Enhanced anti-tumor effect via combination of Triptolide with ionizing radiation

Proc Amer Assoc Cancer Res, Volume 47, 2006 4720 Aim: Pancreas cancer is among the most resistant cancer to any therapy. The lack of effective treatment results in a very low survival rate, with median survivals of only about 6 month. This study is to explore the enhancement of therapeutic effect on human pancreatic cancer via combination of triptolide (TPL) and ionizing radiation (IR). Methods: AsPC-1 human pancreatic cancer cells cultured in 96 well plates were treated in vitro with different concentrations (0, 6, 12, 25 nM) of TPL alone or TPL plus 4 Gy IR one day later. The positive control is gemcitabine, a first line drug against pancreatic cancer. On day 4, the cell proliferation in anchorage-dependent conditions was analyzed by the sulforhodamine B (SRB) method. In addition, AsPC-1 cells treated with TPL or TPL plus IR were analyzed for their clonogenic survival. Finally, the extent of apoptosis induced by the above treatments were analyzed by Annexin V-FITC / propidium iodide (PI) staining as well as other apoptotic indices. In vivo, the AsPC-1 tumor xenografts were treated with 0.2 mg/kg TPL or 10 Gy IR or 100 mg/kg gemcitabine along or TPL plus IR. The tumor size was measured with Vernier calipers twice a week and the tumors were excised and weighed at the end of the experiment. The alterations of VEGF upon different treatments were determined by ELISA. Results: In vitro , TPL exerted a great inhibitory effect on growth of AsPC-1 cells, which was much more potent than gemcitabine (ID50 25 nM vs. >200 nM). The combination of TPL with IR shifted the ID50 of TPL from 25 nM to 12.5 nM (anchorage dependant phase cells) and reduced clonogenic survival of AsPC-1 cells (exponential growth phase cells). Cells treated with TPL plus IR exhibited a greater apoptosis as evidenced by stronger Annexin V/PI staining ratio, higher levels of cleaved caspases and increased DNA fragmentation as compared with TPL or IR alone. In vivo , the tumor growth of AsPC-1 xenografts was much slower in the group treated with TPL plus IR compared to the TPL or IR or gemcitabine alone groups. Importantly, 2 of 5 mice had prolonged complete responses (14 days). The VEGF level of AsPC-1 cells was greatly reduced upon treatment with TPL plus IR. Conclusion: 1) TPL possesses a potent therapeutic effect on pancreatic cancer, which seems superior to a current first line drug, gemcitabine; 2) the combination of TPL with IR enhances the therapeutic effect both in vitro and in vivo ; and 3) the inhibitory effect of TPL plus IR on pancreatic cancer is associated with induction of apoptosis and reduction of VEGF.