PROX1 is an early driver of lineage plasticity in prostate cancer
Zhi Duan, et al.
Zhi Duan, et al.
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Research Article Cell biology Oncology

PROX1 is an early driver of lineage plasticity in prostate cancer

Abstract

Lineage plasticity is recognized as a critical determinant of lethality and resistance to AR pathway inhibitors in prostate cancer. Lineage plasticity is a continuum, ranging from AR activity-low tumors, AR-null tumors that do not express a neuroendocrine prostate cancer (NEPC) program (i.e., double-negative prostate cancer [DNPC]), and AR-null NEPC tumors. Factors upregulated early in lineage plasticity are not well-characterized. The clarification of such factors is essential to identify tumors undergoing lineage plasticity or at risk of this occurring. Our integrative analysis of metastatic prostate cancer patient tumors, patient-derived xenografts, and cell models determined that PROX1 is upregulated early in the lineage plasticity continuum and progressively increases as tumors lose AR activity. We determined DNA methylation is a key regulator of PROX1 expression. PROX1 suppression in DNPC and NEPC reduces cell survival and impacts apoptosis and differentiation, demonstrating PROX1’s functional importance. PROX1 is not directly targetable with standard drug development approaches. However, affinity immunopurification demonstrated histone deacetylases (HDACs) are among the top PROX1-interacting proteins; HDAC inhibition depletes PROX1 and recapitulates PROX1 suppression in DNPC and NEPC. Altogether, our results suggest PROX1 promotes the emergence of lineage plasticity, and HDAC inhibition is a promising approach to treat tumors across the lineage plasticity continuum.

Authors

Zhi Duan, Mingchen Shi, Anbarasu Kumaraswamy, Dong Lin, Dhruv Khokhani, Yong Wang, Chao Zhang, Diana Flores, Eva Rodansky, Olivia A. Swaim, William K. Storck, Hannah N. Beck, Radhika A. Patel, Erolcan Sayar, Brian P. Hanratty, Hui Xue, Xin Dong, Zoe R. Maylin, Rensheng Wan, David A. Quigley, Martin Sjöström, Ya-Mei Hu, Faming Zhao, Zheng Xia, Siyuan Cheng, Xiuping Yu, Felix Y. Feng, Li Zhang, Rahul Aggarwal, Eric J. Small, Visweswaran Ravikumar, Arvind Rao, Karan Bedi, John K. Lee, Colm Morrissey, Ilsa Coleman, Peter S. Nelson, Eva Corey, Aaron M. Udager, Ryan J. Rebernick, Marcin P. Cieslik, Arul M. Chinnaiyan, Joel A. Yates, Michael C. Haffner, Yuzhuo Wang, Joshi J. Alumkal

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Figure 3

PROX1 is epigenetically regulated by DNA methylation.

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PROX1 is epigenetically regulated by DNA methylation. (A) PROX1 promoter...
(A) PROX1 promoter methylation in prostate cancer patient tumors was extracted from the Zhao et al. 2020 dataset (n = 100) (26), and 5-methylcytosine (5-mC) score is shown. Data are reported as the mean ± SD. PROX1 promoter is significantly hypermethylated in NEPC (AR–NE+) tumors as indicated by P values calculated by unpaired 2-sample Wilcoxon’s test with Benjamini-Hochberg correction for multiple comparison. **P < 0.01. (B) Scatterplots and linear fitted lines of PROX1 promoter DNA methylation versus log2 PROX1 expression in samples from the WCDT dataset (22, 26). Spearman’s correlation coefficient (ρ) and P values are shown. (C) Genome tracks from whole-genome bisulfite sequencing analysis of indicated PDX samples indicate hypermethylation of PROX1 promoter region (highlighted in yellow) in adenocarcinoma PDXs (blue) and hypomethylation in NEPC PDXs (red). (D) Methylation-specific PCR (MSPCR) was used to amplify a region of the PROX1 promoter from prostate cancer PDXs and cell models. Methylated (M) and unmethylated (U) specific bands are shown for the indicated samples, which are color-coded: ARPC by blue, amphicrine by orange, AR activity–low by green, DNPC by purple, and NEPC by red. (E) The indicated cell lines were treated with 400 nM dAza (decitabine) daily for 5 days. RT-qPCR was performed to quantify PROX1 expression with β-actin used as an endogenous control. Data are reported as the mean ± SD (n = 3). Statistical significance was calculated with a Student’s t test with Welch’s correction. *P < 0.05. (F) MSPCR was performed using DNA extracted from cells treated in E. Ratio of unmethylated (U) to methylated (M) products from densitometry analysis is shown below the respective bands.