Download - Discovery Novel Allosteric Fragment Inhibitors of HIV-1 Reverse Transcriptase for HIV Prevention
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Discovery Novel Allosteric Fragment Inhibitors of HIV-1 Reverse
Transcriptase for HIV Prevention
A/Prof Gilda Tachedjian Retroviral Biology and Antivirals Laboratory
Centre for Biomedical ResearchBurnet Institute
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Discover Novel Allosteric Inhibitors of HIV-1 RT
Polymerase active site
NNRTI binding pocket
RNase H active site
HIV RT inhibitors approved or being developed for oral and topical PrEP same drug classes used for therapy
Potential for drug resistance in the contextof PrEP use in a real life setting
13 RT inhibitors used in the clinic they onlybelong to two classes: NRTIs and NNRTIs
Conformational flexibility of RT – function
Additional druggable allosteric sites in RT
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• “Fragments” are chemicals MW < 250 Da• More efficient at probing the chemical space – sample greater
chemical diversity by screening a smaller library• Bind with weak affinity • Strategically elaborated into larger high affinity inhibitors• Validated approach – US FDA approved drug Vemurafinib
Fragment Based Drug Discovery (FBDD)
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Three Novel Fragments Inhibit NNRTI Resistant HIV-1 RT
Inhibition of HIV-1 RT DDDP
4A2 inhibits RNase H
IC50 178 µM
Screen
13%
1.1%
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V89 and 4A2 Competes with Template/Primer and Cell culture data
4A2 inhibits HIV-1
EC50 18 ± 4 µM
4A2 Competes with T/P
IC50 93 ± 1 µM
V89 Competes with dNTP
Ki 220 ± 74 µM
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Identified fragments with novel scaffolds and modes of action compared to HIV RT inhibitors used clinically
Structure activity relationship (SAR) and X-ray crystallography studies are in progress to: - identify optimised fragments (more potent than original hits) - identify binding sites on HIV-1 RT - elaborate fragments into potent inhibitors by structure-based drug design
Summary
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Tachedjian Lab
Univ of PittsburghNicolas Sluis-Cremer
Acknowledgments
Rutgers University Eddy ArnoldJoe Bauman
Monash Institute of Pharmaceutical Sciences (MIPS)David ChalmersMartin ScanlonSteve Headey
Jennifer LaCath LathamDavid TyssenAdam Johnson
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•Detects weak binders•Can screen mixtures of compounds (x 5)•Pulse saturates entire protein (receptor) with magnetization transferring to protein bound ligand•Once ligand dissociates, saturated ligands can be detected•Resonances of small fragments not directly affected by pulse
Magnetic Field
RT
fragmentDissociate
Magnetisation transfer to fragment
Detect bound fragments
Saturation Transfer Difference Nuclear Magnetic Resonance (STD NMR)
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• Library of 630 fragments (ave MW 208) - Astex “Rule of Three” to make sure fragment like i.e. mass≤300 Da, ≤3 H-bond acceptors, ≤3H-bond donors, a clogP of ≤3, rotatable bonds ≤3 and a polar surface <60A2
Maybridge Ro3 library
Fragment Library
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Fragments are generally less potent against MoMLV and do not inhibit Klenow DNA pol