Wip1 phosphatase inhibition enhances etoposide mediated apoptosis by increasing p53 phosphorylation in Rhabdoid tumor cells

Protein phosphatase magnesium-dependent 1 delta (PPM1D/Wip1) is a genotoxic stress-activated serine/threonine phosphatase playing a central role in the timely termination of DNA damage response (DDR). Wip1 is frequently mutated, amplified, and overexpressed in common human cancers and is recognized as an oncogene. Rhabdoid tumors (RT) are a type of soft tissue sarcoma being quite aggressive and resistant to chemotherapy. In this study, the role of Wip1 in the regulation of genotoxic stress-induced apoptosis was studied in RT cell line A204. First PPM1D mRNA levels were analyzed in RT compared to normal tissues by in silico analysis utilizing RNA-seq data generated by TARGET from TCGA Pan Cancer. Second, MCF-7, BJ, and A204 cells were used to test the gene copy number and mRNA levels of Wip1 by qRT-PCR. Small molecule inhibitor GSK2830371 and etoposide were used to target Wip1 and to trigger genotoxic stress, respectively. MTT and AnexinV/7AAD analyzes were used to determine the viability and apoptotic response of A204 cells. Inhibition of Wip1 and total and phosphorylated levels of p53 was demonstrated by WB analysis. Thus, here we showed that Wip1 is overexpressed and amplified in the A204 RT cell line. Wip1 overexpressing A204 cells display resistance to etoposide-induced apoptosis due to decreased phosphorylation of p53. Targeting of Wip1 by GSK2830371 increased the phosphorylation of p53 in response to etoposide treatment. Additionally combined use of GSK2830371 and etoposide enhances p53-mediated apoptosis in A204 cells.