Novel Junction-specific and Quantifiable In Situ Detection of AR-V7 and its Clinical Correlates in Metastatic Castration-resistant Prostate Cancer

Zhu, Yezi; Sharp, Adam; Anderson, Courtney; Silberstein, John L.; Taylor, Maritza N.; Lu, Changxue; Zhao, Pei; Marzo, Angelo M. De; Antonarakis, Emmanuel S.; Wang, Mindy; Wu, Xinfeng; Luo, Yuling; Su, Nan; Rodrigues, Daniel Nava; Figueiredo, Ines; Welti, Jonathan; Park, Emily; Xiao, Jun; Coleman, Ilsa M.; Morrissey, Colm; Plymate, Stephen R.; Nelson, Peter S.; Bono, Johann S. de; Luo, Jun

doi:10.1016/j.eururo.2017.08.009

Cited by 57 publications

(70 citation statements)

References 28 publications

(45 reference statements)

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“…In contrast, enzalutamide did not reduce PDX-O growth, consistent with its parental biopsy being from a CRPC patient who had developed resistance to enzalutamide (Figure 6B and C). AR-V7 expression increases as PC patients progress to CRPC and develop treatment resistance (14,15,(22)(23)(24)(25)(26). Consistent with this, castration of CP50 PDX for 14 days significantly increased AR-V7 expression, with a less pronounced impact on AR-FL, C-MYC expression and AR signaling (Figure 6D Figure S13A and B).…”

Section: I-bet151 Regulates Ar-v7 and C-myc Expression And Inhibits supporting

confidence: 63%

“…This has led to advances in antiandrogen therapy, with the successful development of abiraterone and enzalutamide (9)(10)(11)(12)(13). Despite these advances, primary resistance is common and secondary resistance on treatment inevitable in CRPC, in part due to expression of constitutively active AR splice variants that evade current antiandrogen treatment strategies (14,15,(21)(22)(23)(24)(25)(26). One promising strategy undergoing clinical evaluation in CRPC is targeting BET family proteins (ClinicalTrials.gov identifier: NCT02711956, NCT03150056).…”

Section: Discussionmentioning

confidence: 99%

“…In addition to PC cell lines, I-BET151 inhibited the growth of PDOs derived from fresh metastatic CRPC patients who had progressed despite abiraterone and/or enzalutamide treatment. These data suggest that BET inhibitors may provide therapeutic approaches to overcome AR resistant mechanisms in CRPC including AR amplification and AR-V7 expression that associate with worse outcome and resistance to abiraterone and/or enzalutamide (6,14,15,(22)(23)(24)(25)(26)47). To further investigate this, we developed a PDX-O and PDX (CP50) from a patient who had progressed on both abiraterone and enzalutamide with tumor AR amplification and AR-V7 protein expression.…”

Section: Discussionmentioning

confidence: 99%

“…Immunohistochemistry (IHC) for BRD4, AR-V7 and full length androgen receptor (AR-FL) was performed on patient samples and CP50 PDX as previously described (Supplementary Methods) (25,26,43).…”

Section: Tissue Analysismentioning

confidence: 99%

“…However, primary and secondary resistance to both therapies is common and may, in part, be due to the expression of constitutively active AR splice variants of which AR variant 7 (AR-V7) is considered the most significant and best studied (14)(15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27). Consistent with this, recent studies have demonstrated that AR-V7 expression increases as patients progress to CRPC, and associates with resistance to current AR directed therapies (14,15,(22)(23)(24)(25)(26). AR-V7 lacks the AR ligand binding domain (LBD), but contains a cryptic exon 3 (derived from an intron) after AR exon 3 (27).…”

Section: Introductionmentioning

confidence: 99%

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Targeting Bromodomain and Extra-Terminal (BET) Family Proteins in Castration-Resistant Prostate Cancer (CRPC)

Welti

Sharp

Yuan

et al. 2018

Clinical Cancer Research

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118

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Welti et al; Targeting BET family proteins in CRPC 2 Statement of Translational RelevanceAdvanced prostate cancer invariably progresses to lethal castration resistant prostate cancer (CRPC). Resistance to current androgen receptor (AR) targeting therapies is associated with the development of AR aberrations including the constitutively active AR splice variant 7 (AR-V7). Currently, no clinically available therapies effectively inhibit aberrant AR signaling. BRD4, a bromodomain and extraterminal (BET) family protein, is a critical AR coregulator. We show that BRD4 expression associates with patient outcome and AR driven transcription in lethal prostate cancer. Moreover, BET inhibitors (BETi) reduce AR splicing and AR-V7 expression by regulating alternative splicing, abrogating AR signaling and inhibiting growth of CRPC patient derived models. Clinical studies with BETi in CRPC should pursue pharmacodynamics studies evaluating abrogation of AR splicing and persistent AR signaling to optimize the development of these drugs for the treatment of CRPC.Research. Experimental Design: We determined associations between BET expression, AR driven transcription and patient outcome; and the effect and mechanism by which chemical BETi (JQ1 and GSK1210151A; I-BET151) and BET family protein knockdown regulates AR-V7 expression and AR signaling in prostate cancer (PC) models.Results: Nuclear BRD4 protein expression increases significantly (p=<0.01) with castration resistance in same patient treatment naïve (median H-score; interquartile range: 100; 100-170) and CRPC (150; 110-200) biopsies, with higher expression at diagnosis associating with worse outcome (HR 3.25, 95% CI 1.50-7.01; p=<0.001). BRD2, BRD3 and BRD4 RNA expression in CRPC biopsies correlates with AR driven transcription (all p=<0.001). Chemical BETi, and combined BET family protein knockdown, reduce AR-V7 expression and AR signaling. This was not recapitulated by C-MYC knockdown. In addition, we show that BETi regulates RNA processing thereby reducing alternative splicing and AR-V7 expression. Furthermore, BETi reduce growth of PC cells and patient derived organoids with known AR mutations, AR amplification and AR-V7 expression. Finally, BETi, unlike enzalutamide, decreases persistent AR signaling and growth (p=<0.001) of a patient derived xenograft model of CRPC with AR amplification and AR-V7 expression. Conclusion:BETi merit clinical evaluation as inhibitors of AR splicing and function, with trials demonstrating their blockade in proof of mechanism pharmacodynamic studies.

