Prostate Cancer and Prostatic Diseases (2002) 5, 22–27 ß 2002 Nature Publishing Group All rights reserved 1365–7852/02 $25.00 www.nature.com/pcan Polymorphisms of GSTP1 and related and prostate cancer risk

TM Beer1*, AJ Evans1, KM Hough1, BA Lowe2, JE McWilliams1 & WD Henner1 1Division of Hematology and Medical Oncology, Oregon Health Sciences University, Oregon, USA; and 2Division of Urology and Renal Transplantation, Oregon Health Sciences University, Oregon, USA

Glutathione S-transferase P1 (GSTP1) is markedly downregulated in prostate cancer and prostatic intraepithelial neoplasia compared to normal prostate tissue. Downregulation of GSTP1 may, therefore, be an early event in prostate carcino- genesis. An A?G polymorphism at nucleotide 313 results in an amino acid substitution (Ile105Val) in the substrate binding site of GSTP1 and reduces catalytic activity of GSTP1. In a study of 36 prostate cancer patients, Harries et al. reported that the Ile/Ile genotype is associated with a decreased risk of prostate cancer (odds ratio 0.4 (0.17 – 0.82)). We sought to confirm this finding and to examine the impact of this polymorphism together with several related poly- morphisms implicated as risk factors for carcinogen-associated malignancies. One hundred and seventeen patients with prostate adenocarcinoma and 183 population-based controls were recruited to this case – control study. Genotyping of the GSTP1 (Ile105Val), GSTM1 (null), GSTT1 (null) and CYP1A1 (Ile462Val) genes was performed using polymerase chain reaction (PCR) based techniques on DNA prepared from peripheral blood. A questionnaire was used to collect demographic information from each subject. Cases were significantly older (P < 0.0001) and had significantly greater family history of prostate cancer (P < 0.0001), confirming known risk factors for this disease. By w2 analysis, none of the genotype distributions varied among cases and controls. Using a logistic regression model to control for known risk factors we were also unable to demonstrate a significant association with prostate cancer for any of the poly- morphisms tested. This population fails to identify a relationship between the above polymorphisms and prostate adenocarcinoma. Prostate Cancer and Prostatic Diseases (2002) 5, 22–27. DOI: 10.1038/sj/pcan/4500549

