The mechanism of H171T resistance reveals the importance of Nδ-protonated His171 for the binding of allosteric inhibitor BI-D to HIV-1 integrase

Slaughter, Alison; Jurado, Kellie Ann; Deng, Nanjie; Feng, Lei; Kessl, Jacques J.; Shkriabai, Nikoloz; Larue, Ross C.; Fadel, Hind J.; Patel, Pratiq A.; Jena, Nivedita; Fuchs, James R.; Poeschla, Eric M.; Levy, Ronald M.; Engelman, Alan; Kvaratskhelia, Mamuka

doi:10.1186/s12977-014-0100-1

Cited by 40 publications

(54 citation statements)

References 69 publications

(125 reference statements)

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“…Previous studies on point mutants at amino acid residues critical for INLAI binding to IN, that dramatically decrease IN affinity for INLAIs, showed that both ARV activities were either lost concomitantly in the T174I mutant or both strongly affected, although at variable level in the case of the H171T mutant (12,35). These mutants, the T174I in particular, demonstrate that the two activities of INLAIs are linked to their binding to the LEDGF-binding pocket.…”

Section: Discussionmentioning

confidence: 82%

Structure-function analyses unravel distinct effects of allosteric inhibitors of HIV-1 integrase on viral maturation and integration

Bonnard

Rouzic

Eiler

et al. 2018

Journal of Biological Chemistry

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Recently, a new class of HIV-1 integrase (IN) inhibitors with a dual mode of action, called IN-LEDGF/p75 allosteric inhibitors (INLAIs), was described. Designed to interfere with the IN-LEDGF/p75 interaction during viral integration, unexpectedly, their major impact was on virus maturation. This activity has been linked to induction of aberrant IN multimerization, while inhibition of the IN-LEDGF/p75 interaction accounts for weaker antiretroviral effect at integration. Since these dual activities result from INLAI binding to IN at a single binding site, we expected that these activities co-evolved together, driven by the affinity for IN. Using an original INLAI, MUT-A, and its activity on an Ala-125 (A125) IN variant, we found that these two activities on A125 IN can be fully dissociated: MUT-A-induced IN multimerization and the formation of eccentric condensates in viral particles, that are responsible for inhibition of virus maturation, were lost, while inhibition of the IN-LEDGF/p75 interaction and consequently integration, was fully retained. Hence the mere binding of INLAI to A125 IN is insufficient to promote the conformational changes of IN required for aberrant multimerization. By analyzing the Xray structures of MUT-A bound to the IN catalytic core domain (CCD) with or without the A125 polymorphism, we discovered that the loss of IN multimerization is due to stabilization of the A125 IN variant CCD dimer, highlighting the importance of the CCD dimerization energy for IN multimerization. Our study reveals that affinity for the LEDGF/p75-binding pocket is not sufficient to induce INLAI-dependent IN multimerization and the associated inhibition of viral maturation.The integrase (IN) protein of Human Immunodeficiency Virus type 1 (HIV-1) catalyzes the stable insertion of the viral cDNA genome into the host cell chromatin, a step of the viral life cycle that is required for efficient viral gene expression. Integration occurs via a two-step reaction where IN initially cleaves after a conserved CA dinucleotide at the 3' end of the viral cDNA genome to free a 3'-OH group (3' processing), which is next used to carry out a nucleophilic attack on cellular chromosomal DNA (strand transfer).IN is one of the preferred targets for the development of antiretroviral (ARV) drugs. However, given the high genetic variability of HIV-1, IN mutations conferring cross-resistance to the first generation INSTIs, RAL and EVG, were described in patients receiving INSTI-containing regimens (2). The second generation INSTI DTG has a higher genetic barrier and conserves good ARV activity against a number of RAL-and EVGresistant strains. Recent reports showed that Bictegravir, a second generation INSTI still in development from Gilead Sciences, has a resistance profile similar to DTG (3). Nevertheless, DTG and Bictegravir are sensitive to the most detrimental INSTI resistant mutations albeit at lower levels than first generation INSTIs (4). Therefore, the development of small molecule inhibitors impairing IN functions with dis...

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Section: Discussionmentioning

confidence: 82%

Structure-function analyses unravel distinct effects of allosteric inhibitors of HIV-1 integrase on viral maturation and integration

Bonnard

Rouzic

Eiler

et al. 2018

Journal of Biological Chemistry

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“…These findings have suggested that LEDGF/p75 is able to effectively compete with ALLINIs for binding at the IN CCD dimer interface during the initial steps of virus replication. In contrast, in producer cells, neither depletion nor overexpression of LEDGF/p75 influenced the potency of ALLINIs on subsequent virus infectivity (Jurado et al 2013; Balakrishnan et al 2013; Slaughter et al 2014; Fadel et al 2014). These results indicate that the lack of competition between the inhibitor and LEDGF/p75 during virus assembly enables ALLINIs to potently induce aberrant IN multimerization and impair correct core morphogenesis.…”

