Sulforaphane increases the efficacy of anti-androgens by rapidly decreasing androgen receptor levels in prostate cancer cells

Khurana,Namrata; Talwar,Sudha; Chandra,Partha   K.; Sharma,Pankaj; Abdel-Mageed,Asim  B.; Mondal,Debasis; Sikka,Suresh   C.

doi:10.3892/ijo.2016.3641

Sulforaphane increases the efficacy of anti-androgens by rapidly decreasing androgen receptor levels in prostate cancer cells

Authors:
- Namrata Khurana
- Sudha Talwar
- Partha K. Chandra
- Pankaj Sharma
- Asim B. Abdel-Mageed
- Debasis Mondal
- Suresh C. Sikka
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Affiliations: Department of Urology, Tulane University School of Medicine, New Orleans, LA 70112, USA, Department of Pharmacology, Tulane University School of Medicine, New Orleans, LA 70112, USA, Amity Institute of Biotechnology, Amity University, Noida, U.P. 201313, India
Published online on: August 2, 2016 https://doi.org/10.3892/ijo.2016.3641
Pages: 1609-1619

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Abstract

Prostate cancer (PCa) cells utilize androgen for their growth. Hence, androgen deprivation therapy (ADT) using anti-androgens, e.g. bicalutamide (BIC) and enzalutamide (ENZ), is a mainstay of treatment. However, the outgrowth of castration resistant PCa (CRPC) cells remains a significant problem. These CRPC cells express androgen receptor (AR) and utilize the intratumoral androgen towards their continued growth and invasion. Sulforaphane (SFN), a naturally occurring isothiocyanate found in cruciferous vegetables, can decrease AR protein levels. In the present study, we tested the combined efficacy of anti-androgens and SFN in suppressing PCa cell growth, motility and clonogenic ability. Both androgen-dependent (LNCaP) and androgen-independent (C4-2B) cells were used to monitor the effects of BIC and ENZ, alone and in combination with SFN. Co-exposure to SFN significantly (p<0.005) enhanced the anti-proliferative effects of anti-androgens and downregulated expression of the AR-responsive gene, prostate specific antigen (PSA) (p<0.05). Exposure to SFN decreased AR protein levels in a time- and dose-dependent manner with almost no AR detected at 24 h with 15 µM SFN (p<0.005). This rapid and potent AR suppression by SFN occurred by both AR protein degradation, as suggested by cycloheximide (CHX) co-exposure studies, and by suppression of AR gene expression, as evident from quantitative RT-PCR experiments. Pre-exposure to SFN also reduced R1881-stimulated nuclear localization of AR, and combined treatment with SFN and anti-androgens abrogated the mitogenic effects of this AR-agonist (p<0.005). Wound-healing assays revealed that co-exposure to SFN and anti-androgens can significantly (p<0.005) reduce PCa cell migration. In addition, long-term exposures (14 days) to much lower concentrations of these agents, SFN (0.2 µM), BIC (1 µM) and/or ENZ (0.4 µM) significantly (p<0.005) decreased the number of colony forming units (CFUs). These findings clearly suggest that SFN may be used as a promising adjunct agent to augment the efficacy of anti-androgens against aggressive PCa cells.

