Combined BET bromodomain and DNA methyltransferase inhibition targets critical survival pathways in transdifferentiated prostate cancer
William K. Storck, Diana Flores, Anbarasu Kumaraswamy, Zhi Duan, Shrabastee Chakraborty, Chao Zhang, Eva Rodansky, Dhruv Khokhani, Olivia A. Swaim, Karan Bedi, Raymond G. Cavalcante, Canping Chen, Faming Zhao, Ya-Mei Hu, Zheng Xia, Ryan J. Rebernick, Marcin Cieslik, Rahul Mannan, Somnath Mahapatra, Arul M. Chinnaiyan, Aaron M. Udager, Joshua A. Kuleape, Catherine R. Alumkal, Hannah N. Beck, Peter S. Nelson, Colm Morrissey, Michael C. Haffner, Leigh Ellis, Yuzhuo Wang, Joel A. Yates, Joshi J. Alumkal
William K. Storck, Diana Flores, Anbarasu Kumaraswamy, Zhi Duan, Shrabastee Chakraborty, Chao Zhang, Eva Rodansky, Dhruv Khokhani, Olivia A. Swaim, Karan Bedi, Raymond G. Cavalcante, Canping Chen, Faming Zhao, Ya-Mei Hu, Zheng Xia, Ryan J. Rebernick, Marcin Cieslik, Rahul Mannan, Somnath Mahapatra, Arul M. Chinnaiyan, Aaron M. Udager, Joshua A. Kuleape, Catherine R. Alumkal, Hannah N. Beck, Peter S. Nelson, Colm Morrissey, Michael C. Haffner, Leigh Ellis, Yuzhuo Wang, Joel A. Yates, Joshi J. Alumkal
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Research Article Cell biology Oncology

Combined BET bromodomain and DNA methyltransferase inhibition targets critical survival pathways in transdifferentiated prostate cancer

Abstract

Lineage plasticity, or transdifferentiation, is increasingly recognized as a resistance mechanism to androgen receptor (AR) inhibition in prostate cancer. Lineage plasticity is characterized by loss of AR signaling and epithelial differentiation, along with activation of stemness-associated pathways, epithelial-mesenchymal transition, or alternative differentiation programs such as neuroendocrine prostate cancer (NEPC). Loss of the tumor suppressors TP53 and RB1 is common in tumors exhibiting lineage plasticity; however, the mechanisms by which TP53/RB1 loss promotes this phenotype remain poorly understood, and effective treatments are limited. Using multiomic profiling of TP53/RB1-loss prostate cancer models, we identified alterations in chromatin accessibility, DNA methylation, and gene expression associated with lineage plasticity. Importantly, many pathways activated upon TP53/RB1 loss could be blocked through BET bromodomain inhibition. TP53/RB1-deficient cells also harbored widespread DNA methylation changes that silenced pathways linked with restraining lineage plasticity. Combined BET bromodomain and DNA methyltransferase (DNMT) inhibition was more effective than single-agent treatment in suppressing growth of TP53/RB1-loss models exhibiting a stem-like or NEPC program. This was partly explained by abrogation of discrete lineage plasticity pathways modulated by each agent. Altogether, our work suggests combined BET bromodomain and DNMT inhibition is a promising therapeutic approach for prostate tumors exhibiting lineage plasticity.

Authors

William K. Storck, Diana Flores, Anbarasu Kumaraswamy, Zhi Duan, Shrabastee Chakraborty, Chao Zhang, Eva Rodansky, Dhruv Khokhani, Olivia A. Swaim, Karan Bedi, Raymond G. Cavalcante, Canping Chen, Faming Zhao, Ya-Mei Hu, Zheng Xia, Ryan J. Rebernick, Marcin Cieslik, Rahul Mannan, Somnath Mahapatra, Arul M. Chinnaiyan, Aaron M. Udager, Joshua A. Kuleape, Catherine R. Alumkal, Hannah N. Beck, Peter S. Nelson, Colm Morrissey, Michael C. Haffner, Leigh Ellis, Yuzhuo Wang, Joel A. Yates, Joshi J. Alumkal

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Figure 3

Trp53/Rb1 loss leads to widespread chromatin hypo-accessibility and DNA methylation changes that suppress epithelial gene expression.

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Trp53/Rb1 loss leads to widespread chromatin hypo-accessibility and DNA...
(A) PCA plot of ERRBS data from SKO, DKO, DKOcr105, DKOcr298, and TKO mouse cell lines (5) (n = 3 for all cell lines). (B) Schematic depicting analysis integrating differential hypo-accessible chromatin at promoters and enhancers (ATAC-Seq) corresponding with increased DNA methylation (ERRBS) and gene suppression (RNA-Seq) from Trp53/Rb1-deficient (TKO; DKOcr298) versus Trp53/Rb1-intact (SKO) cell lines to determine methylated and silenced genes induced by loss of Trp53 and Rb1. (C) Representative top shared methylated and suppressed gene sets (FDR < 0.1) from MSigDB Hallmark, 16. Chemical and Genetic Perturbations (CGP), and and Gene Ontology Biological Process (GOBP) gene set collections related to stemness, EMT, metastasis, and luminal/epithelial regulation in TKO and DKOcr298 ranked by TKO FDR. Statistical significance was determined using hypergeometric test followed by Benjamini-Hochberg correction. (D) IGV (57) genome browser snapshots showing DNA methylation (ERRBS), chromatin accessibility (ATAC-Seq), H3K27Ac ChIP-Seq enrichment, and gene expression (RNA-Seq) at selected genes associated with epithelial differentiation in SKO, DKOcr298, and TKO cell lines. Gene promoter regions are highlighted. (E) Methylation-specific PCR (MSPCR) analysis of Trp53/Rb1 loss–induced hypermethylated and silenced genes in TKO treated with increasing doses of dAZA. Cells were treated daily for 72 hours. Methylated (M) and unmethylated (U) specific amplicons are shown for each indicated sample. The densitometry ratio of unmethylated to methylated products (U to M) is shown below their respective amplicon pairs. SKO is included as an unmethylated control. Representative samples from biological triplicates are shown for each condition. (F) RNA-qPCR of Trp53/Rb1 loss–induced hypermethylated and silenced genes in TKO with increasing doses of dAZA. RNAs used in this analysis were derived from the same samples as in E. Data presented are the mean ± SD (n = 3 for each treatment condition). Statistical significance was determined using 1-way ANOVA with Dunnett’s multiple-comparison test. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001.