Keywords: GSTP1; polymorphism; risk; prostate cancer

Introduction GSTP14 Family history of prostate cancer is, save for age, the The S-transferase P1 isoenzyme (GSTP1)isa 1–3 phase II detoxification . In contrast to most can- strongest predictor of prostate cancer risk. Only a 5 fraction, approximately a fifth, of inherited prostate cers, prostate carcinogenesis is associated with marked cancer risk can be explained by Mandelian inheritance downregulation of GSTP1. GSTP1 is expressed predomi- of cancer susceptibility genes.4 Genetic polymorphisms nantly in the basal layer of the normal prostate epithe- that alter the function or expression of genes that partici- lium. Although normal prostate secretory cells do not pate in prostate carcinogenesis may also contribute to routinely express GSTP1, they remain capable of expres- sing this enzyme and retain an unmethylated GSTP1 inherited prostate cancer risk. Genes involved in host – 6 carcinogen interactions are the focus of this investigation. promoter. Increased GSTP1 expression is seen in secre- tory cells of atrophic prostate epithelium, supporting the notion that the enzyme remains inducible in this cellular compartment.6 GSTP1 expression is markedly diminished *Correspondence: TM Beer, Division of Hematology and Medical in an overwhelming majority of adenocarcinoma and Oncology, Oregon Health Sciences University, Mail Code L586, 3181 7–9 S.W. Sam Jackson Park Rd., Portland, OR 97201, USA. prostatic intraepithelial neoplasia (PIN) specimens. In E-mail: [email protected] both cancer and PIN, this downregulation occurs at the Received 3 April 2001; accepted 28 August 2001 transcription level and is thought to be mediated by Genetic polymorphisms in prostate cancer risk TM Beer et al 23 hypermethylation of deoxycytidine residues in the CG- CYP1A1 rich promoter region located 50 of the transcriptional start site.7,10 – 13 Events leading up to GSTP1 inactivation A polymorphism exists in the CYP1A1 consisting of during prostate carcinogenesis remain unclear. Several a single base change at codon 462 in exon 7 (Ile462Val) 35 investigators have speculated that the early loss of that codes for either an Ile or a Val. The Val allele codes GSTP1 function leads to increased vulnerability to oxi- for a more inducible form of the enzyme and the Val/Val dant and heterocyclic amine carcinogens, both implicated genotype is associated with an increased risk of prostate in prostate carcinogenesis.14 – 16 Nelson and colleagues cancer (OR ¼ 2.6 (1.11 – 6.25)) and the presence of a single demonstrated that GSTP1 overexpression confers partial Val allele results in a trend towards an increased risk of 29 resistance to the cytotoxic effects of 2-hydroxyamino-1- prostate cancer (OR ¼ 1.4 (0.86 – 2.29)) in Japanese men. methyl-6-phenylimidazo[4,5- b]pyridine, the most abun- In the study reported here, we have investigated a dant mutagenic heterocyclic amine found in cooked largely Caucasian population of cases and controls for meats.17 It is therefore also possible that heritable differ- which we have obtained information concerning other ences in GSTP1 function may also be associated with known risk factors. Our hypothesis was, that because prostate cancer development. GSTP1 is downregulated early in prostate carcinogenesis, An A?G polymorphism at nucleotide 313 results in an the polymorphism, which reduces GSTP1’s enzymatic amino acid substitution (Ile105Val) in the substrate bind- activity, may be associated with increased risk of prostate ing site of GSTP1.18 The substitution of the less bulkier cancer. In addition to GSTP1, we examined several closely and more hydrophobic valine results in substrate-depen- related genes, known to mediate host-carcinogen interac- dent alterations of GSTP1 catalytic activity.18,19 A second tion, because of the possibility that GSTP1 may interact polymorphism, a C?T transition at nucleotide 341 results with these genes during carcinogenesis. in a Ala114Val amino acid substitution. This polymorph- ism is linked to the Ile105Val polymorphism and does not appear to result in significant additional alteration in enzymatic function.18 For this reason, this linked poly- morphism is not examined in this study. Materials and methods In a study of 36 prostate cancer patients, Harries et al. Population samples reported that the Ile/Ile genotype is associated with a decreased risk of prostate cancer (OR ¼ 0.4 (0.17 – Study subjects were recruited from the Portland, Oregon 0.82)).20 Kote-Jarai found a trend towards a protective area. Individuals with adenocarcinoma of the prostate effect of Ile alleles.21 In contrast to this finding, Gsur et al (cases) were recruited from the Urology and Medical found that the Val/Val genotype was protective (OR ¼ 0.24 Oncology clinic at Oregon Health Sciences University (0.09 – 0.61)) in a study of 322 men with either newly and Portland VA Medical Center. The two institutions diagnosed prostate cancer or benign prostatic hypertro- serve a broad mix of referral, general community and phy.22 Three investigators reported no effect of this poly- indigent patients. Controls were recruited primarily by morphism on prostate cancer risk.23 – 25 advertisement. Eligibility criteria for cases was age 18 or greater, histologically proven adenocarcinoma of the prostate and ability to give informed consent. Eligibility for controls was age 18 or greater, male gender, and lack GSTM1 of any malignancy (except non-melanoma skin cancer). After giving informed consent, study subjects were admi- The glutathione S-transferase M1 isoenzyme (GSTM1)is nistered a questionnaire regarding age, race, ethnicity, homozygously deleted in about one-half of the popula- 26 – 28 and family history of cancer. This study was approved tion of Europe and the United States. A trend by the Institutional Review Boards of Oregon Health towards increased risk of prostate cancer was seen in Sciences University and Portland VA Medical Center. Japanese men (OR ¼ 1.3 (0.82 – 2.04)) and appeared to be additive with risk associated with a CYP1A1 polymorph- 29 ism. No association between this polymorphism and DNA preparation prostate cancer risk was seen in several studies of a predominantly Caucasian population.21 – 23,25,30,31 DNA was isolated from peripheral blood lymphocytes using either Ficoll gradient centrifugation followed by Proteinase K digestion and phenol – chloroform extrac- tion36 or by use of the PuregeneTM method (Gentra GSTT1 Systems, Minneapolis, MN, USA). DNA samples were serially numbered upon arrival in the laboratory to There exists a deletion polymorphism (null mutation) of ensure subject confidentiality. GSTT1 with about 38% of the population deficient in the enzyme.32 The available information about GSTT1 null mutations and prostate cancer risk is conflicting. Two Polymerase chain reaction (PCR) techniques studies of predominantly Caucasian population sug- gested a modestly elevated risk (OR ¼ 1.83 (1.19 – PCR was carried out in a total volume of 50 ml, containing 2.80))30 (OR ¼ 2.31 (1.17 – 4.59)),25 while three others 0.2 – 1.0 mg of genomic DNA; 400 mM each of dATP, dCTP, failed to confirm these results.21,22,33 No relationship dGTP and dTTP; 10x PCR buffer containing 1.5 mM of between GSTT1 genotype and prostate cancer risk was MgCl2;50mM of each primer; 1.25 units of Taq DNA found in a study of Japanese men.34 polymerase (Promega) and cycled in a thermal cycler (MJ

