Development of a human immunodeficiency virus vector-based, single-cycle assay for evaluation of anti-integrase compounds.
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.
Bona
Roberta
R
National AIDS Center, Istituto Superiore di Sanità, Viale Regina Elena 299, 00161, Rome, Italy.
Andreotti
Mauro
M
Buffa
Viviana
V
Leone
Pasqualina
P
Galluzzo
Clementina Maria
CM
Amici
Roberta
R
Palmisano
Lucia
L
Mancini
Maria Grazia
MG
Michelini
Zuleika
Z
Di Santo
Roberto
R
Costi
Roberta
R
Roux
Alessandra
A
Pommier
Yves
Y
Marchand
Christophe
C
Vella
Stefano
S
Cara
Andrea
A
D002460:Cell Line; D005822:Genetic Vectors; D019427:HIV Integrase / Q000187:drug effects; D019428:HIV Integrase Inhibitors / Q000494:pharmacology*; D015497:HIV-1 / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics; D006801:Humans; D020128:Inhibitory Concentration 50; D016086:Lentivirus / Q000235:genetics; D007963:Leukocytes, Mononuclear / Q000821:virology; D008156:Luciferases / Q000378:metabolism; D008826:Microbial Sensitivity Tests / Q000379:methods*; D014161:Transduction, Genetic
D019428:HIV Integrase Inhibitors; D008156:Luciferases; D019427:HIV Integrase