tumour on the vulva was excised. In all surgical in­terventions, a conservative approach was followed, i.e. only the tumour with a wide safety margin was removed and not the en­tire vulva. In 19 cases, unilateral nodectomy was performed, whereas in 16 cases, nodec­tomy was bilateral. Results Metastatic nodes Sentinel node biopsies were positive (SNB+) in 10 out of 35 patients (28.6%), which is consistents with the data from the litera­ture. In 4 patients, the sentinel nodes were identified bilaterally, and in two cases, the nodes were metastatic on both sides. In all patients, postoperative care was normal, without serious complications. In one patient, a cluster of clinically metastatic nodes was detected behind the positive sentinel node. Therefore, all nodes Figure 1. Vulvar static lymphoscintigraphies with 99mTc nanocolloid. Radiol Oncol 2007; 41(4): 167-73. down to the femoral canal were removed. The patient underwent also CT examina­tion which detected that deeply seated pel­vic lymph nodes were also metastatic. In 3 patients, reoperation (inguinofemo­ral dissection) was required, and 9 were treated with postoperative irradiation. Six patients died of the disease spread; in 4 Table 1. Positive sentinel node biopsies (SNB+) patients, the disease progressed into the inguinal area; in four patients, no evidence of disease was found. In one patient, local recurrence was observed 43 months after the completed therapy. Local excision was performed and the patient was also irradi­ated. The patient was without evidence of disease after 55 months (Table 1). PT Histology SNB Treatement Age Status (nov 2007) Time (months) 1 Squamous, G1 + L Reoperation + irradiation 57 NED 55 2 Squamous, G2 + L, + R + irradiation 69 DOD 42 3 Squamous, G3 + R + irradiation 58 DOD 10 4 Squamous, G3 + L + irradiation 59 DOD ? 5 Squamous, G2 - R, + L + irradiation 63 NED 36 6 Squamous, G2 +R Reoperation + irradiation 65 DOD 3 7 Squamous + R + irradiation 71 DOD 18 8 Squamous, G2 + R, + L + irradiation 76 DOD 10 9 Squamous + R, - L Reop. (rad. limfadenectomy) 67 NED 25 10 Adenosquamous + L + irradiation 73 NED 11 Table 2. Negative sentinel node biopsies (SNB-) PT FIGO Stage Histology SNB Age STATUS (nov 2007) Time (months) 1 IB Squamous, G1 - L, - R 61 NED 34 2 II Squamous, G2 - R 78 NED 51 Groin reccurence 3 II Squamous, G1 - L, - R after 24 months - reoperation 80 DOD 49 4 I Squamous - L, - R 70 NED 52 I Squamous, G1 - R 70 NED 46 6 I B Squamous - L 63 NED 46 Groin reccurence 7 I Baseo- Squamous - L, - R after 26 months - reoperation 67 NED 43 8 I Planocelular, G3 - L, - R 46 NED 45 9 I Squamous, G1 - L 49 NED 46 I B Squamous, G1 - R 76 NED 35 11 I B Microinvasive, (1,6mm) - L, - R 72 NED 38 12 II Squamous, G1 - R 87 DOC Groin reccurence 13 I B Squamous, G1 - R (scar) after 12 months, reoperation 80 NED 36 14 II Bazeocelular - R 83 NED 31 II Squamous, G2 - R, - L 88 NED 18 16 I Squamous, G1 - L 36 NED 33 17 II Squamous, G1 - R, - L 48 NED 35 18 I B Squamous, G1 - R, - L 53 NED 28 19 Melanoma malignum - R, - L reoperation +irradiation 47 AWD 33 I Squamous, G1 - R 75 NED 25 21 I Squamous, G1 - R, - L 50 NED 18 22 I Squamous, G2 - R 79 NED 2 23 I Squamous, G1 - R 58 NED 14 24 I Squamous - R 63 NED 18 I Squamous, G1 - R, - L 66 NED 15 L = left; R = right Nodes without metastatic involvement Sentinel node biopsies were negative (SNB-) in 25 out of 35 patients (71.4%). Postoperative complications were not ob­served; the patients were therefore dis­missed from the hospital 2 or 3 days after surgery. In three patients, recurrence in the inguinal region was detected 12, 24 and 26 months after the completed primary treat­ment, respectively. In one patient, the cause Radiol Oncol 2007; 41(4): 167-73. of the recurrence could be assigned to the inexperience of the surgeon; in the second, the node, next to the sentinel node, which was active, was not identified; and in the third patient, the active node did not show up because of a scar of an earlier operation in the inguinal region. By the end of 2007, 22 patients are alive, 21 of them with no evidence of disease (NED), one died with groin recurrence and lung metastases after 49 months, one died of other causes, and patient with melano­ma malignum is alive with the disease (liver metastases) (Table 2). One patient (N° 22) was lost in the follow up. Discussion The first who, as early as 1991, assumed that the superficial inguinal lymph nodes in vulvar carcinoma serve as sentinel nodes for the deep-lying ones was DiSaia et al. The first study dealing with this subject was published by Levenback et al in 1994. He and his team detected the sentinel nodes by using blue dye and verified the reliability of the method by complete in-guinofemoral lymphadenectomy performed later on. Three years later, deCesare published the results of his study made on intraop­erative sentinel node detection by using a gamma-ray detector. Both methods were then merged by Hullu who used lympho­scintigraphy to mark the nodes preopera­tively and the blue dye to detect them intra­operatively.1 Both sentinel lymph node marking tech­niques are suitable; however, the best results are usually obtained by the combination of both. From the reported data, it is evident that the methods applying the combination of both yield a 100% success, whereas blue dye applied alone is successful in only 56­88% of cases.5 Conclusions Sentinel node biopsy is a promising di­agnostic tool in the evaluation of vulvar carcinoma and of node status as well as of disease stage. Due to low incidence of squamous cell carcinoma of the vulva, the experiences in the above treatment of this cancer are not so rich and the data in the literature are scarce; therefore, this treat­ment modality is applied more or less as an experimental method as it has not been yet approved as a standard method. In order to recognize the value of the obtained knowl­edge about the new treatment method and to confirm the efficiency or inefficiency of this method, further multicentre research studies need to be performed on larger ran­domized groups of patients. References 1. de Hullu JA, van der Zee AG. Sentinel node teh­niques in cancer of the vulva. Curr Womens Health Rep 2003; 3: 19-26. 2. Sliutz G, Reinthaller A, Lantzsch T, Mende T, Sinzinger H, Kainz C, et al. Lymphatic mapping of sentinel nodes in early vulvar cancer. Gynecol Oncol 2002; 84: 449-52. 3. Puig-Tintore LM, Ordi J, Vidal-Sicart S, Lejárcegui JA, Torné A, Pahisa J, et al. Further data on the usefulness of sentinel node identification and ultrastaging in vulvar squamous cell carcinoma. Gynecol Oncol 2003; 88: 29-34. 4. De Cicco C, Sideri M, Bartolomei M. Sentinel node biopsy in early vulvar cancer. Br J Cancer 2000; 82: 295-9. 5. De Hullu JA, Hollema H, Piers DA, Verheijen RH, van Diest PJ, Mourits MJ, et al. Sentinel lymph node procedure is highly acurate in squamous cell carci­noma of the vulva. J Clin Oncol 2000; 18: 2811-6. 6. De Hullu JA, Van der Avoort IA, Oonk MH, van der Zee AG. Management of vulvar cancers. Eur J Surg Oncol 2006; 32: 825-31 Assessment of differential expression of oncogenes in adenocarcinoma of stomach with fluorescent labeling and simultaneous amplification of gene transcripts Uroš Rajcevic1, Petra Hudler1, Gordan Mijovski2, Gregor Gorjanc3, Georg Hoelzl4, Stanislav Repše5, Robert Juvan5, Milena Kovac3, Christian G. Huber6 and Radovan Komel1 1Medical Center for Molecular Biology, Faculty of Medicine, Institute of Biochemistry, Ljubljana, Slovenia; 2Clinical Department for Anesthesiology and Intensive Care of Operative Branches, University Clinical Center, Ljubljana, Slovenia; 3Department of Animal Science, Biotechnical Faculty, Domzale, Slovenia; 4Institute of Analytical Chemistry and Radiochemistry, Leopold-Franzens University, Innsbruck, Austria; 5Clinical Department for Abdominal Surgery, University Clinical Center, Ljubljana, Slovenia; 6Instrumental Analysis and Bioanalysis, Saarland University, Im Stadtwald, 66123 Saarbruecken, Germany Background. Gastric cancer is one of the leading malignancies with a poor prognosis and low survival rates. Although the mechanisms underlying its development are still unknown, there is a consensus that genetic instability, inactivation of tumor suppressor genes and over-expression of oncogenes are involved in the early and late stages of gastric carcinogenesis. In the present study we wanted to display differential expression of seven oncogenes, namely CCNE1, EGF, ERBB3, FGF4, HRG1, HGFR and TDGF1. Patients and methods. We employed a method based on the multiplex reverse transcription polymerase chain (RT-PCR) method with a fluorescence detection. Results. More than half of patients (74.3%) out of total 74 with gastric adenocarcinoma had over-expressed at least one oncogene, with the exception of FGF4, which was expressed in tumor tissue of less than one third of patients. 56.8% of the patients patients showed over-expression of two or more oncogenes. Conclusions. Patients with precancerous lesions had elevated levels of TDGF1 or cripto-1 (64.9%) and CCNE1 (57.1%), suggesting that they could be used as markers for an early detection of malignant changes in stomach. Finally, the fluorescent multiplex RT-PCR method could be of value for rapid assessment of oncogene mRNA levels in small samples of tumor or precancerous biopsies. Key words: stomach neoplasms – genetics; adenocarcinoma; reverse transcriptase polymerase chain reac­tion; oncogenes Received 16 October 2007 Accepted 26 November 2007 Correspondence to: Prof. Radovan Komel, Ph.D., Medical Center for Molecular Biology, Faculty of Medicine, Institute of Biochemistry, Vrazov trg 2, SI­1000 Ljubljana, Slovenia. Phone: +386 1 543 7644; Fax : +386 1 543 76 41; E-mail: radovan.komel@mf.uni-lj.si Introduction Changes in gene expression are at the core of development and differentiation, and play a decisive role in many pathological processes such as carcinogenesis. Gastric cancer (GC) is a leading cause of cancer mortality worldwide, surpassed only by lung and breast cancer.1 The leading histo­logical form is adenocarcinoma, accounting for more than 9/10 of all cases, which can be subdivided into two clinicopathologi-cal entities: intestinal or well-differentiated and diffuse or poorly differentiated GC.2 Recent molecular analyses have clari­fied many genetic alterations in gastric car-cinogenesis but this is hardly sufficient to understand the underlying mechanism(s).3 Furthermore, the disease shows diverse clinical properties regarding invasiveness, ability to form metastases and responsive­ness to chemotherapy. The transformation of normal gastric epithelial cells into cancer is a multistep process associated with the progressive accumulation of abnormalities in DNA-repair genes, tumor suppressor genes and oncogenes. Increased or exces­sive expression of oncogenes could, under certain conditions, induce a neoplastic transformation of cells.4 A current hypothesis predicts the multistep process of gastric carcinogen-esis, which involves, among other, genetic changes of multiple oncogenes.4-6 Thus, assessment of expression of putative onco­genes might be of clinical and prognostic importance.7-9 Although it was believed that in the sequential multi-step tumori-genesis process, inactivation of a single mutant gene product would be insufficient to obtain tumor regression, recent data proved otherwise.10 In mouse models even a transient inactivation of a transforming oncogene (Ras, Myc or Bcr-Abl) was suf­ficient for reversion of malignant pheno­type.10,11 Targeting oncogenes could thus be a promising approach in treatment and evidence exists about successful results ob­tained in humans treated with antibodies for the Her-2 receptor against metastatic breast cancer, and with Bcr-Abl kinase i inhibitor, imatinib mesylate (Glivec), in chronic myelogenous leukemia (CML) and gastrointestinal stromal tumors (GIST). The evidence of oncogene expression could also be useful for selecting the most effec­tive therapy. Recent studies showed that in lung cancer, c-Met (hepatocyte growth fac­tor receptor, HGFR) amplification leads to gefitinib or erlotinib resistance, however, on the other hand, the research on GC cell lines with this defect showed their extreme sensitivity to another type of chemothera­peutics, specific tyrosine kinase inhibitor PHA-665752.7,8 Therefore, even though the mechanisms of gastric carcinogenesis are still elusive, it seems that in the future a rapid and reliable assessment of certain key oncogenes might be very important for both diagnosis and treatment. Usually, the study of differential expression of indi­vidual oncogenes is a laborious and time consuming procedure. In this work, we developed a diagnostic approach based on fluorescent multiplex quantitative reverse transcription polymerase chain reaction (RT-PCR) assay and capillary electrophore­sis separation, which allows insight and relative quantification of differential ex­pression of a number of oncogenes simulta­neously. With these two simple techniques seven oncogenes associated with molecu­lar genesis of gastric adenocarcinoma (cy­clin E (CCNE1), epidermal growth factor (EGF), fibroblast growth factor 4 (FGF4), teratocarcinoma-derived growth factor 1 or Cripto-1 (TDGF1), hepatocyte growth fac­tor receptor (HGFR) or c-Met, neuregulin 1 or heregulin (HRG1 or NRG1) and v-erb-b2 erythroblastic leukemia viral oncogene ho-molog 3 (avian) or c-erbB3 (ERBB3)) were screened in samples of tumors and cor­responding normal mucosa of 74 patients with adenocarcinoma of the stomach, and in samples of precancerous lesions (chron­ic atrophic gastritis, ulcers and intestinal metaplasia/dysplasia) and corresponding normal tissues of 77 patients. Materials and methods Patients Samples of tumors and corresponding nor­mal mucosae of 74 patients with GC