Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 473
Filtrar
1.
PLoS Pathog ; 16(7): e1008795, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32716975

RESUMO

HSV-1 causes 50% of first-time genital herpes infections in resource-rich countries and affects 190 million people worldwide. A prophylactic herpes vaccine is needed to protect against genital infections by both HSV-1 and HSV-2. Previously our laboratory developed a trivalent vaccine that targets glycoproteins C, D, and E present on the HSV-2 virion. We reported that this vaccine protects animals from genital disease and recurrent virus shedding following lethal HSV-2 challenge. Importantly the vaccine also generates cross-reactive antibodies that neutralize HSV-1, suggesting it may provide protection against HSV-1 infection. Here we compared the efficacy of this vaccine delivered as protein or nucleoside-modified mRNA immunogens against vaginal HSV-1 infection in mice. Both the protein and mRNA vaccines protected mice from HSV-1 disease; however, the mRNA vaccine provided better protection as measured by lower vaginal virus titers post-infection. In a second experiment, we compared protection provided by the mRNA vaccine against intravaginal challenge with HSV-1 or HSV-2. Vaccinated mice were totally protected against death, genital disease and infection of dorsal root ganglia caused by both viruses, but somewhat better protected against vaginal titers after HSV-2 infection. Overall, in the two experiments, the mRNA vaccine prevented death and genital disease in 54/54 (100%) mice infected with HSV-1 and 20/20 (100%) with HSV-2, and prevented HSV DNA from reaching the dorsal root ganglia, the site of virus latency, in 29/30 (97%) mice infected with HSV-1 and 10/10 (100%) with HSV-2. We consider the HSV-2 trivalent mRNA vaccine to be a promising candidate for clinical trials for prevention of both HSV-1 and HSV-2 genital herpes.


Assuntos
Proteção Cruzada/imunologia , Herpes Genital , Herpesvirus Humano 1/imunologia , Herpesvirus Humano 2/imunologia , Vacinas contra Herpesvirus/imunologia , Latência Viral/imunologia , Animais , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Feminino , Herpes Genital/virologia , Camundongos , Camundongos Endogâmicos BALB C , RNA Mensageiro , Proteínas do Envelope Viral/imunologia
2.
PLoS One ; 15(3): e0228163, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32130229

RESUMO

Anti-retroviral therapy (ART) has been highly successful in controlling HIV replication, reducing viral burden, and preventing both progression to AIDS and viral transmission. Yet, ART alone cannot cure the infection. Even after years of successful therapy, ART withdrawal leads inevitably to viral rebound within a few weeks or months. Our hypothesis: effective therapy must control both the replicating virus pool and the reactivatable latent viral reservoir. To do this, we have combined ART and immunotherapy to attack both viral pools simultaneously. The vaccine regimen consisted of DNA vaccine expressing SIV Gag, followed by a boost with live attenuated rubella/gag vectors. The vectors grow well in rhesus macaques, and they are potent immunogens when used in a prime and boost strategy. We infected rhesus macaques by high dose mucosal challenge with virulent SIVmac251 and waited three days to allow viral dissemination and establishment of a reactivatable viral reservoir before starting ART. While on ART, the control group received control DNA and empty rubella vaccine, while the immunotherapy group received DNA/gag prime, followed by boosts with rubella vectors expressing SIV gag over 27 weeks. Both groups had a vaccine "take" to rubella, and the vaccine group developed antibodies and T cells specific for Gag. Five weeks after the last immunization, we stopped ART and monitored virus rebound. All four control animals eventually had a viral rebound, and two were euthanized for AIDS. One control macaque did not rebound until 2 years after ART release. In contrast, there was only one viral rebound in the vaccine group. Three out of four vaccinees had no viral rebound, even after CD8 depletion, and they remain in drug-free viral remission more than 2.5 years later. The strategy of early ART combined with immunotherapy can produce a sustained SIV remission in macaques and may be relevant for immunotherapy of HIV in humans.


Assuntos
Síndrome de Imunodeficiência Adquirida/terapia , Fármacos Anti-HIV/uso terapêutico , Vacinas contra a SAIDS/administração & dosagem , Síndrome de Imunodeficiência Adquirida dos Símios/terapia , Vírus da Imunodeficiência Símia/imunologia , Síndrome de Imunodeficiência Adquirida/sangue , Síndrome de Imunodeficiência Adquirida/imunologia , Síndrome de Imunodeficiência Adquirida/virologia , Animais , Terapia Combinada/métodos , Modelos Animais de Doenças , Esquema de Medicação , Quimioterapia Combinada/métodos , Produtos do Gene gag/genética , Produtos do Gene gag/imunologia , Vetores Genéticos/administração & dosagem , Vetores Genéticos/genética , Humanos , Macaca mulatta , Plasmídeos/administração & dosagem , Plasmídeos/genética , Vírus da Rubéola/imunologia , Vacinas contra a SAIDS/genética , Síndrome de Imunodeficiência Adquirida dos Símios/sangue , Síndrome de Imunodeficiência Adquirida dos Símios/imunologia , Síndrome de Imunodeficiência Adquirida dos Símios/virologia , Vírus da Imunodeficiência Símia/genética , Vírus da Imunodeficiência Símia/isolamento & purificação , Fatores de Tempo , Resultado do Tratamento , Vacinas Atenuadas/administração & dosagem , Vacinas de DNA/administração & dosagem , Vacinas de DNA/genética , Latência Viral/efeitos dos fármacos , Latência Viral/imunologia , Replicação Viral/efeitos dos fármacos , Replicação Viral/imunologia
3.
PLoS Pathog ; 16(3): e1008442, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32196533

