Qian Wang1, Ning Liang2, Tao Yang3, Yuedan Li4, Jing Li5, Qian Huang6, Chen Wu7, Ligang Sun2, Xile Zhou8, Xiaobin Cheng8, Long Zhao8, Gang Wang9, Zhangqian Chen10, Xianli He11, Chaoxu Liu12. 1. Department of Anorectal Surgery, The First Affiliated Hospital, Zhengzhou University, Zhengzhou 450052, China. 2. Department of General Surgery, The 75(th) Group Army Hospital, Dali 671000, China. 3. Department of Pain Treatment, Tangdu Hospital, Air Force Military Medical University, Xi'an 710032, China. 4. Department of Medicinal Chemistry and Pharmaceutical Analysis, School of Pharmacy, Air Force Military Medical University, Xi'an, Shaanxi 710032, China. 5. College and Hospital of Stomatology, Xi'an Jiao Tong University, Xi'an 710000, China. 6. Department of Obstetrics and Gynecology, The 75(th) Group Army Hospital, Dali 671000, China. 7. Department of General Surgery, Huashan Hospital, Fudan University, Shanghai 201907, China. 8. Department of Colorectal Surgery, The First Affiliated Hospital of Zhejiang University, Hangzhou 310006, China. 9. Department of General Surgery, The 74(th) Group Army Hospital, Guangzhou 510318, China. Electronic address: wanggangfmmu@163.com. 10. State Key Laboratory of Cancer Biology, National Clinical Research Center for Digestive Diseases and Xijing Hospital of Digestive Diseases, Air Force Military Medical University, Xi'an 710032, Shaanxi, China. Electronic address: chenzhangqian996@163.com. 11. Department of General Surgery, Tangdu Hospital, Air Force Military Medical University, Xi'an 710032, Shaanxi, China. Electronic address: hexianli999@126.com. 12. Department of Colorectal Surgery, The First Affiliated Hospital of Zhejiang University, Hangzhou 310006, China. Electronic address: 1520003@zju.edu.cn.
Abstract
BACKGROUND & AIMS: Hepatoblastoma (HB) and hepatocellular carcinoma (HCC) both exhibit notable cancer stem cell (CSC) features. Moreover, the development of both diseases is closely associated with the presence of CSCs. We investigated the role of brain-expressed X-linked protein 1 (BEX1) in regulating the CSC properties of HB and a subtype of HCC with high CSC features (CSC-HCC). METHODS: Stemness scores were analyzed in 5 murine HCC models. A subpopulation of BEX1-positive cells and BEX1-negative cells were sorted from HCC cell lines, and subjected to transcriptome analysis. The expression and function of BEX1 was examined via western blotting, sphere formation assays, and xenograft tumor models. RESULTS: We identified BEX1 as a novel CSC marker that was required for the self-renewal of liver CSCs. Furthermore, zebularine, a potent DNMT1 inhibitor, can induce the reactivation of BEX1 by removing epigenetic inhibition. Notably, BEX1 was highly expressed in patients with HB and CSC-HCC, but not in patients with non-CSC HCC. Moreover, DNMT1-mediated methylation of the BEX1 promoter resulted in differential BEX1 expression patterns in patients with HB, CSC-HCC, and non-CSC-HCC. Mechanistically, BEX1 interacted with RUNX3 to block its inhibition of β-catenin transcription, which led to the activation of Wnt/β-catenin signaling, and stemness maintenance in both HB and CSC-HCC. In contrast, downregulated BEX1 expression released RUNX3 and inhibited the activation of Wnt/β-catenin signaling in non-CSC-HCC. CONCLUSION: BEX1, under the regulation of DNMT1, is necessary for the self-renewal and maintenance of liver CSCs through activation of Wnt/β-catenin signaling, rendering BEX1 a potentially valuable therapeutic target in both HB and CSC-HCC. LAY SUMMARY: Cancer stem cells (CSCs) contribute to a high rate of cancer recurrence, as well as resistance to conventional therapies. However, the regulatory mechanisms underlying their self-renewal remains elusive. Herein, we have reported that BEX1 plays a key role in regulating CSC properties in different types of liver cancer. Targeting BEX1-mediated Wnt/β-catenin signaling may help to address the high rate of recurrence, and heterogeneity of liver cancer.
BACKGROUND & AIMS: Hepatoblastoma (HB) and hepatocellular carcinoma (HCC) both exhibit notable cancer stem cell (CSC) features. Moreover, the development of both diseases is closely associated with the presence of CSCs. We investigated the role of brain-expressed X-linked protein 1 (BEX1) in regulating the CSC properties of HB and a subtype of HCC with high CSC features (CSC-HCC). METHODS: Stemness scores were analyzed in 5 murine HCC models. A subpopulation of BEX1-positive cells and BEX1-negative cells were sorted from HCC cell lines, and subjected to transcriptome analysis. The expression and function of BEX1 was examined via western blotting, sphere formation assays, and xenograft tumor models. RESULTS: We identified BEX1 as a novel CSC marker that was required for the self-renewal of liver CSCs. Furthermore, zebularine, a potent DNMT1 inhibitor, can induce the reactivation of BEX1 by removing epigenetic inhibition. Notably, BEX1 was highly expressed in patients with HB and CSC-HCC, but not in patients with non-CSC HCC. Moreover, DNMT1-mediated methylation of the BEX1 promoter resulted in differential BEX1 expression patterns in patients with HB, CSC-HCC, and non-CSC-HCC. Mechanistically, BEX1 interacted with RUNX3 to block its inhibition of β-catenin transcription, which led to the activation of Wnt/β-catenin signaling, and stemness maintenance in both HB and CSC-HCC. In contrast, downregulated BEX1 expression released RUNX3 and inhibited the activation of Wnt/β-catenin signaling in non-CSC-HCC. CONCLUSION: BEX1, under the regulation of DNMT1, is necessary for the self-renewal and maintenance of liver CSCs through activation of Wnt/β-catenin signaling, rendering BEX1 a potentially valuable therapeutic target in both HB and CSC-HCC. LAY SUMMARY: Cancer stem cells (CSCs) contribute to a high rate of cancer recurrence, as well as resistance to conventional therapies. However, the regulatory mechanisms underlying their self-renewal remains elusive. Herein, we have reported that BEX1 plays a key role in regulating CSC properties in different types of liver cancer. Targeting BEX1-mediated Wnt/β-catenin signaling may help to address the high rate of recurrence, and heterogeneity of liver cancer.