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1.
PLoS Pathog ; 18(1): e1010183, 2022 01.
Artículo en Inglés | MEDLINE | ID: mdl-34986207

RESUMEN

Antibodies are principal immune components elicited by vaccines to induce protection from microbial pathogens. In the Thai RV144 HIV-1 vaccine trial, vaccine efficacy was 31% and the sole primary correlate of reduced risk was shown to be vigorous antibody response targeting the V1V2 region of HIV-1 envelope. Antibodies against V3 also were inversely correlated with infection risk in subsets of vaccinees. Antibodies recognizing these regions, however, do not exhibit potent neutralizing activity. Therefore, we examined the antiviral potential of poorly neutralizing monoclonal antibodies (mAbs) against immunodominant V1V2 and V3 sites by passive administration of human mAbs to humanized mice engrafted with CD34+ hematopoietic stem cells, followed by mucosal challenge with an HIV-1 infectious molecular clone expressing the envelope of a tier 2 resistant HIV-1 strain. Treatment with anti-V1V2 mAb 2158 or anti-V3 mAb 2219 did not prevent infection, but V3 mAb 2219 displayed a superior potency compared to V1V2 mAb 2158 in reducing virus burden. While these mAbs had no or weak neutralizing activity and elicited undetectable levels of antibody-dependent cellular cytotoxicity (ADCC), V3 mAb 2219 displayed a greater capacity to bind virus- and cell-associated HIV-1 envelope and to mediate antibody-dependent cellular phagocytosis (ADCP) and C1q complement binding as compared to V1V2 mAb 2158. Mutations in the Fc region of 2219 diminished these effector activities in vitro and lessened virus control in humanized mice. These results demonstrate the importance of Fc functions other than ADCC for antibodies without potent neutralizing activity.


Asunto(s)
Productos del Gen env/inmunología , Anticuerpos Anti-VIH/farmacología , Infecciones por VIH , Carga Viral/efectos de los fármacos , Animales , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales/farmacología , Anticuerpos Anti-VIH/inmunología , VIH-1/inmunología , Humanos , Inmunización Pasiva , Regiones Constantes de Inmunoglobulina , Ratones , Membrana Mucosa
2.
J Virol ; 94(7)2020 03 17.
Artículo en Inglés | MEDLINE | ID: mdl-31941770

RESUMEN

Envelope (Env) glycoprotein of human immunodeficiency virus type 1 (HIV-1) is an important target for the development of an HIV vaccine. Extensive glycosylation of Env is an important feature that both protects the virus from antibody responses and serves as a target for some highly potent broadly neutralizing antibodies. Therefore, analysis of glycans on recombinant Env proteins is highly significant. Here, we present glycosylation profiles of recombinant gp120 proteins from four major clades of HIV-1 (A, B, C, and AE), produced either as research-grade material in 293 and CHO cells or as two independent lots of clinical material under good manufacturing practice (GMP) conditions. Almost all potential N-linked glycosylation sites were at least partially occupied in all proteins. The occupancy rates were largely consistent among proteins produced under different conditions, although a few sites showed substantial variability even between the two GMP lots. Our data confirmed previous studies in the field, showing an abundance of oligomannose on Env protein, with 40 to 50% of glycans being Man5 to Man9 on all four proteins under all production conditions. Overall, the differences in occupancy and glycan forms among different Env subtypes produced under different conditions were less dramatic than anticipated, and antigenicity analysis with a panel of six monoclonal antibodies, including antibodies that recognize glycan forms, showed that all four gp120s maintained their antibody-binding profiles. Such findings have major implications for the final production of a clinical HIV vaccine with Env glycoprotein components.IMPORTANCE HIV-1 Env protein is a major target for the development of an HIV-1 vaccine. Env is covered with a large number of sugar-based glycan forms; about 50% of the Env molecular weight is composed of glycans. Glycan analysis of recombinant Env is important for understanding its roles in viral pathogenesis and immune responses. The current report presents the first extensive comparison of glycosylation patterns of recombinant gp120 proteins from four major clades of HIV-1 produced in two different cell lines, grown either under laboratory conditions or at 50-liter GMP scale in different lots. Information learned in this study is valuable for the further design and production of HIV-1 Env proteins as the critical components of HIV-1 vaccine formulations.