RESUMO

Latency reversal agents (LRA) variably induce HIV re-expression in CD4 T cells but reservoirs are not cleared. Whether HIV epitope presentation is similar between latency reversal and initial infection of CD4 T cells is unknown yet crucial to define immune responses able to detect HIV-infected CD4 T cells after latency reversal. HIV peptides displayed by MHC comes from the intracellular degradation of proteins by proteasomes and post-proteasomal peptidases but the impact of LRAs on antigen processing is not known. Here we show that HDAC inhibitors (HDCAi) reduced cytosolic proteolytic activities while PKC agonists (PKCa) increased them to a lesser extent than that induced by TCR activation. During the cytosolic degradation of long HIV peptides in LRA-treated CD4 T cells extracts, HDACi and PKCa modulated degradation patterns of peptides and altered the production of HIV epitopes in often opposite ways. Beyond known HIV epitopes, HDACi narrowed the coverage of HIV antigenic fragments by 8-11aa degradation peptides while PKCa broadened it. LRAs altered HIV infection kinetics and modulated CD8 T cell activation in an epitope- and time-dependent manner. Interestingly the efficiency of endogenous epitope processing and presentation to CD8 T cells was increased by PKCa Ingenol at early time points despite low levels of antigens. LRA-induced modulations of antigen processing should be considered and exploited to enhance and broaden HIV peptide presentation by CD4 T cells and to improve immune recognition after latency reversal. This property of LRAs, if confirmed with other antigens, might be exploited to improve immune detection of diseased cells beyond HIV.


Assuntos
Apresentação do Antígeno , Antígenos Virais/imunologia , Linfócitos T CD8-Positivos/imunologia , Infecções por HIV/imunologia , HIV-1/fisiologia , Inibidores de Histona Desacetilases/farmacologia , Proteína Quinase C/antagonistas & inibidores , Latência Viral/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/patologia , Linfócitos T CD8-Positivos/patologia , Infecções por HIV/tratamento farmacológico , Infecções por HIV/patologia , Humanos , Proteína Quinase C/imunologia , Latência Viral/imunologia
4.
PLoS Pathog ; 16(2): e1008151, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-32109259

RESUMO

HIV latency is the major barrier to a cure for people living with HIV (PLWH) on antiretroviral therapy (ART) because the virus persists in long-lived non-proliferating and proliferating latently infected CD4+ T cells. Latently infected CD4+ T cells do not express viral proteins and are therefore not visible to immune mediated clearance. Therefore, identifying interventions that can reverse latency and also enhance immune mediated clearance is of high interest. Interferons (IFNs) have multiple immune enhancing effects and can inhibit HIV replication in activated CD4+ T cells. However, the effects of IFNs on the establishment and reversal of HIV latency is not understood. Using an in vitro model of latency, we demonstrated that plasmacytoid dendritic cells (pDC) inhibit the establishment of HIV latency through secretion of type I IFNα, IFNß and IFNω but not IFNε or type III IFNλ1 and IFNλ3. However, once latency was established, IFNα but no other IFNs were able to efficiently reverse latency in both an in vitro model of latency and CD4+ T cells collected from PLWH on suppressive ART. Binding of IFNα to its receptor expressed on primary CD4+ T cells did not induce activation of the canonical or non-canonical NFκB pathway but did induce phosphorylation of STAT1, 3 and 5 proteins. STAT5 has been previously demonstrated to bind to the HIV long terminal repeat and activate HIV transcription. We demonstrate diverse effects of interferons on HIV latency with type I IFNα; inhibiting the establishment of latency but also reversing HIV latency once latency is established.


Assuntos
Linfócitos T CD4-Positivos , Repetição Terminal Longa de HIV/imunologia , HIV-1/fisiologia , Interferon-alfa/imunologia , Transcrição Genética/imunologia , Latência Viral/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/patologia , Linfócitos T CD4-Positivos/virologia , Células Dendríticas/imunologia , Células Dendríticas/patologia , Células Dendríticas/virologia , Células HEK293 , Humanos , NF-kappa B/imunologia , Fatores de Transcrição STAT/imunologia
5.
J Immunol ; 204(5): 1242-1254, 2020 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-31988180

