Lo-Kong Chan1, Daniel Wai-Hung Ho2, Charles Shing Kam2, Elley Yung-Tuen Chiu2, Irene Lai-Oi Lo3, Derek Tsz-Wai Yau4, Elaine Tin-Yan Cheung4, Chung-Ngai Tang5, Victor Wai-Lun Tang6, Terence Kin-Wah Lee7, Carmen Chak-Lui Wong2, Kenneth Siu-Ho Chok8, Albert Chi-Yan Chan8, Tan-To Cheung8, Chun-Ming Wong2, Irene Oi-Lin Ng9. 1. Department of Pathology, The University of Hong Kong, Hong Kong; State Key Laboratory of Liver Research, The University of Hong Kong, Hong Kong. Electronic address: clkchan@pathology.hku.hk. 2. Department of Pathology, The University of Hong Kong, Hong Kong; State Key Laboratory of Liver Research, The University of Hong Kong, Hong Kong. 3. Department of Surgery, Queen Elizabeth Hospital, Hong Kong. 4. Department of Pathology, Queen Elizabeth Hospital, Hong Kong. 5. Department of Surgery, Pamela Youde Nethersole Eastern Hospital, Hong Kong. 6. Department of Pathology, Pamela Youde Nethersole Eastern Hospital, Hong Kong. 7. Department of Applied Biology and Chemical Technology, The Hong Kong Polytechnic University, Hong Kong. 8. State Key Laboratory of Liver Research, The University of Hong Kong, Hong Kong; Department of Surgery, The University of Hong Kong, Hong Kong. 9. Department of Pathology, The University of Hong Kong, Hong Kong; State Key Laboratory of Liver Research, The University of Hong Kong, Hong Kong. Electronic address: iolng@hku.hk.
Abstract
BACKGROUND & AIMS: Mutational profiling of patient tumors has suggested that hepatocellular carcinoma (HCC) development is mainly driven by loss-of-function mutations in tumor suppressor genes. p90 ribosomal S6 kinase 2 (RSK2) functions as a direct downstream kinase of ERK1/2 and elevated RSK2 expression has been reported to support oncogenic functions in some cancers. We investigated if RSK2 was also dysregulated by inactivating mutations in cancers including HCC. METHODS: We performed exome sequencing and targeted DNA sequencing on HBV-associated HCCs to examine recurrent RSK2 mutations. The functional significance and mechanistic consequences of RSK2 mutations were examined in natural RSK2-null HCC cells, and RSK2-knockout HCC cells. The potential downstream pathways underlying RSK2 mutations were investigated by RNA sequencing, qRT-PCR and mass spectrometry. RESULTS: We detected recurrent somatic RSK2 mutations at a rate of 6.3% in our HCC cohorts and revealed that, among many cancer types, HCC was the cancer most commonly harboring RSK2 mutations. The RSK2 mutations were inactivating and associated with a more aggressive tumor phenotype. We found that, functionally, restoring RSK2 expression in natural RSK2-null HBV-positive Hep3B cells suppressed proliferation and migration in vitro and tumorigenicity in vivo. Mechanistically, RSK2-inactivating mutations attenuated a SOS1/2-dependent negative feedback loop, leading to the activation of MAPK signaling. Of note, this RSK2 mutation-mediated MAPK upregulation rendered HCC cells more sensitive to sorafenib, a first-line multi-kinase inhibitor for advanced HCC. Furthermore, such activation of MAPK signaling enhanced cholesterol biosynthesis-related gene expression in HCC cells. CONCLUSIONS: Our findings reveal the mechanistic and functional significance of RSK2-inactivating mutations in HCC. These inactivating mutations may serve as an alternative route to activate MAPK signaling and cholesterol metabolism in HCC. LAY SUMMARY: In this study, we identified and functionally characterized RSK2-inactivating mutations in human hepatocellular carcinoma and demonstrated their association with aggressive tumor behavior. Mutations in RSK2 drive signaling pathways with known oncogenic potential, leading to enhanced cholesterol biosynthesis and potentially sensitizing tumors to sorafenib treatment.
BACKGROUND & AIMS: Mutational profiling of patient tumors has suggested that hepatocellular carcinoma (HCC) development is mainly driven by loss-of-function mutations in tumor suppressor genes. p90 ribosomal S6 kinase 2 (RSK2) functions as a direct downstream kinase of ERK1/2 and elevated RSK2 expression has been reported to support oncogenic functions in some cancers. We investigated if RSK2 was also dysregulated by inactivating mutations in cancers including HCC. METHODS: We performed exome sequencing and targeted DNA sequencing on HBV-associated HCCs to examine recurrent RSK2 mutations. The functional significance and mechanistic consequences of RSK2 mutations were examined in natural RSK2-null HCC cells, and RSK2-knockout HCC cells. The potential downstream pathways underlying RSK2 mutations were investigated by RNA sequencing, qRT-PCR and mass spectrometry. RESULTS: We detected recurrent somatic RSK2 mutations at a rate of 6.3% in our HCC cohorts and revealed that, among many cancer types, HCC was the cancer most commonly harboring RSK2 mutations. The RSK2 mutations were inactivating and associated with a more aggressive tumor phenotype. We found that, functionally, restoring RSK2 expression in natural RSK2-null HBV-positive Hep3B cells suppressed proliferation and migration in vitro and tumorigenicity in vivo. Mechanistically, RSK2-inactivating mutations attenuated a SOS1/2-dependent negative feedback loop, leading to the activation of MAPK signaling. Of note, this RSK2 mutation-mediated MAPK upregulation rendered HCC cells more sensitive to sorafenib, a first-line multi-kinase inhibitor for advanced HCC. Furthermore, such activation of MAPK signaling enhanced cholesterol biosynthesis-related gene expression in HCC cells. CONCLUSIONS: Our findings reveal the mechanistic and functional significance of RSK2-inactivating mutations in HCC. These inactivating mutations may serve as an alternative route to activate MAPK signaling and cholesterol metabolism in HCC. LAY SUMMARY: In this study, we identified and functionally characterized RSK2-inactivating mutations in human hepatocellular carcinoma and demonstrated their association with aggressive tumor behavior. Mutations in RSK2 drive signaling pathways with known oncogenic potential, leading to enhanced cholesterol biosynthesis and potentially sensitizing tumors to sorafenib treatment.
Authors: Jennifer Cable; Duanqing Pei; Lola M Reid; Xin Wei Wang; Sonam Bhatia; Panagiotis Karras; Jan Joseph Melenhorst; Markus Grompe; Justin D Lathia; Erwei Song; Calvin J Kuo; Ning Zhang; Richard M White; Stephanie Ky Ma; Lichun Ma; Y Rebecca Chin; Michael M Shen; Irene Oi Lin Ng; Klaus H Kaestner; Lei Zhou; Shaheen Sikandar; Clemens A Schmitt; Wei Guo; Carmen Chak-Lui Wong; Junfang Ji; Dean G Tang; Anna Dubrovska; Chunzhang Yang; Wolf R Wiedemeyer; Irving L Weissman Journal: Ann N Y Acad Sci Date: 2021-11-30 Impact factor: 6.499