Prostate Cancer and Prostatic Diseases Genetic polymorphisms in prostate cancer risk TM Beer et al

24 0 0 Research Inc., PTC 200). Amplification products were primers 5 -TGTCTCCCTCTGGTTACAGGAAGC-3 and resolved on an agarose gel containing ethidium bromide 50-GCAGGATAGCCAGGAAGAGAAAGAC-30. PCR was and DNA bands were visualized by ultraviolet trans- performed for 35 cycles at 94C for 20 s, 62C for 40 s and illumination. 72C for 40 s. Products were then digested with Bsr.A 140 bp band is formed by the digestion of wild type CYP1A1 while those alleles with the A?G transition GSTP1 (Ile105Val, A313G) polymorphism maintain the 180 bp band. GSTP1 genotyping was done following the methods of 18 Ali-Osman and co-workers. Briefly, exon 5 together Statistical analysis with segments of surrounding introns were amplified by PCR and subsequently digested by Tail which digests Different races are likely to have different genotype the mutant but not the wild type. The primers were 50- distributions for the polymorphisms. Since the study CCAGGCTGGGGCTCACAGACAGC-30 and 50-GGTCA- contained so few non-Caucasian individuals (5.1% GCCCAAGCCACCTGAGG-50. The reactions were cycled non-white), data analysis was restricted to Caucasian for 94C for 1 min, 53.5C for 1 min and 72C for 1 min. subjects. Odds ratios and 95% confidence intervals were calculated by comparing genotype frequencies in the case population to the control population by standard meth- GSTM1 null polymorphism ods.38 The referent genotypes are shown in Table 1. The PCR assay for deletion of GSTM1 presence or dele- Because cases and controls differed significantly with tion was performed as described by Comstock and co- respect to age and family history, both known risk factors workers.37 Co-amplification of exon 4 of the HPRT gene for prostate cancer, further analysis was performed to was used as a positive internal control. The primers were examine if imbalance in these risk factors contributed to 50-CTGCCCTACTTGATTGATGGG-30 and 50CTGGATTG- the outcome. First the odds ratios were calculated sepa- TAGCAGATCATGC-30. The reaction was cycled 35 times rately for cohorts below and above the median age of the at 94C for 20 s, 55C for 1 min and 1 min extension at entire study population in a stratified analysis. Then 72C. multiple logistic regression was used to model the effects of known risk factors (ie age, family history), genotypes for each of the various polymorphisms, and the interac- GSTT1 null polymorphism tions of genotypes and known risk factors. Statistical analyses were performed using the SPSS and Stat View The method for detecting the presence or absence of 5.0 statistical packages. GSTT1 was modified from Pemble et al 32 and utilizes primers 50-TTCCTTACTGGTCCTCACATCTC-30 and 50- TCACCGGATCATGGCCAGCA-30. The reaction was cycled 35 times at 94C for 20 s, 58C for 40 s and at 72C for 40 s. Primers for human apurinic endonuclease Results (APE) (exon 5) were included in the PCR reactions as a positive internal control. Demographics One hundred and seventeen individuals with prostate CYP1A1 (Ile462Val, A4889G) polymorphism cancer (cases) and 183 controls were recruited. Demo- graphic data for the two groups are illustrated in Table 2. We developed a restriction fragment length polymorph- The case subjects were significantly older (P < 0.0001) and ism (RFLP) assay for the genotyping of CYP1A1 alleles. had a more extensive family history (P < 0.0001) than We amplified a 180 bp portion of the CYP1A1 gene using control subjects.

