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Antimicrobial Agents and Chemotherapy, October 2006, p. 3250-3259, Vol. 50, No. 10
0066-4804/06/$08.00+0     doi:10.1128/AAC.00493-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Engineered Vaginal Lactobacillus Strain for Mucosal Delivery of the Human Immunodeficiency Virus Inhibitor Cyanovirin-N

Xiaowen Liu,¹ Laurel A. Lagenaur,¹ David A. Simpson,¹ Kirsten P. Essenmacher,¹ Courtney L. Frazier-Parker,¹ Yang Liu,¹ Daniel Tsai,¹ Srinivas S. Rao,² Dean H. Hamer,³ Thomas P. Parks,¹ Peter P. Lee,^4* and Qiang Xu^1*

Osel, Inc., 4008 Burton Drive, Santa Clara, California 95054,¹ Laboratory Animal Medicine, Vaccine Research Center, National Institutes of Health, Bethesda, Maryland 20892,² Laboratory of Biochemistry, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892,³ Department of Medicine, Stanford University, Stanford, California 94305⁴

Received 20 April 2006/ Returned for modification 7 June 2006/ Accepted 31 July 2006

Women are at significant risk of human immunodeficiency virus (HIV) infection, with the cervicovaginal mucosa serving as a major portal for virus entry. Female-initiated preventatives, including topical microbicides, are urgently needed to help curtail the HIV/AIDS pandemic. Here we report on the development of a novel, live microbicide that employs a natural vaginal strain of Lactobacillus jensenii engineered to deliver the potent HIV inhibitor cyanovirin-N (CV-N). To facilitate efficient expression of CV-N by this bacterium, the L. jensenii 1153 genome was sequenced, allowing identification of native regulatory elements and sites for the chromosomal integration of heterologous genes. A CV-N expression cassette was optimized and shown to produce high levels of structurally intact CV-N when expressed in L. jensenii. Lactobacillus-derived CV-N was capable of inhibiting CCR5-tropic HIV_BaL infectivity in vitro with a 50% inhibitory concentration of 0.3 nM. The CV-N expression cassette was stably integrated as a single copy into the bacterial chromosome and resolved from extraneous plasmid DNA without adversely affecting the bacterial phenotype. This bacterial strain was capable of colonizing the vagina and producing full-length CV-N when administered intravaginally to mice during estrus phase. The CV-N-producing Lactobacillus was genetically stable when propagated in vitro and in vivo. This work represents a major step towards the development of an inexpensive yet durable protein-based microbicide to block the heterosexual transmission of HIV in women.

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* Corresponding author. Mailing address for P. P. Lee: Department of Medicine, Stanford University, Stanford, CA 94305. Phone: (650) 498-7942. Fax: (650) 736-0974. E-mail: ppl{at}stanford.edu. Mailing address for Q. Xu: Osel, Inc., 4008 Burton Dr., Santa Clara, CA 95054. Phone: (408) 986-0012, ext. 205. Fax: (408) 986-0019. E-mail: qxu{at}oselinc.com.

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Antimicrobial Agents and Chemotherapy, October 2006, p. 3250-3259, Vol. 50, No. 10
0066-4804/06/$08.00+0     doi:10.1128/AAC.00493-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

This article has been cited by other articles:

• Liu, X., Lagenaur, L. A., Lee, P. P., Xu, Q. (2008). Engineering of a Human Vaginal Lactobacillus Strain for Surface Expression of Two-Domain CD4 Molecules. Appl. Environ. Microbiol. 74: 4626-4635 [Abstract] [Full Text]