Impact of the Enfuvirtide Resistance Mutation N43D and the Associated Baseline Polymorphism E137K on Peptide Sensitivity and Six-Helix Bundle Structure

Xuefang, Bai; Wilson, Karen L.; Seedorff, Jennifer E.; Ahrens, Douglas; Green, Justin; Davison, Donna K.; Jin, Lei; Stanfield-Oakley, Sherry; Mosier, Sarah; Melby, Thomas; Cammack, Nick; Wang, Zhongmin; Greenberg, Michael; Dwyer, John J.

doi:10.1021/bi702509d

Cited by 30 publications

(32 citation statements)

References 35 publications

(85 reference statements)

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“…In this scenario, the drug-target affinity per se is unaltered, but the opportunity for the drug to access the target site is restricted, for example, because the target site is exposed for a shorter time. This mechanism has been described for HIV-1 fusion inhibitors (48,60). We think that such a window mechanism can also account for the VIR165 resistance mutations.…”

Section: Discussionmentioning

confidence: 52%

HIV-1 Escape from a Peptidic Anchor Inhibitor through Stabilization of the Envelope Glycoprotein Spike

Eggink

Taeye

Bontjer

et al. 2016

J Virol

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The trimeric HIV-1 envelope glycoprotein spike (Env) mediates viral entry into cells by using a spring-loaded mechanism that allows for the controlled insertion of the Env fusion peptide into the target membrane, followed by membrane fusion. Env is the focus of vaccine research aimed at inducing protective immunity by antibodies as well as efforts to develop drugs that inhibit the viral entry process. The molecular factors contributing to Env stability and decay need to be understood better in order to optimally design vaccines and therapeutics. We generated viruses with resistance to VIR165, a peptidic inhibitor that binds the fusion peptide of the gp41 subunit and prevents its insertion into the target membrane. Interestingly, a number of escape viruses acquired substitutions in the C1 domain of the gp120 subunit (A60E, E64K, and H66R) that rendered these viruses dependent on the inhibitor. These viruses could infect target cells only when VIR165 was present after CD4 binding. Furthermore, the VIR165-dependent viruses were resistant to soluble CD4-induced Env destabilization and decay. These data suggest that VIR165-dependent Env proteins are kinetically trapped in the unliganded state and require the drug to negotiate CD4-induced conformational changes. These studies provide mechanistic insight into the action of the gp41 fusion peptide and its inhibitors and provide new ways to stabilize Env trimer vaccines. IMPORTANCEBecause of the rapid development of HIV-1 drug resistance, new drug targets need to be explored continuously. The fusion peptide of the envelope glycoprotein can be targeted by anchor inhibitors. Here we describe virus escape from the anchor inhibitor VIR165. Interestingly, some escape viruses became dependent on the inhibitor for cell entry. We show that the identified escape mutations stabilize the ground state of the envelope glycoprotein and should thus be useful in the design of stabilized envelopebased HIV vaccines. With over 35 million people currently infected, HIV-1 remains a major health problem. Although progress has been made in the development of antiviral drugs, the potential of the virus to acquire resistance remains an issue. Neither a cure nor an effective vaccine is available. HIV-1 enters target cells by using its envelope glycoprotein (Env) spikes on the virus surface. Env is the target for drugs that inhibit the viral entry process and for broadly neutralizing antibodies (bNAbs) that researchers aim to induce with Env-based vaccines.Env is a trimeric complex consisting of three gp41 transmembrane subunits and three noncovalently attached gp120 subunits. The entry process is initiated by binding of gp120 to the CD4 receptor. This interaction induces conformational changes in Env, revealing the binding site for a chemokine coreceptor, generally CXCR4 or CCR5. Additional conformational changes in gp41 involve two heptad repeat regions (HR1 and HR2) in gp41 and the fusion peptide (FP) (1). The FP might be partially exposed in the prefusion, native state of Env. In a com...

