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Autor/a:
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Vamvaka, Evangelia; Evans, Abbey; Ramessar, Koreen; Krumpe, L. R. H.; Shattock, Robin J.; O’Keefe, Barry R.; Christou, Paul; Capell Capell, Teresa
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Notas:
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Cyanovirin-N (CV-N) is a lectin with potent antiviral activity that has been proposed as a component of microbicides for the prevention of infection with Human immunodeficiency virus (HIV). The production of protein-based microbicide components requires a platform that is sufficiently economical and scalable to meet the demands of the large at-risk population, particularly in resource poor developing countries. We, therefore, expressed CV-N in rice endosperm, because the dried seed is ideal for storage and transport and crude extracts could be prepared locally and used as a microbicide component without further purification. We found that crude extracts from rice seeds expressing up to 10 µg CV-N per gram dry seed weight showed dose-dependent gp120 binding activity, confirming that the protein was soluble, correctly folded and active. The recombinant lectin (OSCV-N) reduced the infectivity of HIV-1BaL (an R5 virus strain representing the majority of transmitted infections) by ~90 % but showed only weak neutralization activity against HIV-1RF (representative of X4 virus, rarely associated with transmission), suggesting it would be highly effective for pre-exposure prophylaxis against the vast majority of transmitted strains. Crude extracts expressing OSCV-N showed no toxicity towards human cells at working dilutions indicating that microbicide components produced in rice endosperm are safe for direct application as topical microbicides in humans.
The authors are grateful to Dr. Maite Sabalza Gallue´s for cloning the CV-N gene in the pRP5 plasmid and Ms. Jennifer Wilson for technical assistance with live-virus anti-HIV assays. In addition, the authors would like to acknowledge funding from the Ministerio de Ciencia e Innovacio´n, Spain (BIO2012- 35359), the Center CONSOLIDER on Agrigenomics funded by MICINN, Spain, and COST Action FA0804 (Molecular farming: plants as a production platform for high value proteins). This project was funded in whole or in part with federal funds from the National Cancer Institute, National Institutes of Health, under contract HHSN26120080001E, and supported in part by the Intramural Research Program of the NIH, National Cancer Institute, Center for Cancer Research. The content of this publication does not necessarily reflect the views or policies of the Department of Health and Human Services, nor does mention of trade names, commercial products, or organizations imply endorsement by the US Government. |
