UROLOGICAL ONCOLOGY Role of Steroid Hormone Receptors in Formation and Progression of Bladder Carcinoma: A Case-Control Study Rahil Mashhadi,1 Gholamreza Pourmand,1* Farid Kosari,2 Abdolrasoul Mehrsai,1 Sepehr Salem,1 Mohammad Reza Pourmand,3 Sudabeh Alatab,1 Mehdi Khonsari,1 Fariba Heydari,1 Laleh Beladi,1 Farimah Alizadeh1 Purpose: To compare the expression rate of sex steroid hormone receptors of estrogen (ER), progesterone (PR) and androgen (AR) in normal urothelium and urothelial bladder cancer (UBC) and to evaluate the possible associations of these receptors expression with cancer progression and patient’s survival. Materials and Methods: We evaluated the clinical data and tumor specimens of 120 patients with pathologically confirmed primary UBC with 132 normal healthy controls. Both patients and controls selected from list of subjects who have been referred to Sina Urology clinic, and had a minimum of one year follow-up duration. Data collected from medical cords. For evaluation of expression, immunohistochemistry was performed on paraffin-embedded tissue sections using a monoclonal antibody for androgen, estrogen and progesterone receptors. Presence of at least 10% positive cells defined as positive expression. Results: None of the control subjects showed AR expression, while 22% of the patients were AR-positive. ER/PR expressions were observed in 4.2%/ and 2.5% of the cases and in 2.3% and 1.5% of the controls, respectively. A sta- tistically significant correlation was found between AR expression and tumor stage and grade (P < .001). AR-positive patients showed a significantly poorer prognosis than AR-negative cases (log-rank test, P = .02, hazard ratio = 2.12; 95% confidence interval: 1.36-4.65). Conclusion: AR expression was significantly associated with higher grade and poorly differentiated tumors with unfavorable outcome. AR expression test might be useful as a diagnostic tool for determining the malignancy and outcome of UBC patients. Keywords: receptors; androgen; estrogen; progesterone; tumor markers; biological; urinary bladder neoplasms; mor- tality; neoplasm recurrence; gene expression regulation; survival rate. INTRODUCTION Urothelial bladder cancer (UBC) is one of the most common cancers, as it is ranked the 9th most common cancer worldwide.(1) UBC is responsible for the death of 130,000 people annually worldwide(2) and its incidence is 3 times higher in men than women.(1) This cancer ranked the 7th most common cancer in men and the 17th most common in women.(1,3) UBC is the fourth most incident cancer in the USA and it is the 5th most common cancer in Iran.(1,3,4) According to the 2008 report of Iran’s National Cancer Registry, the incidence of bladder cancer was 13.03 in males and 3.32 in females per 100,000 population.(5) Interestingly, in addition to dissimilarity in incidence, the tumor behavior is also different between two sex- es. The female subjects tend to have more aggressive tumors with less favorable prognosis than male sub- jects(6-8) Although the exact origin of this difference between the genders is unknown, it is assumed that part of this variation comes from higher exposure of male subjects to industrial, environmental and occupational chemicals and also tobacco use.(9,10) However higher incidence in males cannot be fully explained solely by above men- tioned factors.(9) A study by Mir and colleagues showed that, even after adjustment for carcinogenic factors, sex-associated differences in UBC risk was still exist.(10) Some studies suggested the hormonal factors as a po- tential explanation for the gender disparity in the inci- dence and behavior of bladder cancer.(9) To support this hypothesis, some experimental animal studies showed that development of chemically induced UBC was less in female than in male animals.