NOTE 
Annals of Nuclear Medicine Vol. 4, No. 2 71-74, 1990 
Radioimmunoscintigraphy of human pancreatic carcinoma xenografts in nude mice with 131I-labeled monoclonal antibody 
Takatoshi TSUDA H KOSHIBA T USUI,** M. KUBOTA,* Kokichi KIKUCHI*** and Kazuo MORITA* 
*Department of Radiology, **First Department of Surgery ***First Department of Pathology, Sapporo Medical College, Sapporo 
Encouraged by reports of radioimmunoimaging of colorectal carcinomas1-3 and by examining an immunohistochemical report on resected pancreas cancer tissues4 we studied the diag-nostic potential of radioimmunoimaging with the radioiodinelabeled monoclonal antibody to the surface antigen of a pancreas cancer cell line. A monoclonal antibody (MoAb; HC-1) to a human pancreas cancer cell line (HGC25)5 was labeled with radioiodine and injected into athymic nude mice implanted with human pancreas cancer cells. Antibody HC-1 was cleared from the circulation and accumulated significantly in the implanted tumor sites. 
Key words : radioimmunoimaging, pancreas cancer, monoclonal antibody 
INTRODUCTION 
THE DETECTION Of malignant tumors by scintigraphy has become a theme of human cancer research. The development of the hybridoma technology which has allowed production of monospecific antibodies has followed the improvement of this research tech-nique.6 A monoclonal antibody (MoAb; HC-1) to a human pancreatic carcinoma cell line (HGC25) was established previously.4 Briefly, this antibody (HC-1) is an IgG2a which reacts not only with pancreas cancer cell lines but also with other cancer cell lines such as colon, stomach cancer and so on. And it does not detect HLA associated antigens since it failed to react with human myeloid and lymphoid cell lines and normal hematopoietic cells. Immunohistochemical analysis showed that this monoclonal antibody reacted with pancreas cancer tissues, but did not react with normal and other malignant epithelial tissues. This report describes the localization of human pancreatic carcinoma xenograft in nude mice using 131I-labeled HC-1. 
MATERIALS AND METHODS 
Antibody preparation : The monoclonal antibody HC-1 whose biological characterization was reported previously by Usui and Koshiba was isolated from ascitic fluid obtained from hybridoma bearing pristine primed BALB/C mice and purified by DEAE chromatography (0.04 M phosphate buffer, 0.03 M NaCl, pH 8.0) and then by Protain A affinity chromatography (0.1 M citrate-phosphate buffer, pH 4. 5).7 Radioiodination of whole lgG with iodine-131 (131I) was performed by the chloramine-T method.8 Un-bound iodine was removed by gel-filtration on a Sephadex G-50 column (PBS, pH 7.5). The result showed a specific activity of 2.5 uCi/ug. 
Tumor preparation : 0.1 ml (1 x 106/ml in PBS of the cloned pancreatic cancer cell line (HGC25)) was inoculated subcutaneously into the back or abdomen of BALB/c athymic nu/nu mice. The tumors which grew to 1.0-1.5 cm in diameter at 3-4 weeks after the inoculation were used in this study. The administration of non-radioactive iodine to the mice started on the seventh day before the injection of radioiodinated antibody and continued throughout the experiments. 
Biodistribution studies and radioimmunoimaging : For the scintigraphic examination and the bio-distribution study, radioiodinelabeled antibodies at doses of 10 to 20ug (0.925 to 1.85 MBq (25 to 50 uCi)) were injected into the tail vein of nude mice bearing the tumor. figures). 
Scintigrams were obtained with a gamma camera equipped with a pinhole collimator. The tumors and the selected organs from the mice sacrificed at suitable times were removed, weighed and counted. Biodistribution data were expressed as a percentage of the injected dose per gram of tissue normalized to a 20 g mouse. The tumor to blood ratio was also calculated. 
RESULTS 
131I-labeled MoAb HC-1 imaging: Scintigrams were obtained at 3 hours, 1 day, 6 days and 14 days after injection of the 131I labeled monoclonal antibody. It took 6 days to decrease the background radio-activity so that the tumor was well defined (Fig. 1). On the other hand, no significant activity in the transplanted tumors was observed in the scintigrams after intravenous injection of free 131I (not shown in figures). 
Biodistribution studies of labeled antibodies : The results of the in vivo localization of the labeled antibodies are shown in Table 1, whose figures indicate the percentages of injected dose per gram. The tumor-to-blood ratio of the radioiodinated whole IgG (HC-1) progressively increased with time. and the radioactivity of the transplanted tumors is high enough to separate from liver. 
DISCUSSION 
Hybridoma technology has provided numerous murine monoclonal antibodies specific for human malignant tumors. Various degrees of success have been obtained in studies of tumor localization using labeled MoAbs that react with known tumor makers such as carcinoembryonic antigens9-10 and alpha-fetoproteinll and MoAb which react with specific surface antigens of tumors including breast,colon, and other cancers.12 
Some monoclonal antibodies to pancreatic carcinoma have been produced,13,14 but there have been only a few reports on radioimmunoscintigraphy.15,16 We demonstrated here the specific radioimmunolocalization of xenografts of human pancreas carcinoma in nude mice by 131I labeled whole IgG (MoAb ; HC-1). The biological characteristics of HC-1 were previously reported in detail.4 As mentioned in the introductory part of this article, MoAb HC-1 is an IgG2a by which the antigenic determinant is recognized, and is a glycoprotein with a molecular weight of 130 k Daltons found in human pancreatic carcinoma cells but not in tumor extracts. 
For clinical use, 111In has many advantages over 131I, because of the low radiation dose and suitable energy for scintigraphic imaging. Moreover, the labeling of antibodies can be done rapidly and efficiently through chelation with DTPA.9,n As a matter of fact, 111In labeled monoclonal antibodies have been clinically used in imaging of the malignant melanoma and gastrointestinal tumors. However, non specific high tracer accumulation in normal organs, such as the liver, kidneys and spleen is observed, and this may interfere with the detection of abdominal tumors. It seems that the diagnostic value is limited by radioimmunoimaging with 111In conjugated antibody for pancreas cancer and for malignant tumors in the upper and lower abdomen. 
The final goal of this study is to examine and treat the patient with pancreas cancer, so we made mention of in vivo localization of the radioiodinated monoclonal antibody (HC-1). In serial scintigrams, excellent images of the tumor were obtained 6 days after the injection of 131I-IgG (HC-1), and the radioiodine activity in liver, kidneys and spleen was low. The investigation of in vivo localization of the radiolabeled antibody to the membrane associated antigen of one pancreatic carcinoma cell line (HGC25) of pancreatic carcinoma is summarized above. Because the problem has yet to be solved, we could not draw a conclusion on the heterogeniety and the antigenic modulation of pancreas tumor cells in this radioimmunoimaging study. It is necessary to continue the investigation of these problems. The present examination is still at the animal experimental stage. 
A summary of this report has been already presented at the 45th Annual Meeting of Japan Radiological Society, Tokyo, 1986 and the 27th Annual Meeting of the Japanese Society of Nuclear Medicine, Tokyo, 1987. 
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