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Human programmed cell death protein 1 (hPD-1) is an essential receptor in the immune checkpoint pathway. It has played an important role in cancer therapy. However, not all patients respond positively to the PD-1 antibody treatment, and the underlying mechanism remains unknown. PD-1 is a transmembrane glycoprotein, and its extracellular domain (ECD) is reported to be responsible for interactions and signal transduction. This domain contains 4 N-glycosylation sites and 25 potential O-glycosylation sites, which implicates the importance of glycosylation. The structure of hPD-1 has been intensively studied, but the glycosylation of this protein, especially the glycan on each glycosylation site, has not been comprehensively illustrated. In this study, hPD-1 ECD expressed by human embryonic kidney 293 (HEK 293) and Chinese hamster ovary (CHO) cells was analyzed; not only N- and O-glycosylation sites but also the glycans on these sites were comprehensively analyzed using mass spectrometry. In addition, hPD-1 ECD binding to different anti-hPD-1 antibodies was tested, and N-glycans were found functioned differently. All of this glycan information will be beneficial for future PD-1 studies.
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Understanding the facilitator of HIV-1 infection and subsequent latency establishment may aid the discovery of potential therapeutic targets. Here, we report the elevation of plasma transforming growth factor ß (TGF-ß) during acute HIV-1 infection among men who have sex with men (MSM). Using a serum-free in vitro system, we further delineated the role of TGF-ß signaling in mediating HIV-1 infection of activated and resting memory CD4+ T cells. TGF-ß could upregulate both the frequency and expression of the HIV-1 coreceptor CCR5, thereby augmenting CCR5-tropic viral infection of resting and activated memory CD4+ T cells via Smad3 activation. The production of live HIV-1JR-FL upon infection and reactivation was increased in TGF-ß-treated resting memory CD4+ T cells without increasing CD4 expression or inducing T cell activation. The expression of CCR7, a central memory T cell marker that serves as a chemokine receptor to facilitate T cell trafficking into lymphoid organs, was also elevated on TGF-ß-treated resting and activated memory CD4+ T cells. Moreover, the expression of CXCR3, a chemokine receptor recently reported to facilitate CCR5-tropic HIV-1 infection, was increased on resting and activated memory CD4+ T cells upon TGF-ß treatment. These findings were coherent with the observation that ex vivo CCR5 and CXCR3 expression on total resting and resting memory CD4+ T cells in combination antiretroviral therapy (cART)-naive and cART-treated patients were higher than in healthy individuals. Overall, the study demonstrated that TGF-ß upregulation induced by acute HIV-1 infection might promote latency reservoir establishment by increasing infected resting memory CD4+ T cells and lymphoid organ homing of infected central memory CD4+ T cells. Therefore, TGF-ß blockade may serve as a potential supplementary regimen for HIV-1 functional cure by reducing viral latency. IMPORTANCE Incomplete eradication of HIV-1 latency reservoirs remains the major hurdle in achieving a complete HIV/AIDS cure. Dissecting the facilitator of latency reservoir establishment may aid the discovery of druggable targets for HIV-1 cure. This study showed that the T cell immunomodulatory cytokine TGF-ß was upregulated during the acute phase of infection. Using an in vitro serum-free system, we specifically delineated that TGF-ß promoted HIV-1 infection of both resting and activated memory CD4+ T cells via the induction of host CCR5 coreceptor. Moreover, TGF-ß-upregulated CCR7 or CXCR3 might promote HIV-1 latent infection by facilitating lymphoid homing or IP-10-mediated viral entry and DNA integration, respectively. Infected resting and central memory CD4+ T cells are important latency reservoirs. Increased infection of these cells mediated by TGF-ß will promote latency reservoir establishment during early infection. This study, therefore, highlighted the potential use of TGF-ß blockade as a supplementary regimen with cART in acute patients to reduce viral latency.
