Serum- and glucocorticoid- inducible kinase 2, SGK2, is a novel autophagy regulator and modulates platinum drugs response in cancer cells.

For many tumor types chemotherapy still represents the therapy of choice and many standard treatments are based on the use of platinum (PT) drugs. However, de novo or acquired resistance to platinum is frequent and leads to disease progression. In Epithelial Ovarian Cancer (EOC) patients, PT-resistant recurrences are very common and improving the response to treatment still represents an unmet clinical need. To identify new modulators of PT-sensitivity, we performed a loss-of-function screening targeting 680 genes potentially involved in the response of EOC cells to platinum. We found that SGK2 (Serum-and Glucocorticoid-inducible kinase 2) plays a key role in PT-response. We show here that EOC cells relay on the induction of autophagy to escape PT-induced death and that SGK2 inhibition increases PT sensitivity inducing a block in the autophagy cascade due to the impairment of lysosomal acidification. Mechanistically we demonstrate that SGK2 controls autophagy in a kinase-dependent manner by binding and inhibiting the V-ATPase proton pump. Accordingly, SGK2 phosphorylates the subunit V1H (ATP6V1H) of V-ATPase and silencing or chemical inhibition of SGK2, affects the normal autophagic flux and sensitizes EOC cells to platinum. Hence, we identified a new pathway that links autophagy to the survival of cancer cells under platinum treatment in which the druggable kinase SGK2 plays a central role. Our data suggest that blocking autophagy via SGK2 inhibition could represent a novel therapeutic strategy to improve patients' response to platinum.

Splicing factor proline- and glutamine-rich (SFPQ) protein regulates platinum response in ovarian cancer-modulating SRSF2 activity.

In epithelial ovarian cancer (EOC), response to platinum (PT)-based chemotherapy dictates subsequent treatments and predicts patients' prognosis. Alternative splicing is often deregulated in human cancers and can be altered by chemotherapy. Whether and how changes in alternative splicing regulation could impact on the response of EOC to PT-based chemotherapy is still not clarified. We identified the splicing factor proline and glutamine rich (SFPQ) as a critical mediator of response to PT in an unbiased functional genomic screening in EOC cells and, using a large cohort of primary and recurrent EOC samples, we observed that it is frequently overexpressed in recurrent PT-treated samples and that its overexpression correlates with PT resistance. At mechanistic level, we show that, under PT treatment, SFPQ, in complex with p54nrb, binds and regulates the activity of the splicing factor SRSF2. SFPQ/p54nrb complex decreases SRSF2 binding to caspase-9 RNA, favoring the expression of its alternative spliced antiapoptotic form. As a consequence, SFPQ/p54nrb protects cells from PT-induced death, eventually contributing to chemoresistance. Overall, our work unveils a previously unreported SFPQ/p54nrb/SRSF2 pathway that in EOC cells plays a central role in regulating alternative splicing and PT-induced apoptosis and that could result in the design of new possible ways of intervention to overcome PT resistance.

Identification and Characterization of a New Platinum-Induced TP53 Mutation in MDAH Ovarian Cancer Cells.

Platinum-based chemotherapy is the therapy of choice for epithelial ovarian cancer (EOC). Acquired resistance to platinum (PT) is a frequent event that leads to disease progression and predicts poor prognosis. To understand possible mechanisms underlying acquired PT-resistance, we have recently generated and characterized three PT-resistant isogenic EOC cell lines. Here, we more deeply characterize several PT-resistant clones derived from MDAH-2774 cells. We show that, in these cells, the increased PT resistance was accompanied by the presence of a subpopulation of multinucleated giant cells. This phenotype was likely due to an altered progression through the M phase of the cell cycle and accompanied by the deregulated expression of genes involved in M phase progression known to be target of mutant TP53. Interestingly, we found that PT-resistant MDAH cells acquired in the TP53 gene a novel secondary mutation (i.e., S185G) that accompanied the R273H typical of MDAH cells. The double p53S185G/R273H mutant increases the resistance to PT in a TP53 null EOC cellular model. Overall, we show how the selective pressure of PT is able to induce additional mutation in an already mutant TP53 gene in EOC and how this event could contribute to the acquisition of novel cellular phenotypes.

Common biological phenotypes characterize the acquisition of platinum-resistance in epithelial ovarian cancer cells.

Standard of care for Epithelial Ovarian Cancer (EOC) patients relies on platinum-based therapy. However, acquired resistance to platinum occurs frequently and predicts poor prognosis. To understand the mechanisms underlying acquired platinum-resistance, we have generated and characterized three platinum-resistant isogenic EOC cell lines. Resistant cells showed 3-to 5- folds increase in platinum IC50. Cross-resistance to other chemotherapeutic agents commonly used in the treatment of EOC patients was variable and dependent on the cell line utilized. Gene expression profiling (GEP) of coding and non-coding RNAs failed to identify a common signature that could collectively explain the mechanism of resistance. However, we observed that all resistant cell lines displayed a decreased level of DNA platination and a faster repair of damaged DNA. Furthermore, all platinum resistant cell lines displayed a change in their morphology and a higher ability to grown on mesothelium. Overall, we have established and characterized three new models of platinum-resistant EOC cell lines that could be exploited to further dissect the molecular mechanisms underlying acquired resistance to platinum. Our work also suggests that GEP studies alone, at least when performed under basal culture condition, do not represent the optimal way to identify molecular alterations linked to DNA repair pathway defects.