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While ultraviolet radiation (UV) is critical in the pathogenesis of basal cell carcinomas (BCC), its role in determining the phenotypic variation shown by patients is unknown. Thus, patients manifest variation in BCC numbers, patterns of presentation and tumour site. We have used this diversity to classify patients into subgroups that are associated with different risks of developing tumours. Two phenotypes are particularly interesting. Firstly, presentation with clusters of BCC. These patients, termed multiple presentation phenotype (MPP), had two to five BCC at one presentation, suggesting rapid accrual over short periods of time. They comprised 15% of our 1200 BCC patients. A minority of patients demonstrated multiple clustering events, a phenomenon that is associated with a genetic pre-disposition. The second risk phenotype, characterised by tumours on the trunk, is also associated with a pre-disposition. Both phenotypes were characterised by a susceptibility to develop numerous BCC. Thus, all our patients with more than five BCC had one or both of these phenotypes. We are using a candidate gene approach to identify loci associated with risk of these phenotypes (susceptibility genes) and tumour numbers in them (modifier genes). Interestingly, we did not identify differences in UV exposure between patients with high- and low-risk phenotypes.

Non-melanoma skin cancer (NMSC) represents a significant cause of morbidity and mortality among renal transplant recipients, with tumors behaving more aggressively than those in nontransplant patients. Not all immunosuppressed patients develop NMSC, however, and in those that do, the rate of accrual and numbers of lesions vary considerably. Though ultraviolet light is critical, it is unlikely that this alone explains the observed phenotypic diversity, suggesting the possible involvement of genetic factors. Furthermore, although twin studies in nontransplant patients with NMSC suggest a low genetic component, several genes associated with susceptibility and outcome in these patients have been identified. Thus, having previously shown that polymorphism in members of the glutathione S-transferase (GST) supergene family is associated with altered NMSC risk in nontransplant patients, we examined allelism in GSTM1, GSTP1, GSTM3, and GSTT1 in 183 renal transplant recipients. GSTM1 null was associated with increased squamous cell carcinoma (SCC) risk (p = 0.042, OR = 3.1). This remained significant after correction for age, gender, and ultraviolet light exposure (p = 0.012, OR = 8.4) and was particularly strong in patients with higher ultraviolet light exposure (e.g., sunbathing score > 3, p = 0.003, OR = 11.5) and in smokers (p = 0.021, OR = 4.8). Analysis of the interaction between GSTM1 null and sunbathing score showed that the two factors were synergistic and individuals with both risk parameters demonstrated a shorter time from transplantation to development of the first SCC (p = 0.012, hazard ratio = 7.1). GSTP1*Ile homozygotes developed larger numbers of SCC (p = 0.002, rate ratio = 7.6), particularly those with lower ultraviolet light exposure and cigarette consumption. GSTM3 and GSTT1 also demonstrated significant associations, though some genotype frequencies were low. These preliminary data suggest that genetic factors mediating protection against oxidative stress are important in NMSC development in immunosuppressed patients and may be useful in identifying high-risk individuals.

The reproducibility and acceptability of airways resistance measurements using the interrupter technique (MicroRint) obtained using a mouthpiece were compared with those using a face mask.Fifty children aged 4–7 yrs performed four sets of sixRintmeasurements; two using a mouthpiece and two using a face mask with integral mouthpiece.Complete data were obtained from 45 (90%) children using the mouthpiece and 43 (86%) children using the mask. The two methods were equally repeatable with comparable intraclass correlation coefficients (ICC) and coefficients of variation. MeanRintvalues obtained using the mouthpiece were significantly lower than those using the face mask ((mean±sd) mouthpiece=0.81±0.18 kPa·L−1·s, mask=0.88±0.24 kPa·L−1·s p=0.0002). Although the mean paired differences between the two methods were small (0.07 kPa·L−1·s), the ICC and limits of agreement confirmed that the two methods could not be used interchangeably. Sixty-seven per cent of children preferred the face mask but this was more time-consuming (p=0.03). Children did not produce more repeatable results using their preferred method, nor did they improve with practice.Repeatable airway resistance measurements using the interrupter technique can be obtained from young children using either a mouthpiece or a face mask, but there are significant clinical and statistical differences between the results obtained.

Background Basal cell carcinoma (BCC) patients demonstrate considerable phenotypic diversity. The basis of this heterogeneity is poorly understood. We have shown that presentational phenotypes are associated with BCC numbers. Thus, patients with a cluster of new BCC at any presentation comprise a subgroup, termed MPP, that is at increased risk of developing numerous lesions. Patients with more than one cluster (multiple cluster MPP) are at particular risk. Procedure We determined in a cohort of BCC cases, whether: (i) tumor accrual was altered after clustering; and (ii) multiple cluster MPP is associated with characteristics linked with sensitivity to UV or, GSTT1, GSTM1, GSTM3, GSTP1, MC1R, CYP2D6, TNF-α, and VDR genotypes previously associated with BCC presentational phenotypes. Results (i) After clustering BCC accrual increased; and (ii) exposure to UV in single and multiple cluster MPP cases were similar. In multiple cluster cases, mean age at first presentation with a single tumor occurred earlier and, the frequencies of CYP2D6 EM (94.4%) and GSTT1 null (41.2%) were significantly greater (P = 0.028 and P = 0.004) than in single cluster cases (67.1 and 14.3%). The odds ratios for these associations with the multiple cluster MPP were large; 15.5 and 7.39, respectively. Conclusions The finding of clusters of new, primary BCC is a critical event that is followed by markedly increased accrual of further tumors. Clustering occurs at a relatively late age and may be associated with a failure in immune surveillance. We propose the MPP is not the consequence of excessive UV exposure but reflects the presence of a distinct BCC subgroup defined by a combination of risk genes. Med. Pediatr. Oncol. 36:559–563, 2001. © 2001 Wiley-Liss, Inc.

