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Comprehensive proteomic profiling identifies the androgen receptor axis and other signaling pathways as targets of microRNAs suppressed in metastatic prostate cancer

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Abstract

MicroRNAs are important epigenetic regulators of protein expression by triggering degradation of target mRNAs and/or inhibiting their translation. Dysregulation of microRNA expression has been reported in several cancers, including prostate cancer (PC). We comprehensively characterized the proteomic footprint of a panel of 12 microRNAs that are potently suppressed in metastatic PC (SiM-miRNAs: miR-1, miR-133a, miR-133b, miR-135a, miR-143-3p, miR-145-3p, miR-205, miR-221-3p, miR-221-5p, miR-222-3p, miR-24-1-5p, and miR-31) using reverse-phase proteomic arrays. Re-expression of these SiM-miRNAs in PC cells suppressed cell proliferation and targeted key oncogenic pathways, including cell cycle, apoptosis, Akt/mammalian target of rapamycin signaling, metastasis and the androgen receptor (AR) axis. However, only 12%, at most, of these observed protein expression changes could be explained by predicted direct binding of miRNAs to corresponding mRNAs, suggesting that the majority of these proteomic effects result indirectly. AR and its steroid receptor coactivators (SRCs; SRC-1, -2 and -3) were recurrently affected by these SiM-miRNAs. In agreement, we identified inverse correlations between expression of these SiM-miRNAs and early clinical recurrence, as well as with AR transcriptional activity in human PC tissues. We also identified robust induction of miR-135a by androgen and strong direct binding of AR to the miR-135a locus. As miR-135a potently suppresses AR expression, this results in a negative feedback loop that suppresses AR protein expression in an androgen-dependent manner, while de-repressing AR expression upon androgen deprivation. Our results demonstrate that epigenetic silencing of these SiM-miRNAs can result in increased AR axis activity and cell proliferation, thus contributing to disease progression. We further demonstrate that a negative feedback loop involving miR-135a can restore AR expression under androgen-deprivation conditions, thus contributing to the upregulation of AR protein expression in castration-resistant PC. Finally, our unbiased proteomic profiling demonstrates that the majority of actual protein expression changes induced by SiM-miRNAs cannot be explained based on predicted direct interactions.

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Acknowledgements

We acknowledge the joint participation by Adrienne Helis Malvin Medical Research Foundation through its direct engagement in the continuous active conduct of medical research in conjunction with Baylor College of Medicine. This work was also supported by the American Cancer Society RSG-14-218-01-TBG (to NM), the Prostate Cancer Foundation (to BWO and NM), the Conquer Cancer Foundation of the American Society of Clinical Oncology Young Investigator and Career Development Awards (both to NM), NICHD 8818 and Department of Defense Breast Cancer Research Program Innovator Award (to BWO), the Pilot/Feasibility Program of the Diabetes and Endocrinology Research Center (P30-DK079638) at Baylor College of Medicine (to NM), and an Alkek Foundation for Molecular Discovery Pilot grant (to CC). NM is a Dan L. Duncan Scholar, a Caroline Wiess Law Scholar and a member of the Dan L. Duncan Cancer Center (supported by the NCI Cancer Center Support grant P30CA125123) and the Center for Drug Discovery at Baylor College of Medicine. We thank the assistance of the Shared Resources of the Dan L. Duncan Cancer Center (supported by the NCI Cancer Center Support grant P30CA125123) and the Functional Proteomics RPPA Core Facility (The University of Texas M.D. Anderson Cancer Center, Houston, TX, USA).

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Coarfa, C., Fiskus, W., Eedunuri, V. et al. Comprehensive proteomic profiling identifies the androgen receptor axis and other signaling pathways as targets of microRNAs suppressed in metastatic prostate cancer. Oncogene 35, 2345–2356 (2016). https://doi.org/10.1038/onc.2015.295

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