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Targeting the HIV-1 and HBV Capsids, an EnCore

by

William M. McFadden

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Stefan G. Sarafianos

Viruses 2023, 15(4), 896; https://doi.org/10.3390/v15040896 - 31 Mar 2023

Viewed by 722

Abstract

Not many structures are common among all viruses: only nucleic acid and a protein coat [...] Full article

(This article belongs to the Special Issue Capsid-Targeting Antivirals and Host Factors)

Research

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Open AccessArticle

Molecular Dynamics Free Energy Simulations Reveal the Mechanism for the Antiviral Resistance of the M66I HIV-1 Capsid Mutation

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Viruses 2021, 13(5), 920; https://doi.org/10.3390/v13050920 - 15 May 2021

Cited by 8 | Viewed by 2134

Abstract

While drug resistance mutations can often be attributed to the loss of direct or solvent-mediated protein−ligand interactions in the drug-mutant complex, in this study we show that a resistance mutation for the picomolar HIV-1 capsid (CA)-targeting antiviral (GS-6207) is mainly due to the [...] Read more.

While drug resistance mutations can often be attributed to the loss of direct or solvent-mediated protein−ligand interactions in the drug-mutant complex, in this study we show that a resistance mutation for the picomolar HIV-1 capsid (CA)-targeting antiviral (GS-6207) is mainly due to the free energy cost of the drug-induced protein side chain reorganization in the mutant protein. Among several mutations, M66I causes the most suppression of the GS-6207 antiviral activity (up to ~84,000-fold), and only 83- and 68-fold reductions for PF74 and ZW-1261, respectively. To understand the molecular basis of this drug resistance, we conducted molecular dynamics free energy simulations to study the structures, energetics, and conformational free energy landscapes involved in the inhibitors binding at the interface of two CA monomers. To minimize the protein−ligand steric clash, the I66 side chain in the M66I−GS-6207 complex switches to a higher free energy conformation from the one adopted in the apo M66I. In contrast, the binding of GS-6207 to the wild-type CA does not lead to any significant M66 conformational change. Based on an analysis that decomposes the absolute binding free energy into contributions from two receptor conformational states, it appears that it is the free energy cost of side chain reorganization rather than the reduced protein−ligand interaction that is largely responsible for the drug resistance against GS-6207. Full article

(This article belongs to the Special Issue Capsid-Targeting Antivirals and Host Factors)

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Open AccessArticle

Discovery of New Small Molecule Hits as Hepatitis B Virus Capsid Assembly Modulators: Structure and Pharmacophore-Based Approaches

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Viruses 2021, 13(5), 770; https://doi.org/10.3390/v13050770 - 27 Apr 2021

Cited by 12 | Viewed by 1877

Abstract

Hepatitis B virus (HBV) capsid assembly modulators (CpAMs) have shown promise as potent anti-HBV agents in both preclinical and clinical studies. Herein, we report our efforts in identifying novel CpAM hits via a structure-based virtual screening against a small molecule protein-protein interaction (PPI) [...] Read more.

Hepatitis B virus (HBV) capsid assembly modulators (CpAMs) have shown promise as potent anti-HBV agents in both preclinical and clinical studies. Herein, we report our efforts in identifying novel CpAM hits via a structure-based virtual screening against a small molecule protein-protein interaction (PPI) library, and pharmacophore-guided compound design and synthesis. Curated compounds were first assessed in a thermal shift assay (TSA), and the TSA hits were further evaluated in an antiviral assay. These efforts led to the discovery of two structurally distinct scaffolds, ZW-1841 and ZW-1847, as novel HBV CpAM hits, both inhibiting HBV in single-digit µM concentrations without cytotoxicity at 100 µM. In ADME assays, both hits displayed extraordinary plasma and microsomal stability. Molecular modeling suggests that these hits bind to the Cp dimer interfaces in a mode well aligned with known CpAMs. Full article

(This article belongs to the Special Issue Capsid-Targeting Antivirals and Host Factors)

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Open AccessArticle

All-Atom MD Simulations of the HBV Capsid Complexed with AT130 Reveal Secondary and Tertiary Structural Changes and Mechanisms of Allostery

by

Carolina Pérez-Segura

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Boon Chong Goh

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Jodi A. Hadden-Perilla

Viruses 2021, 13(4), 564; https://doi.org/10.3390/v13040564 - 26 Mar 2021

Cited by 9 | Viewed by 2156

Abstract

The hepatitis B virus (HBV) capsid is an attractive drug target, relevant to combating viral hepatitis as a major public health concern. Among small molecules known to interfere with capsid assembly, the phenylpropenamides, including AT130, represent an important antiviral paradigm based on disrupting [...] Read more.