(10) Moreover, these an- 1 Urology Research Center, Tehran University of Medical Sciences, Tehran, Iran. 2 Department of Pathology, Sina Hospital, Tehran University of Medical Science, Tehran, Iran. 3 Department of Pathobiology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran. *Correspondence: Hassan-Abad Square, Sina Hospital, Urology Research Center, Tehran, Iran. Tel: +98 21 6634 8560. Fax: +98 21 6634 8560. E-mail: gh_pourmand@yahoo.com. Received June 2014 & Accepted November 2014 Urological Oncology 1968 imal studies generally demonstrate that hormonal ma- nipulation changes the natural course of the tumors and that animals who have been treated with androgen inhib- itors had better survival and more benign courses.(11,12) In agreement with these observations, some epidemiologic studies demonstrated that postmenopausal women have a greater risk for development of UBC as well as breast cancer than premenopausal women.(13,14) On the basis of these findings, sex steroid hormones and subsequent- ly their receptors have been considered as a potential explanation for the different biologic behavior of UBC between men and women.(10) Sex steroid hormones act by binding to their receptors including androgen receptors (ARs), estrogen receptors (ERs) and progesterone receptors (PRs) in target cells. (7) It is well known that steroid hormone receptors are expressed in normal bladder urothelium,(15,16) although physiological functions of these receptors in the blad- der are not completely understood.(17,18) On the other hand, it has been reported that the expression of ARs, ERs and PRs play an essential role during development, growth and progression of several malignancies.(9,19-22) Shen and colleagues showed that ERs are expressed in human bladder cancers and their expressions augment with increase of the stage and grade of cancer. Their results also demonstrated a strong inhibitory effect of antiestrogen treatment on UBC growth in vitro.(23) Miy- amoto and colleagues used N-butyl-N-(4-hydroxybutyl) nitrosamine to induce bladder cancer in both wild-type and ARs knockout male and female mice, and showed that 92% of the wild-type male, 42% of the wild-type female mice and none of the ARs knockout male and female mice developed tumors.(12) Despite the importance of steroid hormone receptors in the initiation, progression and outcome of bladder can- cer in experimental animal studies, its role in human UBC is still controversial.(24) In this regard, in present re- search we aimed to further clarify the role of sex steroid receptors (AR, PR and ER) expression in development and progression of UBC in human subjects and to in- vestigate whether or not there is an association between grade of the cancer and sex steroid receptor expression. To our knowledge this is the first study from our region in which the role of these receptors in outcome and prognosis of patients with UBC has been addressed. MATERIALS AND METHODS Study Subjects and Tissue Specimens This is a retrospective case-control study, in which 252 subjects including 120 pathologically confirmed UBC patients and 132 non-UBC individuals were recruited. The study protocol was approved by Medical Ethics Committee of Tehran University of Medical Sciences. Participants were drawn from the list of patients who attended the Urology clinic at Sina Hospital, and had a bladder specimen through either transurethral resection of bladder tumor (TURBT), cystectomy or cystoscopy. According to the sample size calculation and the litera- ture review, there should be 133 subjects in each group, however due to nature of retrospective studies, we only could recruit 120 subjects in case group and 132 sub- jects in control group which provides a ratio of control/ case of 1.1/1. All of the subjects were between 18-85 years old. In- dividuals who had pathologically confirmed UBC were placed in patient group. Controls were subjects who did not have pathologically confirmed UBC and have been referred to urology clinic because of other causes such