The objective of this study was to determine whether the association between GSTM1 null/GSTTI null and survival in ovarian cancer is mediated by the influence of these genes on p53 expression. In 81 women with pure invasive ovarian cancer, GSTM1 null and GSTT1 null genotypes were identified using polymerase chain reaction and p53 expression was assessed using immunohistochemistry. The association of these factors with survival was examined using Cox's proportional hazards regression models. Performance status (P < 0.001), operative stage (P = 0.004), residual disease (P = 0.001), histologic subtype (P = 0.05), tumor grade (P = 0.007), and the combined GSTMI null/GSTTl null genotype (P = 0.023) were all individually associated with survival. p53 expression was not associated with survival (P = 0.45). In a multivariate analysis, the effects of GSTM1 null/GSTT1 null on survival were lost when residual disease and tumor grade were included. The effects of p53 expression on survival were unchanged when residual disease, tumor grade, operative stage, and performance score were included. GSTM1 null/GSTT1null did not influence the effects of p53 expression on survival and vice versa. The GSTM1 null/GSTT1 null genotype was associated with response to primary chemotherapy (P = 0.007) but p53 expression was not. We conclude that the association of GSTM1 null/GSTTl null with survival appears to be mediated through different mechanisms to p53 expression in ovarian cancer and in addition, may be a better predictor of outcome.

Dhar KK, Branigan K, Howells REJ, Musgrove C, Jones PW, Strange RC, Fryer AA, Redman CWE, Hoban PR. Prognostic significance of cyclin D1 gene (CCND1) polymorphism in epithelial ovarian cancer. We have investigated the influence of CCND1 genotype on clinical outcome in 138 women with epithelial ovarian cancer. CCND1 genotypes were identified from peripheral blood DNA by polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) analysis. Patient CCND1 genotypes were compared with clinical details including FIGO tumor stage, residual tumor volume, tumor histology and differentiation, response to chemotherapy, progression free interval, and survival. We observed no association between patient CCND1 genotypes and tumor characteristics or response to chemotherapy. There was no significant difference in overall survival and progression free interval (PFI) among women with different CCND1 genotypes. However, analysis of data from patients who responded to postoperative chemotherapy revealed that women with CCND1 AA genotype were associated with early disease progression (P = 0.020, HR 4.58, 95% CI 1.27–16.48) and reduced survival (P = 0.026, HR 4.48, 95% CI 1.19–16.79) compared with those with CCND1 AG and GG genotypes. These data show that CCND1 genotype does not influence overall prognosis in a cohort of epithelial ovarian cancer patients, however, it is associated with disease progression in a subgroup of patients following initial response to chemotherapy.

A single-center, cross-sectional, longitudinal study was conducted to determine the prevalence, annual incidence, and clinical risk factors for skin cancer in a white renal transplant population. One hundred eighty-two white patients (95% of population) with functioning allografts, a mean age at transplantation of 38.9 +/- 15. 6 (SD) years, and a mean follow-up of 8.5 +/- 6.3 years were interviewed and examined between May 1997 and June 1999. All case notes were carefully reviewed. Since transplantation, 16.5% of the patients had developed nonmelanoma skin cancer; 15.4%, actinic keratoses (AK); 53%, viral warts; and 1.6%, lentigo maligna melanoma (n = 3). Thirty-nine percent of the tumors were detected as a consequence of this study, and 20% of these occurred on covered body sites. The squamous cell (SCC)-basal cell carcinoma (BCC) ratio was 3.8:1. Eighty-two percent of the patients were examined a second time 12 months after the initial assessment. Using these data to identify new lesions, the annual incidence was calculated at 6.5%, increasing to 10.5% at more than 10 years posttransplantation. Duration of immunosuppression, older age at transplantation, presence of AK, male sex, and outdoor occupation were significantly associated with both SCC and BCC; SCC alone was associated with a history of having smoked tobacco. Early identification of those at greatest risk using a clinical risk profile may allow the development of more structured preventative and surveillance strategies than currently exist.

Most genetic studies of asthma have concentrated on genes on chromosomes 11q and 5q and their association with the key asthma-related phenotypes of bronchial hyperresponsiveness (BHR) and atopy. Although asthma is characterized by airway inflammation, a critical component of which is oxidative stress, few data exist on genes involved in protecting against this insult. We describe an association study designed to examine whether allelic variation at the glutathione-S-transferase GSTP1 locus influences expression of the BHR and atopy phenotypes in asthma. The enzyme encoded by GSTP1 utilizes a variety of lipid and DNA products of oxidative stress, and polymorphic variants of this gene are associated with altered catalytic function of this enzyme. We found that the frequency of GSTP1 Val(105)/Val(105) was significantly lower in asthmatic than in control subjects. Indeed, the presence of this genotype conferred a sixfold lower risk of asthma than did GSTP1 Ile(105)/Ile(105). Remarkably, asthma risk in Val(105) homozygotes was further reduced (by ninefold) after correction for atopic indices, age, and gender. Trend analysis after stratification according to the degree of bronchial reactivity/obstruction showed that the frequency of GSTP1 Val(105)/Val(105) correlates with decreasing severity of airway dysfunction. Furthermore, subjects with GSTP1 Val(105)/Val(105) have four- and 10-fold lower risks, respectively, of exhibiting atopy defined by skin test positivity and IgE level. These data show that GSTP1 polymorphism is strongly associated with asthma and related phenotypes, and provide an alternative explanation for the linkage of chromosome 11q13 with BHR and atopy.