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Linfocitos T CD4-Positivos , Infecciones por VIH , VIH-1 , Homosexualidad Masculina , Transducción de Señal , Humanos , Masculino , Linfocitos T CD4-Positivos/virología , Infecciones por VIH/tratamiento farmacológico , Seropositividad para VIH , VIH-1/fisiología , Receptores CCR7/metabolismo , Minorías Sexuales y de Género , Factor de Crecimiento Transformador beta , Latencia del Virus/efectos de los fármacos , Replicación Viral , Transducción de Señal/efectos de los fármacosRESUMEN
OBJECTIVE: Immune checkpoint blockade (ICB) has improved cancer treatment, yet why most hepatocellular carcinoma (HCC) patients are resistant to PD-1 ICB remains elusive. Here, we elucidated the role of a programmed cell death protein 1 (PD-1) isoform, Δ42PD-1, in HCC progression and resistance to nivolumab ICB. DESIGN: We investigated 74 HCC patients in three cohorts, including 41 untreated, 28 treated with nivolumab and 5 treated with pembrolizumab. Peripheral blood mononuclear cells from blood samples and tumour infiltrating lymphocytes from tumour tissues were isolated for immunophenotyping. The functional significance of Δ42PD-1 was explored by single-cell RNA sequencing analysis and validated by functional and mechanistic studies. The immunotherapeutic efficacy of Δ42PD-1 monoclonal antibody was determined in HCC humanised mouse models. RESULTS: We found distinct T cell subsets, which did not express PD-1 but expressed its isoform Δ42PD-1, accounting for up to 71% of cytotoxic T lymphocytes in untreated HCC patients. Δ42PD-1+ T cells were tumour-infiltrating and correlated positively with HCC severity. Moreover, they were more exhausted than PD-1+ T cells by single T cell and functional analysis. HCC patients treated with anti-PD-1 ICB showed effective PD-1 blockade but increased frequencies of Δ42PD-1+ T cells over time especially in patients with progressive disease. Tumour-infiltrated Δ42PD-1+ T cells likely sustained HCC through toll-like receptors-4-signalling for tumourigenesis. Anti-Δ42PD-1 antibody, but not nivolumab, inhibited tumour growth in three murine HCC models. CONCLUSION: Our findings not only revealed a mechanism underlying resistance to PD-1 ICB but also identified anti-Δ42PD-1 antibody for HCC immunotherapy.
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Carcinoma Hepatocelular , Neoplasias Hepáticas , Ratones , Animales , Carcinoma Hepatocelular/patología , Neoplasias Hepáticas/patología , Leucocitos Mononucleares , Terapia de Inmunosupresión , Tolerancia Inmunológica , Inmunoterapia , Nivolumab/uso terapéutico , Linfocitos T CD8-positivosRESUMEN
Understanding and identifying new ways of mounting an effective CD8⺠T cell immune response is important for eliminating infectious pathogens. Although upregulated programmed death-1 (PD1) in chronic infections (such as HIV-1 and tuberculosis) impedes T cell responses, blocking this PD1/PD-L pathway could functionally rescue the "exhausted" T cells. However, there exists a number of PD1 spliced variants with unknown biological function. Here, we identified a new isoform of human PD1 (Δ42PD1) that contains a 42-nucleotide in-frame deletion located at exon 2 domain found expressed in peripheral blood mononuclear cells (PBMCs). Δ42PD1 appears to function distinctly from PD1, as it does not engage PD-L1/PD-L2 but its recombinant form could induce proinflammatory cytokines. We utilized Δ42PD1 as an intramolecular adjuvant to develop a fusion DNA vaccine with HIV-1 Gag p24 antigen to immunize mice, which elicited a significantly enhanced level of anti-p24 IgG1/IgG2a antibody titers, and important p24-specific and tetramerâºCD8⺠T cells responses that lasted for ≥7.5 months. Furthermore, p24-specific CD8⺠T cells remain functionally improved in proliferative and cytolytic capacities. Importantly, the enhanced antigen-specific immunity protected mice against pathogenic viral challenge and tumor growth. Thus, this newly identified PD1 variant (Δ42PD1) amplifies the generation of antigen-specific CD8⺠T cell immunity when used in a DNA vaccine.
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Antígeno B7-H1/genética , Linfocitos T CD8-positivos/inmunología , Isoformas de Proteínas/inmunología , Vacunas de ADN/inmunología , Animales , Células Cultivadas , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Femenino , Infecciones por VIH/inmunología , Infecciones por VIH/prevención & control , Humanos , Leucocitos Mononucleares/inmunología , Leucocitos Mononucleares/metabolismo , Ratones , Ratones Endogámicos BALB C , Reacción en Cadena de la Polimerasa , Proteína 2 Ligando de Muerte Celular Programada 1/genética , Vacunas de ADN/química , Vacunas de ADN/genéticaRESUMEN
Oncolytic virotherapy is an innovative approach for cancer treatment. However, recruitment of myeloid-derived suppressor cells (MDSCs) into the tumor microenvironment (TME) after oncolysis-mediated local inflammation leads to tumor resistance to the therapy. Using the murine malignant mesothelioma model, we demonstrated that the in situ vaccinia virotherapy recruited primarily polymorphonuclear MDSCs (PMN-MDSCs) into the TME, where they exhibited strong suppression of cytotoxic T lymphocytes in a reactive oxygen species-dependent way. Single-cell RNA sequencing analysis confirmed the suppressive profile of PMN-MDSCs at the transcriptomic level and identified CXCR2 as a therapeutic target expressed on PMN-MDSCs. Abrogating PMN-MDSC trafficking by CXCR2-specific small molecule inhibitor during the vaccinia virotherapy exhibited enhanced antitumor efficacy in 3 syngeneic cancer models, through increasing CD8+/MDSC ratios in the TME, activating cytotoxic T lymphocytes, and skewing suppressive TME into an antitumor environment. Our results warrant clinical development of CXCR2 inhibitor in combination with oncolytic virotherapy.