were contributed by the Clinical Department for Abdominal Surgery, University Clinical Center Ljubljana. Samples of precancer­ous tissues were obtained at the Clinical Department for Gastroenterology, University Clinical Center Ljubljana. The portions of tumor and precancerous tissues and ad­jacent non-tumor mucosa were carefully sampled and frozen at -70°C for DNA ex­traction. Representative portions of tumor samples were also formalin-fixed and par-affin-embedded for immunohistochemistry and histology. A histological evaluation of these samples was confirmed at the Institute of Oncology Ljubljana. Most GC patients were male with 1.6:1 male-to-female ratio. The mean age at the surgery was 64.5 ± 11.4 years (mean ± standard deviation (SD)). The distribution of tumors based on Lauren’s classification system in the series included 40 (54.8%) intestinal types, 17 (23.3%) dif­fuse and 16 (21.9%) cases of mixed type. With respect to Ming’s classification, the cases were ranked into 24 (32.9%) expan­sive, 35 (47.9%) infiltrative and 13 (17.8%) mixed types. Patients with precancerous lesions were handled as one group, because grouping 11 patients with intestinal meta-plasia/dysplasia, 14 cases with ulcers and 52 with chronic atrophic gastritis was not feasible for relevant statistical analysis. The study was approved by the National Medical Ethics Committee and informed consent was obtained from participants. All samples used in the study were anonymized and la­beled with an appropriate number. Total RNA extraction and purification Total RNA was extracted from pairs of tissue samples using TRIzol reagent (Invitrogen) or RNeasy Mini Kit (Qiagen) according to manufacturer’s instructions. To remove residual impurities such as DNA, proteins and sugars, total RNA was additionally purified using DNAseI (Qiagen), extracted with the mixture of phenol:clorophorm: iso-amyl-alcohol (50:48:2) and precipitated with ethanol. RNA pellet was vacuum-dried and diluted in 8 µl of nuclease free, sterile water (Invitrogen). Purity and concentra­tion were assessed spectrophotometricaly (Beckman UD 7500) at 260 nm and 280 nm. The quality was checked with electrophore­sis on agarose gel. For further procedures samples of RNA with the A260/A280 ratio of equal or more than 1.8 and sufficient consistency were applied. Multiplex RT-PCR Primers for the simultaneous amplification of fragments of seven oncogenes (CCNE1, EGF, ERBB3, FGF4, HGFR, HRG1 and TDGF1) were selected from coding se­quences using Primer3, web-based prim­er selection software (Table 1). All sense primers were 5’ end-labeled with one of the fluorescent dyes, 6-FAM™, TET™ or HEX™ (Applied Biosystems). Primers were selected in a manner that the fragments that they amplify did not overlay.8 Once the PCR conditions of the 7-gene multiplex reaction were established and optimized, semi-quantitative RT-PCR was performed to study the differential expression of se­lected oncogenes in both the tumor and normal mucosa specimens. As internal standard (IS) we used a house-keeping gene, gylceraldehyde-3-phosphate dehy­drogenase (GAPDH).12 For the synthesis of the first chain of complementary DNA (cDNA) 25 pmoles of each of eight non-labeled, antisense primers (CCNE1, EGF, ERBB3, FGF4, HGFR, HRG1, TDGF1) and 1µl (200 U/µl) of Superscript II Reverse transcriptase (Invitrogen) was Radiol Oncol 2007; 41(4): 174-82. Table 1. Sequences of sense and antisense primers of seven oncogene fragments Oncogene Gene name (Entrez Gene database) Primer sequencea Product length (bpb) C-met HGFR TET -5’-gtgctcctgtttaccttg 5’-tagttagtggcggcaccaag 291 CR-1 TDGF1 HEX -5’-gtgtaaatgctggcacggtc 5’-aaaggcagatgccagctag 224 Cyclin e CCNE1 HEX -5’-gtcaagtacaccagccac 5’-ggatagatatagcagcac 214 C-erbB3 ERBB3 FAM -5’-cgatgctgagaaccaata 5’-acttcccatcgtagacct 171 EGF EGF FAM -5’-gagcgaagctttcatatg 5’-tcactgagtcagctccat 159 FGF-4 FGF4 FAM -5’-caccatgaaggtcacccact 5’-cttacacctactcttgca 148 HRG1 NRG1 TET -5’-tcagcagttcagctccttcc 5’-ttgtgttgctgtccacttcc 141 aPrimer sequences, their fluorescent tags and the lengths of fragments they are amplifying (sense primers are fluorescent tagged); bbase pairs. used. The reverse transcription (RT) reac­tion was performed following instructions of the producer’s protocol (Invitrogen). Next, we performed multiplex PCR, using 1 µl of cDNA, 12.34 µl of multiplex primer mix, 2.5 µl 10x PCR Gold buffer (Applied Biosystems), l dNTP mix (2.5 mM each, Promega)), 1 µl 4% DMSO, 0.4 µl Taq Gold polymerase (Applied Biosystems) and 3.76 µl nuclease free H2O. PCR conditions were as follows: initial denaturation 10 minutes at 95°C, followed by 26 reaction cycles; each cycle composed of 30 seconds of de­naturation at 95°C, 30 seconds of anneal­ing at 52°C and 3 minutes of elongation at 72°C. Multiplex reverse transcription and multiplex PCR with fluorescent tagging and simultaneous amplification of gene transcripts were performed separately on GeneAmp PCR system 9600 (Applied Biosystems). The authenticity of fragment amplification with RT-PCR was established by direct sequencing. Following PCR, re­sidual nucleotides, primers, dissociated fluorescent dyes, salts and the enzyme in multiplex RT-PCR reactions were partially purified with QIAquick PCR-product puri­fication kit (Qiagen). Analysis of RT-PCR products by with capillary electrophoresis with fluorescence detection and by IP-RP-HPLC with UV detection Purified, fluorescently tagged multiplex RT-PCR products were separated and an­alyzed with capillary electrophoresis on ABI-PRISM 310 Genetic Analyzer (Applied Biosystems), according to the producer’s in­structions. GeneScan 500® Tamra (Applied Biosystems) was used as an internal stand­ard. Results were evaluated with GeneScan software (Applied Biosystems). Multiplex RT-PCR products were separat­ed, in parallel experiments, also with liquid chromatography. The purpose of this sepa­ration was to assess whether the same prod­ucts can be adequately separated with this alternative technology and whether the re­sults of both techniques would be compara­ble. Purified non-labeled multiplex RT-PCR products of tumor and normal samples of limited number of patients were separated with ion-pair reversed-phase high perform­ance liquid chromatography (IP-RP-HPLC) on DNASepTM columns (Transgenomic) packed with alkylated nonporous particles Table 2. The percentage of patients showing relative expression (RT/RN) of oncogenes in tumors and pre­cancerous lesions Oncogenes Gene name (Entrez Gene database) RT/RN > 1 (% of GCa patients) RT/RN > 5 (% of GC patients) RT/RN > 1 (% of Pcb patients) C-met HGFR 56.4 10.9 26.0 CR-1 TDGF1 36.0 15.6 64.9 Cyclin E CCNE1 42.0 15.8 57.1 EGF EGF 40.0 12.7 - C-erbB-3 ERBB3 44.0 16.0 13.0 FGF-4 FGF4 59.0 -c - HRG1 NRG1 30.3 12.1 24.7 aGC – gastric cancer; bPc – precancerous lesions; c- – no expression detected. of polystyrene-divinylbenzene (PS-DVB­C18).13 The separation solutions were: A = 0.1 M TEAA + 0.1 mM EDTA and B = A + 25% acetonitrile. Chromatographic condi­tions for the separation of the multiplex RT­PCR products were as follows: 10 min lin­ear gradient 20-50% B at a flow-rate of 0.75 ml/min and 50°C column temperature. 15 µl of the sample was injected onto the col­umn in a single injection. Separated ampli­fied fragments of oncogenes were detected with UV-absorbance at 254 nm. Restriction fragments from pBR332 plasmid, digested with HaeIII, were used as size standards. Assessment of differential expression of oncogenes Electropherograms where the height of internal standard/GAPDH was at least 150 relative fluorescence units (RFU) were con­sidered as positive. For relative quantifica­tion of differences in expression of seven oncogenes in the samples of tumors and corresponding normal tissues the follow­ing method was applied: on the electro-pherograms representing normal tissue, the peak area of the specific oncogene fragment (Pn) was compared to the peak area of internal standard expressed in nor­mal tissue (ISn). The ratio coming out of this (RN) was a relative value of a specific oncogene transcript abundance in normal tissue. In the same way the relative value of transcript abundance was determined in tumors (RT=Pt/ISt). To determine the rela­tive ratio of specific oncogene expression in tumors and normal tissues, relative values of expression of specific oncogene were compared (RT/RN= (Pt/ISt)/(Pn/ISn)). Results Reaction conditions for the multiplex PCR were established gradually by altering indi­vidual parameters in the reactions in order to minimize the background on electroph­erograms (Figure 1). Primers were selected in a manner that the fragments that they amplify did not overlay. In addition, differ­ent fluorescent tagging of sense primers served for a better resolution of the ampli­fied fragments. Carefully adjusted melting temperatures of primers and fragments allowed amplification of all the eight frag­ments at the same annealing temperature and in the same reaction tube. With a relative quantification of differ­ences in oncogene expression in tumors and corresponding normal tissue at least one oncogene was over-expressed in tumors in most (74.3%) of 74 patients, whereas in 25.7% we did not detect expression of on­cogenes under investigation. Thirteen pa­tients (17.5%) showed higher levels of one Radiol Oncol 2007; 41(4): 174-82. oncogene and in forty-two (56.8%) cases simultaneous over-expression of two or more oncogenes was observed. In 24.3% of patients we detected 5- or more fold over-expression of individual oncogenes, while 16.2% showed 5- or more fold over-expres­sion of two or more oncogenes simultane­ously. HGFR (c-Met) was over-expressed in 56.4%, TDGF1 in 36%, CCNE1 in 42%, EGF in 40%, ERBB3 in 44%, FGF4 in 59%, and HRG1 (NRG1) in 30.3% of cases (Table 2). In tissues of precancerous lesions we es­tablished the following expression pattern of investigated oncogenes (Table 2): TDGF1 in 64.9%, cyclin E in 57.1%, HGFR in 26.0%, HRG1 in 24.7% and ERBB3 in 13.0% of ex­amined cases. Interestingly, we did not de­tect EGF and FGF4 mRNA. In parallel experiments we separated non-labeled amplicons of 8 selected patients with HPLC. The results obtained were comparable with the capillary electrophoretic separation of fluorescently labeled products, although we observed that the threshold of UV-absorb­ance was lower, which is in accordance with previously published results.14 Nevertheless, we established that separation of PCR prod­uct could be obtained either with HPLC, using non-labeled fragments, thus reducing the cost. However, capillary electrophoresis was more accurate and sensitive, therefore, in our opinion it is a more reliable method, even though there is a need for fluorescently labeled primers, which are slightly more ex­pensive than non-labeled ones. Discussion The assessment of differential expression of multiple oncogenes with multiplex RT­PCR and capillary electrophoresis separa­tion was designed as one of approaches for analysis of changes in oncogenes involved in stomach carcinogenesis. We developed and optimized a method for the assess­ment of differential expression of seven oncogenes with fluorescent tagging and simultaneous amplification of oncogene transcripts. Activated HGFR, encoded by MET proto-oncogene, was found in 26.0% of precancer­ous lesions and in tumors of 56.4% of GC patients. 10.9% of the later over-expressed it for 5- or more fold, which is in accordance with the reports of several authors.5,6,15,16 It has been shown that deregulation of HGFR is involved in the aberrant turning on of the invasive growth, strongly correlating with a higher metastatic potential of cancer cells and with a poor prognosis in several types of cancer, such as gastric, colorectal, breast, liver, kidney and pancreatic.9,10,17 Recently, it was confirmed that HGFR activation pro­tects cells from apoptosis, induces motile phenotype in conjunction with beta-cat­enin and also promotes entry into cell cy­cle.17 Therefore, we could assume that its deregulation is most prominent in the later stages of GC, which was confirmed by our results. Furthermore, MET amplification could constitute an important biomarker for selecting patients for a targeted therapy. Smolen et al. observed that a fraction of GC cell lines appeared to be exquisitely sensi­tive to a specific MET inhibitor.8 TDGF1 or Cripto-1, an epidermal growth factor–CFC (EGF-CFC) family member, was found up-regulated in 36% of our patients with GC and in 65% precancerous lesions. This gene plays an important role in tumor cell pro­liferation and migration and is involved in Nodal and presumably also in Wnt, MAPK and PI3K/AKT signaling pathways.18-21 Recently, it has been found that Cripto-1 protein is detectable in plasma of breast and colon cancer patients, and even in early pre-malignant lesions of colon, breast and stomach, suggesting its potential use as a biomarker for an early diagnosis.19 It is also interesting that Haruma et al. found a corre­lation of TDGF1 expression with the long-term H. pylori infection, which is a known risk factor for the GC development.22 Interestingly, 8 of 11 patients (72.7%) with precancerous lesions with H. pylori infec­tion had also up-regulated this gene, which is in agreement with previously reported results.19,22 The over-expression of cyclin E in gastric carcinoma was reported by sev­eral authors.23-25 We found up-regulated cy­clin E in tumors of 42% of patients; 15.8% of them over-expressed it 5- or more fold, while in the precancerous tissues it was up-regulated in 57.1% of patients. Moreover, its higher mRNA levels were found in 81.8% (9 out of 11) patients with H. pylori infection. Yu et al. also found association of cyclin E over-expression with intestinal metaplasia, although it was also associated with inva­sion, metastasis and prognosis.25,26 Our results correlate with