as hematuria, benign prostatic hyperplasia (BPH), urethral stricture, bladder stones, pelvic trauma or chronic cys- titis. Patients who had concomitant or previous malig- nancies, or a history of hormonal therapy, chemotherapy and/or radiotherapy were excluded from the study. Two groups were sex matched. The anthropometric charac- teristics of patients and medical history were obtained from medical records. Definition for diabetes mellitus was use of anti-diabetic medication or at last two fast blood sugar levels of higher than 120 mg/dL. Defini- tion for hypertension was use of anti-hypertensive med- ication or the average blood pressure in two readings before admission was > 140/90 mmHg. Patients were considered to have dyslipidemia when the serum total cholesterol level was ≥ 200 mg/dL, high density lipo- protein cholestrol was < 40 mg/dL, or triglyceride was ≥ 150 mg/dL. Patients who were taking lipid lowering medications were also categorized in this group. Smok- ers were those who had smoked at least 100 cigarettes in their lifetime, while those who consumed less than 100 cigarettes were defined as non-smokers. Tissue specimens were examined by blinded pathol- ogists to determine the grade, stage and other histo- pathological characteristics. Grading of samples was performed according to the World Health Organization/ International Society of Urologic Pathology classifi- cation of urothelial neoplasia.(25) Pathological T stage (depth of invasion) was assessed according to American Joint Committee on Cancer Classification.(26) Immunohistochemistry Immunohistochemical (IHC) staining was performed on fixed paraffin embedded (3 µm) sections. Briefly, after deparaffinization in xylene and rehydration by graded concentrations of alcohol to distilled water, the specimens were washed with phosphate buffered saline (PBS). Endogenous peroxidase was blocked by 10-15 min incubation of specimens in 5% H 2 O 2 . The spec- imens were washed again with PBS and then antigen retrieval was performed in citrate buffer under 126°C and 2 atmosphere for 30 min. After washing with PBS, in order to decrease nonspecific antibody binding, pro- tein blocking was carried out by incubation in protein block serum-free (code X0909, Dako, Glostrup, Den- mark) for 10 min at room temperature. The sections were then incubated with the following primary anti- bodies: anti-estrogen receptor (clone 1D5, 1/50 dilu- tion, Dako, Glostrup, Denmark), anti-androgen receptor (clone AR441, 1/50 dilution, Dako, Glostrup, Denmark) and anti-progesterone receptor (clone PgR 636, 1/50 di- lution, Dako, Glostrup, Denmark). After washing with PBS, the slides were incubated with a dextran polymer reagent conjugated with peroxidase and secondary anti- body (Envision+, Dako, Glostrup, Denmark) for staining detection for 1 h. We also used 3,3’-diaminobenzidine as a chromogen for color development and subsequently counterstained them with Carazzi’s hematoxylin. Role of Steroid Hormone Receptors in Bladder Carcinoma-Mashhadi et al Vol 11. No 06 Nov-Dec 2014 1969 Immunostained sections were evaluated under a light microscope by two experienced pathologists in blinded fashion. The immunoreactivity was scored on a four- point scale as follows: negative ( <10% of cells with nuclear staining), weak (10-50% of cells with nuclear staining), moderate (51-80% of cells with nuclear stain- ing) and strong (> 80% of cells with nuclear staining). Statistical Analysis For statistical analyses, we used Statistical Package for the Social Science (SPSS Inc, Chicago, Illinois, USA) version 21. Data were presented as mean ± SD. Fisher’s exact test, chi-square test and the independent sample t-test were used to investigate the association between steroid hormone receptors expression with pathological and clinical factors. In all statistical analyzes, P values less than .05 was considered as statistically significant. RESULTS The mean age