title: Homogeneous high-throughput screening assays for HIV-1 integrase 3beta-processing and strand transfer activities.
abstract: HIV-1 integrase (HIV-IN) is a well-validated antiviral drug target catalyzing a multistep reaction to incorporate the HIV-1 provirus into the genome of the host cell. Small molecule inhibitors of HIV-1 integrase that specifically target the strand transfer step have demonstrated efficacy in the suppression of virus propagation. However, only few specific strand transfer inhibitors have been identified to date, and the need to screen for novel compound scaffolds persists. Here, the authors describe 2 homogeneous time-resolved fluorescent resonance energy transfer-based assays for the measurement of HIV-1 integrase 3'-processing and strand transfer activities. Both assays were optimized for high-throughput screening formats, and a diverse library containing more than 1 million compounds was screened in 1536-well plates for HIV-IN strand transfer inhibitors. As a result, compounds were found that selectively affect the enzymatic strand transfer reaction over 3beta processing. Moreover, several bioactive molecules were identified that inhibited HIV-1 reporter virus infection in cellular model systems. In conclusion, the assays presented herein have proven their utility for the identification of mechanistically interesting and biologically active inhibitors of HIV-1 integrase that hold potential for further development into potent antiviral drugs.
authors.affiliation: Genomics Institute of the Novartis Research Foundation, San Diego, CA 92121, USA.
authors.forename: Yu
authors.initials: Y
authors.lastname: Wang
authors.forename: Heath
authors.initials: H
authors.lastname: Klock
authors.forename: Hong
authors.initials: H
authors.lastname: Yin
authors.forename: Karen
authors.initials: K
authors.lastname: Wolff
authors.forename: Kimberly
authors.initials: K
authors.lastname: Bieza
authors.forename: Kirk
authors.initials: K
authors.lastname: Niswonger
authors.forename: Jason
authors.initials: J
authors.lastname: Matzen
authors.forename: Drew
authors.initials: D
authors.lastname: Gunderson
authors.forename: Joanna
authors.initials: J
authors.lastname: Hale
authors.forename: Scott
authors.initials: S
authors.lastname: Lesley
authors.forename: Kelli
authors.initials: K
authors.lastname: Kuhen
authors.forename: Jeremy
authors.initials: J
authors.lastname: Caldwell
authors.forename: Achim
authors.initials: A
authors.lastname: Brinker
mesh_terms: D000998:Antiviral Agents / Q000494:pharmacology; D002460:Cell Line; D045744:Cell Line, Tumor; D003001:Cloning, Molecular; D004305:Dose-Response Relationship, Drug; D004353:Drug Evaluation, Preclinical / Q000295:instrumentation / Q000379:methods*; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D019427:HIV Integrase / Q000235:genetics* / Q000378:metabolism*; D019428:HIV Integrase Inhibitors / Q000494:pharmacology; D006801:Humans; D020128:Inhibitory Concentration 50; D008957:Models, Genetic; D010713:Phosphatidylcholines / Q000494:pharmacology; D013997:Time Factors
chemical_list: D000998:Antiviral Agents; D019428:HIV Integrase Inhibitors; D010713:Phosphatidylcholines; C044819:1,2-diarachidonoyl-glycero-3-phosphocholine; D019427:HIV Integrase