300. Gene Therapy for Gastric Cancer Cells Via CEA-Targeted Adv-FZ33

Abstract

IntroductionAt present, adenoviral vectors have been widely used for gene transfer into tumor cells primarily because they can be produced in high titers and can infect many different cell types. However, a major problem when utilizing the adenoviral vectors for gene therapy applications is thought to be related to low transduction efficiency due to weak expression of the adenovirus receptor, coxsackie– adenovirus receptor (CAR) in cancer cells, or side effects due to expression of CAR in normal cells. There is an urgent requirement to improve the specificity of gene delivery via adenoviral vectors in the context of cancer gene therapy.Experimental designWe constructed a fiber modified Ad5 vector (Adv-FZ33) in which a IgG-binding domain (Z33) derived from staphylococcal protein A was inserted into the HI loop of knob protein. We then evaluated on both in vitro and in vivo levels the extent of retargeting towards and therapeutic effectiveness against CEA-positive gastric cancers when using the fully human CEA antibody (C2-45) complex with this modified vector.ResultsIn vitro LacZ or EGFP reporter gene expression after infection with Adv-FZ33 vectors conjugated to anti-CEA mAb was approximately 20 times higher than that obtained with vector conjugated to the control mAb (human IgG4). We generated Ax3CAUP-FZ33, which is an Adv-FZ33 derivative vector expressing a therapeutic gene, namely E. coli uracil phosphoribosyltransferase (UPRT) which converts 5FU directly to 5-fluorouridine monophosphate. When conjugated with the anti-CEA mAb, C2-45, Ax3CAUP-FZ33 enhanced the cytotoxicity of 5FU (19.7-fold in terms of IC50 values) against gastric cancer cells. In a nude mouse peritoneal dissemination model, use of Ax3CAUP- FZ33 conjugated with anti-CEA mAb C2-45, in combination with 5FU administration, resulted in greatly enhanced anti-tumor effects against CEA-positive cancer cells. Tumor growth in mice treated with Ax3UP-FZ33/C2-45/5FU was significantly suppressed, and tumor volumes were less than a quarter of those of the control IgG4 group (total weight, 0.12 ± 0.11 g; P < 0.05). Furthermore, we examined the survival effects of Ax3UP-FZ33 conjugated with anti- CEA mAb plus 5FU in mice with peritoneal disseminated gastric cancer. The median survival time of the Ax3CAUP-FZ33/C2-45/5FU group was significantly longer than those of the PBS and 5FU only treatment groups (p<0.01, versus PBS and 5FU only groups). On the other hand, no significant difference was present between the Ax3CAUP-FZ33/5FU or Ax3CAUP-FZ33/IgG4/5FU groups and the PBS or 5FU only groups.ConclusionsThese data suggest that CEA-targeted FZ33-mutant adenovirus- mediated gene delivery offers a strong and selective therapeutic modality against CEA-producing cancers. This novel strategy may be a promising tool for cancer gene therapy.