Commercial human pulmonary artery endothelial cells have in-vitro behavior that varies by sex.

It is unknown whether biological sex influences phenotypes of commercially available human pulmonary artery endothelial cells (HPAECs). Ten lots of commercial HPAECs were used (Lonza Biologics; PromoCell). Five (50%) were confirmed to be genotypically male (+) and five (50%) were confirmed to be female (-).

Culture of pulmonary artery endothelial cells from pulmonary artery catheter balloon tips: considerations for use in pulmonary vascular disease.

Endothelial dysfunction is a hallmark of pulmonary arterial hypertension (PAH) but there are no established methods to study pulmonary artery endothelial cells (PAECs) from living patients. We sought to culture PAECs from pulmonary artery catheter (PAC) balloons used during right-heart catheterisation (RHC) to characterise successful culture attempts and to describe PAEC behaviour.PAECs were grown in primary culture to confluence and endothelial cell phenotype was confirmed.

Strain-Dependent Activation and Inhibition of Human Immunodeficiency Virus Entry by a Specific PF-68742 Stereoisomer.

Human immunodeficiency virus (HIV-1) entry into cells is mediated by the viral envelope glycoprotein (Env) trimer, which consists of three gp120 exterior glycoproteins and three gp41 transmembrane glycoproteins. When gp120 binds sequentially to the receptors CD4 and CCR5 on the target cell, the metastable Env trimer is triggered to undergo entry-related conformational changes. PF-68742 is a small molecule that inhibits the infection of a subset of HIV-1 strains by interfering with an Env function other than receptor binding.

A CD4-mimetic compound enhances vaccine efficacy against stringent immunodeficiency virus challenge.

The envelope glycoprotein (Env) trimer ((gp120/gp41)) mediates human immunodeficiency virus (HIV-1) entry into cells. The "closed," antibody-resistant Env trimer is driven to more open conformations by binding the host receptor, CD4. Broadly neutralizing antibodies that recognize conserved elements of the closed Env are potentially protective, but are elicited inefficiently.

Comparison of Uncleaved and Mature Human Immunodeficiency Virus Membrane Envelope Glycoprotein Trimers.

The mature envelope glycoprotein (Env) spike on the surfaces of human immunodeficiency virus type 1 (HIV-1)-infected cells and virions is derived from proteolytic cleavage of a trimeric gp160 glycoprotein precursor. In these studies, we compared the conformations of cleaved and uncleaved membrane Envs with truncated cytoplasmic tails to those of stabilized soluble gp140 SOSIP.664 Env trimers.

A Small-Molecule CD4-Mimetic Compound Protects Bone Marrow-Liver-Thymus Humanized Mice From HIV-1 Infection.

Background: Small-molecule CD4-mimetic compounds (CD4mc) inhibit human immunodeficiency virus (HIV-1) entry by blocking binding to the CD4 receptor and by premature triggering of the viral envelope glycoprotein (Env) spike.

Methods: The efficacy of a CD4mc in protecting bone marrow-liver-thymus (BLT) humanized mice from vaginal HIV-1 challenge was evaluated.

Results: Intravaginal application of the CD4mc JP-III-48, either before or simultaneously with virus challenge, protected BLT humanized mice from HIV-1JR-CSF infection in a dose- dependent manner.

Evaluation of the contribution of the transmembrane region to the ectodomain conformation of the human immunodeficiency virus (HIV-1) envelope glycoprotein.

Background: The human immunodeficiency virus (HIV-1) envelope glycoprotein (Env), a Type 1 transmembrane protein, assembles into a trimeric spike complex that mediates virus entry into host cells. The high potential energy of the metastable, unliganded Env trimer is maintained by multiple non-covalent contacts among the gp120 exterior and gp41 transmembrane Env subunits. Structural studies suggest that the gp41 transmembrane region forms a left-handed coiled coil that contributes to the Env trimer interprotomer contacts.

Residues in the gp41 Ectodomain Regulate HIV-1 Envelope Glycoprotein Conformational Transitions Induced by gp120-Directed Inhibitors.

Interactions between the gp120 and gp41 subunits of the human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein (Env) trimer maintain the metastable unliganded form of the viral spike. Binding of gp120 to the receptor, CD4, changes the Env conformation to promote gp120 interaction with the second receptor, CCR5 or CXCR4. CD4 binding also induces the transformation of Env into the prehairpin intermediate, in which the gp41 heptad repeat 1 (HR1) coiled coil is assembled at the trimer axis.

Activation and Inactivation of Primary Human Immunodeficiency Virus Envelope Glycoprotein Trimers by CD4-Mimetic Compounds.

Unlabelled: Human immunodeficiency virus type 1 (HIV-1) entry into cells is mediated by the viral envelope glycoproteins (Env), a trimer of three gp120 exterior glycoproteins, and three gp41 transmembrane glycoproteins. The metastable Env is triggered to undergo entry-related conformational changes when gp120 binds sequentially to the receptors, CD4 and CCR5, on the target cell. Small-molecule CD4-mimetic compounds (CD4mc) bind gp120 and act as competitive inhibitors of gp120-CD4 engagement.

