Although the Ki-ras gene is frequently mutated in human colon cancer, the Ha-ras protooncogene is maximally expressed in the most differentiated cells of intestinal mucosa.
A mutation in c-K-ras (KRAS2) has long been implicated as one of the important early events in the development of a large proportion of human colon cancers.
Linkage analysis shows that these susceptibility genes are different from the mouse homologs of the genes known to be somatically mutated in human colon cancer (KRAS2, TP53, DCC, MCC, APC, MSH2, and probably also MLH1).
These results, together with our previous observations of a relative lack of Ki-ras gene mutations in the same tumors, are similar to those found in human UC-associated colon cancer, suggest a common pathway in these two systems, although they are different in their implication of p53 mutations.
A point mutation at codon 12 of the K-ras gene was found, while no alterations were noted in the p53 gene, whose mutations are frequent in colon cancers.
We used a hammerhead ribozyme (KrasRz) against mutated K-ras gene transcripts (codon 12, GTT) to inactivate mutant K-ras function in the colon cancer cell line SW480, harbouring a mutant K-ras gene.
To develop a new therapy by regulating the activated K-ras gene in colon cancers, we prepared synthetic DNA encoding the ribozyme (catalytic RNA), and inserted it into an expression vector (LNCX) originated from a retrovirus.
KRAS exon 1 sequences from DNA isolated from 191 sporadic colon cancers were PCR amplified using one primer labeled with fluorescein and a second primer extended by a GC-clamp.
Previous reports in the literature have shown a relationship between different KRAS mutations and prognosis and response to medical treatment in colon cancer patients.
In addition, Pyrosequencing proved superior to dideoxy sequencing in the detection of KRAS mutations from DNA mixtures of paraffin-embedded colon cancer and normal tissue as well as from paraffin-embedded pancreatic cancers.
We conclude that polymorphisms in some insulin-related genes are associated with an increased risk of colon cancer with specific KRAS2 and TP53 mutations, implying a link between these genetic changes and specific mutational pathways in carcinogenesis.