Development of protein-based antiviral drugs for human papillomaviruses.
Recently, we designed artificial zinc-finger proteins (AZPs) that prevent a viral replication protein, E2, of human papillomavirus type 18 (HPV-18) from binding to its replication origin and demonstrated that the gene-delivered AZPs inhibited HPV-18 DNA replication in mammalian cells. In the present study, we examined another approach to delivery of AZPs. We constructed cell-permeable AZPs by fusing an AZP previously generated for inhibition of HPV-18 DNA replication with a cell-penetrating peptide (CPP), and confirmed that these CPP-AZP fusions reduced the replication rate in transient replication assays when added to the culture medium. In particular, 250 nM CPP-AZP reduced HPV-18 DNA replication to 3% of that of a control experiment. Western blot analysis detected 7% of the CPP-AZP added to the culture medium in the cell lysates, and demonstrated that greater internalization of CPP-AZP into mammalian cells causes greater inhibition of viral DNA replication. Furthermore, CPP-AZP did not show any significant cytotoxixity in MTT assays. Thus, our results demonstrate that cell-permeable AZPs could serve as potent protein-based antiviral drugs.
Mino
Takashi
T
Department of Synthetic Chemistry and Biological Chemistry, Graduate School of Engineering, Kyoto University, Kyotodaigaku-Katsura, Nishikyo-ku, Kyoto 615-8510, Japan.
Mori
Tomoaki
T
Aoyama
Yasuhiro
Y
Sera
Takashi
T
D000998:Antiviral Agents / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism / Q000494:pharmacology; D052161:Human papillomavirus 18 / Q000187:drug effects* / Q000502:physiology; D009856:Oncogene Proteins, Viral / Q000037:antagonists & inhibitors*; D010455:Peptides / Q000737:chemistry; D014779:Virus Replication / Q000187:drug effects; D016335:Zinc Fingers
D000998:Antiviral Agents; D004268:DNA-Binding Proteins; D009856:Oncogene Proteins, Viral; D010455:Peptides; C069102:oncogene protein E2, Human papillomavirus type 18