Influence of genetic polymorphisms on DNA repair, p53 mutations and cancer risk

Individuals have different susceptibility towards environmental exposures that may cause or contribute to disease. This inter-individual variation is partly due to genetic polymorphisms in genes involved in DNA repair and metabolism, which have potential to modulate the function of the encoded proteins.

This thesis is a contribution to the knowledge about the importance of polymorphisms, and how they might interfere with health. More specifically, the aims have been to investigate (1) if and how the mutational spectra of the tumour suppressor gene p53 in lung and bladder tumours is affected by genetic polymorphisms, (2) if polymorphisms can influence lung cancer risk in a study population with a high frequency of women and non-smokers, and (3) to develop a method to study the functionality of DNA repair polymorphisms in vitro.

Papers I-II are based on a lung cancer case-control study population, with approximately 50 % nonsmokers and 70 % women. Papers III-IV are based on a bladder cancer cohort that includes all bladder cancer cases in the Stockholm county between 1995-1996. In paper V, lymphocytes from healthy nonsmoking volunteers were collected to study the functionality of DNA repair polymorphisms, using the Comet assay. PCR-RFLP and Taq-Man allelic discrimination were used for genotyping, SSCP and sequencing for identification of p53 mutations in tumours.

The impact of genetic polymorphisms on the p53 gene was investigated in study I, III and IV. In study I we observed an association between the XPD 751G1n allele and an increased frequency of p53 transversion mutations in lung tumours, especially among smokers, suggesting that XPD 751G1n carriers have a reduced DNA repair proficiency. In study III we found that p53 transversion mutations were more common in bladder tumours of GSTM1 null individuals, and also the GSTP1 105Val allele was associated with a higher frequency of p53 transversion mutations than the wild type allele. GSTP1 105 Val was also associated with a higher frequency of p53 transitions at CpG sites. These results indicated that an impaired glutathione conjugation might affect the p53 mutational spectrum.

In study IV, the cyclin D1 variant allele homozygotes and the XRCC3 241 Met homozygotes were associated with an increased p53 mutation frequency, while the NQ01 187Ser and 139Trp alleles and the XPC 939GIn allele were associated with an increased frequency of p53 transversion mutations. Thus, in general, these polymorphisms seem to have an impact on the type of mutations in the p53 gene in lung- and bladdertumours, while their impact on the frequency of mutation is small. This increased risk for specific types of mutations probably reflects a higher susceptibility to environmental exposures following the impaired metabolism (glutathione conjugation), or less efficient DNA repair, among carriers of the susceptible alleles.

The influence of polymorphisms on lung cancer risk was investigated in study II, where the XRCC1 399GIn and the XRCC3 241 Met alleles were found to be associated with decreased risk for lung cancer among non-smoking- and smoking- women respectively. The NBS1 185GIn allele was associated with an increased risk among non-smoking and low-dose smoking women. The effects of the two HR-proteins, XRCC3 and NBS1 enhanced each other. Our results suggest that these polymorphisms influence the cancer risk through their effect on the repair capacity, which may alter the risk of having mutations fixed in the genome.

In study V, we used a modified comet assay to study the influence of DNA repair polymorphisms on repair of MMS-induced DNA damage in peripheral blood lymphocytes in vitro. The investigated APEX1 Asp 148Glu, XRCC3 Thr241Met and NBS1 Glu 185Gln polymorphisms were shown to have effect on the early stages of DNA repair, and were associated with the rate of transitions of cells from a more damaged to a less damaged state. This type of functional study of DNA repair enzyme polymorphisms will contribute to a better understanding of the significance of gene association studies in cancer research.

List of scientific papers

I. Hou SM, Ryk C, Kannio A, Angelini S, Falt S, Nyberg F, Husgafvel-Pursiainen K (2003). Influence of common XPD and XRCC1 variant alleles on p53 mutations in lung tumors. Environ Mol Mutagen. 41(1): 37-42.
https://doi.org/10.1002/em.10128

II. Ryk C, Kumar R, Thirumaran KR, Hou S (2006). Polymorphisms in the DNA repair genes XRCC1, APEX1, XRCC3 and NBS1, and the risk for Lung Cancer in never- and ever-smokers. Lung Cancer. [Accepted]
https://doi.org/10.1016/j.lungcan.2006.08.004

III. Ryk C, Berggren P, Kumar R, Hemminki K, Larsson P, Steineck G, Lambert B, Hou SM (2005). Influence of GSTM1, GSTT1, GSTP1 and NAT2 genotypes on the p53 mutational spectrum in bladder tumours. Int J Cancer. 113(5): 761-8.
https://doi.org/10.1002/ijc.20650

IV. Ryk C, Kumar R, Sanyal S, Verdier PJ, Hemminki K, Larsson P, Steineck G, Hou SM (2005). Influence of polymorphism in DNA repair and defence genes on p53 mutations in bladder tumours. Cancer Lett. [Accepted]
https://doi.org/10.1016/j.canlet.2005.10.025

V. Ryk C, Routledge MN, Allan JM, Wild CP, Kumar R, Lambert B, Hou S (2006). Influence of polymorphisms in DNA repair genes on repair kinetics, as measured by the Comet assay. [Submitted]