Accumulation of K‐Rascodon 12 mutations in the F344 rat distal colon following azoxymethane exposure

Abstract

Azoxymethane (AOM) administration to F344 male rats is a widely used model of human colon carcinogenesis. The current study investigates quantitatively the accumulation of K-Ras codon 12 mutations following AOM exposure. Male, 6-week-old F344 rats were treated subcutaneously with 30 mg/kg body weight of AOM, and colon tissue was collected at 1, 8, 24, and 32 weeks after treatment. The K-Ras codon 12 GGT to GAT and GGT to GTT mutant fractions (MFs) were measured using allele-specific competitive blocker polymerase chain reaction (ACB-PCR). Between 1 and 32 weeks after AOM treatment, the K-Ras codon 12 GGT to GAT geometric mean MF in the rat colon increased significantly from 12.9 × 10(-5) to 145 × 10(-5) , and the GGT to GTT geometric mean MF increased significantly from 5.26 × 10(-5) to 180 × 10(-5) . K-Ras codon 12 GGT to GAT MF also increased significantly in AOM-treated rat colon tissue at 1 week compared to controls (4.44 × 10(-5) ). The accumulation of the GGT to GAT MF long after the DNA adduct repair phase suggests that a portion of cells containing this mutation have a proliferative advantage, allowing them to accumulate as nascent tumors progress. Also, the GGT to GAT background MF increased in untreated rats, indicating that there is accumulation with age. The ACB-PCR assay generates quantitative data of cancer-related mutations and thus provides insight into pathological processes following carcinogen exposure.