Synergistic efficacy of O 6benzylguanine and 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) in a human colon cancer xenograft completely resistant to BCNU alone

Abstract

The DNA repair protein O 6alkylguanine-DNA alkyltransferase (alkyltransferase) repairs cytotoxic DNA damage formed by 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU). High levels of this repair protein cause tumor drug resistance to nitrosoureas. To investigate the ability of a direct alkyltransferase inhibitor, O 6-benzylguanine, to reverse the nitrosourea resistance of human colon cancer cells, we studied the VACO 6 cell line which has high alkyltransferase and is completely resistant to BCNU at maximal tolerated doses in the xenograft model. O 6-Benzylguanine at 0.5 μg mL for 1 hr inactivated VACO 6 alkyltransferase by > 98% and reduced the ic 50 of BCNU by 3- to 4-fold. Further analysis indicated that these two agents act in a highly synergistic fashion. In xenograft bearing athymic mice, dose-dependent depletion of hepatic and tumor alkyltransferase was noted. To maintain alkyltransferase depletion in the xenograft for at least 24 hr, two doses of 60 mg kg O 6-benzylguanine were given 1 hr prior and 7 hr after BCNU. Under these conditions, VACO 6 xenografts became responsive to BCNU with significant reductions (P < 0.001) in the tumor growth rate. The combination increased toxicity to the host, reducing the maximum tolerated dose of BCNU by approximately 50%. This study provides definitive evidence that high alkyltransferase activity is responsible for BCNU resistance in human colon cancer xenografts and that with careful drug scheduling, O 6-benzylguanine can sensitize a tumor which is completely unresponsive to BCNU alone. Further studies which optimize the therapeutic index of BCNU and O 6-benzylguanine in vivo will define the schedule to be used in broader preclinical studies.