RESUMO

In people living with HIV on antiretroviral therapy, HIV latency is the major barrier to a cure. HIV persists preferentially in CD4+ T cells expressing multiple immune checkpoint (IC) molecules, including programmed death (PD)-1, T cell Ig and mucin domain-containing protein 3 (TIM-3), lymphocyte associated gene 3 (LAG-3), and T cell immunoreceptor with Ig and ITIM domains (TIGIT). We aimed to determine whether these and other IC molecules have a functional role in maintaining HIV latency and whether blocking IC molecules with Abs reverses HIV latency. Using an in vitro model that establishes latency in both nonproliferating and proliferating human CD4+ T cells, we show that proliferating cells express multiple IC molecules at high levels. Latent infection was enriched in proliferating cells expressing PD-1. In contrast, nonproliferating cells expressed IC molecules at significantly lower levels, but latent infection was enriched in cells expressing PD-1, TIM-3, CTL-associated protein 4 (CTLA-4), or B and T lymphocyte attenuator (BTLA). In the presence of an additional T cell-activating stimulus, staphylococcal enterotoxin B, Abs to CTLA-4 and PD-1 reversed HIV latency in proliferating and nonproliferating CD4+ T cells, respectively. In the absence of staphylococcal enterotoxin B, only the combination of Abs to PD-1, CTLA-4, TIM-3, and TIGIT reversed latency. The potency of latency reversal was significantly higher following combination IC blockade compared with other latency-reversing agents, including vorinostat and bryostatin. Combination IC blockade should be further explored as a strategy to reverse HIV latency.


Assuntos
Anticorpos Monoclonais Murinos/farmacologia , Linfócitos T CD4-Positivos , Proliferação de Células/efeitos dos fármacos , Enterotoxinas/farmacologia , HIV-1/fisiologia , Modelos Imunológicos , Latência Viral , Antígenos CD/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/patologia , Linfócitos T CD4-Positivos/virologia , Feminino , Células HEK293 , Receptor Celular 2 do Vírus da Hepatite A/antagonistas & inibidores , Receptor Celular 2 do Vírus da Hepatite A/imunologia , Humanos , Ativação Linfocitária/efeitos dos fármacos , Masculino , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/imunologia , Receptores Imunológicos/antagonistas & inibidores , Receptores Imunológicos/imunologia , Latência Viral/efeitos dos fármacos , Latência Viral/imunologia
6.
Arch Virol ; 165(2): 321-330, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31828511

RESUMO

Persistence of human immunodeficiency virus 1 (HIV-1) latency and residual immune activation remain major barriers to treatment in patients receiving highly active antiretroviral therapy (HAART). In the present study, we investigated the molecular mechanisms of persistent HIV infection and residual immune activation in HAART-treated patients. We showed that the expression level of B-cell CLL/lymphoma 11B (BCL11B) was significantly increased in CD4+T cells from HIV-infected patients undergoing HAART, and this was accompanied by increased expression of BCL11B-associated chromatin modifiers and inflammatory factors in comparison to healthy controls and untreated patients with HIV. In vitro assays showed that BCL11B significantly inhibited HIV-1 long terminal repeat (LTR)-mediated transcription. Knockdown of BCL11B resulted in the activation of HIV latent cells, and dissociation of BCL11B and its related chromatin remodeling factors from the HIV LTR. Our findings suggested that increased expression of BCL11B and its related chromatin modifiers contribute to HIV-1 transcriptional silencing, and alteration of BCL11B levels might lead to abnormal transcription and inflammation.


Assuntos
Montagem e Desmontagem da Cromatina/genética , Infecções por HIV/genética , HIV-1/genética , Proteínas Repressoras/genética , Proteínas Repressoras/imunologia , Transcrição Genética/imunologia , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/imunologia , Adulto , Terapia Antirretroviral de Alta Atividade/métodos , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/virologia , Cromatina/genética , Cromatina/virologia , Montagem e Desmontagem da Cromatina/efeitos dos fármacos , Montagem e Desmontagem da Cromatina/imunologia , Feminino , Infecções por HIV/tratamento farmacológico , Infecções por HIV/imunologia , Infecções por HIV/virologia , Repetição Terminal Longa de HIV/genética , Repetição Terminal Longa de HIV/imunologia , HIV-1/efeitos dos fármacos , HIV-1/imunologia , Humanos , Masculino , Transcrição Genética/efeitos dos fármacos , Latência Viral/genética , Latência Viral/imunologia
7.
J Virol ; 94(6)2020 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-31852788

RESUMO

High rates of wild-type (WT) herpes simplex virus 1 (HSV-1) latency reactivation depend on the anti-apoptotic activities of latency-associated transcript (LAT). Replacing LAT with the baculovirus inhibitor of apoptosis protein (cpIAP) or cellular FLIP (FLICE-like inhibitory protein) gene restored the WT latency reactivation phenotype to that of a LAT-minus [LAT(-)] virus, while similar recombinant viruses expressing interleukin-4 (IL-4) or interferon gamma (IFN-γ) did not. However, HSV-1 recombinant virus expressing cpIAP did not restore all LAT functions. Recently, we reported that a similar recombinant virus expressing CD80 in place of LAT had higher latency reactivation than a LAT-null virus. The present study was designed to determine if this CD80-expressing recombinant virus can restore all LAT functions as observed with WT virus. Our results suggest that overexpression of CD80 fully rescues LAT function in latency reactivation, apoptosis, and immune exhaustion, suggesting that LAT and CD80 have multiple overlapping functions.IMPORTANCE Recurring ocular infections caused by HSV-1 can cause corneal scarring and blindness. A major function of the HSV-1 latency-associated transcript (LAT) is to establish high levels of latency and reactivation, thus contributing to the development of eye disease. Here, we show that the host CD80 T cell costimulatory molecule functions similarly to LAT and can restore the ability of LAT to establish latency, reactivation, and immune exhaustion as well as induce the expression of caspase 3, caspase 8, caspase 9, and Bcl2. Our results suggest that, in contrast to several other previously tested genes, CD80-expressing virus can completely compensate for all known and tested LAT functions.