Table 1 Distribution and odds ratios and power calculations for the gene polymorphisms studied

Polymorphism Cases Controls Odds ratio (95% CI) P-value OR detectable with 80% power

GSTP1 total 109 146 0.86 (0.52 – 1.42)* 0.56 2.15 Ile/Ile 51 63 Ile/Val 45 69 Val/Val 13 14 GSTM1 total 111 147 1.24 (0.75 – 2.03) 0.40 2.1 present 50 74 deleted 61 73 GSTT1 total 111 146 1.16 (0.65 – 2.06) 0.62 2.75 present 83 113 deleted 28 33 CYP1A1 total 110 146 0.68 (0.29 – 1.58)** 0.36 2.55 Ile/Ile 101 129 Ile/Val 717 Val/Val 20

*OR comparing Ile/Val and Val/Val to Ile/Ile. **OR comparing Ile/Val and Val/Val to Ile/Ile.

Prostate Cancer and Prostatic Diseases Genetic polymorphisms in prostate cancer risk TM Beer et al 25 Polymorphism allele frequencies Table 3 Logistic regression: analysis of family history and genotypes adjusted for age DNA was analysable from all of the samples though the exact numbers vary for each assay due to insufficient P-value OR* 95% Confidence intervals DNA or failure to amplify by PCR. The genotype dis- Age (years) < 0.0001 1.15 1.11 – 1.18 tribution for control subjects and the calculated allele or Family history in first 0.0002 7.90 2.65 – 23.53 genotype frequencies did not differ from previously degree relatives described frequencies in Caucasian populations. For the GSTP1 0.2117 0.67 0.36 – 1.26 two single nucleotide polymorphisms (SNPs), the control GSTM1 0.2235 1.47 0.79 – 2.73 GSTT1 0.9959 1.00 0.48 – 2.08 group genotype frequencies did not differ significantly CYP1A1 0.3503 0.62 0.23 – 1.69 from that predicted by Hardy-Weinberg equilibrium.39 *Estimated odds ratio from logistic coefficients. Genotype comparisons as described in Table 2. Polymorphism frequencies in cases and controls P > 0.1). Finally, to look for potentially important syner- 2 Individual genes were first analysed using w analysis and gies between polymorphisms, each of the polymorphisms these results are summarized in Table 1. None of the was added in combination to the analysis using both polymorphism frequencies differed significantly between additive and multiplicative models (data not shown). the cases and controls. For those polymorphisms where Again, there were no significant associations observed multiple comparisons were possible (GSTP1 and between any of the combined genotypes and the risk of CYP1A1) all possible comparisons were examined and adenocarcinoma of the prostate. none yielded a significant difference between cases and controls (data not shown). The minimum odds ratios detectable with 80% power are also shown in Table 1. Discussion Stratified analysis and logistic regression In this population, we examined and were unable to identify an increased risk for prostate cancer conferred The control population in this study was a convenience by several polymorphisms that have been reported to sample and cases and controls were not balanced for increase the risk of other malignancies. other, non-genetic risk factors known to be important in In the process of identifying and confirming genetic prostate cancer risk. When each genotype was examined and other risk factors for prostate cancer, it is important in strata below and above the median age of the entire that initial reports undergo confirmation. In follow-up study population (age 59), the odds ratios where consis- studies, it is important that the data obtained include tent across strata and consistent with results for the entire information regarding other known non-genetic risk fac- study population (data not shown). To explore this issue tors for prostate cancer. In the case of prostate cancer, further, we performed a logistic regression analysis of our these risk factors include most importantly race, age and data to control for the effect of known risk factors. In our family history of prostate cancer. initial analysis we examined age and family history in The previous report that the Ile/Ile genotype of the first degree relatives, which have been previously asso- glutathione S-transferase, GSTP1, was associated with a 40 ciated with prostate cancer. As expected, age and family reduced risk of prostate cancer was based on only a small history were predictive of case status (Table 3, P < 0.0001 population of 36 prostate cancer patients. In the larger for age and P ¼ 0.0002 for family history). population of cases and controls reported here, no sig- The information regarding genotype for each of the nificant effect of GSTP1 genotype on prostate cancer risk polymorphisms tested was then tested to determine its was identified. Likewise, no effect of polymorphisms in effect adjusted for age. Each polymorphism was added to the GSTM1, GSTT1 or CYP1A1 genes was detected. the analysis individually. The results of this analysis Polymorphisms in several of the human Phase II detox- indicated no significant association of any of the tested ification have been associated with altered risk polymorphisms in predicting case/control status (Table 3, of cancer, particularly tobacco-associated cancers such as lung and bladder. In the case of tobacco-associated can- Table 2 Demographic data for enrolled case and control subjects cers such effects are plausible because genetic changes in the activity of detoxification enzymes should alter cellular Cases Controls P-value* exposure to carcinogens. In the case of adenocarcinoma of Number of subjects 117 183 the prostate there is no clear link between exposure to Age (mean Æ s.d.) 66.9 Æ 9.6 52.2 Æ 10.5 < 0.0001 tobacco products or other carcinogens and subsequent Number with a family history 21 7 < 0.0001 risk of prostate cancer. of prostate cancer in first There is, however, a biological link between prostate degree relatives Race carcinogenesis and GSTP1 expression. This link made it Caucasian 111 150 particularly important to attempt to confirm whether Asian/Pacific Islander 1 1 GSTP1 genotypes alter prostate cancer risk. This link Black 2 17 provides a plausible biological mechanism by which a Hispanic 3 5 GSTP1 polymorphism might alter prostate carcinogen- Native American 0 4 esis. During development of prostate cancer, GSTP1 Unknown 0 6 expression is markedly down-regulated. The exact *Significance tests were by Student’s t-test for age and by w2 for family history. mechanism of this downregulation of GSTP1 expression

