The Pharmacogenomics Journal (2004) 4, 251–259 & 2004 Nature Publishing Group All rights reserved 1470-269X/04 $30.00 www.nature.com/tpj ORIGINAL ARTICLE A case-based evaluation of SRD5A1, SRD5A2, AR, and ADRA1A as candidate genes for severity of BPH M Klotsman1,2 ABSTRACT 3 In men with a clinical diagnosis of benign prostatic hyperplasia (BPH), CR Weinberg polytomous logistic regression analysis was conducted to evaluate associa- 1,4 K Davis tions between two silent polymorphisms in SRD5A1 (codon positions 30 and CG Binnie5 116), two polymorphisms in SRD5A2 (Val89Leu substitution and C to T KE Hartmann1 transition in intron 1), a trinucleotide (CAG)n repeat in androgen receptor (AR), and an Arg492Cys substitution in ADRA1A and clinical parameters that 1Department of Epidemiology, University of characterize severity of BPH. Candidate gene selection was based on two North Carolina School of Public Health, Chapel mechanistic pathways targeted by pharmacotherapy for BPH: (1) androgen 2 Hill, NC, USA; Genetics Research, metabolic loci contributing to prostate growth (static obstruction); and (2) GlaxoSmithKline, RTP, NC, USA; 3Biostatistics Branch, National Institute of Environmental factors affecting smooth muscle tone (dynamic obstruction). Polymorphisms Health Sciences, RTP, NC, USA; 4World-Wide in SRD5A2 were not associated with severity of BPH; however, SRD5A1 Epidemiology, GlaxoSmithKline, RTP, NC, USA; polymorphisms were associated with severity of BPH. The process(es) in 5 Discovery Genetics, GlaxoSmithKline, RTP, NC, which these silent single-nucleotide polymorphisms (SNPs) influence BPH USA phenotypes is unknown and additional studies will be needed to assess Correspondence: whether these SNPs have direct functional consequences. The characteriza- Dr M Klotsman, PO Box 13398, Five Moore tion of additional molecular factors that contribute to static and dynamic Dr, RTP, NC 27709-3398, USA. obstruction may help predict response to pharmacotherapy and serve to Tel: þ 1 919 483 5249 identify novel drug targets for the clinical management of BPH. Fax: þ 1 919 315 0311 E-mail: [email protected] The Pharmacogenomics Journal (2004) 4, 251–259. doi:10.1038/sj.tpj.6500248 Published online 11 May 2004 Keywords: BPH; prostate; testosterone 5-alpha-reductase; adrenergic alpha-an- tagonists INTRODUCTION Benign prostatic hyperplasia (BPH) is an extremely common condition among aging men that typically manifests as a constellation of lower urinary tract symptoms (LUTS) which are bothersome, impair psychological and functional well-being, and interfere with activities of daily living. Prevalence estimates for BPH vary, but it is clear that BPH is nearly ubiquitous in older men and is a significant health concern.1 Histologic evidence of BPH was found in almost 90% of men in their 80s,2 and it is estimated that nearly one in four men receive treatment for LUTS by the eighth decade of life.3 Despite significant morbidity caused by BPH, the etiology and determinants of severity of this condition remain poorly understood. Received: 24 November 2003 The current BPH literature suggests that LUTS generally result from static Revised: 27 January 2004 Accepted: 19 February 2004 (mechanical) and/or dynamic component(s). The static component is a Published online 11 May 2004 consequence of the benign enlargement of the prostate, which increases pressure Evaluation of candidate genes for severity of BPH M Klotsman et al 252 on the urethra with subsequent obstruction of urine flow. RESULTS Prostate growth is regulated, in part, by a defined androgen- Baseline Distributions dependent pathway,4 which begins with the diffusion of Genetic data were available for 87% of the study participants serum testosterone (TES) into the prostate. In the prostatic (n ¼ 2428 out of 2802) for whom baseline data were available stroma, TES is irreversibly converted to the more active (Table 1). Genotypes were missing primarily because of dihydrotestosterone (DHT) by the enzyme 5a-reductase type failed assays or subject refusal to submit a blood sample for II. Converted DHT then enters epithelial cells and selectively genetic analysis. The baseline characteristics for the subset binds to the androgen receptor (AR), a ligand-dependent of men in whom genetic data were unavailable appeared transcription factor, forming a DHT/AR complex. This similar to those of the study group as a whole (data not complex is translocated into the nucleus and promotes shown). prostatic growth by binding to androgen response elements in the regulatory region of target genes5,6 (eg, androgen regulation of prostate-specific antigen (PSA) expression is mediated by AR).7 Table 1 Baseline clinical parameters and genotype distribu- By contrast, dynamic obstruction is caused by dysfunction tions among 2201 Caucasians of smooth muscle tone in the prostatic capsule surrounding Parameter Mean (7SD) the prostate. Prostatic smooth muscle tone varies according to the sympathetic stimulation of a -adrenoceptors, in 1 Age (yr) 66.577.6 a 8–10 particular, the 