these findings. In addition, TDGF1 and cyclin E mRNA lev­els were among the highest in our precan­cerous patients, indicating their possible involvement in early changes affecting gas­tric mucosa, especially in patients with H. pylori. Therefore, these two genes are pos­sible candidates for biomarkers for an early diagnosis of malignant changes in stomach epithelium. EGF and HRG1 are members of a large family of EGF-like growth factors that influ­ence a variety of cellular events, including proliferation, migration and survival. Both these factors bind ERBB3 or ERBB4 through heterodimerization mechanism, producing potent activators of cell transformation.27 It has been demonstrated that EGF could activate ERBB3 (and ERBB4) receptors in­directly through EGFR/ERBB3 (or EGFR/ ERBB4) heterodimerization by a transphos­phorylation, while on the other hand, HRG proteins act directly as ligands for ERBB family of receptor tyrosine kinases, specifi­cally for ERBB3 and ERBB4.27 The expres­sion of ERBB3 and EGF was reported to be elevated in gastric tumors.25,28,29 There are few data of HRG1 over-expression in stom­ach adenocarcinoma, but neoplastic pathol­ogies with HRG1 expression were found in breast, ovary, prostate, small intestine and brain.27 The co-expression of these onco­genes might play important roles in ma­lignant potential of GC and precancerous cells. HRGs, ERBB3 and EGF are involved in the regulation of cellular proliferation, differentiation, migration, apoptosis and survival and angiogenesis, therefore, the fine tuning of this cascade most probably directs cell proliferation and differentiation upon the growth factor stimulation, while its deregulation probably plays an impor­tant role in the tumor growth.27 The elevat­ed levels of all three genes in our GC pa­tients and slightly elevated mRNAs of only ERBB3 and HRG1 in patients with precan­cerous lesions could also indicate that EGF deregulation appears in the later stages of GC development. We found slightly higher levels of FGF4 in 59% of GC patients but in none of the ex­amined cases for 5- or more times. However, none of the precancerous patients had el­evated FGF4, which is in accordance with previous results.25 This oncogene might be probably deregulated in the later stages of GC, because it was found that it is mainly implicated in angiogenesis and metasta­sis.25 Multiplex RT-PCR analysis using capil­lary electrophoresis and laser-induced fluo­rescence allowed us to quantify a relatively small amounts of mRNAs with high sensi­tivity. This seems important when dealing with tissue samples obtained from surgery and biopsies. Furthermore, we observed a deregulation of cripto and cyclin E in precan­cerous patients, thus indicating a possible use of these markers for an early identifica­tion of pre-malignant lesions. Researchers have already confirmed the possible use of cripto as a serologic marker for breast and colon carcinoma.18 Our method might be Radiol Oncol 2007; 41(4): 174-82. helpful for a rapid and reliable assessment of expression profiles of selected oncogenes and thus could be used as a tool for select­ing appropriate chemotherapy and for the prognosis. Acknowledgements This work was supported by research grant P0-0527-0381 from the Slovenian Ministry for Education, Science and Sports, by ICGEB Cooperative Research Project CRP/ SLO 98-02, and by the Austrian Science Fund (P-14133-PHY), which are gratefully acknowledged. References 1. Ferlay J. Globocan 2000: Cancer incidence, mortality, and prevalence worldwide. Lyon: IARC Press; 2001: 1 CD-ROM. 2. Lauren P. The two histological main types of gastric carcinoma: diffuse and so-called intesti­nal-type carcinoma. An attempt at a histo-clinical classification. Acta Pathol Microbiol Scand 1965; 64: 31-49. 3. Zheng L, Wang L, Ajani J, Xie K. Molecular basis of gastric cancer development and progression. Gastric Cancer 2004; 72: 61-77. 4. Kufe DW, Holland JF, Frei E, Abramson DH, Dang CT, DeAngelis LM, et al. Cancer medicine 6. Hamilton, Ont.: BC Decker; 2003. 5. Barletta C, Scillato F, Sega FM, Mannella E. Genetic alteration in gastrointestinal cancer. A molecular and cytogenetic study. Anticancer Res 1993; 136A: 2325-9. 6. Tahara E. Genetic pathways of two types of gastric cancer. IARC Sci Publ 2004; 157: 327-49. 7. Engelman JA, Zejnullahu K, Mitsudomi T, Song Y, Hyland C, Park JO, et al. MET amplification leads to gefitinib resistance in lung cancer by activating ERBB3 signaling. Science 2007; 316(5827): 1039-43. 8. Smolen GA, Sordella R, Muir B, Mohapatra G, Barmettler A, Archibald H, et al. Amplification of MET may identify a subset of cancers with extreme sensitivity to the selective tyrosine kinase inhibitor PHA-665752. Proc Natl Acad Sci USA 2006; 103(7): 2316-21. 9. Uen YH, Lin SR, Wu CH, Hsieh JS, Lu CY, Yu FJ, et al. Clinical significance of MUC1 and c-Met RT-PCR detection of circulating tumor cells in pa­tients with gastric carcinoma. Clin Chim Acta 2006; 367(1-2): 55-61. 10. Corso S, Comoglio PM, Giordano S. Cancer ther­apy: can the challenge be MET? Trends Mol Med 2005; 11(6): 84-92. 11. Felsher DW. Cancer revoked: oncogenes as thera­peutic targets. Nat Rev Cancer 2003; 3(5): 375-80. 12. Kataoka H, Joh T, Kasugai K, Okayama N, Moriyama A, Asai K, et al. Expression of mRNA for heregulin and its receptor, ErbB-3 and ErbB-4, in human upper gastrointestinal mucosa. Life Sci 1998; 63(7): 553-64. 13. Bachmair F, Huber C, Daxenbichler G. Quanititation of gene expression by means of HPLC analysis of RT-PCR products. Clin Chim Acta 1999; 279(1-2): 25-34. 14. Huber CG. Micropellicular stationary phases for high-performance liquid chromatography of double-stranded DNA. J Chromatogr A 1998; 806: 3-30. 15. Heideman DA, Snijders PJ, Bloemena E, Meijer CJ, Offerhaus GJ, Meuwissen SG, et al. Absence of tpr-met and expression of c-met in human gastric mucosa and carcinoma. J Pathol 2001; 194(4): 428­35. 16. Nakajima M, Sawada H, Yamada Y, Watanabe A, Tatsumi M, Yamashita J, et al. The prognostic sig­nificance of amplification and overexpression of c-met and c-erb B-2 in human gastric carcinomas. Cancer 1999; 85(9): 1894-902. 17. Rasola A, Fassetta M, De Bacco F, D’Alessandro L, Gramaglia D, Di Renzo MF, et al. A positive feedback loop between hepatocyte growth fac­tor receptor and beta-catenin sustains colorectal cancer cell invasive growth. Oncogene 2007; 26(7): 1078-87. 18. Bianco C, Adkins HB, Wechselberger C, Seno M, Normanno N, De Luca A, et al. Cripto-1 activates nodal- and ALK4-dependent and -independent signaling pathways in mammary epithelial Cells. Mol Cell Biol 2002; 22(8): 2586-97. 19. Bianco C, Strizzi L, Mancino M, Rehman A, Hamada S, Watanabe K, et al. Identification of cripto-1 as a novel serologic marker for breast and colon cancer. Clin Cancer Res 2006; 12(17): 5158­64. 20. Hamada S, Watanabe K, Hirota M, Bianco C, Strizzi L, Mancino M, et al. beta-Catenin/TCF/ LEF regulate expression of the short form hu­man Cripto-1. Biochem Biophys Res Commun 2007; 355(1): 240-4. 21. Minchiotti G. Nodal-dependant Cripto signal­ing in ES cells: from stem cells to tumor biology. Oncogene 2005; 24(37): 5668-75. 22. Haruma K, Ito M, Kohmoto K, Kamada T, Kitada Y, Yasui W, et al. Expression of cell cycle regula­tors and growth factor/receptor systems in gas­tric carcinoma in young adults: association with Helicobacter pylori infection. Int J Mol Med 2000; 5(2): 185-90. 23. Akama Y, Yasui W, Yokozaki H, Kuniyasu H, Kitahara K, Ishikawa T, et al. Frequent amplifica­tion of the cyclin E gene in human gastric carcino­mas. Jpn J Cancer Res 1995; 86(7): 617-21. 24. Tenderenda M. A study on the prognostic value of cyclins D1 and E expression levels in resectable gastric cancer and on some correlations between cyclins expression, histoclinical parameters and selected protein products of cell-cycle regulatory genes. J Exp Clin Cancer Res 2005; 24(3): 405-14. 25. Yasui W, Oue N, Aung PP, Matsumura S, Shutoh M, Nakayama H. Molecular-pathological prognos­tic factors of gastric cancer: a review. Gastric Cancer 2005; 8(2): 86-94. 26. Yu J, Miehlke S, Ebert MP, Szokodi D, Wehvnignh B, Malfertheiner P, et al. Expression of cyclin genes in human gastric cancer and in first degree rela­tives. Chin Med J (Engl) 2002; 115(5): 710-5. 27. Breuleux M. Role of heregulin in human cancer. Cell Mol Life Sci 2007; 64(18): 2358-77. 28. Kobayashi M, Iwamatsu A, Shinohara-Kanda A, Ihara S, Fukui Y. Activation of ErbB3-PI3-kinase pathway is correlated with malignant phenotypes of adenocarcinomas. Oncogene 2003; 22(9): 1294­301. 29. Slesak B, Harlozinska A, Porebska I, Bojarowski T, Lapinska J, Rzeszutko M, et al. Expression of epidermal growth factor receptor family proteins (EGFR, c-erbB-2 and c-erbB-3) in gastric cancer and chronic gastritis. Anticancer Res 1998; 18(4A): 2727-32. Radiol Oncol 2007; 41(4): 174-82. Role of the cancer registries in determining cancer mortality in Asia Balkrishna B.Yeole Mumbai Cancer Registry, Indian Cancer Society, Mumbai, India Background. In the absence of dependable data from the Civil Registration System (CRS) many countries have developed their own Sample Registration System (SRS). Under this scheme in India the SRS collects the information on fertility and mortality indicators at state and national levels. Conclusions. In Asia in general and in India in particular cancer registries have played a crucial role in providing the improved cancer mortality data. Key words: cancer registry; cancer mortality; sample registration system Introduction The burden of disease can be assessed through a number of epidemiological pa­rameters such as incidence, prevalence, mortality and disability caused by the dis­ease. The global burden of disease study conducted by WHO, World Bank and Harvard School of Public in 1990s, further developed the sophisticated epidemiologi­cal parameters on mortality, morbidity and disability to provide a composite index on the burden of disease like YLD (Years of Life Lived with Disability) and DALY (composite Index of Burden of Disease). However, data on incidence, prevalence, Received 16 October 2007 Accepted 30 October 2007 Correspondence to: Dr. Balkrishna B.Yeole, M.Sc., Ph.D., Director, Mumbai Cancer Registry, Indian Cancer Society, 74, Jerbai Wadia Road, Parel, Mumbai- 400 012, India; Phone: +91-22-24122351; Fax: +91-22­24122351; E-mail: bcrics@vsnl.com Paper was presented at the 27th Conference of International Association of Cancer Registries, Ljubljana, Slovenia, 17-20 September 2007. and disease specific mortality are frequent­ly incomplete, not very reliable or are lack­ing in many countries particularly in Asia and Africa. In the absence of dependable data from the Civil Registration System (CRS) many countries have developed their own Sample Registration System (SRS). Under this scheme in India the SRS collects the information on fertility and mortality in­dicators at state and national levels. The SRS mechanism involves collections of data through two different procedures. It is performed by continuous enumeration and retrospective half yearly survey by a process of matching two records and sub­sequent field verification of unmatched and partially matched events. The meth­odology provides a cross check on the correctness of events of births and deaths listed in both records.1 In order to estimate the cancer mortality from other than cancer registries there is a paucity of adequate data on the one hand and the complex pathogenesis of disease on the other one which make for complexi­ty, particularly in rural population. In India, major death registration sources are neither reliable nor complete. A large percentage of cases goes unregistered and out of reg­istered cases only, 10% of deaths are medi­cally certified.2 In this contest it is important to mention that quality of death reporting to the system in less developed countries including India is very poor. Due to several socio-econom­ic constraints the cause is not adequately noted in the death certificates. A sample registration system practice in India helps in this but for correlating with cancer regis­try data this is not optimally helpful. When cancer morbidity figures from SRS system and cancer registry are compared the SRS figures are at low levels.2 The procedure adopted by cancer registries in determining cancer mortality There are three main sources of cancer deaths to be collected in the registry: Vital statistics departments of Municipal Corporation, Medical records department of collaborative hospitals and Active fol­low-up through telephone, postal enquiries and house visits. In cancer registries, completeness of reg­istrations, certification of deaths, disease coding practices, cause of death, basic in­formation like address and demographic information and especially the duration of stay and primary cause of death are the main problems to be encountered. There are number of reasons for the un­der registration of cancer deaths in cancer registries. The death certificates are not available at source of information. The death might have occurred out side the area of registration. Deaths may not have been registered at the vital statistics department of Municipal Corporation. If the cancer has long survival and death was not due to can­cer, there is a high probability that the in­formation that the disease had cancer had been totally forgotten. The migration also plays an important role. The death registra­tion system may be defective at the vital sta­tistics department. The cause of deaths may be erroneously reported like old age, etc. Mumbai Cancer Registry was established in 1963 for Mumbai Municipal Corporation area. At present this registry covers about 12 million population having 437.7 sq km area. The death registration system in Mumbai is quite complete i.e. 98.7%.3 Reporting of cause of death