The hepatitis B virus (HBV) capsid is an attractive drug target, relevant to combating viral hepatitis as a major public health concern. Among small molecules known to interfere with capsid assembly, the phenylpropenamides, including AT130, represent an important antiviral paradigm based on disrupting the timing of genome packaging. Here, all-atom molecular dynamics simulations of an intact AT130-bound HBV capsid reveal that the compound increases spike flexibility and improves recovery of helical secondary structure in the spike tips. Regions of the capsid-incorporated dimer that undergo correlated motion correspond to established sub-domains that pivot around the central chassis. AT130 alters patterns of correlated motion and other essential dynamics. A new conformational state of the dimer is identified, which can lead to dramatic opening of the intradimer interface and disruption of communication within the spike tip. A novel salt bridge is also discovered, which can mediate contact between the spike tip and fulcrum even in closed conformations, revealing a mechanism of direct communication across these sub-domains. Altogether, results describe a dynamical connection between the intra- and interdimer interfaces and enable mapping of allostery traversing the entire core protein dimer. Full article

(This article belongs to the Special Issue Capsid-Targeting Antivirals and Host Factors)

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Open AccessArticle

Design, Synthesis and Characterization of HIV-1 CA-Targeting Small Molecules: Conformational Restriction of PF74

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Rajkumar Lalji Sahani

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Raquel Diana-Rivero

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Sanjeev Kumar V. Vernekar

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Lei Wang

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Haijuan Du

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Huanchun Zhang

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Andres Emanuelli Castaner

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Viruses 2021, 13(3), 479; https://doi.org/10.3390/v13030479 - 15 Mar 2021

Cited by 9 | Viewed by 2185

Abstract

Small molecules targeting the PF74 binding site of the HIV-1 capsid protein (CA) confer potent and mechanistically unique antiviral activities. Structural modifications of PF74 could further the understanding of ligand binding modes, diversify ligand chemical classes, and allow identification of new variants with [...] Read more.

Small molecules targeting the PF74 binding site of the HIV-1 capsid protein (CA) confer potent and mechanistically unique antiviral activities. Structural modifications of PF74 could further the understanding of ligand binding modes, diversify ligand chemical classes, and allow identification of new variants with balanced antiviral activity and metabolic stability. In the current work, we designed and synthesized three series of PF74-like analogs featuring conformational constraints at the aniline terminus or the phenylalanine carboxamide moiety, and characterized them using a biophysical thermal shift assay (TSA), cell-based antiviral and cytotoxicity assays, and in vitro metabolic stability assays in human and mouse liver microsomes. These studies showed that the two series with the phenylalanine carboxamide moiety replaced by a pyridine or imidazole ring can provide viable hits. Subsequent SAR identified an improved analog 15 which effectively inhibited HIV-1 (EC₅₀ = 0.31 μM), strongly stabilized CA hexamer (ΔTm = 8.7 °C), and exhibited substantially enhanced metabolic stability (t_1/2 = 27 min for 15 vs. 0.7 min for PF74). Metabolic profiles from the microsomal stability assay also indicate that blocking the C5 position of the indole ring could lead to increased resistance to oxidative metabolism. Full article

(This article belongs to the Special Issue Capsid-Targeting Antivirals and Host Factors)

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Open AccessArticle

A New Generation of Functional Tagged Proteins for HIV Fluorescence Imaging

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Viruses 2021, 13(3), 386; https://doi.org/10.3390/v13030386 - 28 Feb 2021

Cited by 5 | Viewed by 1836

Abstract

During the last decade, there was a marked increase in the development of tools and techniques to study the molecular mechanisms of the HIV replication cycle by using fluorescence microscopy. Researchers often apply the fusion of tags and fluorophores to viral proteins, surrogate [...] Read more.

During the last decade, there was a marked increase in the development of tools and techniques to study the molecular mechanisms of the HIV replication cycle by using fluorescence microscopy. Researchers often apply the fusion of tags and fluorophores to viral proteins, surrogate proteins, or dyes to follow individual virus particles while they progress throughout infection. The inclusion of such fusion motifs or surrogates frequently disrupts viral infectivity or results in a change of the wild-type phenotype. Here, we detail the construction and functional characterization of two new constructs where we fused fluorescent proteins to the N-terminus of HIV-1 Integrase. In the first, IN is recruited into assembling particles via a codon optimized Gag to complement other viral constructs, while the second is fused to a Gag-Pol expression vector fully capable of integration. Our data shows that N-terminal tagged IN is functional for integration by both recovery of integration of catalytically inactive IN and by the successful infectivity of viruses carrying only labeled IN. These tools will be important to study the individual behavior of viral particles and associate such behavior to infectivity. Full article

