Targeted sequencing reveals the mutational landscape responsible for sorafenib therapy in advanced hepatocellular carcinoma
Jing Tang, Chengjun Sui, Dongfang Wang, Xinyuan Lu, Guijuan Luo, Qing Zhao, Qiuyu Lian, Seogsong Jeong, Ximeng Lin, Yanjing Zhu, Bo Zheng, Rui Wu, Qing Wang, Xiaolong Liu, Jingfeng Liu, Qiang Xia, Gang Wu, Jin Gu, Hongyang Wang, Lei Chen
Abstract
The existence of primary and acquired drug resistance is the main obstacle for the effect of multi-kinase inhibitor sorafenib and regorafenib in advanced hepatocellular carcinoma (HCC). However, plenty of patients did not significantly benefit from sorafenib treatment and little is known about the mechanism of drug resistance. Methods: Laser capture microdissection was used to acquire matched normal liver and tumor tissues on formalin-fixed paraffin-embedded specimens collected before sorafenib therapy from the first surgery of 119 HCC patients. Ultra-deep sequencing (~1000) targeting whole exons of 440 genes in microdissected specimens and siRNA screen in 7 cell lines were performed to find mutations associated with differential responses to sorafenib. Patient-derived xenograft models were employed to determine the role of TP53 in response to sorafenib. Lentiviruses harboring wild-type and c.G52C-mutant OCT4 were applied to explore the function of OCT4 in resistance to sorafenib. ChIP-PCR assay for analysis of OCT4 transcriptional activity was performed to explore the affinity with the KITLG promoter. Statistical analyses were used to associate levels of p53 and OCT4 with tumor features and patient outcomes.