The protein O6-methylguanine-DNA methyltransferase (MGMT) is able to repair the mutagenic O6-methylguanine (O6-MeG) adduct back to guanine. In this context, it may protect against colorectal cancer formation associated with N-nitroso compounds. Such compounds may be endogenously formed by nitrosylation of amino acids, which can give rise to mutagenic O6-MeG and O6-carboxymethylguanine (O6-CMG) adducts. It is well established that O6-MeG is repaired by MGMT. However, up to now, whether O6-CMG is repaired by this enzyme remains unresolved. Therefore, the aim of the present study was to analyze the fate of both types of O6-guanine adducts in the presence and absence of MGMT activity. To this end, MGMT activity was efficiently blocked by its chemical inhibitor O6-benzylguanine in human colon epithelial cells (HCECs). Exposure of cells to azaserine (AZA) caused significantly higher levels of both O6-MeG and O6-CMG adducts in MGMT-inhibited cells, with O6-CMG as the more abundant DNA lesion. Interestingly, MGMT inhibition did not result in higher levels of AZA-induced DNA strand breaks in spite of elevated DNA adduct levels. In contrast, MGMT inhibition significantly increased DNA strand break formation after exposure to temozolomide (TMZ), a drug that exclusively generates O6-MeG adducts. In line with this finding, the viability of the cells was moderately reduced by TMZ upon MGMT inhibition, whereas no clear effect was observed in cells treated with AZA. In conclusion, our study clearly shows that O6-CMG is repaired by MGMT in HCEC, thereby suggesting that MGMT might play an important role as a tumor suppressor in diet-mediated colorectal cancer.

中文翻译：

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O6-甲基鸟嘌呤-DNA甲基转移酶在人结肠上皮细胞中修复O6-羧甲基鸟嘌呤加合物。

蛋白质 O6-甲基鸟嘌呤-DNA 甲基转移酶 (MGMT) 能够将诱变的 O6-甲基鸟嘌呤 (O6-MeG) 加合物修复回鸟嘌呤。在这种情况下，它可以防止与 N-亚硝基化合物相关的结直肠癌形成。此类化合物可以通过氨基酸的亚硝基化内源性形成，这会产生诱变的 O6-MeG 和 O6-羧甲基鸟嘌呤 (O6-CMG) 加合物。众所周知，O6-MeG 是由 MGMT 修复的。然而，到目前为止，O6-CMG是否被这种酶修复还没有定论。因此，本研究的目的是分析两种类型的 O6-鸟嘌呤加合物在 MGMT 活性存在和不存在的情况下的命运。为此，MGMT 活性被其化学抑制剂 O6-苄基鸟嘌呤在人结肠上皮细胞 (HCEC) 中有效阻断。细胞暴露于氮杂丝氨酸 (AZA) 导致 MGMT 抑制细胞中 O6-MeG 和 O6-CMG 加合物的水平显着升高，其中 O6-CMG 作为更丰富的 DNA 损伤。有趣的是，尽管 DNA 加合物水平升高，MGMT 抑制并未导致更高水平的 AZA 诱导的 DNA 链断裂。相比之下，MGMT 抑制显着增加了暴露于替莫唑胺 (TMZ) 后 DNA 链断裂的形成，TMZ 是一种专门生成 O6-MeG 加合物的药物。与这一发现一致，在 MGMT 抑制后，TMZ 适度降低了细胞的活力，而在用 AZA 处理的细胞中没有观察到明显的影响。总之，我们的研究清楚地表明 O6-CMG 在 HCEC 中被 MGMT 修复，从而表明 MGMT 可能在饮食介导的结直肠癌中作为肿瘤抑制因子发挥重要作用。