Endocrine Abstracts

Metformin has recently emerged as a potential therapeutic tool for different tumour pathologies, including prostate cancer (PCa)¹, one of the leading causes of cancer-deaths in men worldwide. However, the molecular mechanisms underlying the antitumor effects of metformin in PCa are not fully elucidated. In this context, previous reports have suggested a relationship between metformin and the process of alternative splicing in some cell-types. Nevertheless, the potential role that the splicing machinery [-spliceosome components (SCs) and splicing factors (SFs)] might play in the antitumor effects of metformin in PCa remains unknown. Therefore, the aim of this study was to examine the effects and implications of metformin on the expression levels of key splicing machinery components and splicing variants in PCa. Specifically, expression levels of 14 SCs and 28 SFs were measured by a microfluidic-based qPCR array in androgen-dependent and independent PCa-derived cell lines (LNCaP and PC-3, respectively) and in PC-3-induced xenograft tumours under low- and high-fat diets conditions in response to metformin (5 mM in vitro; 250 mg/kg/day in vivo). Moreover, different mechanistic (qPCR/Western-Blot) and functional (cell-proliferation) assays in response to metformin and/or in response to the silencing of certain SCs/SFs (using specific siRNAs) were implemented. Metformin reduced the expression level of 21% (3/14) of the SCs and 43% (12/28) of the SFs studied in LNCaP and PC-3 cells. Likewise, downregulation of certain SCs and SFs in response to metformin treatment was also observed in vivo in PC-3 induced xenografts. Interestingly, some of these changes were diet (low-fat vs high-fat) dependent, while others were diet-independent. Remarkably, three elements of the splicing machinery (NOVA1, SF3B1 and SRRM1) were consistently reduced in response to metformin in both in vitro and in vivo models. In fact, silencing of these three elements completely blocked the anti-proliferative actions of metformin in PCa cells. Consistently, metformin reduced the expression levels of certain splicing variants related to PCa development or aggressiveness (e.g.: KLF6-SV1, RAC1B, SST₅TMD4) in PCa cells. Finally, we evaluated whether these changes exerted by metformin were AMPK-dependent or -independent using adenoviruses encoding either a dominant-negative or constitutively-active isoform of AMPKa. Altogether, our results suggest that the antitumor effect exerted by metformin in PCa might be mediated, at least in part, by the modulation of certain splicing machinery elements (NOVA1/SF3B1/SRRM1) and splicing variants(KLF6-SV1/RAC1B/SST₅TMD4).

Fundings: ISCIII (PI16-00264, PI17-02287), MINECO (FPU18/02485, FPU16/06190, BFU2016-80360-R), Junta de Andalucía (BIO-0139), CIBERobn.

Reference

1. Sarmento-Cabral A, et al.. Mol Cancer Res 2017 15 (7) 862–874.