Screening of novel inhibitors of HIV-1 reverse transcriptase with a reporter ribozyme assay.
"Highly active anti-retroviral therapy (HAART)" is currently the standard treatment for human immunodeficiency virus (HIV). This treatment consists of a cocktail of two reverse transcriptase (RT) inhibitors and a protease inhibitor. Despite the success of this regimen, there is a continuing need for innovative drug to overcome problems with tolerability and the emergence of viral resistance. The present protocol describes a novel strategy to rapidly screening a new class of small molecule HIV-1 RT inhibitors, which bind to the primer/template binding site of RT, as yet an unexplored site for small molecule interference on this target. The assay is based on aptamer-displacement which is visualized by applying a rationally designed HIV-1 RT responsive ribozyme. The handiness of the assay procedure permits automation, compatible with high-throughput screening (HTS). Subsequently, the identified hit compounds have been evaluated by an in vitro enzymatic assay to test the inhibitory potential. The strategy provides a powerful and efficient screening format for site-directed inhibitors with biological activity.
Yamazaki
Satoko
S
Life and Medical Sciences, University of Bonn, Bonn, Germany.
Famulok
Michael
M
D000494:Allosteric Regulation; D019380:Anti-HIV Agents / Q000737:chemistry / Q000494:pharmacology*; D052157:Aptamers, Nucleotide / Q000737:chemistry / Q000378:metabolism*; D015195:Drug Design; D015658:HIV Infections / Q000188:drug therapy; D054303:HIV Reverse Transcriptase / Q000037:antagonists & inhibitors* / Q000378:metabolism; D015497:HIV-1 / Q000201:enzymology / Q000235:genetics; D006801:Humans; D016337:RNA, Catalytic / Q000737:chemistry / Q000378:metabolism*; D018894:Reverse Transcriptase Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D054852:Small Molecule Libraries
D019380:Anti-HIV Agents; D052157:Aptamers, Nucleotide; D016337:RNA, Catalytic; D018894:Reverse Transcriptase Inhibitors; D054852:Small Molecule Libraries; C514824:reverse transcriptase, Human immunodeficiency virus 1; D054303:HIV Reverse Transcriptase