Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 74
Filtrar
Más filtros

Banco de datos
País/Región como asunto
Tipo del documento
Intervalo de año de publicación
1.
Proc Natl Acad Sci U S A ; 120(13): e2222073120, 2023 03 28.
Artículo en Inglés | MEDLINE | ID: mdl-36961924

RESUMEN

Binding to the host cell receptors, CD4 and CCR5/CXCR4, triggers large-scale conformational changes in the HIV-1 envelope glycoprotein (Env) trimer [(gp120/gp41)3] that promote virus entry into the cell. CD4-mimetic compounds (CD4mcs) comprise small organic molecules that bind in the highly conserved CD4-binding site of gp120 and prematurely induce inactivating Env conformational changes, including shedding of gp120 from the Env trimer. By inducing more "open," antibody-susceptible Env conformations, CD4mcs also sensitize HIV-1 virions to neutralization by antibodies and infected cells to antibody-dependent cellular cytotoxicity (ADCC). Here, we report the design, synthesis, and evaluation of novel CD4mcs based on an indoline scaffold. Compared with our current lead indane scaffold CD4mc, BNM-III-170, several indoline CD4mcs exhibit increased potency and breadth against HIV-1 variants from different geographic clades. Viruses that were selected for resistance to the lead indane CD4mc, BNM-III-170, are susceptible to inhibition by the indoline CD4mcs. The indoline CD4mcs also potently sensitize HIV-1-infected cells to ADCC mediated by plasma from HIV-1-infected individuals. Crystal structures indicate that the indoline CD4mcs gain potency compared to the indane CD4mcs through more favorable π-π overlap from the indoline pose and by making favorable contacts with the vestibule of the CD4-binding pocket on gp120. The rational design of indoline CD4mcs thus holds promise for further improvements in antiviral activity, potentially contributing to efforts to treat and prevent HIV-1 infection.


Asunto(s)
Infecciones por VIH , Seropositividad para VIH , VIH-1 , Humanos , Citotoxicidad Celular Dependiente de Anticuerpos , Proteína gp120 de Envoltorio del VIH , Antígenos CD4/metabolismo , Anticuerpos Anti-VIH/farmacología
3.
J Virol ; 97(11): e0117123, 2023 Nov 30.
Artículo en Inglés | MEDLINE | ID: mdl-37888980

RESUMEN

IMPORTANCE: CD4-mimetic compounds (CD4mcs) are small-molecule inhibitors of human immunodeficiency virus (HIV-1) entry into host cells. CD4mcs target a pocket on the viral envelope glycoprotein (Env) spike that is used for binding to the receptor, CD4, and is highly conserved among HIV-1 strains. Nonetheless, naturally occurring HIV-1 strains exhibit a wide range of sensitivities to CD4mcs. Our study identifies changes distant from the binding pocket that can influence the susceptibility of natural HIV-1 strains to the antiviral effects of multiple CD4mcs. We relate the antiviral potency of the CD4mc against this panel of HIV-1 variants to the ability of the CD4mc to activate entry-related changes in Env conformation prematurely. These findings will guide efforts to improve the potency and breadth of CD4mcs against natural HIV-1 variants.


Asunto(s)
Fármacos Anti-VIH , Antígenos CD4 , Proteína gp120 de Envoltorio del VIH , VIH-1 , Imitación Molecular , Receptores del VIH , Humanos , Fármacos Anti-VIH/química , Fármacos Anti-VIH/farmacología , Sitios de Unión/efectos de los fármacos , Antígenos CD4/química , Antígenos CD4/metabolismo , Proteína gp120 de Envoltorio del VIH/química , Proteína gp120 de Envoltorio del VIH/metabolismo , VIH-1/química , VIH-1/clasificación , VIH-1/efectos de los fármacos , VIH-1/metabolismo , Unión Proteica/efectos de los fármacos , Receptores del VIH/metabolismo , Internalización del Virus/efectos de los fármacos
5.
J Virol ; 96(17): e0063622, 2022 09 14.
Artículo en Inglés | MEDLINE | ID: mdl-35980207