Asunto(s)
Vacunas contra el SIDA/química , Proteína gp120 de Envoltorio del VIH/química , Polisacáridos/química , Animales , Anticuerpos Monoclonales/inmunología , Anticuerpos Neutralizantes/inmunología , Linfocitos T CD4-Positivos/citología , Células CHO , Cricetulus , Epítopos/inmunología , Glicosilación , Células HEK293 , VIH-1 , Humanos , Inmunoglobulina G/inmunología , Dominios Proteicos , Proteínas Recombinantes/química
3.
J Biol Chem ; 291(51): 26377-26387, 2016 Dec 16.
Artículo en Inglés | MEDLINE | ID: mdl-27784784

RESUMEN

Once it enters the host cell, herpes simplex virus type 1 (HSV-1) recruits a series of host cell factors to facilitate its life cycle. Here, we demonstrate that serine/arginine-rich splicing factor 2 (SRSF2), which is an important component of the splicing speckle, mediates HSV-1 replication by regulating viral gene expression at the transcriptional and posttranscriptional levels. Our results indicate that SRSF2 functions as a transcriptional activator by directly binding to infected cell polypeptide 0 (ICP0), infected cell polypeptide 27 (ICP27), and thymidine kinase promoters. Moreover, SRSF2 participates in ICP0 pre-mRNA splicing by recognizing binding sites in ICP0 exon 3. These findings provide insight into the functions of SRSF2 in HSV-1 replication and gene expression.


Asunto(s)
Regulación Viral de la Expresión Génica/fisiología , Herpesvirus Humano 1/fisiología , Precursores del ARN/biosíntesis , Empalme del ARN/fisiología , ARN Viral/biosíntesis , Factores de Empalme Serina-Arginina/metabolismo , Transcripción Genética/fisiología , Replicación Viral/fisiología , Animales , Chlorocebus aethiops , Células HeLa , Humanos , Proteínas Inmediatas-Precoces/biosíntesis , Proteínas Inmediatas-Precoces/genética , Precursores del ARN/genética , ARN Viral/genética , Factores de Empalme Serina-Arginina/genética , Ubiquitina-Proteína Ligasas/biosíntesis , Ubiquitina-Proteína Ligasas/genética , Células Vero
4.
Bioorg Med Chem ; 25(1): 27-37, 2017 01 01.
Artículo en Inglés | MEDLINE | ID: mdl-27769671

RESUMEN

By merging the critical pharmacophore of EGFR/HER2 and HDAC inhibitors into one compound, a novel series of EGFR, HER-2, and HDAC multitarget inhibitors were synthesized. Compounds 9a-l contained 4-anilinoquinazolines with C-6 triazole-linked long alkyl chains of hydroxamic acid and displayed excellent inhibition against these enzymes (compound 9d exhibited the best inhibitory potency on wild-type EGFR, HDAC1, and HDAC6 with IC50 values 0.12nM, 0.72nM and 3.2nM individually). Furthermore, compounds 9b and 9d potently inhibited proliferation of five human cancer cell lines (with IC50 values between 0.49 and 8.76µM). Further mechanistic study revealed that compound 9d also regulated the phosphorylation of EGFR and HER2 and histone H3 hyperacetylation on the cellular level and induced remarkable apoptosis in BT-474 cells. Therefore, our study suggested that a system network-based multi-target drug design strategy might provided an alternate drug design method, by taking into account the synergy effect of EGFR, HER-2 and HDAC.