Recent reports, primarily in patients with secondary progressive and relapsing-remitting multiple sclerosis have suggested that the apolipoprotein (APOE) e4 allele may be associated with a worse prognosis.1-3 The e class of APOE is involved in lipid transport and plays a part in brain development, synapse repair, and response to neuronal injury. It has been postulated that patients possessing the e4 allele might have a reduced capacity for neuronal remodelling after multiple sclerosis relapses.3 There is general agreement that patients with primary progressive multiple sclerosis have a worse outcome both in relation to development of disability over time and in mortality. Such patients do not have a relapsing course. Moreover, there are pathological and imaging differences between primary progressive and secondary progressive multiple sclerosis suggesting that primary progressive disease may be a less inflammatory form of the disease.4 The relation between APOE and outcome has not been specifically considered in patients with primary progressive multiple sclerosis, although 11 patients with …

Tumour necrosis factor alpha (TNF-α) appears important in ultraviolet-induced immunosuppression, suggesting that it is a susceptibility candidate for cutaneous basal cell carcinoma (BCC). We now describe data on the association between TNF microsatellite polymorphisms, first on susceptibility in 202 controls and 133 cases each having two to 30 BCCs, and secondly, within the cases, on BCC numbers. The data show that the proportions of individuals with TNF a1- and a7-containing genotypes were significantly different (P = 0·0271, P = 0·0393, respectively) between cases and controls. Secondly, within the cases, TNF alleles d4 (P = 0·023) and d6 (P = 0·006) alone, and the TNF a2–b4–d5 haplotype (P = 0·007), were significantly associated with the number of BCC lesions. These preliminary data provide the first evidence that TNF microsatellite polymorphism may influence the pathogenesis of multiple BCC.

Background: Oxidative stress has been implicated in inflammatory demyelination. The glutathione S-transferase (GST) supergene family encodes isoenzymes that appear to be critical in protection against oxidative stress. Certain GST loci are polymorphic, demonstrating alleles that are null (GSTM1/GSTT1), encode low activity variants (GSTP1), or are associated with variable inducibility (GSTM3). Objectives: To investigate the association between clinical outcome in MS and allelic variants of GSTM1, GSTM3, GSTT1, and GSTP1. Methods: Four hundred patients with clinically definite MS were studied. Disability was measured using the Kurtzke Expanded Disability Status Scale (EDSS). Disability was graded as mild (EDSS 0–4), moderate (4.5–5.5), or severe (EDSS 6–10). PCR-based genotyping was performed using DNA extracted from lymphocytes. Significant associations between GST genotypes and clinical outcome were corrected for gender, onset age, and disease duration using logistic regression. Results: We found that the GSTM3 AA genotype was associated with severe disability in patients with a disease duration of more than 10 years ( p = 0.027, n = 177, OR = 2.4, 95% CI = 1.1–5.0). Homozygosity for both GSTM1*0 and GSTP1*Ile 105 containing allele was associated with severe disability in patients with a disease duration greater than 10 years ( p = 0.022, n = 179, OR = 5.0, 95% CI = 1.3–19.8). Conclusions: Our results suggest that long-term prognosis in MS is influenced by a genetically determined ability to remove the toxic products of oxidative stress.

Glutathione S-transferase GSTM1 B and GSTT1 null, and cytochrome P450 CYP2D6 EM have been associated with cutaneous basal cell carcinoma (BCC) numbers, although their quantitative effects show that predisposition to many BCC is determined by an unknown number of further loci. We speculate that other loci that determine response to oxidative stress, such as NAD(H):quinone oxidoreductase (NQO1) are candidates. Accordingly, we assessed the association between NQO1 null and BCC numbers primarily to rank NQO1 null in a model that included genotypes already associated with BCC numbers. We found that only 14 out of 457 cases (3.1%) were NQO1 null. This frequency did not increase in cases with characteristics linked with BCC numbers including gender, skin type, a truncal lesion or more than one new BCC at any presentation (MPP). However, the mean number of BCC in NQO1 * 0 homozygotes was greater than in wild-type allele homozygotes and heterozygotes, although the difference was not quite significant (P = 0.06). These data reflect the link between NQO1 null and BCC numbers in the 42 MPP cases rather than the whole case group. We identified an interaction between NQO1 null and GSTT1 null that was associated with more BCC (P = 0.04), although only four cases had this combination. The relative influence of NQO1 null was studied in a multivariate model that included: (i) 241 patients in whom GSTM1 B, GSTT1 null and CYP2D6 EM genotype data were available, and (ii) 101 patients in whom these genotypes, as well as data on GSTM3, CYP1A1 and melanocyte-stimulating hormone receptor (MC1R) genotypes were available. NQO1 null (P = 0.001) and MC1R asp294/asp294 (P = 0.03) were linked with BCC numbers, and the association with CYP2D6 EM approached significance (P = 0.08). In a stepwise regression model only these genotypes were significantly associated with BCC numbers with NQO1 null being the most powerful predictor.

Allelic variants in the melanocyte stimulating hormone receptor (MC1R) gene are susceptibility/outcome candidates for cutaneous basal cell carcinoma (BCC). We identified the val92met (V92M) and asp294his (A294H) alleles in 311 cases and 190 controls. The cases included four homo- and 53 heterozygotes for V92M and 12 heterozygotes for A294H and two compound heterozygotes (V92M/A294H). Allele frequencies were similar in controls. In the cases, we found no association between the alleles and skin type though A294H was more common in those with red hair (4/19) than with other hair colours (6/163) (P=0.012). V92M was not associated with BCC numbers. Cases with A294H had fewer BCC in comparison with those without the allele though the difference was not significant. After inclusion of red hair in the model, A294H was significantly associated with fewer tumours. While MC1R alleles are attractive candidates for BCC, the variants studied did not influence susceptibility. The association with outcome was relatively weak. The large number of MC1R alleles and their low frequency, make assessment of the importance of this gene in the pathogenesis of skin cancers difficult.