1A-adrenoceptor subtype. In a healthy PSA (ng/ml) 3.872.0 male, smooth muscle in the capsule contracts to propel AUA-SI 17.276.0 7 prostatic secretions from storage into the prostatic urethra Qmax (ml/s) 10.4 3.6 during ejaculation. In a BPH disease state, increased smooth Post-void residual volume 79775 muscle tone in the outer prostatic capsule is believed to Total prostate volume (cm3) 54.9723.0 bring on LUTS by increasing urethral resistance. Conse- Transition zone volume (cm3) 26.8717.0 quently, the dynamic component of BPH is determined, at Serum testosterone (pg/ml) 392871489 7 least partially, by a -adrenoceptor activity and prostatic Serum dihydrotestosterone 404 202 1A 7 smooth muscle tone.11 Body mass index 27.8 4.1 Systematic screens of genes encoding for 5a-reductase SRD5A1: Silent Arg 30a type I and II (SRD5A1 and SRD5A2), AR, and a -adreno- 1A G/G 625 (28.6%) ceptor (ADRA1A) have identified polymorphisms that may G/C 1104 (50.6%) influence protein production and/or integrity. These poly- C/C 454 (20.8%) morphic loci can be operationally compartmentalized into one of two pathways that contribute to the severity of BPH: SRD5A1: Silent Ala 116a (1) an androgen-dependent pathway, involving 5a-reductase A/A 874 (40.0%) and AR activity, contributing to static obstruction; and (2) a A/G 1004 (46.1%) G/G 304 (13.9%) non-growth-dependent pathway, mediated by a1A-adreno- ceptor, that may be instrumental in the onset of dynamic b obstruction. The polymorphisms evaluated in this study SRD5A2: Val89Leu Val/Val 1110 (50.9%) included two silent polymorphisms in SRD5A1 (codon 12 Val/Leu 885 (40.6%) positions 30 and 116), two polymorphisms in SRD5A2 Leu/Leu 184 (8.4%) 13,14 (Val89Leu and C to T transition in intron 1), a (CAG)n 15,16 repeat in AR, and Arg492Cys substitution in SRD5A2: Intron 1 C-Tb ADRA1A.17,18 C/C 1065 (49.7%) Pharmacotherapy with a-blockers and 5a-reductase inhi- C/T 880 (41.1%) bitors has become commonplace for the medical manage- T/T 196 (9.2%) ment of BPH.19 Alpha blockers alleviate dynamic obstruction by interrupting motor sympathetic adrenergic ADRA1A: Arg492Leu nerve stimulation in the prostate,20 while 5a-reductase Arg/Arg 630 (28.7%) Arg/Cys 1099 (50.1%) inhibitors limit DHT production resulting in involution of Cys/Cys 464 (21.2%) the prostate.21 Given the importance of a-blockers and 5a- reductase inhibitors in the treatment of BPH, this study was AR: (CAG)n repeats undertaken to evaluate associations between polymorph- (CAG)0–18 82 (6.0%) isms in candidate genes whose effects are targeted by (CAG)19–22 970 (43.9%) pharmacotherapy and clinical parameters used to character- (CAG)423 1103 (50.1%) ize the severity of BPH. Further characterization of poly- a morphic loci hypothesized to play a role in static and The SRD5A1 Arg30 and Ala116 SNPs were not strongly linked (correlation, R ¼ 0.6775). dynamic obstruction precipitated by BPH may help to shed bThe SRD5A2 V89L and Intron 1 C-T SNPs appear to be in linkage disequilibrium light on etiologic mechanisms predisposing men to BPH. (correlation, R ¼ 1). The Pharmacogenomics Journal Evaluation of candidate genes for severity of BPH M Klotsman et al 253 Allele and Genotype Frequencies tion were observed (Table 2). Chi-square statistics were re- Regression analyses performed to evaluate whether candi- calculated for men under the age of 60, to allow for the date genetic markers are associated with severity of BPH possibility that gene-by-gene interaction contributes to an were restricted to Caucasian subjects. The two single- earlier age of onset. In this younger group of men, statistical nucleotide polymorphisms (SNPs) in 5a-reductase type II significance was not reached for any of the gene-by-gene (SRD5A2) are physically close (intron 1 and exon 1) and pairs evaluated (data not shown). were in linkage disequilibrium (LD) (correlation, R ¼ 1). Results based on fitting logistic regression models to relate Concordant SRD5A2 genotype pairs were observed in 93% of three-level genotypes at each polymorphic loci and BPH- study participants, and predominant allele frequencies were related clinical parameters are shown in Table 3. Polymorph- nearly identical (71.2% and 70.3% for 89-Val and intron-C isms in 5a-reductase type II (SRD5A2) did not appear to be alleles, respectively). The SNPs in SRD5A1 were in partial LD associated with the severity of BPH; however, polymorph- (correlation, R ¼ 0.6775). These LD estimates are based on a isms in 5a-reductase type I were associated with severity. case-only population and may not be comparable to LD one The silent Arg30 polymorphism in 5a-reductase type I would observe in the general population. (SRD5A1) was associated with serum TES and DHT levels (P ¼ 0.03 and P ¼ 0.04, respectively), and there was limited Genetic Analysis evidence for an association with maximum urinary flow A total of eight gene-by-gene combinations were evaluated (Qmax; P ¼ 0.09), and post-void residual volume (PVR; to assess the possible synergistic genetic effects of pairs of P ¼ 0.10) (Table 3).