Search Results (15)

Objective: The six-helix bundle (6-HB) is critical for HIV-1 membrane fusion. To disrupt this process, peptide inhibitors have been meticulously designed to target interactions within the 6-HB regions, thereby blocking membrane fusion and exerting inhibitory effects. Current peptide inhibitors like Enfuvirtide suffer from drug resistance and short in vivo half-life. This study aims to design novel anti-HIV-1 peptides by integrating heptad-repeat rules and membrane-anchor strategies. Methods: Artificial peptides were designed using HR rules from the HIV-1 gp41 6-HB motif and membrane-anchor modifications. Results: EK35S-Palm has emerged as a highly promising candidate for HIV-1 inhibition, exhibiting robust binding affinity to the target and effectively impeding the 6-HB spontaneous formation. Discussion: HR-based design avoids viral sequence homology, and membrane anchoring enhances local agent concentration, improving pharmacokinetics. The HR binding and membrane stabilization of EK35S-Palm provide synergistic inhibition. Conclusions: Integrating HR structural design with membrane-anchor strategies yields potent HIV-1 fusion inhibitors. EK35S-Palm demonstrates superior efficacy and stability over current therapies. These approaches hold great potential for overcoming the current therapy limitations and advancing the more effective and durable HIV-1 fusion inhibitors. Full article

Human immunodeficiency virus type 1 (HIV-1) is characterized by high variability and drug resistance. This has necessitated the development of antivirals with a new chemotype and therapy. We previously identified an artificial peptide with non-native protein sequence, AP3, with the potential to inhibit HIV-1 fusion through targeting hydrophobic grooves on the N-terminal heptad repeat trimer of viral glycoprotein gp41. Here, a small-molecule HIV-1 inhibitor targeting chemokine coreceptor CCR5 on the host cell was integrated into the AP3 peptide, producing a novel dual-target inhibitor with improved activity against multiple HIV-1 strains including those resistant to the currently used anti-HIV-1 drug enfuvirtide. Its superior antiviral potency in comparison with the respective pharmacophoric moieties is in consonance with the dual binding of viral gp41 and host factor CCR5. Therefore, our work provides a potent artificial peptide-based bifunctional HIV-1 entry inhibitor and highlights the multitarget-directed ligands approach in the development of novel therapeutic anti-HIV-1 agents. Full article

Currently, it is estimated that 1–2 million people worldwide are infected with HIV-2, accounting for 3–5% of the global burden of HIV. The course of HIV-2 infection is longer compared to HIV-1 infection, but without effective antiretroviral therapy (ART), a substantial proportion of infected patients will progress to AIDS and die. Antiretroviral drugs in clinical use were designed for HIV-1 and, unfortunately, some do not work as well, or do not work at all, for HIV-2. This is the case for non-nucleoside reverse transcriptase inhibitors (NNRTIs), the fusion inhibitor enfuvirtide (T-20), most protease inhibitors (PIs), the attachment inhibitor fostemsavir and most broadly neutralizing antibodies. Integrase inhibitors work well against HIV-2 and are included in first-line therapeutic regimens for HIV-2-infected patients. However, rapid emergence of drug resistance and cross-resistance within each drug class dramatically reduces second-line treatment options. New drugs are needed to treat infection with drug-resistant isolates. Here, we review the therapeutic armamentarium available to treat HIV-2-infected patients, as well as promising drugs in development. We also review HIV-2 drug resistance mutations and resistance pathways that develop in HIV-2-infected patients under treatment. Full article

In this study, the HIV-1 P18I10 CTL peptide derived from the V3 loop of HIV-1 gp120 and the T20 anti-fusion peptide of HIV-1 gp41 were inserted into the HPV16 L1 capsid protein to construct chimeric HPV:HIV (L1:P18I10 and L1:T20) VLPs by using the mammalian cell expression system. The HPV:HIV VLPs were purified by chromatography. We demonstrated that the insertion of P18I10 or T20 peptides into the DE loop of HPV16 L1 capsid proteins did not affect in vitro stability, self-assembly and morphology of chimeric HPV:HIV VLPs. Importantly, it did not interfere either with the HIV-1 antibody reactivity targeting sequential and conformational P18I10 and T20 peptides presented on chimeric HPV:HIV VLPs or with the induction of HPV16 L1-specific antibodies in vivo. We observed that chimeric L1:P18I10/L1:T20 VLPs vaccines could induce HPV16- but weak HIV-1-specific antibody responses and elicited HPV16- and HIV-1-specific T-cell responses in BALB/c mice. Moreover, could be a potential booster to increase HIV-specific cellular responses in the heterologous immunization after priming with rBCG.HIVA vaccine. This research work would contribute a step towards the development of the novel chimeric HPV:HIV VLP-based vaccine platform for controlling HPV16 and HIV-1 infection, which is urgently needed in developing and industrialized countries. Full article