RESUMEN

Binding to the host cell receptors CD4 and CCR5/CXCR4 triggers conformational changes in the human immunodeficiency virus (HIV-1) envelope glycoprotein (Env) trimer that promote virus entry. CD4 binding allows the gp120 exterior Env to bind CCR5/CXCR4 and induces a short-lived prehairpin intermediate conformation in the gp41 transmembrane Env. Small-molecule CD4-mimetic compounds (CD4mcs) bind within the conserved Phe-43 cavity of gp120, near the binding site for CD4. CD4mcs like BNM-III-170 inhibit HIV-1 infection by competing with CD4 and by prematurely activating Env, leading to irreversible inactivation. In cell culture, we selected and analyzed variants of the primary HIV-1AD8 strain resistant to BNM-III-170. Two changes (S375N and I424T) in gp120 residues that flank the Phe-43 cavity each conferred an ~5-fold resistance to BNM-III-170 with minimal fitness cost. A third change (E64G) in layer 1 of the gp120 inner domain resulted in ~100-fold resistance to BNM-III-170, ~2- to 3-fold resistance to soluble CD4-Ig, and a moderate decrease in viral fitness. The gp120 changes additively or synergistically contributed to BNM-III-170 resistance. The sensitivity of the Env variants to BNM-III-170 inhibition of virus entry correlated with their sensitivity to BNM-III-170-induced Env activation and shedding of gp120. Together, the S375N and I424T changes, but not the E64G change, conferred >100-fold and 33-fold resistance to BMS-806 and BMS-529 (temsavir), respectively, potent HIV-1 entry inhibitors that block Env conformational transitions. These studies identify pathways whereby HIV-1 can develop resistance to CD4mcs and conformational blockers, two classes of entry inhibitors that target the conserved gp120 Phe-43 cavity. IMPORTANCE CD4-mimetic compounds (CD4mcs) and conformational blockers like BMS-806 and BMS-529 (temsavir) are small-molecule inhibitors of human immunodeficiency virus (HIV-1) entry into host cells. Although CD4mcs and conformational blockers inhibit HIV-1 entry by different mechanisms, they both target a pocket on the viral envelope glycoprotein (Env) spike that is used for binding to the receptor CD4 and is highly conserved among HIV-1 strains. Our study identifies changes near this pocket that can confer various levels of resistance to the antiviral effects of a CD4mc and conformational blockers. We relate the antiviral potency of a CD4mc against this panel of HIV-1 variants to the ability of the CD4mc to activate changes in Env conformation and to induce the shedding of the gp120 exterior Env from the spike. These findings will guide efforts to improve the potency and breadth of small-molecule HIV-1 entry inhibitors.


Asunto(s)
Antígenos CD4 , Farmacorresistencia Viral , Glicoproteínas , Guanidinas , Indenos , Mutación , Productos del Gen env del Virus de la Inmunodeficiencia Humana , Sitios de Unión/genética , Antígenos CD4/química , Antígenos CD4/metabolismo , Farmacorresistencia Viral/genética , Glicoproteínas/química , Glicoproteínas/genética , Glicoproteínas/metabolismo , Guanidinas/química , Guanidinas/farmacología , Proteína gp120 de Envoltorio del VIH/química , Proteína gp120 de Envoltorio del VIH/genética , Proteína gp120 de Envoltorio del VIH/metabolismo , Proteína gp41 de Envoltorio del VIH/química , Proteína gp41 de Envoltorio del VIH/genética , Proteína gp41 de Envoltorio del VIH/metabolismo , Inhibidores de Fusión de VIH/química , Inhibidores de Fusión de VIH/farmacología , Infecciones por VIH/tratamiento farmacológico , Infecciones por VIH/virología , VIH-1/química , VIH-1/efectos de los fármacos , VIH-1/metabolismo , Humanos , Indenos/química , Indenos/farmacología , Conformación Proteica/efectos de los fármacos , Receptores del VIH/química , Receptores del VIH/metabolismo , Productos del Gen env del Virus de la Inmunodeficiencia Humana/química , Productos del Gen env del Virus de la Inmunodeficiencia Humana/genética , Productos del Gen env del Virus de la Inmunodeficiencia Humana/metabolismo
6.
PLoS Pathog ; 16(5): e1008577, 2020 05.
Artículo en Inglés | MEDLINE | ID: mdl-32392227

RESUMEN

The HIV-1 envelope glycoprotein (Env) mediates viral entry via conformational changes associated with binding the cell surface receptor (CD4) and coreceptor (CCR5/CXCR4), resulting in subsequent fusion of the viral and cellular membranes. While the gp120 Env surface subunit has been extensively studied for its role in viral entry and evasion of the host immune response, the gp41 transmembrane glycoprotein and its role in natural infection are less well characterized. Here, we identified a primary HIV-1 Env variant that consistently supports >300% increased viral infectivity in the presence of autologous or heterologous HIV-positive plasma. However, in the absence of HIV-positive plasma, viruses with this Env exhibited reduced infectivity that was not due to decreased CD4 binding. Using Env chimeras and sequence analysis, we mapped this phenotype to a change Q563R, in the gp41 heptad repeat 1 (HR1) region. We demonstrate that Q563R reduces viral infection by disrupting formation of the gp41 six-helix bundle required for virus-cell membrane fusion. Intriguingly, antibodies that bind cluster I epitopes on gp41 overcome this inhibitory effect, restoring infectivity to wild-type levels. We further demonstrate that the Q563R change increases HIV-1 sensitivity to broadly neutralizing antibodies (bNAbs) targeting the gp41 membrane-proximal external region (MPER). In summary, we identify an HIV-1 Env variant with impaired infectivity whose Env functionality is restored through the binding of host antibodies. These data contribute to our understanding of gp41 residues involved in membrane fusion and identify a mechanism by which host factors can alleviate a viral defect.