Prostate Cancer and Prostatic Diseases Genetic polymorphisms in prostate cancer risk TM Beer et al 26 and the role that decreased GSTP1 expression plays in 11 Brooks JD et al. CG island methylation changes near the GSTP1 prostate cell tumorigenesis is as yet unclear. The GSTP1 gene in prostatic intraepithelial neoplasia. Cancer Epidemiol Bio- protein appears to have other functions than to catalyze markers Prev 1998; 7: 531 – 536. 12 Millar DS et al. Detailed methylation analysis of the glutathione S- conjugation of nucleophiles to glutathione. Recently, transferase pi (GSTP1) gene in prostate cancer. Oncogene. 1999; 18: Adler and co-workers have reported that GSTP1 can act 1313 – 1324. to alter intracellular signaling through an interaction with 13 Singal R, van Wert J, Bashambu M. Cytosine methylation Jun N-terminal kinase (JNK).41 Decreased GSTP1 expres- represses glutathione S-transferase P1 (GSTP1) sion might therefore alter intracellular signaling and this in human prostate cancer cells. Cancer Res 2001; 61: 4820 – effect would provide a plausible biochemical mechanism 4826. 14 De Marzo AM, Nelson WG, Meeker AK, Coffey DS. Stem cell to explain a role in prostate carcinogenesis. The descrip- features of benign and malignant prostate epithelial cells. J Urol tion of the effect of TER 199, a highly specific GSTP1 1998; 160(6 Pt 2): 2381 – 2392. inhibitor, in stimulating growth of granulocytes further 15 DeWeese TL, Hruszkewycz AM, Marnett LJ. Oxidative stress in suggests a potentially important regulatory role for the chemoprevention trials. Urology 2001; 57(Suppl): 137 – 140. GSTP1 protein.42 16 Nelson WG, De Marzo AM, DeWeese TL. The molecular patho- In summary, in order to address the effect that different genesis of prostate cancer: implications for prostate cancer pre- genotypes may have on altering the risk of developing vention. Urology 2001; 57(Suppl): 39 – 45. 17 Nelson CP et al. Protection against 2-hydroxyamino-1-methyl-6- prostate cancer, we conducted a case – control study of phenylimidazo[4,5-b]pyridine cytotoxicity and DNA adduct for- four genetic polymorphisms and their association with mation in human prostate by glutathione S-transferase P1. Cancer prostate cancer. We were unable to demonstrate a sig- Res 2001; 61: 103 – 109. nificant association for the development of prostate 18 Ali-Osman F et al. Molecular cloning, characterization, and cancer in individuals bearing the Val substitutions in expression in Escherichia coli of full-length cDNAs of three GSTP1, the null alleles of GSTM1 and GSTT1, or the Val human glutathione S-transferase Pi gene variants. Evidence for differential catalytic activity of the encoded proteins. J Biol Chem encoding alleles of the CYP1A1 gene. Finally, using 1997; 272: 10004 – 10012. logistic regression analysis, we found no significant inter- 19 Sundberg K et al. Differences in the catalytic efficiencies of allelic actions between any of the polymorphisms tested. variants of glutathione transferase P1-1 towards carcinogenic diol epoxides of polycyclic aromatic hydrocarbons. Carcinogenesis 1998; 19: 433 – 436. 20 Harries LW et al. Identification of genetic polymorphisms at the Acknowledgements glutathione S-transferase Pi locus and association with suscept- ibility to bladder, testicular and prostate cancer. Carcinogenesis We would like to thank Dr Emily Harris for her sugges- 1997; 18: 641 – 644. tions and Dr Gary Sexton for statistical advice. This work 21 Kote-Jarai Z et al. Relationship between glutathione S-transferase was supported in part by grant 1R01 CA 72792-01 from M1, P1 and T1 polymorphisms and early onset prostate cancer. the National Institutes of Health. Pharmacogenetics 2001; 11: 325 – 330. 