Identification and evaluation of inhibitors targeting the HIV-1 integrase-LEDGF/p75 interaction

• Main Author: Harrison, Angela Theresa
• Format: Others
• Language: en
• Published: 2014
• Online Access: http://hdl.handle.net/10539/15433

A dissertation submitted to the Faculty of Health Sciences, University of the Witwatersrand, in fulfilment of the requirements for the degree of Master of Science in Medicine === The human immunodeficiency virus type 1 (HIV-1) integrase (IN) facilitates the irreversible integration of the viral chromosome into the host DNA in a two-step reaction process consisting of 3’ end processing and strand transfer. The pre-integration complex (PIC) is a nucleoprotein complex consisting of HIV-1 IN, reverse transcriptase, matrix, nucleocapsid, viral protein R (Vpr) and various cellular host proteins. The dominant host protein that forms part of the PIC is known as lens epithelium derived growth factor (LEDGF/p75) that is ubiquitously expressed in the nucleus. The integration of HIV-1 into chromatinised template is stimulated by LEDGF/p75, while LEDGF/p75 stabilizes HIV-1 IN subunit-subunit interactions and promotes HIV-1 IN tetramerisation. This study aimed to identify and evaluate potential inhibitors of the HIV-1 IN-LEDGF/p75 interaction. A compound library (NCC-202) comprised of 281 compounds obtained from the NIH clinical collection was screened through an HIV-1 IN-LEDGF/p75 molecular model prepared on Accelrys Discovery StudioTM 3.1. Recombinant HIV-1 IN and LEDGF/p75 were expressed in bacterial cells and purified by nickel affinity and cation exchange chromatography, and used to establish an AlphaScreen assay for compound screening. A total of twelve compounds were identified as possible HIV-1 IN-LEDGF/p75 inhibitors. Biochemical screening using an AlphaScreen assay was used and a total of six of the twelve inhibitors displaying inhibition above 50 % were identified. The best compound with an IC50 of 1.97 μM was identified as lovastatin. Lovastatin was found to have an EC50 of 6.54 μM in the antiviral assay but activity was likely attributed to cytotoxicity with a CC50 of 5.31 μM. Based on information from structurally similar statins, we hypothesised that a closed lactone ring in combination with a methyl group on the naphthalene structure elicited the likely inhibitory profile and that the lactone ring induced a large part of the cytotoxic effect of the compounds screened. Overall, we identified statins as potential inhibitors of the HIV-1 IN-LEDGF/p75 interaction, however the overlap of toxicity with antiviral effects renders statins in their current form unsuitable for antiretroviral treatment.