Asunto(s)
Antineoplásicos/química , Antineoplásicos/farmacología , Ácidos Hidroxámicos/química , Ácidos Hidroxámicos/farmacología , Quinazolinas/química , Quinazolinas/farmacología , Antineoplásicos/síntesis química , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Diseño de Fármacos , Ensayos de Selección de Medicamentos Antitumorales , Proteínas Fluorescentes Verdes/antagonistas & inhibidores , Inhibidores de Histona Desacetilasas/síntesis química , Inhibidores de Histona Desacetilasas/química , Inhibidores de Histona Desacetilasas/farmacología , Humanos , Ácidos Hidroxámicos/síntesis química , Simulación del Acoplamiento Molecular , Quinazolinas/síntesis química , Receptor ErbB-2/antagonistas & inhibidores , Relación Estructura-Actividad , Triazoles/síntesis química , Triazoles/química , Triazoles/farmacología
5.
Nat Commun ; 15(1): 4301, 2024 May 21.
Artículo en Inglés | MEDLINE | ID: mdl-38773089

RESUMEN

The vaccine elicitation of HIV tier-2-neutralization antibodies has been a challenge. Here, we report the isolation and characterization of a CD4-binding site (CD4bs) specific monoclonal antibody, HmAb64, from a human volunteer immunized with a polyvalent DNA prime-protein boost HIV vaccine. HmAb64 is derived from heavy chain variable germline gene IGHV1-18 and light chain germline gene IGKV1-39. It has a third heavy chain complementarity-determining region (CDR H3) of 15 amino acids. On a cross-clade panel of 208 HIV-1 pseudo-virus strains, HmAb64 neutralized 20 (10%), including tier-2 strains from clades B, BC, C, and G. The cryo-EM structure of the antigen-binding fragment of HmAb64 in complex with a CNE40 SOSIP trimer revealed details of its recognition; HmAb64 uses both heavy and light CDR3s to recognize the CD4-binding loop, a critical component of the CD4bs. This study demonstrates that a gp120-based vaccine can elicit antibodies capable of tier 2-HIV neutralization.


Asunto(s)
Vacunas contra el SIDA , Anticuerpos Neutralizantes , Antígenos CD4 , Anticuerpos Anti-VIH , VIH-1 , Humanos , Vacunas contra el SIDA/inmunología , VIH-1/inmunología , Anticuerpos Anti-VIH/inmunología , Anticuerpos Neutralizantes/inmunología , Antígenos CD4/inmunología , Antígenos CD4/metabolismo , Vacunas de ADN/inmunología , Anticuerpos Monoclonales/inmunología , Infecciones por VIH/prevención & control , Infecciones por VIH/inmunología , Infecciones por VIH/virología , Microscopía por Crioelectrón , Proteína gp120 de Envoltorio del VIH/inmunología , Proteína gp120 de Envoltorio del VIH/química , Sitios de Unión , Regiones Determinantes de Complementariedad/inmunología , Regiones Determinantes de Complementariedad/química
6.
Res Sq ; 2023 Oct 20.
Artículo en Inglés | MEDLINE | ID: mdl-37886518

RESUMEN

The vaccine elicitation of HIV-neutralizing antibodies with tier-2-neutralization breadth has been a challenge. Here, we report the isolation and characteristics of a CD4-binding site specific monoclonal antibody, HmAb64, from a human volunteer immunized with a polyvalent gp120 DNA prime-protein boost vaccine. HmAb64 derived from heavy chain variable germline gene IGHV1-18, light chain germline gene IGKV1-39, and had a 3rd heavy chain complementarity determining region (CDR H3) of 15 amino acids. On a cross-clade panel of 208 HIV-1 pseudo-virus strains, HmAb64 neutralized 21 (10%), including tier-2 neutralization resistant strains from clades B, BC, C, and G. The cryo-EM structure of the antigen-binding fragment of HmAb64 bound to a conformation between prefusion closed and occluded open forms of envelope trimer, using both heavy and light CDR3s to recognize the CD4-binding loop, a critical component of the CD4-binding site. A gp120 subunit-based vaccine can thus elicit an antibody capable of tier 2-HIV neutralization.

