Identification of miR-30b-3p and miR-30d-5p as direct regulators of androgen receptor signaling in prostate cancer by complementary functional microRNA library screening.
The Androgen Receptor (AR) plays a key role in prostate biology and in the progression of prostate cancer (PCa) to castration resistance. The role of microRNAs (miRNAs) in aberrant AR signaling have not been fully characterized. Here we screened a library of 810 miRNA mimics to identify miRNAs that alter AR activity in complementary functional assays including protein lysate microarray (LMA) quantification of AR and PSA protein levels, AR transcriptional reporter activity, and AR-positive PCa cell viability. Candidate AR-regulating miRNAs were verified through AR transcriptional reporter and cell viability assays. MiRNA binding sites were found within the AR 3'-untranslated region (UTR) and within the AR and AR-V7 coding regions. MiRNA activity was characterized by western blotting, 3'-UTR reporter assay, and AR-GFP and AR-V7-GFP reporter assays. Results uncovered miR-30 family members as direct AR inhibitors. Inhibition of endogenous miR-30b-3p and miR-30d-5p enhanced AR expression and androgen-independent cell growth. Droplet digital RT-PCR quantification of miR-30c-5p and miR-30d-5p revealed significantly reduced levels in metastatic castration resistant PCa (CRPC), when compared to healthy prostate tissues. MiR-30d-5p levels were inversely correlated with AR activity, as measured by PSA mRNA, in metastatic CRPC. Collectively, these studies provide a comprehensive evaluation of AR-regulating miRNAs in PCa.
The James Buchanan Brady Urologic Institute and Department of Urology, Johns Hopkins School of Medicine, Baltimore, MD, USA.
Binod
B
Kumar
The James Buchanan Brady Urologic Institute and Department of Urology, Johns Hopkins School of Medicine, Baltimore, MD, USA.
Salar
S
Khaleghzadegan
The James Buchanan Brady Urologic Institute and Department of Urology, Johns Hopkins School of Medicine, Baltimore, MD, USA.
Brian
B
Mears
The James Buchanan Brady Urologic Institute and Department of Urology, Johns Hopkins School of Medicine, Baltimore, MD, USA.
Koji
K
Hatano
The James Buchanan Brady Urologic Institute and Department of Urology, Johns Hopkins School of Medicine, Baltimore, MD, USA.
Tarana A
TA
Kudrolli
The James Buchanan Brady Urologic Institute and Department of Urology, Johns Hopkins School of Medicine, Baltimore, MD, USA.
Wasim H
WH
Chowdhury
The James Buchanan Brady Urologic Institute and Department of Urology, Johns Hopkins School of Medicine, Baltimore, MD, USA.
David B
DB
Yeater
The James Buchanan Brady Urologic Institute and Department of Urology, Johns Hopkins School of Medicine, Baltimore, MD, USA.
Charles M
CM
Ewing
The James Buchanan Brady Urologic Institute and Department of Urology, Johns Hopkins School of Medicine, Baltimore, MD, USA.
Jun
J
Luo
The James Buchanan Brady Urologic Institute and Department of Urology, Johns Hopkins School of Medicine, Baltimore, MD, USA.|The department of Oncology, Sidney Kimmel Comprehensive Cancer Center, School of Medicine, Johns Hopkins University, Baltimore, MD, USA.
William B
WB
Isaacs
The department of Oncology, Sidney Kimmel Comprehensive Cancer Center, School of Medicine, Johns Hopkins University, Baltimore, MD, USA.
Luigi
L
Marchionni
The James Buchanan Brady Urologic Institute and Department of Urology, Johns Hopkins School of Medicine, Baltimore, MD, USA.|The department of Oncology, Sidney Kimmel Comprehensive Cancer Center, School of Medicine, Johns Hopkins University, Baltimore, MD, USA.
Shawn E
SE
Lupold
D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000502:physiology; D015723:Gene Library; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D008297:Male; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism*; D011471:Prostatic Neoplasms / Q000235:genetics* / Q000378:metabolism*; D064129:Prostatic Neoplasms, Castration-Resistant / Q000235:genetics* / Q000378:metabolism*; D011944:Receptors, Androgen / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D014162:Transfection
C410798:AR protein, human; C546086:MIRN30b microRNA, human; D035683:MicroRNAs; D011944:Receptors, Androgen
miR-30; miRNA; microRNA; androgen receptor; prostate cancer; castration resistant prostate cancer
Oncogene. 2012 Oct 11;31(41):4460-71
22266859
Cancer Res. 2013 Feb 1;73(3):1232-44
23233736
Cancer Res. 2009 Jan 1;69(1):16-22
19117982
Br J Cancer. 2013 Apr 30;108(8):1668-76
23571738
Br J Cancer. 2011 Mar 29;104(7):1168-77
21407217
Cancer Res. 2011 Mar 1;71(5):1956-67
21343391
Cancer Res. 2009 Apr 15;69(8):3245-8
19351814
JAMA. 2005 Jul 13;294(2):238-44
16014598
Cell. 2005 Jan 14;120(1):15-20
15652477
N Engl J Med. 2004 Oct 7;351(15):1488-90
15470210
Cancer Res. 2008 Mar 15;68(6):1777-85
18339858
Genome Biol. 2010;11(8):R90
20799968
Eur Urol. 2015 Oct;68(4):581-8
25556023
Mol Endocrinol. 1991 Dec;5(12):1921-30
1724287
Oncogene. 2013 Sep 26;32(39):4694-701
23085757
Carcinogenesis. 2015 Aug;36(8):858-66
25969144
Cancer Cell. 2010 Jul 13;18(1):11-22
20579941
Nucleic Acids Res. 2015 Apr 30;43(8):4075-86
25845598
Nat Cell Biol. 2015 Sep;17(9):1145-57
26302406
Oncogene. 2014 May 8;33(19):2495-503
23728339
Eur Urol. 2016 Jun;69(6):1120-8
26489476
Nat Genet. 1995 Apr;9(4):401-6
7795646
Br J Cancer. 2015 Sep 29;113(7):1055-65
26325107
Cancer Res. 2009 Mar 15;69(6):2305-13
19244107
Oncotarget. 2012 Nov;3(11):1455-71
23231923
Oncogene. 2016 May 5;35(18):2345-56
26364608
Mol Ther. 2007 Aug;15(8):1495-503
17565351
Science. 2009 Nov 27;326(5957):1230
19933109
Nat Rev Clin Oncol. 2014 Jun;11(6):365-76
24840076
Horm Cancer. 2014 Aug;5(4):207-17
24798453
CA Cancer J Clin. 1972 Jul-Aug;22(4):232-40
4625049
Cancer Res. 2009 Sep 15;69(18):7165-9
19738047
Int J Cancer. 2011 Aug 15;129(4):810-9
21710544
EMBO Mol Med. 2012 Mar;4(3):143-59
22351564
Cancer Treat Rev. 2014 Apr;40(3):426-33
24139549
Proc Natl Acad Sci U S A. 1988 Nov;85(21):8151-5
3186717
Cancer Res. 2007 Jul 1;67(13):6130-5
17616669
Mol Cell Biochem. 2013 May;377(1-2):121-30
23417242
Anal Biochem. 2000 Jun 15;282(1):158-61
10860516
Mol Endocrinol. 1997 Feb;11(2):148-61
9013762
Oncogene. 2013 Aug 29;32(35):4130-8
23069658
Cancer Res. 2008 Jul 1;68(13):5469-77
18593950
J Clin Invest. 2010 Aug;120(8):2715-30
20644256
Nat Rev Urol. 2015 Mar;12(3):137-44
25666893
PLoS One. 2012;7(5):e36535
22590558
Eur Urol. 2016 Aug;70(2):312-22
26806656
Adv Pharmacol. 2014;70:327-66
24931201
Mol Med Rep. 2015 Jun;11(6):4625-32
25673182
Science. 2008 Dec 12;322(5908):1695-9
19008416
Nat Genet. 2010 Aug;42(8):668-75
20601956