Assuntos
Apoptose/imunologia , Antígeno B7-1/imunologia , Herpesvirus Humano 1/fisiologia , MicroRNAs/imunologia , RNA Viral/imunologia , Ativação Viral/imunologia , Latência Viral/imunologia , Animais , Apoptose/genética , Antígeno B7-1/genética , Camundongos , MicroRNAs/genética , RNA Viral/genética , Ativação Viral/genética , Latência Viral/genética
8.
mBio ; 10(6)2019 12 03.
Artigo em Inglês | MEDLINE | ID: mdl-31796538

RESUMO

Human cytomegalovirus (HCMV) latency is an active process which remodels the latently infected cell to optimize latent carriage and reactivation. This is achieved, in part, through the expression of viral genes, including the G-protein-coupled receptor US28. Here, we use an unbiased proteomic screen to assess changes in host proteins induced by US28, revealing that interferon-inducible genes are downregulated by US28. We validate that major histocompatibility complex (MHC) class II and two pyrin and HIN domain (PYHIN) proteins, myeloid cell nuclear differentiation antigen (MNDA) and IFI16, are downregulated during experimental latency in primary human CD14+ monocytes. We find that IFI16 is targeted rapidly during the establishment of latency in a US28-dependent manner but only in undifferentiated myeloid cells, a natural site of latent carriage. Finally, by overexpressing IFI16, we show that IFI16 can activate the viral major immediate early promoter and immediate early gene expression during latency via NF-κB, a function which explains why downregulation of IFI16 during latency is advantageous for the virus.IMPORTANCE Human cytomegalovirus (HCMV) is a ubiquitous herpesvirus which infects 50 to 100% of humans worldwide. HCMV causes a lifelong subclinical infection in immunocompetent individuals but is a serious cause of mortality and morbidity in the immunocompromised and neonates. In particular, reactivation of HCMV in the transplant setting is a major cause of transplant failure and related disease. Therefore, a molecular understanding of HCMV latency and reactivation could provide insights into potential ways to target the latent viral reservoir in at-risk patient populations.


Assuntos
Infecções por Citomegalovirus/genética , Citomegalovirus/imunologia , Interferons/genética , Latência Viral/genética , Diferenciação Celular/genética , Diferenciação Celular/imunologia , Linhagem Celular , Infecções por Citomegalovirus/imunologia , Infecções por Citomegalovirus/virologia , Regulação para Baixo/genética , Regulação para Baixo/imunologia , Expressão Gênica/genética , Expressão Gênica/imunologia , Regulação Viral da Expressão Gênica/genética , Regulação Viral da Expressão Gênica/imunologia , Células HEK293 , Humanos , Interferons/imunologia , Monócitos/imunologia , Monócitos/virologia , Células Mieloides/imunologia , Células Mieloides/virologia , NF-kappa B/genética , NF-kappa B/imunologia , Regiões Promotoras Genéticas/genética , Regiões Promotoras Genéticas/imunologia , Proteômica/métodos , Receptores Acoplados a Proteínas-G/imunologia , Células THP-1 , Proteínas Virais/genética , Proteínas Virais/imunologia , Ativação Viral/genética , Ativação Viral/imunologia , Latência Viral/imunologia
9.
PLoS Pathog ; 15(10): e1008074, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31609991

RESUMO

Studies have demonstrated that intensive ART alone is not capable of eradicating HIV-1, as the virus rebounds within a few weeks upon treatment interruption. Viral rebound may be induced from several cellular subsets; however, the majority of proviral DNA has been found in antigen experienced resting CD4+ T cells. To achieve a cure for HIV-1, eradication strategies depend upon both understanding mechanisms that drive HIV-1 persistence as well as sensitive assays to measure the frequency of infected cells after therapeutic interventions. Assays such as the quantitative viral outgrowth assay (QVOA) measure HIV-1 persistence during ART by ex vivo activation of resting CD4+ T cells to induce latency reversal; however, recent studies have shown that only a fraction of replication-competent viruses are inducible by primary mitogen stimulation. Previous studies have shown a correlation between the acquisition of effector memory phenotype and HIV-1 latency reversal in quiescent CD4+ T cell subsets that harbor the reservoir. Here, we apply our mechanistic understanding that differentiation into effector memory CD4+ T cells more effectively promotes HIV-1 latency reversal to significantly improve proviral measurements in the QVOA, termed differentiation QVOA (dQVOA), which reveals a significantly higher frequency of the inducible HIV-1 replication-competent reservoir in resting CD4+ T cells.


