Molecular genetics of hereditary non-polyposis colorectal cancer

Abstract

Colorectal cancer (CRC) is one of the most prevalent malignancies in the Western World and one of the most predominant causes of death by cancer. There is a subgroup of syndromes with a high incidence of CRC which is transmitted in an autosomal dominant fashion. The most common of these syndromes is hereditary non-polyposis colorectal cancer (HNPCC) which has an estimated incidence of 5%. Families with HNPCC are also at risk of developing extracolonic tumors of the endometrium, stomach, ovary, small intestine, kidney and ureter.

To identify genes predisposing to HNPCC, linkage studies were performed using RFLP marker from all chromosomes and microsatellite markers chromosome 3p. Linkage was found in one Swedish family with the marker D3S1029 on chromosome 3p (paper I). The calculated lod score for the particular family was 3.01. We also observed that tumor DNA tested with microsatellite markers frequently showed gain of extra alleles, not present in the constitutional DNA. The instability phenomenon, later called RER, for replication error, gave a clue to the function of the localized gene (paper I). The candidate locus on the short arm of chromosome 3 was further sublocalized using 19 microsatellite markers. The markers were mapped in relation to each other by the use of rodent cell hybrids containing the human chromosome 3 with various deletions. Using two families linked to the 3p locus for haplotype analysis, the putative disease gene location was mapped to 3p21.3-23 (paper II).

The hMLHl gene, involved in DNA mismatch repair, was located to chromosome 3p and identified to cause HNPCC. Denaturing gradient gel electrophoresis (DGGE) was used to screen 39 Swedish HNPCC families for disease causing mutations in the hMLHl gene. Eight germline mutations and 3 polymorphisms were identified. Two of the mutations were splice mutations resulting in a deletion of an exon in the hMLH1 transcript and 6 were missense mutations, changing one amino acid for another in the protein (paper III). The same set of 39 families, as well as 22 new families, were tested for the most common Finnish founder mutation. Three families, all of Finnish origin, were shown to have this large genomic deletion of hMLHl (paper IV).

Colorectal tumors from patients with HNPCC were shown to have the same frequency of APC and K-RAS mutations as sporadic CRC. RER was shown to be very frequent, 96%, and TGFB type II receptor (T,BRII) mutations occur in 56% of the CRC. Tumors from patients with known hMLHl gerrnline mutations had lost one allele, indicating that inactivation of both alleles is necessary for defective mismatch repair (paper V). Extracolonic cancers, breast and other tumors, from HNPCC also display a high frequency (69%) of RER, suggesting that these cancers belong to the HNPCC syndrome. One woman with breast cancer, has a constitutional missense mutation in exon 2 of the hMLH1 gene and also a missense mutation in exon 12, which was inherited from her father who also developed breast cancer. To further study this genotype-phenotype correlation we screened 218 breast cancer families for mutations in exon 12 of hMLHl. However none of the families presented such a mutation (paper VI).