An Msh2 conditional knockout mouse for studying intestinal cancer and testing anticancer agents.

BACKGROUND & AIMS Mutations in the DNA mismatch repair (MMR) gene MSH2 cause Lynch syndromes I and II and sporadic colorectal cancers. Msh2(null) mice predominantly develop lymphoma and do not accurately recapitulate the colorectal cancer phenotype. METHODS We generated and examined mice with a conditional Msh2 disruption (Msh2(LoxP)), permitting tissue-specific gene inactivation. ECMsh2(LoxP/LoxP) mice carried an EIIa-Cre transgene, and VCMsh2(LoxP/LoxP) mice carried a Villin-Cre transgene. We combined the VCMsh2(LoxP) allele with either Msh2(Delta7null) (VCMsh2(LoxP/null)) or Msh2(G674D) mutations (VCMsh2(LoxP/G674D)) to create allelic phase mutants. These mice were given cisplatin or 5-fluorouracil/leucovorin and oxaliplatin (FOLFOX), and their tumors were measured by magnetic resonance imaging. RESULTS Embryonic fibroblasts from ECMsh2(LoxP/LoxP) mice do not express MSH2 and are MMR deficient. Reverse transcription, polymerase chain reaction, and immunohistochemistry from VCMsh2(LoxP/LoxP) mice demonstrated specific loss of Msh2 messenger RNA and protein from epithelial cells of the intestinal tract. Microsatellite instability was observed in all VCMsh2 strains and limited to the intestinal mucosa. Resulting adenomas and adenocarcinomas had somatic truncation mutations to the adenomatous polyposis coli (Apc) gene. VCMsh2(LoxP/LoxP) mice did not develop lymphoma. Comparison of allelic phase tumors revealed significant differences in multiplicity and size. When treated with cisplatin or FOLFOX, tumor size was reduced in VCMsh2(LoxP/G674D) but not VCMsh2(LoxP/null) tumors. The apoptotic response to FOLFOX was partially sustained in the intestinal mucosa of VCMsh2(LoxP/G674D) animals. CONCLUSIONS Msh2(LoxP/LoxP) mice in combination with appropriate Cre recombinase transgenes have excellent potential for preclinical modeling of Lynch syndrome, MMR-deficient tumors of other tissue types, and use in drug development.

[1]  R. Kucherlapati,et al.  Loss of Rb1 in the gastrointestinal tract of Apc1638N mice promotes tumors of the cecum and proximal colon , 2008, Proceedings of the National Academy of Sciences.

[2]  A. Rosenwald,et al.  Distinct effects of the recurrent Mlh1G67R mutation on MMR functions, cancer, and meiosis , 2008, Proceedings of the National Academy of Sciences.

[3]  Kathleen R. Cho,et al.  Mouse model of colonic adenoma-carcinoma progression based on somatic Apc inactivation. , 2007, Cancer research.

[4]  R. Kucherlapati,et al.  Inactivation of conditional Rb by Villin-Cre leads to aggressive tumors outside the gastrointestinal tract. , 2006, Cancer research.

[5]  T. Maudelonde,et al.  Six novel heterozygous MLH1, MSH2, and MSH6 and one homozygous MLH1 germline mutations in hereditary nonpolyposis colorectal cancer. , 2004, Cancer genetics and cytogenetics.

[6]  Richard D Kolodner,et al.  The mismatch repair complex hMutS alpha recognizes 5-fluorouracil-modified DNA: implications for chemosensitivity and resistance. , 2004, Gastroenterology.

[7]  J. Jiricny,et al.  Mismatch repair and DNA damage signalling. , 2004, DNA repair.

[8]  Daniel Metzger,et al.  Tissue‐specific and inducible Cre‐mediated recombination in the gut epithelium , 2004, Genesis.

[9]  P. Peltomäki,et al.  HNPCC mutation MLH1 P648S makes the functional protein unstable, and homozygosity predisposes to mild neurofibromatosis type 1 , 2004, Genes, chromosomes & cancer.

[10]  R. Kucherlapati,et al.  An Msh2 Point Mutation Uncouples DNA Mismatch Repair and Apoptosis , 2004, Cancer Research.

[11]  G. Meijer,et al.  A Homozygous MSH6 Mutation in a Child with Café-au-Lait Spots, Oligodendroglioma and Rectal Cancer , 2004, Familial Cancer.