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Células Supresoras de Origen Mieloide , Viroterapia Oncolítica , Vaccinia , Animales , Ratones , Línea Celular Tumoral , Células Supresoras de Origen Mieloide/patología , Linfocitos T Citotóxicos , Microambiente Tumoral , Vaccinia/patología , Virus VacciniaRESUMEN
Regardless of the route of transmission, R5-tropic HIV-1 predominates early in infection, rendering C-C chemokine receptor type 5 (CCR5) antagonists as attractive agents not only for antiretroviral therapy but also for prevention. Here, we report the specificity, potency, and underlying mechanism of action of a novel small molecule CCR5 antagonist, TD-0680. TD-0680 displayed the greatest potency against a diverse group of R5-tropic HIV-1 and SIV strains when compared with its prodrug, TD-0232, the Food and Drug Administration-approved CCR5 antagonist Maraviroc, and TAK-779, with EC(50) values in the subnanomolar range (0.09-2.29 nm). Importantly, TD-0680 was equally potent at blocking envelope-mediated cell-cell fusion and cell-mediated viral transmission as well as the replication of a TAK-779/Maraviroc-resistant HIV-1 variant. Interestingly, TD-0232 and TD-0680 functioned differently despite binding to a similar transmembrane pocket of CCR5. Site-directed mutagenesis, drug combination, and antibody blocking assays identified a novel mechanism of action of TD-0680. In addition to binding to the transmembrane pocket, the unique exo configuration of this molecule protrudes and sterically blocks access to the extracellular loop 2 (ECL2) region of CCR5, thereby interrupting the interaction between virus and its co-receptor more effectively. This mechanism of action was supported by the observations of similar TD-0680 potency against CD4-dependent and -independent SIV strains and by molecular docking analysis using a CCR5 model. TD-0680, therefore, merits development as an anti-HIV-1 agent for therapeutic purposes and/or as a topical microbicide for the prevention of sexual transmission of R5-tropic HIV-1.
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Fármacos Anti-VIH/farmacología , Antagonistas de los Receptores CCR5 , Infecciones por VIH/tratamiento farmacológico , VIH-1/fisiología , Sulfonamidas/farmacología , Tropanos/farmacología , Internalización del Virus/efectos de los fármacos , Replicación Viral/efectos de los fármacos , Amidas/farmacología , Sitios de Unión , Línea Celular , Ciclohexanos/farmacología , Infecciones por VIH/metabolismo , Infecciones por VIH/transmisión , Humanos , Maraviroc , Estructura Secundaria de Proteína , Compuestos de Amonio Cuaternario/farmacología , Receptores CCR5/metabolismo , Triazoles/farmacología , Replicación Viral/fisiologíaRESUMEN
The delivery of nucleic acid vaccine to stimulate host immune responses against Coronavirus disease 2019 shows promise. However, nucleic acid vaccines have drawbacks, including rapid clearance and poor cellular uptake, that limit their therapeutic potential. Microrobots can be engineered to sustain vaccine release and further control the interactions with immune cells that are vital for robust vaccination. Here, the 3D fabrication of biocompatible and biodegradable microrobots via the two-photon polymerization of gelatin methacryloyl (GelMA) and their proof-of-concept application for DNA vaccine delivery is reported. Programmed degradation and drug release by varying the local exposure dose in 3D laser lithography and further functionalized the GelMA microspheres with polyethyleneimine for DNA vaccine delivery to dendritic cell and primary cells is demonstrated. In mice, the DNA vaccine delivered by functionalized microspheres elicited fast, enhanced, and durable antigen expression, which may lead to prolonged protection. Furthermore, we demonstrate the maneuverability of microrobots by fabricating GelMA microspheres on magnetic skeletons. In conclusion, GelMA microrobots may provide an efficient vaccination strategy by controlling the expression duration of DNA vaccines.