Despite the enormous efforts made to develop other fusion inhibitors for HIV, the enfuvirtide (known as T20) peptide is the only approved HIV-1 inhibitory drug so far. Investigating the role of potential residues of the T20 peptide’s conformational dynamics could help us to understand the role of potential residues of the T20 peptide. We investigated T20 peptide conformation and binding interactions with the HIV-1 receptor (i.e., gp41) using MD simulations and docking techniques, respectively. Although the mutation of E143 into alanine decreased the flexibility of the E143A mutant, the conformational compactness of the mutant was increased. This suggests a potential role of E143 in the T20 peptide’s conformation. Interestingly, the free energy landscape showed a significant change in the wild-type T20 minimum, as the E143A mutant produced two observed minima. Finally, the docking results of T20 to the gp41 receptor showed a different binding interaction in comparison to the E143A mutant. This suggests that E143 residue can influence the binding interaction with the gp41 receptor. Overall, the E143 residue showed a significant role in conformation and binding to the HIV-1 receptor. These findings can be helpful in optimizing and developing HIV-1 inhibitor peptides. Full article

In our previous work, we replaced the TRM (tryptophan-rich motif) of T20 (Enfuvirtide) with fatty acid (C16) to obtain the novel lipopeptide LP-40, and LP-40 displayed enhanced antiviral activity. In this study, we investigated whether the C16 modification could enhance the high-resistance barrier of the inhibitor LP-40. To address this question, we performed an in vitro simultaneous screening of HIV-1_NL4-3 resistance to T20 and LP-40. The mechanism of drug resistance for HIV-1 Env was further studied using the expression and processing of the Env glycoprotein, the effect of the Env mutation on the entry and fusion ability of the virus, and an analysis of changes to the gp41 core structure. The results indicate that the LP-40 activity is enhanced and that it has a high resistance barrier. In a detailed analysis of the resistance sites, we found that mutations in L33S conferred a stronger resistance, except for the well-recognized mutations in amino acids 36–45 of gp41 NHR, which reduced the inhibitory activity of the CHR-derived peptides. The compensatory mutation of eight amino acids in the CHR region (NDQEEDYN) plays an important role in drug resistance. LP-40 and T20 have similar resistance mutation sites, and we speculate that the same resistance profile may arise if LP-40 is used in a clinical setting. Full article

Recently, a series of highly effective peptide- or protein-based HIV fusion inhibitors have been identified. However, due to their short half-life, their clinical application is limited. Therefore, the development of long-acting HIV fusion inhibitors is urgently needed. Here, we designed and constructed a protein-based, long-acting HIV fusion inhibitor, termed FLT (FN3-L35-T1144), consisting of a monobody, FN3, which contains an albumin-binding domain (ABD), a 35-mer linker (L35), and a peptide-based HIV fusion inhibitor, T1144. We found that FLT bound, via its FN3 component, with human serum albumin (HSA) in a reversible manner, thus maintaining the high efficiency of T1144 against infection by both HIV-1 IIIB (X4) and Bal (R5) strains with IC₅₀ of 11.6 nM and 15.3 nM, respectively, and remarkably prolonging the half-life of T1144 (~27 h in SD rats). This approach affords protein-based HIV fusion inhibitors with much longer half-life compared to enfuvirtide, a peptide-based HIV fusion inhibitor approved for use in clinics. Therefore, FLT is a promising candidate as a new protein-based anti-HIV drug with an improved pharmacokinetic profile. Full article

The clinical application of HIV fusion inhibitor, enfuvirtide (T20), was limited mainly because of its short half-life. Here we designed and synthesized two PEGylated C34 peptides, PEG2kC34 and PEG5kC34, with the PEG chain length of 2 and 5 kDa, respectively, and evaluated their anti-HIV-1 activity and mechanisms of action. We found that these two PEGylated peptides could bind to the HIV-1 peptide N36 to form high affinity complexes with high α-helicity. The peptides PEG2kC34 and PEG5kC34 effectively inhibited HIV-1 Env-mediated cell–cell fusion with an effective concentration for 50% inhibition (EC₅₀) of about 36 nM. They also inhibited infection of the laboratory-adapted HIV-1 strain NL4-3 with EC₅₀ of about 4–5 nM, and against 47 HIV-1 clinical isolates circulating in China with mean EC₅₀ of PEG2kC34 and PEG5kC34 of about 26 nM and 32 nM, respectively. The plasma half-life (t_1/2) of PEG2kC34 and PEG5kC34 was 2.6 h and 5.1 h, respectively, and the t_1/2 of PEGylated C34 was about 2.4-fold and 4.6-fold longer than C34 (~1.1 h), respectively. These findings suggest that PEGylated C34 with broad-spectrum anti-HIV-1 activity and prolonged half-life can be further developed as a peptide fusion inhibitor-based long-acting anti-HIV drug for clinical use to treat HIV-infected patients who have failed to respond to current anti-retrovirus drugs. Full article

