SRD5A gene polymorphism in Japanese men predicts prognosis of metastatic prostate cancer with androgen-deprivation therapy

European Journal of Cancer (2015) xxx, xxx– xxx

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SRD5A gene polymorphism in Japanese men predicts prognosis of metastatic prostate cancer with androgen-deprivation therapy Masaki Shiota a,1, Naohiro Fujimoto b,1, Akira Yokomizo a,⇑, Ario Takeuchi a, Momoe Itsumi a, Junichi Inokuchi a, Katsunori Tatsugami a, Takeshi Uchiumi c, Seiji Naito a a

Department of Urology, Graduate School of Medical Sciences, Kyushu University, Fukuoka 812-8582, Japan Department of Urology, School of Medicine, University of Occupational and Environmental Health, Kitakyushu 807-8556, Japan c Department of Clinical Chemistry and Laboratory Medicine, Graduate School of Medical Sciences, Kyushu University, Fukuoka 812-8582, Japan b

Received 30 January 2015; received in revised form 24 May 2015; accepted 17 June 2015

KEYWORDS Androgen-deprivation therapy Castration-resistant prostate cancer Prostate cancer SNP SRD5A

Abstract Aim: De novo androgen synthesis is thought to be involved in the progression to castration-resistant prostate cancer (CRPC) during androgen-deprivation therapy (ADT). During androgen synthesis, 5a-reductase encoded by SRD5A catalyses testosterone into more active dihydrotestosterone and may be involved in the progression to CRPC. Then, this study aimed to reveal the association between genetic variations in SRD5A and the prognosis in metastatic prostate cancer. Methods: We studied the polymorphisms rs518673 and rs166050 in SRD5A1, and rs12470143, rs523349, rs676033 and rs2208532 in SRD5A2 as well as the time to CRPC progression and overall survival in 104 patients with metastatic prostate cancer that had undergone primary ADT. The association between the polymorphisms and the progression to CRPC as well as overall survival was examined.

⇑ Corresponding author at: Department of Urology, Graduate School of Medical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-ku, Fukuoka 812-8582, Japan. Tel.: +81 92 642 5603; fax: +81 92 642 5618. E-mail address: [email protected] (A. Yokomizo). 1 These authors contributed equally to this work.

http://dx.doi.org/10.1016/j.ejca.2015.06.122 0959-8049/Ó 2015 Elsevier Ltd. All rights reserved.

Please cite this article in press as: Shiota M. et al., SRD5A gene polymorphism in Japanese men predicts prognosis of metastatic prostate cancer with androgen-deprivation therapy, Eur J Cancer (2015), http://dx.doi.org/10.1016/j.ejca.2015.06.122

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Results: Patients carrying the more active GG genotype in SRD5A2 rs523349 exhibited a higher risk of the progression (hazard ration [95% confidence interval], 1.93 [1.14–3.14], p = 0.016) and death (hazard ration [95% confidence interval], 2.14 [1.16–3.76], p = 0.016), compared with less active GC/CC genotypes in SRD5A2 rs523349. Conclusions: High 5a-reductase activity due to the polymorphism in SRD5A2 may contribute to resistance to ADT. Furthermore, SRD5A2 rs523349 polymorphism may be a promising biomarker for metastatic prostate cancer patients treated with primary ADT and a molecular target for advanced prostate cancer. Ó 2015 Elsevier Ltd. All rights reserved.