as far as cancer is concerned is quite good – non site-specific deaths are less than 1%. At present 9500 incidence cas­es are registered at this registry. From the department of Vital Statistics of Municipal Corporation 6200 cancer deaths are col­lected out of which 5000 are residents, 800 are non residents and 400 are not known residents.4 The method of collection of cancer death information in this registry is that the staff of the registry visits the Municipal Corporation’s office to scrutinize all the deaths and copies information on deaths mentioning cancer of tumour which is the primary or secondary cause of death. Residents’ deaths to incidence cases give M/I Ratio. For Mumbai it is 52.6% which is quite comparable with European and devel­oped countries. The municipal Corporation also pub­lishes the annual report on vital statistics.5 When the comparison is made for the year 2000, the information about the number of cancer deaths has two sources: the Municipal report has reported only 4320 cancer deaths, whereas the registry col­lected 6200 cancer deaths. This implies that the registry has recorded 1880 more cancer deaths (30%) than the published report by the Municipal Corporation. Radiol Oncol 2007; 41(4): 183-7. Reasons for less cancer deaths reporting in the Municipal Corporation report may be due to the fact that they have been looked only at the primary cause of death and over­looked the secondary or underlying cause of death. When the site specific deaths are com­pared in both reports, deaths due to second­ary sites, glands, brain tumours, leukaemia, were very minimally reported in the corpo­ration reports. This may be due to the lack of training of the coder of the vital statistics department. In short, all the deaths record­ed at Vital statistics should be scrutinized by the trained registry staff. In India, other than Mumbai registry, the method applied for few registries for the improving cancer mortality is described below. It is well known that in India except Mumbai the death registration system is quite incomplete and the cause of death re­porting system is not at satisfactory level. When the registry started functioning, the M/I ratios were for Chennai in 1982 23%, for Bangalore in 1982 17%, for Bhopal in 1987 19% and for Delhi in 1987 19%. The Chennai registry was established in 1982 by the cancer institute. At present, it covers the population of 4.3 million having an area of 170 sq km. Having the incomplete reporting of cancer deaths and poor notifi­cation of cause of deaths this registry has improved the problem of under registration by the following way. This registry records all the deaths regardless of the cause of death from the vital statistics department and hospital records. Then, all the deaths are computerized. All this mortality data are matched with morbidity data. Matched deaths are then updated in morbidity data. Unmatched cancer deaths are then traced back by house visits. Cases with no other details are registered as “DCO’s”. By this method the M/I ratio of this registry has been improved from 24% to 54%.6 Indian Council of Medical Research, New Delhi has established the population based registry at Bhopal, in 1986 with the aim to evaluate the cancerogenic effect of Mythel Isocynate and cancer. This registry covers 285 sq km area having the population of 1.4 million. A death registration system is far from adequate resulting in under registra­tion of the cancer mortality. This registry has identified burial grounds and crema­toriums for the death registration system implementing the same methodology as of Chennai PBCR. It has been shown that M/ I ratio which was around 19% initially has gone up to 36%.7 Tata Memorial Hospital, Mumbai, in col­laboration with Indian Council of Medical Research, New Delhi, established the first rural cancer registry at Barshi in Solapur district of Maharashtra in 1987. At present it covers 0.4 million population with an area 3717 sq km. Information on deaths is col­lected from the village death records and also from the local community. As death records are not generally medically certi­fied, the relatives of all diseased are con­tacted to collect the relevant information to assist in “follow-track” to the medical records in the treating hospital or physician to identify the proven cancer cases. In this registry the M/I ratio is 79%.8 Many registries collect the follow-up in­formation for the survival studies. This pro­cedure is also helpful to improve the cancer mortality in registries. Mumbai cancer reg­istry collects the follow-up information for most of the major sites after 5 years for each case. To get the follow-up information, the methodology is used, firstly, to match with the cancer deaths collected from the vital statistics department (50%). The follow-up information from the remaining patients are done by telephone and postal enquiries (15%) and by house visits (10%). Due to this procedure there have been improvements in cancer mortality about 10%.9 Special cross sectional surveys in regis­tration areas are also helpful in improving cancer mortality statistics of the registry. Tata Institute of Fundamental Research, Mumbai has carried out a special health survey for Mumbai City population during 1991-94. In this survey the information on deaths has been also collected when survey data and Mumbai Registry data has been matched for the cancer mortality. It has been observed that there has been an im­provement of 4.2% in cancer mortality. It has been shown that there is an ef­fective use of cancer registries for cancer survivorship research.10 Two hospital can­cer registries in USA were used to recruit a large sample of breast cancer survivors for a study examining the late reproductive ef­fects of breast cancer treatments. These two participative cancer registries were an ex­cellent source of identifying a large sample for long term for breast cancer survivors. Although there are some limitations to this approach including a non response of a sig­nificant number of breast cancer survivors, tumour registries represent an important resource for the rapid identification of can­cer survivors for research studies. Findings from this study suggest several enhance­ments for the future study that may increase the yield from registry recruitments. Cancer mortality through cancer reg­istries in Asia, Africa, may be improved as follows: it is well known that the usual method of mortality data collection as in the west will not give reliable and complete data. It is absolutely necessary that the im­provements in the system of registration of deaths include the implementation of standard core information mortality form in all hospital-nursing homes in registra­tion area and at birth and death registration units of the vital statistics department and at burial grounds and crematoriums. The improvements in the system of certification of the cause of death stress on the underly­ing and antecedent cause of death should be given. The medical personnel should be educated on the method of certifying the cause of death. The cancer registration topic should be introduced in curriculum of the final year MBBS at least one question on this topic in any clinical subject. Verbal au­topsies have to be more rigorous and stand­ardized procedures before the exact cause of death can be ascertained. Conclusions The cancer mortality assess is an impor­tant function of any cancer control pro-gramme. Cancer registries of the sound system are used for evaluating cancer mor­tality. Because of the scientific discipline in cancer registration system, the mortality rates obtained through cancer registry will be optimally productive. In Asia in general and in India in particular cancer registries have played a crucial role in providing the improved cancer mortality data. References 1. Sample Registration System. Statistical Report – 1998. New Delhi, India: Registrar General; 2000. 2. Ramanakumar AV, Yeole BB. Assessing cancer burden in rural India: An Analysis by cause of death statistics. Asian Pac J Cancer Prev 2005; 6: 221-3. 3. Gupta RB, Rao GR. Effect of elimination of differ­ent causes of death on expectation of life-Bombay, 1960-61. Indian J Med Res 1973; 61: 950-61. 4. Kurkure AP, Yeole BB, Sunny L, Koyande SS. Cancer incidence and mortality in Greater Mumbai, 2001. Mumbai, India: Indian Cancer Society: 2005. 5. Municipal Corporation of Greater Mumbai. Annual Report- 2001. Mumbai, India, 2003. 6. Shanta V, Swaminathan R, Cancer incidence and mortality in Chennai, 2001. Chennai, India: Cancer Institute; 2004. Radiol Oncol 2007; 41(4): 183-7. 7. Bharadwaj AK, Shrivastav A, Cancer incidence and mortality in Bhopal. Annual Report 2001. Bhopal, India; 2005. 8. Dinshaw KA, Nene BM. Cancer incidence and mor­tality in Barshi –2001. Annual Report. Barshi, India; 2005. 9. Sankarnarayan R, Black RJ, Parkin DM. Cancer survival in developing countries. IARC Scientific Publication No.145. Lyon, France: IARC. 1998. 10. Pakitil AT, Khan BA, Petersen L, Abraham LS, Greendale GA, Ganj PA. Making effective use of tumour registries in cancer survivorship. Cancer 2001; 92: 1305-14. Retrospective analysis of dose delivery in intra-operative high dose rate brachytherapy Moonseong Oh, Jaiteerth S. Avadhani, Harish K. Malhotra, Barbara Cunningham, Patrick Tripp, Wainwright Jaggernauth, Matthew B. Podgorsak Department of Radiation Medicine, Roswell Park Cancer Institute, Buffalo, New York, USA Background. This study was performed to quantify the inaccuracy in clinical dose delivery due to the incomplete scatter conditions inherent in intra-operative high dose rate (IOHDR) brachytherapy. Methods. Treatment plans of 10 patients previously treated in our facility, which had irregular shapes of treated areas, were used. Treatment geometries reflecting each clinical case were simulated using a phantom assembly with no added build-up on top of the applicator. The treatment planning geometry (full scatter surrounding the applicator) was subsequently simulated for each case by adding bolus on top of the ap­plicator. Results. For geometries representing the clinical IOHDR incomplete scatter environment, measured doses at the 5 mm and 10 mm prescription depths were lower than the corresponding prescribed doses by about 7.7% and 11.1%, respectively. Also, for the two prescription methods, an analysis of the measured dose distributions and their corresponding treatment plans showed average decreases of 1.2 mm and 2.2 mm in depth of prescription dose, respectively. Conclusions. Dosimetric calculations with the assumption of an infinite scatter environment around the applicator and target volume have shown to result in dose delivery errors that significantly decrease the prescription depth for IOHDR treatment. Key words: intraoperative period; brachytherapy; radiotherapy dosage Introduction Intra-operative radiation therapy (IORT) is the delivery of a relatively high dose of radiation to the tumour bed or residual disease at the time of surgical resection. The benefit of this technique is the po- Received 12 October 2007 Accepted 19 October 2007 Correspondence to: Matthew B. Podgorsak, Department of Radiation Medicine, Roswell Park Cancer Institute, Elm & Carlton Streets, Buffalo, NY 14263, USA; Phone: +1 716 845 1536; Fax: +1 716 845 7616; E-mail: moonseong.oh@roswellpark.org tential to shield or displace normal tis­sues thus minimally exposing them to radiation. Clinically, IORT has been used as an adjuvant to surgery and/or fraction­ated external beam radiation therapy for locally advanced cancers of the abdomen, pelvis, head and neck, brain, thorax, and extremities.1-7 Historically, linear accelera­tors employing electron beams were used for IORT.5, 8-10 However, there has been an interest in applying high dose rate (HDR) brachytherapy for this purpose.11-14 In intra-operative high dose rate (IOHDR) brachytherapy, applicators are Table 1. Foreshortening of the depth of prescription dose Patients Intended prescription depth (mm) Prescription dose (cGy) d (Prescription distance from the plan (mm)) d’ (Actual distance (mm)) d-d’ (mm) A 10 1500 9.8 7.8 2.0 B 10 1000 10 7.6 2.4 C 10 500 10 7.7 2.3 D 10 1000 9.9 8.0 1.9 E 10 1250 10 7.9 2.1 F 10 750 10 7.8 2.2 G 10 1250 10 7.8 2.2 H 10 750 10 7.7 2.3 I 5 1250 5 3.8 1.2 J 5 1500 5 3.7 1.3 secured directly to the residual tumour or tumour bed. The region anterior to the ap­plicators is mostly air with significantly less scattering properties than tissue. The dose computation algorithm in commercial treat­ment planning system assumes that the ap­plicators are surrounded by an infinite scat­ter medium.15 This assumption, however, is strictly valid only in cases of interstitial and intracavitary brachytherapy and may lead to an over-estimation of the dose in the case of IOHDR brachytherapy. In a recent publication,16 we have shown that this lack of scatter from one side of the applicator has the potential of leading to significant underdosage during treatments. Our measurements showed that underdos-ages at two planned prescription depths (5 mm and 10 mm) were 8.5% and 12.5% for each of the conventional treatment ge­ometries studied (applicators with surface areas of 4, 7, and 12 cm2). In a clinical envi­ronment, IOHDR brachytherapy treatments typically involve irregular surface areas and there has been concern whether the previ­ous published results with standard irradia­tion geometries can be ported to these clini­cal situations as well. In the present study, we have used an experimental approach to quantify the magnitude of underdosage in clinical cases with irregularly shaped appli­cators. Materials and methods In this retrospective study, the treatment plans of 10 consecutive patients previously treated at our facility were analyzed. Eight patients had a