Elsevier

Bioorganic & Medicinal Chemistry

Volume 18, Issue 21, 1 November 2010, Pages 7515-7521
Bioorganic & Medicinal Chemistry

Small molecules targeting the interaction between HIV-1 integrase and LEDGF/p75 cofactor

https://doi.org/10.1016/j.bmc.2010.08.051Get rights and content

Abstract

The search of small molecules as protein–protein interaction inhibitors represents a new attractive strategy to develop anti-HIV-1 agents. We previously reported a computational study that led to the discovery of new inhibitors of the interaction between enzyme HIV-1 integrase (IN) and the nuclear protein lens epithelium growth factor LEDGF/p75.1

Herein, we describe new findings about the binding site of LEDGF/p75 on IN employing a different computational approach. In this way further structural requirements, helpful to disrupt LEDGF/p75-IN binding, have been identified. The main result of this work was the exploration of a relevant hydrophobic region. So we planned the introduction of suitable and simple chemical modifications on our previously reported ‘hit’ and the new synthesized compounds were subjected to biological tests.

The results obtained demonstrate that the hydrophobic pocket could play a key role in improving inhibitory efficacy thus opening new suggestions to design active ligands.

Graphical abstract

GRID approach was used to decipher structural requirements helpful to design and synthesize new inhibitors able to disrupt LEDGF/p75-IN binding.

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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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