(This article belongs to the Special Issue Capsid-Targeting Antivirals and Host Factors)

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Open AccessArticle

Studies on the Efficacy, Potential Cardiotoxicity and Monkey Pharmacokinetics of GLP-26 as a Potent Hepatitis B Virus Capsid Assembly Modulator

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Viruses 2021, 13(1), 114; https://doi.org/10.3390/v13010114 - 15 Jan 2021

Cited by 10 | Viewed by 1868

Abstract

While treatment options are available for hepatitis B virus (HBV), there is currently no cure. Anti-HBV nucleoside analogs and interferon-alpha 2b rarely clear HBV covalently closed circular DNA (cccDNA), requiring lifelong treatment. Recently, we identified GLP-26, a glyoxamide derivative which modulates HBV capsid [...] Read more.

While treatment options are available for hepatitis B virus (HBV), there is currently no cure. Anti-HBV nucleoside analogs and interferon-alpha 2b rarely clear HBV covalently closed circular DNA (cccDNA), requiring lifelong treatment. Recently, we identified GLP-26, a glyoxamide derivative which modulates HBV capsid assembly. The impact of GLP-26 on viral replication and integrated DNA was assessed in an HBV nude mouse model bearing HBV transfected AD38 xenografts. At day 45 post-infection, GLP-26 reduced HBV titers by 2.3–3 log₁₀ versus infected placebo-treated mice. Combination therapy with GLP-26 and entecavir reduced HBV log₁₀ titers by 4.6-fold versus placebo. Next, we examined the pharmacokinetics (PK) in cynomolgus monkeys administered GLP-26 via IV (1 mg/kg) or PO (5 mg/kg). GLP-26 was found to have 34% oral bioavailability, with a mean input time of 3.17 h. The oral dose produced a mean peak plasma concentration of 380.7 ng/mL, observed 0.67 h after administration (~30-fold > in vitro EC₉₀ corrected for protein binding), with a mean terminal elimination half-life of 2.4 h and a mean area under the plasma concentration versus time curve of 1660 ng·hr/mL. GLP-26 was 86.7% bound in monkey plasma. Lastly, GLP-26 demonstrated a favorable toxicity profile confirmed in primary human cardiomyocytes. Thus, GLP-26 warrants further preclinical development as an add on to treatment for HBV infection. Full article

(This article belongs to the Special Issue Capsid-Targeting Antivirals and Host Factors)

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Open AccessArticle

HIV-1 Uncoating and Nuclear Import Precede the Completion of Reverse Transcription in Cell Lines and in Primary Macrophages

by

Ashwanth C. Francis

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Mariana Marin

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Mathew J. Prellberg

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Kristina Palermino-Rowland

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Gregory B. Melikyan

Viruses 2020, 12(11), 1234; https://doi.org/10.3390/v12111234 - 30 Oct 2020

Cited by 26 | Viewed by 3258

Abstract

An assembly of capsid proteins (CA) form the mature viral core enclosing the HIV-1 ribonucleoprotein complex. Discrepant findings have been reported regarding the cellular sites and the extent of core disassembly (uncoating) in infected cells. Here, we combined single-virus imaging and time-of-drug-addition assays [...] Read more.

An assembly of capsid proteins (CA) form the mature viral core enclosing the HIV-1 ribonucleoprotein complex. Discrepant findings have been reported regarding the cellular sites and the extent of core disassembly (uncoating) in infected cells. Here, we combined single-virus imaging and time-of-drug-addition assays to elucidate the kinetic relationship between uncoating, reverse transcription, and nuclear import of HIV-1 complexes in cell lines and monocyte-derived macrophages (MDMs). By using cyclophilin A-DsRed (CDR) as a marker for CA, we show that, in contrast to TZM-bl cells, early cytoplasmic uncoating (loss of CDR) is limited in MDMs and is correlated with the efficiency of reverse transcription. However, we find that reverse transcription is dispensable for HIV-1 nuclear import, which progressed through an uncoating step at the nuclear pore. Comparison of the kinetics of nuclear import and the virus escape from inhibitors targeting distinct steps of infection, as well as direct quantification of viral DNA synthesis, revealed that reverse transcription is completed after nuclear import of HIV-1 complexes. Collectively, these results suggest that reverse transcription is dispensable for the uncoating step at the nuclear pore and that vDNA synthesis is completed in the nucleus of unrelated target cells. Full article