prescription depth of 10 mm where the therapy was delivered out us­ing applicators (Freiburg Flap Applicator, Freiburg, Germany) consisting of a contigu­ous array of 5 mm radius plastic spherical beads, which have a provision to insert multiple nylon catheters separated by 10 mm. The remaining two patients were treated without an applicator, resulting in a prescription depth of 5 mm. In our practice, the prescription depth is defined as the distance between the center of the source dwell positions and the treatment plane. The clinical set up of a representative IOHDR brachytherapy treatment is shown in Figure 1. In this study, the prescribed dose varied from 5 Gy to 15 Gy (Table 1). The treated areas were irregular in shape, covering surface areas of 8 cm2 to 180 cm2 Radiol Oncol 2007; 41(4): 188-95. Figure 1. Clinical setup of a representative IOHDR case. and consisted of 3 to 16 catheters, depend­ing on the size of the target volumes. The clinical treatment plans for each pa­tient were restored to the planning system (Plato, v. 14.2, Nucletron, Columbia, MD) and were renormalized to deliver a dose of 200 cGy to the original prescription depth. The measurement setup is shown in Figure 2. For patients with a 10 mm prescription depth, the treatment delivery was simulated Figure 2. The measurement setup: (a) Full scatter environment, (b) No scatter environment. Radiol Oncol 2007; 41(4): 188-95. by inserting 5 mm bolus material posterior to the applicator to achieve the prescribed distance from the center of the source to a piece of radiographic film. EDR2 films used to get dose profiles were placed on top of a solid water phantom (30 cm x 30 cm x 15 cm) and full scatter conditions were ob­tained by putting 15 cm of bolus material on the top of the applicator. The H & D curve for EDR2 film for Ir­192 was generated using a reference appli­cator having a treatment area of 7 x 7 cm2 and a prescription depth of 10 mm with the prescribed dose varying from 0 to 400 cGy (Figure 3). Fifteen centimeters bolus on top of the applicator was used to simulate the full scattering environment.16 H & D curve films as well as the measurement films were processed in quick succession to re­duce processor dependent uncertainties. A Kodak RP X-OMAT film processor (Model M6B) was used for developing all films. These films were then scanned using a Vidar VXR-16 Dosimetry PROTM film scan­ner (Vidar Systems Corporation, VA) and analyzed using the RIT 113 film Dosimetry system (version 3.14). Results Figure 4 shows the optimized dose distribu­tions in transverse planes for 2 representa­tive patients having a prescription dose of 15 & 7.5 Gy delivered to the prescription depths of 5 and 10 mm, respectively. In the case of incomplete scatter, the prescription dose was actually delivered to a point that was shorter than the intended prescription depth. The magnitude of this shift was found to be a function of the prescription depth and was about 1.3 mm and 2.2 mm for the prescription depths of 5 mm and 10 mm, respectively, as shown in Table 1. Also, Table 1 shows the average distance differences between the measured and planned prescription dose lines. As can be Figure 4. Optimized dose distribution in an axial plane (outer line: prescribed dose line, inner line: actual depth of prescription dose): (a) For the plan with a 5 mm prescription depth of patient J, (b) for the plan with a 10 mm prescription depth of patient F. Table 2. Measured doses in clinical (no added scatter) and treatment planning (full scatter) geometries Delivered Dose (cGy) Prescription Intended Dose Patients Underdosage (%) depth (mm) (cGy) No Scatter Full Scatter A 10 200 174.15 194.47 10.45 B 10 200 178.21 198.31 10.14 C 10 200 180.99 205.6 11.97 D 10 200 173.79 194.06 10.45 E 10 200 174.48 197.49 11.65 F 10 200 172.92 197.44 12.42 G 10 200 175.13 196.32 10.79 H 10 200 172.91 194.51 11.10 Average dose for 8 patients 175.32 197.28 11.13 I 5 200 184.15 199.32 7.61 J 5 200 181.91 197.07 7.69 Average dose for 2 patients 183.03 198.2 7.65 seen from Table 1, the distance between the planned and the measured isodose line is a function of the prescription depth. As expected, the actual doses delivered to the prescription points were 10.5% to 12.4% 102 100 98 96 94 92 90 88 86 Normalized Dose (%) (average 11.1%) lower than prescribed doses for the cases with the prescription depth of 10 mm and were about 7.7% lower with the prescription depth of 5 mm (Table 2 and Figure 5). The magnitude of this under- Figure 5. Comparison of measured doses for both 10 mm and 5 mm prescription depths; the doses are normalized to the corresponding doses with full scattering. Radiol Oncol 2007; 41(4): 188-95. dosage was dependent on the prescription depth and independent of the treatment area. In our earlier paper,16 we had reported on this underdosage for square applica­tor geometries (4 x 4, 7 x 7 & 12 x 12 cm2). Our present work, using the actual patient plans with irregular geometry of applica­tors/source configurations, has also given similar results, thereby, further strengthen­ing our earlier observation that the under-dosage is a function of prescription depth and does not depend on either the shape or the size of the treatment area. This under-dosage was also uniformly spread through­out the treatment area as can be seen from Figure 6 where delivered dose profiles with and without backscatter materials are plot­ted for a representative patient. For this particular patient, the prescription was 10 mm. As shown in Table 2, measured doses with the full scattering were very close to the intended dose of 200 cGy, with aver­ages of 197.3 cGy and 198.2 cGy for 10 mm and 5 mm prescription depths, respectively. Incidentally the measured dose values are within the ±3% uncertainty expected from film dosimetry17, 18 when carried out in a controlled environment. Figure 6. Delivered dose profiles with and without scatter in 10 mm prescription depth for patient B. Discussion In case of IOHDR treatments which usu­ally have a single plane and no crossing at the end of catheters, this results in plans in which the dwell positions at the periphery get higher dwell times than at the center. Thus, this type of inhomo­geneous dwell time distribution results in optimized treatment plans which give more homogeneous dose throughout the desired target area. In case of IOHDR brachytherapy treatments, this target is an area encompassed by the prescription point. In arriving at the optimized plan, the planning system assumes that the applicators are surrounded by infinite scatter material. However, this assump­tion becomes invalid when one side of the applicator is exposed only to air, as is fre­quently the case in IOHDR brachytherapy. Therefore, a significant underdosage due to the lack of scatter would potentially be delivered to the target volume. In most IOHDR treatments, typical ge­ometry does not include full scatter on the opposite side of the applicator from the treatment area. This results in an over-es­timation of the delivered dose in the target volume by the treatment planning system due to the over-simplistic assumption of the full scatter during the dose computa­tion. One can think of correcting for this underdosage by augmenting the scatter en­vironment by placing a scattering medium on the top of the applicators during IOHDR procedure. However, the addition of bo­lus material during the IORT is not always feasible. A real danger is the weight of the added bolus compressing critical structures in and around the target area. A simple ap­proach will be to account for this underdos-age during the treatment planning stage by either prescribing to an appropriate isodose level or by shifting the prescription points deeper into the target. We have demonstrated that without the typical scatter environment, actual dose de­livered can be different from prescription dose. But how this “underdosing” would potentially change clinical outcome is less clear. IOHDR clinical experience has been built on long-established techniques that do not account for the suboptimal scatter environment. While our measurements suggest that typical HDR treatment plan­ning systems underestimate the actual dose delivered by 7.7% at 5 mm and 11.1% at 10 mm, we do not know whether this “under-dosage” would change clinical outcome. Our study has demonstrated that the his­torical data available using electron IORT cannot be compared with IOHDR with­out factoring this underdosage. Peripheral nerve and other normal structure IOHDR recommended tolerance doses have been established by clinical experience. For target prescription dose and for normal structure dose tolerance, we hope that our proposed more accurate dosimetry would be clinically meaningful. But the best test of the benefits of improved dosimetric ac­curacy - an advantage in local control or survival - would require years of follow-up to observe. Conclusions Dosimetry calculations for IOHDR brachy-therapy are typically carried out with treat­ment planning systems that assume an infinite scatter environment around the applicator and the target volume. We have shown this assumption results in signifi­cant shortening of the prescription depth, thereby leading to substantial underdosage to the tumor volume. The underdosage was found to be a function of the prescription depth and was found to be independent of the shape and area of the treated volume. It may be clinically relevant to correct for these errors by augmenting the scatter environment or, preferably, by appropri­ately modifying the prescription dose or by moving the dose prescription points downstream from the catheters during the treatment planning itself. References 1. Palta JR, Biggs PJ, Hazle JD, Huq MS, Dahl RA, Ochran TG, et al. Intraoperative electron beam radiation ther­apy: Technique, dosimetry, and dose specification: Report of Task Force 48 of the radiation therapy com­mittee, American Association of Physicists in Medicine. Int J Radiat Oncol Biol Phys 1995; 33: 725-46. 2. Carvo FA, Micaily B, Brady LW. Intraoperative ra­diotherapy. A positive view. Am J Clin Oncol 1993; 16: 418-23. 3. JohnstonePA,SindelarWF,KinsellaTJ.Experimental and clinical studies of intraoperative radiation thera­py. Curr Probl Cancer 1994; 18: 249-90. 4. Willett CG. Intraoperative radiation therapy. Int J Clin Oncol 2001; 6: 209-14. 5. Konski A, Neisler J, Phibbs G, Bronn D, Dobelbower RR Jr.. The use of intraoperative electron beam radiation therapy in the treatment of para-aortic metastases from gynecologic tumors. Am J Clin Oncol 1993; 16: 67-71. 6. Goldson AL. Past, present, and future prospects of intraoperative radiotherapy (IORT). Semin Oncol 1981; 8: 59-64. 7. Toita T, Nakano M. Takizawa Y. Sueyama H, Kakihana Y, Kushi A, et al. Intraoperative radia­tion therapy (IORT) for head and Neck. Int J Radiat Oncol Biol Phys 1994; 30: 1219-24. 8. Dobelbower RR Jr., Konski AA, Merrick HW 3rd, Bronn DG, Schifeling D, Kamen C. Intraoperative electron beam radiation therapy (IOEBRT) for car­cinoma of the exocrine pancreas. Int J Radiat Oncol Biol Phys 1991; 20: 113-19. 9. McCullough EC, Anderson JA. The dosimetric properties of an applicator system for intraopera­tive electron-beam therapy utilizing a Clinac-18 ac­celerator. Med Phys 1982; 9: 261-68. 10. Gieschen HL, Spiro IJ, Suit HD, Ott MJ, Rattner DW, Ancukiewicz M, et al. Long-term results of intraoperative electron beam radiotherapy for pri­mary and recurrent retroperitoneal soft tissue sar­coma. Int J Radiat Oncol Biol Phys 2001; 50: 127-31. Radiol Oncol 2007; 41(4): 188-95. 11. Nag S, Hu KS. Intraoperative high-dose-rate brach­ytherapy. Surg Oncol Clin N Am 2003; 12: 1079-97. 12. Nag S, Lukas P, Thomas DS, Harison L. Intraoperative high dose rate remote brachyther­apy. In: Nag S, Editor. High dose rate brachy-therapy: A textbook. Armonk, New York: Futura Publishing Company; 1994. p. 427-45. 13. Harrison LB, Minsky BD, Enker WE, Mychalczak B, Guillem J, Paty PB, et al. High dose rate intra-operative radiation therapy (HDR-IORT) as part of the management strategy for locally advanced primary and recurrent rectal cancer. Int J Radiat Oncol Biol Phys 1998; 42: 325-30. 14. Hu KS, Enker WE, Harrison LB. High-dose-rate in-traoperative irradiation: current status and future directions. Semin Radiat Oncol 2002; 12: 62-80. 15. Nath R, Anderson LL, Luxton G, Weaver KA, Williamson JF, Meigooni AS. Dosimetry of inter­stitial brachytherapy sources: Recommendations of the AAPM Radiation Therapy Committee Task Group No. 43. Med Phys 1995; 22: 209-34. 16. Raina S, Avadhani JS, Oh M, Malhotra MK, Jaggernauth W, Kuettel MR, et al. Quantifying IOHDR brachytherapy underdosage resulting from an incomplete scatter environment. Int J Radiat Oncol Biol Phys 2005; 61: 1582-86. 17. McDermott PN, He T, DeYoung A. Dose calcula­tion accuracy of lung planning with a commercial IMRT treatment planning system. J Appl Clin Med Phys 2003; 4: 341-51. 18. Jacqueline E, Sasa M, William BH, Dempsey F, Low DA. Dosimetry of therapeutic photon beams using an extended dose range film. Med Phys 2002; 29: 2438-45. In vivo dosimetry with diodes in rectal cancer patients Andrej Strojnik Institute of Oncology Ljubljana, Department of Radiophysics, Ljubljana, Slovenia Background. Success of radiotherapy relies on accurate dose delivery. In vivo dosimetry improves control of treatment quality. Patients and methods. In vivo dosimetry with commercial diodes was performed in 209 rectal cancer patients treated with four-field box technique. The diodes measured either entrance or exit dose in each treatment field. The results were compared to the planned values and the dose delivered to the isocenter was calculated. Tolerance levels were set to 5% for entrance dose and 8% for exit dose. Results. 