Asunto(s)
Anticuerpos Neutralizantes/farmacología , Anticuerpos Anti-VIH/farmacología , Proteína gp41 de Envoltorio del VIH , Infecciones por VIH/inmunología , VIH-1/inmunología , Internalización del Virus/efectos de los fármacos , Anticuerpos Neutralizantes/inmunología , Antígenos CD4/inmunología , Células HEK293 , Anticuerpos Anti-VIH/inmunología , Proteína gp41 de Envoltorio del VIH/antagonistas & inhibidores , Proteína gp41 de Envoltorio del VIH/inmunología , Infecciones por VIH/tratamiento farmacológico , Infecciones por VIH/patología , Humanos
7.
J Virol ; 94(10)2020 05 04.
Artículo en Inglés | MEDLINE | ID: mdl-32161177

RESUMEN

During human immunodeficiency virus type 1 (HIV-1) entry into cells, the viral envelope glycoprotein (Env) trimer [(gp120/gp41)3] binds the receptors CD4 and CCR5 and fuses the viral and cell membranes. CD4 binding changes Env from a pretriggered (state-1) conformation to more open downstream conformations. BMS-378806 (here called BMS-806) blocks CD4-induced conformational changes in Env important for entry and is hypothesized to stabilize a state-1-like Env conformation, a key vaccine target. Here, we evaluated the effects of BMS-806 on the conformation of Env on the surface of cells and virus-like particles. BMS-806 strengthened the labile, noncovalent interaction of gp120 with the Env trimer, enhanced or maintained the binding of most broadly neutralizing antibodies, and decreased the binding of poorly neutralizing antibodies. Thus, in the presence of BMS-806, the cleaved Env on the surface of cells and virus-like particles exhibits an antigenic profile consistent with a state-1 conformation. We designed novel BMS-806 analogues that stabilized the Env conformation for several weeks after a single application. These long-acting BMS-806 analogues may facilitate enrichment of the metastable state-1 Env conformation for structural characterization and presentation to the immune system.IMPORTANCE The envelope glycoprotein (Env) spike on the surface of human immunodeficiency virus type 1 (HIV-1) mediates the entry of the virus into host cells and is also the target for antibodies. During virus entry, Env needs to change shape. Env flexibility also contributes to the ability of HIV-1 to evade the host immune response; many shapes of Env raise antibodies that cannot recognize the functional Env and therefore do not block virus infection. We found that an HIV-1 entry inhibitor, BMS-806, stabilizes the functional shape of Env. We developed new variants of BMS-806 that stabilize Env in its natural state for long periods of time. The availability of such long-acting stabilizers of Env shape will allow the natural Env conformation to be characterized and tested for efficacy as a vaccine.


Asunto(s)
Glicoproteínas/química , Glicoproteínas/efectos de los fármacos , Proteína gp120 de Envoltorio del VIH/química , Proteína gp120 de Envoltorio del VIH/efectos de los fármacos , VIH-1/inmunología , Piperazinas/farmacología , Internalización del Virus/efectos de los fármacos , Células A549 , Anticuerpos Neutralizantes/inmunología , Antígenos CD4/efectos de los fármacos , Antígenos CD4/metabolismo , Glicoproteínas/genética , Células HEK293 , Proteína gp120 de Envoltorio del VIH/genética , Proteína gp120 de Envoltorio del VIH/metabolismo , VIH-1/efectos de los fármacos , VIH-1/genética , Humanos , Ligandos , Modelos Moleculares , Conformación Proteica
8.
J Virol ; 93(21)2019 11 01.
Artículo en Inglés | MEDLINE | ID: mdl-31391272

RESUMEN

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. Determinants of HIV-1 resistance to PF-68742 map to the disulfide loop and fusion peptide of gp41. Of the four possible PF-68742 stereoisomers, only one, MF275, inhibited the infection of CD4-positive CCR5-positive cells by some HIV-1 strains. MF275 inhibition of these HIV-1 strains occurred after CD4 binding but before the formation of the gp41 six-helix bundle. Unexpectedly, MF275 activated the infection of CD4-negative CCR5-positive cells by several HIV-1 strains resistant to the inhibitory effects of the compound in CD4-positive target cells. In contrast to CD4 complementation by CD4-mimetic compounds, activation of CD4-independent infection by MF275 did not depend upon the availability of the gp120 Phe 43 cavity. Sensitivity to inhibitors indicates that MF275-activated virus entry requires formation/exposure of the gp41 heptad repeat (HR1) as well as CCR5 binding. MF275 apparently activates a virus entry pathway parallel to that triggered by CD4 and CD4-mimetic compounds. Strain-dependent divergence in Env conformational transitions allows different outcomes, inhibition or activation, in response to MF275. Understanding the mechanisms of MF275 activity should assist efforts to optimize its utility.IMPORTANCE Envelope glycoprotein (Env) spikes on the surface of human immunodeficiency virus (HIV-1) bind target cell receptors, triggering changes in the shape of Env. We studied a small molecule, MF275, that also induced shape changes in Env. The consequences of MF275 interaction with Env depended on the HIV-1 strain, with infection by some viruses inhibited and infection by other viruses enhanced. These studies reveal the strain-dependent diversity of HIV-1 Envs as they undergo shape changes in proceeding down the entry pathway. Appreciation of this diversity will assist attempts to develop broadly active inhibitors of HIV-1 entry.