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1	Yap TA, Zivi A, Omlin A and de Bono JS: The changing therapeutic landscape of castration-resistant prostate cancer. Nat Rev Clin Oncol. 8:597–610. 2011. View Article : Google Scholar : PubMed/NCBI
2	Hodgson MC, Bowden WA and Agoulnik IU: Androgen receptor footprint on the way to prostate cancer progression. World J Urol. 30:279–285. 2012. View Article : Google Scholar :
3	Chang KH, Ercole CE and Sharifi N: Androgen metabolism in prostate cancer: From molecular mechanisms to clinical consequences. Br J Cancer. 111:1249–1254. 2014. View Article : Google Scholar : PubMed/NCBI
4	Ryan CJ, Smith A, Lal P, Satagopan J, Reuter V, Scardino P, Gerald W and Scher HI: Persistent prostate-specific antigen expression after neoadjuvant androgen depletion: An early predictor of relapse or incomplete androgen suppression. Urology. 68:834–839. 2006. View Article : Google Scholar : PubMed/NCBI
5	Harris WP, Mostaghel EA, Nelson PS and Montgomery B: Androgen deprivation therapy: Progress in understanding mechanisms of resistance and optimizing androgen depletion. Nat Clin Pract Urol. 6:76–85. 2009. View Article : Google Scholar : PubMed/NCBI
6	Godbole AM and Njar VC: New insights into the androgen-targeted therapies and epigenetic therapies in prostate cancer. Prostate Cancer. 2011:9187072011. View Article : Google Scholar : PubMed/NCBI
7	Kim W and Ryan CJ: Androgen receptor directed therapies in castration-resistant metastatic prostate cancer. Curr Treat Options Oncol. 13:189–200. 2012. View Article : Google Scholar : PubMed/NCBI
8	Liu C, Lou W, Zhu Y, Yang JC, Nadiminty N, Gaikwad NW, Evans CP and Gao AC: Intracrine androgens and AKR1C3 activation confer resistance to enzalutamide in prostate cancer. Cancer Res. 75:1413–1422. 2015. View Article : Google Scholar : PubMed/NCBI
9	Schrader AJ, Schrader MG and Cronauer MV: Words of wisdom. Re: Androgen receptor splice variants mediate enzalutamide resistance in castration-resistant prostate cancer cell lines. Eur Urol. 64:169–170. 2013. View Article : Google Scholar : PubMed/NCBI
10	Arora VK, Schenkein E, Murali R, Subudhi SK, Wongvipat J, Balbas MD, Shah N, Cai L, Efstathiou E, Logothetis C, et al: Glucocorticoid receptor confers resistance to anti-androgens by bypassing androgen receptor blockade. Cell. 155:1309–1322. 2013. View Article : Google Scholar : PubMed/NCBI
11	Joseph JD, Lu N, Qian J, Sensintaffar J, Shao G, Brigham D, Moon M, Maneval EC, Chen I, Darimont B, et al: A clinically relevant androgen receptor mutation confers resistance to second-generation anti-androgens enzalutamide and ARN-509. Cancer Discov. 3:1020–1029. 2013. View Article : Google Scholar : PubMed/NCBI
12	Korpal M, Korn JM, Gao X, Rakiec DP, Ruddy DA, Doshi S, Yuan J, Kovats SG, Kim S, Cooke VG, et al: An F876L mutation in androgen receptor confers genetic and phenotypic resistance to MDV3100 (enzalutamide). Cancer Discov. 3:1030–1043. 2013. View Article : Google Scholar : PubMed/NCBI
13	Kawata H, Ishikura N, Watanabe M, Nishimoto A, Tsunenari T and Aoki Y: Prolonged treatment with bicalutamide induces androgen receptor overexpression and androgen hypersensitivity. Prostate. 70:745–754. 2010. View Article : Google Scholar : PubMed/NCBI
