oxalates and HIV-Infections

Current therapeutic armamentarium for treatment of HIV-1 infection is based on the use of highly active antiretroviral therapy that, unfortunately, does not act as a curative remedy. Moreover, duration of the therapy often results in lack of compliance with the consequent emergence of multidrug resistance. Finally, drug toxicity issues also arise during treatments. In the attempt to achieve a curative effect, in addition to invest substantial resources in finding new anti-HIV-1 agents and in optimizing antiviral lead compounds and drugs currently available, additional efforts should be done to deplete viral reservoir located within host CD4

Topics: Anti-HIV Agents; Antiretroviral Therapy, Highly Active; HIV Envelope Protein gp120; HIV Envelope Protein gp41; HIV Infections; HIV-1; Humans; Organophosphates; Oxalates; Piperazines; Protein Binding; Protein Conformation; Structure-Activity Relationship; Virus Internalization

CD4-mimetic HIV-1 entry inhibitors are small sized molecules which imitate similar conformational flexibility, in gp120, to the CD4 receptor. However, the mechanism of the conformational flexibility instigated by these small sized inhibitors is little known. Likewise, the effect of the antibody on the function of these inhibitors is also less studied. In this study, we present a thorough inspection of the mechanism of the conformational flexibility induced by a CD4-mimetic inhibitor, NBD-557, using Molecular Dynamics Simulations and free energy calculations. Our result shows the functional importance of Asn425 in substrate induced conformational dynamics in gp120. The MD simulations of Asn425Gly mutant provide a less dynamic gp120 in the presence of NBD-557 without incapacitating the binding enthalpy of NBD-557. The MD simulations of complexes with the antibody clearly show the enhanced affinity of NBD-557 due to the presence of the antibody, which is in good agreement with experimental Isothermal Titration Calorimetry results (

Topics: Anti-HIV Agents; HIV Antibodies; HIV Envelope Protein gp120; HIV Infections; HIV-1; Humans; Molecular Dynamics Simulation; Oxalates; Piperidines; Protein Binding; Protein Conformation; Thermodynamics

We previously identified two small-molecule CD4 mimetics--NBD-556 and NBD-557--and synthesized a series of NBD compounds that resulted in improved neutralization activity in a single-cycle HIV-1 infectivity assay. For the current investigation, we selected several of the most active compounds and assessed their antiviral activity on a panel of 53 reference HIV-1 Env pseudoviruses representing diverse clades of clinical isolates. The selected compounds inhibited tested clades with low-micromolar potencies. Mechanism studies indicated that they act as CD4 agonists, a potentially unfavorable therapeutic trait, in that they can bind to the gp120 envelope glycoprotein and initiate a similar physiological response as CD4. However, one of the compounds, NBD-09027, exhibited reduced agonist properties, in both functional and biophysical studies. To understand the binding mode of these inhibitors, we first generated HIV-1-resistant mutants, assessed their behavior with NBD compounds, and determined the X-ray structures of two inhibitors, NBD-09027 and NBD-10007, in complex with the HIV-1 gp120 core at ∼2-Å resolution. Both studies confirmed that the NBD compounds bind similarly to NBD-556 and NBD-557 by inserting their hydrophobic groups into the Phe43 cavity of gp120. The basic nitrogen of the piperidine ring is located in close proximity to D368 of gp120 but it does not form any H-bond or salt bridge, a likely explanation for their nonoptimal antagonist properties. The results reveal the structural and biological character of the NBD series of CD4 mimetics and identify ways to reduce their agonist properties and convert them to antagonists.

Topics: Anti-HIV Agents; CD4 Antigens; Cell Line, Tumor; Crystallography, X-Ray; HeLa Cells; HIV Envelope Protein gp120; HIV Infections; HIV-1; Humans; Oxalates; Piperidines; Protein Binding

Plasma lactate measurements are typically performed in real time, limiting their usefulness in multicenter or longitudinal studies. To determine the stability of lactate specimens, blood was drawn in sodium fluoride/potassium oxalate tubes from 13 volunteers before and after 5 min of handgrip exercise to intentionally increase lactate concentrations. Plasma was stored at -70 degrees C. Aliquots were assayed in real time and after 1, 3, 6, 9, 12, 18, and 24 months. Real-time lactate concentrations measured at baseline ranged from 0.52 to 2.23 mmol/L before and from 2.91 to 11.04 mmol/L after handgrip exercise. Using a linear mixed model, the estimated change from baseline at month 24 was 1.67% (95% confidence interval, -0.70% to 4.03%) for pre-exercise samples and 0.39% (95% CI, -1.13% to 1.91%) for post-exercise samples. Stored serial specimens from 232 HIV-infected subjects in a multicenter trial of antiretroviral therapy were also assayed centrally. Among those, median plasma lactate increased from baseline to 64 weeks by 0.4 mmol/L with zidovudine+lamivudine treatment and by 0.6 mmol/L with didanosine+stavudine (each p<0.001 from baseline; p=0.04 for difference between groups over time). When performed as in this study, frozen storage with central batch lactate analysis is appropriate for prospectively collected samples in multicenter trials.

Topics: Acidosis, Lactic; Anti-HIV Agents; Blood Preservation; Blood Specimen Collection; Didanosine; HIV Infections; Humans; Lactates; Lamivudine; Multicenter Studies as Topic; Oxalates; Randomized Controlled Trials as Topic; Reverse Transcriptase Inhibitors; Sodium Fluoride; Specimen Handling; Stavudine; Time Factors; Zidovudine

Gene amplification of virus-specific sequences is widely used as a method to detect or confirm human immunodeficiency virus (HIV) infection. In this study we used an enzyme-linked affinity assay to quantify polymerase chain reaction products from whole blood, plasma, and separated mononuclear cells collected in the presence of four common anticoagulants: acid citrate dextrose, sodium EDTA, potassium oxalate, and sodium heparin. Attenuation of the product signal was observed after amplification of nucleic acid extraction from whole blood, washed mononuclear cells, and plasma from specimens collected in sodium heparin. These inhibitory effects on gene amplification could be reversed with heparinase. The addition of as little as 0.05 U of heparin completely inhibited amplification of an HLA-DQa sequence from placental DNA. We conclude that heparin can cause attenuation or inhibition of gene amplification. Acid citrate dextrose and EDTA, which lack inhibitory activity, are the most appropriate anticoagulants for clinical blood samples when polymerase chain reaction amplification is anticipated.

Topics: Citric Acid; DNA, Viral; Edetic Acid; Enzyme-Linked Immunosorbent Assay; Gene Amplification; Genes, gag; Glucose; Heparin; HIV; HIV Infections; HLA-DQ Antigens; Humans; Leukocytes, Mononuclear; Oxalates; Placenta; Polymerase Chain Reaction