Short Communication: Small-Molecule CD4 Mimetics Sensitize HIV-1-Infected Cells to Antibody-Dependent Cellular Cytotoxicity by Antibodies Elicited by Multiple Envelope Glycoprotein Immunogens in Nonhuman Primates.

Recent studies have linked antibody Fc-mediated effector functions with control of human immunodeficiency virus type 1 (HIV-1) and simian immunodeficiency virus infections. Interestingly, the presence of antibodies with potent antibody-dependent cellular cytotoxicity (ADCC) activity in RV144 vaccine trial participants correlated inversely with HIV-1 acquisition risk. These antibodies were recently found to recognize epitopes on the HIV-1 envelope (Env) glycoprotein exposed upon Env-CD4 binding.

Small-Molecule CD4-Mimics: Structure-Based Optimization of HIV-1 Entry Inhibition.

The optimization, based on computational, thermodynamic, and crystallographic data, of a series of small-molecule ligands of the Phe43 cavity of the envelope glycoprotein gp120 of human immunodeficiency virus (HIV) has been achieved. Importantly, biological evaluation revealed that the small-molecule CD4 mimics (4-7) inhibit HIV-1 entry into target cells with both significantly higher potency and neutralization breadth than previous congeners, while maintaining high selectivity for the target virus. Their binding mode was characterized via thermodynamic and crystallographic studies.

Antibodies Elicited by Multiple Envelope Glycoprotein Immunogens in Primates Neutralize Primary Human Immunodeficiency Viruses (HIV-1) Sensitized by CD4-Mimetic Compounds.

Unlabelled: The human immunodeficiency virus (HIV-1) envelope glycoproteins (Env) mediate virus entry through a series of complex conformational changes triggered by binding to the receptors CD4 and CCR5/CXCR4. Broadly neutralizing antibodies that recognize conserved Env epitopes are thought to be an important component of a protective immune response. However, to date, HIV-1 Env immunogens that elicit broadly neutralizing antibodies have not been identified, creating hurdles for vaccine development.

Discovery and optimization of novel small-molecule HIV-1 entry inhibitors using field-based virtual screening and bioisosteric replacement.

With the emergence of drug-resistant strains and the cumulative toxicities associated with current therapies, demand remains for new inhibitors of HIV-1 replication. The inhibition of HIV-1 entry is an attractive, yet underexploited therapeutic approach with implications for salvage and preexposure prophylactic regimens, as well as topical microbicides. Using the combination of a field-derived bioactive conformation template to perform virtual screening and iterative bioisosteric replacements, coupled with in silico predictions of absorption, distribution, metabolism, and excretion, we have identified new leads for HIV-1 entry inhibitors.

CD4-mimetic small molecules sensitize human immunodeficiency virus to vaccine-elicited antibodies.

Unlabelled: Approaches to prevent human immunodeficiency virus (HIV-1) transmission are urgently needed. Difficulties in eliciting antibodies that bind conserved epitopes exposed on the unliganded conformation of the HIV-1 envelope glycoprotein (Env) trimer represent barriers to vaccine development. During HIV-1 entry, binding of the gp120 Env to the initial receptor, CD4, triggers conformational changes in Env that result in the formation and exposure of the highly conserved gp120 site for interaction with the coreceptors, CCR5 and CXCR4.

Structure-Based Design and Synthesis of an HIV-1 Entry Inhibitor Exploiting X-Ray and Thermodynamic Characterization.

The design, synthesis, thermodynamic and crystallographic characterization of a potent, broad spectrum, second-generation HIV-1 entry inhibitor that engages conserved carbonyl hydrogen bonds within gp120 has been achieved. The optimized antagonist exhibits a sub-micromolar binding affinity (110 nM) and inhibits viral entry of clade B and C viruses (IC geometric mean titer of 1.7 and 14.

Inhibiting early-stage events in HIV-1 replication by small-molecule targeting of the HIV-1 capsid.

The HIV-1 capsid (CA) protein plays essential roles in both early and late stages of virl replication and has emerged as a novel drug target. We report hybrid structure-based virtual screening to identify small molecules with the potential to interact with the N-terminal domain (NTD) of HIV-1 CA and disrupt early, preintegration steps of the HIV-1 replication cycle. The small molecule 4,4'-[dibenzo[b,d]furan-2,8-diylbis(5-phenyl-1H-imidazole-4,2-diyl)]dibenzoic acid (CK026), which had anti-HIV-1 activity in single- and multiple-round infections but failed to inhibit viral replication in peripheral blood mononuclear cells (PBMCs), was identified.

Structure-based design, synthesis, and characterization of dual hotspot small-molecule HIV-1 entry inhibitors.