Assuntos
Linfócitos T CD4-Positivos/virologia , HIV-1/imunologia , HIV-1/fisiologia , Memória Imunológica/imunologia , Latência Viral/imunologia , Idoso , Antirretrovirais/uso terapêutico , Linfócitos T CD4-Positivos/imunologia , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Infecções por HIV/imunologia , HIV-1/crescimento & desenvolvimento , Humanos , Masculino , Pessoa de Meia-Idade , Provírus/crescimento & desenvolvimento , Carga Viral/efeitos dos fármacos , Latência Viral/efeitos dos fármacos , Replicação Viral/efeitos dos fármacos
10.
J Immunol ; 203(6): 1665-1674, 2019 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-31434708

RESUMO

EBV infects and immortalizes B cells in vitro and in vivo. It is the causative agent of most immune deficiency-related lymphoproliferative disorders and is associated with various lymphomas. EBV latency III-transformed B cells are known to express two immunosuppressive molecules, IL-10 and PD-L1, two characteristics of regulatory B cells (Bregs). In this study, we show that, in addition to secretion of the Breg immunosuppressive cytokines IL-10, IL-35, and TGF-ß1, EBV latency III-transformed B cells were able to repress proliferation of their autologous T cells preactivated by CD2, CD3, and CD28. This inhibitory effect was likely caused by CD4+ T cells because EBV latency III-transformed B cells induced a strong proliferation of isolated autologous CD8 T cells. Indeed, EBV was able to promote expansion of autologous FOXP3+ CD39high CTLA4+, Helios+, GITR+, LAG3+ CD4 T cells (i.e., regulatory T cells [Tregs]). Two types of Tregs were induced: unconventional CD25neg and conventional CD25pos Tregs. These Tregs expressed both the latency-associated peptide (LAP) and the PD-1 receptor, two markers of functional Tregs. Expansion of both Treg subtypes depended on PD-L1, whose expression was under the control of LMP1, the main EBV oncogene. These results demonstrate that, like Bregs, EBV latency III-transformed B cells exhibit strong immunoregulatory properties. These data provide clues to the understanding of how after EBV primo-infection, EBV-proliferating B cells can survive in an aggressive immunological environment and later emerge to give rise to EBV-associated B cell lymphomas such as in elderly patients.


Assuntos
Linfócitos B/imunologia , Antígeno B7-H1/imunologia , Herpesvirus Humano 4/imunologia , Linfócitos T Reguladores/imunologia , Latência Viral/imunologia , Antígenos CD/imunologia , Apirase/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Antígeno CTLA-4/imunologia , Linhagem Celular , Fatores de Transcrição Forkhead/imunologia , Proteína Relacionada a TNFR Induzida por Glucocorticoide/imunologia , Humanos , Subunidade alfa de Receptor de Interleucina-2/imunologia
11.
PLoS Pathog ; 15(8): e1007991, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31425551

RESUMO

Latency reversal agents (LRAs) have proven to induce HIV-1 transcription in vivo but are ineffective at decreasing the size of the latent reservoir in antiretroviral treated patients. The capacity of the LRAs to perturb the viral reservoir present in distinct subpopulations of cells is currently unknown. Here, using a new RNA FISH/flow ex vivo viral reactivation assay, we performed a comprehensive assessment of the viral reactivation capacity of different families of LRAs, and their combinations, in different CD4+ T cell subsets. We observed that a median of 16.28% of the whole HIV-reservoir induced HIV-1 transcripts after viral reactivation, but only 10.10% of these HIV-1 RNA+ cells produced the viral protein p24. Moreover, none of the LRAs were powerful enough to reactivate HIV-1 transcription in all CD4+ T cell subpopulations. For instance, the combination of Romidepsin and Ingenol was identified as the best combination of drugs at increasing the proportion of HIV-1 RNA+ cells, in most, but not all, CD4+ T cell subsets. Importantly, memory stem cells were identified as highly resistant to HIV-1 reactivation, and only the combination of Panobinostat and Bryostatin-1 significantly increased the number of cells transcribing HIV within this subset. Overall, our results validate the use of the RNA FISH/flow technique to assess the potency of LRAs among different CD4+ T cell subsets, manifest the intrinsic differences between cells that encompass the latent HIV reservoir, and highlight the difficulty to significantly impact the latent infection with the currently available drugs. Thus, our results have important implications for the rational design of therapies aimed at reversing HIV latency from diverse cellular reservoirs.