421 entrance dose and 415 exit dose measurements were performed. The average difference from expected values was 0.9% for entrance dose (SD 2.1%) and -0.5% for exit dose (SD 3.3%). In 209 patients, the average absorbed dose in the isocenter differed from the planned values by 0.2% (SD 1.4%). Measurement results exceeded the tolerance levels in two patients. Conclusion. Smaller standard deviation of absorbed dose to the isocenter (1.4%), compared to those of entrance (2.1%) and exit dose measurements (3.3%), confirms a correlation between the entrance and exit dose deviations of pairs of opposed fields. The fact that during this study in vivo dosimetry exposed two cases of potentially inaccurate treatments proves its necessity. Key words: rectal neoplasms – radiotherapy; radiotherapy dosage Introduction Effectiveness of radiotherapy greatly de­pends on the accuracy of the absorbed dose to the tumor and the surrounding tissue. Complementary to portal imag­ing which helps verifying the position and shape of the treatment fields, in vivo dosimetry provides dosimetric information Received 9 December 2007 Accepted 22 December 2007 Correspondence to: Andrej Strojnik, Department of Radiophysics, Institute of Oncology Ljubljana, Zaloska cesta 2, SI – 1000 Ljubljana, Slovenia. Phone: +386 1 5879 631; E-mail: astrojnik@onko-i.si regarding actual treatment delivery. It is understandably considered an indispen­sable quality assurance procedure and a safety measure in the treatment process.1-3 This paper describes the calibration of dosimetric diodes and presents the results of in vivo dosimetry in 209 rectal cancer patients.4 Materials and methods Calibration Two dosimetric diodes EDP-20, manufac­tured by Scanditronix, were calibrated against an ionization chamber in a 15 MV photon beam from a Varian 2100CD linear accelerator. In reference conditions, each diode was taped to a plastic water phantom (dimensions: 20 cm x 30 cm x 30 cm) at a distance of 100 cm from the accelerator fo­cus, in the center of an open treatment field measuring 10 cm x 10 cm and with gantry angle set to 0°. The ionization chamber was irradiated with the same treatment pa­rameters at depth dose maximum, 2.5 cm below the phantom surface. In addition to the calibration factor also correction factors accounting for non-reference conditions (different focus surface distances, field sizes, wedged filters and exit dose meas­urement) were determined.5 During each set of measurements all other parameters, apart from the one in question, were kept at reference values. In the case of exit dose correction factor, the gantry was rotated to 180° and the focus surface distance to the near side of the slab was set to 100 cm. The signal dependence on the gantry angle was investigated up to 20°. Throughout the calibration and in vivo measurements, each diode was connected to a dedicated channel on an emX Scanditronix electrometer. The electrometer was connected to a computer running DPD12-pc software al­so provided by Scanditronix Wellhofer. Dark current drift and offset of the assembly were measured and accounted for. In vivo measurements The two diodes were used in routine meas­urements in rectal cancer patients treated with four-field box technique with the iso-center in the center of the planning target volume. With this technique, the beams delivered the dose to the target from four directions, with the gantry angle values of 270°, 0°, 90° and 180°. Such configuration allowed the diode taped to the patient’s skin on the 0° beam’s axis to measure not only the 0° beam’s entrance dose, but also the 180° beam’s exit dose. The same princi­ple applied to the 90° and 270° beams. The clinical routine for each patient was as fol­lows: the patient was set-up in the correct treatment position as established at the CT simulator and the diodes were taped to the patient’s skin at the entrance points of 0° and 90° beams. As the treatment started with the accelerator gantry at 270°, the measurement of the 270° beam’s exit dose was first performed, followed by the meas­urement of the 0° beam’s entrance dose, then of the 90° beam’s entrance dose and concluding with that of the 180° beam’s exit dose. The measurement readings were multiplied by appropriate calibration and correction factors and compared to the values calculated by the planning system. If the difference exceeded the tolerance level of 5% for entrance dose or 8% for exit dose, a thorough investigation of treatment parameters was performed together with a scrupulous review of the treatment plan; in vivo dosimetry was repeated at the next treatment session and focus skin distanc­es were carefully measured to verify the correct placement of the dosimeters with respect to the accelerator’s focus. If the problem persisted, the treating radiation oncologist was consulted: if portal images of the treated area were satisfactory, the number of monitor units of the problematic treatment field was adapted and another session of in vivo dosimetry was required. From the deviations of entrance and exit dose measurements from expected values, the deviation of absorbed dose in the iso-center was estimated for each patient. This was accomplished by averaging the devia­tions of all four fields. The rationale for this was that the beams were weighed equally in the isocenter within a few percent. The proof is as follows: In all patients, in vivo dosimetry was per­formed at the second treatment session (portal imaging at the first session). Measurements in the same patient were repeated only in cases of exceeded tolerances due to primary beam attenuation by the diode build-up cap which increases the skin dose and reduces the dose at greater depths.6 Results Calibration Diodes were calibrated for the clinical dose rates of approximately 3 Gy/min. The sig­nal remained constant throughout the dose rate interval between 1 Gy/min and 6 Gy/min. The diode response linearity was Radiol Oncol 2007; 41(4): 196-202. tested for the clinical doses between 10 cGy and 10 Gy with the results within the meas­urement error well below 1%. The diodes were irradiated in geometric conditions which are commonly encountered in the treatment of patients. These include focus skin distances from 75 cm to 115 cm with the field sizes between 5 cm x 5 cm and 30 cm x 30 cm. The diodes were also irradiated with wedged beams, nominal inclinations of hard wedges being 15°, 30° and 45°. Correction factors are presented in Tables 1,2 and 3. Exit dose measurements were performed with the thicknesses of the plastic water phantom ranging from 20 cm to 35 cm simulating different patient thick­nesses. The exit dose correction factor was found to be almost constant (within 1%) and was established to be 1.10 and 1.12 for the diodes 1 and 2, respectively. The influ­ence of the gantry angles of up to 20° was below 1%. Due to the applied four-field box technique and pelvic area topology, the beams were assumed to be perpendicular to the patient’s skin surface on which the diode was taped. Gantry angle correction factors were therefore omitted. Temperature dependence was not inves­tigated. It is practically impossible to moni­tor the temperature of the detector during the treatment of patients and apply an ad­equate correction (it takes several minutes for a diode, after being taped on the patient, to reach thermal equilibrium with the skin7). According to some guidelines2, the influence of temperature may be neglected. It was estimated that during the time of use the diodes absorbed approximately 200 Gy. No sensitivity degradation was ob­served. In vivo measurements In vivo dosimetry was conducted in 209 pa­tients. In 6 (3%) out of 209 patients, in vivo measurements exceeded the tolerances. In 2 Table 1. Focus skin distance correction factors Focus Skin Distance Correction factor (cm) 75 0.94 * and 0.95 ** 80 0.96 85 0.98 90 0.99 95 0.99 100 1 105 1.01 110 1.01 115 1.02 * Diode 1 and ** Diode 2 Table 2. Field size correction factors Field Size (cm x cm) Correction factor 5 x 5 – 20 x 20 1 25 x 25 – 30 x 30 1.01 Table 3. Wedge correction factors Wedge (°) Correction factor 15 1.01 30 1.02 45 1.02 (33%) of the 6 even repeated measurement results were beyond acceptable levels. In the first of the above two patients, a closer inspection revealed a false CT image set had been assigned to the patient. A new therapy plan was later created with the correct CT image set. In the second of the above two patients the source of error proved to be a set of incomplete CT images: due to the size of the patient the outmost parts of the patient’s hips had not been captured by the CT scanner. To rectify the problem, the focus skin distances in the lateral fields were measured and the number of monitor units was adapted. After the corrections, in vivo dosimetry was repeated and the results were within the tolerance levels in both cases. Excluding 8 treatment sessions with the measurement results outside the accept- Difference between measured and planned EXIT dose Figure 2. Frequencies of differences between measured and expected exit doses. In 415 measurements, the average difference was -0.5% with the standard deviation of 3.3%. 4 values between -10% and -8% were excluded from the chart. Repeated measurement results were inside the acceptable limits. Radiol Oncol 2007; 41(4): 196-202. Difference between delivered and planned dose IN ISOCENTER Figure 3. Frequencies of differences between delivered and expected doses in isocenter. In 209 patients, the average difference was 0.2% with the standard deviation of 1.4%. able intervals, 421 entrance dose and 415 exit dose measurements were carried out (3 exit dose measurements were overlooked by mistake and replaced by entrance dose measurements in the next treatment ses­sion). The average deviation from expected values was 0.9% for entrance dose (SD 2.1%) and -0.5% for exit dose (SD 3.3%). In 209 patients, the average absorbed dose in the isocenter differed from the planned values by 0.2% (SD 1.4%). Difference distributions are presented in Figures 1,2 and 3. Discussion The wider spread of exit dose deviations (SD 3.3%) in comparison to the entrance dose deviations (SD 2.1%) was suspected to be due to various bowel fillings of patients; however, no further investigation was con­ducted. The lesser standard deviation of the differences between actual and ex­pected dose in isocenter (1.4%) confirmed a correlation between the entrance and exit dose deviations of any pair of opposed beams: if a diode was closer to the accelera­tor focus than expected when measuring the entrance dose, it was also farther than expected when measuring the exit dose – and vice versa. In the group of 209 patients, in vivo dosimetry revealed and prevented two cas­es of inaccurate treatment. In both cases, the cause of error was traced to geometrical inconsistencies related to incorrect CT data used by the planning system. Both prob­lems occurred as a consequence of human errors and no equipment malfunction was discovered. Such discrepancies could have also been detected by the optical distance indicator! Therefore a quick focus skin dis­tance check could prove a valuable quality assurance procedure in the departments that have not yet been outfitted with in vivo dosimetric equipment. Acknowledgment The study was done partly within a nation­al program Development and Evaluation of New Approaches to Cancer Treatment P3­0003 financially supported by the Slovenian Research Agency (ARRS). References 1. Van Dam J, Marinello G. Methods for in vivo dosim­etry in external radiotherapy. ESTRO Booklet No. 1, Leuven: Garant, 1994. 2. Huyskens DP, Bogaerts R, Verstraete J, Lf M, Nystr H, Fiorino C, et al. Practical guidelines for the implementation of in vivo dosimetry with diodes in external radiotherapy with photon beams (entrance dose). ESTRO Booklet No. 5, Leuven: Garant, 2001. 3. AAPM Report No. 87: Diode in vivo dosimetry for patients receiving external beam radiation therapy. Medical Physics Publishing, 2005. 4. Rikner G, Grusell E. General specifications for silicon semiconductors for use in radiation dosimetry. Phys Med Biol 1987; 32: 1109-17. 5. Leunens G, Van Dam J, Dutreix A, van der Schueren E. Quality assurance in radiotherapy by in vivo do­simetry. 1. Entrance dose measurements, a reliable procedure. Radiother Oncol 1990; 17: 141-51. 6. Nilsson B, Ruden BI, Sorcini B. Characteristics of silicon diodes as patient dosemeters in external radia­tion therapy. Radiother Oncol 1988; 11: 279-88. 7. Grusell E, Rikner G. Evaluation of temperature ef­fects in p-type silicon detectors. Phys Med Biol 1986; 31: 527-34. Radiol Oncol 2007; 41(4): 196-202. Radio/ Oncol 2007; 41(4): 161-5. Tamponada srca kot prvi znak adenokarcinoma pljuc Prikaz primera in pregled literature Letonja M, Debeljak A Izhodišca. Maligna obolenja lahko povzrocijo pericarditis, ki se kaže kot akutni perikarditis, perikardialni izliv, konstriktivni pericarditis z izlivom ali kot tamponada srca. Pri vecini bolnikov z malignim obolenjem in perikarditisom pa v casu bolezni nikoli ne najdemo kli­nicnih znakov perikarditisa. Prikaz primera. 