[Proc Amer Assoc Cancer Res, Volume 47, 2006]
Tumor Biology 33: Genetically Engineered Animal Models for Human Cancers
A rat knock out model of human familial adenomatous polyposis
Michael Gould and
University of Wisconsin, Madison, WI
Humans with mutations in the Adenomatous Polyposis Coli (APC) gene get hundreds to thousands of adenomas in the colon and develop colorectal cancer, however current mouse models of human intestinal cancer, including the ApcMin model developed at the McArdle Laboratory, develop lesions primarily in the small intestine and have never been observed to metastasize. We have generated a rat knock out of Apc in which the animals develop primarily colonic tumors by 90 days of age. We have termed this allele Pirc for polyposis in rat colon. Our group has recently developed the technology to knock out rat genes of interest through mutagenesis with N-ethyl-N-nitrosourea (ENU). We screened 2530 base pairs of exon 15 of the rat Apc gene from genomic DNA in the offspring of mutagenized Fisher-344 inbred rats using a universal vector gap repair yeast truncation assay (Biotechniques. 2004 Sep;37(3):383-8). Screening of 1360 progeny yielded a single mutant with an A to T transversion changing a lysine to a stop codon at amino acid position 1137. Upon abstract submission the founder male is 260 days old, remains fertile, and has successfully transmitted the mutation to 60 progeny. Ten of the resulting mutant N1 animals were brother-sister mated and their offspring confirmed that the ApcPirc allele is homozygous lethal. Necropsy of six N1 ApcPirc heterozygotes, three at 90 days and three at 130 days of age, identified an average of 6 visible adenomatous polyps in the colon, while six wild type age-matched littermates were tumor-free. Histological immunofluorescence staining of four of the tumors for ß-catenin confirmed the tumor-associated relocalization of ß-catenin from the lateral membrane to the cytoplasm and nucleus. The affected animals showed neither visible lesions nor ß-catenin relocalization in the small intestine. Pirc/+ animals treated somatically with ENU at 2 weeks of age develop in excess of 100 microadenomas in the distal small intestine and colon by 60 days post injection. No lesions were seen in the ENU treated non-carriers. ApcPirc was successfully crossed to Wistar-Furth and intercrossing of the F1 carriers determined that the ApcPirc allele was homozygous lethal in the F2 progeny. Ongoing studies include the examination of tumor number and distribution in older carrier N1 and F1 animals as well as the effects of a high fat diet on N1 animals. The primary localization of tumors to the colon recapitulates human intestinal cancer, addressing one of the major drawbacks of ApcMin and other mouse models that have a primarily small-intestinal distribution. The colonic phenotype and longer life span of the ApcPirc rat combined with the growing resources of rat genetics, genomics and biology indicate that the rat may be an advantageous model for human tumor progression and experimental treatment of colon cancer.
Copyright © 2006 by the American Association for Cancer Research.