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AIM: Inhibitors of nerve growth factor (NGF) reduce pain in several chronic pain indications. NGF signals through tyrosine kinase receptors of the tropomyosin-related kinase (Trk) family and the unrelated p75 receptor. PF-06273340 is a small molecule inhibitor of Trks A, B and C that reduces pain in nonclinical models, and the present study aimed to investigate the pharmacodynamics of this first-in-class molecule in humans. METHODS: A randomized, double-blind, single-dose, placebo- and active-controlled five-period crossover study was conducted in healthy human subjects (NCT02260947). Subjects received five treatments: PF-06273340 50 mg, PF-06273340 400 mg, pregabalin 300 mg, ibuprofen 600 mg and placebo. The five primary endpoints were the pain detection threshold for the thermal pain tests and the pain tolerance threshold for the cold pressor, electrical stair and pressure pain tests. The trial had predefined decision rules based on 95% confidence that the PF-06273340 effect was better than that of placebo. RESULTS: Twenty subjects entered the study, with 18 completing all five periods. The high dose of PF-06273340 met the decision rules on the ultraviolet (UV) B skin thermal pain endpoint [least squares (LS) mean vs. placebo: 1.13, 95% confidence interval: 0.64-1.61], but not on the other four primary endpoints. The low dose did not meet the decision criteria for any of the five primary endpoints. Pregabalin (cold pressor and electrical stair tests) and ibuprofen (UVB thermal pain) showed significant analgesic effects on expected endpoints. CONCLUSIONS: The study demonstrated, for the first time, the translation of nonclinical effects into man in an inflammatory pain analgesic pharmacodynamic endpoint using a pan-Trk inhibitor.
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Analgésicos/uso terapêutico , Hiperalgesia/tratamento farmacológico , Glicoproteínas de Membrana/antagonistas & inibidores , Pirimidinas/uso terapêutico , Pirróis/uso terapêutico , Receptor trkA/antagonistas & inibidores , Receptor trkB/antagonistas & inibidores , Receptor trkC/antagonistas & inibidores , Adolescente , Adulto , Analgésicos/administração & dosagem , Analgésicos/farmacocinética , Estudos Cross-Over , Relação Dose-Resposta a Droga , Método Duplo-Cego , Voluntários Saudáveis , Humanos , Hiperalgesia/enzimologia , Masculino , Pessoa de Meia-Idade , Limiar da Dor/efeitos dos fármacos , Pirimidinas/administração & dosagem , Pirimidinas/farmacocinética , Pirróis/administração & dosagem , Pirróis/farmacocinética , Adulto JovemRESUMO
BACKGROUND: The therapeutic potential of phosphodiesterase-5 inhibitor PF-03049423 was evaluated in a phase 2, multicenter, randomized, double-blind, placebo-controlled study of subjects with acute ischemic stroke ( CLINICAL TRIAL REGISTRATION INFORMATION: http://www.clinicaltrials.gov, unique identifier: NCT01208233; http://www.clinicaltrialsregister.eu, EudraCT number: 2010-021414-32). MATERIALS AND METHODS: Subjects (N = 70) received PF-03049423 6 mg (or placebo, N = 67) once daily, orally, commencing between 24 and 78 hours of stroke onset, and continuing for 90 days. Postbaseline efficacy assessments were performed on Days 7, 14, 30, 60, and 90. Modified Rankin Scale (mRS), Barthel Index, National Institutes of Health Stroke Scale, Box and Blocks Test, Hand-Grip Strength Test, 10-Meter Walk Test, Repeatable Battery Assessment of Neuropsychological Status Naming and Coding Subtests, Line Cancellation Test, and Recognition Memory Test were administered to evaluate poststroke recovery. The primary endpoint was the mRS responder rate (score 0-2 at Day 90). The study included a planned interim analysis of efficacy data. RESULTS: The primary efficacy analysis using logistic regression showed no statistically significant difference between PF-03049423 6 mg and placebo (responder rate of 42.6% and 46.2%, respectively). Although PF-03049423 showed a satisfactory safety and tolerability profile, no signal of efficacy emerged from any of the outcome measures. CONCLUSIONS: PF-03049423 showed no therapeutic potential for acute ischemic stroke.