22 Gsur A et al. Polymorphisms of glutathione-S-transferase genes (GSTP1, GSTM1 and GSTT1) and prostate-cancer risk. Int J Cancer 2001; 95: 152 – 155. References 23 Autrup JL et al. Glutathione S-transferases as risk factors in prostate cancer. Eur J Cancer Prev 1999; 8: 525 – 532. 1 Cannon L, Bishop DT, Skolnick M. Genetic epidemiology of 24 Shepard TF et al. No association between the I105V polymorph- prostate cancer in the Utah Mormon genealogy. Cancer Surv ism of the glutathione S-transferase P1 gene (GSTP1) and prostate 1982; 1:47– 69. cancer risk: a prospective study. Cancer Epidemiol Biomarkers Prev 2 Steinberg GD et al. Family history and the risk of prostate cancer. 2000; 9: 1267 – 1268. Prostate 1990; 17: 337 – 347. 25 Steinhoff C et al. Glutathione transferase isozyme genotypes in 3 Spitz MR et al. Familial patterns of prostate cancer: a case-control patients with prostate and bladder carcinoma. Arch Toxicol 2000; analysis. J Urol, 1991; 146: 1305 – 1307. 74: 521 – 526. 4 Bastacky SI et al. Pathological features of hereditary prostate 26 Board PG. Biochemical genetics of glutathione-S-transferase in cancer. J Urol 1995; 153(3 Pt 2): 987 – 992. man. Am J Hum Genet 1981; 33:36– 43. 5 Tsuchida S, Sato K. Glutathione transferases and cancer. Crit Rev 27 Seidegard J, Guthenberg C, Pero RW, Mannervik B. The trans- Biochem Mol Biol 1992; 27: 337 – 384. stilbene oxide-active glutathione transferase in human mono- 6 De Marzo AM, Marchi VL, Epstein JI, Nelson WG. Proliferative nuclear leucocytes is identical with the hepatic glutathione trans- inflammatory atrophy of the prostate: implications for prostatic ferase mu. Biochem J 1987; 246: 783 – 785. carcinogenesis. Am J Pathol 1999; 155: 1985 – 1992. 28 Seidegard J, Pero RW, Miller DG, Beattie EJ. A glutathione 7 Lee WH et al. Cytidine methylation of regulatory sequences near transferase in human leukocytes as a marker for the susceptibility the pi-class glutathione S-transferase gene accompanies human to lung cancer. Carcinogenesis 1986; 7: 751 – 753. prostatic carcinogenesis. Proc Natl Acad Sci USA 1994; 91: 11733 – 29 Murata M et al. Cytochrome P4501A1 and glutathione S-transfer- 11737. ase M1 genotypes as risk factors for prostate cancer in Japan. Jpn J 8 Cookson MS, Reuter VE, Linkov I, Fair WR. Glutathione S- Clin Oncol 1998; 28: 657 – 660. transferase PI (GST-pi) class expression by immunohistochemis- 30 Rebbeck TR et al. Glutathione S-transferase-mu (GSTM1) and try in benign and malignant prostate tissue. J Urol 1997; 157: -theta (GSTT1) genotypes in the etiology of prostate cancer. 673 – 676. Cancer Epidemiol Biomarkers Prev 1999; 8(4 Pt 1): 283 – 287. 9 Moskaluk CA et al. Immunohistochemical expression of pi-class 31 Wadelius M et al. Polymorphisms in NAT2, CYP2D6, CYP2C19 glutathione S-transferase is down-regulated in adenocarcinoma and GSTP1 and their association with prostate cancer. Pharmaco- of the prostate. Cancer 1997; 79: 1595 – 1599. genetics 1999; 9: 333 – 340. 10 Lee WH, Isaacs WB, Bova GS, Nelson WG. CG island methyla- 32 Pemble S et al. Human glutathione S-transferase theta (GSTT1): tion changes near the GSTP1 gene in prostatic carcinoma cells cDNA cloning and the characterization of a genetic polymorph- detected using the polymerase chain reaction: a new prostate ism. Biochem J 1994; 300: 271 – 276. cancer biomarker. Cancer Epidemiol Biomarkers Prev 1997; 6: 33 Autrup JL et al. Glutathione S-transferases as risk factors in 443 – 450. prostate cancer. Eur J Cancer Prev 1999; 8: 525 – 532.

