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CRISPR/Cas9 Editing of the Polyomavirus Tumor Antigens Inhibits Merkel Cell Carcinoma Growth In Vitro

Laboratory of Virology and Chemotherapy, Department of Microbiology, Immunology and Transplantation, Rega Institute for Medical Research, KU Leuven, 3000 Leuven, Belgium
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Cancers 2019, 11(9), 1260; https://doi.org/10.3390/cancers11091260
Received: 16 July 2019 / Revised: 21 August 2019 / Accepted: 24 August 2019 / Published: 28 August 2019
Merkel cell carcinoma (MCC) is an aggressive type of skin cancer whose main causative agent is Merkel cell polyomavirus (MCPyV). MCPyV is integrated into the genome of the tumor cells in most MCCs. Virus-positive tumor cells constitutively express two viral oncoproteins that promote cell growth: the small (sT) and the large (LT) tumor antigens (TAs). Despite the success of immunotherapies in patients with MCC, not all individuals respond to these treatments. Therefore, new therapeutic options continue to be investigated. Herein, we used CRISPR/Cas9 to target the viral oncogenes in two virus-positive MCC cell lines: MS-1 and WAGA. Frameshift mutations introduced in the target sequence upon repair of the Cas9-induced DNA break resulted in decreased LT protein levels, which subsequently impaired cell proliferation, caused cell cycle arrest, and led to increased apoptosis. Importantly, a virus-negative non-MCC cell line (HEK293T) remained unaffected, as well as those cells expressing a non-targeting single-guide RNA (sgRNA). Thus, we presumed that the noted effects were not due to the off-target activity of the TAs-targeting sgRNAs. Additionally, WAGA cells had altered levels of cellular proteins involved in cell cycle regulation, supporting the observed cell cycle. Taken together, our findings provide evidence for the development of a CRISPR/Cas9-based therapeutic option for virus-positive MCC. View Full-Text
Keywords: Merkel cell carcinoma; Merkel cell polyomavirus; viral tumor antigens; CRISPR/Cas9; cell cycle regulation; novel therapeutics Merkel cell carcinoma; Merkel cell polyomavirus; viral tumor antigens; CRISPR/Cas9; cell cycle regulation; novel therapeutics
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Temblador, A.; Topalis, D.; Andrei, G.; Snoeck, R. CRISPR/Cas9 Editing of the Polyomavirus Tumor Antigens Inhibits Merkel Cell Carcinoma Growth In Vitro. Cancers 2019, 11, 1260.

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