Application of highly active antiretroviral drugs (ARDs) effectively reduces morbidity and mortality in HIV-infected individuals. However, the emergence of multiple drug-resistant strains has led to the increased failure of ARDs, thus calling for the development of anti-HIV drugs with targets or mechanisms of action different from those of the current ARDs. The first peptide-based HIV entry inhibitor, enfuvirtide, was approved by the U.S. FDA in 2003 for treatment of HIV/AIDS patients who have failed to respond to the current ARDs, which has stimulated the development of several series of protein- and peptide-based HIV entry inhibitors in preclinical and clinical studies. In this review, we highlighted the properties and mechanisms of action for those promising protein- and peptide-based HIV entry inhibitors targeting the HIV-1 gp120 or gp41 and discussed their advantages and disadvantages, compared with the current ARDs. Full article

Identifying drug resistance mutations is important for the clinical use of antivirals and can help define both a drug’s mechanism of action and the mechanistic basis of resistance. Resistance mutations are often identified one-at-a-time by studying viral evolution within treated patients or during viral growth in the presence of a drug in cell culture. Such approaches have previously mapped resistance to enfuvirtide, the only clinically approved HIV-1 fusion inhibitor, to enfuvirtide’s binding site in the N-terminal heptad repeat (NHR) of the Envelope (Env) transmembrane domain as well as a limited number of allosteric sites. Here, we sought to better delineate the genotypic determinants of resistance throughout Env. We used deep mutational scanning to quantify the effect of all single-amino-acid mutations to the subtype A BG505 Env on resistance to enfuvirtide. We identified both previously characterized and numerous novel resistance mutations in the NHR. Additional resistance mutations clustered in other regions of Env conformational intermediates, suggesting they may act during different fusion steps by altering fusion kinetics and/or exposure of the enfuvirtide binding site. This complete map of resistance sheds light on the diverse mechanisms of enfuvirtide resistance and highlights the utility of using deep mutational scanning to comprehensively map potential drug resistance mutations. Full article

Enfuvirtide (T20) is the first U.S. FDA-approved HIV fusion inhibitor-based anti-HIV drug. Its clinical application is limited because of its low potency and short half-life. We previously reported that peptide HP23-E6-IDL, containing both N- and C-terminal anchor-tails, exhibited stronger potency and a better resistance profile than T20. Here we designed an analogous peptide, YIK, by introducing a mutation, T639I, and then a lipopeptide, YIK-C16, by adding palmitic acid (C16) at the C-terminus of YIK. We found that YIK-C16 was 4.4- and 3.6-fold more potent than HP23-E6-IDL and YIK against HIV-1_IIIB infection and 13.3- and 10.5-fold more effective than HP23-E6-IDL and YIK against HIV-1_Bal infection, respectively. Consistently, the ex vivo anti-HIV-1_IIIB activity, as determined by the highest dilution-fold of the serum causing 50% inhibition of HIV-1 infection, of YIK-C16 in the sera of pretreated mice was remarkably higher than that of YIK or HP23-E6-IDL. The serum half-life (t_1/2 = 5.9 h) of YIK-C16 was also significantly longer than that of YIK (t_1/2 = 1.3 h) and HP23-E6-IDL (t_1/2 = 1.0 h). These results suggest that the lipopeptide YIK-C16 shows promise for further development as a new anti-HIV drug with improved anti-HIV-1 activity and a prolonged half-life. Full article