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COVID-19 , Vacunas de ADN , Animales , Ratones , Sistemas de Liberación de Medicamentos , Gelatina , Vacunación , HidrogelesRESUMEN
The potency of cancer vaccines is often compromised by a variety of immunoinhibitory mechanisms, including stimulation of the programmed cell death protein 1 (PD-1)/programmed death ligand 1 (PD-L1) immune checkpoint pathway. Here, to overcome inhibition, we determined the potential of recombinant adeno-associated virus (rAAV)-vectored, PD1-based vaccination in the tumor microenvironment (TME) to activate antigen-specific T cell responses in the immune-competent murine mesothelioma model. We found that our rAAV-soluble PD1 (sPD1)-TWIST1 vaccine elicited and maintained TWIST1-specific cytotoxic T lymphocyte (CTL) responses and the PD-1 blocker systemically against lethal mesothelioma challenge after intramuscular injection, which was more effective than rAAV-TWIST1 or rAAV-sPD1 alone. More importantly, intratumoral injection of rAAV-sPD1-TWIST1 significantly enhanced immune surveillance by inducing TWIST1-specific CTL responses against vaccine-encoded TWIST1 and bystander gp70-AH1 epitopes, increasing CTL infiltration into the TME and decreasing tumor-associated immunosuppression, leading to complete elimination of established mesothelioma in 5 of 8 tumor-bearing mice. In addition, direct oncosuppression synergized with recruitment of T cells after localized rAAV-sPD1-TWIST1 treatment in a humanized mouse model to inhibit growth of REN human mesothelioma. Our results warrant clinical development of the rAAV-sPD1-TWIST1 vaccine to enhance immunotherapy against a wide range of TWIST1-expressing tumors.
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Effective immunotherapy treats cancers by eradicating tumourigenic cells by activated tumour antigen-specific and bystander CD8+ T-cells. However, T-cells can gradually lose cytotoxicity in the tumour microenvironment, known as exhaustion. Recently, DNA methylation, histone modification, and chromatin architecture have provided novel insights into epigenetic regulations of T-cell differentiation/exhaustion, thereby controlling the translational potential of the T-cells. Thus, developing strategies to govern epigenetic switches of T-cells dynamically is critical to maintaining the effector function of antigen-specific T-cells. In this mini-review, we 1) describe the correlation between epigenetic states and T cell phenotypes; 2) discuss the enzymatic factors and intracellular/extracellular microRNA imprinting T-cell epigenomes that drive T-cell exhaustion; 3) highlight recent advances in epigenetic interventions to rescue CD8+ T-cell functions from exhaustion. Finally, we express our perspective that regulating the interplay between epigenetic changes and transcriptional programs provides translational implications of current immunotherapy for cancer treatments.
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Insufficient T cell infiltration into noninflamed tumors, such as hepatocellular carcinoma (HCC), restricts the effectiveness of immune-checkpoint blockade (ICB) for a subset of patients. Epigenetic therapy provides further opportunities to rewire cancer-associated transcriptional programs, but whether and how selective epigenetic inhibition counteracts the immune-excluded phenotype remain incompletely defined. Here, we showed that pharmacological inhibition of histone deacetylase 8 (HDAC8), a histone H3 lysine 27 (H3K27)-specific isozyme overexpressed in a variety of human cancers, thwarts HCC tumorigenicity in a T cell-dependent manner. The tumor-suppressive effect of selective HDAC8 inhibition was abrogated by CD8+ T cell depletion or regulatory T cell adoptive transfer. Chromatin profiling of human HDAC8-expressing HCCs revealed genome-wide H3K27 deacetylation in 1251 silenced enhancer-target gene pairs that are enriched in metabolic and immune regulators. Mechanistically, down-regulation of HDAC8 increased global and enhancer acetylation of H3K27 to reactivate production of T cell-trafficking chemokines by HCC cells, thus relieving T cell exclusion in both immunodeficient and humanized mouse models. In an HCC preclinical model, selective HDAC8 inhibition increased tumor-infiltrating CD8+ T cells and potentiated eradication of established hepatomas by anti-PD-L1 therapy without evidence of toxicity. Mice treated with HDAC8 and PD-L1 coblockade were protected against subsequent tumor rechallenge as a result of the induction of memory T cells and remained tumor-free for greater than 15 months. Collectively, our study demonstrates that selective HDAC8 inhibition elicits effective and durable responses to ICB by co-opting adaptive immunity through enhancer reprogramming.
