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EBNA1-targeted probe for the imaging and growth inhibition of tumours associated with the Epstein–Barr virus

Nature Biomedical Engineering volume 1, Article number: 0042 (2017) Cite this article

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Abstract

Epstein–Barr nuclear antigen 1 (EBNA1), a dimeric oncoprotein of the Epstein–Barr virus (EBV), is essential for both viral-genome maintenance and the survival of infected cells. Despite EBNA1’s potential as a therapeutic target, tools for the direct monitoring of EBNA1 in vitro and in vivo are lacking. Here, we show that a peptide-based inhibitor that luminesces when bound to EBNA1 inside the nucleus of EBV+ cells can regulate EBNA1 homodimer formation and selectively inhibit the growth of EBV+ tumours of nasopharyngeal carcinoma cells (C666-1 and NPC43) and Burkitt’s lymphoma Raji cells. We also show that the peptide-based probe leads to 93% growth inhibition of EBV+ tumours in mice. Our findings support the hypothesis that selective inhibition of EBNA1 dimerization can be used to afford better EBV-related cancer differentiation, and highlight the potential application of the probe as a new generation of biotracers for investigating the fundamental biological function of EBNA1 and for exploring its application as a therapeutic target.

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Figure 1: Rational design of probes and MD simulations of their binding to EBNA1.
Figure 2: Selectivity and emission of L2P4 on addition of EBNA1.
Figure 3: In vitro imaging of probes in EBV-positive and EBV-negative cells.
Figure 4: Homodimerization and cytotoxicity assays.
Figure 5: In vivo studies of P4 and L2P4 as EBV-specific anti-cancer agents.

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Acknowledgements

This work is funded by the Hong Kong Baptist University (FRG2/14-15/013013), Hong Kong Polytechnic University (HKPolyU), Hong Kong Research Grants Council (HKBU 20301615), Hong Kong Polytechnic University Central Research Grant (G-UC08), Hong Kong Research Grants Council (PolyU 153012/15P), University Research Facility for Chemical and Environmental Analysis (UCEA) and Area of Excellent Grants (1-ZVGG) of Hong Kong Polytechnic University, ECS-Grant - RGC (PolyU 253002/14P), HK PolyU (PolyU 5096/13P), HKBU and HKPolyU Joint Research Programme (RC-ICRS/15-16/02F-WKL02F-WKL), the EPSRC (Durham University, DTA award) and Research Grants Council of the Hong Kong SAR for the NPC Area of Excellence (AoE/M 06/08 Center for Nasopharyngeal Carcinoma Research).

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Authors and Affiliations

  1. Department of Chemistry, Hong Kong Baptist University, Kowloon Tong, Hong Kong SAR, China

    Lijun Jiang, Rongfeng Lan, Hongguang Li, Wai-Lun Chan & Ka-Leung Wong

  2. Clinical Division, School of Chinese Medicine, Hong Kong Baptist University, Kowloon Tong, Hong Kong SAR, China

    Tao Huang & Zhao-Xiang Bian

  3. Department of Applied Biology and Chemical Technology, State Key Laboratory of Chiroscience, Hong Kong Polytechnic University, Hung Hum, Hong Kong SAR, China

    Chi-Fai Chan, Wing-Yan Wong, Magnolia Muk-Lan Lee, Brandon Dow Chan, Wai-Sum Lo, William C. S. Tai, Ga-Lai Law & Wing-Tak Wong

  4. Department of Chemistry, Durham University, Durham, DH1 3LE, UK

    Sam Lear, Jingyi Zong & Steven L. Cobb

  5. Department of Biology, Hong Kong Baptist University,

    Nai-Ki Mak & Hong Lok Lung

  6. Centre for Nasopharyngeal Carcinoma Research, University of Hong Kong, Hong Kong SAR, China

    Nai-Ki Mak, Maria Li Lung, Hong Lok Lung & Sai Wah Tsao

  7. Department of Clinical Oncology, University of Hong Kong, Hong Kong SAR, China

    Maria Li Lung

  8. School of Biomedical Sciences, University of Hong Kong, Hong Kong SAR, China

    Sai Wah Tsao

  9. School of Cancer Sciences, University of Birmingham, Vincent Drive, Birmingham, B15 2TT, UK

    Graham S. Taylor

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  1. Lijun Jiang
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Contributions

N.-K.M. and K.-L.W. conceived and supervised the project. L.J. performed the synthesis, characterization, spectroscopic properties measurements and cytotoxicity assays in normal and some EBV-negative cells; H.L. contributed the photophysical data analysis. R.L. purified the WT-EBNA1 and EBNA1 mutants, and conducted the dimerization assay; T.H., Z.-X.B. and W.-T.W. contributed to the MD simulations; C.-F.C. carried out the confocal imaging, co-localization and some cytotoxicity assays; S.L., J.Z. and S.L.C. performed the peptide synthesis and purified the peptide conjugates; W.-Y.W., M.M.-L.L., B.D.C. and W.C.S.T. carried out the in vivo inhibition. M.L.L., H.L.L., S.W.T. and G.S.T. contributed to the in vitro cytotoxicity in EBV-positive and EBV-negative pairs of cancer cell lines; G.-L.L. participated in the cell work and molecular design; L.J., W.-L.C., W.-S.L., G.-L.L., S.L.C. and K.-L.W. wrote the manuscript.

Corresponding authors

Correspondence to Nai-Ki Mak, Ga-Lai Law, Wing-Tak Wong, Steven L. Cobb or Ka-Leung Wong.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Information

Supplementary methods, figures, tables and references. (PDF 11131 kb)

Supplementary Video 1

Dynamic distribution of L2P4 in EBV-negative HeLa cells. (MOV 2419 kb)

Supplementary Video 2

Dynamic distribution of L2P4 in EBV-positive C666-1 cells. (MOV 2723 kb)

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Jiang, L., Lan, R., Huang, T. et al. EBNA1-targeted probe for the imaging and growth inhibition of tumours associated with the Epstein–Barr virus. Nat Biomed Eng 1, 0042 (2017). https://doi.org/10.1038/s41551-017-0042

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