Small molecules targeting the interaction between HIV-1 integrase and LEDGF/p75 cofactor
Graphical abstract
GRID approach was used to decipher structural requirements helpful to design and synthesize new inhibitors able to disrupt LEDGF/p75-IN binding.
Introduction
To date, 25 compounds for the treatment of HIV infections have been formally licensed. They belong to different categories: reverse transcriptase inhibitors (RTIs), protease inhibitors (PIs); fusion inhibitors (FIs), coreceptor inhibitors (CRIs), and integrase inhibitors (INIs).2 However, even the combination of these drugs is not sufficient enough to stop the progression of AIDS partly due to the emergence of drug-resistant HIV-1 mutants as well as the severe side effects.3, 4, 5 Therefore, alternative strategies need to be considered and new targets need to be identified to combat the growing AIDS pandemic. Currently, small-molecule inhibitors of the direct protein–protein interactions (PPIs), that mediate important biological processes, are an emerging and interesting area in drug design. In particular, there is evidence that various proteins play auxiliary roles in the HIV-1 life cycle and that some host cellular cofactors specifically control the integration process of viral DNA. The disruption of these essential cofactor-IN protein–protein interactions could represent a new frontier to design and develop novel anti-HIV-1 agents for anti-AIDS therapy.6
Amongst these cofactors, the ubiquitous lens epithelium growth factor LEDGF/p75 seems to play a pivotal role in the transcriptional regulation, as well as in cell survival and in prevention of apoptosis. In particular, cells lacking of LEDGF/p75 showed a severe defect in HIV-1 infection characterized by decreased levels of integrated viral DNA.7, 8 This evidence pointed out the possibility that LEDGF/p75 could be a promising target to inhibit DNA integration of HIV-1 life cycle. The LEDGF/p75 acts through a tethering mechanism as a potent cofactor for HIV-1 integration with the N-terminal Pro-Trp-Trp-Pro (PWWP) and A/T-hook elements binding to chromatin, and a C-terminal integrase-binding domain (IBD) binding to IN.9, 10
It is well known that these two regions of LEDGF/p75 are crucial for cofactor activity.
The specific protein–protein interaction between HIV-1 IN catalytic core domain (CCD) and IBD has been well described by X-ray analysis (Protein Data Bank file 2B4J) as well as by site-directed mutation studies, and it is characterized by IBD residues Ile365, Asp366, Phe406, and Val408.11 In a previous paper, we focused our interest on key residues Ile365-Asp366 and used a computational approach to describe the pharmacophoric requirements needed to disrupt the interaction between HIV-1 IN CCD and LEDGF/p75 IBD.1 This study led to the identification of some small molecules mimicking the Ile-Asp dipeptide; the most active molecule was the 2-hydroxy-4-(4-hydroxy-1H-indol-3-yl)-4-oxobut-2-enoic acid (CHIBA-3003, I, Fig. 1) that inhibited the IN-LEDGF/p75 interaction showing IC50 value of 35 μM in AlphaScreen assay.
We therefore considered that CHIBA-3003 was a promising ‘hit compound’ for the development of new and more potent inhibitors preventing IN-LEDGF/75 interaction. Considering that very few inhibitors of this interaction are known and with the aim of achieving other information about the IN/LEDGF protein interaction and identify new important chemical frames improving the inhibitory potency of CHIBA-3003, in this study we have explored new binding pocket regions through GRID molecular interaction fields (MIFs) and synthesized several new small molecules targeting this PPI. Furthermore, docking experiments furnished new suggestions about the putative interactions within the binding site of these inhibitors, which hopefully could advance into novel anti-HIV drug targets.
Section snippets
Rational design
It is well known that a typical protein–protein interaction occurs between the IBD of LEDGF/p75 and the integrase catalytic core domain (INCCD). The IBD of LEDGF/p75 forms a specific interaction with the binding pocket at the interface of two monomers of the INCCD. The IBD residues, involved in the interaction, are located in its interhelical loop regions and 4 relevant amino acid residues, Ile365, Asp366, Phe406, and Val408, are considered essential for the direct contact with INCCD residues.11
Conclusion
In conclusion the importance of an additional lipophilic feature for IN-LEDGF/p75 inhibition has been suggested following the identification of a new hydrophobic region in IBD binding site. Novel small molecules active as PPIs have been designed, synthesized and tested. The obtained information could be helpful for the discovery of new inhibitors of this emerging and promising PPI target particularly important for anti-AIDS drug design.
Molecular modeling
Both the GRID analysis and docking were performed using IN structure retrieved by X-ray crystallographic of the dimeric catalytic core domain of HIV-1 IN complexed with LEDGF/p75 IBD deposited in the RCSB Protein Data Bank (entry code 2 B4J).11
GRID calculations
The calculations were performed with version 22 of the GRID software.14 The GRID box dimensions were chosen to encompass all residues able to interact with IBD; and the NPLA (Number of Planes per Å for the grid box) was set to 2. The probes applied were
Acknowledgment
This work was supported by the European Commission (HEALTH-F3-2008-201032) (THINC project).
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