7.
NPJ Vaccines ; 7(1): 18, 2022 Feb 09.
Artículo en Inglés | MEDLINE | ID: mdl-35140230

RESUMEN

Since publishing our original reports on the safety and immunogenicity of a polyvalent DNA prime-protein boost HIV vaccine (PDPHV) which elicited high titer antibody responses with broad specificity, neutralizing activities to multiple HIV-1 subtypes, as well as poly-functional T cell responses, accumulated findings from other HIV vaccine studies indicated the important roles of Ig isotype distribution, Fc medicated functions and the persistence of memory immune responses which were not studied in previous PDPHV related reports. The current report provides further detailed characterization of these parameters in human volunteers receiving the PDPHV regimen. Antibody responses were assessed using IgG isotype and gp70-V1V2-binding ELISAs, peptide arrays, and antibody-dependent cellular cytotoxicity (ADCC) assays. B cell ELISPOT was used to detect gp120-specific memory B cells. Our results showed that the gp120-specific antibodies were primarily of the IgG1 isotype. HIV-1 envelope protein variable regions V1 and V2 were actively targeted by the antibodies as determined by specific binding to both peptide and V1V2-carrying scaffolds. The antibodies showed potent and broad ADCC responses. Finally, the B cell ELISPOT analysis demonstrated persistence of gp120-specific memory B cells for at least 6 months after the last dose. These data indicate that broadly reactive binding Abs and ADCC responses as well as durable gp120-specific memory B cells were elicited by the polyvalent heterologous prime-boost vaccination regimens and showed great promise as a candidate HIV vaccine.

8.
Emerg Microbes Infect ; 10(1): 833-841, 2021 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-33853515

RESUMEN

While DNA prime-protein boost vaccination approach has been widely used in preclinical and clinical studies especially in the field of HIV vaccine development, the exact role of DNA immunization has not been fully identified. Our previous work demonstrated that DNA immunization was able to elicit T follicular helper (Tfh) cell responses and germinal center (GC) B cell development in a mouse model. In the current report, a mouse immunogenicity study was conducted to further ask whether DNA immunization is able to elicit antigen-specific B cell responses. Using HIV-1 Env as model antigen delivered in the form of DNA prime-protein boost, our data demonstrated that DNA prime was able to enhance the antigen-specific B cell responses for both Env-specific antibody secreting cells (ASC) and memory B cells. Furthermore, the DNA priming can greatly reduce the need of including an adjuvant as part of the recombinant protein vaccine boost formulation. Our findings revealed one mechanism that supports the value of DNA priming in assisting the inductin of high affinity and long lasting antigen specific antibody responses.


Asunto(s)
Células Productoras de Anticuerpos/inmunología , Proteína gp120 de Envoltorio del VIH/genética , Infecciones por VIH/prevención & control , Vacunas de ADN/administración & dosificación , Vacunas contra el SIDA/administración & dosificación , Vacunas contra el SIDA/inmunología , Animales , Células HEK293 , Anticuerpos Anti-VIH/sangre , Proteína gp120 de Envoltorio del VIH/inmunología , Infecciones por VIH/inmunología , Humanos , Inmunización Secundaria , Ratones , Vacunas de ADN/inmunología
9.
Emerg Microbes Infect ; 10(1): 342-355, 2021 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-33555988

RESUMEN

The current study aims to develop a safe and highly immunogenic COVID-19 vaccine. The novel combination of a DNA vaccine encoding the full-length Spike (S) protein of SARS-CoV-2 and a recombinant S1 protein vaccine induced high level neutralizing antibody and T cell immune responses in both small and large animal models. More significantly, the co-delivery of DNA and protein components at the same time elicited full protection against intratracheal challenge of SARS-CoV-2 viruses in immunized rhesus macaques. As both DNA and protein vaccines have been proven safe in previous human studies, and DNA vaccines are capable of eliciting germinal center B cell development, which is critical for high-affinity memory B cell responses, the DNA and protein co-delivery vaccine approach has great potential to serve as a safe and effective approach to develop COVID-19 vaccines that provide long-term protection.


