Dual‐Specificity Phosphatase 9 Regulates Cellular Proliferation and Predicts Recurrence After Surgery in Hepatocellular Carcinoma

Abstract

Hepatocellular carcinoma (CC) is a common and deadly cancer with complex molecular pathogenesis. Little is known about dual‐specificity phosphatases (DUSPs) in HCC. We investigated DUSP9 expression in human HCC, associations between DUSP9 and patient outcomes, and effects of altered DUSP9 expression on HCC biology. We studied public data sets as well as 196 patients at our institution who had HCC resections. Quantitative real‐time reverse transcription polymerase chain reaction and western blot demonstrated that DUSP9 expression was increased >10‐fold in HCC compared to adjacent liver and healthy controls (P = 0.005). Kaplan‐Meier and multivariable regression analyses revealed that higher DUSP9 expression was associated with shorter disease‐free survival (high DUSP9, 1.6; 95% confidence interval, 0.9‐2.3 vs. low DUSP9, 3.4; 95% confidence interval, 1.8‐5.0 years; P = 0.04) and increased risk of recurrence (hazard ratio 1.55; 95% confidence interval, 1.01‐2.67; P = 0.05) after resection. DUSP9 complementary DNA (cDNA) was cloned using rapid amplification of cDNA ends, revealing two DUSP9 isoforms in human HCC cells. Studies of transcriptional regulation using promoter‐luciferase reporter constructs suggested that DUSP9 transcription is regulated by E26 transformation‐specific transcription factors. Proliferation of hepatic cells in vitro was enhanced by lentiviral‐mediated overexpression of DUSP9. In contrast, DUSP9 knockout HCC cells generated using clustered regularly interspaced short palindromic repeats (CRISPR) demonstrated decreased HCC proliferation and doxorubicin resistance in vitro and impaired xenograft growth in vivo. RNA sequencing, gene set enrichment, and network/pathway analysis revealed that DUSP9 knockout is associated with activation of protein kinase activity and apoptosis. Conclusion: DUSP9 regulates cell proliferation and predicts recurrence after surgery in HCC. DUSP9 may represent a novel prognostic candidate and therapeutic target. Additional studies are warranted to further explore the role and regulation of DUSP9 in HCC.