Prostate Cancer and Prostatic Diseases Genetic polymorphisms in prostate cancer risk TM Beer et al

34 Murata M et al. Genetic polymorphisms in cytochrome P450 38 Schlesselman J. Case – Control Studies: Design, Conduct, Analysis 27 (CYP) 1A1, CYP1A2, CYP2E1, glutathione S-transferase (GST) Oxford University Press: New York, 1982. M1 and GSTT1 and susceptibility to prostate cancer in the 39 Thompson MW, McInnes RR, Willard HF, Thompson JS. Thomp- Japanese population. Cancer Lett 2001; 165: 171 – 177. son & Thompson Genetics in Medicine 5th ed. W.B. Saunders Co: 35 Kawajiri K et al. The CYP1A1 gene and cancer susceptibility. Crit Philadelphia, 1991. Rev Oncol Hematol 1993; 14:77– 87. 40 Pienta KJ, Esper PS. Risk factors for prostate cancer. Ann Intern 36 Kawasaki ES. Sample preparation from blood, cells, and other Med 1993; 118: 793 – 803. fluids. In: Innis MA, Gelfand DH, Sninsky JJ, White TJ (eds). PCR 41 Adler V et al. Regulation of JNK signaling by GSTp. EMBO J 1999; Protocols: A Guide to Methods and Application. Academic Press: San 18: 1321 – 1334. Diego, 1990, pp 146 – 152. 42 Kauvar LM, Morgan AS, Sanderson PE, Henner WD. Glutathione 37 Comstock KE, Sanderson BJ, Claflin G, Henner WD. GST1 gene based approaches to improving cancer treatment. Chem Biol deletion determined by polymerase chain reaction. Nucleic Acids Interact 1998; 111/112: 225 – 238. Res 1990; 18: 3670.

Prostate Cancer and Prostatic Diseases