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Section: I-bet151 Regulates Ar-v7 and C-myc Expression And Inhibits supporting

confidence: 63%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Tissue Analysismentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Targeting Bromodomain and Extra-Terminal (BET) Family Proteins in Castration-Resistant Prostate Cancer (CRPC)

Welti

Sharp

Yuan

et al. 2018

Clinical Cancer Research

Self Cite

118

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Detection and Quantification of Multiple RNA Sequences Using Emerging Ultrasensitive Fluorescent In Situ Hybridization Techniques

Erben

Buonanno

2019

CP Neuroscience

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Fluorescent detection of transcripts using RNAscope has quickly become a standard in situ hybridization (ISH) approach in neuroscience with over 400 publications since its introduction in 2012. RNAscope's sensitivity and specificity allow the simultaneously detection of up to three low abundance mRNAs in single cells (i.e., multiplexing) and, in contrast to other ISH techniques, RNAscope is performed in 1 day. BaseScope, a newer ultrasensitive platform, uses improved amplification chemistry of single oligonucleotide probe pairs (∼50 bases). This technique allows discrimination of single nucleotide polymorphisms or splice variants that differ by short exons. A present limitation of BaseScope is that expression analysis is limited to a single gene (i.e., single‐plexing). This article outlines detailed protocols for both RNAscope and BaseScope in neuronal tissue. We discuss how to perform ISH experiments using either fresh‐frozen or formalin‐fixed paraffin‐embedded sections, as well as dissociated cultured neurons. We also outline how to obtain quantitative data from hybridized tissue sections. © 2019 by John Wiley & Sons, Inc.

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A Coiled‐Coil Domain Containing 50 Splice Variant Is Modulated by Serine/Arginine‐Rich Splicing Factor 3 and Promotes Hepatocellular Carcinoma in Mice by the Ras Signaling Pathway

Wang

Zhang

et al. 2018

Hepatology

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Deregulation of alternative splicing contributes to the malignant progression of cancer. Little is known about the significant alternative splicing events in hepatocellular carcinoma (HCC). High through-put sequencing revealed that CCDC50 pre-mRNA is aberrantly spliced in 50% of our HCC cases. A BaseScope assay was performed to examine the expression of CCDC50S (a truncated oncogenic splice variant) in HCC tissues. Compared with liver benign tumors and several types of solid tumors, CCDC50S mRNA was up-regulated in HCC, with a diagnostic potential (sensitivity: 0.711, specificity: 0.793). High expression of CCDC50S mRNA in HCC was significantly correlated with poor tumor differentiation, advanced TNM stage and unfavorable prognosis. Overexpression of CCDC50S exerted tumorigenic activities that promoted HCC growth and metastasis via activation of Ras/Foxo4 signaling. Either suppression of MEK/ERK phosphorylation or overexpression of Foxo4 markedly attenuated CCDC50S-mediated phenotypes. Furthermore, serine and arginine rich splicing factor 3 (SRSF3) directly bound to CCDC50S mRNA to maintain its stability in the cytoplasm. The cytosolic retention of SRSF3 was mediated by the interaction of HBx and 14-3-3β. Ectopic HBx expression induced the expression of cytosolic SRSF3 and CCDC50S. Our study provided compelling evidence that up-regulation of CCDC50S was modulated by HBx/SRSF3/14-3-3β complex, and enhanced the oncogenic progression of HCC via the Ras/Foxo4 signaling pathway. These data suggest that CCDC50S may serve as a novel diagnostic and prognostic biomarker, and probably a promising therapeutic target in HCC. This article is protected by copyright. All rights reserved.

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Novel Junction-specific and Quantifiable In Situ Detection of AR-V7 and its Clinical Correlates in Metastatic Castration-resistant Prostate Cancer

Abstract: Higher AR-V7 levels detected and quantified using a novel method were associated with poorer response to abiraterone or enzalutamide in prostate cancer.

Cited by 57 publications

References 28 publications

Targeting Bromodomain and Extra-Terminal (BET) Family Proteins in Castration-Resistant Prostate Cancer (CRPC)

Targeting Bromodomain and Extra-Terminal (BET) Family Proteins in Castration-Resistant Prostate Cancer (CRPC)

Detection and Quantification of Multiple RNA Sequences Using Emerging Ultrasensitive Fluorescent In Situ Hybridization Techniques

A Coiled‐Coil Domain Containing 50 Splice Variant Is Modulated by Serine/Arginine‐Rich Splicing Factor 3 and Promotes Hepatocellular Carcinoma in Mice by the Ras Signaling Pathway

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