T. C. Hsu

Genetic susceptibility to head and neck cancer : Interaction between nutrition and mutagen sensitivity

The development of head and neck cancer may depend not only on exposure to environmental carcinogens but also on a genetically based susceptibility to carcinogen‐induced damage. This thesis presents a case‐control study that demonstrates the significance of mutagen sensitivity, a measure of an individuals intrinsic DNA repair capacity against free radical damage, as a risk factor for the disease. As part of the case‐control analysis, 167 previously untreated patients and 177 age‐ and sex‐matched healthy controls were assessed for various lifestyle factors including tobacco and alcohol habits, occupational exposures, and diet. Mutagen sensitivity expressed by each individual was determined by quantifying bleomycin‐induced chromosomal breaks within peripheral blood lymphocytes in vitro. Consistent with our initial observations and those of others, mutagen hypersensitivity was strongly associated with increased risk of head and neck cancer (odds ratio, 4.95; 95% confidence interval, 2.67 to 9.17) after adjusting for age, sex, and race. Low intake of vitamins C and E was also associated with an increased risk of disease and was interactive with mutagen sensitivity in risk estimates. Individuals with both a low intake of various antioxidants and increased chromosomal sensitivity to oxidant‐induced DNA damage were at greatest risk. This study supports the concept that the risk of head and neck cancer is determined by a balance of factors that either enhance or protect against free radical oxygen damage, including innate capacities for DNA repair.

Family history of cancer, mutagen sensitivity, and increased risk of head and neck cancer

To evaluate individual cancer susceptibility, 170 previously untreated patients with pathologically-confirmed squamous cell carcinoma of the oral cavity, pharynx, and larynx, and 175 age- and sex-matched health controls were investigated for the occurrence of cancer in first-degree relatives along with other established risk factors for head and neck cancer. More than 54% of these subjects were assayed for mutagen sensitivity by quantifying in-vitro bleomycin-induced chromosomal breaks within peripheral blood lymphocytes. After adjusting for age, gender, education, family income, tobacco and alcohol consumption, the odds ratio associated with three or more first-degree relatives with cancer at any site was 3.79 (95% CI 0.9-15.9) with a linearly-increased trend in risk (P = 0.040). Significantly elevated risk was found to be associated with a history of cancer within siblings (OR = 2.61, 1.2-5.6, P = 0.014). Patients with a family cancer history and mutagen sensitivity were at greatest risk (OR = 7.88, 2.5-25.3, P = 0.005), indicating an additive interactive effect. The findings suggested that genetic familial influence is important in the causation of head and neck cancer.

Mitosis-arresting effects of cigarette smoke condensate on human lymphoid cell lines

Whole-smoke condensates from the University of Kentucky reference cigarettes induced partial mitotic arrest in 4 human lymphoid cell lines. Treatment of cells for 3 h with 100 and 200 micrograms of cigarette-smoke condensate/ml of culture medium increased the frequency of metaphases and decreased the proportion of anaphases in the treated cell populations. Cytoskeletal studies using antitubulin immunofluorescence techniques and transmission electron microscopic studies demonstrated that in early stages of mitosis the formation of aster and the separation of centrosomes in condensate-treated cells were comparable to those of untreated control cells, but the poleward migration of centrosomes was inhibited. Arrested metaphases revealed two centrosomes surrounded by aggregated chromosomes in the center of each cell but the structure of the centrioles, microtubules and the kinetochores appeared normal. The results demonstrate the presence of antimitotic compounds in the tobacco-smoke condensate.

