Stromal epigenetic alterations drive metabolic and neuroendocrine prostate cancer reprogramming
Rajeev Mishra, … , Edwin M. Posadas, Neil A. Bhowmick
Rajeev Mishra, … , Edwin M. Posadas, Neil A. Bhowmick
Published October 1, 2018; First published July 26, 2018
Citation Information: J Clin Invest. 2018;128(10):4472-4484. https://doi.org/10.1172/JCI99397.
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Categories: Research Article Cell biology Metabolism

Stromal epigenetic alterations drive metabolic and neuroendocrine prostate cancer reprogramming

Abstract

Prostate cancer is an androgen-dependent disease subject to interactions between the tumor epithelium and its microenvironment. Here, we found that epigenetic changes in prostatic cancer-associated fibroblasts (CAF) initiated a cascade of stromal-epithelial interactions. This facilitated lethal prostate cancer growth and development of resistance to androgen signaling deprivation therapy (ADT). We identified a Ras inhibitor, RASAL3, as epigenetically silenced in human prostatic CAF, leading to oncogenic Ras activity driving macropinocytosis-mediated glutamine synthesis. Interestingly, ADT further promoted RASAL3 epigenetic silencing and glutamine secretion by prostatic fibroblasts. In an orthotopic xenograft model, subsequent inhibition of macropinocytosis and glutamine transport resulted in antitumor effects. Stromal glutamine served as a source of energy through anaplerosis and as a mediator of neuroendocrine differentiation for prostate adenocarcinoma. Antagonizing the uptake of glutamine restored sensitivity to ADT in a castration-resistant xenograft model. In validating these findings, we found that prostate cancer patients on ADT with therapeutic resistance had elevated blood glutamine levels compared with those with therapeutically responsive disease (odds ratio = 7.451, P = 0.02). Identification of epigenetic regulation of Ras activity in prostatic CAF revealed RASAL3 as a sensor for metabolic and neuroendocrine reprogramming in prostate cancer patients failing ADT.

Authors

Rajeev Mishra, Subhash Haldar, Veronica Placencio, Anisha Madhav, Krizia Rohena-Rivera, Priyanka Agarwal, Frank Duong, Bryan Angara, Manisha Tripathi, Zhenqiu Liu, Roberta A. Gottlieb, Shawn Wagner, Edwin M. Posadas, Neil A. Bhowmick

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Figure 2

Ras activation stimulates macropinocytosis in prostatic fibroblasts and potentiates growth of adjacent epithelial.

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Ras activation stimulates macropinocytosis in prostatic fibroblasts and ...
(A) Representative Western blots of RASAL3, Ras, and Ras effectors in prostatic human NAF/CAF (n = 3). See complete unedited blots in the supplemental material. (B) TMR-dextran (red) uptake by CAF shows elevated levels of macropinocytosis compared with NAF. DAPI staining (blue) identifies nuclei. (C) CAF that were coincubated with fluorescent DQ-BSA (green) and TMR-dextran (red) were fixed after 1-hour chase. The fluorescent signal emanating from DQ-BSA with TMR-dextran–positive staining (arrowheads) indicates albumin uptake by macropinosomes and subsequent breakdown. (D) Representative Western blots of RASAL3, total Ras, Ras-GTP, and phosphorylated ERK (phos-ERK) expression by Rasal3-KO and Cas9 control mouse prostatic fibroblasts are shown (n = 3). (E) Representative images show TMR-dextran–positive macropinosomes (arrowheads) in Rasal3-KO prostatic mouse fibroblasts compared with Cas9. (F) Representative Western blots of mouse prostatic fibroblasts, WT and expressing RasV12, indicate total Ras, Ras-GTP, and phosphorylated ERK status (n = 3). (G) Representative images show TMR-dextran–positive macropinosomes (arrowheads) in RasV12 prostatic mouse fibroblasts (expressing GFP) compared with their WT counterparts. (A–G) Representative images of 3 independent experiments are shown. (H) Schematic illustrates orthotopic tissue recombinant xenograft models of CWR22Rv1 and RasV12 mouse fibroblasts allowed to grow for 2 weeks, followed by treatment with vehicle or 10 mg/kg EIPA (n = 8). Representative gross tumor images are shown with a graph of all the tumor volumes. P value was calculated using 2-tailed Student’s t test. Data are represented as mean ± SD. **P < 0.01. Scale bars: 30 μm (B, C, E, G); 3 mm (H).