14	Colabufo NA, Pagliarulo V, Berardi F, Contino M, Inglese C, Niso M, Ancona P, Albo G, Pagliarulo A and Perrone R: Bicalutamide failure in prostate cancer treatment: Involvement of Multi Drug Resistance proteins. Eur J Pharmacol. 601:38–42. 2008. View Article : Google Scholar : PubMed/NCBI
15	Bohl CE, Gao W, Miller DD, Bell CE and Dalton JT: Structural basis for antagonism and resistance of bicalutamide in prostate cancer. Proc Natl Acad Sci USA. 102:6201–6206. 2005. View Article : Google Scholar : PubMed/NCBI
16	Saad F, Adachi JD, Brown JP, Canning LA, Gelmon KA, Josse RG and Pritchard KI: Cancer treatment-induced bone loss in breast and prostate cancer. J Clin Oncol. 26:5465–5476. 2008. View Article : Google Scholar : PubMed/NCBI
17	Salturk Z, Çakır O, Kumral TL, Yıldırım G, Ötünçtemur A, Aydoğdu Ï and Uyar Y: Subjective and objective effects of androgen ablation therapy on voice. J Voice. 29:490–493. 2015. View Article : Google Scholar : PubMed/NCBI
18	McCarty MF, Hejazi J and Rastmanesh R: Beyond androgen deprivation: Ancillary integrative strategies for targeting the androgen receptor addiction of prostate cancer. Integr Cancer Ther. 13:386–395. 2014. View Article : Google Scholar : PubMed/NCBI
19	Lamont KR and Tindall DJ: Minireview: Alternative activation pathways for the androgen receptor in prostate cancer. Mol Endocrinol. 25:897–907. 2011. View Article : Google Scholar : PubMed/NCBI
20	Brooke GN and Bevan CL: The role of androgen receptor mutations in prostate cancer progression. Curr Genomics. 10:18–25. 2009. View Article : Google Scholar : PubMed/NCBI
21	Armstrong CM and Gao AC: Drug resistance in castration resistant prostate cancer: Resistance mechanisms and emerging treatment strategies. Am J Clin Exp Urol. 3:64–76. 2015.PubMed/NCBI
22	Elbarbry F and Elrody N: Potential health benefits of sulforaphane: A review of the experimental, clinical and epidemiological evidences and underlying mechanisms. J Med Plants Res. 5:473–484. 2011.
23	Zhang Y and Tang L: Discovery and development of sulforaphane as a cancer chemopreventive phytochemical. Acta Pharmacol Sin. 28:1343–1354. 2007. View Article : Google Scholar : PubMed/NCBI
24	Clarke JD, Dashwood RH and Ho E: Multi-targeted prevention of cancer by sulforaphane. Cancer Lett. 269:291–304. 2008. View Article : Google Scholar : PubMed/NCBI
25	Cheung KL and Kong AN: Molecular targets of dietary phenethyl isothiocyanate and sulforaphane for cancer chemoprevention. AAPS J. 12:87–97. 2010. View Article : Google Scholar :
26	Traka MH, Melchini A and Mithen RF: Sulforaphane and prostate cancer interception. Drug Discov Today. 19:1488–1492. 2014. View Article : Google Scholar : PubMed/NCBI
27	Keum YS, Khor TO, Lin W, Shen G, Kwon KH, Barve A, Li W and Kong AN: Pharmacokinetics and pharmacodynamics of broccoli sprouts on the suppression of prostate cancer in transgenic adenocarcinoma of mouse prostate (TRAMP) mice: Implication of induction of Nrf2, HO-1 and apoptosis and the suppression of Akt-dependent kinase pathway. Pharm Res. 26:2324–2331. 2009. View Article : Google Scholar : PubMed/NCBI
28	Shapiro TA, Fahey JW, Dinkova-Kostova AT, Holtzclaw WD, Stephenson KK, Wade KL, Ye L and Talalay P: Safety, tolerance, and metabolism of broccoli sprout glucosinolates and isothiocyanates: a clinical phase I study. Nutr Cancer. 55:53–62. 2006. View Article : Google Scholar : PubMed/NCBI
29	Petri N, Tannergren C, Holst B, Mellon FA, Bao Y, Plumb GW, Bacon J, O’Leary KA, Kroon PA, Knutson L, et al: Absorption/metabolism of sulforaphane and quercetin, and regulation of phase II enzymes, in human jejunum in vivo. Drug Metab Dispos. 31:805–813. 2003. View Article : Google Scholar : PubMed/NCBI
30	Singh SV, Srivastava SK, Choi S, Lew KL, Antosiewicz J, Xiao D, Zeng Y, Watkins SC, Johnson CS, Trump DL, et al: Sulforaphane-induced cell death in human prostate cancer cells is initiated by reactive oxygen species. J Biol Chem. 280:19911–19924. 2005. View Article : Google Scholar : PubMed/NCBI
31	Xiao D, Powolny AA, Antosiewicz J, Hahm ER, Bommareddy A, Zeng Y, Desai D, Amin S, Herman-Antosiewicz A and Singh SV: Cellular responses to cancer chemopreventive agent D,L-sulforaphane in human prostate cancer cells are initiated by mitochondrial reactive oxygen species. Pharm Res. 26:1729–1738. 2009. View Article : Google Scholar : PubMed/NCBI
32	Pei Y, Wu B, Cao Q, Wu L and Yang G: Hydrogen sulfide mediates the anti-survival effect of sulforaphane on human prostate cancer cells. Toxicol Appl Pharmacol. 257:420–428. 2011. View Article : Google Scholar : PubMed/NCBI
33	Zhang C, Su ZY, Khor TO, Shu L and Kong AN: Sulforaphane enhances Nrf2 expression in prostate cancer TRAMP C1 cells through epigenetic regulation. Biochem Pharmacol. 85:1398–1404. 2013. View Article : Google Scholar : PubMed/NCBI
34	Schultz MA, Hagan SS, Datta A, Zhang Y, Freeman ML, Sikka SC, Abdel-Mageed AB and Mondal D: Nrf1 and Nrf2 transcription factors regulate androgen receptor transactivation in prostate cancer cells. PLoS One. 9:e872042014. View Article : Google Scholar : PubMed/NCBI
35	Kensler TW, Egner PA, Agyeman AS, Visvanathan K, Groopman JD, Chen JG, Chen TY, Fahey JW and Talalay P: Keap1-nrf2 signaling: A target for cancer prevention by sulforaphane. Top Curr Chem. 329:163–177. 2013. View Article : Google Scholar :
36	Kim SH and Singh SV: D,L-Sulforaphane causes transcriptional repression of androgen receptor in human prostate cancer cells. Mol Cancer Ther. 8:1946–1954. 2009. View Article : Google Scholar : PubMed/NCBI
37	Wiczk A, Hofman D, Konopa G and Herman-Antosiewicz A: Sulforaphane, a cruciferous vegetable-derived isothiocyanate, inhibits protein synthesis in human prostate cancer cells. Biochim Biophys Acta. 1823:1295–1305. 2012. View Article : Google Scholar : PubMed/NCBI
38	Gibbs A, Schwartzman J, Deng V and Alumkal J: Sulforaphane destabilizes the androgen receptor in prostate cancer cells by inactivating histone deacetylase 6. Proc Natl Acad Sci USA. 106:16663–16668. 2009. View Article : Google Scholar : PubMed/NCBI
39	Wu HC, Hsieh JT, Gleave ME, Brown NM, Pathak S and Chung LW: Derivation of androgen-independent human LNCaP prostatic cancer cell sublines: Role of bone stromal cells. Int J Cancer. 57:406–412. 1994. View Article : Google Scholar : PubMed/NCBI
40	Uygur B and Wu WS: SLUG promotes prostate cancer cell migration and invasion via CXCR4/CXCL12 axis. Mol Cancer. 10:1392011. View Article : Google Scholar : PubMed/NCBI
41	Chou TC: Drug combination studies and their synergy quantification using the Chou-Talalay method. Cancer Res. 70:440–446. 2010. View Article : Google Scholar : PubMed/NCBI
42	Horoszewicz JS, Leong SS, Chu TM, Wajsman ZL, Friedman M, Papsidero L, Kim U, Chai LS, Kakati S, Arya SK, et al: The LNCaP cell line - a new model for studies on human prostatic carcinoma. Prog Clin Biol Res. 37:115–132. 1980.
43	Cunningham D and You Z: In vitro and in vivo model systems used in prostate cancer research. J Biol Methods. 2:pii. e172015. View Article : Google Scholar : PubMed/NCBI
44	Agoulnik IU, Vaid A, Bingman WE, Erdeme H, Frolov A, Smith CL, Ayala G, Ittmann MM and Weigel NL: Role of SRC-1 in the promotion of prostate cancer cell growth and tumor progression. Cancer Res. 65:7959–7967. 2005.PubMed/NCBI
45	He S, Zhang C, Shafi AA, Sequeira M, Acquaviva J, Friedland JC, Sang J, Smith DL, Weigel NL, Wada Y, et al: Potent activity of the Hsp90 inhibitor ganetespib in prostate cancer cells irrespective of androgen receptor status or variant receptor expression. Int J Oncol. 42:35–43. 2013.
46	Ai J, Wang Y, Dar JA, Liu J, Liu L, Nelson JB and Wang Z: HDAC6 regulates androgen receptor hypersensitivity and nuclear localization via modulating Hsp90 acetylation in castration-resistant prostate cancer. Mol Endocrinol. 23:1963–1972. 2009. View Article : Google Scholar : PubMed/NCBI
47	Myzak MC, Tong P, Dashwood WM, Dashwood RH and Ho E: Sulforaphane retards the growth of human PC-3 xenografts and inhibits HDAC activity in human subjects. Exp Biol Med (Maywood). 232:227–234. 2007.
48	Shiota M, Yokomizo A and Naito S: Oxidative stress and androgen receptor signaling in the development and progression of castration-resistant prostate cancer. Free Radic Biol Med. 51:1320–1328. 2011. View Article : Google Scholar : PubMed/NCBI
49	Demir A, Cecen K, Karadag MA, Kocaaslan R and Turkeri L: The course of metastatic prostate cancer under treatment. Springerplus. 3:7252014. View Article : Google Scholar
50	Lee JG, Zheng R, McCafferty-Cepero JM, Burnstein KL, Nanus DM and Shen R: Endothelin-1 enhances the expression of the androgen receptor via activation of the c-myc pathway in prostate cancer cells. Mol Carcinog. 48:141–149. 2009. View Article : Google Scholar
51	Mostaghel EA and Nelson PS: Intracrine androgen metabolism in prostate cancer progression: Mechanisms of castration resistance and therapeutic implications. Best Pract Res Clin Endocrinol Metab. 22:243–258. 2008. View Article : Google Scholar : PubMed/NCBI
52	Mostaghel EA, Page ST, Lin DW, Fazli L, Coleman IM, True LD, Knudsen B, Hess DL, Nelson CC, Matsumoto AM, et al: Intraprostatic androgens and androgen-regulated gene expression persist after testosterone suppression: Therapeutic implications for castration-resistant prostate cancer. Cancer Res. 67:5033–5041. 2007. View Article : Google Scholar : PubMed/NCBI
53	Huo C, Kao YH and Chuu CP: Androgen receptor inhibits epithelial-mesenchymal transition, migration, and invasion of PC-3 prostate cancer cells. Cancer Lett. 369:103–111. 2015. View Article : Google Scholar : PubMed/NCBI
54	Singh SV, Warin R, Xiao D, Powolny AA, Stan SD, Arlotti JA, Zeng Y, Hahm ER, Marynowski SW, Bommareddy A, et al: Sulforaphane inhibits prostate carcinogenesis and pulmonary metastasis in TRAMP mice in association with increased cytotoxicity of natural killer cells. Cancer Res. 69:2117–2125. 2009. View Article : Google Scholar : PubMed/NCBI
55	Aapro MS, Eliason JF, Krauer F and Alberto P: Colony formation in vitro as a prognostic indicator for primary breast cancer. J Clin Oncol. 5:890–896. 1987.PubMed/NCBI
56	Kallifatidis G1, Rausch V, Baumann B, Apel A, Beckermann BM, Groth A, Mattern J, Li Z, Kolb A, Moldenhauer G, et al: Sulforaphane targets pancreatic tumour-initiating cells by NF-κB-induced antiapoptotic signaling. Gut. 58:949–963. 2009. View Article : Google Scholar