69-letna bolnica je bila sprejeta na intenzivni oddelek zaradi akutno nastale dispneje in tahikardije, ki so ju spremljale razširjene jugularne vene, hepatomegalija in de­snostranski plevralni izliv. Radiogram prsnega koša je potrdil desnostranski plevralni izliv in razkril povecano srcno senco. Z ehokardiografijo smo potrdili diagnozo tamponade srca. Pri bolnici smo opravili terapevtsko perikardiocentezo, po kateri so klinicni simptomi in znaki tamponade srca izzveneli. S citološko analizo perikardialnega punktata smo ugotovili metastatski adenokarcinom. CT toraksa je prikazal desnostranski plevralni izliv in solitarno okroglo lezijo v desnem spodnjem režnju pljuc. Bronhoskopija z biopsijo in krtacenjem je odkrila adenokarcinom pljuc. Zakljucki. Tamponada srca je redka kot prvi znak maligne bolezni, prav tako je izjemen maligen perikardialni izliv kot posledica adenokarcinoma pljuc pri nekadilkah. Opisana bolnica je imela zelo dolgo preživetje po odkritju bolezni, saj je pricakovana življenjska doba bolnikov z malignim perikarditisom ne glede na zdravljenje kratka. Slovenian abstracts Radio/ Oncol 2007; 41(4): 167-73. Vloga biopsije varovalne bezgavke pri raku vulve. Izkušnje na Onkološkem inštitutu Ljubljana Vakselj A, Bebar S Izhodišca. Biopsija varovalne bezgavke (SNB) je obetajoc diagnosticni poseg za ugotovitev stadija bolezni in mocno zmanjša postoperativno obolevnost. Trenutno še ni sprejeta kot standardni nacin zdravljenja. Bolnice in metode. Od marca 2003 do konca leta 2006 smo na Onkološkem inštitutu Ljubljana naredili pri 35 bolnicah z rakom vulve SNB pred operacijo. Srednja starost bolnic je bila 65,8 let (36-88). 32 bolnic je imelo skvamoznocelicni karcinom, ena maligni melanom, ena bazalnocelicni karcinom in ena adeno-skvamozni karcinom. SNB je bila narejena z izotopom 99mTc, ki smo ga uporabili za staticno in dinamicno limfoscintigrafijo ter z intra­dermalnim injiciranjem metilenskega modrila. Rezultati. Pri 25 bolnicah nismo našli metastaz (71,4%). Pri 3 od 25 bolnic je prišlo do ponovitve bolezni ingvinalno (po 12, 24 in 26 mesecih). Pri prvi bolnici je bil vzrok verjetno pomanjkanje izkušenj, pri drugi nam ni uspelo identificirati druge aktivne bezgavke, pri tretji pa se le ta ni prikazala zaradi predhodne operacije v ingvinalnem predelu (brazgotina). Od SNB negativnih je ob koncu leta 2007 živih 22 bolnic brez znakov bolezni, ena je umrla po 45 mesecih zaradi recidiva v ingvinalnem predelu, ena zaradi drugih vzrokov, bolnica z malignim melanomom je živa z jetrnimi metastazami. O eni bolnici ni novih podatkov. Zakljucki. Da bi potrdili vrednost nove diagnosticne metode, predvsem njeno ucinkovitost pri nacrtovanju zdravljenja, potrebujemo multicentricne randomizirane raziskave pri vecjemu številu bolnic. Radio/ Oncol 2007; 41(4): I-VI. Radio/ Oncol 2007; 41(4): 174-82. Dolocanje diferencialnega izražanja onkogenov pri želodcnih adenokarcinomih z metodo socasnega fluorescentnega oznacevanja in pomnoževanja genskih prepisov Rajcevic U, Hudler P, Mijovski G, Gorjanc G, Hoelzl G, Repše S, Juvan R, Kovac M, Huber CG in Komel R Izhodišca. Rak želodca sodi med obolenja, za katera je znacilna slaba prognoza in nizko preživetje. Ceprav je o mehanizmih njegovega nastanka zelo malo znanega, je ocitno, da so v njegov razvoj vpleteni genetska nestabilnost, inaktivacija tumorje zaviralnih genov in prekomerno izražanje onkogenov. Namen raziskave je bil dolociti diferencialno izražanje sedmih onkogenov: CCNEl, EGF, ERBB3, FGF4, HRGl, HGFR and TDGFl. Bolniki in metode. Uporabili smo metodo hkratnega obratnega prepisovanja in pomnoževanja s polimerazo (angl. Reverse Transcription Polymerase Chain Reaction, RT-PCR). Uporabljali smo fluorescentna barvila, ki so nam omogocila enostavno detekcijo produktov. Rezultati. Vec kot polovica bolnikov (74, %) od 74-ih z adenokarcinomom želodca je pre­komerno izražalo vsaj enega od onkogenov, 56,% pa jih je prekomerno izražalo dva ali vec onkogenov. Povišano izražanje gena FGF4 smo zaznali pri manj kot tretjini bolnikov. Zakljucki. Pri bolnikih s prekanceroznimi lezijami smo zaznali prekomerno izražanje genov TDGFl ali cripto-1 (64,%) in CCNEl (57 ,%). Ta gena bi lahko bila uporabna kot oznacevalca za zgodnjo diagnostiko malignih sprememb v želodcu. Metoda fluorescentnega hkratnega RT-PCR pa se je izkazala kot zelo uporabna za hitro dolocanje kolicine mRNA v majhnih vzorcih tumorskega tkiva in prekanceroz. Radio/ Oncol 2007; 41(4): 183-7. Vloga registrov raka pri ugotavljanju smrtnosti zaradi raka v Aziji Yeole BB Izhodišca. Zaradi odsotnosti zanesljivih podatkov o smrtnosti, ki jih pridobiva država, so v mnogih deželah razvili svoj nacin zajemanja podatkov (Sample Registration System -SRS). Tako v Indiji s SRS zbirajo podatke o rodnosti in smrtnosti v posamezni zvezni državi pa tudi na državnem nivoju. Zakljucki. V Aziji, zlasti v Indiji imajo registri raka kljucno vlogo za izboljšanje zajemanja podatkov o smrtnosti zaradi raka. Radio/ Oncol 2007; 41(4): 1-VI. Radio/ Oncol 2007; 41(4): 188-95. Retrospektivna analiza klinicne dozimetrije pri intraoperativni brahiterapiji z visoko hitrostjo sevalne doze Oh M, Avadhani JS, Malhotra HK, Cunningham B, Tripp P, J aggernauth W, Podgorsak MB Izhodišca. Z raziskavo smo želeli kolicinsko ovrednotiti odstopanja v klinicni dozimetriji zaradi omejenega sipanja fotonov, ki so prisotna v intraoperativni brahiterapiji z visoko hitrostjo sevalne doze. Metode. Uporabili smo 10 obsevalnih nacrtov bolnikov, ki smo jih zdravili na naši ustanovi. Vsi so imeli nepravilne oblike obsevalnih podrocij. Obsevalne geometrije vsakega klinicne­ga primera so bile simulirane s fantomom brez bolusa nad vstavljenimi aplikatorji. Z dodat­kom bolusa nad aplikatorje je bila predstavljena tudi možnost terapevtskega nacrtovanja obsevanja z upoštevanjem polnega sipanja fotonov. Rezultati. V merilnih primerih, ki so predstavljali nepopolno sipanje fotonov, so bile absor­birane doze na predpisanih globinah 5mm in 10mm v tkivu nižje za 7,7% in 11,1 %. V obeh primerih je analiza porazdelitve doz iz nacrtov obsevanja pokazala zmanjšanje dosega doze v globino fantoma za 1,2 mm in 2,2 mm. Zakljucki. Dozimetricni izracuni, ki upoštevajo neskoncno sipalno obmocje okoli aplika­torja in tarcnega volumna, kažejo na dozimetricno napako, ki pomeni znatno zmanjšanje pricakovane predpisane globine obsevanja v IOHDR terapiji. 5/ovenian abstracts Radio/ Onco/ 2007; 41(4): 196-202. In vivo dozimetrija z diodami pri bolnikih s karcinomom rektuma Strojnik A Izhodišca. Ucinkovitost radioterapije je odvisna od natancnosti obsevanja. Z dozimetrijo na bol­niku zagotovimo dodaten nadzor nad kvaliteto obsevanja. Bolniki in metode. Pri 209 bolnikih s karcinomom rektuma, ki so se zdravili z obsevalno tehniko štirih polj, smo opravili in vivo dozimetrijo z dozimetricnima diodama. Diodi sta v slehernem obsevalnem polju merili bodisi vstopno bodisi izstopno dozo. Rezultate smo primerjali z nacrto­vanimi vrednostmi in izracunali absorbirano dozo v izocentru. Dopustili smo odstopanja do 5% pri vstopni in do 8% pri izstopni dozi. Rezultati. Izvedli smo 421 meritev vstopne in 415 meritev izstopne doze. Povprecno odstopanje od pricakovanih vrednosti je bilo 0,9% pri vstopni dozi (SD 2, 1 % ) in -0,5% pri izstopni dozi (SD 3,3%). Pri 209 bolnikih se je doza v izocentru v povprecju razlikovala za 0,2% (SD 1,4%) od nacrtovane. Meritve so prekoracile dopustna odstopanja pri dveh bolnikih. Zakljucki. Dejstvo, da smo v okviru te raziskave odkrili in preprecili dve netocni zdravljenji, ponovno dokazuje nujnost in vivo dozimetrije. Manjši standardni odklon absorbirane doze v izocentru (1,4%) od standardnih odklonov vstopne (2,1 %) in izstopne doze (3,3%) potrjuje kore­lacijo med odstopanji vstopne in izstopne doze pri parih opozitnih polj. Radio/ Onco/ 2007; 41(4): I-VI. Notices Notices submitted far publication slwuld contain a mailing address, phone and/ or fax number and/or e-mail oj a Contact person or department. Oncology February 21-23, 2008 The "8th Targeted Therapy Meeting" will take place in Santa Monica, CA, USA-See http://www.iaslc.org Oncology March 5-9, 2008 The NCCN 13th Annual Conference "Clinical Practice Guidelines & Quality Cancer Care™" will be offered in Hollywood, Florida, USA See http://www.nccn.org Lung cancer April 23-26, 2008 The first lung cancer conference in Europe will be held in Geneva, Switzerland. E-mail pia.hirsch@uchsc.edu Lungcancer April 24-26, 2008 The "IASLC/ESMO Lung Cancer Conference" will be offered in Geneva, Switzerland. See http://www.iask.org Lungcancer June 12-14, 2008 The "11 th Central European Lung Cancer Conference" will be offered in Ljubljana, Slovenia. Contact Conference secretariat, Ms. Ksenia Potocnik, Department of Thoracic Surgery, Medica! Centre Ljubljana, Slovenia; or call +386 1 522 2485; or fax +386 1 522 3968; or e-mail ksenia.potocnik @kclj.si; or see http://www.ce-lung2008.org/ Oncology October 9-10, 2008 The "3rd Latin American Cancer Conference" will take place in Vina del Mar, Chile. E mail nisehy@uol.com.br or rodrigo.arriagada@ki.se Oncology November 13-15, 2008 The Chicago/IASLC/ ASCO/ ASTRO symposiums "Malignancies of the Chest and Head and Neck" will be offered in Chicago. E-mail: evokes@medicine. bsd. uchicago.edu Lung cancer 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.iask.orgiumages/ 12worldconfannounce. pdf Oncology September 4-8, 2009 The "34th ESMO Congress" will take place in Vienna, Austria. Contact ESMO Head Office, Congress Department, Via La Santa 7, CH-6962 Viganello-Lugano, Switzerland; or +41 (0)91 973 19 19; or fax +41 (0)91 973 19 18; or e­mail congress@esmo.org; or see http://www.esmo.org 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. vm Authors Index 2007 Austin L: 2/72-79 Avadhani J: 4/188-95 Bebar S: 4/167-73 Belaj N: 1/48-55 Belusic-Gobic M: 2/57-62 Berger D: 3/133-43 Carlone M: 2/90-98 Cerovic R: 2/57-62 Cesar R: 2/80-85 Cunningham B: 4/188-95 Cufer T: 3/115-122 Debeljak A: 4/161-5 Debevec L: 2/80-85; 3/144-51 Delic U: 1/1-12 Faj Z: 1/48-55; 1/48-55 Fallone BG: 1/41-7; 2/90-98 Filipic M: 1/15-22 Fromm S: 3/133-43 Garaj-Vrhovac V: 3/123-32 Gobic D: 2/57-62 Golubovic V: 2/57-62 Gorjanc G: 4/174-82 Hoelzl G: 4/174-82 Horvat-Žnidaršic I: 1/15-22 Huber C: 4/174-82 Hudler P: 4/174-82 Jaggernauth W: 4/188-95 Jamar B: 2/86-89 Jancar B: 1/13-14; 2/63-63; 3/113-4; 4/166-166 Jezeršek Novakovic B: 1/23-32 Juretic M: 2/57-62 Juvan R: 4/174-82 Južnic Šetina T: 1/23-32 Kasabašic M: 1/48-55 Kern I: 2/80-85 Klancevic M: 1/1-12 Kocijancic I: 1/33-8; 3/99-106; 3/99-106 Kolaric Z: 3/107-12 Komel R: 4/174-82 Konstadinova-Kunovska S: 3/152-60 Kotnik V: 1/23-32 Kovac M: 4/174-82 Kukuljan M: 2/57-62 Letonja M: 4/161-5 Lincender L: 1/1-12 Magaš Z: 3/107-12 Malhotra H: 4/188-95 Markov K: 1/41-7 Marotti M: 3/107-12 Matos E: 3/115-122 Mijovski G: 4/174-82 Miller R: 2/72-79 Milutinovic A: 1/15-22 Nikšic G: 3/123-32 Novakovic S: 1/23-32 Ocvirk J: 1/39-40 Oh M: 4/188-95 Pal A: 3/107-12 Pavcec Z: 3/107-12 Pinn M: 2/72-79 Pirker R: 3/133-43 Podgorsak M: 4/188-95 Pokrajac B: 3/133-43 Potter R: 3/133-43 Rajcevic U: 4/174-82 Rajer M: 2/64-71 Repše S: 4/174-82 Rottenfusser A: 3/133-43 Rozman I: 3/144-51 Radio/ Oncol 2007; 41(4): VIII-XI. Saghir H: 3/107-12 Samardziski M: 3/152-60 Schinkel C: 1/41-7; 2/90-98 Schomas D: 2/72-79 Sedmak B: 1/15-22 Sefic-Pašic I: 1/1-12 Stavrev P: 1/41-7; 2/90-98 Stavreva N: 1/41-7; 2/90-98 Strojnik A: 4/196-202 Šavli M: 2/86-89 Šeruga B: 1/39-40 Šuput D: 1/15-22 Tolevska C: 3/152-60 Tomaš I: 1/ 48-55 Tripp P: 4/188-95 Vakselj A: 4/167-73 Vasileva V: 3/152-60 Vegar-Zubovic S: 1/1-12 Vovk M: 1/23-32 Vrcic D: 1/1-12 Vrhovac I: 3/123-32 Yeole B: 4/183-7 Zafiroski G: 3/152-60 Žegura B: 1/15-22 Žokalj I: 3/107-12 Supplement 1/2007 Bartenjev I: S18-S21 Bracko M: S13-S17 Cemažar M: S37-S42 Hocevar M: S22-S27 Kranjc S:S37-S42 Ocvirk J: G-G; S37-S42; S43-S45 Paulin Košir S: S37-S42 Primic Žakelj M: Sl-S12 Reberšek M: S46-S52 Rudolf Z: S37-S42 Serša G: S37-S42 Slekovec-Kolar B: S37-S42 Snoj M: G-G; S37-S42; S53-S55 Strojan P: G-G; S28-S36 Zadnik V: Sl-S12 Žagar T: Sl-S12 Subject Index 2007 adenocarcinoma: 4/161-5; 4/174-82 adenocarcinoma -radiography: 2/86-89 adult: 3/107-12 antibodies, monoclonal: 1/23-32; 3/115-122 antigens, CD 20: 1/23-32 bacterial toxins: 1/15-22 brachytherapy: 4/188-95 breast neoplasms -therapy: 3/115-122 cancer mortality: 4/183-7 cancer registry: 4/183-7 carcinoma, non -small -cell lung ­ radiotherapy: 3/133-43 carcinoma, renal cell -epidemiology ­ diagnosis -therapy: 2/64-71 cardiac tamponade: 4/161-5 catheter ablation: 1/33-8 colonic neoplasms-diagnosis, tomography, X-ray computed: 1/1-12 comet assay: 1/15-22 computer -assisted: 3/133-43 cyanobacteria: 1/15-22 DNA damage: 1/15-22 dose -response relationship, radiation: 1/41-7 dose-response relationship, models, statistical, TCP: 2/90-98 empyema, pleural: 2/57-62 ethics, medical: 2/80-85 IFN alfa: supl/S43-S45 ileocecal valve: 3/107-12 intestinal obstruction: 2/86-89; 3/107-12 intraoperative period: 4/188-95 intussusception: 3/107-12 jejunal neoplasms: 2/86-89 lung neoplasms: 4/161-5 lung neoplasms -pathology, biopsy needle: 3/99-106 Radio/ Oncol 2007; 41(4): VIII-XI. lung neoplasms -surgery: 1/33-8; 3/144-51 lymphatic metastasis -radionuclide imaging: 4/167-73 lymphoma, B-cell -therapy: 1/23-32 magnetic resonance imaging: 1/1-12 mediastinitis: 2/57-62 microsatellite repeats: 3/123-32 neoplasms metastasis: 1/39-40 neoplasms, multiple primary: 2/80-85 nephrectomy: 2/64-71 NTCP, iso-effect, DVH: 1/41-7 oncogenes: 4/174-82 osteosarcoma -diagnosis -therapy: 3/152-60 pancreatic neoplasms: 1/39-40 pelvic neoplasms -radiotherapy: 1/48-55 periapical abscess -complications: 2/57-62 pericardial effusion: 4/161-5 periosteum: 3/152-60 pneumothorax: 3/99-106 Poisson distribution: 2/90-98 prognosis: 2/80-85 quality of life: 3/144-51 radiation effects: 3/123-32 radiation ionizing: 3/123-32 radiotherapy dosage: 2/90-98; 4/188-95; 4/196-202 radiotherapy dosage -methods: 1/48-55 radiotherapy planning: 3/133-43 radiotherapy, conformal: 3/133-43 rats, inbred F344: 1/15-22 receptor, ERB-2 -antagonists and inhibitors: 3/115-122 rectal neoplasms -radiotherapy: 4/196-202 reverse transcriptase chain reaction: 4/174-82 sample registration system: 4/183-7 sentinel lymph node biopsy: 4/167-73 stomach neoplasms -genetics: 4/174-82 survival analysis: 2/64-71 testicular neoplasms -secondary: 1/39-40 thoracotomy: 3/144-51 vulvar neoplasms: 4/167-73 vulvar neoplasms -radiotherapy -drug therapy -surgery -Bartholin gland: 2/72-79 Supplement 1/2007 dakarbazin: S46-S52 dejavniki tveganja: S1-S12 dermoskopija: S18-S21 