Lie Cheng

Glutathione-S-transferase polymorphisms and risk of squamous-cell carcinoma of the head and neck

Differences in genetic susceptibility to tobacco‐induced carcinogenesis appear to modulate an individuals risk of squamous‐cell carcinoma of the head and neck (SCCHN). Risk for SCCHN may be associated with the null alleles of the carcinogen‐metabolizing genes glutathione‐S‐transferase (GST) T1 and GSTM1. In this study, we evaluated the association between GSTM1 and GSTT1 null genotypes and risk of SCCHN in a matched case‐control study of 162 patients with SCCHN and 315 healthy controls. Our results showed that 53.1% of cases and 42.9% of controls were null for GSTM1, whereas 32.7% of cases and 17.5% of controls were null for GSTT1 (p < 0.05 and p < 0.001, respectively). Furthermore, 19.8% of cases but only 7.9% of controls were null for both genes (p < 0.001). Multivariate analysis using logistic regression models, including age, sex, ethnicity, smoking status, alcohol status and GST genotypes, showed that both of these genotypes remained independent risk factors for disease [adjusted odds ratios (ORs) = 1.50 and 2.27, respectively; 95% confidence intervals (CIs) = 1.01–2.23 and 1.43–3.60, respectively). When the genotypes were divided into neither null, either null or both null, there was a dose‐response relationship (adjusted OR = 1.50, 95% CI = 0.98–2.30) for the either‐null group and (adjusted OR = 3.64, 95% CI = 1.94–6.84) for the both‐null group (p < 0.001, trend test). Our findings suggest that the GSTM1 and GSTT1 null genotypes are independent risk factors for SCCHN and markers for genetic susceptibility to tobacco‐induced carcinogenesis. Int. J. Cancer (Pred. Oncol.) 84:220–224, 1999.

Fas A670G polymorphism, apoptotic capacity in lymphocyte cultures, and risk of lung cancer

Tobacco carcinogens can damage DNA, leading to apoptosis. There may be individual variation in apoptotic capacity (AC), and this variation may explain difference in AC associated with risk of lung cancer, if genome integrity is not restored by efficient DNA repair. To test the hypothesis that genetically determined AC is associated with risk of lung cancer, we conducted a pilot case-control study of 68 patients with newly diagnosed, untreated lung cancer and 74 cancer-free controls. We measured the AC of their cultured peripheral blood lymphocytes in response to in vitro exposure to an ultimate tobacco carcinogen, benzo[a]pyrene diol epoxide (BPDE), by using terminal dUTP nucleotide end labeling and flow cytometry. We also investigated the frequency of the -A670G polymorphism in Fas, a gene involved in controlling the apoptotic pathway, by using polymerase chain reaction-restriction fragment length polymorphism analysis. After exposing the cells to 4 microM BPDE for 5 h, we observed a significantly lower AC in lung cancer patients (155.2+/-143.9%) than in the controls (216.6+/-184.6%) (P<0.05). Low AC was an independent risk factor (adjusted odds ratio (OR)=2.69, 95% confidence interval (CI)=1.18-6.15) for lung cancer after adjustment for age, sex, ethnicity, smoking status and apoptotic baseline in a logistic regression model. Although the Fas -A670G polymorphism was not an independent risk factor for lung cancer, it appeared to modulate the risk. The adjusted ORs for lung cancer risk associated with lower AC were 4.00 (95% CI=1.48-10.80) among those with the Fas -670 AG and GG genotypes and 0.97 (95% CI=0.18-5.30) among those with the Fas -670AA genotype. These data suggest that alteration in the apoptotic pathway may be a risk factor for lung cancer and this risk may be modulated by the Fas -A670G polymorphism. Larger prospective studies are needed to verify these findings.