Assuntos
Fármacos Anti-HIV/farmacologia , Linfócitos T CD4-Positivos/imunologia , Infecções por HIV/imunologia , HIV-1/imunologia , Ativação Viral/imunologia , Latência Viral/imunologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/virologia , Depsipeptídeos/farmacologia , Diterpenos/farmacologia , Infecções por HIV/tratamento farmacológico , Infecções por HIV/virologia , HIV-1/efeitos dos fármacos , Humanos , Carga Viral , Ativação Viral/efeitos dos fármacos , Latência Viral/efeitos dos fármacos
12.
Int J Mol Sci ; 20(15)2019 Jul 24.
Artigo em Inglês | MEDLINE | ID: mdl-31344940

RESUMO

Human cytomegalovirus (HCMV) is ubiquitously prevalent. HCMV infection is typically asymptomatic and controlled by the immune system in healthy individuals, yet HCMV can be severely pathogenic for the fetus during pregnancy and in immunocompromised persons, such as transplant recipients or HIV infected patients. HCMV has co-evolved with the hosts, developed strategies to hide from immune effector cells and to successfully survive in the human organism. One strategy for evading or delaying the immune response is maintenance of the viral genome to establish the phase of latency. Furthermore, HCMV immune evasion involves the downregulation of human leukocyte antigens (HLA)-Ia molecules to hide infected cells from T-cell recognition. HCMV expresses several proteins that are described for downregulation of the HLA class I pathway via various mechanisms. Here, we review the wide range of immune evasion mechanisms of HCMV. Understanding the mechanisms of HCMV immune evasion will contribute to the development of new customized therapeutic strategies against the virus.


Assuntos
Infecções por Citomegalovirus/imunologia , Citomegalovirus/imunologia , Imunidade Celular/genética , Células Matadoras Naturais/imunologia , Citomegalovirus/patogenicidade , Infecções por Citomegalovirus/virologia , Genoma Viral/imunologia , Antígenos HLA/imunologia , Humanos , Interferon gama/genética , Interferon gama/imunologia , Linfócitos T/imunologia , Linfócitos T/virologia , Latência Viral/imunologia
13.
PLoS Comput Biol ; 15(7): e1007229, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31339888

RESUMO

Antiretroviral therapy (ART) effectively controls HIV infection, suppressing HIV viral loads. Suspension of therapy is followed by rebound of viral loads to high, pre-therapy levels. However, there is significant heterogeneity in speed of rebound, with some rebounds occurring within days, weeks, or sometimes years. We present a stochastic mathematical model to gain insight into these post-treatment dynamics, specifically characterizing the dynamics of short term viral rebounds (≤ 60 days). Li et al. (2016) report that the size of the expressed HIV reservoir, i.e., cell-associated HIV RNA levels, and drug regimen correlate with the time between ART suspension and viral rebound to detectable levels. We incorporate this information and viral rebound times to parametrize our model. We then investigate insights offered by our model into the underlying dynamics of the latent reservoir. In particular, we refine previous estimates of viral recrudescence after ART interruption by accounting for heterogeneity in infection rebound dynamics, and determine a recrudescence rate of once every 2-4 days. Our parametrized model can be used to aid in design of clinical trials to study viral dynamics following analytic treatment interruption. We show how to derive informative personalized testing frequencies from our model and offer a proof-of-concept example. Our results represent first steps towards a model that can make predictions on a person living with HIV (PLWH)'s rebound time distribution based on biomarkers, and help identify PLWH with long viral rebound delays.


Assuntos
Fármacos Anti-HIV/administração & dosagem , Infecções por HIV/tratamento farmacológico , Infecções por HIV/virologia , Modelos Biológicos , Algoritmos , Biomarcadores/sangue , Biologia Computacional , Infecções por HIV/imunologia , Humanos , Funções Verossimilhança , RNA Viral/sangue , Processos Estocásticos , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Linfócitos T/virologia , Fatores de Tempo , Carga Viral/efeitos dos fármacos , Viremia/tratamento farmacológico , Viremia/imunologia , Viremia/virologia , Latência Viral/efeitos dos fármacos , Latência Viral/imunologia , Suspensão de Tratamento
14.
Mol Immunol ; 111: 145-151, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31054408

RESUMO

Tuberculosis (TB) is a global epidemic with devastating consequences. Emerging evidence suggests that B-cells have the ability to modulate the immune response and understanding these roles during Mycobacterium tuberculosis (M.tb) infection can help to find new strategies to treat TB. The immune system of individuals with pulmonary TB form granulomas in the lung which controls the infection by inhibiting the M.tb growth and acts as a physical barrier. Thereafter, surviving M.tb become dormant and in most cases the host's immunity prevents TB reactivation. B-cells execute several immunological functions and are regarded as protective regulators of immune responses by antibody and cytokine production, as well as presenting antigen. Some of these B-cells, or regulatory B-cells, have been shown to express death-inducing ligands, such as Fas ligand (FasL). This expression and binding to the Fas receptor leads to apoptosis, a major immune regulation mechanism, in addition to the ability to induce T-cell tolerance. Here, I discuss the relevance of B-cells, in particular their non-humoral functions by addressing their regulatory properties during M.tb infection.


