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Journal of Virology, February 2003, p. 2640-2650, Vol. 77, No. 4
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.4.2640-2650.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Developing Novel Oncolytic Adenoviruses through Bioselection

Wen Yan, Galila Kitzes, Farid Dormishian, Lynda Hawkins, Adam Sampson-Johannes, Josh Watanabe, Jenny Holt, Vivian Lee, Thomas Dubensky, Ali Fattaey, Terry Hermiston, Allan Balmain,^|| and Yuqiao Shen^*

ONYX Pharmaceuticals, Inc., Richmond, California 94806

Received 23 August 2002/ Accepted 12 November 2002

Mutants of human adenovirus 5 (Ad5) with enhanced oncolytic activity were isolated by using a procedure termed bioselection. Two mutants, ONYX-201 and ONYX-203, were plaque purified from a pool of randomly mutagenized Ad5 that was repeatedly passaged in the human colorectal cancer cell line HT29, and they were subsequently characterized. ONYX-201 and ONYX-203 replicated more rapidly in HT29 cells than wild-type Ad5, and they lysed HT29 cells up to 1,000-fold more efficiently. The difference was most profound when cells were infected at a relatively low multiplicity of infection, presumably due to the compounding effects of multiple rounds of infection. This enhanced cytolytic activity was observed not only in HT29 cells but also in many other human cancer cell lines tested. In contrast, the cytotoxicity of the bioselected mutants in a number of normal primary human cells was similar to that of wild-type Ad5, thus enhancing the therapeutic index (cytotoxicity in tumor cells versus that in normal cells) of these oncolytic agents. Both ONYX-201 and -203 contain seven single-base-pair mutations when compared with Ad5, four of which were common between ONYX-201 and -203. The mutation at nucleotide 8350, shared by both mutant viruses, was shown to be essential for the observed phenotypes. This mutation was mapped to the i-leader region of the major late transcription unit, resulting in the truncation of 21 amino acids from the C terminus of the i-leader protein. This work demonstrates that bioselection is a powerful tool for developing novel tumor-selective oncolytic viruses. Other potential applications of this technology are discussed.

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* Corresponding author. Mailing address: Onyx Pharmaceuticals, Inc., 3031 Research Dr., Richmond, CA 94806. Phone: (510) 243-3679. Fax: (510) 222-9758. E-mail: jshen{at}onyx-pharm.com.

Present address: Genetic Therapy, Inc., Gaithersburg, MD 20878.

Present address: Cerus Corporation, Concord, CA 94520.

Present address: Berlex Laboratory, Inc., Richmond, CA 94804.

^|| Present address: UCSF Comprehensive Cancer Center, San Francisco, CA 94115.

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Journal of Virology, February 2003, p. 2640-2650, Vol. 77, No. 4
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.4.2640-2650.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

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• Subramanian, T., Vijayalingam, S., Chinnadurai, G. (2006). Genetic Identification of Adenovirus Type 5 Genes That Influence Viral Spread. J. Virol. 80: 2000-2012 [Abstract] [Full Text]