Section: Small Molecules Promote Aberrant Higher Order Hiv-1 Integmentioning

confidence: 83%

“…Experiments with LEDGF/p75 knockdown and knockout cells helped to explain these observations (Jurado et al 2013; Wang et al 2012; Schrijvers et al 2012; Balakrishnan et al 2013; Fadel et al 2014; Slaughter et al 2014). ALLINI BI-D potency during the early stages of viral replication increased ~29-fold and ~17-fold in LEDGF/p75 knockdown and knockout cells, respectively (Jurado et al 2013; Slaughter et al 2014). These findings have suggested that LEDGF/p75 is able to effectively compete with ALLINIs for binding at the IN CCD dimer interface during the initial steps of virus replication.…”

Section: Small Molecules Promote Aberrant Higher Order Hiv-1 Integmentioning

confidence: 97%

“…The dynamic light scattering (DLS) experiments have been instrumental to monitor IN multimerization in the presence of ALLINIs and allowed for the proposal of the mechanism depicted in Fig. 3e (Sharma et al 2014; Slaughter et al 2014). In the absence of the inhibitor, individual IN subunits are in dynamic equilibrium between monomers, dimers, and tetramers.…”

Section: The Complexity Of In Subunit–subunit Interactionsmentioning

confidence: 99%

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HIV-1 Integrase Multimerization as a Therapeutic Target

Feng

Larue

Slaughter

et al. 2015

Current Topics in Microbiology and Immunology

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Multimeric HIV-1 integrase (IN) plays an essential, multifunctional role in virus replication and serves as an important therapeutic target. Structural and biochemical studies have revealed the importance of the ordered interplay between IN molecules for its function. In the presence of viral DNA ends, individual IN subunits assemble into a tetramer and form a stable synaptic complex (SSC), which mediates integration of the reverse transcribed HIV-1 genome into chromatin. Cellular chromatin-associated protein LEDGF/p75 engages the IN tetramer in the SSC and directs HIV-1 integration into active genes. A mechanism to deregulate the productive interplay between IN subunits with small molecule inhibitors has recently received considerable attention. Most notably, allosteric IN inhibitors (ALLINIs) have been shown to bind to the IN dimer interface at the LEDGF/p75 binding pocket, stabilize interacting IN subunits, and promote aberrant, higher order IN multimerization. Consequently, these compounds impair formation of the SSC and associated LEDGF/p75-independent IN catalytic activities as well as inhibit LEDGF/p75 binding to the SSC in vitro. However, in infected cells, ALLINIs more potently impaired correct maturation of virus particles than the integration step. ALLINI treatments induced aberrant, higher order IN multimerization in virions and resulted in eccentric, non-infectious virus particles. These studies have suggested that the correctly ordered IN structure is important for virus particle morphogenesis and highlighted IN multimerization as a plausible therapeutic target for developing new inhibitors to enhance treatment options for HIV-1-infected patients.

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“…The resistance mutation IN A128T is responsible for an 8.65-to 11-fold change of the EC50 of CX05045 (4) and CX014442 (5), respectively while A128N and L102F induce a 64-and 60-fold change of the EC50 of BI224436 (9). Other single resistance mutations identified are the Y99H, Y99N A129T and H171T 78,86 . Although LEDGINs block early replication at the integration step, no resistance to INSTIs was observed with LEDGIN-resistant viruses.…”

Section: Mechanism Of Inhibitors Targeting the Ledgf/p75mentioning

confidence: 99%

Targeting Virus-host Interactions of HIV Replication

Weydert¹,

Rijck²,

Christ³

et al. 2015

CTMC

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Cellular proteins that are hijacked by HIV in order to complete its replication cycle, form attractive new targets for antiretroviral therapy. In particular, the protein-protein interactions between these cellular proteins (cofactors) and viral proteins are of great interest to develop new therapies. Research efforts have led to the validation of different cofactors and some successes in therapeutic applications. Maraviroc, the first cofactor inhibitor approved for human medicinal use, provided a proof of concept. Furthermore, compounds developed as Integrase-LEDGF/p75 interaction inhibitors (LEDGINs) have advanced to early clinical trials. Other compounds targeting cofactors and cofactor-viral protein interactions are currently under development. Likewise, interactions between cellular restriction factors and their counteracting HIV protein might serve as interesting targets in order to impair HIV replication. In this respect, compounds targeting the Vif-APOBEC3G interaction have been described. In this review, we focus on compounds targeting the Integrase-LEDGF/p75 interaction, the Tat-P-TEFb interaction and the Vif-APOBEC3G interaction. Additionally we give an overview of currently discovered compounds presumably targeting cellular cofactor-HIV protein interactions.

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The mechanism of H171T resistance reveals the importance of Nδ-protonated His171 for the binding of allosteric inhibitor BI-D to HIV-1 integrase

Cited by 40 publications

References 69 publications

Structure-function analyses unravel distinct effects of allosteric inhibitors of HIV-1 integrase on viral maturation and integration

Structure-function analyses unravel distinct effects of allosteric inhibitors of HIV-1 integrase on viral maturation and integration

HIV-1 Integrase Multimerization as a Therapeutic Target

Targeting Virus-host Interactions of HIV Replication

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