Assuntos
Linfócitos B/imunologia , Mycobacterium tuberculosis/imunologia , Tuberculose Pulmonar/imunologia , Latência Viral/imunologia , Apoptose/imunologia , Proteína Ligante Fas/imunologia , Ativação Linfocitária/imunologia , Receptor fas/imunologia
15.
PLoS Pathog ; 15(5): e1007802, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31116788

RESUMO

A major barrier to curing HIV-1 is the long-lived latent reservoir that supports re-emergence of HIV-1 upon treatment interruption. Targeting this reservoir will require mechanistic insights into the establishment and maintenance of HIV-1 latency. Whether T cell signaling at the time of HIV-1 infection influences productive replication or latency is not fully understood. We used a panel of chimeric antigen receptors (CARs) with different ligand binding affinities to induce a range of signaling strengths to model differential T cell receptor signaling at the time of HIV-1 infection. Stimulation of T cell lines or primary CD4+ T cells expressing chimeric antigen receptors supported HIV-1 infection regardless of affinity for ligand; however, only signaling by the highest affinity receptor facilitated HIV-1 expression. Activation of chimeric antigen receptors that had intermediate and low binding affinities did not support provirus transcription, suggesting that a minimal signal is required for optimal HIV-1 expression. In addition, strong signaling at the time of infection produced a latent population that was readily inducible, whereas latent cells generated in response to weaker signals were not easily reversed. Chromatin immunoprecipitation showed HIV-1 transcription was limited by transcriptional elongation and that robust signaling decreased the presence of negative elongation factor, a pausing factor, by more than 80%. These studies demonstrate that T cell signaling influences HIV-1 infection and the establishment of different subsets of latently infected cells, which may have implications for targeting the HIV-1 reservoir.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/virologia , Infecções por HIV/imunologia , Infecções por HIV/virologia , HIV-1/imunologia , Provírus/imunologia , Latência Viral/imunologia , Humanos , Células Jurkat , Transdução de Sinais , Ativação Viral/imunologia , Replicação Viral/imunologia
17.
Sci Adv ; 5(3): eaav6322, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30944862

RESUMO

A proposed strategy to cure HIV uses latency-reversing agents (LRAs) to reactivate latent proviruses for purging HIV reservoirs. A variety of LRAs have been identified, but none has yet proven effective in reducing the reservoir size in vivo. Nanocarriers could address some major challenges by improving drug solubility and safety, providing sustained drug release, and simultaneously delivering multiple drugs to target tissues and cells. Here, we formulated hybrid nanocarriers that incorporate physicochemically diverse LRAs and target lymphatic CD4+ T cells. We identified one LRA combination that displayed synergistic latency reversal and low cytotoxicity in a cell model of HIV and in CD4+ T cells from virologically suppressed patients. Furthermore, our targeted nanocarriers selectively activated CD4+ T cells in nonhuman primate peripheral blood mononuclear cells as well as in murine lymph nodes, and substantially reduced local toxicity. This nanocarrier platform may enable new solutions for delivering anti-HIV agents for an HIV cure.


Assuntos
Fármacos Anti-HIV/administração & dosagem , Linfócitos T CD4-Positivos/efeitos dos fármacos , Infecções por HIV/tratamento farmacológico , HIV-1/efeitos dos fármacos , Nanoestruturas/administração & dosagem , Latência Viral/efeitos dos fármacos , Animais , Fármacos Anti-HIV/química , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/virologia , Células Cultivadas , Fenômenos Químicos , Portadores de Fármacos/química , Infecções por HIV/imunologia , Infecções por HIV/virologia , HIV-1/imunologia , HIV-1/fisiologia , Humanos , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/virologia , Ativação Linfocitária/efeitos dos fármacos , Ativação Linfocitária/imunologia , Macaca , Camundongos Endogâmicos C57BL , Nanoestruturas/química , Latência Viral/imunologia
18.
EBioMedicine ; 43: 295-306, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30952614

RESUMO

BACKGROUND: Despite the success of antiretroviral therapy (ART), latent HIV-1 continues to persist in a long-lived population of resting memory CD4+ T cells within those who are infected. Finding a safe and effective means to induce latency reversal (LR) during ART to specifically expose this latent HIV-1 cellular reservoir for immune elimination has been a major barrier to a functional cure. METHODS: In this study, we test the use of antigen-presenting type 1-polarized, monocyte-derived dendritic cells (MDC1) generated from chronic HIV-1-infected individuals on ART as a means to induce HIV-1 latency reversal in autologous CD4+ T cells harboring replication-competent provirus. We use the same MDC1 for ex-vivo generation of autologous HIV-1 antigen-specific CD8+ cytotoxic T cells (CTL) and test their effector responses against the MDC1-exposed HIV-1- infected CD4+ T cell targets. FINDINGS: MDC1 presentation of either HIV-1 or cytomegalovirus (CMV) antigens to CD4+ T cells facilitated HIV-1 LR. This antigen-driven MDC1-mediated LR was sharply diminished with blockade of the CD40L/CD40 'helper' signaling pathway. Importantly, these antigen-presenting MDC1 also activated the expansion of CTL capable of killing the exposed HIV-1-infected targets. INTERPRETATION: Inclusion of virus-associated MHC class II 'helper' antigens in MDC1-based HIV-1 immunotherapies could serve both as a targeted means to safely unmask antigen-specific CD4+ T cells harboring HIV-1, and to support CTL responses that can effectively target the MDC1-exposed HIV-1 cellular reservoir as a functional cure strategy. FUND: This study was supported by the NIH-NAID grants R21-AI131763, U01-AI35041, UM1-AI126603, and T32-AI065380.