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Carcinoma Hepatocelular , Inhibidores de Histona Desacetilasas , Inhibidores de Puntos de Control Inmunológico , Neoplasias Hepáticas , Animales , Linfocitos T CD8-positivos , Carcinoma Hepatocelular/tratamiento farmacológico , Carcinoma Hepatocelular/inmunología , Línea Celular Tumoral , Inhibidores de Histona Desacetilasas/farmacología , Histona Desacetilasas , Humanos , Inhibidores de Puntos de Control Inmunológico/farmacología , Neoplasias Hepáticas/tratamiento farmacológico , Neoplasias Hepáticas/inmunología , Ratones , Proteínas RepresorasRESUMEN
We synthesized a series of acylhydrazone compounds bearing naturally occurring amino acids' side chains as HIV assembly inhibitors. Biological evaluation indicated that the compounds had anti-SIV and capsid assembly inhibitory activities. The structure-activity relationship (SAR) study showed that compounds bearing proper aromatic side chains had potential antiviral activities. The molecular modeling experiments revealed the molecular mechanism that they could bind to CA in the same manner as CAP-1 and occupy two more grooves.
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Antivirales/farmacología , VIH-1/efectos de los fármacos , Hidrazonas/farmacología , Antivirales/síntesis química , Antivirales/química , Cristalografía por Rayos X , Hidrazonas/síntesis química , Hidrazonas/química , Pruebas de Sensibilidad Microbiana , Modelos Moleculares , Relación Estructura-ActividadRESUMEN
Checkpoint immunotherapy is a major breakthrough for cancer treatment, yet its efficacy is often limited against many types of malignancies, including malignant mesothelioma. Considering that the immunotherapeutic efficacy depends on immunosurveillance, we sought to develop an active immunization method to break immune tolerance to tumor self-antigen. Here, we demonstrated that TWIST1, the basic helix-loop-helix transcription factor, was associated with human mesothelioma tumorigenesis and required for the invasion and metastasis of mesothelioma in the immune-competent murine AB1 model. When conventional TWIST1 vaccines were not effective in vivo, programmed cell death protein 1 (PD1)-based vaccination provided prophylactic control by inducing long-lasting TWIST1-specific T cell responses against both subcutaneous and metastatic mesothelioma lethal challenges. Furthermore, while CTLA-4 blockade alone did not show any immunotherapeutic efficacy against established mesothelioma, its combination with PD1-based vaccination resulted in 60% complete remission. Mechanistically, these functional T cells recognized a novel highly conserved immunodominant TWIST1 epitope, exhibited cytotoxic activity and long-term memory, and led to durable tumor regression and survival benefit against established AB1 mesothelioma and 4T1 breast cancer. We concluded that PD1-based vaccination controls mesothelioma by breaking immune tolerance to the tumor self-antigen TWIST1. Our results warrant clinical development of the PD1-based vaccination to enhance immunotherapy against a wide range of TWIST1-expressing tumors.
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BACKGROUND: Memory CD8+T cell responses play an essential role in protection against persistent infection. However, HIV-1 evades vaccine-induced memory CD8+T cell response by mechanisms that are not fully understood. METHODS: We analyzed the temporal dynamics of CD8+T cell recall activity and function during EcoHIV infection in a potent PD1-based vaccine immunized immunocompetent mice. FINDINGS: Upon intraperitoneal EcoHIV infection, high levels of HIV-1 GAG-specific CD8+T lymphocytes recall response reduced EcoHIV-infected cells significantly. However, this protective effect diminished quickly after seven days, followed by a rapid reduction of GAG-specific CD8+T cell number and activity, and viral persistence. Mechanistically, EcoHIV activated dendritic cells (DCs) and myeloid cells. Myeloid cells were infected and rapidly expanded, exhibiting elevated PD-L1/-L2 expression and T cell suppressive function before day 7, and were resistant to CD8+T cell-mediated apoptosis. Depletion of myeloid-derived suppressor cells (MDSCs) reduced EcoHIV infection and boosted T cell responses. INTERPRETATION: This study provides an overview of the temporal interplay of persistent virus, DCs, MDSCs and antigen-specific CD8+T cells during acute infection. We identify MDSCs as critical gatekeepers that restrain antiviral T cell memory responses, and highlight MDSCs as an important target for developing effective vaccines against chronic human infections. FUNDING: Hong Kong Research Grant Council (T11-709/18-N, HKU5/CRF/13G), General Research Fund (17122915 and 17114114), Hong Kong Health and Medical Research Fund (11100752, 14130582, 16150662), Grant RGC-ANR A-HKU709/14, the San-Ming Project of Medicine (SZSM201512029), University Development Fund of the University of Hong Kong and Li Ka Shing Faculty of Medicine Matching Fund to HKU AIDS Institute.