of the patients was 66.2 ± 12.10 years in case group and 60.4 ± 15.54 years in control group (P = .001). Over all the majority of subjects (85.7%, n = 216) were male. This rate in case and control groups was 87.5% (n = 105) and 84% (n = 111), respectively, however the ratio of male to female in case and con- trol groups was not significantly different (P = .47). The mean follow-up period was 24.5 months (range, 12-60 months). Figure 1 shows examples of AR/PR/ER positive cases and four-point scale of staining score is shown in Figure 2. Evaluation of steroid hormone re- ceptor expression revealed that only AR expression is significantly different between case and control groups (P = .0001). Moreover, we could not find any significant difference between AR, PR and ER expression in two genders in both case and control groups. The anthropo- metric and clinical characteristics of patients as well as steroid hormone receptor expression are presented in Table 1. Clinical and demographic characteristics of patients with UBC are summarized in Table 2. We also assessed the possible associations between steroid hormone receptors expression and tumor re- currence, tumor progression, tumor metastasis, tumor grades and stages, death because of UBC and family history of UBC. We found that only AR expression had a significant association with tumor stage (P < .001) and Variables Control (n = 132) Case (n= 120) P Value Age (year), mean ± SD 60.4 ± 15.54 66.2 ± 12.10 .001 Male/female 111/21 (84/16) 105/15 (87.5/12.5) .47 Smokers 67 (50.8) 73 (60.8) .12 Hypertension 43 (32.6) 52 (43.3) .09 Hyperlipidemia 7 (5.3) 15 (12.5) .04 Diabetes mellitus 35 (26.5) 42 (35) .17 Family history of cancer 1 (0.8) 5 (4.2) .10 AR expression 0 26 (21.7) <.0001 PR expression 3 (2.3) 5 (4.2) .48 ER expression 2 (1.5) 3 (2.5) .67 Table 1. Anthropometric characteristics and steroid hormone expression in study groups.* Abbreviations: AR, androgen receptor; PR, progesterone receptor; ER, estrogen receptor. * Data are presented as no (%). Variables Values Stage T1 61 (50.8) T2 21 (17.5) T3 18 (15) T4 20 (16.7) Grade Low 20 (16.7) High 100 (83.3) Recurrence Yes 57 (47.5) No 63 (52.5) Metastasis Yes 14 (11.7) No 106 (88.3) Mortality Yes 10 (8.3) No 110 (91.7) Chemotherapy Yes 44 (36.7) No 76 (63.3) Radiotherapy Yes 11 (9.2) No 109 (90.8) Table 2. Tumor characteristics (n = 120).* * Data are presented as no (%). Figure 1. Immunohistochemical staining of steroid hormone receptors in primary bladder cancer. (A) Estrogen receptor, (B) progesterone receptor, (C) androgen receptor. Role of Steroid Hormone Receptors in Bladder Carcinoma-Mashhadi et al Urological Oncology 1970 tumor grade (P < .0001). Moreover, the rate of AR ex- pression was higher in patients with family history of UBC (P = .04). Interestingly, AR/PR-positive patients had a higher rate of metastasis in comparison to AR/ PR-negative patients (P < .05). Also, AR-positive pa- tients showed a poorer prognosis than AR-negative cas- es (log rank test, P = .08), while survival was not affect- ed by PR/ER expression. DISCUSSION The incidence of bladder cancer is higher in men (about three times) than women.(1) It seems that this difference is primarily due to a difference in chemical exposure and smoking.(1,9) However, animal studies have shown that chemically and spontaneous bladder cancer de- velopment is significantly higher in male rats than fe- males.(12) Epidemiological studies have also shown that the development of bladder cancer in postmenopausal women is more common than women who are premeno- pausal.(13) It has been shown that in bladder cancer cells with AR-positive, cell growth is promoted by andro- gens,(12,16,18) or in another study, it has been found that antiestrogens can inhibit urothelial carcinoma of the bladder in ER-positive bladder cancer cell lines.