Objectives. To investigate whether the therapeutic response of rheumatoid arthritis (RA) patients to d-penicillamine is associated with polymorphisms in genes of the glutathione S-transferase (GST ) supergene family. Methods. Disease activity in 81 patients with RA treated with d-penicillamine monotherapy was assessed using the Stoke Index, a validated index of disease activity, prior to treatment and at 6 months. GST typing was performed using a polymerase chain reaction-based approach and a logistic regression model was used to investigate any possible association between the therapeutic response to d-penicillamine and the GST genotype. Results. A poor therapeutic response was associated with the GSTM1 null genotype [odds ratio (OR) 3.94], and in particular with the GSTM1*0/GSTM3*A haplotype (OR 7.63). Conclusions. Our results suggest that GST polymorphisms may influence the response to d-penicillamine in RA, and that patients in possession of the GSTM1*0/GSTM3*A haplotype are significantly less likely to show a beneficial response to the drug.

OBJECTIVE—To determine whether glutathione S-transferase GSTM1, GSTM3, GSTT1, and GSTP1 genotypes influence susceptibility or outcome in rheumatoid arthritis (RA). METHODS—277 RA patients were compared with 577 controls to examine any associations between GST genotypes and susceptibility to RA. The effect of genotypes on outcome (Larsen and functional scores) and time integrated acute phase responses (erythrocyte sedimentation rate and C reactive protein) was assessed in 122 patients with disease duration of 5-10 years. GST and HLA-DRB1 genotypes were determined using polymerase chain reaction based assays. Data were analysed using multiple regression analysis with correction for age, sex, disease duration, and the DRB1 associated shared epitope (SE) and rheumatoid factor (RF) positivity where appropriate. RESULTS—The GSTM1*A/*B genotype was less common in RA cases (3 of 276) than in controls (22 of 591) (exact p=0.047), though significance was lost when adjustment was made for multiple comparisons. The Larsen score was higher (p=0.039) in the GSTM1 null patients (89.9) than those with other GSTM1 genotypes (74.7), and this was independent of the SE. Again, correction for multiple testing resulted in loss of significance. The difference in Larsen scores between patients homozygous or negative for the SE (87.9 v 74.3) was similar to that between GSTM1 null and non-null patients. No associations between GSTM3 or GSTT1 genotypes and disease markers were identified although the association between GSTP1*B/*B and Larsen score approached significance (p=0.096). CONCLUSION—It is proposed that certain GSTs may influence susceptibility and radiological progression in RA and that this is independent of the effect of the HLA-DRB1 associated SE. The mechanism for this effect is presumed to be because of differences in the ability of various GST enzymes to utilise the cytotoxic products of oxidant stress. Although significance was lost after correction for multiple testing, the data indicate that further studies may be of value in RA to determine the influence of the GST and other genes involved in cellular protection against oxidative stress. Keywords: rheumatoid arthritis; glutathione S-transferase; polymorphism; shared epitope

While cigarette smoking and alcohol consumption are recognized covariates for head and neck squamous cell carcinoma (SCC), the role of genetic factors in determining individual susceptibility is unknown. The human tumour necrosis factor (TNF) region on chromosome 6p21 within the major histocompatibility complex (MHC) includes a number of immunologically important genes. Recently, five microsatellite markers have been described in the TNF locus. TNF levels vary with different TNF microsatellite alleles, and associations of these microsatellite markers with autoimmune diseases and different types of cancer have been shown. Therefore, the TNF locus represents candidate susceptibility genes for head and neck cancer. This study describes the influence of TNF a-d microsatellite polymorphisms on susceptibility to head and neck cancer by comparing the allele frequencies of 269 patients suffering from laryngeal cancer and 123 patients suffering from oral cavity pharyngeal cancer and 113 German controls. DNA was extracted from peripheral blood samples, amplified by polymerase chain reaction with fluorescently labelled primers for TNF microsatellite (a- d) and electrophoresed on polyacrylamide gels using an automated DNA sequencer. The data showed no differences in allele frequencies between controls and pharyngeal cancer patients. By contrast, the TNF b3 allele was associated with altered risk for laryngeal cancer (p = 0.0006, odds ratio 2.2). Homozygosity for TNF b3/b3 resulted in an increased risk of developing laryngeal cancer (p = 0.004, odds ratio 5.3). Susceptibility to supraglottic SCC and multiple primary tumours was mediated by the absence of the a11 allele. The data provide the first evidence that allelism at the TNF microsatellite markers alter the risk of developing SCC of the larynx

Allelism has been found in human glutathione S-transferase (GST) genes of the alpha, mu, theta and pi families with the best characterised examples being those in mu class GSTM1 and theta class GSTT1. Isoenzymes encoded by these genes catalyse the detoxification of various reactive toxic and mutagenic compounds including epoxides resulting from the cytochrome P450-mediated metabolism of polycyclic aromatic hydrocarbons as well as lipid and DNA products of oxidative stress (Hayes and Strange, 1995, Smith et al, 1995). Homozygosity for null alleles or those encoding low activity variants are likely therefore, to be associated with a biochemical consequence. However, while accumulating evidence suggests the importance of different GST, it remains unclear precisely which in vivo processes are influenced by these polymorphisms. In this chapter we discuss firstly, a new polymorphism in GSTM3 and secondly, the role of GST polymorphisms in determining cancer susceptibility with particular reference to allelism at GSTMI, GSTT1 and GSTM3 and their interactions with cytochrome P450 (CYP) genotypes in basal cell carcinoma of skin (BCC).

