23 research outputs found

    Dual-acting stapled peptides target both HIV-1 entry and assembly

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    Background: Previously, we reported the conversion of the 12-mer linear and cell-impermeable peptide CAI to a cell-penetrating peptide NYAD-1 by using an i,i + 4 hydrocarbon stapling technique and confirmed its binding to the C-terminal domain (CTD) of the HIV-1 capsid (CA) protein with an improved affinity (Kd ~ 1 μM) compared to CAI (Kd ~ 15 μM). NYAD-1 disrupts the formation of both immature- and mature-like virus particles in in vitro and cell-based assembly assays. In addition, it displays potent anti-HIV-1 activity in cell culture against a range of laboratory-adapted and primary HIV-1 isolates.<p></p> Results: In this report, we expanded the study to i,i + 7 hydrocarbon-stapled peptides to delineate their mechanism of action and antiviral activity. We identified three potent inhibitors, NYAD-36, -66 and -67, which showed strong binding to CA in NMR and isothermal titration calorimetry (ITC) studies and disrupted the formation of mature-like particles. They showed typical α-helical structures and penetrated cells; however, the cell penetration was not as efficient as observed with the i,i + 4 peptides. Unlike NYAD-1, the i,i + 7 peptides did not have any effect on virus release; however, they impaired Gag precursor processing. HIV-1 particles produced in the presence of these peptides displayed impaired infectivity. Consistent with an effect on virus entry, selection for viral resistance led to the emergence of two mutations in the gp120 subunit of the viral envelope (Env) glycoprotein, V120Q and A327P, located in the conserved region 1 (C1) and the base of the V3 loop, respectively.<p></p> Conclusion: The i,i + 7 stapled peptides derived from CAI unexpectedly target both CA and the V3 loop of gp120. This dual-targeted activity is dependent on their ability to penetrate cells as well as their net charge. This mechanistic revelation will be useful in further modifying these peptides as potent anti-HIV-1 agents.<p></p&gt

    Recent progress in antiretrovirals - lessons from resistance

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    Recent failures in efforts to develop an effective vaccine against HIV-1 infection have emphasized the importance of antiretroviral therapy in treating HIV-1-infected patients. Thus far, inhibitors of two viral enzymes, reverse transcriptase and protease, have had a profoundly positive impact on the survival of HIV-1-infected patients. However, new inhibitors that act at diverse steps in the viral replication cycle are urgently needed because of the development of resistance to currently available antiretrovirals. This review summarizes recent progress in antiretroviral drug discovery and development by specifically focusing on novel inhibitors of three phases of replication: viral entry, integration of the viral DNA into the host cell genome and virus particle maturation

    Human immunodeficiency virus type 1 assembly, release, and maturation

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    A detailed understanding of human immunodeficiency virus type 1 (HIV-1) assembly, release, and maturation is fundamental to our knowledge of the HIV-1 replication cycle and has the potential to inform the development of new antiretroviral strategies. The structural protein Gag plays a central role in these pathways and drives production of a mature infectious particle through protein–protein, protein–RNA, and protein–lipid interactions. These interactions facilitate multimerization of Gag to form the structural shell of the particle, encapsidation of the RNA genome, trafficking of the virion components to the site of assembly, acquisition of a lipid bilayer and associated envelope glycoproteins, hijacking host cell machinery to facilitate virus release, and proteolytic maturation of the nascent virion. In this review, we describe the significant progress that has been achieved in understanding these processes and highlight key areas that remain unclear. Finally, we discuss how this knowledge is being applied to develop new anti-HIV drugs, an important research priority due to rapid emergence of HIV-1 isolates resistant to currently approved antiretroviral drugs

    Hydrazone- and hydrazide-containing N-substituted glycines as peptoid surrogates for expedited library synthesis: applications to the preparation of Tsg101-directed HIV-1 budding antagonists

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    [Image: see text] Replacing the Pro6 in the p6(Gag)-derived 9-mer “P-E-P-T-A-P-P-E-E” with N-substituted glycine (NSG) residues is problematic. However, incorporation of hydrazone amides (“peptoid hydrazones”) can be readily achieved in library fashion. Furthormore, reduction of these hydrazones to N-substituted “peptoid hydrazides” affords a facile route to library diversification. This approach is demonstrated by application to Tsg101-binding compounds designed as potential HIV budding antagonists

    Antiviral activity of alpha-helical stapled peptides designed from the HIV-1 capsid dimerization domain, Retrovirology

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    Extent: 18p.Background: The C-terminal domain (CTD) of HIV-1 capsid (CA), like full-length CA, forms dimers in solution and CTD dimerization is a major driving force in Gag assembly and maturation. Mutations of the residues at the CTD dimer interface impair virus assembly and render the virus non-infectious. Therefore, the CTD represents a potential target for designing anti-HIV-1 drugs. Results: Due to the pivotal role of the dimer interface, we reasoned that peptides from the a-helical region of the dimer interface might be effective as decoys to prevent CTD dimer formation. However, these small peptides do not have any structure in solution and they do not penetrate cells. Therefore, we used the hydrocarbon stapling technique to stabilize the a-helical structure and confirmed by confocal microscopy that this modification also made these peptides cell-penetrating. We also confirmed by using isothermal titration calorimetry (ITC), sedimentation equilibrium and NMR that these peptides indeed disrupt dimer formation. In in vitro assembly assays, the peptides inhibited mature-like virus particle formation and specifically inhibited HIV-1 production in cell-based assays. These peptides also showed potent antiviral activity against a large panel of laboratory-adapted and primary isolates, including viral strains resistant to inhibitors of reverse transcriptase and protease. Conclusions: These preliminary data serve as the foundation for designing small, stable, a-helical peptides and small-molecule inhibitors targeted against the CTD dimer interface. The observation that relatively weak CA binders, such as NYAD-201 and NYAD-202, showed specificity and are able to disrupt the CTD dimer is encouraging for further exploration of a much broader class of antiviral compounds targeting CA. We cannot exclude the possibility that the CA-based peptides described here could elicit additional effects on virus replication not directly linked to their ability to bind CA-CTD.Hongtao Zhang, Francesca Curreli, Xihui Zhang, Shibani Bhattacharya, Abdul A Waheed, Alan Cooper, David Cowburn, Eric O Freed and Asim K Debnat

    Determinants of activity of the HIV-1 maturation inhibitor PA-457

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    Abstract3-O-(3′,3′-dimethylsuccinyl) betulinic acid, also termed PA-457 or DSB, is a novel HIV-1 inhibitor that blocks virus maturation by disrupting cleavage of the capsid precursor, CA-SP1. To better define the molecular target for PA-457, we prepared a panel of mutant viruses with point deletions spanning the CA-SP1 cleavage domain and characterized each of these viruses for PA-457 sensitivity. Our results indicate that amino acid residues in the N-terminal half of SP1 serve as determinants of PA-457 activity, while residues in the C-terminal half of SP1 were not involved in compound activity. These findings support and extend previous observations that PA-457 is a specific inhibitor of CA-SP1 cleavage and identify the CA-SP1 domain as the primary viral determinant for this novel inhibitor of HIV-1 replication
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