(This article belongs to the Special Issue Capsid-Targeting Antivirals and Host Factors)

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Review

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Open AccessReview

The Role of Capsid in the Early Steps of HIV-1 Infection: New Insights into the Core of the Matter

by

Nawal AlBurtamani

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Alwin Paul

and

Ariberto Fassati

Viruses 2021, 13(6), 1161; https://doi.org/10.3390/v13061161 - 17 Jun 2021

Cited by 6 | Viewed by 2582

Abstract

In recent years, major advances in research and experimental approaches have significantly increased our knowledge on the role of the HIV-1 capsid in the virus life cycle, from reverse transcription to integration and gene expression. This makes the capsid protein a good pharmacological [...] Read more.

In recent years, major advances in research and experimental approaches have significantly increased our knowledge on the role of the HIV-1 capsid in the virus life cycle, from reverse transcription to integration and gene expression. This makes the capsid protein a good pharmacological target to inhibit HIV-1 replication. This review covers our current understanding of the role of the viral capsid in the HIV-1 life cycle and its interaction with different host factors that enable reverse transcription, trafficking towards the nucleus, nuclear import and integration into host chromosomes. It also describes different promising small molecules, some of them in clinical trials, as potential targets for HIV-1 therapy. Full article

(This article belongs to the Special Issue Capsid-Targeting Antivirals and Host Factors)

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Open AccessReview

Addressing Antiretroviral Drug Resistance with Host-Targeting Drugs—First Steps towards Developing a Host-Targeting HIV-1 Assembly Inhibitor

by

Jaisri R. Lingappa

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Vishwanath R. Lingappa

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Jonathan C. Reed

Viruses 2021, 13(3), 451; https://doi.org/10.3390/v13030451 - 10 Mar 2021

Cited by 10 | Viewed by 1808

Abstract

The concerning increase in HIV-1 resistance argues for prioritizing the development of host-targeting antiviral drugs because such drugs can offer high genetic barriers to the selection of drug-resistant viral variants. Targeting host proteins could also yield drugs that act on viral life cycle [...] Read more.

The concerning increase in HIV-1 resistance argues for prioritizing the development of host-targeting antiviral drugs because such drugs can offer high genetic barriers to the selection of drug-resistant viral variants. Targeting host proteins could also yield drugs that act on viral life cycle events that have proven elusive to inhibition, such as intracellular events of HIV-1 immature capsid assembly. Here, we review small molecule inhibitors identified primarily through HIV-1 self-assembly screens and describe how all act either narrowly post-entry or broadly on early and late events of the HIV-1 life cycle. We propose that a different screening approach could identify compounds that specifically inhibit HIV-1 Gag assembly, as was observed when a potent rabies virus inhibitor was identified using a host-catalyzed rabies assembly screen. As an example of this possibility, we discuss an antiretroviral small molecule recently identified using a screen that recapitulates the host-catalyzed HIV-1 capsid assembly pathway. This chemotype potently blocks HIV-1 replication in T cells by specifically inhibiting immature HIV-1 capsid assembly but fails to select for resistant viral variants over 37 passages, suggesting a host protein target. Development of such small molecules could yield novel host-targeting antiretroviral drugs and provide insight into chronic diseases resulting from dysregulation of host machinery targeted by these drugs. Full article

(This article belongs to the Special Issue Capsid-Targeting Antivirals and Host Factors)

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Open AccessReview

Interactions of HIV-1 Capsid with Host Factors and Their Implications for Developing Novel Therapeutics

by

Shentian Zhuang

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Bruce E. Torbett

Viruses 2021, 13(3), 417; https://doi.org/10.3390/v13030417 - 05 Mar 2021

Cited by 14 | Viewed by 3335

Abstract

The Human Immunodeficiency Virus type 1 (HIV-1) virion contains a conical shell, termed capsid, encasing the viral RNA genome. After cellular entry of the virion, the capsid is released and ensures the protection and delivery of the HIV-1 genome to the host nucleus [...] Read more.