While cigarette smoking and alcohol consumption have been linked to laryngeal squamous cell carcinoma (SCC), the role of genetic factors in determining individual susceptibility is unknown. We describe the role of allelism at the glutathione S-transferase GSTM1, GSTM3, GSTT1 and cytochrome P450 CYP1A1, CYP2E1, CYP2D6 loci in determining individual susceptibility to laryngeal SCC. Enzyme genotypes were determined using polymerase chain reaction and restriction enzyme digestion of leukocyte DNA collected from 269 patients with T1-T4 laryngeal carcinomas and 216 controls. While the frequencies of the heterozygote GSTM1 A/B genotype and the homozygote GSTM3 B/B genotype were statistically significantly lower in the patients with tumors than in controls, the frequency of the GSTT1 null genotype was higher in the patients than in controls. The data suggest that allelism at GST loci mediates susceptibility to SCC of the larynx. GSTM1 A/B and GSTM3 B/B appear to be associated with reduced risk, while GSTT1 null may confer increased risk. These findings are compatible with the view that genetic predisposition is important in determining risk for this cancer.

The factors that determine development of single and multiple primary cutaneous basal cell carcinomas (BCCs) are unclear. We describe a case-control study firstly, to examine the influence of allelism at the glutathione S-transferase GSTM1 and GSTT1 and cytochrome P450 CYP2D6 loci on susceptibility to these tumours and, secondly, to identify interactions between genotypes and relevant individual characteristics, such as skin type and gender. Frequency distributions for GSTM1 genotypes in cases and controls were not different, although the frequency of GSTM1 A/B was significantly lower (P = 0.048) in the multiple BCCs than in controls. We found no significant differences in the frequencies of GSTT1 and CYP2D6 genotypes in cases and controls. Interactions between genotypes were studied by comparing multinomial frequency distributions in mutually exclusive groups. These identified no differences between cases and controls for combinations of the putatively high risk GSTM1 null, GSTT1 null, CYP2D6 EM genotypes. Interactions between GSTM1 A/B and the CYP2D6 PM and GSTT1-positive genotypes were also not different. Frequency distributions of GSTM1 A/B with CYP2D6 EM in controls and multiple BCCs were significantly different (P = 0.033). The proportion of males in the multiple BCC group (61.3%) was greater than in controls (47.0%) and single BCC (52.2%), and the frequency of the combination GSTM1 null/male gender was significantly greater in patients with multiple tumours (P = 0.002). Frequency distributions of GSTM1 null/skin type 1 were also significantly different (P = 0.029) and the proportion of subjects who were GSTM1 null with skin type 1 was greater (P = 0.009) in the multiple BCC group. We examined the data for interactions between GSTM1 null/skin type 1/male gender by comparing frequency distributions of these factors in the single and multiple BCC groups. The distributions were almost significantly different (exact P = 0.051). No significant interactions between GSTT1 null or CYP2D6 EM and skin type 1 were identified. Comparisons of frequency distributions of smoking with the GSTM1 null, GSTT1 null and CYP2D6 EM genotypes identified no differences between patients with single and multiple tumours.

Allelism in the glutathione S-transferase, GSTM3 gene has been identified using PCR with specific primers to exon 6/exon 7. Sequencing showed the mutant GSTM3*B allele to have a three-base deletion in intron 6 with a frequency of 0.158. The mutation generates a recognition sequence, 5′-AAGATA-3′, for the negative transcription factor YY1. GSTM3*B was significantly associated with GSTM1*A.

The polymorphic glutathione S-transferase GSTM1 and GSTT1 loci are attractive candidates for susceptibility to ulcerative colitis and Crohn's disease because their enzymes catalyse the detoxification of products of oxidative stress and tobacco-derived carcinogens. We have tested this hypothesis by comparing genotype frequencies in 245 ulcerative colitis, 112 Crohn's disease patients and 373 controls. Polymerase chain reaction approaches were used to identify the GSTM1 A, GSTM1 B, GSTM1 A B , GSTM1 null and GSTT1 null and GSTT1 positive genotypes. The ages and smoking characteristics of ulcerative colitis and Crohn's patients were significantly different. Frequencies of GSTM1 genotypes in distal and total ulcerative colitis and controls were not different though frequency of GSTT1 null was significantly greater in the total disease (34.6%) than in the distal disease (17.5%) and control groups (17.7%). The age-corrected odds ratio for GSTT1 null was 2.40 (P = 0.0077). The frequency of GSTT1 null in the Crohn's patients and controls was similar though the frequency of GSTM1 B (7.3%) was significantly reduced compared with controls (15.0%) and patients with distal ulcerative colitis (19.9%). These data indicate that GSTM1 and GSTT1 mediate risk of certain inflammatory pathologies but are not general markers of susceptibility to such diseases. They also contribute to data showing that susceptibility to distal and total ulcerative colitis and Crohn's disease are influenced by different genes.

We describe studies to determine if susceptibility to pituitary tumours is associated with the putatively high risk GSTM1 null and CYP2D6 EM genotypes. Frequency distributions of these genotypes were similar in cases and controls though the frequency of CYP2D6 PM and GSTM1 B tended to be lower (P = 0.072 and P = 0.095 respectively) in the tumour group. Immunopositivity for p53 was found in 18/97 tumours. In these samples GSTM1 null (39%) and CYP2D6 EM (56%) frequencies were not different to those in controls. The frequencies of CYP2D6 PM and GSTM1 B in the p53 immunonegative tumours tended to be lower (P = 0.055 and P = 0.1185 respectively) than in controls. Mutations in gsp and ras were studied using the polymerase chain reaction and allele specific oligonucleotide analysis. Eight of 19 somatotrophinomas demonstrated mutations in gsp; frequencies of GSTM1 null and CYP02D6 EM were similar to controls. No ras mutations were identified in 55-tumour studies. The data indicate the GSTM1 null and CYP2D6 EM genotypes are not associated with altered expression of p53 or, mutation of gsp and ras in these adenomas and, suggest the CYP2D6 PM genotype is associated with a reduced risk of pituitary adenomas and, that GSTM1*B confers greater protection than GSTM1*A.