Asunto(s)
Proteína gp120 de Envoltorio del VIH/metabolismo , Proteína gp41 de Envoltorio del VIH/metabolismo , Infecciones por VIH/tratamiento farmacológico , VIH-1/clasificación , VIH-1/efectos de los fármacos , Piridonas/farmacología , Sulfonamidas/farmacología , Internalización del Virus/efectos de los fármacos , Antivirales/farmacología , Linfocitos T CD4-Positivos/efectos de los fármacos , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/metabolismo , Proteína gp120 de Envoltorio del VIH/química , Proteína gp120 de Envoltorio del VIH/genética , Proteína gp41 de Envoltorio del VIH/química , Proteína gp41 de Envoltorio del VIH/genética , Infecciones por VIH/metabolismo , Infecciones por VIH/virología , Humanos , Unión Proteica , Conformación Proteica , Multimerización de Proteína , Piridonas/química , Receptores CCR5/genética , Receptores CCR5/metabolismo , Estereoisomerismo , Sulfonamidas/química , Replicación Viral
9.
J Virol ; 92(12)2018 06 15.
Artículo en Inglés | MEDLINE | ID: mdl-29618643

RESUMEN

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. Deletion of the gp41 cytoplasmic tail did not significantly affect the sensitivity of viruses with the HIV-1AD8 Env to inhibition by antibodies or a CD4-mimetic compound. After glutaraldehyde fixation and purification from membranes, a cleaved Env exhibited a hydrodynamic radius of ∼10 nm and an antibody-binding profile largely consistent with that expected based on virus neutralization sensitivity. The purified cleaved Env trimers exhibited a hollow architecture with a central void near the trimer axis. Uncleaved Env, cross-linked and purified in parallel, exhibited a hydrodynamic radius similar to that of the cleaved Env. However, the uncleaved Env was recognized by poorly neutralizing antibodies and appeared by negative-stain electron microscopy to sample multiple conformations. Compared with membrane Envs, stabilized soluble gp140 SOSIP.664 Env trimers appear to be more compact, as reflected in their smaller hydrodynamic radii and negative-stain electron microscopy structures. The antigenic features of the soluble gp140 SOSIP.664 Env trimers differed from those of the cleaved membrane Env, particularly in gp120 V3 and some CD4-binding-site epitopes. Thus, proteolytic maturation allows the membrane-anchored Env to achieve a conformation that retains functional metastability but masks epitopes for poorly neutralizing antibodies.IMPORTANCE The entry of human immunodeficiency virus type 1 (HIV-1) into host cells is mediated by the envelope glycoprotein (Env) spike on the surface of the virus. Host antibodies elicited during natural HIV-1 infection or by vaccination can potentially recognize the Env spike and block HIV-1 infection. However, the changing shape of the HIV-1 Env spike protects the virus from antibody binding. Understanding the shapes of natural and man-made preparations of HIV-1 Envs will assist the development of effective vaccines against the virus. Here, we evaluate the effects of several Env modifications commonly used to produce Env preparations for vaccine studies and the determination of structure. We found that the cleavage of the HIV-1 Env precursor helps Env to assume its natural shape, which resists the binding of many commonly elicited antibodies. Stabilized soluble Envs exhibit more compact shapes but expose some Env elements differently than the natural Env.


Asunto(s)
Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/inmunología , Anticuerpos Anti-VIH/inmunología , VIH-1/inmunología , Productos del Gen env del Virus de la Inmunodeficiencia Humana/inmunología , Animales , Línea Celular Tumoral , Perros , Glutaral/química , Células HEK293 , Proteína gp120 de Envoltorio del VIH/inmunología , Proteína gp120 de Envoltorio del VIH/metabolismo , Proteína gp41 de Envoltorio del VIH/genética , Humanos , Conformación Proteica , Multimerización de Proteína , Productos del Gen env del Virus de la Inmunodeficiencia Humana/metabolismo
10.
Nat Chem Biol ; 13(10): 1115-1122, 2017 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-28825711

RESUMEN

The HIV-1 envelope (Env) spike is a conformational machine that transitions between prefusion (closed, CD4- and CCR5-bound) and postfusion states to facilitate HIV-1 entry into cells. Although the prefusion closed conformation is a potential target for inhibition, development of small-molecule leads has been stymied by difficulties in obtaining structural information. Here, we report crystal structures at 3.8-Å resolution of an HIV-1-Env trimer with BMS-378806 and a derivative BMS-626529 for which a prodrug version is currently in Phase III clinical trials. Both lead candidates recognized an induced binding pocket that was mostly excluded from solvent and comprised of Env elements from a conserved helix and the ß20-21 hairpin. In both structures, the ß20-21 region assumed a conformation distinct from prefusion-closed and CD4-bound states. Together with biophysical and antigenicity characterizations, the structures illuminate the allosteric and competitive mechanisms by which these small-molecule leads inhibit CD4-induced structural changes in Env.


Asunto(s)
Proteína gp120 de Envoltorio del VIH/química , Proteína gp41 de Envoltorio del VIH/química , Piperazinas/química , Bibliotecas de Moléculas Pequeñas/química , Triazoles/química , Internalización del Virus/efectos de los fármacos , Cristalografía por Rayos X , Proteína gp120 de Envoltorio del VIH/antagonistas & inhibidores , Proteína gp41 de Envoltorio del VIH/antagonistas & inhibidores , Modelos Moleculares , Piperazinas/farmacología , Bibliotecas de Moléculas Pequeñas/farmacología , Relación Estructura-Actividad , Triazoles/farmacología
11.
J Infect Dis ; 218(3): 471-475, 2018 07 02.
Artículo en Inglés | MEDLINE | ID: mdl-29617845

RESUMEN

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. Conclusion: The direct antiviral effects of a CD4mc prevent HIV-1 infection in a murine model of sexual transmission.