Heterochromatin and interstitial telomeric DNA homology

Abstract. The purpose of this investigation was twofold. The first objective was to demonstrate that, in most of ten mammalian species commonly used in biomedical research, not all constitutive heterochromatin (C-bands) represents telomeric DNA. For example, the C-bands in human chromosomes, the long arm of the X and the entire Y chromosome of Chinese hamster, and most of the short arms of Peromyscus and Syrian hamster chromosomes are not telomeric DNA. In addition to the usual terminal telomeric DNA in the chromosomes of these mammalian species, the pericentromeric regions of seven or eight Syrian hamster chromosomes and all Chinese hamster chromosomes except pair one have pericentromeric regions that hybridize with telomeric DNA, some in C-bands and some not. The second objective was to describe a simple fluorescence in situ hybridization (FISH) reverse-printing procedure to produce black-and-white microphotographs of metaphase and interphase cells showing locations of telomeric DNA with no loss of resolution. Thus, at least three different types of heterochromatin (telomeric heterochromatin, nontelomeric heterochromatin and a combination of both) are present in these mammalian species, and this simple black-and-white reverse printing of telomeric FISH preparations can depict them economically without sacrificing clarity.

Effects of N-acetyl-l-cysteine and ascorbic acid on mutagen-induced chromosomal sensitivity in patients with head and neck cancers

The protective effect of N-acetyl-L-cysteine (NAC) and ascorbic acid on mutagen-induced chromosomal breakage was determined using human lymphoblastoid cell lines as well as freshly cultured lymphocytes from patients with head and neck malignancies and healthy control subjects. Mutagen sensitivity was determined using the previously described bleomycin exposure assay. The toxicities of different concentrations of NAC and ascorbic acid, as well as both the preincubation and dose-dependent protective effects of these two agents, were analyzed. Both test drugs proved to be effective in diminishing mutagen-induced chromatid breakage in established lymphocyte cell lines. In freshly cultured lymphocytes, NAC given in doses ranging from 0.1 to 10 mmol/L decreased the number of mutagen-induced breaks per cell in a range from 23% to 73%, and ascorbic acid decreased chromosomal breakage by 21% to 58% in a dose range from 0.01 to 1 mmol/L. The results of this study demonstrate the protective effect mediated in vitro by both NAC and ascorbic acid against mutagen-induced chromosomal damage. A similar in vivo phenomenon may explain the differences in occurrence of head and neck cancer between populations with different dietary backgrounds.

Mutagen sensitivity in humans: A comparison between two nomenclature systems for recording chromatid breaks

Currently, there are two systems in use for recording chromatid breaks, either spontaneously occurring or induced by mutagens: The International System for Human Cytogenetic Nomenclature (ISCN) and the Chatham Barrs Inn Conference (CBIC) recommendation. The former system considers that a chromatid break is recognized only when the chromatid fragment is displaced to the other side of its sister chromatid, while all others, regardless of the distance between the two broken ends, are called chromatid gaps. The CBIC system recognizes a chromatid break when the intervening achromatic segment is longer than or equal to the diameter of the chromatid, whether the fragment is displaced or not. Minor lesions are called chromatid gaps. We conducted experiments using bleomycin treatment of human cells (primary cultures or lymphoblastoid cell lines) and read the chromatid lesions both ways. We conclude that the CBIC system appears to have more direct biologic relevance than the ISCN system.

Three Measures of Mutagen Sensitivity in a Cancer-Free Population

Different individuals appear to respond differently to the same carcinogen, and different mutagens act differently on cells. We conducted mutagen sensitivity assays by using three mutagens (bleomycin, a radiomimetic agent; 4-nitroquinoline-1-oxide [4-NQO], an ultraviolet light mimetic agent; and benzo[a]pyrene diol epoxide [BPDE], a tobacco mutagen) in parallel in healthy human subjects to determine the relationships among these assays. Our results showed that the mean breaks per cell values (b/c) (+/- SD) for bleomycin, 4-NQO, and BPDE sensitivity were 0.49 (+/- 0.26), 0.53 (+/- 0.30), and 0.66 (+/- 0.41), respectively. Age, sex, smoking status, and family history of cancer were not correlated with any of these mutagen sensitivities. Also, there was no correlation between bleomycin and 4-NQO or 4-NQO and BPDE sensitivity, but a weak correlation between bleomycin and BPDE was observed (correlation coefficient = 0.289; P = 0.001). When the 75th percentile of b/c was used as a cutoff point in each assay, only one individual (1.8%) was sensitive to all three mutagens. Ten individuals (17.9%) were sensitive to two mutagens, 20 (35.7%) to one mutagen, and 25 (44.6%) to none of three mutagens. Our study suggests that these three mutagens may involve different DNA damage and repair pathways. The lack of correlation between the assay results may indicate the necessity of using a battery of mutagen sensitivity tests to refine our ability to identify a subpopulation at high cancer risk.

