MUTATION ANALYSIS IN MOUSE MODELS OF COLON CANCER

The tumor suppressor gene Apc is mutated in 85% of human colorectal cancers and loss of function of the Apc protein is thought to be an initiating event in tumor formation.   A variety of mouse strains have been made that recapitulate the effects of Apc loss, including the Apc1638N strain made by Dr. Raju Kucherlapati and colleagues at Albert Einstein College of Medicine (1).  These mice are heterozygous for a mutant form of Apc and are susceptible to spontaneous intestinal tumor formation if the remaining ‘wild-type’ copy of Apc in cells is inactivated by mutation.  Such mice provide powerful experimental models for investigating the genetic and environmental risk factors that increase the incidence of colorectal cancer.  One of the known genetic risk factors is an inherited defect in a gene required for DNA mismatch repair (MMR), a crucial repair mechanism that normally corrects errors in DNA sequences and so reduces the risk of mutation.  Inherited defects in MMR are responsible for the familial colon cancer syndrome known as HNPCC (Hereditary Non-Polyposis Colon Cancer) and in many cases this is due to mutations in the DNA repair gene Mlh1. 

Kucherlapati, together with Dr. Winfried Edelman at Einstein and colleagues at Strang Cancer Prevention Center, has made mouse models of HNPCC by ‘knocking out’ DNA repair genes such as Mlh1.  When crossed with the cancer-prone Apc1638N strain, these Mlh1-deficient mice showed a more than 8-fold increase in tumor incidence (2).  The mechanism underlying this effect has been investigated by Dr. Mari Kuraguchi, working with Dr. Anthony Brown of Strang Cancer Prevention Center and Weill Medical College of Cornell University.  Specifically, Dr. Kuraguchi has asked whether Apc in these mice is a primary target for mutations induced by the MMR deficiency.  Using an assay for truncation mutations in Apc based on PCR amplification of tumor DNA, followed by in vitro translation of the cloned DNA fragments, she has found that 84% of the intestinal tumors in the Apc1638N Mlh1^-/- harbor truncation mutations in Apc.  Moreover, sequencing of these mutations reveals a characteristic mutational ‘signature’ of Mlh1 deficiency, in the form of particular base substitutions and frameshift mutations in short DNA repeats (3).  These results indicate that the increase in tumor formation due to MMR deficiency in these mice is attributable to an increased rate of mutations in the Apc gene.  Dr. Kuraguchi has also been characterizing Apc mutations in mice defective for two other DNA repair genes, Msh3 and Msh6.  Analyses of this sort help in recognizing the underlying mutational mechanisms responsible for DNA defects in particular cancers and may be useful in diagnosing specific types of HNPCC.  As animal models of HNPCC, these mouse strains also offer great potential for evaluating prevention strategies that may reduce the incidence of colon cancer in human patients with genetic predisposition to the disease.  This may lead to improved methods of diminishing the risk of colon cancer.

1.  Fodde, R., Edelmann, W., Yang, K., van Leeuwen, C., Carlson, C., Renault, B., Breukel, C., Alt, E., Lipkin, M., Khan, P. M., and Kucherlapati, R.  A targeted chain-termination mutation in the mouse Apc gene results in multiple intestinal tumors. Proc Natl Acad Sci U S A, 91: 8969-73, 1994.

2.  Edelmann, W., Yang, K., Kuraguchi, M., Heyer, J., Lia, M., Kneitz, B., Fan, K., Brown, A. M. C., Lipkin, M., and Kucherlapati, R. Tumorigenesis in Mlh1 and Mlh1/Apc1638N mutant mice. Cancer Res, 59: 1301-7, 1999.

3.  Kuraguchi, M., Edelmann, W., Yang, K., Lipkin, M., Kucherlapati, R., and Brown, A. M. C. Tumor-associated Apc mutations in Mlh1^-/- Apc1638N mice reveal a mutational signature of Mlh1 deficiency.  Oncogene, 19: 5755-5763, 2000.

Mari Kuraguchi, Ph.D., is a postdoctoral scientist at Strang Cancer Prevention Center and recipient of a Fellowship in Cancer Prevention from the American Association of Cancer Research.  Anthony M.C. Brown, Ph.D., is Associate Director of Research at Strang Cancer Prevention Center and Associate Professor of Cell Biology at Weill Medical College of Cornell University.