(23) In another study on breast cancer, PR and ER were consid- ered as prognostic factors which play a role in the iden- tification of patients who may benefit from hormonal therapy.(14) It should be noted that previous studies that assess the relationship between AR/PR/ER expression and his- topathological characteristics of the tumors have led to conflicting results.(7) So, the results of such studies cannot be clearly identified prognostic significance of AR/PR/ER expression in patients with bladder cancer. Variability seen in results of such studies could be due to differences in sample size, study methods or interpre- tation of the results.(7) Moreover, there is no sufficient or strong evidence to establish epidemiological links be- tween steroid hormone receptors and observed gender differences in development of cancer.(24) In the present study, we found that AR/PR/ER expres- sions were similar in both sexes with UBC. Similar re- sults have also been reported in other studies.(24,27-29) In this study there is no correlation between AR/PR/ER ex- pression and gender differences in subjects with UBC. In a study by Kirkali and colleagues loss of AR expres- sion in malignant bladder urothelium is reported. They concluded that AR did not have a direct role in malig- nant transformation.(30) Boorjian and colleagues reported that loss of AR expression might lead to invasive blad- der cancer as they found a decreased AR expression in high stage tumors; also they observed AR expression in 53% of urothelial carcinoma and in 86% of normal urothelium cases.(27) Another study showed a significant decrease in the expression of AR in bladder cancer com- pared to nonneoplastic bladder specimens.(7) In present study AR was positive in 22% of the 120 patients with BC, but none of the 132 normal urotheliums showed AR-positivity as observed in the studies by Tuygun and colleagues,(24) and Ruizeveld de Winter and colleagues. (31) Birtle and colleagues studied AR expression in 17 cases of high grade transitional cell carcinoma (TCC) of the bladder. They showed that AR staining was nega- tive in all areas of normal urothelium, although AR was positive in 9/17 (52%) cases.(32) Although, Zhuang and colleagues reported nuclear immunoreactivity of AR in 7/9 (77%) urinary bladder cancers, they failed to detect positive immunohistochemical staining in normal uri- nary bladder. They mentioned that AR expression can be used as a diagnostic marker.(17) According to the available data, the prognostic role of the AR expression in bladder cancer is controversial. Tuygun and colleagues reported a significant decrease in AR expression in higher grades and invasive tumors, which is consistent with the findings of Boorjian and colleagues and Miyamoto and colleagues.(17,24,27) In contrast, Mir and colleagues in a study involving 472 patients, showed that AR-positivity was higher in mus- cle–invasive tumors (15%) compared to non-muscle invasive one (9%).(10) In another study with 33 superfi- cial bladder cancers, authors reported that patients with high AR expression tended to have a higher recurrence rate, compared to patients with low AR expression.(12) In present study, we found a significant correlation be- tween AR expression and high grade and high stage tu- mors (P < .0001 and P < .001, respectively). Also, the present study demonstrated a significantly higher rate of metastasis in AR-positive patients compared to AR-neg- ative patients (P = .009). Moreover, relapse-free surviv- al in AR-positive patients was lower than AR-negative patients (log-rank test, P = .08). Therefore AR could be used as a prognostic factor (hazard ratio: 2.12; 95% con- fidence interval: 1.36-4.65). In this study, PR and ER were positive in 2.5% and 4.2% of the UBC specimens, respectively. Our results confirmed that PR/ER expression is not associated with aggressiveness of UBC. Similar to our findings, Bolenz and colleagues reported that PR expression cannot be a prognostic factor in patients with UBC. In their study PR was not expressed in any of the UBC specimens.