1. Introduction Prostate cancer is the most common non-cutaneous cancer among males and the second leading cause of cancer-caused death in developed countries [1]. Androgen-deprivation therapy (ADT) is the gold-standard therapy for recurrent or advanced prostate cancer [2]. Initially, most prostate cancers are dependent on androgens and respond well to ADT. However, most eventually recur in a castration-resistant manner during ADT, and are defined as castration-resistant prostate cancer (CRPC) [3]. Although several molecular mechanisms of castration resistance have been identified [4,5], de novo androgen synthesis is thought to be a common mechanism, under which most CRPCs are supplied with adequate androgen under castration conditions [6]. CYP17, a critical regulator of de novo androgenesis, is upregulated in CRPC [6,7]. The CYP17 inhibitor abiraterone acetate has shown significant survival benefit for patients with CRPC [8]. In addition, 5a-reductase encoded by the SRD5A gene is another critical enzyme in androgen metabolism, and converts testosterone to dihydrotestosterone (DHT), the most potent AR agonist in prostate cells [9]. The SRD5A genes SRD5A1 and SRD5A2 encode 5a-reductase type 1 and type 2, respectively. While SRD5A2 is dominantly expressed in non-malignant prostate tissues, SRD5A1 expression has been shown in prostate cancer [10,11]. Finasteride, the 5a-reductase type 2 inhibitor, and dutasteride, the dual 5a-reductase inhibitor for both 5a-reductase type 1 and type 2 enzymes, have been tested as chemo-preventative agents in the large randomised PCPT and REDUCE trials, respectively. Intriguingly, these trials have shown similar controversial results, in which the inhibitors reduced prostate cancer incidence, but increased the risk of aggressive prostate cancer [12,13]. In addition to prostate carcinogenesis, 5a-reductase is involved in prostate cancer progression. Several SRD5A1 and SRD5A2 single nucleotide polymorphisms (SNPs), including rs518673 and rs166050 in SRD5A1 and rs12470143, rs523349 (V89L; more active V variant encoded by G allele), rs676033, rs2208532 and rs4952197 in SRD5A2, were significantly or marginally associated with biochemical recurrence after radical

prostatectomy [14]. In the subsequent study, the above SNPs in SRD5A genes except rs4952197 were differentially correlated with androgen levels [15]. In the REDEEM trial, dutasteride was shown to reduce pathological prostate cancer progression or therapeutic intervention compared with active surveillance, suggesting that 5a-reductase functions in promoting the progression of localised prostate cancer [16]. The relevance of 5a-reductase in the progression to castration resistance remains unclear. The prediction of response to treatment and progression remains a challenge as prostate cancer is a heterogeneous disease. To predict the oncological outcome in patients with prostate cancer treated with primary ADT, J-CAPRA score utilising clinical TNM stage, prostatic-specific antigen (PSA) value at diagnosis and pathological grading by Gleason score has been developed and validated [17,18]. However, the predictive accuracy could be improved by incorporating other predictive parameters such as a genetic biomarker. Presently, the effect of SRD5A polymorphisms linked to progression and prognosis in advanced prostate cancer patients remains unknown. In this study, we examined the impact of the polymorphisms in SRD5A genes on the oncological outcome in metastatic prostate cancer patients. 2. Materials and methods 2.1. Patients All patients were histopathologically diagnosed with adenocarcinoma of the prostate. Clinical staging was determined according to the unified TNM criteria based on the results of a digital rectal examination, transrectal ultrasound, computed tomography, magnetic resonance imaging and bone scan [19]. Primarily ADT was conducted with surgical or medical castration using a luteinizing hormone-releasing hormone agonist (goserelin acetate or leuprorelin acetate) and/or an antiandrogen agent (bicalutamide, flutamide or chlormadinone acetate). The oncological outcome was defined as described previously [20]. Briefly, time to progression was defined as the duration from ADT initiation to the date of progression to CRPC. CRPC was defined according to three

Please cite this article in press as: Shiota M. et al., SRD5A gene polymorphism in Japanese men predicts prognosis of metastatic prostate cancer with androgen-deprivation therapy, Eur J Cancer (2015), http://dx.doi.org/10.1016/j.ejca.2015.06.122