Asunto(s)
Biomimética , Antígenos CD4/administración & dosificación , Inhibidores de Fusión de VIH/administración & dosificación , Infecciones por VIH/prevención & control , VIH-1/efectos de los fármacos , Administración Intravaginal , Animales , Médula Ósea , Modelos Animales de Enfermedad , Femenino , Hígado , Ratones SCID , Timo , Resultado del Tratamiento
12.
J Virol ; 91(3)2017 Feb 01.
Artículo en Inglés | MEDLINE | ID: mdl-27881646

RESUMEN

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. Some CD4mc have been shown to trigger Env prematurely, initially activating Env function, followed by rapid and irreversible inactivation. Here, we study CD4mc with a wide range of anti-HIV-1 potencies and demonstrate that all tested CD4mc are capable of activating as well as inactivating Env function. Biphasic dose-response curves indicated that the occupancy of the protomers in the Env trimer governs viral activation versus inactivation. One CD4mc bound per Env trimer activated HIV-1 infection. Envs with two CD4mc bound were activated for infection of CD4-negative, CCR5-positive cells, but the infection of CD4-positive, CCR5-positive cells was inhibited. Virus was inactivated when all three Env protomers were occupied by the CD4mc, and gp120 shedding from the Env trimer was increased in the presence of some CD4mc. Env reactivity and the on rates of CD4mc binding to the Env trimer were found to be important determinants of the potency of activation and entry inhibition. Cross-sensitization of Env protomers that do not bind the CD4mc to neutralization by an anti-V3 antibody was not evident. These insights into the mechanism of antiviral activity of CD4mc should assist efforts to optimize their potency and utility. IMPORTANCE: The trimeric envelope glycoproteins of human immunodeficiency virus type 1 (HIV-1) mediate virus entry into host cells. Binding to the host cell receptors, CD4 and CCR5, triggers changes in the conformation of the HIV-1 envelope glycoprotein trimer important for virus entry. Small-molecule CD4-mimetic compounds inhibit HIV-1 infection by multiple mechanisms: (i) direct blockade of the interaction between the gp120 exterior envelope glycoprotein and CD4; (ii) premature triggering of conformational changes in the envelope glycoproteins, leading to irreversible inactivation; and (iii) exposure of cryptic epitopes to antibodies, allowing virus neutralization. The consequences of the binding of the CD4-mimetic compound to the HIV-1 envelope glycoproteins depends upon how many of the three subunits of the trimer are bound and upon the propensity of the envelope glycoproteins to undergo conformational changes. Understanding the mechanistic factors that influence the activity of CD4-mimetic compounds can help to improve their potency and coverage of diverse HIV-1 strains.


Asunto(s)
Antígenos CD4/química , Proteína gp120 de Envoltorio del VIH/química , Imitación Molecular , Multimerización de Proteína , Fármacos Anti-VIH/química , Fármacos Anti-VIH/farmacología , Anticuerpos Neutralizantes/farmacología , Antígenos CD4/genética , Antígenos CD4/metabolismo , Línea Celular , Células Gigantes , Anticuerpos Anti-VIH/farmacología , Proteína gp120 de Envoltorio del VIH/agonistas , Proteína gp120 de Envoltorio del VIH/antagonistas & inhibidores , Proteína gp120 de Envoltorio del VIH/genética , Infecciones por VIH/genética , Infecciones por VIH/metabolismo , Infecciones por VIH/virología , VIH-1/efectos de los fármacos , VIH-1/fisiología , Humanos , Concentración 50 Inhibidora , Pruebas de Sensibilidad Microbiana , Mutación , Fenotipo , Unión Proteica , Receptores CCR5/química , Receptores CCR5/genética , Receptores CCR5/metabolismo , Internalización del Virus
13.
J Virol ; 91(15)2017 08 01.
Artículo en Inglés | MEDLINE | ID: mdl-28490588

RESUMEN

The envelope glycoproteins (Envs) on the surfaces of HIV-1 particles are targeted by host antibodies. Primary HIV-1 isolates demonstrate different global sensitivities to antibody neutralization; tier-1 isolates are sensitive, whereas tier-2 isolates are more resistant. Single-site mutations in Env can convert tier-2 into tier-1-like viruses. We hypothesized that such global change in neutralization sensitivity results from weakening of intramolecular interactions that maintain Env integrity. Three strategies commonly applied to perturb protein structure were tested for their effects on global neutralization sensitivity: exposure to low temperature, Env-activating ligands, and a chaotropic agent. A large panel of diverse tier-2 isolates from clades B and C was analyzed. Incubation at 0°C, which globally weakens hydrophobic interactions, causes gradual and reversible exposure of the coreceptor-binding site. In the cold-induced state, Envs progress at isolate-specific rates to unstable forms that are sensitive to antibody neutralization and then gradually lose function. Agents that mimic the effects of CD4 (CD4Ms) also induce reversible structural changes to states that exhibit isolate-specific stabilities. The chaotropic agent urea (at low concentrations) does not affect the structure or function of native Env. However, urea efficiently perturbs metastable states induced by cold and CD4Ms and increases their sensitivity to antibody neutralization and their inactivation rates Therefore, chemical and physical agents can guide Env from the stable native state to perturbation-sensitive forms and modulate their stability to bestow tier-1-like properties on primary tier-2 strains. These concepts can be applied to enhance the potency of vaccine-elicited antibodies and microbicides at mucosal sites of HIV-1 transmission.IMPORTANCE An effective vaccine to prevent transmission of HIV-1 is a primary goal of the scientific and health care communities. Vaccine-elicited antibodies target the viral envelope glycoproteins (Envs) and can potentially inhibit infection. However, the potency of such antibodies is generally low. Single-site mutations in Env can enhance the global sensitivity of HIV-1 to neutralization by antibodies. We found that such a hypersensitivity phenotype can also be induced by agents that destabilize protein structure. Exposure to 0°C or low concentrations of Env-activating ligands gradually guides Env to metastable forms that expose cryptic epitopes and that are highly sensitive to neutralization. Low concentrations of the chaotropic agent urea do not affect native Env but destabilize perturbed states induced by cold or CD4Ms and increase their neutralization. The concept of enhancing antibody sensitivity by chemical agents that affect the structural stability of proteins can be applied to increase the potency of topical microbicides and vaccine-elicited antibodies.