Cellular infection by HIV-1 is initiated with a binding event between the viral envelope glycoprotein gp120 and the cellular receptor protein CD4. The CD4-gp120 interface is dominated by two hotspots: a hydrophobic gp120 cavity capped by Phe43(CD4) and an electrostatic interaction between residues Arg59(CD4) and Asp368(gp120). The CD4 mimetic small-molecule NBD-556 (1) binds within the gp120 cavity; however, 1 and related congeners demonstrate limited viral neutralization breadth.

Liposomes for HIV prophylaxis.

There are approximately 33.4 million adults living with HIV worldwide of which an estimated 15.7 million are women.

Contribution of intrinsic reactivity of the HIV-1 envelope glycoproteins to CD4-independent infection and global inhibitor sensitivity.

Human immunodeficiency virus (HIV-1) enters cells following sequential activation of the high-potential-energy viral envelope glycoprotein trimer by target cell CD4 and coreceptor. HIV-1 variants differ in their requirements for CD4; viruses that can infect coreceptor-expressing cells that lack CD4 have been generated in the laboratory. These CD4-independent HIV-1 variants are sensitive to neutralization by multiple antibodies that recognize different envelope glycoprotein epitopes.

Design, synthesis and biological evaluation of small molecule inhibitors of CD4-gp120 binding based on virtual screening.

The low-molecular-weight compound JRC-II-191 inhibits infection of HIV-1 by blocking the binding of the HIV-1 envelope glycoprotein gp120 to the CD4 receptor and is therefore an important lead in the development of a potent viral entry inhibitor. Reported here is the use of two orthogonal screening methods, gold docking and ROCS shape-based similarity searching, to identify amine-building blocks that, when conjugated to the core scaffold, yield novel analogs that maintain similar affinity for gp120. Use of this computational approach to expand SAR produced analogs of equal inhibitory activity but with diverse capacity to enhance viral infection.

Hydrocarbon double-stapling remedies the proteolytic instability of a lengthy peptide therapeutic.

The pharmacologic utility of lengthy peptides can be hindered by loss of bioactive structure and rapid proteolysis, which limits bioavailability. For example, enfuvirtide (Fuzeon, T20, DP178), a 36-amino acid peptide that inhibits human immunodeficiency virus type 1 (HIV-1) infection by effectively targeting the viral fusion apparatus, has been relegated to a salvage treatment option mostly due to poor in vivo stability and lack of oral bioavailability. To overcome the proteolytic shortcomings of long peptides as therapeutics, we examined the biophysical, biological, and pharmacologic impact of inserting all-hydrocarbon staples into an HIV-1 fusion inhibitor.

Transitions to and from the CD4-bound conformation are modulated by a single-residue change in the human immunodeficiency virus type 1 gp120 inner domain.

Binding to the primary receptor CD4 induces conformational changes in the human immunodeficiency virus type 1 (HIV-1) gp120 envelope glycoprotein that allow binding to the coreceptor (CCR5 or CXCR4) and ultimately trigger viral membrane-cell membrane fusion mediated by the gp41 transmembrane envelope glycoprotein. Here we report the derivation of an HIV-1 gp120 variant, H66N, that confers envelope glycoprotein resistance to temperature extremes. The H66N change decreases the spontaneous sampling of the CD4-bound conformation by the HIV-1 envelope glycoproteins, thus diminishing CD4-independent infection.

Soluble CD4 and CD4-mimetic compounds inhibit HIV-1 infection by induction of a short-lived activated state.

Binding to the CD4 receptor induces conformational changes in the human immunodeficiency virus (HIV-1) gp120 exterior envelope glycoprotein. These changes allow gp120 to bind the coreceptor, either CCR5 or CXCR4, and prime the gp41 transmembrane envelope glycoprotein to mediate virus-cell membrane fusion and virus entry. Soluble forms of CD4 (sCD4) and small-molecule CD4 mimics (here exemplified by JRC-II-191) also induce these conformational changes in the HIV-1 envelope glycoproteins, but typically inhibit HIV-1 entry into CD4-expressing cells.

Small-molecule CD4 mimics interact with a highly conserved pocket on HIV-1 gp120.

Human immunodeficiency virus (HIV-1) interaction with the primary receptor, CD4, induces conformational changes in the viral envelope glycoproteins that allow binding to the CCR5 second receptor and virus entry into the host cell. The small molecule NBD-556 mimics CD4 by binding the gp120 exterior envelope glycoprotein, moderately inhibiting virus entry into CD4-expressing target cells and enhancing CCR5 binding and virus entry into CCR5-expressing cells lacking CD4. Studies of NBD-556 analogs and gp120 mutants suggest that (1) NBD-556 binds within the Phe 43 cavity, a highly conserved, functionally important pocket formed as gp120 assumes the CD4-bound conformation; (2) the NBD-556 phenyl ring projects into the Phe 43 cavity; (3) enhancement of CD4-independent infection by NBD-556 requires the induction of conformational changes in gp120; and (4) increased affinity of NBD-556 analogs for gp120 improves antiviral potency during infection of CD4-expressing cells.