Polymorphisms and probable lack of mutation in the WAF1-CIP1 gene in colorectal cancer
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Y J Li1, P Laurent-Puig, R J Salmon
1Laboratoire de Génétique des Tumeurs, Institut Curie, Paris, France.
Abstract:
WAF1/CIP1, a gene up-regulated by p53 encodes an inhibitor of cyclin-dependent kinases. Induction of WAF1/CIP1 in cells with intact p53 is believed to be instrumental in cell cycle arrest and apoptosis caused by DNA damage. In a model system, WAF1/CIP1 has been shown to have tumor suppressive activity. It is not known however whether WAF1/CIP1 is mutated in human primary tumors. Cells from colorectal cancer have been shown to acquire a series of genetic alterations, including frequent p53 mutations. Thus colorectal tumors, particularly those without identified p53 mutations, are good candidate to search for putative WAF1/CIP1 mutations. DNA extracted from 45 tumors, (including 28 tumors for which p53 mutations had previously been searched for and not found) were PCR amplified for exon 2 of WAF1/CIP1. A search for point mutations was performed in each amplified product using a denaturing gradient gel electrophoresis (DGGE) technique which enables the efficient screening of codons 9 to 139 (i.e. 80% of the WAF1/CIP1 coding sequence). Two different DNA variants were identified and shown to be present in constitutional DNAs of the corresponding patients. The first variant, a C to A transversion at codon 31, changes a serine for an arginine and was detected in eight tumors (18% of the cases). The second variant, detected in a single case (2%) is a silent A to T transversion at the third base of codon 91. DNA extracted from 70 unrelated members from the Centre d'Etude du Polymorphisme Humain (CEPH) was screened for these polymorphisms. The ser/arg polymorphism of codon 31 was detected in seven cases (10%) thus suggesting that it is not associated with a marked colorectal cancer predisposition. The polymorphism on codon 91 was not detected. Two additional variants (arginine to histidine at position 67 and threonine to methionine at position 80) were observed once each in the CEPH family members. Somatic mutation of the WAF1/CIP1 gene was not observed, indicating that, unless there are hot spots for mutations outside the screened region, this gene is not a frequent site of point mutation in colorectal cancer.
Insights
The WAF1/CIP1 gene, linked to cell cycle control and tumor suppression, was investigated for mutations in colorectal cancers. Researchers found no evidence of somatic mutations in the screened region, suggesting WAF1/CIP1 is not frequently mutated in this cancer type.
Area of Science:
- Molecular Biology
- Cancer Genetics
Background:
- The WAF1/CIP1 gene, regulated by p53, encodes a cyclin-dependent kinase inhibitor crucial for cell cycle arrest and apoptosis.
- WAF1/CIP1 exhibits tumor-suppressive activity in model systems, but its mutation status in human primary tumors remains largely unknown.
- Colorectal cancers frequently accumulate genetic alterations, including p53 mutations, making them a key focus for studying other tumor suppressor genes.
Purpose of the Study:
- To investigate the presence of WAF1/CIP1 gene mutations in primary colorectal tumors.
- To determine if WAF1/CIP1 mutations are associated with colorectal cancer development, particularly in tumors lacking p53 mutations.
Main Methods:
- PCR amplification of exon 2 of the WAF1/CIP1 gene from 45 colorectal tumor DNA samples.
- Denaturing Gradient Gel Electrophoresis (DGGE) was used to screen for point mutations in codons 9-139 (80% of the coding sequence).
- Screening of identified variants in constitutional DNA and in 70 unrelated individuals from the CEPH panel.
Main Results:
- Two DNA variants were identified in tumor DNA, both also present in constitutional DNA, indicating they are polymorphisms.
- A C to A transversion at codon 31 (Ser/Arg) was found in 18% of tumors and 10% of controls, suggesting no strong association with colorectal cancer predisposition.
- A silent A to T transversion at codon 91 was found in 2% of tumors.
- No somatic mutations were detected in the screened region of WAF1/CIP1 in the analyzed colorectal tumors.
Conclusions:
- The WAF1/CIP1 gene does not appear to be a frequent site of point mutation in colorectal cancer within the screened region.
- The identified variants are likely constitutional polymorphisms rather than somatic mutations driving tumorigenesis.
- Further investigation may be warranted for potential mutations in other regions of the WAF1/CIP1 gene.