Multiple allelism at loci encoding detoxicating enzymes is associated with cancer risk. We have studied genetic variation at the glutathione S-transferase GSTM1 locus to see whether phenotypes confer altered susceptibility to basal cell carcinoma (BCC), squamous cell carcinoma (SCC), malignant melanoma (MM), or multiple skin tumours of different histological types. The frequency of GSTM1 null in cases and controls (52%) was similar, except for patients with two or more tumours of different types (71%, p = 0.033). GSTM1 A/B was reduced in frequency (p < 0.05) in patients with single or multiple BCC. Thus GSTM1 A/B may be protective, and effectiveness of detoxication may be a factor determining susceptibility to skin cancer.

Although it has been known for centuries that there are compounds in exhaled breath that are altered in disease, it is only in the last few decades that it has been possible to measure them with sufficient accuracy and precision to make them clinically useful. The clinical utility of breath analysis has also been limited by the practical difficulties of collecting representative breath samples, free from contaminants. More recent methods of breath analysis have allowed real-time analysis of breath, eliminating the need for sample collection, and therefore potentially allowing the rapid feedback of results to patient and clinician. One possible future application of breath analysis may be the monitoring of metabolic control in patients with diabetes mellitus. This perspective article provides an overview of the studies of breath analysis in diabetes, focusing on the breath metabolites; acetone, isoprene and also methyl nitrate that have previously been reported to be altered in diabetes, highlighting the factors that may potentially confound their interpretation. Specific attention is given to selected ion flow tube mass spectrometry (SIFT-MS) and proton transfer reaction mass spectrometry (PTR-MS), because they are techniques that have been developed specifically for the absolute quantification of breath metabolites in real time, although reference is made to some of the alternative techniques, including sensors and optical devices. Whilst breath analysis, using SIFT-MS, PTR-MS and other sensitive techniques, can potentially be used for the non-invasive monitoring of metabolic conditions that may include diabetes mellitus, further work is required in terms of the clinical and analytical validation. Furthermore, it is unclear at present what breath metabolites should be monitored and what factors may confound their interpretation. Although a non-invasive method of monitoring glycaemic control is clearly desirable, it will be important to demonstrate its analytical comparability with the well-established and validated methods for blood glucose measurement.

We describe the identification of the GSTM1 null, GSTM1 A, GSTM1 B and GSTM1 A,B polymorphisms at the glutathione S-transferase GSTM1 locus using a single-step PCR method. Target DNA was amplified using primers to intron 6 and exon 7 with site-directed mutagenesis being used to introduce a restriction site in DNA amplified from GSTM1 *A, thereby allowing differentiation of this allele and GSTM1 *B. The accuracy of this approach in identifying the GSTM1 A, GSTM1 B, GSTM1 A,B and GSTM1 null polymorphisms was confirmed by comparison with, firstly, an established PCR method that distinguishes GSTM1 *0 homozygotes from individuals with the other GSTM1 genotypes and, secondly, GSTM1 phenotypes determined using chromatofocusing.

Studies were carried out to test the hypothesis that the GSTM1 null phenotype at the mu (mu) class glutathione S-transferase 1 locus is associated with an increased predisposition to primary biliary cirrhosis. Starch gel electrophoresis was used to compare the prevalence of GSTM1 null phenotype 0 in patients with end stage primary biliary cirrhosis and a group of controls without evidence of liver disease. The prevalence of GSTM1 null phenotype in the primary biliary cirrhosis and control groups was similar; 39% and 45% respectively. In the primary biliary cirrhosis group all subjects were of the common GSTM1 0, GSTM1 A, GSTM1 B or GSTM1 A, B phenotypes while in the controls, one subject showed an isoform with an anodal mobility compatible with it being a product of the putative GSTM1*3 allele. As the GSTM1 phenotype might be changed by the disease process, the polymerase chain reaction was used to amplify the exon 4-exon 5 region of GSTM1 and show that in 13 control subjects and 11 patients with primary biliary cirrhosis, GSTM1 positive and negative genotypes were associated with corresponding GSTM1 expressing and non-expressing phenotypes respectively. The control subject with GSTM1 3 phenotype showed a positive genotype.

The risk of non-melanoma skin cancer is increased after organ transplantation, with a prevalence and annual incidence for the United Kingdom reported at 16.5% and 7.1-10.6% respectively. 1 2 Non-melanoma skin cancer presents at an earlier age and spreads more rapidly in people who have received a transplant than in the general population, and it often occurs at more than one site. These factors result in substantial morbidity and a sevenfold increase in mortality from the disease, although absolute death rates are low.3 The high incidence, rapid growth, and increased metastatic potential of non-melanoma skin cancer in transplant recipients justifies a surveillance programme. 1 3 We did a survey to establish current practice in skin cancer surveillance in UK centres managing renal transplant recipients. View this table: Skin cancer surveillance and education of transplant recipients followed up in 61 surgical and nephrology centres. Values are numbers (percentages) of centres unless stated otherwise We sent a questionnaire to …

Summary A positive association between latitude and prostate cancer mortality has been interpreted to indicate that ultraviolet radiation (UVR) protects against development of this cancer. We aimed to examine this hypothesis. We compared exposure between 210 cases and 155 controls. Childhood sunburn (odds ratio 0·18, 95% CI 0·08–0·38), regular foreign holidays (0·41, 0·25–0·68), sunbathing score (0·83, 0·76–0·89), and low exposure to UVR (3·03, 1·59–5·78) were associated with development of prostate cancer. Furthermore, cases with low UVR exposure developed cancer at a younger median age (67·7 years, IQR 61·5–74·6) than cases with higher exposure (72·1 years, 67·5–76·4); p=0·006. These findings are compatible with UVR having a protective role against prostate cancer.

