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Antimicrobial Agents and Chemotherapy, October 2006, p. 3407-3417, Vol. 50, No. 10
0066-4804/06/$08.00+0     doi:10.1128/AAC.00517-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Development of a Human Immunodeficiency Virus Vector-Based, Single-Cycle Assay for Evaluation of Anti-Integrase Compounds

Roberta Bona,1,{dagger} Mauro Andreotti,2,{dagger} Viviana Buffa,2 Pasqualina Leone,1 Clementina Maria Galluzzo,2 Roberta Amici,2 Lucia Palmisano,2 Maria Grazia Mancini,2 Zuleika Michelini,2 Roberto Di Santo,3 Roberta Costi,3 Alessandra Roux,3 Yves Pommier,4 Christophe Marchand,4 Stefano Vella,2 and Andrea Cara2*

National AIDS Center,1 Department of Drug Research and Evaluation, Istituto Superiore di Sanità, Viale Regina Elena 299, 00161 Rome, Italy,2 Istituto Pasteur-Fondazione Cenci Bolognetti, Dipartimento di Studi Farmaceutici, P.le Aldo Moro 5, 00185 Rome, Italy,3 Laboratory of Molecular Pharmacology, National Institutes of Health, Bethesda, Maryland 208924

Received 27 April 2006/ Returned for modification 13 June 2006/ Accepted 13 July 2006

Therapeutic strategies aimed at inhibiting human immunodeficiency virus type 1 (HIV-1) replication employ a combination of drugs targeted to two viral enzymes (reverse transcriptase and protease) and to the viral entry/fusion step. However, the high propensity of HIV-1 to develop resistance makes the development of novel compounds targeting different steps of the HIV-1 life cycle essential. Among these, integrase (IN) inhibitors have successfully passed the early phases of clinical development. By preventing integration, IN inhibitors preclude viral replication while allowing production of extrachromosomal forms of viral DNA (E-DNA). Here, we describe an improved and standardized assay aimed at evaluating IN inhibitors by taking advantage of the transcriptional activity of E-DNA produced by HIV-derived vectors in the absence of replication-competent virus. In this context, the use of the firefly luciferase gene as a reporter gene provides a rapid and quantitative measure of viral-vector infectivity, thus making it a safe and cost-effective assay for evaluating novel IN inhibitors.

* Corresponding author. Mailing address: Department of Drug Research and Evaluation, Istituto Superiore di Sanità, Viale Regina Elena 299, 00161, Rome, Italy. Phone: (39)06.4990.3503. Fax: (39)06.4938.7199. E-mail: acara{at}iss.it.

{dagger} R. Bona and M. Andreotti contributed equally to this work.

Antimicrobial Agents and Chemotherapy, October 2006, p. 3407-3417, Vol. 50, No. 10
0066-4804/06/$08.00+0     doi:10.1128/AAC.00517-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



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