Asunto(s)
Anticuerpos Neutralizantes/inmunología , Anticuerpos Anti-VIH/inmunología , VIH-1/inmunología , Productos del Gen env del Virus de la Inmunodeficiencia Humana/inmunología , Biomimética , Antígenos CD4/metabolismo , Frío , Epítopos/inmunología , Infecciones por VIH/virología , VIH-1/efectos de los fármacos , VIH-1/genética , VIH-1/aislamiento & purificación , VIH-1/efectos de la radiación , Humanos , Pruebas de Neutralización , Urea/metabolismo , Productos del Gen env del Virus de la Inmunodeficiencia Humana/genética
14.
J Virol ; 91(5)2017 03 01.
Artículo en Inglés | MEDLINE | ID: mdl-28003492

RESUMEN

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. In nature, HIV-1 Envs must balance the requirements to maintain the noncovalent association of gp120 with gp41 and to evade the host antibody response with the need to respond to CD4 binding. Here we show that the gp41 HR1 region contributes to gp120 association with the unliganded Env trimer. Changes in particular amino acid residues in the gp41 HR1 region decreased the efficiency with which Env moved from the unliganded state. Thus, these gp41 changes decreased the sensitivity of HIV-1 to cold inactivation and ligands that require Env conformational changes to bind efficiently. Conversely, these gp41 changes increased HIV-1 sensitivity to small-molecule entry inhibitors that block Env conformational changes induced by CD4. Changes in particular gp41 HR1 amino acid residues can apparently affect the relative stability of the unliganded state and CD4-induced conformations. Thus, the gp41 HR1 region contributes to the association with gp120 and regulates Env transitions from the unliganded state to downstream conformations.IMPORTANCE The development of an efficient vaccine able to prevent HIV infection is a worldwide priority. Knowledge of the envelope glycoprotein structure and the conformational changes that occur after receptor engagement will help researchers to develop an immunogen able to elicit antibodies that block HIV-1 transmission. Here we identify residues in the HIV-1 transmembrane envelope glycoprotein that stabilize the unliganded state by modulating the transitions from the unliganded state to the CD4-bound state.


Asunto(s)
Proteína gp120 de Envoltorio del VIH/química , Proteína gp41 de Envoltorio del VIH/química , VIH-1/efectos de los fármacos , Secuencias de Aminoácidos , Animales , Fármacos Anti-VIH/química , Fármacos Anti-VIH/farmacología , Perros , Células HEK293 , VIH-1/fisiología , Humanos , Piperazinas/química , Piperazinas/farmacología
15.
Proc Natl Acad Sci U S A ; 112(20): E2687-94, 2015 May 19.
Artículo en Inglés | MEDLINE | ID: mdl-25941367

RESUMEN

HIV-1-infected cells presenting envelope glycoproteins (Env) in the CD4-bound conformation on their surface are preferentially targeted by antibody-dependent cell-mediated cytotoxicity (ADCC). HIV-1 has evolved a sophisticated mechanism to avoid exposure of ADCC-mediating Env epitopes by down-regulating CD4 and by limiting the overall amount of Env at the cell surface. Here we report that small-molecule CD4-mimetic compounds induce the CD4-bound conformation of Env, and thereby sensitize cells infected with primary HIV-1 isolates to ADCC mediated by antibodies present in sera, cervicovaginal lavages, and breast milk from HIV-1-infected individuals. Importantly, we identified one CD4 mimetic with the capacity to sensitize endogenously infected ex vivo-amplified primary CD4 T cells to ADCC killing mediated by autologous sera and effector cells. Thus, CD4 mimetics hold the promise of therapeutic utility in preventing and controlling HIV-1 infection.