diagnostika: S53-S55 dopolnilno zdravljenje: S43-S45 elektrokemoterapija: S37-S42 epidemiologija: S1-S12 incidenca: S1-S12 indikacije: S22-S27; S28-S36; S37-S42; S43-S45 kirurgija: S22-S27 maligni melanom: S1-S12; S13-S17 melanom: S18-S21; S22-S27; S28-S36; S37­S42; S43-S45; S46-S52; S53-S55 patologija: S13-S17 preventiva: S18-S21 radioterapija: S28-S36 sistemsko zdravljenje: S46-S52 stranski ucinki: S22-S27; S28-S36; S43-S45 teledermoskopija: S18-S21 umrljivost: S1-S12 zdravljenje: S53-S55 zgodnje odkrivanje: S18-S21 FUNDACIJA "DOCENT DR. J. CHOLEWA" JE NEPROFITNO, NEINSTITUCIONALNO IN NESTRANKARSKO ZDRUŽENJE POSAMEZNIKOV, USTANOV IN ORGANIZACIJ, KI ŽELIJO MATERIALNO SPODBUJATI IN POGLABLJATI RAZISKOVALNO DEJAVNOST V ONKOLOGIJI. DUNAJSKA 106 1000 LJUBLJANA ŽR: 02033-0017879431 Activity of "Dr. J. Cholewa" Foundation for Cancer Research and Education ­a report for the final quarter of 2007 The Dr. J. Cholewa Foundation for Cancer Research and Education continues to focus its activities and attention to cancer research and education in Slovenia and continues to deal carefully and with professional vigour with the requests and proposals for research grants and scholarships. The Foundation members with clinical and research experience in cancer and members with important experi­ence in finance will continue to be instrumental in this activity. The Dr. J. Cholewa Foundation for Cancer Research and Education continues to support the regular publication of "Radiology and Oncology" international medical scientific journal in 2007. This journal is edited, published and printed in Ljubljana, Slovenia. The support for "Radiology and Oncology" emphasizes and addresses the need for the spread of information and knowledge about advances in cancer among professionals and too many interested individuals in lay public and others in Slovenia and elsewhere. "Radiology and Oncology" is now an open access journal, available on its own website, thus allowing its users and readers to freely access, use and re -use it. The Foundation continues to attach weight to the support of the publication of the results from cancer research in Slovenia in respectable international scientific journal worldwide. Tomaž Benulic, MD Borut Štabuc, MD, PhD Andrej Plesnicar, MD Sanolabor 5th Conference on Experimental and Translational Oncology Kranjska gora, Slovenia, March, 26-30, 2008 Organized by: Janko Kos, Tamara Lah and Gregor Serša Topics: ¦ Carcinogenesis ¦ Mechanisms of Tumour Progression ¦ Stem Cells in Cancer ¦ Tumour Markers ¦ Delivery Systems in Cancer Therapy ¦ New Drugs and Therapeutic Markers Location: Hotel Kompas Kranjska gora, Slovenia http://www.hoteli-kompas.si Correspondence: Phone: +386 1 241 29 70 Fax: +386 1 241 29 80 Email: ceto@nib.si http://www.onko-i.si/ceto Organized under patronage of Association of Radiology and Oncology ••:• k o t t e r m a n n . • 1 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, transfuzijo, patologijo in sodno medicino CORNING Corning (Amerika): specialna laboratorijska plastika za aplikacijo v imunologiji, mikrobiologiji, virologiji, ipd., mehanske eno-in veckanalne pipete in nastavki MICRON!C Micronic (Nizozemska): sistemi za shranjevanje vzorcev, pipete, nastavki za pipete Tmvkudech· Ihue's No Rrason to Opuate with Anyone E1se lmplantech (Amerika): obrazni in glutealni vsadki Biomerica (Amerika): hitri testi za diagnostiko, EIA /RIA testi LA&ORMED d.o.u. 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, mano-in poliklonalna protitelesa rB Sakura finetek (Evropa): aparati za pripravo histoloških preparatov: mikro­inkriotomi, zalivalci, tkivni procesorji, barvalci, pokrivalci IBS INTEGRA WWi11'1if1t? lntegra 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 lnfo@lobol'med.sl Tel.: (0)143649 01 Fax: (0)1 436 49 05 SIEMENS SiemensMed,cal.com oncology SEEK-FIND-ACT-FOLLOW -the Continuum of Oncology Care™ Siemens oncology portfoho comprises comprehens,ve max1m1zed ut1hzat1on po1ent1al, and patient-friendly des1gn workflow solutions integrating the full spectrum of care and features from screen,ng/early detection and d1agnos1s through therapy and follow-up Ali from one prov,der -w1th over Every day 1n the United States alone, 29,000 cancer 100 years h1story of 1nnovat1on 1n medica! technology pa11ents rece1ve rad1a11on therapy del1vered by Siemens l1near accelerators As clin1cal protocols trans1t1on to Siemens proven chnical methods can help you to ach,eve include IMRT and IGRT. Siemens seamlessly 1n egrates more successful outcomes How 7 Through industry­the d,agnosilc and treatment modaht,es That's what we leading technology, 1ncreased produc11v1ty measures for call Best Practice Oncology Care ® rimidex anastrozol Kratka Informacija o zdravilu Ime zdravila Arimidex 1 mg filmsko obložene tablete Sestava Ena tableta vsebuje 1 mg anastrozola. Indikacije Adjuvantno zdravljenje žensk po menopavzi, ki imajo zgodnji invazivni rak dojke s pozitivnimi estrogenskimi receptorji. Zdravljenje napredo­valega raka dojke pri ženskah po menopavzi. Ucinkovitost pri bolnicah z negativnimi estrogenskimi receptorji ni bila dokazana razen pri tistih, ki so imele predhodno pozitiven klinicni odgovor na tamoksifen. Odmerjanje in nacin uporabe 1 tableta po 1 mg peroralno, enkrat na dan. Pri zgodnjem raku je priporocljivo trajanje zdravljenja 5 let. Kontraindikacije Arimidex je kontraindiciran pri: ženskah pred menopavzo, nosecnicah in dojecih materah, bolnicah s hujšo ledvicno odpovedjo (ocistek kreatinina manj kot 20 ml/min (oziroma 0,33 ml/s)), bolnicah z zmernim do hudim jetrnim obolenjem, bolnicah, ki imajo znano preobcutljivost za anastrozol ali za katerokoli drugo sestavino zdravila. Posebna opozorila In previdnostni ukrepi Menopavzo je potrebno biokemicno dolociti pri vseh bolnicah, kjer obstaja dvom o hormonskem statusu. Ni podatkov o varni uporabi Arimidexa pri bolnicah z zmerno ali hudo jetrno okvaro ali hujšo ledvicno odpovedjo (ocistek kreatinina manj kakor20 ml/min (oziroma 0,33 ml/s)). Pri ženskah z osteoporozo ali pri ženskah s povecanim tveganjem za razvoj osteoporoze je treba dolociti njihovo mineralno gostoto kosti z denzitometrijo, na primer s slikanjem DEXA na zacetku zdravljenja, pozneje pa v rednih intervalih. Po potrebi je treba zaceti z zdravljenjem ali preprecevanjem osteoporoze in to skrbno nadzorovati. Povzetek glavnih neželenih ucinkov Zelo pogosti (2: 10 %): navali vrocine, obicajno blagi do zmerni Pogosti (2: 1 % in < 10 %): astenija, bolecine / okorelost v sklepih, suhost vagine, razredcenje las, izpušcaji, slabost, diareja, glavobol (vsi obicajno blagi do zmerni) Arimidex znižuje nivo estrogena v obtoku, zato lahko povzroci zmanjšanje mineralne kostne gostote, kar pomeni za nekatere bolnike zvecano tveganje za zlome. Medsebojno delovanje z drugimi zdravili Zdravila, ki vsebujejo estrogen, ne smete dajati socasno z Arimidexom, ker bi se njegovo farmakološko delovanje iznicilo. Tamoksifena se ne sme uporabljati skupaj z Arimidexom, ker lahko pride do zmanjšanja njegovega delovanja. Režim Izdajanja zdravila Rp/Spec Datum priprave informacije Februar 2007 Pred predpisovanjem, prosimo, preberite celoten povzetek glavnih znacilnosti zdravila. Dodatne informacije in literatura so na voljo pri: AstraZeneca UK Llmited Podružnica v Sloveniji Verovškova 55, Ljubljana in na spletnih straneh: www.breastcancersource.com www.arlmldex.net AstraZeneca ® ERBITUX CETUKSIMAB Erbitux S mg/ml raztopina za infundiranje (skrajšana navodila za uporabo) Cetuksimab je monoklonsko lgG1 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 citotoksicni terapiji, ki je vkljucevala tudi irinotekan. Zdravilo Erbituxje v kombinaciji z radioterapijo indicirano za zdravljenje bolnikov z lokalno napredovalim rakom skvamoznih celic glave in vratu. Odmerjanje in nacin uporabe: Zdravilo Erbitux pri vseh indikacijah infundirajte enkrat na teden. Zacetni odmerek je 400 mg cetuksimaba na m' telesne površine. Vsi naslednji tedenski odmerki so vsak po 250 mg/m'. Kontraindikacije: Zdravilo Erbitux je kontraindicirano pri bolnikih z znano hudo preobcutljivostno reakcijo (3. ali 4. stopnje) na cetuksimab. Posebna opozorila in previdnostni ukrepi: ce pri bolniku nastopi blaga ali zmerna reakcija, povezana z infundiranjem, lahko zmanjšate hitrost infundiranja. Priporocljivo je, da ostane hitrost infundiranja na nižji vrednosti tudi pri vseh naslednjih infuzijah. ce se pri bolniku pojavi huda kožna reakcija(. 3. stopnje po kriterijih US National Cancer Institute, Common Toxicity Criteria; NCI-CTC), morate prekiniti terapijo s cetuksimabom. Z zdravljenjem smete nadaljevati le, ce se je reakcija pomirila do 2. stopnje. Priporoca se dolocanje koncentracije elektrolitov v serumu pred zdravljenjem in periodicno med zdravljenjem s cetuksimabom. Po potrebi se priporoca nadomešcanje elektrolitov. Posebna previdnost je potrebna pri oslabljenih bolnikih in pri tistih z obstojeco srcno-pljucno boleznijo. Neželeni ucinki: Zelo pogosti(. 1/10): dispneja, blago do zmerno povecanje jetrnih encimov, kožne reakcije, blage ali zmerne reakcije povezane z infundiranjem, blag do zmeren mukozitis. Pogosti(. 1/100, < 1/10): konjunktivitis, hude reakcije povezane z infundiranjem. Pogostost ni znana: Opazili so progresivno zniževanje nivoja magnezija v serumu, ki pri nekaterih bolnikih povzroca hudo hipomagneziemijo. Glede na resnost so opazili tudi druge elektrolitske motnje, vecinoma hipokalciemijo ali hipokaliemijo. Posebna navodila za shranjevanje: Shranjujte v hladilniku (2 •c -8 °C). Ne zamrzujte. Vrsta ovojnine in vsebina: 1 viala po 20 ml ali 100 ml. Imetnik dovoljenja za promet: Merek KGaA, 64271 Darmstadt, Nemcija. Podrobne informacije o zdravilu so objavljene na spletni strani Evropske agencije za zdravila (EMEA) http://www.emea.europa.eu. 1 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 II 1 ,MERCK www.oncology.merck.de 11 1 SERONO -malignim gliomom, na primer multiformnim gliob astt·oci1:om,om, . Odmerjanje in nacin uporabe Temodal smejo predpisati le zdravniki, ki imajo izkušnje z zdravljenjem možganskih tumorjev. Odrasli bolniki z novo diagnosticiranim glioblastomom multiforme Temodal se uporablja v kombinaciji z žarišcno radioterapijo (faza socasne terapije), temu pa sledi do 6 ciklov monoterapije z temozolomidom. Faza socasne terapije Zdravilo Temodal naj bolnik jemlje peroralno v odmerku 75 mg/m2 na dan 42 dni, socasno z žarišcno radioterapijo (60 Gy, dani · .••·•••• · . . bol11lkjeJT1ljfvesoc.;42:dnevhega•oodo1JjaosgcasH ,; ;;E. 1,5.;xy109/l;::števJlo,.onibocitovl. 1O0 1/10): glavobol, slabost, zaprtje. Pogosti(< 1/10, >1/100): zmanjšan apetit, driska, bruhanje, bolecine v trebuhu, astenija, bolecine, vroc­ina. Redki(< 1/1.000, >1/10.000): aritmija, bolecine v prsih, patološko delovanje jeter, zvecanje jetrnih transaminaz. VRSTA OVOJNINE IN VSEBINA: Škatla s S filmsko ob­loženimi tabletami. NACIN IZDAJE ZDRAVILA: Na zdravniški recept. IMETNIK DO­VOLJENJA ZA PROMET: Lek farmacevtska družba d.d., Verovškova 57, Ljubljana, Slovenija. INFORMACIJA PRIPRAVLJENA: april 2007 @lek clan skupine Sandoz Lek farmacevtska družba d. d. Verovškova 57. 1526 Liubliana. Slovenii,a • wwwJP.k.,si Radiologi; and Oncologi; Editorial policy Editorial policy of the journal Radiology and Oncology is to publish original scientific papers, professional papers, review arti­cles, case reports and varia (editorials, re­views, short communications, professional information, book reviews, letters, etc.) pertinent to diagnostic and intervention­al radiology, computerized tomography, magnetic resonance, ultrasound, nuclear medicine, radiotherapy, clinical and experi­mental oncology, radiobiology, radiophys­ics and radiation protection. The Editorial Board requires that the paper has not been published or submitted for publica­tion elsewhere: the authors are responsible for all statements in their papers. Accepted articles become the property of the journal and therefore cannot be published else­where without written permission from the editorial board. Papers concerning the work on humans, must comply with the principles of the declaration of Helsinki (1964). The approval of the ethical commit­tee must then be stated on the manuscript. Papers with questionable justification will be rejected. Manuscript written in English should be submitted to the Editorial Office in tripli­cate (the original and two copies), including the illustrations: Radiology and Oncology, Institute of Oncology, Zaloska 2, SI-1000 Ljubljana, Slovenia; (Phone: +386 (0)1 5879 369, Tel./Fax: +386 (0)1 5879 434, E-mail: gsersa@onko-i.si). Authors are also asked to submit their manuscripts electronically, either by E-mail or on CD rom. The type of computer and word-processing package should be specified (Word for Windows is preferred). All articles are subjected to editorial re­view and review by independent referee selected by the editorial board. Manuscripts which do not comply with the technical re­quirements stated herein will be returned to the authors for correction before peer­review. Rejected manuscripts are generally returned to authors, however, the journal cannot be held responsible for their loss. The editorial board reserves the right to ask authors to make appropriate changes in the contents as well as grammatical and stylistic corrections when necessary. The expenses of additional editorial work and requests for reprints will be charged to the authors. General instructions• • Radiology and Oncology will consider manuscripts prepared according to the Vancouver Agreement (N Engl J Med 1991; 324: 424-8, BMJ 1991; 302: 6772; JAMA 1997; 277: 927­34.). Type the manuscript double spaced on one side with a 4 cm margin at the top and left hand side of the sheet. Write the paper in grammatically and stylistically correct language. Avoid abbreviations un­less previously explained. The technical