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consecutive increments in PSA resulting in two 50% increases over the nadir despite consecutive ADT, or progression or appearance of two lesions on a bone scan and soft tissue lesions. In all of 59 (73.8%) men among 80 patients with CRPC, serum testosterone level was evaluated and confirmed as castrated level (<50 ng/dl). Overall survival was calculated from the ADT initiation to death or the last follow-up day. Blood samples were obtained from patients with prostate cancer that had undergone primary ADT from 1993 to 2005 when initially diagnosed as prostate cancer. Written informed consent was obtained from all patients. This study was performed in accordance with the principles described in the Declaration of Helsinki and the Ethical Guidelines for Epidemiological Research enacted by the Japanese Government, and approved by the institutional review board. 2.2. Polymorphism genotyping Genomic DNA was immediately extracted from whole blood of patients and stored at 80 °C until analysis. Then, rs518673 and rs166050 in SRD5A1, and rs12470143, rs523349, rs676033 and rs2208532 in SRD5A2 were selected as target SNPs based on previous reports [14,15]. Genotyping was performed on a CFX Connect Real-Time System (Bio-Rad, Hercules, CA, United States of America (USA)) with pre-designed Taqman SNP Genotyping Assays (Life Technologies, Carlsbad, CA, USA) for SRD5A1, rs518673 (C_9766953_10), rs10493113 (C_2935721_10) and SRD5A2, rs12470143 (C_345976_10), rs523349 (C_2362601_10), rs676033 (C_2362605_10), rs2208532 (C_2791128_10) and TaqMan Gene Expression Master Mix (Life Technologies Corporation), according to the manufacturer’s protocol. Genetic linkages between the gene polymorphisms in each gene were shown previously [14], indicating most linkage disequilibrium was less than 0.5, except linkage disequilibrium between rs523349 and rs676033, which was more than 0.9. 2.3. Statistical analysis Statistical analyses were performed using JMP9 software (SAS Institute, Cary, NC, USA). Comparisons between groups were analysed by Wilcoxon or Pearson tests. Survival analyses were conducted by Kaplan– Meier method and the log-rank test. Cox proportional hazards model was used to estimate the hazard ratios (HRs). The continuous data were assessed using the Student’s t-test. Levels of statistical significance were set at p < 0.05. 3. Results The oncological outcome in prostate cancer patients treated with primary ADT is extremely different between

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non-metastatic and metastatic prostate cancer patients, irrespective of the individual’s background. Therefore, we only analysed metastatic prostate cancer patients. Supplementary Table 1 shows the clinicopathological characteristics and treatments of the 104 patients analysed in this study. Median age was 73 years and PSA level at diagnosis was 244.0 ng/ml. Gleason score at biopsy was over seven and clinical stages were advanced stages (T3 and T4) in most patients. High PSA level at diagnosis, high Gleason score at biopsy and advanced clinical T-stage reflected the characteristics of metastatic prostate cancer. Fifty-one (56.0%) and 94 (90.4%) patients had regional lymph node and distant metastasis at diagnosis, respectively. Most patients (88.5%) were treated with combined androgen blockade using castration and antiandrogen agent as initial treatment. Of the 104 patients, 80 (76.9%) men progressed to CRPC and 60 (57.7%) men died during the median follow-up duration of 44.2 (interquartile range; IQR, 19.0–91.8) months. Among the investigated gene polymorphisms, as shown in Table 1, GG versus GC/CC in SRD5A2 rs523349 which causes a non-synonymous mutation in 5a-reductase type 2 (V89L) was significantly associated with high risk of progression to CRPC (HR [95% confidence interval; CI], 1.72 [1.07–2.72], p = 0.026). In addition, although AA versus AG/GG in SRD5A2 rs518673 was also marginally associated with high risk of progression to CRPC (HR [95% CI], 2.38 [0.91–5.10], p = 0.073), the number of AA alleles in SRD5A2 rs518673 was only six (5.6%), suggesting a lower utility in the clinical setting (Supplementary Table 2). Therefore, we focused on the major GG homozygote in SRD5A2 rs523349. As shown in Fig. 1A, the GC and CC alleles in SRD5A2 rs523349 were associated with longer progression-free survival during ADT (median [95% CI]; CC, 15 [7–33] months; GC, 24 [13–38] months), compared with the GG allele (median [95% CI]; GG, 11 [6–18] months) (p = 0.041; GG versus GC, p = 0.014; GG versus CC, p = 0.52; GC versus CC, p = 0.28). Next, the major homozygote versus other alleles and minor homozygote versus other alleles in SRD5A2 rs523349 were compared. The progression-free survival in the major GG homozygote was significantly worse compared with GC/CC alleles (median [95% CI]; GG, 11 [6–18] months; GC/CC, 19 [13–34] months, p = 0.019) (Fig. 1B, Supplementary Table 2). Conversely, the progression-free survival between the minor homozygote CC versus GG/GC was equivalent (median [95% CI]; CC, 15 [7–33] months; GG/GC, 18 [13–30] months, p = 0.63) (Fig. 1C, Supplementary Table 2). With regard to overall survival, as shown in Table 1, the major GG homozygotes in SRD5A2 rs523349 were significantly associated with high risk of all-caused death compared with GC/CC alleles (HR [95% CI], 1.86 [1.07–3.15], p = 0.029). As shown in Fig. 2A, the