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Section: Discussionmentioning

confidence: 52%

HIV-1 Escape from a Peptidic Anchor Inhibitor through Stabilization of the Envelope Glycoprotein Spike

Eggink

Taeye

Bontjer

et al. 2016

J Virol

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“…Moreover, results showed that N28Fd was much more resistant to proteinase digestion than T20, prolonging the existence of N28Fd in the human circulatory system. Finally, N36Fd and N28Fd potently inhibit a series of T20-resistant strains, although we cannot exclude the possibility that these NHR trimers may loose antiviral activity against the HIV-1 variants with compensatory mutations in the CHR region (42)(43)(44). Because N28Fd and T20 target different regions of gp41, combining them in clinical practice should help to avoid generating viruses otherwise resistant to either one alone.…”

Section: Discussionmentioning

confidence: 98%

Novel Recombinant Engineered gp41 N-terminal Heptad Repeat Trimers and Their Potential as Anti-HIV-1 Therapeutics or Microbicides

Chen

et al. 2010

Journal of Biological Chemistry

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Peptides derived from N-terminal heptad repeat (NHR) of the HIV-1 gp41 are generally poor inhibitors of HIV-1 entry, because they tend to aggregate and do not form a trimeric coiled-coil. In this study, we have fused portions of gp41 NHR, e.g. N36 or N28, to the T4 fibritin trimerization domain, Foldon (Fd), thus constructing novel NHR trimers, designated N36Fd or N28Fd, which could be expressed in Escherichia coli cells. The purified N36Fd and N28Fd exhibited SDS-resistant trimeric coiled-coil conformation with improved ␣-helicity compared with the corresponding N-peptides. They could interact with a C-peptide (e.g. C34) to form stable six-helix bundle and possessed potent anti-HIV-1 activity against a broad spectrum of HIV-1 strains. N28Fd was effective against T20-resistant HIV-1 variants and more resistant to proteinase K compared with T20 (enfuvirtide), a C-peptide-based HIV fusion inhibitor. Therefore, N28Fd trimer has great potentials for further development as an affordable therapeutic or microbicide for treatment and prevention of HIV-1 infection.

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“…When introduced by themselves, such HR-2 mutations typically have either no impact or only a modest impact on ENF sensitivity (27,36). In some instances, HR-2 mutations have been shown to enhance the stability of the six-helix bundle in either structural-modeling or peptide binding studies (1,14). By using a variety of in vitro assays, we were able to probe the functional consequences of HR-2 mutations either in isolation or in conjunction with their corresponding HR-1 mutations that evolved in ENF-treated patients.…”

Section: Discussionmentioning

confidence: 99%

HR-2 Mutations in Human Immunodeficiency Virus Type 1 gp41 Restore Fusion Kinetics Delayed by HR-1 Mutations That Cause Clinical Resistance to Enfuvirtide

Ray

Blackburn

Doms

2009

J Virol

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Enfuvirtide (ENF) prevents the entry of human immunodeficiency virus type 1 (HIV-1) into cells by binding to the HR-1 region of the viral envelope (Env) protein gp41 subunit. Resistance to ENF arises via mutations in the drug binding site in HR-1. In addition, HR-2 mutations are commonly observed in ENF-resistant Env proteins, though their role remains unclear. We explored the mechanistic basis for clinical resistance to ENF and the role of HR-2 mutations. Using panels of ENF resistance-associated mutants for two patients, we found that mutations in HR-1 slowed the fusion kinetics and that mutations in HR-2 restored fusion rates. We assessed the differences in the rates of fusion of these mutants from a temperature-arrested state and observed similar trends, suggesting that the step of delay occurs after coreceptor engagement. Sensitivity to neutralizing antibodies was unchanged by the HR-1 and HR-2 mutants in each panel. Since this result was in contrast to those of a previous in vitro analysis where enhanced sensitivity to neutralization was demonstrated for heterologous Envs with ENF resistance-associated HR-1 changes, we examined the context dependence of HR-1 and HR-2 mutations by transferring the mutations seen in one patient into the Env context of another. These studies revealed that some, but not all, HR-1 mutations, when placed out of context (i.e., in a patient Env where they did not originally arise), enhance sensitivity to neutralizing antibodies. However, in most cases, HR-1 mutations in ENF-treated patients evolve in a manner that preserves pretreatment neutralization sensitivity so as to evade the pressures of the immune system.

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Impact of the Enfuvirtide Resistance Mutation N43D and the Associated Baseline Polymorphism E137K on Peptide Sensitivity and Six-Helix Bundle Structure

Cited by 30 publications

References 35 publications

HIV-1 Escape from a Peptidic Anchor Inhibitor through Stabilization of the Envelope Glycoprotein Spike

HIV-1 Escape from a Peptidic Anchor Inhibitor through Stabilization of the Envelope Glycoprotein Spike

Novel Recombinant Engineered gp41 N-terminal Heptad Repeat Trimers and Their Potential as Anti-HIV-1 Therapeutics or Microbicides

HR-2 Mutations in Human Immunodeficiency Virus Type 1 gp41 Restore Fusion Kinetics Delayed by HR-1 Mutations That Cause Clinical Resistance to Enfuvirtide

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