Asunto(s)
Citotoxicidad Celular Dependiente de Anticuerpos/inmunología , Antígenos CD4/inmunología , Infecciones por VIH/prevención & control , Infecciones por VIH/transmisión , VIH-1/inmunología , Antígenos CD4/metabolismo , Linfocitos T CD4-Positivos/inmunología , Citometría de Flujo , Células HEK293 , Humanos , Mutagénesis Sitio-Dirigida , Conformación Proteica , Proteínas Recombinantes/inmunología , Proteínas Recombinantes/metabolismo , Productos del Gen env del Virus de la Inmunodeficiencia Humana/metabolismo
16.
J Virol ; 90(10): 5031-5046, 2016 05 15.
Artículo en Inglés | MEDLINE | ID: mdl-26962221

RESUMEN

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. Small-molecule CD4-mimetic compounds engage the CD4-binding pocket on the gp120 exterior Env and induce Env conformations that are highly sensitive to neutralization by antibodies, including antibodies directed against the conserved Env region that interacts with CCR5/CXCR4. Here, we show that CD4-mimetic compounds sensitize primary HIV-1 to neutralization by antibodies that can be elicited in monkeys and humans within 6 months by several Env vaccine candidates, including gp120 monomers. Monoclonal antibodies directed against the gp120 V2 and V3 variable regions were isolated from the immunized monkeys and humans; these monoclonal antibodies neutralized a primary HIV-1 only when the virus was sensitized by a CD4-mimetic compound. Thus, in addition to their direct antiviral effect, CD4-mimetic compounds dramatically enhance the HIV-1-neutralizing activity of antibodies that can be elicited with currently available immunogens. Used as components of microbicides, the CD4-mimetic compounds might increase the protective efficacy of HIV-1 vaccines. IMPORTANCE: Preventing HIV-1 transmission is a high priority for global health. Eliciting antibodies that can neutralize transmitted strains of HIV-1 is difficult, creating problems for the development of an effective vaccine. We found that small-molecule CD4-mimetic compounds sensitize HIV-1 to antibodies that can be elicited in vaccinated humans and monkeys. These results suggest an approach to prevent HIV-1 sexual transmission in which a virus-sensitizing microbicide is combined with a vaccine.


Asunto(s)
Fármacos Anti-VIH/farmacología , Anticuerpos Neutralizantes/inmunología , Antígenos CD4/inmunología , Anticuerpos Anti-VIH/inmunología , VIH-1/inmunología , Productos del Gen env del Virus de la Inmunodeficiencia Humana/inmunología , Animales , Fármacos Anti-VIH/química , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales/aislamiento & purificación , Anticuerpos Neutralizantes/biosíntesis , Antígenos CD4/metabolismo , Epítopos/inmunología , Proteína gp120 de Envoltorio del VIH/inmunología , Infecciones por VIH/prevención & control , Infecciones por VIH/transmisión , Infecciones por VIH/virología , Haplorrinos , Humanos , Inmunización , Imitación Molecular , Fragmentos de Péptidos/inmunología
17.
Virol J ; 14(1): 33, 2017 02 16.
Artículo en Inglés | MEDLINE | ID: mdl-28209172

RESUMEN

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. Here we evaluate the contribution of the gp41 transmembrane region to the folding and stability of Env trimers. METHODS: Multiple polar/charged amino acid residues, which hypothetically disrupt the stop-transfer signal, were introduced in the proposed lipid-interactive face of the transmembrane coiled coil, allowing release of soluble cleavage-negative Envs containing the modified transmembrane region (TMmod). We also examined effects of cleavage, the cytoplasmic tail and a C-terminal fibritin trimerization (FT) motif on oligomerization, antigenicity and functionality of soluble and membrane-bound Envs. RESULTS: The introduction of polar/charged amino acids into the transmembrane region resulted in the secretion of soluble Envs from the cell. However, these TMmod Envs primarily formed dimers. By contrast, control cleavage-negative sgp140 Envs lacking the transmembrane region formed soluble trimers, dimers and monomers. TMmod and sgp140 trimers were stabilized by the addition of a C-terminal FT sequence, but still exhibited carbohydrate and antigenic signatures of a flexible ectodomain structure. On the other hand, detergent-solubilized cleaved and uncleaved Envs isolated from the membranes of expressing cells exhibited "tighter" ectodomain structures, based on carbohydrate modifications. These trimers were found to be unstable in detergent solutions, but could be stabilized by the addition of a C-terminal FT moiety. The C-terminal FT domain decreased Env cleavage and syncytium-forming ability by approximately three-fold; alteration of the FT trimerization interface restored Env cleavage and syncytium formation to near-wild-type levels. CONCLUSION: The modified transmembrane region was not conducive to trimerization of soluble Envs. However, for HIV-1 Env ectodomains that are minimally modified, membrane-anchored Envs exhibit the most native structures and can be stabilized by appropriately positioned FT domains.