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Fármacos Neuroprotetores/uso terapêutico , Inibidores da Fosfodiesterase 5/uso terapêutico , Acidente Vascular Cerebral/tratamento farmacológico , Adulto , Idoso , Idoso de 80 Anos ou mais , Método Duplo-Cego , Feminino , Humanos , Cooperação Internacional , Masculino , Pessoa de Meia-Idade , Pirazinas/uso terapêutico , Piridinas/uso terapêutico , Índice de Gravidade de Doença , Fatores de TempoRESUMO
The expression of endogenous retrotransposable elements, including long interspersed nuclear element 1 (LINE-1 or L1) and human endogenous retrovirus, accompanies neoplastic transformation and infection with viruses such as HIV. The ability to engender immunity safely against such self-antigens would facilitate the development of novel vaccines and immunotherapies. In this article, we address the safety and immunogenicity of vaccination with these elements. We used immunohistochemical analysis and literature precedent to identify potential off-target tissues in humans and establish their translatability in preclinical species to guide safety assessments. Immunization of mice with murine L1 open reading frame 2 induced strong CD8 T cell responses without detectable tissue damage. Similarly, immunization of rhesus macaques with human LINE-1 open reading frame 2 (96% identity with macaque), as well as simian endogenous retrovirus-K Gag and Env, induced polyfunctional T cell responses to all Ags, and Ab responses to simian endogenous retrovirus-K Env. There were no adverse safety or pathological findings related to vaccination. These studies provide the first evidence, to our knowledge, that immune responses can be induced safely against this class of self-antigens and pave the way for investigation of them as HIV- or tumor-associated targets.
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Vacinas contra a AIDS/administração & dosagem , Vacinas contra a AIDS/imunologia , Vacinas Anticâncer/administração & dosagem , Vacinas Anticâncer/imunologia , Elementos de DNA Transponíveis/imunologia , Retrovirus Endógenos/imunologia , Vacinas contra a AIDS/genética , Adulto , Sequência de Aminoácidos , Animais , Vacinas Anticâncer/genética , Elementos de DNA Transponíveis/genética , Modelos Animais de Doenças , Retrovirus Endógenos/genética , Retrovirus Endógenos/metabolismo , Feminino , Humanos , Macaca mulatta , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Produtos do Gene env do Vírus da Imunodeficiência Humana/genética , Produtos do Gene env do Vírus da Imunodeficiência Humana/imunologia , Produtos do Gene gag do Vírus da Imunodeficiência Humana/genética , Produtos do Gene gag do Vírus da Imunodeficiência Humana/imunologiaRESUMO
BACKGROUND: We evaluated the immunological responses of African green monkeys immunized with multiple F and G protein-based vaccines and assessed protection against the Memphis 37 strain of respiratory syncytial virus (RSV). METHODS: Monkeys were immunized with F and G proteins adjuvanted with immunostimulatory (CpG) oligodeoxyribonucleotides admixed with either Alhydrogel or ISCOMATRIX adjuvant. Delivery of F and G proteins via replication incompetent recombinant vesicular stomatitis viruses (VSVs) and human adenoviruses was also evaluated. Mucosally or parenterally administered recombinant adenoviruses were used in prime-boost regimens with adjuvanted proteins or recombinant DNA. RESULTS: Animals primed by intranasal delivery of recombinant adenoviruses, and boosted by intramuscular injection of adjuvanted F and G proteins, developed neutralizing antibodies and F/G protein-specific T cells and were protected from RSV infection. Intramuscular injections of Alhydrogel (plus CpG) adjuvanted F and G proteins reduced peak viral loads in the lungs of challenged monkeys. Granulocyte numbers were not significantly elevated, relative to controls, in postchallenge bronchoalveolar lavage samples from vaccinated animals. CONCLUSIONS: This study has validated the use of RSV (Memphis 37) in an African green monkey model of intranasal infection and identified nonreplicating vaccines capable of eliciting protection in this higher species challenge model.
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Infecções por Vírus Respiratório Sincicial/prevenção & controle , Vacinas contra Vírus Sincicial Respiratório/farmacologia , Vírus Sinciciais Respiratórios/imunologia , Adenovírus Humanos/genética , Adenovírus Humanos/imunologia , Adjuvantes Imunológicos/farmacologia , Animais , Anticorpos Antivirais/imunologia , Lavagem Broncoalveolar/métodos , Chlorocebus aethiops , Granulócitos/imunologia , Granulócitos/virologia , Imunização/métodos , Pulmão/imunologia , Pulmão/virologia , Distribuição Aleatória , Infecções por Vírus Respiratório Sincicial/imunologia , Infecções por Vírus Respiratório Sincicial/virologia , Vacinas contra Vírus Sincicial Respiratório/genética , Vacinas contra Vírus Sincicial Respiratório/imunologia , Vírus Sinciciais Respiratórios/genética , Linfócitos T/imunologia , Linfócitos T/virologia , Vesiculovirus/genética , Vesiculovirus/imunologia , Proteínas Virais de Fusão/genética , Proteínas Virais de Fusão/imunologia , Carga Viral/imunologia , Proteínas Virais/genética , Proteínas Virais/imunologia , Replicação Viral/genética , Replicação Viral/imunologiaRESUMO
BACKGROUND: Elevated BCAA levels are strongly associated with diabetes, but how diabetes affects BCAA, branched-chain ketoacids (BCKAs), and the broader metabolome after a meal is not well known. OBJECTIVE: To compare quantitative BCAA and BCKA levels in a multiracial cohort with and without diabetes after a mixed meal tolerance test (MMTT) as well as to explore the kinetics of additional metabolites and their associations with mortality in self-identified African Americans. METHODS: We administered an MMTT to 11 participants without obesity or diabetes and 13 participants with diabetes (treated with metformin only) and measured the levels of BCKAs, BCAAs, and 194 other metabolites at 8 time points across 5 h. We used mixed models for repeated measurements to compare between group metabolite differences at each timepoint with adjustment for baseline. We then evaluated the association of top metabolites with different kinetics with all-cause mortality in the Jackson Heart Study (JHS) (N = 2441). RESULTS: BCAA levels, after adjustment for baseline, were similar at all timepoints between groups, but adjusted BCKA kinetics were different between groups for α-ketoisocaproate (P = 0.022) and α-ketoisovalerate (P = 0.021), most notably diverging at 120 min post-MMTT. An additional 20 metabolites had significantly different kinetics across timepoints between groups, and 9 of these metabolites-including several acylcarnitines-were significantly associated with mortality in JHS, irrespective of diabetes status. The highest quartile of a composite metabolite risk score was associated with higher mortality (HR:1.57; 1.20, 2.05, P = 0.00094) than the lowest quartile. CONCLUSIONS: BCKA levels remained elevated after an MMTT among participants with diabetes, suggesting that BCKA catabolism may be a key dysregulated process in the interaction of BCAA and diabetes. Metabolites with different kinetics after an MMTT may be markers of dysmetabolism and associated with increased mortality in self-identified African Americans.
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Aminoácidos de Cadeia Ramificada , Diabetes Mellitus , Humanos , Aminoácidos de Cadeia Ramificada/metabolismo , Fatores de Risco , Obesidade/metabolismo , MetabolomaRESUMO
Background: An urgent need remains for antiviral therapies to treat patients hospitalized with COVID-19. PF-07304814-the prodrug (lufotrelvir) and its active moiety (PF-00835231)-is a potent inhibitor of the SARS-CoV-2 3CL protease. Method: Eligible participants were 18 to 79 years old and hospitalized with confirmed COVID-19. This first-in-human phase 1b study was designed with 2 groups: single ascending dose (SAD) and multiple ascending dose (MAD). Participants could receive local standard-of-care therapy. In SAD, participants were randomized to receive a 24-hour infusion of lufotrelvir/placebo. In MAD, participants were randomized to receive a 120-hour infusion of lufotrelvir/placebo. The primary endpoint was to assess the safety and tolerability of lufotrelvir. The secondary endpoint was to evaluate the pharmacokinetics of lufotrelvir and PF-00835231. Results: In SAD, participants were randomized to receive 250 mg lufotrelvir (n = 2), 500 mg lufotrelvir (n = 2), or placebo (n = 4) by continuous 24-hour infusion. In MAD, participants were randomized to receive 250 mg lufotrelvir (n = 7), 500 mg lufotrelvir (n = 6), or placebo (n = 4) by continuous 120-hour infusion. No adverse events or serious adverse events were considered related to lufotrelvir. At doses of 250 and 500â mg, concentrations for the prodrug lufotrelvir and active moiety PF-00835231 increased in a dose-related manner. Unbound concentrations of the lufotrelvir active metabolite reached steady state approximately 2- and 4-fold that of in vitro EC90 following 250- and 500-mg doses, respectively. Conclusions: These safety and pharmacokinetic findings support the continued evaluation of lufotrelvir in clinical studies. Clinical Trials Registration. ClinicalTrials.gov NCT04535167.
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Gout is the most common form of inflammatory arthritis and is a multifactorial disease typically characterized by hyperuricemia and monosodium urate crystal deposition predominantly in, but not limited to, the joints and the urinary tract. The prevalence of gout and hyperuricemia has increased in developed countries over the past two decades and research into the area has become progressively more active. We review the current field of knowledge with emphasis on active areas of hyperuricemia research including the underlying physiology, genetics and epidemiology, with a focus on studies which suggest association of hyperuricemia with common comorbidities including cardiovascular disease, renal insufficiency, metabolic syndrome and diabetes. Finally, we discuss current therapies and emerging drug discovery efforts aimed at delivering an optimized clinical treatment strategy.
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Age-related macular degeneration (AMD) remains a major cause of blindness, with dysfunction and loss of retinal pigment epithelium (RPE) central to disease progression. We engineered an RPE patch comprising a fully differentiated, human embryonic stem cell (hESC)-derived RPE monolayer on a coated, synthetic basement membrane. We delivered the patch, using a purpose-designed microsurgical tool, into the subretinal space of one eye in each of two patients with severe exudative AMD. Primary endpoints were incidence and severity of adverse events and proportion of subjects with improved best-corrected visual acuity of 15 letters or more. We report successful delivery and survival of the RPE patch by biomicroscopy and optical coherence tomography, and a visual acuity gain of 29 and 21 letters in the two patients, respectively, over 12 months. Only local immunosuppression was used long-term. We also present the preclinical surgical, cell safety and tumorigenicity studies leading to trial approval. This work supports the feasibility and safety of hESC-RPE patch transplantation as a regenerative strategy for AMD.
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Células-Tronco Embrionárias Humanas/transplante , Degeneração Macular/terapia , Epitélio Pigmentado da Retina/transplante , Acuidade Visual/fisiologia , Idoso , Animais , Membrana Basal/diagnóstico por imagem , Membrana Basal/crescimento & desenvolvimento , Diferenciação Celular/genética , Feminino , Humanos , Degeneração Macular/diagnóstico por imagem , Degeneração Macular/patologia , Masculino , Camundongos , Pessoa de Meia-Idade , Epitélio Pigmentado da Retina/diagnóstico por imagem , Epitélio Pigmentado da Retina/crescimento & desenvolvimento , Transplante de Células-Tronco/efeitos adversos , Suínos , Tomografia de Coerência ÓpticaRESUMO
The objective of these clinical studies was to assess the safety and urate lowering activity of a novel urate transporter 1 (URAT1)/ xanthine oxidase (XO) inhibitor PF-06743649 in healthy subjects and gout patients. Escalating doses of PF-06743649 or placebo were given to healthy young subjects, healthy elderly subjects and gout patients. Serum uric acid (sUA) and urinary pharmacodynamic markers were assayed, and safety was assessed by collection of adverse events and assessment of safety labs, ECGs and vital signs. Administration of PF-06743649 led to rapid decrease in sUA in all cohorts; in gout patients, a change from baseline of 69 % was observed for the 40 mg dose. Urinary and serum biomarkers were consistent with inhibition of both URAT1 and XO. Although dosing was otherwise well tolerated, two subjects experienced serious adverse events of acute kidney injury. Both subjects exhibited increased serum creatinine and blood urea nitrogen in the first 3 days post first dose and were hospitalised. One subject exhibited oliguria for the first 24 h. Both subjects made a complete recovery with minimal intervention. PF-06743649 was effective at rapidly lowering sUA, but further development was terminated for an identified renal safety risk.
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Injúria Renal Aguda/induzido quimicamente , Supressores da Gota/efeitos adversos , Supressores da Gota/farmacocinética , Gota/tratamento farmacológico , Transportadores de Ânions Orgânicos/antagonistas & inibidores , Proteínas de Transporte de Cátions Orgânicos/antagonistas & inibidores , Xantina Oxidase/antagonistas & inibidores , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores/análise , Método Duplo-Cego , Feminino , Humanos , Hiperuricemia/sangue , Masculino , Pessoa de Meia-Idade , Estados Unidos , Ácido Úrico/sangue , Adulto JovemRESUMO
Objective. Ischemic stroke is a serious medical condition with limited therapeutic options. The evaluation of the therapeutic potential of novel pharmacological interventions is carried-out in Phase II trials. The study design, primarily intended to evaluate efficacy and safety, is a balance between utilizing as few patients as possible to minimize safety risk and enrolling sufficient patients to detect unambiguous efficacy signals. We sought to determine whether post-stroke recovery outcomes based on behavioral measures of cognitive and motor impairment yielded additional information beyond that of clinician-based methods. Design. This was a multicenter, multinational, randomized, parallel group, controlled versus placebo, efficacy, and safety study of PF-03049423 for treatment of acute ischemic stroke. Settings and participants. Our study subjects were acute ischemic stroke inpatients. Measurements. Outcome measures were derived from rating scales (Modified Rankin Scale, Barthel Index, and National Institutes of Health Stroke Scale) and behavioral tests (Box and Blocks Test, Hand Grip Strength Test, 10-Meter Walk Test, Repeatable Battery Assessment of Neuropsychological Status Naming and Coding Subtests, Line Cancellation Test, and Recognition Memory Test). Assessments were performed at Days 7, 14, 30, 60, and 90. Post-hoc analyses of correlations among the outcome measures at each measurement time point on a cohort of 137 subjects were conducted. Results. Results support the validity of measures from Box and Blocks Test, Hand Grip Strength Test, 10-Meter Walk Test, and Repeatable Battery Assessment of Neuropsychological Status Coding Subtests to monitor post-stroke recovery in clinical trial settings. Notably, the Recognition Memory Test did not show a correlation with the Modified Rankin Scale, and, in fact, did not show improvement over time. Conclusion. The behavioral measures of cognitive and motor functions included in this study may extend the evaluation of the therapeutic potential of new treatments for stroke recovery. The lack of correlation between Recognition Memory Test and the traditional efficacy endpoints, at least in part due to absence of any improvement in recognition memory, suggests that there may be cognitive elements not detected by the Modified Rankin Scale. This is clinically relevant and memory improvement has potential as an endpoint in future trials aiming to improve certain aspects of cognition.
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The social and economic consequences of drug addiction are immense. Although many methods are adopted to treat addiction, including therapeutic intervention and counseling, the long-term success rate has been limited and there continues to be a need for more effective treatments. A novel approach that has sparked a significant degree of interest recently is the use of vaccines designed to raise specific antibodies against drugs of abuse. Antibodies that prevent addictive substances crossing the blood-brain barrier may prove to be an effective mechanism that will help prevent relapse during efforts to abstain from the drug. Proof-of-principle for this approach has been established in numerous animal models. Currently a cocaine vaccine is in phase II clinical trials and, more recently, two vaccines to nicotine have entered phase I trials. Key efficacy trials are required to establish the true potential of these therapeutic vaccines.
