Lipin Expression Correlates to Differences in Phospholipid Profiles in Prostate Cancer Progression

Abstract

PURPOSE The association between clinical outcomes of prostate cancer progression and lipid remodeling is not well understood. Although it is known that increases in select circulating lipids correlate to decreased patient survival, the mechanisms mediating alterations in these lipids are not fully explained. We addressed this gap-in-knowledge using in vitro models of non-cancerous, hormone-sensitive, and castration-resistant prostate cancer (CRPC) cell lines combined with quantitative HPLC-ESI-Orbitrap-MS lipidomic analysis. HYPOTHESIS We hypothesized that the expression of lipin (phosphatidic acid phosphatase), an enzyme mediating the conversion of phosphatidic acid to diacylglycerol, correlates to changes in lipidomic profiles in prostate cancer cells. We further hypothesized that changes in this enzyme influences the overall survivability of prostate cancer patients. METHODS Lipids were isolated using the Bligh-Dyer extraction from non-cancerous, hormone-sensitive, and CRPC cell lines. Lipids present in both cells and media were initially analyzed using an untargeted shotgun approach (ESI-MS), followed by a targeted-based quantitative HPLC-ESI-Orbitrap-MS approach. Post data acquisition for both shotgun and HPLC-ESI/MS/MS lipidomics were based on multiple methods including isotope, carbon number (to internal standards) and ionization efficiency-based corrections. Online resources and software's utilized included MetaboAnalyst, LIPIDMATCH, LIPID MAPS, XCMS, and MZmine. Gene alteration frequency and overall survival data were generated using cBioPortal for Cancer Genomics and GEPIA 2. Lipin expression in cells was determined using immunoblot analysis. RESULTS Our data showed significant changes in the abundance of lipid species between the various cell lines and a correlation between select lipid species and the progression of prostate cancer. Prominent among these were the levels of phosphatidic acid (PA) and diacylglycerol (DAG), a process controlled by lipin. Most notably, PA levels in cells tended to inversely correlate with DAG levels. These levels correlated to the decreased expression on lipin, specifically lipin-1, suggesting that decreased lipin-1 expression correlates to more aggressive prostate cancer. This hypothesis was supported by analysis of alteration in lipin genes expressed in numerous cohorts of prostate cancer patients, demonstrating that lipin mutations indicated a decrease in the overall survival of prostate cancer patients. CONCLUSIONS Distinct phospholipid classes were observed to have increased levels in prostate cancer cells, as compared to non-cancer cells. Preliminary data shows a correlation between the abundance of PA and DAG with lipin expression in cells representing more aggressive prostate cancers, which is supported by data from patient populations, suggesting that altered lipin expression correlates to decreased survival.