Abstract 1445: Emergence of frameshift mutations during tumorigenesis in VCMsh2 mouse model of Lynch syndrome

Abstract

Abstract Monoallelic germline mutations in MMR genes (e.g., MLH1, MSH2, MSH6, and PMS2) predispose individuals to Lynch syndrome (LS) with microsatellite instability (MSI) throughout the genome. Mutations at coding mononucleotide repeats (MNR) usually result in frameshift mutations (FSMs). Recurrent FSMs are thought to play a central role in the increased risk of different types of cancer. Neoantigens produced by FSMs have been shown to elicit immune responses, which is the basis for frameshift neoantigen-based vaccines. To develop a prophylactic vaccine and prevention strategy for this high-risk population, we assessed FSMs during tumorigenesis from histologically normal mucosa and fecal DNA of a LS mouse model using targeted sequencing and a panel of FSMs in MNR regions. Msh2LoxP/LoxP;Villin-Cre mice (VCMsh2) started developing detectable tumors at 7-8 months and median survival was 11.5 months with 100% penetrance. Interestingly, FSMs were detectable not only in tumors and mucosa at 7-8 months, but also in young mice (1 month), embryos and pups although FSMs detected and variant allele frequency (VAF) were very low, indicating that FSMs accumulated over time, and FSMs alone may be not sufficient for tumorigenesis since tumors emerge at older age. To determine whether Msh2 was absent in embryos and pups, immunohistochemical (IHC) analysis was performed. Msh2 was lost in almost all intestine epithelial cells in 2-month-old mice but still present in young pups and embryos with very few cells absent of Msh2, indicating that emerging FSMs in these young pups may be due to decreased Msh2 expression because of delayed Cre function. To determine whether low Msh2 expression could lead to the emergence of FSMs, intestine mucosa from 8-, 10-, and 12-month-old Msh2LoxP/+;Villin-Cre heterozygous mice was sequenced. FSMs were detectable with low VAF, although they didn’t develop tumors. IHC staining revealed that Msh2 was absent in only a few intestine epithelial cells in these heterozygous mice, suggesting that loss of heterozygosity was a late event and rare at 12 months. To determine whether MSI emerged in young VCMsh2 pups and heterozygous mice due to haploinsufficiency of Msh2, MSI was assessed using seven markers and fragment size analysis. One marker (mBat67) showed instability in the intestinal mucosa of heterozygous mice and three showed instability in fecal DNA of one month old VCMsh2 mice. Interestingly, mucosal and fecal samples from a time course study in VCMsh2 mice showed progressive increase in MSI with a good correlation between MSI and FSMs during the tumorigenesis process. In summary, FSMs emerged at an early stage of tumorigenesis in VCMsh2 mice, which correlated with MSI status. Our data indicates that FSMs and MSI status can be used to monitor the tumor development of LS colorectal cancer. Funded by the National Cancer Institute, National Institutes of Health, under Contract No. HHSN261201500003I Citation Format: Yurong Song, Lei Wei, Shaneen S. Baxter, Brandon Somerville, Holli Loomans-Kropp, Chelsea Sanders, Ryan N. Baugher, Stephanie D. Mellott, Todd B. Young, Heidi E. Lawhorn, Teri M. Plona, Qiang Hu, Song Liu, Alan Hutson, Simone Difilippantonio, Ligia Pinto, Steven M. Lipkin, Matthias Kloor, Shizuko Sei, Robert H. Shoemaker. Emergence of frameshift mutations during tumorigenesis in VCMsh2 mouse model of Lynch syndrome [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1445.