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Linfocitos T CD8-positivos/inmunología , Interacciones Huésped-Patógeno/inmunología , Memoria Inmunológica , Infecciones por Lentivirus/inmunología , Infecciones por Lentivirus/virología , Lentivirus/inmunología , Células Supresoras de Origen Mieloide/inmunología , Animales , Presentación de Antígeno/inmunología , Células Presentadoras de Antígenos/inmunología , Células Presentadoras de Antígenos/metabolismo , Biomarcadores , Linfocitos T CD8-positivos/metabolismo , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Modelos Animales de Enfermedad , Femenino , Infecciones por VIH/inmunología , Infecciones por VIH/metabolismo , Infecciones por VIH/virología , VIH-1/genética , VIH-1/inmunología , Humanos , Inmunocompetencia , Inmunomodulación , Lentivirus/genética , Infecciones por Lentivirus/metabolismo , Activación de Linfocitos/inmunología , Depleción Linfocítica , Ratones , Ratones Transgénicos , Células Supresoras de Origen Mieloide/metabolismo , Carga Viral , Vacunas Virales/inmunologíaRESUMEN
HIV-1 capsid protein (CA) plays important roles in the viral replication cycle. A number of acylhydrazone derivatives that act as inhibitors of HIV-1 CA assembly, were designed and synthesized. The synthesized compounds were tested for their antiviral activities and cytotoxicities using CEM cells. Some derivatives also were assayed for their ability to inhibit HIV-1 CA assembly in vitro. Among them, compounds 14f and 14i display the most promising potency with EC(50) values of 0.21 and 0.17 microM respectively.
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Fármacos Anti-VIH/síntesis química , Antivirales/síntesis química , Proteínas de la Cápside/antagonistas & inhibidores , Proteínas de la Cápside/metabolismo , Sistemas de Liberación de Medicamentos/métodos , VIH-1/efectos de los fármacos , Hidrazonas/síntesis química , Fármacos Anti-VIH/farmacología , Antivirales/farmacología , Sitios de Unión/efectos de los fármacos , Sitios de Unión/fisiología , Proteínas de la Cápside/fisiología , Línea Celular Tumoral , VIH-1/fisiología , Humanos , Hidrazonas/farmacologíaRESUMEN
Thirty-two quinoline derivatives were designed and synthesized as HIV-1 Tat-TAR interaction inhibitors. All the compounds showed high antiviral activities in inhibiting the formation of SIV-induced syncytium in CEM174 cells. Nine of them with low cytotoxicities were evaluated by Tat dependent HIV-1 LTR-driven CAT gene expression colorimetric enzyme assay in human 293T cells, indicating effective inhibitory activities of blocking the Tat-TAR interaction. Molecular modeling experiments indicated that these compounds may inhibit Tat-TAR interaction by binding to Tat protein instead of TAR RNA.
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Fármacos Anti-VIH/síntesis química , Fármacos Anti-VIH/farmacología , Duplicado del Terminal Largo de VIH/efectos de los fármacos , VIH-1/efectos de los fármacos , Quinolinas/síntesis química , Quinolinas/farmacología , Productos del Gen tat del Virus de la Inmunodeficiencia Humana/antagonistas & inhibidores , Fármacos Anti-VIH/química , Línea Celular , Diseño de Fármacos , Humanos , Quinolinas/química , Relación Estructura-Actividad , Productos del Gen tat del Virus de la Inmunodeficiencia Humana/metabolismoRESUMEN
HIV-1 assembly and disassembly (uncoating) processes are critical for the HIV-1 replication. HIV-1 capsid (CA) and human cyclophilin A (CypA) play essential roles in these processes. We designed and synthesized a series of thiourea compounds as HIV-1 assembly and disassembly dual inhibitors targeting both HIV-1 CA protein and human CypA. The SIV-induced syncytium antiviral evaluation indicated that all of the inhibitors displayed antiviral activities in SIV-infected CEM cells at the concentration of 0.6-15.8 microM for 50% of maximum effective rate. Their abilities to bind CA and CypA were determined by ultraviolet spectroscopic analysis, fluorescence binding affinity and PPIase inhibition assay. Assembly studies in vitro demonstrated that the compounds could potently disrupt CA assembly with a dose-dependent manner. All of these molecules could bind CypA with binding affinities (Kd values) of 51.0-512.8 microM. Fifteen of the CypA binding compounds showed potent PPIase inhibitory activities (IC(50) values<1 microM) while they could not bind either to HIV-1 Protease or to HIV-1 Integrase in the enzyme assays. These results suggested that 15 compounds could block HIV-1 replication by inhibiting the PPIase activity of CypA to interfere with capsid disassembly and disrupting CA assembly.