Please cite this article in press as: Shiota M. et al., SRD5A gene polymorphism in Japanese men predicts prognosis of metastatic prostate cancer with androgen-deprivation therapy, Eur J Cancer (2015), http://dx.doi.org/10.1016/j.ejca.2015.06.122

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Table 1 The associations between clinicopathological parameters and progression-free survival or overall survival. Variable

Age Prostatic-specific antigen (PSA) at diagnosis Biopsy Gleason score <7 7 P8 Clinical stage cT2 cT3 cT4 Clinical stage N0 N1 Clinical stage M0 M1 Hormonal therapy Combined androgen blockade Castration rs523349 in SRD5A2 (V89L) GG GC/CC *

Progression-free survival

Overall survival

Hazard ratio (HR)

95% confidence interval (CI)

P-value

HR

95% CI

P-value

1.53 3.50

0.46–5.47 0.83–10.93

0.50 0.083

3.99 1.00

0.98–17.53 0.11–5.78

0.054 1.00

0 Ref 1.60

0–3.46 – 0.98–2.73

0.28 – 0.061

5.27 Ref 1.32

0.29–27.37 – 0.74–2.43

0.20 – 0.35

Ref 0.76 0.98

– 0.38–1.67 0.45–2.29

– 0.47 0.96

Ref 1.43 2.03

– 0.57–4.82 0.74–7.14

– 0.48 0.18

Ref 1.25

0.76–2.08

0.38

Ref 1.05

0.59–1.89

0.87

Ref 1.85

– 0.87–4.77

– 0.11

Ref 1.83

– 0.74–6.10

– 0.21

Ref 0.81

– 0.37–1.53

– 0.53

Ref 1.07

– 0.44–2.21

– 0.86

Ref 0.58

– 0.37–0.94

– 0.026*

Ref 0.54

– 0.32–0.94

– 0.029*

Statistically significant.

overall survival rate in CC, GC and GG alleles in SRD5A2 rs523349 were CC, 46 [14–98] months; GC, 95 [82–105] months; and GG, 31 [19–48] months (median [95% CI]), respectively (p = 0.022; GG versus GC, p = 0.0098; GG versus CC, p = 0.68; GC versus CC, p = 0.072). Next, the major homozygote versus other alleles and minor homozygote versus other alleles in SRD5A2 rs523349 were compared. The overall survival in the major homozygote GG was significantly worse than GC/CC (median [95% CI]: GG, 31 [19–48] months; GC/CC, 93 [51–102] months, p = 0.021) (Fig. 2B, Supplementary Table 3). Conversely, the overall survival between minor homozygote CC versus GG/GC was equivalent (median [95% CI]: CC, 46 [14–98] months; GC/GG, 84 [46–102] months, p = 0.29) (Fig. 2C, Supplementary Table 3). Thus, although the SRD5A2 rs523349 gene polymorphism was associated with progression and death risks, there was no difference in the background among gene polymorphisms in SRD5A2 rs523349 (Supplementary Table 1). Conversely, no other clinicopathological factor correlated with progression-free survival and overall survival, probably because of the small sample number (Table 1). Actually, SRD5A2 rs523349 polymorphism was more or equivalently potential prognosis factor of progression and survival than major clinicopathological factors such as PSA at diagnosis, Gleason score and clinical stage even on multivariate analysis (data not shown). Allele frequencies in the investigated SNPs are shown in Table 2. The GG, GC and CC genotypes in SRD5A2

rs523349 corresponded to 28.8%, 59.6% and 11.5% of patients, respectively. According to the Hardy– Weinberg principle, frequencies of the major G allele and minor C allele were 0.59 and 0.41, respectively. 4. Discussion Numerous mechanisms have been reported to be involved in the progression of hormone-naı¨ve prostate cancer to CRPC. Among these mechanisms, de novo androgen synthesis is considered one of major mechanisms promoting the progression to CRPC [6]. Although CYP17, HSD3B and AKR1C enzymes are critical regulators in de novo androgenesis, 5a-reductase also contributes to augmentation of androgen activity in prostate cancer cells under suppressed androgen milieu by castration. Consistent with this hypothesis, this study showed that the non-synonymous SRD5A2 rs523349 polymorphism correlated with durability of tumour response to ADT and overall survival in metastatic prostate cancer. To the best of our knowledge, this study is the first report on the effect of SRD5A polymorphisms in advanced prostate cancer. This study showed that the major GG homozygote in SRD5A2 rs523349 corresponding to the more active VV variant-type of 5a-reductase type 2 was a risk factor of the progression to CRPC and poor prognosis [21,22]. Previous studies demonstrated upregulated SRD5A1 and SRD5A2 expressions in CRPC [23–25]. With regard to intratumour androgen levels, increased testosterone and DHT were reported in CRPC xenograft tumours [24] and

Please cite this article in press as: Shiota M. et al., SRD5A gene polymorphism in Japanese men predicts prognosis of metastatic prostate cancer with androgen-deprivation therapy, Eur J Cancer (2015), http://dx.doi.org/10.1016/j.ejca.2015.06.122

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A

5

A rs523349 CC (n=12) GC (n=62) GG (n=30)

rs523349 CC (n=12) GC (n=62) GG (n=30) P = 0.041*

P = 0.022*

B

B rs523349 GC/CC (n=74) GG (n=30)

rs523349 GC/CC (n=74) GG (n=30)

P = 0.019*

P = 0.021*

C rs523349 CC (n=12) GC/GG (n=92)

C rs523349 CC (n=12) GC/GG (n=92)

P = 0.63

P = 0.29

Fig. 1. SRD5A gene polymorphism correlates with progression-free survival in metastatic prostate cancer that had undergone primary androgen-deprivation therapy. (A–C) Progression-free survival rate in patients with metastatic prostate cancer treated with primary androgen-deprivation therapy stratified by gene polymorphism (rs523349) in SRD5A2 gene is shown.

Fig. 2. Gene polymorphism in SRD5A correlates with overall survival in metastatic prostate cancer treated with primary androgen-deprivation therapy. (A–C) Overall survival rate in patients with metastatic prostate cancer undergone primary androgen-deprivation therapy stratified by gene polymorphism (rs523349) in SRD5A2 gene is shown.

CRPC tissues [6], although increased testosterone and decreased DHT and 5a-reductase activity in CRPC cells were also reported [25]. Thus, although the findings remain controversial, these data suggest that more active 5a-reductase promotes cellular resistance to ADT through augmented synthesis of about 10-fold higher DHT compared to testosterone. In addition, our results show that progression and prognosis in the LL variant-type was equivalent or worse, but not significant, than the more active VL variant-type, suggesting that the threshold effect brings such similarity between VL and LL variant-types. Otherwise, the worse prognosis in LL variant-type patients may derive from the intrinsic

aggressive phenotype as shown in localised cancer [14,26,27], less androgen dependency because of lower 5a-reductase activity, or higher rate of castration monotherapy, which is less effective than combined androgen blockade [28] among LL variant-type cases. Our findings suggested that inhibition of 5a-reductase may prevent the progression to CRPC in metastatic prostate cancer. The dual 5a-reductase inhibitor dutasteride effectively prevented prostate cancer tumour growth combined with castration in a mouse xenograft model [29]. In addition, dutasteride combined with the novel anti-androgen enzalutamide could suppress cancer proliferation in hormone-naı¨ve cancer as well as CRPC

Please cite this article in press as: Shiota M. et al., SRD5A gene polymorphism in Japanese men predicts prognosis of metastatic prostate cancer with androgen-deprivation therapy, Eur J Cancer (2015), http://dx.doi.org/10.1016/j.ejca.2015.06.122

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Table 2 Allele frequency of SRD5A polymorphisms. Variable gene

SNP

SRD5A1

SRD5A2

Major/minor allele

Alleles

rs518673

G/A

rs166050

A/G

rs12470143

C/T

rs523349 (V89L)

G/C (V/L)

GG GA AA AA AG GG CC CT TT GG

48 (46.2%) 50 (48.1%) 6 (5.8%) 91 (87.5%) 12 (11.5%) 1 (1.0%) 37 (35.6%) 52 (50.0%) 15 (14.4%) 30 (28.8%)

CG CC CC CT TT AA AG GG

62 12 33 61 10 25 62 17

rs676033

C/T

rs2208532

A/G

n (%)

(59.6%) (11.5%) (31.7%) (58.7%) (9.6%) (24.0%) (59.6%) (16.3%)

Major/minor allele frequencies This study

HapMap, CEPH

HapMap, YRI

HapMap, JPT

HapMap, CHB

0.70/0.30

0.66/0.34

0.75/0.25

0.78/0.22

0.72/0.28

0.93/0.07

0.78/0.22

Not available

0.89/0.11

0.86/0.14

0.61/0.39

0.51/0.49

0.66/0.34

0.62/0.38

0.73/0.27

0.59/0.41

0.81/0.19

0.77/0.23

0.64/0.36

0.52/0.48

0.61/0.39

0.80/0.20

0.75/0.25

0.61/0.39

0.56/0.44

0.54/0.46

0.61/0.39

0.70/0.30

0.59/0.41

0.53/0.47

SNP, single-nucleotide polymorphism.

[30]. Although this hypothesis has not been investigated in humans, it should be tested in the future. In addition, 5a-reductase type 3 was recently shown to be expressed in benign and malignant prostate [23,30], and overexpressed in CRPC tissues [31]. Therefore, a 5a-reductase inhibitor targeting all isoforms might be a more powerful modality combined with castration, and this hypothesis should be examined in preclinical studies. The SRD5A2 rs523349 genetic polymorphism may be a promising biomarker predicting progression and prognosis among patients treated with primary ADT for metastatic prostate cancer. Studies have shown that persistent circulating androgen in serum leads to higher probabilities of recurrence and death in patients treated with primary ADT [32,33]. Although the prediction ability of the newly developed J-CAPRA score has been validated [18], Incorporation of SRD5A2 rs523349 into the J-CAPRA score might help improve diagnostic accuracy in cases treated with primary ADT. Furthermore, the improved prediction of treatment failure in primary ADT would also help identify patients requiring more intensive treatment in addition to conventional ADT monotherapy such as up-front docetaxel chemotherapy combined with ADT, as indicated by the CHAARTED study [34]. In addition, since novel therapeutic agent CYP17 inhibitor abiraterone acetate substantially suppresses de novo androgen synthesis, more intensive hormone therapy by up-front use of abiraterone acetate with conventional ADT in hormone-naı¨ve state may improve the prognosis of metastatic prostate cancer, which is under investigation in STAMPEDE trial [35]. Also, SRD5A2 rs523349 may help identify putative candidates for targeting

therapeutics using 5a-reductase inhibitor. Because SNP analysis is an established and less invasive technique, analysis of SRD5A2 rs523349 is feasible for use in clinical setting. This study also showed relative low GG allele frequency of SRD5A2 rs523349 in the Japanese population with metastatic prostate cancer that was consistent with previous reports indicating a lower rate of GG allele frequency in Asian populations compared with that in Caucasians and African Americans. So far, several studies have reported the survival of Asians has been shown to be better than that of Caucasians or African Americans [36–38]. Based on the result in this study, the different frequencies of the functional SRD5A2 rs523349 genotype in different ethnic populations may partially explain the ethnic disparities of ADT response. This study includes several limitations. The sample size was relatively small, and the study is retrospective and included only a Japanese population. Validation would require analyses using a larger sample size including other ethnic groups. In addition, multiple analyses on six gene polymorphisms in this study could lead to false results by multiple comparisons. Also, since the patients in this study were treated in pre-abiraterone era, employing abiraterone acetate may diminish the prognostic impact of SRD5A2 polymorphism because of substantial suppressive effect of abiraterone acetate on androgen synthesis. This study did not determine serum and intratumour androgen levels, which provide proof of concept that the activity of 5a-reductase affects the efficiency of ADT. So far, several studies have reported no significant change in serum testosterone levels between genotypes in SRD5A2 rs523349 [15,39–42], and no report has examined serum DHT levels as well as intratumour

Please cite this article in press as: Shiota M. et al., SRD5A gene polymorphism in Japanese men predicts prognosis of metastatic prostate cancer with androgen-deprivation therapy, Eur J Cancer (2015), http://dx.doi.org/10.1016/j.ejca.2015.06.122

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testosterone and DHT levels. Therefore, further study investigating the differences of these levels between the SRD5A2 rs523349 polymorphisms is required. In conclusion, the SRD5A2 rs523349 polymorphism was associated with recurrence and prognosis in patients with metastatic prostate cancer that underwent primary ADT possibly by altering androgen metabolism. Then, the SRD5A2 rs523349 polymorphism could be a novel biomarker predicting the prognosis of ADT. Also, these findings help to unravel the mechanism of castration resistance, and suggest that it may be informative to examine SRD5A2 polymorphism among patients in randomised trials comparing enhanced ADT versus standard ADT.

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Funding This work was supported by Kakenhi grants (25462483, 25462484 and 26861273) from The Ministry of Education, Culture, Sports, Science, and Technology of Japan, Japan; National Cancer Center Research and Development Fund (H26-A-4), Japan; and Researcher Promotion Grant from Japanese Urological Association, Japan.

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[13]

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Conflict of interest statement None declared.

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Acknowledgements We would like to thank Edanz Group Japan for editorial assistance as well as Ms. Noriko Hakoda and Ms. Eriko Gunshima for technical assistance.

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Appendix A. Supplementary data Supplementary data associated with this article can be found, in the online version, at http://dx.doi.org/10. 1016/j.ejca.2015.06.122. References [1] Siegel R, Ma J, Zou Z, Jemal A. Cancer statistics, 2014. CA Cancer J Clin 2014;64:9–29. [2] Sharifi N, Gulley JL, Dahut WL. An update on androgen deprivation therapy for prostate cancer. Endocr Relat Cancer 2010;17:R305–15. [3] Sadar MD. Small molecule inhibitors targeting the “achilles’ heel” of androgen receptor activity. Cancer Res 2011;71:1208–13. [4] Shiota M, Yokomizo A, Naito S. Increased androgen receptor transcription: a cause of castration-resistant prostate cancer and a possible therapeutic target. J Mol Endocrinol 2011;47:R25–41. [5] Shiota M, Yokomizo A, Fujimoto N, Naito S. Androgen receptor cofactors in prostate cancer: potential therapeutic targets of castration-resistant prostate cancer. Curr Cancer Drug Targets 2011;11:870–81. [6] Montgomery RB, Mostaghel EA, Vessella R, et al. Maintenance of intratumoral androgens in metastatic prostate cancer: a

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Please cite this article in press as: Shiota M. et al., SRD5A gene polymorphism in Japanese men predicts prognosis of metastatic prostate cancer with androgen-deprivation therapy, Eur J Cancer (2015), http://dx.doi.org/10.1016/j.ejca.2015.06.122

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Please cite this article in press as: Shiota M. et al., SRD5A gene polymorphism in Japanese men predicts prognosis of metastatic prostate cancer with androgen-deprivation therapy, Eur J Cancer (2015), http://dx.doi.org/10.1016/j.ejca.2015.06.122