Ethanol-induced formation of colorectal tumours and precursors in a mouse model of Lynch syndrome
Lynch Syndrome (LS) confers an inherited cancer predisposition, particularly for colorectal cancer, due to germline mutations in one of the DNA mismatch repair (MMR) genes, such as MSH2. MMR is a DNA damage repair pathway involved in the removal of base mismatches and insertion/deletion loops, caused by several endogenous and exogenous factors. Loss of MMR through somatic alteration of the wild-type MMR allele in LS results in defective MMR (dMMR). Lifestyle factors can modify cancer risk for LS and sporadic patients. Ethanol and its metabolite acetaldehyde, are classified as group one carcinogens by the International Agency for Research on Cancer, and are risk factors for sporadic cancers of the upper aerodigestive tract, liver, breast and colorectum. Acetaldehyde is metabolised to acetate by the Aldh family of enzymes, particularly Aldh1b1 in the intestines. Acetaldehyde is very reactive and may cause a range of DNA lesions. However, DNA repair pathways responsible for correcting such lesions remain unknown. It was hypothesized that MMR plays a role in protecting intestinal cells from ethanol/acetaldehyde-induced DNA damage. This study aimed to determine if there is a gene/environment interaction between dMMR and ethanol/acetaldehyde that accelerates colorectal tumour development and progression.A conditional Msh2 knockout (“Msh2-LS”) mouse model with one deleted and one conditional knockout Msh2 allele was used, as it mimics the LS patients’ pattern of MMR gene inactivation. The Msh2-LS model mice were fed either with 20% ethanol in drinking water or normal drinking water. Long-term ethanol consumption led to large intestinal mucosal epithelial hyperproliferation and adenoma formation in 65% (15/23) mice and, in some cases, invasive adenocarcinoma (5/23 mice, 21.7%) within 6 months (mostly in the proximal and mid-colon), compared with 0% (0/23 mice) at 6 months and only one colonic tumour after 15 months in the water-treated mice (p<0.0001). No small intestinal tumours were observed. Additionally, a conditional Aldh1b1 knockout (Aldh1b1flox/flox) Msh2-LS mouse model and a constitutive Aldh1b1 knockout (Aldh1b1-/-) Msh2-LS mouse model were generated, in which the lack of Aldh1b1 enzyme caused increased acetaldehyde levels and acetaldehyde-induced DNA damage. In these Aldh1b1-deficient mice, long-term ethanol consumption led to increased numbers of colorectal adenomas per mouse (4.2, 21 neoplasms in 5 tumour-bearing conditional Aldh1b1fl/fl Msh2-LS mice; and 4.8, 35 neoplasms in 8 tumour-bearing constitutive Aldh1b1-/- Msh2-LS mice) compared with 2.4 (36 neoplasms in 15 tumour-bearing mice) colorectal adenomas per mouse observed in the Msh2-LS mouse model with wild-type Aldh1b1 (p=0.0319 and p=0.0103) , but no colonic tumours were observed in water-treated controls.Precursor lesions were observed as dMMR crypts in the murine colon in all of these mouse models, and their quantification showed increased numbers of dMMR crypt foci in ethanol-treated mice compared with water-treated controls (p=0.0029 in Aldh1b1wt Msh2-LS mice, p=0.0006 in Aldh1b1fl/fl Msh2-LS mice and p<0.0001 in Aldh1b1-/- Msh2-LS mice). A significant increase in DNA damage was detected in the large intestinal epithelium of ethanol-treated mice of all genotypes compared with the respective water-treated controls (p=0.0009 in Aldh1b1wt Msh2-LS mice, p<0.0001 in Aldh1b1fl/fl Msh2-LS mice and in Aldh1b1-/- Msh2-LS mice), along with increased plasma acetaldehyde levels in ethanol-treated mice and acetaldehyde levels were higher in the plasma of Aldh1b1flox/flox Msh2-LS mice and Aldh1b1-/- Msh2-LS mice than in the plasma of Msh2-LS mice.In this study, evidence was provided for a role for Msh2 in protecting the MMR-proficient colonic epithelial cells against ethanol/acetaldehyde-induced DNA damage by activating DNA mismatch repair, triggering cell cycle arrest or cell death by apoptosis. A key role for Aldh1b1 was confirmed for protecting the large intestines from acetaldehyde-induced DNA damage and tumour formation. Long-term ethanol/acetaldehyde exposure was shown to accelerate dMMR-driven intestinal tumour formation and this is proposed to act via promoting proliferation (mucosal epithelial hyperproliferation) and suppressing apoptosis, thus enhancing survival of aberrant dMMR intestinal epithelial cells/crypts relative to MMR-proficient intestinal epithelial cells/crypts, leading to adenoma development (with microsatellite instability) with some progressing to adenocarcinomas. In conclusion, there is strong evidence for a gene/environment interaction between acetaldehyde/dMMR, causing the acceleration of dMMR-driven intestinal tumour formation upon ethanol exposure.