LAUSR

Introduction: Therapy of metastasized prostate cancer (PCA) largely depends on androgen deprivation therapy (ADT). Despite initial response, tumors inevitably reach castration resistance (CRPC), a stage at which chemotherapy and second-generation antiandrogens (e.g. Enzalutamide) represent the main therapeutic options. Acquired resistance to these drugs is an emerging problem, raising a need for new approaches. Deletion or mutation of the Chromodomain Helicase DNA binding protein 1 (CHD1) gene, encoding an ATP-dependent chromatin remodeler, is more prevalent in CRPC compared to earlier tumor stages. Therefore, we investigated the effects of CHD1 depletion in androgen-sensitive LNCaP cells. Experimental procedures: LNCaP cells were grown in media with either 10% fetal bovine serum (FBS), 10% charcoal-stripped serum (CSS) or 10% CSS plus 1nM testosterone. Growth was visualized by image-based cell cytometry. After 10 days of growth, RNA sequencing (RNA-seq) was performed to investigate changes in gene expression (between the different growth conditions with and without CHD1 depletion). Results were verified by qRT-PCR and Western blot. The identified treatment options were tested with small-molecule inhibitors in different concentrations. Finally, we performed chromatin immunoprecipitation followed by next-generation sequencing (ChIP-seq) in order to identify the epigenetic mechanisms underlying the differential regulation. Results: We identified a correlation between androgen-independent growth and CHD1 depletion which could be compensated by addition of 1 nM testosterone. Moreover, we identified CHD1-dependent regulation of Enhancer Of Zeste Homolog-2 (EZH2), which could be confirmed at the protein level. Treatment with 1 µM JQ-EZ (an EZH2 inhibitor) showed a stronger effect on the proliferation of CHD1-knockdown cells compared to controls. ChIP-seq and RNA-seq analysis helped to identify a subset of EZH2-bound genes depending on CHD1 knockdown and androgen deprivation. Among those genes, we identified the I-SMAD genes SMAD6 and SMAD7 which are regulators of TGFβ and BMP signaling to be upregulated after androgen-deprivation and downregulated with CHD1 knockdown. We also showed differential regulation of PMEPA1, a TGFβ-dependent inhibitor of PCA bone metastasis. Coherently, we found upregulation of mesenchymal markers such as CDH2, pointing towards a role in EMT. Conclusion: We identified a new regulatory mechanism for CRPC harboring a CHD1-deletion leading to a more mesenchymal state in an EZH2-dependent manner. First experiments with an EZH2 inhibitor in our system indicate a possible therapeutic use. CHD1 could hereby be used both as a biomarker for early metastasis as well as for therapeutic stratification. Citation Format: Simon J. Bocker, Simon J. Baumgart, Annabell Roth, Vijayalakshmi Kari, Steven A. Johnsen, Oliver Hahn. CHD1 depletion leads to androgen-independent growth of prostate cancer cells lines and regulates the TGFβ system in an EZH2-dependent manner [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1917.