Asunto(s)
Proteína gp120 de Envoltorio del VIH/metabolismo , Proteína gp41 de Envoltorio del VIH/metabolismo , Pliegue de Proteína , Multimerización de Proteína , Sustitución de Aminoácidos , Proteína gp120 de Envoltorio del VIH/química , Proteína gp120 de Envoltorio del VIH/genética , Proteína gp41 de Envoltorio del VIH/química , Proteína gp41 de Envoltorio del VIH/genética , Humanos , Proteínas Mutantes/química , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Conformación Proteica
18.
J Virol ; 88(12): 6542-55, 2014 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-24696475

RESUMEN

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. The DMJ compounds (+)-DMJ-I-228 and (+)-DMJ-II-121 bind gp120 within the conserved Phe 43 cavity near the CD4-binding site, block CD4 binding, and inhibit HIV-1 infection. Here we show that the DMJ compounds sensitize primary HIV-1, including transmitted/founder viruses, to neutralization by monoclonal antibodies directed against CD4-induced (CD4i) epitopes and the V3 region, two gp120 elements involved in coreceptor binding. Importantly, the DMJ compounds rendered primary HIV-1 sensitive to neutralization by antisera elicited by immunization of rabbits with HIV-1 gp120 cores engineered to assume the CD4-bound state. Thus, small molecules like the DMJ compounds may be useful as microbicides to inhibit HIV-1 infection directly and to sensitize primary HIV-1 to neutralization by readily elicited antibodies. IMPORTANCE: Preventing HIV-1 transmission is a priority for global health. Eliciting antibodies that can neutralize many different strains of HIV-1 is difficult, creating problems for the development of a vaccine. We found that certain small-molecule compounds can sensitize HIV-1 to particular antibodies. These antibodies can be elicited in rabbits. These results suggest an approach to prevent HIV-1 sexual transmission in which a virus-sensitizing microbicide is combined with a vaccine.


Asunto(s)
Vacunas contra el SIDA/inmunología , Antivirales/farmacología , Antígenos CD4/inmunología , Anticuerpos Anti-VIH/inmunología , Proteína gp120 de Envoltorio del VIH/antagonistas & inhibidores , Infecciones por VIH/inmunología , VIH-1/efectos de los fármacos , Vacunas contra el SIDA/administración & dosificación , Animales , Antígenos CD4/genética , Línea Celular , Femenino , Proteína gp120 de Envoltorio del VIH/inmunología , Infecciones por VIH/tratamiento farmacológico , Infecciones por VIH/virología , VIH-1/genética , VIH-1/inmunología , Humanos , Pruebas de Neutralización , Conejos
19.
Acc Chem Res ; 47(4): 1228-37, 2014 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-24502450

RESUMEN

This Account provides an overview of a multidisciplinary consortium focused on structure-based strategies to devise small molecule antagonists of HIV-1 entry into human T-cells, which if successful would hold considerable promise for the development of prophylactic modalities to prevent HIV transmission and thereby alter the course of the AIDS pandemic. Entry of the human immunodeficiency virus (HIV) into target T-cells entails an interaction between CD4 on the host T-cell and gp120, a component of the trimeric envelope glycoprotein spike on the virion surface. The resultant interaction initiates a series of conformational changes within the envelope spike that permits binding to a chemokine receptor, formation of the gp41 fusion complex, and cell entry. A hydrophobic cavity at the CD4-gp120 interface, defined by X-ray crystallography, provided an initial site for small molecule antagonist design. This site however has evolved to facilitate viral entry. As such, the binding of prospective small molecule inhibitors within this gp120 cavity can inadvertently trigger an allosteric entry signal. Structural characterization of the CD4-gp120 interface, which provided the foundation for small molecule structure-based inhibitor design, will be presented first. An integrated approach combining biochemical, virological, structural, computational, and synthetic studies, along with a detailed analysis of ligand binding energetics, revealed that modestly active small molecule inhibitors of HIV entry can also promote viral entry into cells lacking the CD4 receptor protein; these competitive inhibitors were termed small molecule CD4 mimetics. Related congeners were subsequently identified with both improved binding affinity and more potent viral entry inhibition. Further assessment of the affinity-enhanced small molecule CD4 mimetics demonstrated that premature initiation of conformational change within the viral envelope spike, prior to cell encounter, can lead to irreversible deactivation of viral entry machinery. Related congeners, which bind the same gp120 site, possess different propensities to elicit the allosteric response that underlies the undesired enhancement of CD4-independent viral entry. Subsequently, key hotspots in the CD4-gp120 interface were categorized using mutagenesis and isothermal titration calorimetry according to the capacity to increase binding affinity without triggering the allosteric signal. This analysis, combined with cocrystal structures of small molecule viral entry agonists with gp120, led to the development of fully functional antagonists of HIV-1 entry. Additional structure-based design exploiting two hotspots followed by synthesis has now yielded low micromolar inhibitors of viral entry.


Asunto(s)
Fármacos Anti-VIH/química , Fármacos Anti-VIH/farmacología , Antígenos CD4/metabolismo , VIH-1/efectos de los fármacos , Antígenos CD4/química , Cristalografía por Rayos X , Diseño de Fármacos , Proteína gp120 de Envoltorio del VIH/química , Proteína gp120 de Envoltorio del VIH/metabolismo , VIH-1/patogenicidad , Interacciones Huésped-Patógeno , Humanos , Imitación Molecular , Conformación Proteica , Bibliotecas de Moléculas Pequeñas/química , Bibliotecas de Moléculas Pequeñas/farmacología , Relación Estructura-Actividad
20.
Bioorg Med Chem Lett ; 24(23): 5439-45, 2014 Dec 01.
Artículo en Inglés | MEDLINE | ID: mdl-25454268

RESUMEN

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.


Asunto(s)
Inhibidores de Fusión de VIH/farmacología , VIH-1/efectos de los fármacos , Descubrimiento de Drogas , Humanos , Conformación Molecular
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA