Jun Pang1, Yue-Wu Yang2, Yiling Huang3,4, Jun Yang3,5, Hao Zhang1, Ruibao Chen3,5, Liang Dong3, Yan Huang3, Dongying Wang3, Jihong Liu5, Benyi Li1,3,4. 1. Department of Urology, The Third Affiliated Hospital, Sun Yat-Sen University, Guangzhou, China. 2. Department of Traditional Chinese Medicine, The Third Affiliated Hospital, Sun Yat-Sen University, Guangzhou, China. 3. Department of Urology, The University of Kansas Medical Center, Kansas City, Kansas. 4. Department of Pathology, China Three Gorges University School of Medicine, Yichang, China. 5. Department of Urology, Tongji Hospital, Huanzhong University of Science and Technology, Wuhan, China.
Abstract
INTRODUCTION AND AIMS: Epigenetic alteration plays a major role in the development and progression of human cancers, including prostate cancer. Histones are the key factors in modulating gene accessibility to transcription factors and post-translational modification of the histone N-terminal tail including methylation is associated with either transcriptional activation (H3K4me2) or repression (H3K9me3). Furthermore, phosphoinositide 3-kinase (PI3 K) signaling and the androgen receptor (AR) are the key determinants in prostate cancer development and progression. We recently showed that prostate-targeted nano-micelles loaded with PI3 K/p110beta specific inhibitor TGX221 blocked prostate cancer growth in vitro and in vivo. Our objective of this study was to determine the role of PI3 K signaling in histone methylation in prostate cancer, with emphasis on histone H3K4 methylation. METHODS: PI3 K non-specific inhibitor LY294002 and p110beta-specific inhibitor TGX221 were used to block PI3 K/p110beta signaling. The global levels of H3K4 and H3K9 methylation in prostate cancer cells and tissue specimens were evaluated by Western blot assay and immunohistochemical staining. A synthetic androgen R1881 was used to stimulate AR activity in prostate cancer cells. A castration-resistant prostate cancer (CRPC) specific human tissue microarray (TMA) was used to assess the global levels of H3K4me2 methylation by immunostaining approach. RESULTS: Our data revealed that H3K4me2 levels were significantly elevated after androgen stimulation. With RNA silencing and pharmacology approaches, we further defined that inhibition of PI3 K/p110beta activity through gene-specific knocking down and small chemical inhibitor TGX221 abolished androgen-stimulated H3K4me2 methylation. Consistently, prostate cancer-targeted delivery of TGX221 in vivo dramatically reduced the global levels of H3K4me2 as assessed by immunohistochemical staining on tissue section of mouse xenografts from CRPC cell lines 22RV1 and C4-2. Finally, immunostaining data revealed a strong H3K4me2 immunosignal in CRPC tissues compared to primary tumors and benign prostate tissues. CONCLUSIONS: Taken together, our results suggest that PI3 K/p110beta-dependent signaling is involved in androgen-stimulated H3K4me2 methylation in prostate cancer, which might be used as a novel biomarker for disease prognosis and targeted therapy. Prostate 77:299-308, 2017.
INTRODUCTION AND AIMS: Epigenetic alteration plays a major role in the development and progression of humancancers, including prostate cancer. Histones are the key factors in modulating gene accessibility to transcription factors and post-translational modification of the histone N-terminal tail including methylation is associated with either transcriptional activation (H3K4me2) or repression (H3K9me3). Furthermore, phosphoinositide 3-kinase (PI3 K) signaling and the androgen receptor (AR) are the key determinants in prostate cancer development and progression. We recently showed that prostate-targeted nano-micelles loaded with PI3 K/p110beta specific inhibitor TGX221 blocked prostate cancer growth in vitro and in vivo. Our objective of this study was to determine the role of PI3 K signaling in histone methylation in prostate cancer, with emphasis on histone H3K4 methylation. METHODS: PI3 K non-specific inhibitor LY294002 and p110beta-specific inhibitor TGX221 were used to block PI3 K/p110beta signaling. The global levels of H3K4 and H3K9 methylation in prostate cancer cells and tissue specimens were evaluated by Western blot assay and immunohistochemical staining. A synthetic androgen R1881 was used to stimulate AR activity in prostate cancer cells. A castration-resistant prostate cancer (CRPC) specific human tissue microarray (TMA) was used to assess the global levels of H3K4me2 methylation by immunostaining approach. RESULTS: Our data revealed that H3K4me2 levels were significantly elevated after androgen stimulation. With RNA silencing and pharmacology approaches, we further defined that inhibition of PI3 K/p110beta activity through gene-specific knocking down and small chemical inhibitor TGX221 abolished androgen-stimulated H3K4me2 methylation. Consistently, prostate cancer-targeted delivery of TGX221 in vivo dramatically reduced the global levels of H3K4me2 as assessed by immunohistochemical staining on tissue section of mouse xenografts from CRPC cell lines 22RV1 and C4-2. Finally, immunostaining data revealed a strong H3K4me2 immunosignal in CRPC tissues compared to primary tumors and benign prostate tissues. CONCLUSIONS: Taken together, our results suggest that PI3 K/p110beta-dependent signaling is involved in androgen-stimulated H3K4me2 methylation in prostate cancer, which might be used as a novel biomarker for disease prognosis and targeted therapy. Prostate 77:299-308, 2017.