DNA Repair Capacity in Healthy Medical Students During and After Exam Stress

There has been extensive research into the effects of stress on immune function but little on the effects of stress on DNA repair capacity (DRC), a process central to maintaining a normal cell cycle. Defective DRC is one of the factors responsible for carcinogenesis. In the present study we assessed DRC in healthy medical students during times of high and low stress. Sixteen medical students were evaluated during the third day of a 5-day exam period and then again 3 weeks later, after vacation. At both time points, participants underwent a brief physical examination, had venous blood drawn, and completed questionnaires to identify subjective stress levels. The DRC was assessed by the host-cell reaction assay, which measures nucleotide excision repair capacity. Participants reported significantly higher levels of subjective stress during the exam period than after vacation. DRC was also significantly higher during the exam period than after vacation, suggesting a positive association between subject stress levels and DRC. The results are discussed in relation to previous findings and implications for cancer research.

EXPRESSION OF FIVE SELECTED HUMAN MISMATCH REPAIR GENES SIMULTANEOUSLY DETECTED IN NORMAL AND CANCER CELL LINES BY A NONRADIOACTIVE MULTIPLEX REVERSE TRANSCRIPTION-POLYMERASE CHAIN REACTION

Abnormalities in at least 1 of 5 mismatch repair (MMR) genes (hMSH2, hMLH1, hPMS1, hPMS2 and GTBP/hMSH6) are found in hereditary nonpolyposis colon cancer and sporadic colon cancers. We used a single-reaction multiplex reverse transcription (RT)-polymerase chain reaction (PCR), with the beta-actin gene as an internal control, to simultaneously evaluate expression of these 5 known human MMR genes in normal and tumor cell lines with known or uncharacterized mutations in MMR genes. The relative quantitation of the transcripts is demonstrated by controlling the number of PCR cycles and titrating cDNA with a dose-curve. The 13 normal cell lines tested were derived from normal lymphocytes, skin, thymus, breast, lung, colon, liver and kidney. The 26 cancer cell lines were derived from melanoma and cancers of the brain, breast, lung, colon, pancreas and prostate. All 5 MMR genes were ubiquitously expressed in all normal cell lines tested, suggesting their housekeeping roles. Aberrant MMR gene expression was only observed in the colon cancer cell lines. Two previously uncharacterized colon cancer cell lines did not express hMLH1. These data suggest that this nonradioactive multiplex RT-PCR assay for MMR gene expression may be useful for fast screening for genetic alterations that may affect gene expression and so may aid molecular analysis of MMR-related colon cancer.

Cryopreserving whole blood for functional assays using viable lymphocytes in molecular epidemiology studies

There is an increasing need for viable lymphocytes in performing phenotypic assays for biomarker studies. Both fresh and cryopreserved lymphocytes have been used for cell culture-based functional assays. However, fresh lymphocytes do not allow assays to be done in batches and cryopreservation of isolated lymphocytes results in a considerable loss of viable cells. To investigate the feasibility of using cryopreserved whole blood as a source of viable lymphocytes in molecular epidemiology studies, two well-established biomarkers, the host-cell reactivation (HCR) and mutagen sensitivity assays, were used to compare the method of cryopreserving whole blood with the traditional methods. In 25 paired blood samples assayed for DNA repair capacity (DRC) by the HCR assay, the DRC values of frozen whole blood (mean +/- SD, 11.59 +/- 3.07) were similar to those of frozen isolated lymphocytes (11.08 +/- 3.50). The correlation between the paired DRC values was 0.77 (P < 0.001). In 31 paired blood samples assayed for the gamma-radiation-induced chromatid breaks by the mutagen sensitivity assay, there was no significant difference between the baseline level of chromatid breaks in lymphocytes from frozen blood (0.05 +/- 0.03) and fresh blood (0.06 +/- 0.03). The blastogenic rate and mitotic index of the cells used for the two assays were compared between the different processing methods. The lymphocytes from frozen whole blood were more sensitive to gamma-radiation, with a higher mean level of chromatid breaks (0.68 +/- 0.21) than that in fresh blood (0.42 +/- 0.12, P < 0.01), and the correlation between the numbers of chromatid breaks in the paired samples was statistically significant (r = 0.61, P < 0.001). These data suggest that within the limits of the parameters investigated here, cryopreserved whole blood is a good source of viable lymphocytes for biomarker assays in molecular epidemiological studies.