The Human Immunodeficiency Virus type 1 (HIV-1) virion contains a conical shell, termed capsid, encasing the viral RNA genome. After cellular entry of the virion, the capsid is released and ensures the protection and delivery of the HIV-1 genome to the host nucleus for integration. The capsid relies on many virus–host factor interactions which are regulated spatiotemporally throughout the course of infection. In this paper, we will review the current understanding of the highly dynamic HIV-1 capsid–host interplay during the early stages of viral replication, namely intracellular capsid trafficking after viral fusion, nuclear import, uncoating, and integration of the viral genome into host chromatin. Conventional anti-retroviral therapies primarily target HIV-1 enzymes. Insights of capsid structure have resulted in a first-in-class, long-acting capsid-targeting inhibitor, GS-6207 (Lenacapavir). This inhibitor binds at the interface between capsid protein subunits, a site known to bind host factors, interferes with capsid nuclear import, HIV particle assembly, and ordered assembly. Our review will highlight capsid structure, the host factors that interact with capsid, and high-throughput screening techniques, specifically genomic and proteomic approaches, that have been and can be used to identify host factors that interact with capsid. Better structural and mechanistic insights into the capsid–host factor interactions will significantly inform the understanding of HIV-1 pathogenesis and the development of capsid-centric antiretroviral therapeutics. Full article

(This article belongs to the Special Issue Capsid-Targeting Antivirals and Host Factors)

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Open AccessReview

Visualizing HIV-1 Capsid and Its Interactions with Antivirals and Host Factors

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Morganne Wilbourne

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Peijun Zhang

Viruses 2021, 13(2), 246; https://doi.org/10.3390/v13020246 - 04 Feb 2021

Cited by 8 | Viewed by 2485

Abstract

Understanding of the construction and function of the HIV capsid has advanced considerably in the last decade. This is due in large part to the development of more sophisticated structural techniques, particularly cryo-electron microscopy (cryoEM) and cryo-electron tomography (cryoET). The capsid is known [...] Read more.

Understanding of the construction and function of the HIV capsid has advanced considerably in the last decade. This is due in large part to the development of more sophisticated structural techniques, particularly cryo-electron microscopy (cryoEM) and cryo-electron tomography (cryoET). The capsid is known to be a pleomorphic fullerene cone comprised of capsid protein monomers arranged into 200–250 hexamers and 12 pentamers. The latter of these induce high curvature necessary to close the cone at both ends. CryoEM/cryoET, NMR, and X-ray crystallography have collectively described these interactions to atomic or near-atomic resolutions. Further, these techniques have helped to clarify the role the HIV capsid plays in several parts of the viral life cycle, from reverse transcription to nuclear entry and integration into the host chromosome. This includes visualizing the capsid bound to host factors. Multiple proteins have been shown to interact with the capsid. Cyclophilin A, nucleoporins, and CPSF6 promote viral infectivity, while MxB and Trim5α diminish the viral infectivity. Finally, structural insights into the intra- and intermolecular interactions that govern capsid function have enabled development of small molecules, peptides, and truncated proteins to disrupt or stabilize the capsid to inhibit HIV replication. The most promising of these, GS6207, is now in clinical trial. Full article

(This article belongs to the Special Issue Capsid-Targeting Antivirals and Host Factors)

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Open AccessReview

HIV Capsid and Integration Targeting

by

Alan N. Engelman

Viruses 2021, 13(1), 125; https://doi.org/10.3390/v13010125 - 18 Jan 2021

Cited by 20 | Viewed by 3286

Abstract

Integration of retroviral reverse transcripts into the chromosomes of the cells that they infect is required for efficient viral gene expression and the inheritance of viral genomes to daughter cells. Before integration can occur, retroviral reverse transcription complexes (RTCs) must access the nuclear [...] Read more.

Integration of retroviral reverse transcripts into the chromosomes of the cells that they infect is required for efficient viral gene expression and the inheritance of viral genomes to daughter cells. Before integration can occur, retroviral reverse transcription complexes (RTCs) must access the nuclear environment where the chromosomes reside. Retroviral integration is non-random, with different types of virus-host interactions impacting where in the host chromatin integration takes place. Lentiviruses such as HIV efficiently infect interphase cells because their RTCs have evolved to usurp cellular nuclear import transport mechanisms, and research over the past decade has revealed specific interactions between the HIV capsid protein and nucleoporin (Nup) proteins such as Nup358 and Nup153. The interaction of HIV capsid with cleavage and polyadenylation specificity factor 6 (CPSF6), which is a component of the cellular cleavage and polyadenylation complex, helps to dictate nuclear import as well as post-nuclear RTC invasion. In the absence of the capsid-CPSF6 interaction, RTCs are precluded from reaching nuclear speckles and gene-rich regions of chromatin known as speckle-associated domains, and instead mis-target lamina-associated domains out at the nuclear periphery. Highlighting this area of research, small molecules that inhibit capsid-host interactions important for integration site targeting are highly potent antiviral compounds. Full article

(This article belongs to the Special Issue Capsid-Targeting Antivirals and Host Factors)

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