[12]  R. Kucherlapati,et al.  A panel of repeat markers for detection of microsatellite instability in murine tumors , 2003, Molecular carcinogenesis.

[13]  Z. Siddik,et al.  Cisplatin: mode of cytotoxic action and molecular basis of resistance , 2003, Oncogene.

[14]  Daniel J Sargent,et al.  Tumor microsatellite-instability status as a predictor of benefit from fluorouracil-based adjuvant chemotherapy for colon cancer. , 2003, The New England journal of medicine.

[15]  T. Kunkel,et al.  Inactivation of Exonuclease 1 in mice results in DNA mismatch repair defects, increased cancer susceptibility, and male and female sterility. , 2003, Genes & development.

[16]  N. Copeland,et al.  A highly efficient recombineering-based method for generating conditional knockout mutations. , 2003, Genome research.

[17]  R. Fishel,et al.  The selection for mismatch repair defects in hereditary nonpolyposis colorectal cancer: revising the mutator hypothesis. , 2001, Cancer research.

[18]  D. Court,et al.  A highly efficient Escherichia coli-based chromosome engineering system adapted for recombinogenic targeting and subcloning of BAC DNA. , 2001, Genomics.

[19]  M. Meyers,et al.  Role of the hMLH 1 DNA Mismatch Repair Protein in Fluoropyrimidine-mediated Cell Death and Cell Cycle Responses 1 , 2001 .

[20]  R. Kucherlapati,et al.  Tumor-associated Apc mutations in Mlh1−/−Apc1638N mice reveal a mutational signature of Mlh1 deficiency , 2000, Oncogene.

[21]  R. Kucherlapati,et al.  Somatic Apc mutations are selected upon their capacity to inactivate the β‐catenin downregulating activity , 2000, Genes, chromosomes & cancer.

[22]  Reynaldo Sequerra,et al.  High-efficiency deleter mice show that FLPe is an alternative to Cre-loxP , 2000, Nature Genetics.

[23]  C. Boland,et al.  Mismatch repair proficiency and in vitro response to 5-fluorouracil. , 1999, Gastroenterology.

[24]  S Srivastava,et al.  A National Cancer Institute Workshop on Microsatellite Instability for cancer detection and familial predisposition: development of international criteria for the determination of microsatellite instability in colorectal cancer. , 1998, Cancer research.

[25]  R. Scott,et al.  Inhibition of nonsense-mediated messenger RNA decay in clinical samples facilitates detection of human MSH2 mutations with an in vivo fusion protein assay and conventional techniques. , 1997, Cancer research.

[26]  P. Karran,et al.  Selective recognition of a cisplatin-DNA adduct by human mismatch repair proteins. , 1997, Nucleic acids research.

[27]  T. Mak,et al.  MSH2 deficiency contributes to accelerated APC-mediated intestinal tumorigenesis. , 1996, Cancer research.

[28]  F. Alt,et al.  Efficient in vivo manipulation of mouse genomic sequences at the zygote stage. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[29]  G. Marsischky,et al.  Redundancy of Saccharomyces cerevisiae MSH3 and MSH6 in MSH2-dependent mismatch repair. , 1996, Genes & development.

[30]  H. Griesser,et al.  MSH2 deficient mice are viable and susceptible to lymphoid tumours , 1995, Nature Genetics.

[31]  P. Stanley,et al.  WW6: an embryonic stem cell line with an inert genetic marker that can be traced in chimeras. , 1995, Proceedings of the National Academy of Sciences of the United States of America.

[32]  M. Radman,et al.  Inactivation of the mouse Msh2 gene results in mismatch repair deficiency, methylation tolerance, hyperrecombination, and predisposition to cancer , 1995, Cell.

[33]  T. Kunkel,et al.  Measurement of Heteroduplex Repair in Human Cell Extracts , 1995 .

[34]  Eric S. Lander,et al.  A genetic map of the mouse with 4,006 simple sequence length polymorphisms , 1994, Nature Genetics.

[35]  L. Aaltonen,et al.  Genomic instability in colorectal cancer: relationship to clinicopathological variables and family history. , 1993, Cancer research.

[36]  N. Copeland,et al.  The human mutator gene homolog MSH2 and its association with hereditary nonpolyposis colon cancer. , 1993, Cell.

[37]  T. Kunkel,et al.  Heteroduplex repair in extracts of human HeLa cells. , 1991, The Journal of biological chemistry.