Assuntos
Células Dendríticas/imunologia , Epitopos/imunologia , Infecções por HIV/imunologia , Infecções por HIV/virologia , HIV-1/fisiologia , Interações Hospedeiro-Patógeno/imunologia , Latência Viral/imunologia , Antígenos Virais , Biomarcadores , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD4-Positivos/virologia , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Células Dendríticas/metabolismo , Infecções por HIV/tratamento farmacológico , Infecções por HIV/metabolismo , Humanos , Interferon gama/metabolismo , RNA Viral , Linfócitos T Citotóxicos/imunologia , Replicação Viral
19.
J Virol ; 93(10)2019 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-30842333

RESUMO

Combination anti-retroviral drug therapy (ART) potently suppresses HIV-1 replication but does not result in virus eradication or a cure. A major contributing factor is the long-term persistence of a reservoir of latently infected cells. To study this reservoir, we established a humanized mouse model of HIV-1 infection and ART suppression based on an oral ART regimen. Similar to humans, HIV-1 levels in the blood of ART-treated animals were frequently suppressed below the limits of detection. However, the limited timeframe of the mouse model and the small volume of available samples makes it a challenging model with which to achieve full viral suppression and to investigate the latent reservoir. We therefore used an ex vivo latency reactivation assay that allows a semiquantitative measure of the latent reservoir that establishes in individual animals, regardless of whether they are treated with ART. Using this assay, we found that latently infected human CD4 T cells can be readily detected in mouse lymphoid tissues and that latent HIV-1 was enriched in populations expressing markers of T cell exhaustion, PD-1 and TIGIT. In addition, we were able to use the ex vivo latency reactivation assay to demonstrate that HIV-specific TALENs can reduce the fraction of reactivatable virus in the latently infected cell population that establishes in vivo, supporting the use of targeted nuclease-based approaches for an HIV-1 cure.IMPORTANCE HIV-1 can establish latent infections that are not cleared by current antiretroviral drugs or the body's immune responses and therefore represent a major barrier to curing HIV-infected individuals. However, the lack of expression of viral antigens on latently infected cells makes them difficult to identify or study. Here, we describe a humanized mouse model that can be used to detect latent but reactivatable HIV-1 in both untreated mice and those on ART and therefore provides a simple system with which to study the latent HIV-1 reservoir and the impact of interventions aimed at reducing it.


Assuntos
HIV-1/imunologia , Latência Viral/imunologia , Latência Viral/fisiologia , Animais , Antirretrovirais/farmacologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD4-Positivos/virologia , Modelos Animais de Doenças , Infecções por HIV/virologia , Soropositividade para HIV/tratamento farmacológico , HIV-1/patogenicidade , Humanos , Camundongos , Receptor de Morte Celular Programada 1/imunologia , Receptores Imunológicos/imunologia , Nucleases dos Efetores Semelhantes a Ativadores de Transcrição/imunologia , Ativação Viral , Replicação Viral
20.
EBioMedicine ; 42: 97-108, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30824386

RESUMO

BACKGROUND: The latent HIV-1 reservoir in treated patients primarily consists of resting memory CD4+ T cells. Stimulating the T-cell receptor (TCR), which facilitates transition of resting into effector T cells, is the most effective strategy to purge these latently infected cells. Here we supply evidence that TCR-stimulated effector T cells still frequently harbor latent HIV-1. METHODS: Primary HIV-1 infected cells were used in a latency assay with or without dendritic cells (DCs) and reversion of HIV-1 latency was determined, in the presence or absence of specific pathway inhibitors. FINDINGS: Renewed TCR-stimulation or subsequent activation with latency reversing agents (LRAs) did not overcome latency. However, interaction of infected effector cells with DCs triggered further activation of latent HIV-1. When compared to TCR-stimulation only, CD4+ T cells from aviremic patients receiving TCR + DC-stimulation reversed latency more frequently. Such a "one-two punch" strategy seems ideal for purging the reservoir. We determined that DC contact activates the PI3K-Akt-mTOR pathway in CD4+ T cells. INTERPRETATION: This insight could facilitate the development of a novel class of potent LRAs that purge latent HIV beyond levels reached by T-cell activation.


Assuntos
Infecções por HIV/metabolismo , Infecções por HIV/virologia , HIV-1/fisiologia , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Latência Viral , Adulto , Idoso , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD4-Positivos/virologia , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Feminino , Infecções por HIV/tratamento farmacológico , Infecções por HIV/imunologia , HIV-1/efeitos dos fármacos , Humanos , Ativação Linfocitária/imunologia , Masculino , Pessoa de Meia-Idade , Modelos Biológicos , NF-kappa B/metabolismo , Fosforilação , Ligação Proteica , Proteínas Proto-Oncogênicas c-fos/química , Proteínas Proto-Oncogênicas c-fos/metabolismo , Proteínas Proto-Oncogênicas c-jun/química , Proteínas Proto-Oncogênicas c-jun/metabolismo , Transdução de Sinais , Latência Viral/imunologia
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA