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Norcantharidin Inhibits Pre-Replicative Complexes Assembly of HepG2 Cells

Sansan Chen

Institute of Biotherapy, School of Biotechnology, Southern Medical University, Guangzhou, China

Department of Urology, Nanfang Hospital, Southern Medical University, Guangzhou, China

These authors contributed equally to this work.

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Xinming Qu

Science and Technology Office, Changchun University of Chinese Medicine, Changchun, China

These authors contributed equally to this work.

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Pei Wan

Department of Urology, Nanfang Hospital, Southern Medical University, Guangzhou, China

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Qing Wen Li

Institute of Biotherapy, School of Biotechnology, Southern Medical University, Guangzhou, China

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Ziyi Wang

Department of Biochemistry and Center for Cancer Research, Hong Kong University of Science and Technology, Hong Kong, China

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Fangfang Guo

Institute of Biotherapy, School of Biotechnology, Southern Medical University, Guangzhou, China

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Li Bai

Institute of Biotherapy, School of Biotechnology, Southern Medical University, Guangzhou, China

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Zhiming Hu

Institute of Biotherapy, School of Biotechnology, Southern Medical University, Guangzhou, China

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Wanlong Tan

Department of Urology, Nanfang Hospital, Southern Medical University, Guangzhou, China

Correspondence to: Dr. Wanlong Tan, Department of Urology, Nanfang Hospital, Southern Medical University, 1838 Guangzhou Avenue, Guangzhou 510515, China. Tel: (+86) 20-6164-1762.

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Jinlong Li

Institute of Biotherapy, School of Biotechnology, Southern Medical University, Guangzhou, China

Correspondence to: Dr. Jinlong Li, Institute of Biotherapy, School of Biotechnology, Southern Medical University, 1838 Guangzhou Dadao Bei, Guangzhou 510515, China. Tel: (+86) 20-6278-9135.

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https://doi.org/10.1142/S0192415X13500468Cited by:13 (Source: Crossref)

Abstract

Norcantharidin (NCTD) is currently used for anticancer therapy but the exact mechanism of action remains unknown. Pre-replicative complexes (pre-RCs) are essential for cell DNA replication and highly related to malignant proliferation. Here, we examined the inhibitory effect of NCTD on pre-RC components in HepG2 cells. We showed that NCTD induced degradation of Cdc6 and Mcm2 in a dose-dependent manner. Under 100 μM NCTD concentration, about 70% of Cdc6 and 50% of Mcm2 were degraded. In addition, the nuclear translocation of Mcm6 was inhibited by NCTD. Further studies aiming at G1 synchronous cells showed that, NCTD reduced the chromatin-bound Cdc6, Mcm2 and Mcm6. Moreover, the cells were blocked from entering the S phase and accumulated at the G1 phase when released synchronously into the cell cycle. Consistently, the DNA replication was inhibited by NCTD. Finally, the combination NCTD with Cdc6 depletion lead to more severe cytotoxicity (88%) than NCTD (52%) and Cdc6 depletion (39%) alone. A synergic cytotoxicity was observed between Cdc6 depletion and NCTD. In conclusion, our results demonstrate that NCTD inhibits pre-RC assembly; subsequently blocks the G1 to S transition; and inhibits DNA replication in HepG2 cells. Pre-RCs are an intriguing target for cancer therapy, which merits further investigations for anticancer development.

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