This study was designed to evaluate the in vitro cytotoxicity and in vivo efficacy of TRA-8, a mouse monoclonal antibody that binds to the DR5 death receptor for tumor necrosis factor-related apoptosis-inducing ligand (TRAIL/Apo2L), alone and in combination with CPT-11 against human colon cancer cells and xenografts.
DR5 expression was assessed on human colon cancer cell lines using flow cytometry, and cellular cytotoxicity following TRA-8 treatment, alone and in combination with SN-38 was determined by measuring cellular ATP levels. Tumor growth inhibition and regression rates of well-established s.c. COLO 205, SW948, HCT116 and HT-29 colon cancer xenografts in athymic nude mice treated with TRA-8 or CPT-11 alone and in combination were determined. 99mTc-TRA-8 was utilized to examine tumor localization of TRA-8 in animals bearing each of the 4 xenografts. In addition, whole body biodistribution and imaging was carried out in COLO 205 bearing animals using in vivo SPECT imaging and tissue counting.
DR5 expression was highest on HCT116, intermediate on SW948 and COLO 205 cells, and lowest on HT-29 cells. COLO 205 cells were the most sensitive to TRA-8-induced cytotoxicity in vitro, SW948 and HCT116 cell lines were moderately sensitive, and HT-29 cells were resistant. Combination treatment with TRA-8 and SN-38 produced additive to synergistic cytotoxicity against all cell lines compared to either single agent. The levels of apoptosis in all cell lines, including HT-29, were increased by combination treatment with SN-38. In vivo, combination therapy with TRA-8 and CPT-11 was superior to either single agent regimen for three of the xenografts; COLO 205, SW948, and HCT116. COLO 205 tumors were most responsive to therapy with 73% complete regressions following combination therapy. HT-29 cells derived no anti-tumor efficacy from TRA-8 therapy. Tumor xenografts established from the 4 colon cancer cell lines had comparable specific localization of 99mTc-TRA-8.
In vitro and in vivo effects of TRA-8 anti-DR5 monoclonal antibody on four different colon cancer cell lines and xenografts were quite variable. The HT-29 cell line had low surface DR5 expression and was resistant to TRA-8 both in vitro and in vivo. Three cell lines (COLO 205, SW948, and HCT116) exhibited moderate to high sensitivity to TRA-8 mediated cytotoxicity which was further enhanced by the addition of SN-38, the active metabolite of CPT-11. In vivo, the combination of TRA-8 and CPT-11 treatment produced the highest anti-tumor efficacy against xenografts established from the three TRA-8 sensitive tumor cell lines. All 4 colon cancer xenografts had comparable localization of 99mTc-TRA-8. These studies support the strategy of TRA-8/CPT-11 combined treatment in human colon cancer clinical trials.