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Fármacos Anti-VIH/farmacología , Proteínas de la Cápside/metabolismo , Ciclofilina A/metabolismo , VIH-1/fisiología , Fármacos Anti-VIH/química , Cápside , Ciclofilina A/química , Diseño de Fármacos , Farmacorresistencia Viral , Humanos , Modelos Moleculares , Ensamble de Virus/efectos de los fármacos , Replicación Viral/efectos de los fármacosRESUMEN
Antitumor cytotoxic T lymphocytes (CTLs) are essential for immune surveillance, yet the blockade of eliciting such CTLs during oncolytic virotherapy remains incompletely understood. Here, we show that oncolysis of mesothelioma by modified vaccinia Tiantan (MVTT) induces damage-associated molecular patterns exposure. Although MVTT leads to regression of established mesothelioma dose-dependently, antitumor CTLs are rarely induced. Mechanistically, MVTT virotherapy generates C-X-C chemokines that recruit CXCR2-expressing polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) into tumor microenvironment, where they suppress dendritic cells (DCs) by producing IL-10 and halt CTL responses. During the virotherapy, however, depletion of PMN-MDSCs but not of monocytic (M)-MDSCs results in the induction of potent antitumor CTLs that not only eradicate established mesothelioma but also prevent the second tumor challenge. Our findings suggest that vaccinia virotherapy may combine strategies that prevent the chemotactic recruitment of PMN-MDSCs, block their suppression on DCs or deplete PMN-MDSCs in order to induce potent CTLs for tumor eradication.
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Newly emerging viruses, such as severe acute respiratory syndrome coronavirus (SARS-CoV), Middle Eastern respiratory syndrome CoVs (MERS-CoV), and H7N9, cause fatal acute lung injury (ALI) by driving hypercytokinemia and aggressive inflammation through mechanisms that remain elusive. In SARS-CoV/macaque models, we determined that anti-spike IgG (S-IgG), in productively infected lungs, causes severe ALI by skewing inflammation-resolving response. Alveolar macrophages underwent functional polarization in acutely infected macaques, demonstrating simultaneously both proinflammatory and wound-healing characteristics. The presence of S-IgG prior to viral clearance, however, abrogated wound-healing responses and promoted MCP1 and IL-8 production and proinflammatory monocyte/macrophage recruitment and accumulation. Critically, patients who eventually died of SARS (hereafter referred to as deceased patients) displayed similarly accumulated pulmonary proinflammatory, absence of wound-healing macrophages, and faster neutralizing antibody responses. Their sera enhanced SARS-CoV-induced MCP1 and IL-8 production by human monocyte-derived wound-healing macrophages, whereas blockade of FcγR reduced such effects. Our findings reveal a mechanism responsible for virus-mediated ALI, define a pathological consequence of viral specific antibody response, and provide a potential target for treatment of SARS-CoV or other virus-mediated lung injury.
