TY - JOUR
T1 - Analysis of PMEPA1 isoforms (A and b) as selective inhibitors of androgen and tgf-β signaling reveals distinct biological and prognostic features in prostate cancer
AU - Sharad, Shashwat
AU - Sztupinszki, Zsófia M.
AU - Chen, Yongmei
AU - Kuo, Claire
AU - Ravindranath, Lakshmi
AU - Szallasi, Zoltan
AU - Petrovics, Gyorgy
AU - Sreenath, Taduru L.
AU - Dobi, Albert
AU - Rosner, Inger L.
AU - Srinivasan, Alagarsamy
AU - Srivastava, Shiv
AU - Cullen, Jennifer
AU - Li, Hua
N1 - Funding Information:
Author Contributions: conceptualization, S.S. (Shashwat Sharad), S.S. (Shiv Srivastava) and H.L.; design, S.S. (Shashwat Sharad), S.S. (Shiv Srivastava), A.S., T.L.S., and H.L.; methodology, S.S. (Shashwat Sharad), Z.M.S., the whole blot figures. C.K., Y.C., L.R., and H.L.; validation, L.R., S.S. (Shashwat Sharad) and H.L.; formal analysis, S.S. (Shashwat Sharad), Z.M.S., Y.C., C.K., and H.L.; investigation, S.S. (Shashwat Sharad), J.C., and H.L.; resources, S.S. (Shiv Srivastava), J.C., I.L.R., A.D., G.P.; data curation, I.L.R, Z.S., J.C., S.S. (Shiv Srivastava); writing—original draft C.K., Y.C., L.R., and H.L.; validation, L.R., S.S. (Shashwat Sharad) and H.L.; formal analysis, S.S. (Shashwat preparation, S.S. (Shashwat Sharad) and H.L.; writing—review and editing, A.D., A.S., and H.L., ; visualization, J.C., S.S. (Shashwat Sharad), H.L., A.S., S.S. (Shiv Srivastava), H.L.; supervision, S.S. (Shashwat Sharad), and pHr.eLp.a; rpartoiojenc,tS a.Sd.m(SihnaissthrwatiaotnS,h Aar.aDd.,) aGn.Pd.,H S.L.S.;. w(Srhitiivn gS—rivreavstiaevwa)a, nAd.eSd., itJi.nCg.,, AS..SD. .(,SAh.aSs.,hawnadt HSh.La.r;avdis) uaanlidzaHtio.Ln.;, J.C., S.S. (Shashwat Sharad), H.L., A.S., S.S. (Shiv Srivastava), H.L.; supervision, S.S. (Shashwat Sharad), and H.L.; project administration, A.D., G.P., S.S. (Shiv Srivastava), A.S., J.C., S.S. (Shashwat Sharad) and H.L.; funding aFcuqnudisiintigo:n T,hIi.Ls .wRo.,rakn hdaSs .bSe. e(Snh siuvpSproivratesdta bvya )f.unding to Center for Prostate Disease Research, Uniformed Services University for the Health Sciences (HU0001-17-2-2019 and HU0001-10-2-0002 to I. Rosner and S. Srivastava) from Funding: This work has been supported by funding to Center for Prostate Disease Research, Uniformed Services University for the Health Sciences (HU0001-17-2-2019 and HU0001-10-2-0002 to I. Rosner and S. Srivastava) from tahnedo MffiacteeorifeCl Conogmrmesasinodn a(UllySADMireRcMteCd)M. edical Research Programs (CDMRP) of the US Army Medical Research and Materiel Command (USAMRMC). Acknowledgments: The authors thank the patients who participated in this study. Allisa Dillman at CPDR for the initial bioinformatic analyses, Wei Huang at CPDR for providing cell culture models. The Cancer Genome the initial bioinformatic analyses, Wei Huang at CPDR for providing cell culture models. The Cancer Genome Atlas (TCGA) data set supported by NIH. Atlas (TCGA) data set supported by NIH.
Funding Information:
aggressive disease progression, we conducted a comparative study to assess the effects of PMEPA1 isoforms (a and b) on prostate cancer cell growth and signaling pathways of AR and TGF-β. In PC3 isoforms (a and b) on prostate cancer cell growth and signaling pathways of AR and TGF-β. In PC3 cells (TGF-β signaling positive but AR negative), PMEPA1-a promoted cell growth, whereas silencing cells (TGF-β signaling positive but AR negative), PMEPA1-a promoted cell growth, whereas silencing of PMEPA1-a resulted in cell growth inhibition (Figure 4A,D). Interestingly, ectopic PMEPA1-b of PMEPA1-a resulted in cell growth inhibition (Figure 4A,D). Interestingly, ectopic PMEPA1-b resulted resulted in decreased proliferation of PC3 cells. These observations were supported by the results of in decreased proliferation of PC3 cells. These observations were supported by the results of cell cell plating efficiency (Figure 4B,E) and soft agar colony formation assays (Figure 4C,F). Further, plating efficiency (Figure 4B,E) and soft agar colony formation assays (Figure 4C,F). Further, depletion depletion of TGFBR1 resulted in increased cell growth and colony formation of PC3 cells, and there was no enhancement of cell growth in response to PMEPA1-a expression in TGFBR1 depleted PC3 cells (Figure 4G,H). In contrast, PMEPA1-b mediated cell growth inhibition was not affected by TGFRI depletion. These findings underscored that the selective effect of PMEPA1-a on the cell growth depletion. These findings underscored that the selective effect of PMEPA1-a on the cell growth was TGF-β signaling dependent.
Funding Information:
Center for Prostate Disease Research, Department of Surgery, Uniformed Services University of the Health Sciences and the Walter Reed National Military Medical Center, 6720A Rockledge Drive, Suite 300, Bethesda, MD 20817, USA; cymclg@gmail.com (Y.C.); ckuo@cpdr.org (C.K.); lravindranath@cpdr.org (L.R.); gpetrovics@cpdr.org (G.P.); tsreenath@cpdr.org (T.L.S.); adobi@cpdr.org (A.D.); inger.l.rosner.mil@mail.mil (I.L.R.); alagarsamy.srinivasan@gmail.com (A.S.); shsr629@gmail.com (S.S.); jcullen@cpdr.org (J.C.) John P. Murtha Cancer Center, Walter Reed National Military Medical Center, Bethesda, MD 20817, USA Henry Jackson Foundation for the Advancement of Military Medicine, 6720A Rockledge Dr, Suite 100, Bethesda, MD 20817, USA Danish Cancer Society Research Center, 2100 Copenhagen, Denmark; sztup@hotmail.com (Z.M.S.); Zoltan.Szallasi@childrens.harvard.edu (Z.S.) Computational Health Informatics Program, Boston Children’s Hospital, Harvard Medical School, Boston, MA 02115, USA SE-NAP Brain Metastasis Research group, 2nd Department of Pathology, Semmelweis University, 1085 Budapest, Hungary Department of Urology, Walter Reed National Military Medical Center, Bethesda, MD 20814, USA Correspondence: ssharad@cpdr.org (S.S.); hli@cpdr.org (H.L.); Tel.: +1-240-694-4931 (S.S.); +1-240-694-4944 (H.L.)
Publisher Copyright:
© 2019 by the authors. Licensee MDPI, Basel, Switzerland.
PY - 2019/12
Y1 - 2019/12
N2 - Dysfunctions of androgen/TGF-β signaling play important roles in prostate tumorigenesis. Prostate Transmembrane Protein Androgen Induced 1 (PMEPA1) inhibits androgen and TGF-β signaling via a negative feedback loop. The loss of PMEPA1 confers resistance to androgen signaling inhibitors and promotes bone metastasis. Conflicting reports on the expression and biological functions of PMEPA1 in prostate and other cancers propelled us to investigate isoform specific functions in prostate cancer (PCa). One hundred and twenty laser capture micro-dissection matched normal prostate and prostate tumor tissues were analyzed for correlations between quantitative expression of PMEPA1 isoforms and clinical outcomes with Q-RT-PCR, and further validated with a The Cancer Genome Atlas (TCGA) RNA-Seq dataset of 499 PCa. Cell proliferation was assessed with cell counting, plating efficiency and soft agar assay in androgen responsive LNCaP and TGF-β responsive PC3 cells. TGF-β signaling was measured by SMAD dual-luciferase reporter assay. Higher PMEPA1-a mRNA levels indicated biochemical recurrence (p = 0.0183) and lower PMEPA1-b expression associated with metastasis (p = 0.0173). Further, lower PMEPA1-b and a higher ratio of PMEPA1-a vs.-b were correlated to higher Gleason scores and lower progression free survival rate (p < 0.01). TGF-β-responsive PMEPA1-a promoted PCa cell growth, and androgen-responsive PMEPA1-b inhibited cancer cell proliferation. PMEPA1 isoforms-a and-b were shown to be promising candidate biomarkers indicating PCa aggressiveness including earlier biochemical relapse and lower disease specific life expectancy via interrupting androgen/TGF-β signaling.
AB - Dysfunctions of androgen/TGF-β signaling play important roles in prostate tumorigenesis. Prostate Transmembrane Protein Androgen Induced 1 (PMEPA1) inhibits androgen and TGF-β signaling via a negative feedback loop. The loss of PMEPA1 confers resistance to androgen signaling inhibitors and promotes bone metastasis. Conflicting reports on the expression and biological functions of PMEPA1 in prostate and other cancers propelled us to investigate isoform specific functions in prostate cancer (PCa). One hundred and twenty laser capture micro-dissection matched normal prostate and prostate tumor tissues were analyzed for correlations between quantitative expression of PMEPA1 isoforms and clinical outcomes with Q-RT-PCR, and further validated with a The Cancer Genome Atlas (TCGA) RNA-Seq dataset of 499 PCa. Cell proliferation was assessed with cell counting, plating efficiency and soft agar assay in androgen responsive LNCaP and TGF-β responsive PC3 cells. TGF-β signaling was measured by SMAD dual-luciferase reporter assay. Higher PMEPA1-a mRNA levels indicated biochemical recurrence (p = 0.0183) and lower PMEPA1-b expression associated with metastasis (p = 0.0173). Further, lower PMEPA1-b and a higher ratio of PMEPA1-a vs.-b were correlated to higher Gleason scores and lower progression free survival rate (p < 0.01). TGF-β-responsive PMEPA1-a promoted PCa cell growth, and androgen-responsive PMEPA1-b inhibited cancer cell proliferation. PMEPA1 isoforms-a and-b were shown to be promising candidate biomarkers indicating PCa aggressiveness including earlier biochemical relapse and lower disease specific life expectancy via interrupting androgen/TGF-β signaling.
KW - (I)(I)Protein levels ofofAR and
KW - AR
KW - Biochemical recurrence
KW - Cells with PMEPA1-b over-expression (G) and PMEPA1-b depletion (H)
KW - Isoform
KW - Metastasis
KW - PMEPA1
KW - Prostate cancer
KW - TGF-β
UR - http://www.scopus.com/inward/record.url?scp=85076457676&partnerID=8YFLogxK
U2 - 10.3390/cancers11121995
DO - 10.3390/cancers11121995
M3 - Article
AN - SCOPUS:85076457676
SN - 2072-6694
VL - 11
JO - Cancers
JF - Cancers
IS - 12
M1 - 1995
ER -