Dna repair by mgmt, but not aag, causes a threshold in alkylation-induced colorectal carcinogenesis

Abstract

Epidemiological studies indicate that N-nitroso compounds (NOC) are causally linked to colorectal cancer (CRC). NOC induce DNA alkylations, including O6-methylguanine (O6-MeG) and N-methylated purines, which are repaired by O6-MeGDNA methyltransferase (MGMT) and N-alkyladenine-DNA glycosylase (AAG)-initiated base excision repair, respectively. In view of recent evidence of nonlinear mutagenicity for NOC-like compounds, the question arises as to the existence of threshold doses in CRC formation. Here, we set out to determine the impact of DNA repair on the dose-response of alkylation-induced CRC. DNA repair proficient (WT) and deficient (Mgmt-/-, Aag-/- and Mgmt-/-/Aag-/-) mice were treated with azoxymethane (AOM) and dextran sodium sulfate to trigger CRC. Tumors were quantified by non-invasive mini-endoscopy. A non-linear increase in CRC formation was observed in WT and Aag-/- mice. In contrast, a linear dose-dependent increase in tumor frequency was found in Mgmt-/- and Mgmt-/-/Aag-/- mice. The data were corroborated by hockey stick modeling, yielding similar carcinogenic thresholds for WT and Aag-/- and no threshold for MGMT lacking mice. O6-MeG levels and depletion of MGMT correlated well with the observed dose-response in CRC formation. AOM induced dose-dependently DNA double-strand breaks in colon crypts including Lgr5-positive colon stem cells, which coincided with ATR-Chk1-p53 signaling. Intriguingly, Mgmt-/- mice displayed significantly enhanced levels of γ-H2AX, suggesting the usefulness of γ-H2AX as an early genotoxicity marker in the colorectum. This study demonstrates for the first time a non-linear dose-response for alkylation-induced colorectal carcinogenesis and reveals DNA repair by MGMT, but not AAG, as a key node in determining a carcinogenic threshold.