Establishment of Novel Gastric Cancer Patient-Derived Xenografts and Cell Lines: Pathological Comparison between Primary Tumor, Patient-Derived, and Cell-Line Derived Xenografts.

Patient-derived xenograft (PDX) models have been recognized as being more suitable for predicting therapeutic efficacy than cell-culture models. However, there are several limitations in applying PDX models in preclinical studies, including their availability-especially for cancers such as gastric cancer-that are not frequently encountered in Western countries. In addition, the differences in morphology between primary, PDX, and tumor cell line-derived xenograft (CDX) models have not been well established. In this study, we aimed to establish a series of gastric cancer PDXs and cell-lines from a relatively large number of gastric cancer patients. We also investigated the clinicopathological factors associated with the establishment of PDX and CDX models, and compared the histology between the primary tumor, PDX, and CDX that originated from the same patient. We engrafted 232 gastric cancer tissues into immune-deficient mice subcutaneously and successfully established 35 gastric cancer PDX models (15.1% success rate). Differentiated type adenocarcinomas (DAs, 19.4%) were more effectively established than poorly differentiated type adenocarcinomas (PDAs, 10.8%). For establishing CDXs, the success rate was less influenced by histological differentiation grade (DA vs. PDA, 12.1% vs. 9.8%). In addition, concordance of histological differentiation grade between primary tumors and PDXs was significant (p < 0.01), while concordance between primary tumors and CDXs was not. Among clinicopathological factors investigated, pathological nodal metastasis status (pN) was significantly associated with the success rate of PDX establishment. Although establishing cell lines from ascites fluid was more efficient (41.2%, 7/17) than resected tissues, it should be noted that all CDXs from ascites fluid had the PDA phenotype. In conclusion, we established 35 PDX and 32 CDX models from 249 gastric cancer patients; among them, 21 PDX/CDX models were established from the same patients. Our findings may provide helpful insights for establishing PDX and CDX models not only from gastric but from other cancer types, as well as select preclinical models for developing new therapeutics.

AZD2171 shows potent antitumor activity against gastric cancer over-expressing fibroblast growth factor receptor 2/keratinocyte growth factor receptor.

PURPOSE: AZD2171 is an oral, highly potent, and selective vascular endothelial growth factor signaling inhibitor that inhibits all vascular endothelial growth factor receptor tyrosine kinases. The purpose of this study was to investigate the activity of AZD2171 in gastric cancer. EXPERIMENTAL DESIGN: We examined the antitumor effect of AZD2171 on the eight gastric cancer cell lines in vitro and in vivo. RESULTS: AZD2171 directly inhibited the growth of two gastric cancer cell lines (KATO-III and OCUM2M), with an IC(50) of 0.15 and 0.37 micromol/L, respectively, more potently than the epidermal growth factor receptor tyrosine kinase inhibitor gefitinib. Reverse transcription-PCR experiments and immunoblotting revealed that sensitive cell lines dominantly expressed COOH terminus-truncated fibroblast growth factor receptor 2 (FGFR2) splicing variants that were constitutively phosphorylated and spontaneously dimerized. AZD2171 completely inhibited the phosphorylation of FGFR2 and downstream signaling proteins (FRS2, AKT, and mitogen-activated protein kinase) in sensitive cell lines at a 10-fold lower concentration (0.1 micromol/L) than in the other cell lines. An in vitro kinase assay showed that AZD2171 inhibited kinase activity of immunoprecipitated FGFR2 with submicromolar K(i) values ( approximately 0.05 micromol/L). Finally, we assessed the antitumor activity of AZD2171 in human gastric tumor xenograft models in mice. Oral administration of AZD2171 (1.5 or 6 mg/kg/d) significantly and dose-dependently inhibited tumor growth in mice bearing KATO-III and OCUM2M tumor xenografts. CONCLUSIONS: AZD2171 exerted potent antitumor activity against gastric cancer xenografts overexpressing FGFR2. The results of these preclinical studies indicate that AZD2171 may provide clinical benefit in patients with certain types of gastric cancer.

A photon counting technique for quantitatively evaluating progression of peritoneal tumor dissemination.

We recently established a mouse model of peritoneal dissemination of human gastric carcinoma, including the formation of ascites, by orthotopic transplantation of cultured gastric carcinoma cells. To clarify the processes of expansion of the tumors in this model, nude mice were sacrificed and autopsied at different points of time after the orthotopic transplantation of the cancer cells for macroscopic and histopathologic examination of the tumors. The cancer cells grew actively in the gastric submucosa and invaded the deeper layers to reach the serosal plane. The tumor cells then underwent exfoliation and became free followed by the formation of metastatic lesions initially in the greater omentum and subsequent colonization and proliferation of the tumors on the peritoneum. Although this model allowed the detection of even minute metastases, it was not satisfactory from the viewpoint of quantitative and objective evaluation. To resolve these problems, we introduced a luciferase gene into this tumor cell line with a high metastasizing potential and carried out in vivo photon counting analysis. This photon counting technique was found to allow objective and quantitative evaluation of the progression of peritoneal dissemination on a real-time basis. This animal metastatic model is useful for monitoring the responses of tumors to anticancer agents.

Development and biological analysis of peritoneal metastasis mouse models for human scirrhous stomach cancer.

The number of published studies on peritoneal dissemination of scirrhous gastric carcinoma is very small as a result of the unavailability of highly reproducible animal models. Orthotopic implantation of HSC-44PE and HSC-58 (scirrhous gastric carcinoma-derived cell lines) cells into nude mice led to dissemination of the tumor cells to the greater omentum, mesenterium, peritoneum and so on, and caused ascites in a small number of animals. Cycles of isolation of the ascitic tumor cells and orthotopic inoculation of these cells were repeated in turn to animals. This was to isolate highly metastatic cell lines with a strong capability of inducing the formation of ascites (44As3 from HSC-44PE; 58As1 and 58As9 from HSC-58). All three cell lines induced tumor formation at the site of orthotopic injection, and caused fatal cancerous peritonitis and bloody ascites in 90-100% of the animals approximately 3-5 weeks after the inoculation. When the parent cells were implanted, the animals became moribund in approximately 12-18 weeks, however, none of the animals developed ascites. Complementary DNA microarray and immunohistochemical analyses revealed differences in the expression levels of genes coding for the matrix proteinase, cell adhesion, motility, angiogenesis and proliferation between the highly metastatic- and parent-cell lines. The usefulness of this model for the evaluation of drugs was assessed by analyzing the stability of the metastatic potential of the cells and the reproducibility. Animals intravenously treated with CPT-11 and GEM showed suppressed tumor growth and significantly prolonged survival. The metastatic cell lines and the in vivo model established in the present study are expected to serve as a model of cancerous peritonitis developing from primary lesions, and as a useful means of clarifying the pathophysiology of peritoneal dissemination of scirrhous gastric carcinoma and the development of drugs for its treatment.