Molecular Pathogenesis and Immune Biology of Intestinal MSI Tumors in a DNA Mismatch Repair-Deficient Mouse Model

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Microsatellite instability (MSI) occurs in almost all Lynch syndrome cancers and in about 15% of sporadic colorectal cancers due to functional inactivation of the cellular DNA mismatch repair (MMR) system. MSI colorectal tumorigenesis in Lynch syndrome patients is assumed to proceed through multiple steps which in comparison to the sporadic forms include an accelerated progression from adenomas to carcinomas. MMR deficiency manifests early during adenoma development and even occurs in normal-appearing mucosa. Instability at coding mononucleotide repeats (cMNRs) leads to frameshift mutations in some genes that can drive MSI tumorigenesis.The functional role and clinical utility of cMNR instability in murine MMR deficient tumors has not been examined yet. Previous MMR-deficient mice do not accurately recapitulate the human situation because homozygous mutant MMR mice predominantly develop lymphomas and only few intestinal tumors. However, recently established conditional MMR gene knockout mice for the first time provide the opportunity to address research questions related to the specific implications of defined cMNR mutations in vivo. In preliminary work cMNR mutations in mouse genes have been identified. We plan to extend our initial experiments to a conditional MMR k.o. mouse line (VCMsh2loxP), a preclinical model of Lynch syndrome. These mice lack intestine-specific Msh2 expression and develop intestinal tumors. Whole intestines from VCMsh2loxP mice at different ages will be examined to identify preneoplastic and neoplastic lesions as well as potential early histological alterations in tumor-adjacent and -distant normal mucosa. Histological and MMR immunostaining analyses of whole intestines from heterozygous VCMsh2loxP mice will resolve whether MMR-deficient crypt foci as recently observed in human Lynch syndrome patients also exist in these mice. For all histologically characterized normal mucosa and tumor tissues, mutation profiles of bioinformatically identified cMNR candidate genes will be determined and based on these mutation data tumor-relevant MSI target genes be sought. Potential correlations between mutations in MSI target genes and age-dependent development of MSI neoplasms and preneoplasia will be investigated. Comparison of human and mouse mutation profiles in MSI target genes and corresponding signaling pathways will define differences and overlaps in murine and human MSI carcinogenesis. Immunohistochemical analyses will determine the type and pattern of immune infiltration with emphasis on the identity of the cells involved. These studies will provide important mechanistic insights into the etiology and progression of MSI tumors. Uncovering tumor-specific murine MSI target genes enables prediction of frameshift peptides derived thereof and a molecular basis to test preventive frameshift peptide-based vaccination strategies for MSI tumors in an animal model.