5-FU in combination with PARP inhibitor ABT-888 deregulates MGMT-dependent mismatch repair (MMR) pathway in MMR-proficient colorectal cancer stem cells by modulating MGMT/PARP1/MSH6 complex.

BACKGROUND: Previous study showed the PARP inhibitor ABT-888 potentiates the cytotoxicity of 5-fluorouracil (5-FU) by inhibiting PARP1-mediated mismatch repair (MMR) pathway via MSH6 deregulation in MMR-proficient colorectal cancer stem cells (CRC-CSCs). Emerging evidence indicates 5-FU regulates O6 -methylguanine-DNA-methyltransferase (MGMT) activity, but the mechanistic basis of MGMT involvement in PARP1-mediated MMR pathway following 5-FU treatment remains complex and poorly defined.

RESEARCH DESIGN AND METHODS: This study delineates the role of MGMT in 5-FU-induced MMR pathway activation and evaluates 5-FU+ABT-888 combination effects on MGMT modulation in CRC-CSCs. The molecular mechanism has been studied by using colocalization, western blot, co-immunoprecipitation, MGMT gene-knockdown, and molecular docking in in vitro, in silico, and ex vivo preclinical models.

RESULTS: 5-FU treatment induced PARylated-PARP1 in CRC-CSCs, promoting PARP1-MGMT-MSH6 interactions that activated MMR. ABT-888 inhibited PARylation in 5-FU-pre-exposed CSCs. Therefore, PARP1 could not physically interact with both MGMT and MSH6, and complete abolishment of MGMT and MSH6, and MMR protein downregulation were observed in combination treatment. MGMT silencing confirmed its critical role in PARP1-mediated MMR activation. Similar findings were obtained in in silico and ex vivo models.

CONCLUSIONS: 5-FU+ABT-888 enhanced CRC-CSCs death by inhibiting the PARP1-MGMT-MSH6 interaction and simultaneously inhibiting the MGMT-dependent PARP1-mediated MMR pathway in MMR-proficient CRC-CSCs.