Asunto(s)
Vacunas contra la COVID-19/inmunología , COVID-19/prevención & control , Glicoproteína de la Espiga del Coronavirus/inmunología , Vacunas de ADN/inmunología , Vacunas de Subunidad/inmunología , Animales , Anticuerpos Neutralizantes/sangre , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/sangre , Anticuerpos Antivirales/inmunología , Línea Celular , ADN/inmunología , Células HEK293 , Humanos , Recuento de Linfocitos , Macaca mulatta , Ratones , Ratones Endogámicos C57BL , Plásmidos/genética , Conejos , Proteínas Recombinantes/inmunología , SARS-CoV-2/genética , SARS-CoV-2/inmunología , Linfocitos T/inmunología
10.
J Ethnopharmacol ; 250: 112528, 2020 Mar 25.
Artículo en Inglés | MEDLINE | ID: mdl-31884038

RESUMEN

ETHNOPHARMACOLOGICAL RELEVANCE: A traditional Chinese medicine (TCM) prescription follows the principle of compatibility (peiwu) to achieve the fundamental purpose: to increase efficacy and reduce toxicity. Rhei rhizoma, commonly known as Chinese rhubarb, is the most frequently used herb with Radix Scutellariaee. This classic fixed compatibility is considered for heat-clearing, qi regulation and detoxifying to gain better efficacy and reduce cytotoxicity with respect to unilateral medicine. With this in mind, we propose it is highly promising to find ingredients in rhubarb to increase the bioavailability of baicalin. AIM OF STUDY: In the present study, effect of rhien on pharmacokinetic profile of baicalin in rat plasma was investigated, and the underlying mechanisms were partly dissected through intestinal absorption, metabolism and biliary excretion with in vivo, in vitro and in situ assays. MATERIALS AND METHODS: Pharmacokinetic analysis in rats was first performed to provide a general overview of the in vivo exposure of baicalin and rhein after co-administration, while the biliary excretion study provided insight to the effect of rhein on the transport of baicalin from hepatocytes to bile. In vitro incubation and inhibition studies in human/rat liver microsome and human/rat intestinal S9 fraction were conducted to elucidate the role of uridine diphosphate-glucuronosyltransferases (UGTs) on the hepatic and intestinal metabolism of baicalein (the aglycone of baicalin), and to determine whether rhein can affect the UGT-mediated glucuronidation of baicalein. In situ intestinal perfusion study was designed to investigate the effect of rhein on intestinal absorption of baicalin, and breast cancer resistance protein (BCRP) inhibitor was co-perfused as positive control to demonstrate the role of the efflux transporter, while BCRP-MDCK II cell(Madin-Daby canine kidney cell) model was used as an in vitro approach to further confirm the conclusion. RESULTS: The AUC and Cmax of baicalin were increased to 189.93% and 305.73%, respectively, and the clearance of baicalin was significantly decreased from 4.17 ± 2.40 to 1.65 ± 0.79 L/h/kg following oral co-administration of rhein. The AUC of baicalin was markedly increased and the biliary clearance was significantly decreased when baicalin and rhein were co-administered intravenously. The effect of rhein on the glucuronidation of baicalein in various subcellular fractions was examined, and it was found that rhein did not affect the UGT-mediated glucuronidation of baicalein. Results of in situ intestinal perfusion revealed that co-perfusion with Ko143 (a potent BCRP inhibitor) or rhein significantly reduced the cumulative excretion amount of baicalin, from 9.27 ± 2.79 to 2.80 ± 0.97 or 4.84 ± 0.60 nM, respectively. Additionally, the efflux ratio Papp(BL-AP)/Papp(AP-BL) of baicalin in BCRP-MDCK II was decreased significantly in the presence of rhein or Ko143, which meant rhein could inhibit the BCRP-mediated efflux transport of baicalin. CONCLUSIONS: These results indicated that rhein can increase the bioavailability of baicalin by inhibiting BCRP-mediated efflux transport of baicalin in enterocytes and hepatocytes rather than by affecting the activity of UGT enzyme.