Background: Novel strategies to increase the efficacy of antiretroviral (ARV) drugs will be of crucial importance. We hypothesize that membranes of HIV-1-infected cells and enveloped HIV-1 particles may be preferentially targeted by the phytopeptide, cycloviolacin O2 (CyO2) to significantly enhance ARV efficacy. Methods: Physiologically safe concentrations of CyO2 were determined via red blood cell (RBC) hemolysis. SYTOX-green dye-uptake and radiolabeled saquinavir (³H-SQV) uptake assays were used to measure pore-formation and drug uptake, respectively. ELISA, reporter assays and ultracentrifugation were conducted to analyze the antiviral efficacy of HIV-1 protease and fusion inhibitors alone and co-exposed to CyO2. Results: CyO2 concentrations below 0.5 μM did not show substantial hemolytic activity, yet these concentrations enabled rapid pore-formation in HIV-infected T-cells and monocytes and increased drug uptake. ELISA for HIV-1 p24 indicated that CyO2 enhances the antiviral efficacy of both SQV and nelfinavir. CyO2 (< 0.5 μM) alone decreases HIV-1 p24 production, but it did not affect the transcription regulatory function of the HIV-1 long terminal repeat (LTR). Ultracentrifugation studies clearly showed that CyO2 exposure disrupted viral integrity and decreased the p24 content of viral particles. Furthermore, direct HIV-1 inactivation by CyO2 enhanced the efficacy of enfuvirtide. Conclusions: The membrane-active properties of CyO2 may help suppress viral load and augment antiretroviral drug efficacy. Full article

Peptides derived from the C-terminal heptad repeat (CHR) of human immunodeficiency virus type 1 (HIV-1) envelope protein transmembrane subunit gp41, such as T20 (enfuvirtide), can bind to the N-terminal heptad repeat (NHR) of gp41 and block six-helix bundle (6-HB) formation, thus inhibiting HIV-1 fusion with the target cell. However, clinical application of T20 is limited because of its low potency and genetic barrier to resistance. HP23, the shortest CHR peptide, exhibits better anti-HIV-1 activity than T20, but the HIV-1 strains with E49K mutations in gp41 will become resistant to it. Here, we modified HP23 by extending its C-terminal sequence using six amino acid residues (E6) and adding IDL (Ile-Asp-Leu) to the C-terminus of E6, which is expected to bind to the shallow pocket in the gp41 NHR N-terminal region. The newly designed peptide, designated HP23-E6-IDL, was about 2- to 16-fold more potent than HP23 against a broad spectrum of HIV-1 strains and more than 12-fold more effective against HIV-1 mutants resistant to HP23. These findings suggest that addition of an anchor–tail to the C-terminus of a CHR peptide will allow binding with the pocket in the gp41 NHR that may increase the peptide’s antiviral efficacy and its genetic barrier to resistance. Full article

T-20 and T-1249 fusion inhibitor peptides were shown to interact with 1-palmitoyl-2-oleyl-phosphatidylcholine (POPC) (liquid disordered, ld) and POPC/cholesterol (1:1) (POPC/Chol) (liquid ordered, lo) bilayers, and they do so to different extents. Although they both possess a tryptophan-rich domain (TRD), T-20 lacks a pocket binding domain (PBD), which is present in T-1249. It has been postulated that the PBD domain enhances FI interaction with HIV gp41 protein and with model membranes. Interaction of these fusion inhibitor peptides with both the cell membrane and the viral envelope membrane is important for function, i.e., inhibition of the fusion process. We address this problem with a molecular dynamics approach focusing on lipid properties, trying to ascertain the consequences and the differences in the interaction of T-20 and T-1249 with ld and lo model membranes. T-20 and T-1249 interactions with model membranes are shown to have measurable and different effects on bilayer structural and dynamical parameters. T-1249’s adsorption to the membrane surface has generally a stronger influence in the measured parameters. The presence of both binding domains in T-1249 appears to be paramount to its stronger interaction, and is shown to have a definite importance in membrane properties upon peptide adsorption. Full article

The hydrophobic pocket in the HIV-1 gp41 N-terminal heptad repeat (NHR) domain plays an important role in viral fusion and entry into the host cell, and serves as an attractive target for development of HIV-1 fusion/entry inhibitors. The peptide anti-HIV drug targeting gp41 NHR, T-20 (generic name: enfuvirtide; brand name: Fuzeon), was approved by the U.S. FDA in 2003 as the first HIV fusion/entry inhibitor for treatment of HIV/AIDS patients who fail to respond to the current antiretroviral drugs. However, because T20 lacks the pocket-binding domain (PBD), it exhibits low anti-HIV-1 activity and short half-life. Therefore, several next-generation HIV fusion inhibitory peptides with PBD have been developed. They possess longer half-life and more potent antiviral activity against a broad spectrum of HIV-1 strains, including the T-20-resistant variants. Nonetheless, the clinical application of these peptides is still limited by the lack of oral availability and the high cost of production. Thus, development of small molecule compounds targeting the gp41 pocket with oral availability has been promoted. This review describes the main approaches for identification of HIV fusion/entry inhibitors targeting the gp41 pocket and summarizes the latest progress in developing these inhibitors as a new class of anti-HIV drugs. Full article