MUTAGEN SENSITIVITY AS A MARKER OF CANCER SUSCEPTIBILITY

Modulation of environmental exposures by host genetic factors may explain interindividual variation in susceptibility to carcinogenesis. One determinant of susceptibility is mutagen sensitivity measured by the frequency of bleomycin‐induced breaks in an in vitro lymphocyte assay. Mutagen sensitivity is a significant predictor of aerodigestive tract cancer risk. In this case‐control study of lung‐cancer susceptibility markers, 54% of 132 lung‐cancer cases had mutagen‐sensitivity scores greater than or equal to 1 break/cell, compared with only 22% of 232 controls. The mean breaks/cell value (±SE) for the 88 African‐American cases was 1.11 (±0.60), compared with 0.82 (±0.49) for the 121 controls (P < 0.001). For the 44 Mexican‐American cases and 111 controls, the comparable values were 1.11 (±0.52) and 0.76 (±0.38), respectively. The overall odds ratio (OR) for mutagen sensitivity (dichotomized at ≥ 1 break/cell), after adjusting for ethnicity and smoking status, was 3.62 (95% confidence limits [CL] = 2.2, 5.9). For current smokers the adjusted risk associated with mutagen sensitivity was 2.52 (1.2, 5.3). For former smokers, the comparable OR (95% CL) was 6.19 (2.7, 14.1). The risk estimate for those under 61 years of age was 4.85 (2.3, 10.4), compared with 2.85 (1.5, 5.6) for older subjects. The risk also appeared to be higher for lighter smokers (<20 cigarettes daily) than heavier smokers (ORs = 5.72 and 3.20, respectively). The ethnicity‐adjusted ORs by quartile of breaks/cell were 1.0, 1.40, 2.46, and 4.80; the trend test was significant at P < 0.001. The joint effects of mutagen sensitivity and former smoking, current smoking, or heavy smoking were greater than additive, although the interaction terms were not statistically significant in the logistic model. Mutagen sensitivity may therefore be a useful member of a panel of susceptibility markers for defining high‐risk subgroups for chemoprevention trials. J. Cell. Biochem: 25S:80–84.

A Short-Term Cytogenetic Test for Genetic Instability in Humans

It has been postulated by various investigators that 70 to 90% of all human cancers can be attributed to environmental causes (Boyland, 1969; Higginson, 1976). These environmental causes may be due to the agents that persons are exposed to in the atmosphere or agents that are ingested. However, not all people exposed to the similar environment develop cancer. Genetic factors may be responsible for some of the variations in susceptibility from individual to individual. The genetic factors may be (1) the ability of an individual to metabolize (activate or inactivate) mutagenic agents as well as promoters, cocarcinogens etc.; (2) the ability of an individual to repair the induced damage; and (3) the immunocompetence of an individual to recognize the potential cancer cells.

Comparative efficacy as antioxidants between ascorbic acid and epigallocatechin gallate on cells of two human lymphoblastoid lines

Both ascorbic acid and epigallocatechin gallate, which is one of the key polyphenols contained in green tea leaves, have been considered excellent antioxidants. The present study compared the efficacy as antioxidants between the two agents on an equimolar basis. Cells of two lymphoid lines were used as test material to determine the reduction of chromosome damage induced by the radiomimetic antibiotic bleomycin. Without bleomycin, both agents, at concentrations of 10(-7), 10(-6), 10(-5), and 10(-4) M, showed chromosome damage similar to the untreated controls. With bleomycin, the weakest concentration of both showed no protective effect. At concentrations of 10(-6) and 10(-5) M, especially the latter, a significant reduction in frequencies of chromatid breaks was recorded. However, at the highest concentration, 10(-4) M, the chromatid break frequencies rose to the same level as that of cells treated with bleomycin alone, suggesting that both behaved like pro-oxidants.