Asunto(s)
Lesión Pulmonar Aguda/inmunología , Anticuerpos Antivirales/inmunología , Inmunoglobulina G/inmunología , Síndrome Respiratorio Agudo Grave/inmunología , Coronavirus Relacionado al Síndrome Respiratorio Agudo Severo/inmunología , Glicoproteína de la Espiga del Coronavirus/inmunología , Lesión Pulmonar Aguda/sangre , Lesión Pulmonar Aguda/virología , Adulto , Anciano , Anciano de 80 o más Años , Animales , Anticuerpos Neutralizantes/administración & dosificación , Anticuerpos Neutralizantes/sangre , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/administración & dosificación , Anticuerpos Antivirales/sangre , Línea Celular , Modelos Animales de Enfermedad , Femenino , Humanos , Inmunización Pasiva/métodos , Inmunoglobulina G/administración & dosificación , Inmunoglobulina G/sangre , Pulmón/inmunología , Pulmón/patología , Pulmón/virología , Macaca mulatta , Macrófagos Alveolares/inmunología , Macrófagos Alveolares/metabolismo , Masculino , Persona de Mediana Edad , Coronavirus Relacionado al Síndrome Respiratorio Agudo Severo/patogenicidad , Síndrome Respiratorio Agudo Grave/sangre , Síndrome Respiratorio Agudo Grave/virología , Glicoproteína de la Espiga del Coronavirus/genética , Virus Vaccinia/genética , Virus Vaccinia/inmunología , Vacunas Virales/administración & dosificación , Vacunas Virales/inmunología , Adulto JovenRESUMEN
DNA vaccines have been extensively studied as preventative and therapeutic interventions for various infectious diseases such as tuberculosis, HIV/AIDS and influenza. Despite promising progresses made, improving the immunogenicity of DNA vaccine remains a technical challenge for clinical development. In this study, we investigated a tuberculosis DNA vaccine BERopt, which contained a codon-optimized fusion immunogen Ag85B-ESAT-6-Rv2660c for enhanced mammalian cell expression and immunogenicity. BERopt immunization through in vivo electroporation in BALB/c mice induced surprisingly high frequencies of Ag85B tetramer+ CD8+ T cells in peripheral blood and IFN-γ-secreting CD8+ T cells in splenocytes. Meanwhile, the BERopt vaccine-induced long-lasting T cell immunity protected BALB/c mice from high dose viral challenge using a modified vaccinia virus Tiantan strain expressing mature Ag85B protein (MVTT-m85B) and the virulent M. tb H37Rv aerosol challenge. Since the BERopt DNA vaccine does not induce anti-vector immunity, the strong immunogenicity and protective efficacy of this novel DNA vaccine warrant its future development for M. tb prevention and immunotherapy to alleviate the global TB burden.
Asunto(s)
Aciltransferasas/administración & dosificación , Antígenos Bacterianos/administración & dosificación , Proteínas Bacterianas/administración & dosificación , Electroquimioterapia/métodos , Inmunogenicidad Vacunal , Mycobacterium tuberculosis/patogenicidad , Vacunas contra la Tuberculosis/administración & dosificación , Tuberculosis Pulmonar/prevención & control , Aciltransferasas/genética , Aciltransferasas/inmunología , Aerosoles , Animales , Antígenos Bacterianos/genética , Antígenos Bacterianos/inmunología , Proteínas Bacterianas/genética , Proteínas Bacterianas/inmunología , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/microbiología , Células Cultivadas , Codón , Modelos Animales de Enfermedad , Femenino , Inmunización , Exposición por Inhalación , Interferón gamma/inmunología , Ratones Endogámicos BALB C , Mycobacterium tuberculosis/genética , Mycobacterium tuberculosis/inmunología , Bazo/inmunología , Bazo/microbiología , Factores de Tiempo , Vacunas contra la Tuberculosis/genética , Vacunas contra la Tuberculosis/inmunología , Tuberculosis Pulmonar/inmunología , Tuberculosis Pulmonar/microbiología , Vacunas de ADN/administración & dosificación , Virus Vaccinia/genética , Virus Vaccinia/inmunologíaRESUMEN
A key focus in cancer immunotherapy is to investigate the mechanism of efficacious vaccine responses. Using HIV-1 GAG-p24 in a model PD1-based DNA vaccine, we recently reported that vaccine-elicited CD8+ T cells conferred complete prevention and therapeutic cure of AB1-GAG malignant mesothelioma in immunocompetent BALB/c mice. Here, we further investigated the efficacy and correlation of protection on the model vaccine-mediated antigen spreading against wild-type AB1 (WT-AB1) mesothelioma. We found that this vaccine was able to protect mice completely from three consecutive lethal challenges of AB1-GAG mesothelioma. Through antigen spreading these animals also developed tumor-specific cytotoxic CD8+ T cells, but neither CD4+ T cells nor antibodies, rejecting WT-AB1 mesothelioma. A majority of these protected mice (90%) were also completely protected against the lethal WT-AB1 challenge. Adoptive cell transfer experiments further demonstrated that antigen spreading-induced CD8+ T cells conferred efficacious therapeutic effects against established WT-AB1 mesothelioma and prevented the increase of exhausted PD-1+ and Tim-3+ CD8+ T cells. A significant inverse correlation was found between the frequency of functional PD1-Tim3- CD8+ T cells and that of MDSCs or tumor mass in vivo. Mechanistically, we found that WT-AB1 mesothelioma induced predominantly polymorphonuclear (PMN) MDSCs in vivo. In co-cultures with efficacious CD8+ T cells, a significant number of PMN-MDSCs underwent apoptosis in a dose-dependent way. Our findings indicate that efficacious CD8+ T cells capable of eliminating both tumor cells and MDSCs are likely necessary for fighting wild-type malignant mesothelioma.