Asunto(s)
Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2/metabolismo , Antraquinonas/farmacología , Flavonoides/farmacocinética , Glucuronosiltransferasa/metabolismo , Administración Oral , Animales , Antraquinonas/administración & dosificación , Área Bajo la Curva , Disponibilidad Biológica , Transporte Biológico , Perros , Interacciones Farmacológicas , Enterocitos/metabolismo , Flavonoides/administración & dosificación , Hepatocitos/metabolismo , Humanos , Absorción Intestinal , Células de Riñón Canino Madin Darby , Masculino , Microsomas Hepáticos/metabolismo , Ratas , Ratas Sprague-Dawley , Rheum/química
11.
Hum Vaccin Immunother ; 13(12): 2996-3009, 2017 12 02.
Artículo en Inglés | MEDLINE | ID: mdl-28933684

RESUMEN

Our previous preclinical studies and a Phase I clinical trial DP6-001 have indicated that a polyvalent Env formulation was able to elicit broadly reactive antibody responses including low titer neutralizing antibody responses against viral isolates of subtypes A, B, C and AE. In the current report, a panel of 62 gp120 immunogens were screened in a rabbit model to identify gp120 immunogens that can elicit improved binding and neutralizing antibody responses and some of them can be included in the next polyvalent formulation. Only about 19% of gp120 immunogens in this panel were able to elicit neutralizing antibodies against greater than 50% of the viruses included in a high throughput PhenoSense neutralization assay when these immuongens were tested as a DNA prime followed by a fixed 5-valent gp120 protein vaccine boost. The new polyvalent formulation, using five gp120 immunogens selected from this subgroup, elicited improved quality of antibody responses in rabbits than the previous DP6-001 formulation. More significantly, this new polyvalent formulation elicited higher antibody responses against a panel of gp70V1/V2 antigens expressing V1/V2 sequences from diverse subtypes. Bioinformatics analysis supports the design of a 4-valent or 5-valent formulation using gp120 immunogens from this screening study to achieve a broad coverage against 16 HIV-1 subtypes.


Asunto(s)
Vacunas contra el SIDA/inmunología , Anticuerpos Neutralizantes/sangre , Anticuerpos Anti-VIH/sangre , Proteína gp120 de Envoltorio del VIH/inmunología , Vacunas de ADN/inmunología , Vacunas contra el SIDA/administración & dosificación , Vacunas contra el SIDA/genética , Animales , Evaluación Preclínica de Medicamentos , Proteína gp120 de Envoltorio del VIH/genética , Esquemas de Inmunización , Pruebas de Neutralización , Conejos , Vacunas de ADN/administración & dosificación , Vacunas de ADN/genética , Vacunas de Subunidad/administración & dosificación , Vacunas de Subunidad/genética , Vacunas de Subunidad/inmunología
12.
Anal Chim Acta ; 934: 186-93, 2016 Aug 31.
Artículo en Inglés | MEDLINE | ID: mdl-27506359

RESUMEN

Since most of the central nervous system (CNS) drug candidates show poor permeability across the blood-brain barrier (BBB), development of a reliable platform for permeability assay will greatly accelerate drug discovery. Herein, we constructed a microfluidic BBB model to mimic drug delivery into the brain to induce cytotoxicity at target cells. To reconstitute the in vivo BBB properties, human cerebral microvessel endothelial cells (hCMEC/D3) were dynamically cultured in a membrane-based microchannel. Sunitinib, a model drug, was then delivered into the microchannel and forced to permeate through the BBB model. The permeated amount was directly quantified by an electrospray ionization quadrupole time-of-flight mass spectrometer (ESI-Q-TOF MS) after on-chip SPE (µSPE) pretreatment. Moreover, the permeated drug was incubated with glioma cells (U251) cultured inside agarose gel in the downstream to investigate drug-induced cytotoxicity. The resultant permeability of sunitinib was highly correlated with literature reported value, and it only required 30 min and 5 µL of sample solution for each permeation experiment. Moreover, after 48 h of treatment, the survival rate of U251 cells cultured in 3D scaffolds was nearly 6% higher than that in 2D, which was in accordance with the previously reported results. These results demonstrate that this platform provides a valid tool for drug permeability and cytotoxicity assays which have great value for the research and development of CNS drugs.