Objective The objective of this study was to investigate whether the functional rs25531 promoter polymorphism in the serotonin transporter gene is associated with premenstrual dysphoric disorder. Study Design The study sample comprised 53 women with clinically diagnosed premenstrual dysphoric disorder (age range, 27–46 years; mean, 37.7 years) and 52 healthy control subjects (age range, 22–48 years; mean, 36.2 years). The rs25531 polymorphism was genotyped in both groups. Because of its close proximity to rs25531, the 5-HTTLPR promoter polymorphism was also genotyped. Genotype and allele frequencies for rs25531 and for the composite 5-HTTLPR/rs25531 marker were analyzed by χ 2 test. Results There was no significant association between any genotype and clinical category and no significant allele distribution profiles for rs25531 or 5-HTTLPR/rs25531 in either the premenstrual dysphoric disorder or the control groups. Conclusion These findings do not support a major role for rs25531, either in isolation or combined with 5-HTTLPR, in contributing to susceptibility to premenstrual dysphoria.

Objectives Periconceptual folate supplementation is recommended to reduce the risk of fetal neural tube defect and other congenital defects. Folate is a source of methyl donors, and experimental data on rodents suggest it could influence DNA methylation and alter gene expression. Previously the authors demonstrated a statistically significant inverse correlation (p=0.001) between cord blood global DNA methylation and plasma homocysteine, a recognised inverse indicator of methyl donor supply. The aim of this study was to determine if these observed changes in global methylation were also apparent in putative gene promoters. Materials and methods Cord blood DNA from babies with different levels of global DNA methylation was used for Methylated DNA Immunoprecipitation. Microarray analysis of enriched methylated DNA was used to examine the methylation status of more than 28 000 gene associted CpG islands. For array identified methylated regions, results were validated by bisulphite sequencing. Results The microarray data demonstrated that, increasing global DNA methylation is accompanied by elevated methylation in promoters/CpG islands of a number of genes. Bioinformatics analysis of array data showed that mean methylation of CpG islands and surrounding shores increased with the increase in level of Global methylation, however, this was higher in the CpG shores compared to the islands. Conclusion This is the first study demonstrating effects of prenatal maternal dietary exposure to methyl groups on fetal gene specific DNA methylation. Changes in DNA methylation can potentially play significant role in fetal programming and may determine the risk of adult-onset diseases.

We describe expression of alpha, mu and pi class glutathione S-transferases (GST) in brain tissue from 21 controls and uninfiltrated and tumour tissue from 17 glioma patients. GST were sequentially resolved by chromatofocusing into the GST2, GST1, GST5, GST2 (5.5), GST3, GST6 sets and the contribution of each to total activity determined. The immunological identity of these isoforms was studied using immunoblotting. The pi class GST3 isoform was the major contributor to activity in control tissue (70.9%) and, uninfiltrated (75.1%) and tumour samples (82.4%). Expression was significantly greater in the tumours (P less than 0.05). Expression of alpha isoforms GST2 and GST2 (5.5) was variable with most subjects demonstrating no detectable GST2 (B1 and B2 chromatofocused monomers). An isoform termed GST2 (5.5) chromatofocussed at pH 5.5 and cross-reacted with antisera to B1. It was detected in most control and glioma patients and comprised about 5% of total activity. The contribution of GST2 and GST2 (5.5) to activity was similar in control, uninfiltrated and tumour tissue. Two mu class enzymes, GST1 and GST5, were identified. GST1 isoforms were detected in 9 of 21 control samples, the phenotype of these and matched liver samples were identical. GST1 isoforms were detected in 4 of 16 tumour samples, a significantly lower incidence than in a previously established control group. GST5 was expressed in most samples, the contribution of this locus to activity was significantly reduced in the tumours (5.2%) compared with control samples (14.5%).

Total protein, alphafetoprotein, albumin, prealbumin, alpha-1-antitrypsin, transferrin and ceruloplasmin levels were measured in samples of human fetal and neonatal cerebrospinal fluid (CSF) (97 cases), obtained between 12 and 41 weeks of gestation. In 39 of these cases (13 to 40 weeks gestation) plasma was also available for comparative analysis. CSF was collected from lateral ventricles in the first half of gestation and from the lumbar region in the second. Since these CSF samples were obtained from different sites, the protein levels in the lateral ventricle (early) samples could not be compared directly with those in the lumbar (later) samples. However, the mean protein levels in the lumbar samples were lower than those in the ventricular samples, which is in accord with the decline in CSF protein levels described in maturing animal fetuses. Despite a wide scatter of results, particularly in the first half of gestation, significant decline in the level of CSF alphafetoprotein was demonstrated during both first and second halves of gestation, and of albumin and prealbumin in the second half. No sex differences were found except for ceruloplasmin in lumbar CSF later in gestation, when males had higher levels than females. In fetal plasma, protein levels increased with increasing gestation apart from alphafetoprotein and prealbumin which both declined progressively. CSF/plasma ratios were dissimilar for different proteins, and changed with increasing gestation. These findings support the concept that the human fetal blood brain barrier matures early.

Skin cancer is a well-recognized risk of prolonged immunosuppression, for example, following renal transplantation. These tumours contrast with idiopathic lesions in that squamous cell, rather than basal cell carcinomas usually predominate. We report a Caucasian female who developed multiple basal cell carcinomas following protracted cytotoxic therapy for acute myeloid leukaemia and subsequently chronic lymphocytic leukaemia. No other clinical risk factors nor relevant polymorphisms of genes encoding for detoxifying enzymes were identified. Immune suppression is a well-recognized cause of multiple skin tumours, the most striking increase usually being of squamous cell carcinomas. We believe this woman is representative of a subgroup of immunosuppressed patients who, for as yet poorly understood reasons, have a predisposition to basal cell, rather than squamous cell carcinoma accrual.

Most common diseases appear to result from complex, poorly understood interactions between genetic and environmental factors. Relatively few factors have been unequivocally linked with disease risk or outcome. Evidence from various studies using different experimental approaches has been interpreted as showing that, apart from its harmful effects on the pathogenesis of the common skin cancers, ultraviolet radiation (UVR) may exert a beneficial effect on development of various internal cancers and other pathologies. This concept is supported by parallel studies showing that hypovitaminosis D is linked with increased risk of various diseases including insulin resistance and multiple sclerosis. These findings suggest that, first, host factors such as skin pigmentation that affect UVR-induced synthesis of vitamin D and, second, polymorphism in genes that mediate the effectiveness of vitamin D action are susceptibility candidates for a variety of diseases. Collectively, these data suggest the hypothesis that, via effects on vitamin D synthesis, UVR exposure has beneficial effects on susceptibility and outcome to a variety of complex diseases. We describe evidence from studies in various diseases, but mainly from prostate cancer patients, that supports this hypothesis, but we emphasize that, although supportive data are available, the concept is unproven. Indeed, other explanations are possible. However, given the potentially important public health implications of the hypothesis and the potential for the development of novel therapeutic modalities, we believe the concept is worthy of further investigation.

UI - 97000815 LA - eng RN - 0 (Autoantibodies) RN - 0 (Autoantigens) RN - 0 (Ribonucleoproteins) RN - 0 (SS-A antigen) RN - EC 2.5.1.18 (Glutathione Transferase) PT - Journal Article DA - 19961112 IS - 0004-3591 SB - AIM SB - IM CY - UNITED STATES

EDITOR,—Graham Bentham shows that the incidence of non-Hodgkin's lymphoma is positively associated with exposure to solar ultraviolet radiation,1 giving insights into the previous observation of a link between non-Hodgkin's lymphoma, malignant melanoma, and squamous cell skin cancer.2 He did not, however, assess any association with basal cell carcinoma, the commonest malignancy in white people. Exposure to ultraviolet radiation is recognised as a critical factor in the pathogenesis of basal cell carcinoma, …

The glutathione S-transferase (GST) enzyme GSTP1 utilizes byproducts of oxidative stress. We previously showed that alleles of GSTP1 that encode the Ile105→Val105 substitution are associated with the asthma phenotypes of atopy and bronchial hyperresponsiveness (BHR). However, a further polymorphic site (Ala114→Val114) has been identified that results in the following alleles: GSTP1 * A (wild-type Ile105→Ala114), GSTP1 * B (Val105→Ala114), GSTP1 * C (Val105→Val114) and GSTP1 * D (Ile105→Val114). Because full identification of GSTP1 alleles may identify stronger links with asthma phenotypes, we describe an amplification refractory mutation system (ARMS) assay that allows identification of all genotypes. We explored whether the GSTP1 substitutions influence susceptibility to asthma, atopy and BHR. Among 191 atopic nonasthmatic, atopic asthmatic and nonatopic nonasthmatic individuals, none had the BD, CD, or DD genotypes. GSTP1 BC was significantly associated with reduced risk for atopy (P = 0.031). Compared with AA, trend test analysis identified a significant decrease in the frequency of GSTP1 BC with increasing severity of BHR (P = 0.031). Similarly, the frequency of GSTP1 AA increased with increasing BHR. These data suggest that GSTP1 * B and possibly GSTP1 * C are protective against asthma and related phenotypes.

Objective To determine whether polymorphism in the manganese superoxide dismutase (MnSOD) gene is associated with susceptibility or disease outcome in rheumatoid arthritis (RA). Methods We used a case–control approach with 153 RA patients and 218 control subjects to examine for any associations between MnSOD genotypes and susceptibility to RA. We also investigated the influence of genotypes on radiologic outcome, as measured using the Larsen score for radiographs of the hands and feet, and on functional outcome, as assessed by the Health Assessment Questionnaire. MnSOD typing was carried out using polymerase chain reaction–based methods. Results were analyzed using multiple regression analysis, with adjustment for age, sex, and disease duration. In separate analyses, we corrected for rheumatoid factor (RF) status and/or the presence of the HLA–DRB1 “shared epitope” (SE). We also examined whether radiologic outcome was influenced by interactions between MnSOD and glutathione S-transferase (GST) genes. Results No association between MnSOD genotype and development of RA was found. The MnSOD VV genotype was associated with a significantly higher (P = 0.04) Larsen score (104.4) than MnSOD AA (83.0), while MnSOD AV was associated with an intermediate score (91.8). Correction for RF status had no significant effect on the results of the analysis, but significance was lost (P = 0.09) after correction for the presence of the SE. There was evidence of interaction between the GSTT1 and MnSOD genotypes, with the MnSOD VV/GSTT1-null combination being associated with the highest Larsen score (142.1; P = 0.007 after correction for the SE). Conclusion Polymorphism in the MnSOD gene is not associated with susceptibility to RA. Our data suggest that MnSOD VV is associated with more severe radiologic outcome, although this relationship may not be independent of the effect of the SE. However, interaction between MnSOD and GST genes appears to influence radiologic outcome independently of the SE.