USP29-mediated HIF1a stabilization promotes Sorafenib resistance of hepatocellular carcinoma cells by upregulating glycolysis. USP29-mediated HIF1a stabilization promotes Sorafenib resistance of hepatocellular carcinoma cells by upregulating glycolysis

Understanding the mechanisms underlying evasive resistance in cancer is an unmet medical need to improve the efficacy of current therapies. In hepatocellular carcinoma (HCC), aberrant expression of hypoxia inducible factor 1 a (HIF1a) and increased aerobic glycolysis metabolism represent drivers of the development of resistance to therapy with the multi-kinase inhibitor Sorafenib. However, it has remained unknown how HIF1a is activated and how its activity and the subsequent induction of aerobic glycolysis promotes Sorafenib resistance in HCC. Here, we report the ubiquitin-specific peptidase USP29 as a new regulator of HIF1a and of aerobic glycolysis during the development of Sorafenib resistance in HCC. In particular, we have identified USP29 as a critical deubiquitylase (DUB) of HIF1a, which directly deubiquitinates and stabilizes HIF1a and, thus, promotes its transcriptional activity. Among the transcriptional targets of HIF1a is the gene encoding for hexokinase 2 (HK2), a key enzyme of the glycolytic pathway. The absence of USP29, and thus of HIF1a transcriptional activity, reduces the levels of aerobic glycolysis and reinstalls the sensitivity to Sorafenib treatment in Sorafenib-resistant HCC cells in vitro and in xenograft transplantation mouse models in vivo. Notably, the absence of USP29 and high HK2 expression levels correlate with the response of HCC patients to Sorafenib therapy. Together, the data demonstrate that, as a DUB of HIF1a, USP29 promotes Sorafenib resistance in HCC cells by upregulating glycolysis, thereby opening new avenues for therapeutically targeting Sorafenib-resistant HCC in patients. Overall design: RNA-Seq of Huh7-Parental and established Huh7-IR, Huh7-CR cell lines

阐明癌症逃避性耐药的潜在机制，是改善当前疗法疗效过程中尚未满足的临床需求。在肝细胞癌（hepatocellular carcinoma, HCC）中，缺氧诱导因子1α（hypoxia inducible factor 1 α, HIF1α）的异常表达与有氧糖酵解代谢增强，是多激酶抑制剂索拉非尼（multi-kinase inhibitor Sorafenib）治疗耐药发生的驱动因素。然而，目前仍不清楚HIF1α的激活机制，以及其活性与后续诱导的有氧糖酵解如何促进肝细胞癌对索拉非尼的耐药性。本研究发现泛素特异性蛋白酶29（ubiquitin-specific peptidase USP29, USP29）是肝细胞癌索拉非尼耐药进程中，HIF1α与有氧糖酵解的新型调控因子。具体而言，本研究证实USP29是HIF1α的关键去泛素化酶（deubiquitylase, DUB），可直接对HIF1α进行去泛素化修饰并稳定其蛋白水平，进而增强其转录活性。HIF1α的转录靶基因包括编码糖酵解通路关键酶己糖激酶2（hexokinase 2, HK2）的基因。在体外索拉非尼耐药肝细胞癌细胞系以及体内异种移植小鼠模型中，缺失USP29进而抑制HIF1α转录活性，可降低有氧糖酵解水平，并恢复细胞对索拉非尼治疗的敏感性。值得注意的是，肝细胞癌患者中USP29缺失与HK2高表达的表型，与患者对索拉非尼治疗的临床应答密切相关。综上，本研究数据证实，作为HIF1α的去泛素化酶，USP29通过上调糖酵解进程促进肝细胞癌细胞对索拉非尼的耐药性，这为临床靶向治疗索拉非尼耐药肝细胞癌患者提供了全新思路。研究整体设计：对亲本Huh7细胞系及已构建的Huh7-IR、Huh7-CR细胞系进行RNA测序（RNA-Seq）。