Asunto(s)
Barrera Hematoencefálica/efectos de los fármacos , Indoles/farmacología , Técnicas Analíticas Microfluídicas , Modelos Biológicos , Pirroles/farmacología , Encéfalo/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Sistemas de Liberación de Medicamentos , Evaluación Preclínica de Medicamentos , Humanos , Procedimientos Analíticos en Microchip , Técnicas Analíticas Microfluídicas/instrumentación , Permeabilidad/efectos de los fármacos , Sunitinib
13.
Cancer Biol Ther ; 17(8): 790-8, 2016 08 02.
Artículo en Inglés | MEDLINE | ID: mdl-27260617

RESUMEN

Hypoxia is a general event in solid tumor growth. Therefore, induced cellular responses by hypoxia are important for tumorigenesis and tumor growth. MicroRNAs (miRNAs) have recently emerged as important regulators of hypoxia induced cellular responses. Here we report that miR-147a is a novel and crucial hypoxia induced miRNA. HIF-1α up-regulates the expression of miR-147a, and miR-147a in turn stabilizes and accumulates HIF-1α protein via directly targeting HIF-3α, a dominant negative regulator of HIF-1α. Subsequent studies in xenograft mouse model reveal that miR-147a is capable of inhibiting tumor growth. Collectively, these data demonstrate a positive feedback loop between HIF-1α, miR-147a and HIF-3α, which provide a new insight into the mechanism of miR-147a induced cell proliferation arrest under hypoxia.


Asunto(s)
Hipoxia de la Célula/fisiología , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , MicroARNs/metabolismo , Secuencia de Aminoácidos , Animales , Apoptosis/fisiología , Proliferación Celular/fisiología , Células HeLa , Xenoinjertos , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , MicroARNs/biosíntesis , Transfección , Regulación hacia Arriba
14.
Emerg Microbes Infect ; 4(7): e44, 2015 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-26251831

RESUMEN

The fourth conserved region (C4) in the HIV-1 envelope glycoprotein (Env) gp120 is a structural element that is important for its function, as it binds to both the receptor CD4 and the co-receptor CCR5/CXCR4. It has long been known that this region is highly immunogenic and that it harbors B-cell as well as T-cell epitopes. It is the target of a number of antibodies in animal studies, which are called CD4-blockers. However, the mechanism by which the virus shields itself from such antibody responses is not known. Here, we determined the crystal structure of R53 in complex with its epitope peptide using a novel anti-C4 rabbit monoclonal antibody R53. Our data show that although the epitope of R53 covers a highly conserved sequence (433)AMYAPPI(439), it is not available in the gp120 trimer and in the CD4-bound conformation. Our results suggest a masking mechanism to explain how HIV-1 protects this critical region from the human immune system.


Asunto(s)
Anticuerpos Monoclonales/química , Anticuerpos Monoclonales/inmunología , Antígenos CD4/inmunología , Epítopos/inmunología , Proteína gp120 de Envoltorio del VIH/inmunología , Animales , Anticuerpos Monoclonales/metabolismo , Sitios de Unión de Anticuerpos/inmunología , Unión Competitiva , Antígenos CD4/metabolismo , Cristalización , Epítopos/química , Epítopos/metabolismo , Proteína gp120 de Envoltorio del VIH/metabolismo , Fragmentos Fab de Inmunoglobulinas/aislamiento & purificación , Fragmentos Fab de Inmunoglobulinas/metabolismo , Conejos , Receptores CCR5/inmunología , Receptores CCR5/metabolismo , Receptores CXCR4/inmunología , Receptores CXCR4/metabolismo , Ovinos
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