(9) In a study by Basakci and colleagues ER was positive in 12.4% of the superficial TCC specimens. They conclud- ed that ER does not play any direct role on the prognosis of superficial bladder TCC.(33) Also in our study, ER and PR expression do not have any direct roles in formation and progression of UBCs. This study has some limitations. First, since this study was a retrospective one, some of the data were not avail- Figure 2. Staining score of androgen receptor in primary bladder cancer. (A) < 10% positive tumor cells, (B) 10-50% positive tumor cells, (C) 51-80% positive tumor cells, (D) > 80% positive tumor cells. Role of Steroid Hormone Receptors in Bladder Carcinoma-Mashhadi et al Vol 11. No 06 Nov-Dec 2014 1971 able. Also, due to complex nature of the malignancies, there might be some confounders and effect modifiers that might interfere with the obtained results. We tried to control and limit the bias by using the blinded patholo- gist and including the control subjects from same popu- lation cohort. Finally two groups were not age matched. CONCLUSION We concluded that there was no significant difference in steroid hormone receptors expression between two sexes. Of studied steroid hormone receptors, only AR expression had significant association with stage and grade of bladder cancer. Based on study results, AR could be used as a prognostic factor in UBC. ACKNOWLEDGEMENTS This research has been sponsored by Tehran University of Medical Sciences, Tehran, Iran. The authors wish to thank Mrs. B. Pourmand for valuable helps in this study. CONFLICT OF INTEREST None declared. REFERENCES 1. Yavari P, Sadrolhefazi B, Mohagheghi MA, Mehrazin R. A descriptive retrospective stu- dy of bladder cancer at a hospital in Iran (1973-2003). Asian-Pac J Cancer Prev. 2009;10:681-4. 2. Karimianpour N, Mousavi-Shafaei P, Ziaee AA, et al. Mutations of RAS Gene Family in Specimens of Bladder Cancer. Urol J. 2008;5:237-42. 3. Ploeg M, Aben KK, Kiemeney LA. The pres- ent and future burden of urinary bladder can- cer in the world. World J Urol. 2009;27:289- 93. 4. Nanda MS, Sameer AS, Syeed N, et al. Gen- etic aberrations of the K-RAS proto-oncoge- ne in bladder cancer in Kashmiri population. Urol J. 2010;7:168-173. 5. Report of National Cancer Registration Iran. Is lamic Republic of Iran. Ministry of Health and Medical Education, Office Health Depu- ty, Center for Disease Control and Preven- tion. 2007-2008. 6. Scosyrev E, Noyes K, Feng C, Messing E. Sex and racial differences in bladder cancer presentation and mortality in the US. Cancer. 2009;115:68-74. 7. Miyamoto H, Yao J.L, Chaux A, et al. Expr- ession of androgen and oestrogen receptors and its prognostic significance in urothelial neoplasm of the urinary bladder. BJU Int. 2012;109:1716-26. 8. Jemal A, Siegel R, Xu J, Ward E. Cancer sta- tistics,2010. CA Cancer J Clin. 2010;60:277- 300. 9. Bolenz C, Lotan Y, Ashfaq R, Shariat SF. Es- trogen and progesterone hormonal receptor expression in urothelial carcinoma of the bladder. Eur Urol. 2009;56:1093-5. 10. Mir C, Shariat SF, Van der Kwast TH, et al. Loss of androgen receptor expression is not associated with pathological stage, grade, gender or outcome in bladder cancer: a large multi-institutional study. BJU Int. 2011;108:24-30. 11. Nam JK, Park SW, Lee SD, Chung MK. Prognostic Value of Sex-Hormone Receptor Expression in Non-Muscle-Invasive Bladder Cancer. Yonsei Med J. 2014;55:1214-21. 12. Miyamoto H, Yang Z, Chen YT, et al. Pro- motion of bladder cancer development and progression by androgen receptor signals. J Natl Cancer Inst. 2007;99:558-68. 13. McGrath M, Michaud DS, De Vivo I. Hor- monal and reproductive factors and the risk of bladder cancer in women. Am J Epi- demiol. 2006;163:236. 14. Fortner RT, Eliassen AH, Spiegelman D, Willett WC, Barbieri RL, Hankinson SE. Premenopausal endogenous steroid hor- mones and breast cancer risk: results from the Nurses’Health Study II. Breast Cancer Research. 2013;15:R19. 15. Chavalmane AK, Comeglio P, Morelli A, et al. Sex steroid receptors in male human blad- der: expression and biological function. J Sex Med. 2010;7:2698-713. 16. Liu Z, Li X, Liu S, et al. Flavokawain A inhib- its urinary bladder carcinogenesis in the UP II-SV40T transgenic mouse bladder cancer model. Proceedings of the 103rd Annual Me- eting of the American Association for Cancer Research. Abstract. 2012;619. 17. Zhuang Y-H, Blauer M, Tammela T, Tuohimaa P. Immunodetection of androgen receptor in human urinary bladder cancer. Histopathology. Histopathology. 1997;30: 556-62. 18. Johnson AM, O’Connell MJ, Miyamoto H, et al. Androgenic dependence of exophytic tu- mor growth in a transgenic mouse model of bladder cancer: a role for thrombospodin-1. B MC. Urol. 2008;8:7. 19. Lamb CA, Helguero LA, Giulianelli S, et al. Antisense oligonucleotides targeting the pro- gesterone receptor inhibit hormone-indepen- dent breast cancer growth in mice. Breast Cancer Res. 2005;7:R1111-21. 20. Rahmani AH, Alzohairy M, Babiker AY, Khan AA, Aly SM, Rizvi MA. Implication of androgen receptor in urinary bladder cancer: a critical mini-review. Int J Mol Epidemiol Genet. 2013;4:150-5. 21. Chang C, Lee SO, Yeh S, Chang TM. Andro- gen receptor (AR) differential roles in hor- mone-related tumors including prostate, Role of Steroid Hormone Receptors in Bladder Carcinoma-Mashhadi et al Urological Oncology 1972 bladder, kidney, lung, breast and liver. Onco gene. 2014;33:3225-34. 22. Li Y, Izumi K, Miyamoto H. The role of the androgen receptor in the development and progression of bladder cancer. Jpn J Clin On- col. 2012;42:569-77. 23. Shen SS, Smith CL, Hsieh JT, et al. Expression of estrogen receptors-alpha and -beta in bladder cancer cell lines and human bladder tumor tissue. Cancer.2006;106:2610-6. 24. Tuygun C, Kankaya D, Imamoglu A, et al. Sex-specific hormone receptors in urothelial carcinomas of the human urinary bladder: a comparative analysis of clinicopathological features and survival outcomes according to receptor expression. Urol Oncol. 2011;29: 43–51. 25. Epstein JI, Amin MB, Reuter VR, Mostofi FK. The World Health Organization/Inter national Society of Urological Pathology consensus classification of urothelial (transi- tional cell) neoplasms of the urinary bladder. Bladder Consensus Conference Committee. Am J Surg Pathol. 1998;22:1435-48. 26. Greene FL, Page DL, Fleming ID, et al. AJCC Cancer Staging Manual, 6th ed. New York (NY): Springer-Verlag, 2002. 27. Boorjian S, Ugras S, Mongan NP, et al. An- drogen receptor expression is inversely cor- related with pathologic tumor stage in blad- der cancer. Urology. 2004;64:383-8. 28. Boorjian SA, Heemers HV, Frank I, et al. Ex- pression and significance of androgen recep- tor coactivators in urothelial carcinoma of the bladder. Endocr Relat Cancer. 2009;16:123- 37. 29. Kauffman EC, Robinson BD, Downes MJ, et al. Role of androgen receptor and associated lysine-demethylase coregulators, LSD1 and JMJD2A, in localized and advanced human bladder cancer. Mol Carcinog. 2011;50:931- 44. 30. Kirkali Z, Cowan S, Leake RE. Androgen receptors in transitional cell carcinoma. Int Urol Nephrol. 1990;22:231-4. 31. Ruizeveld de Winter JA, Trapman J, Vermey M, Mulder E, Zegers ND, van der Kwast TH. Androgen receptor expression in human tis- sues: An immunohistochemical study. J His tochem Cytochem. 1991;39:927-36. 32. Birtle AJ ,Freeman A, Munson P. The andro- gen receptor revisited in urothelial carcino- ma. Histopathology. 2004;45:P98-9. 33. Basakci A , Kirkali Z , Tuzel E , Yorukoglu K , Mungan MU , Sade M. Prognostic signif- icance of estrogen receptor expression in superficial transitional cell carcinoma of the urinary bladder. Eur Urol. 2002;41:342-5. Role of Steroid Hormone Receptors in Bladder Carcinoma-Mashhadi et al Vol 11. No 06 Nov-Dec 2014 1973