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Transtornos Relacionados ao Uso de Substâncias/terapia , Vacinas/uso terapêutico , Animais , Ensaios Clínicos como Assunto , Transtornos Relacionados ao Uso de Cocaína/imunologia , Transtornos Relacionados ao Uso de Cocaína/terapia , Humanos , Modelos Imunológicos , Transtornos Relacionados ao Uso de Substâncias/imunologia , Tabagismo/imunologia , Tabagismo/terapiaRESUMO
Orthopoxviruses remain a threat as biological weapons and zoonoses. The licensed live-virus vaccine is associated with serious health risks, making its general usage unacceptable. Attenuated vaccines are being developed as alternatives, the most advanced of which is modified-vaccinia virus Ankara (MVA). We previously developed a gene-based vaccine, termed 4pox, which targets four orthopoxvirus antigens, A33, B5, A27 and L1. This vaccine protects mice and non-human primates from lethal orthopoxvirus disease. Here, we investigated the capacity of the molecular adjuvants GM-CSF and Escherichia coli heat-labile enterotoxin (LT) to enhance the efficacy of the 4pox gene-based vaccine. Both adjuvants significantly increased protective antibody responses in mice. We directly compared the 4pox plus LT vaccine against MVA in a monkeypox virus (MPXV) nonhuman primate (NHP) challenge model. NHPs were vaccinated twice with MVA by intramuscular injection or the 4pox/LT vaccine delivered using a disposable gene gun device. As a positive control, one NHP was vaccinated with ACAM2000. NHPs vaccinated with each vaccine developed anti-orthopoxvirus antibody responses, including those against the 4pox antigens. After MPXV intravenous challenge, all control NHPs developed severe disease, while the ACAM2000 vaccinated animal was well protected. All NHPs vaccinated with MVA were protected from lethality, but three of five developed severe disease and all animals shed virus. All five NHPs vaccinated with 4pox/LT survived and only one developed severe disease. None of the 4pox/LT-vaccinated animals shed virus. Our findings show, for the first time, that a subunit orthopoxvirus vaccine delivered by the same schedule can provide a degree of protection at least as high as that of MVA.
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Vacina Antivariólica/administração & dosagem , Animais , Feminino , Macaca mulatta , Masculino , Plasmídeos , Vacina Antivariólica/genéticaRESUMO
DNA vaccines expressing HSV-2 gD, gB, ICP27, VP22 and VP13/14 were shown to be immunogenic in mice; gD and gB elicited neutralising antibody, and all five antigens induced T cell responses measured by IFNγ ELISPOT. In murine HSV-2 challenge studies, gD and gB provided moderate to high levels of protection while ICP27 provided a lower level of protection depending on the model (intravaginal or intranasal) and the challenge dose. Combining vaccines expressing gB or gD with vaccines expressing ICP27 provided greater protection than any antigen alone. We conclude that the addition of ICP27 to enhance the anti-viral T cell response can improve the efficacy of gD- and gB-based vaccines.
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Infecções por Herpesviridae/prevenção & controle , Herpesvirus Humano 2/imunologia , Vacinas contra Herpesvirus/imunologia , Proteínas Imediatamente Precoces/imunologia , Vacinas de DNA/imunologia , Proteínas do Envelope Viral/imunologia , Proteínas Virais/imunologia , Animais , Anticorpos Antivirais/sangue , Modelos Animais de Doenças , Feminino , Infecções por Herpesviridae/imunologia , Infecções por Herpesviridae/patologia , Herpesvirus Humano 2/genética , Vacinas contra Herpesvirus/administração & dosagem , Proteínas Imediatamente Precoces/genética , Camundongos , Camundongos Endogâmicos BALB C , Índice de Gravidade de Doença , Linfócitos T/imunologia , Vacinas de DNA/administração & dosagem , Proteínas do Envelope Viral/genética , Proteínas Virais/genéticaRESUMO
Candidate DNA vaccines for hemorrhagic fever with renal syndrome expressing the envelope glycoprotein genes of Hantaan (HTNV) or Puumala (PUUV) viruses were evaluated in an open-label, single-center Phase 1 study consisting of three vaccination groups of nine volunteers. The volunteers were vaccinated by particle-mediated epidermal delivery (PMED) three times at four-week intervals with the HTNV DNA vaccine, the PUUV DNA vaccine or both vaccines. At each dosing, the volunteers received 8 µg DNA/4 mg gold. There were no study-related serious adverse events, and all injection site pain was graded as mild. The most commonly reported systemic adverse events were fatigue, headache, malaise, myalgia, and lymphadenopathy. Blood samples were collected on days 0, 28, 56, 84, 140, and 180, and assayed for the presence of neutralizing antibodies. In the single vaccine groups, neutralizing antibodies to HTNV or PUUV were detected in 30% or 44% of individuals, respectively. In the combined vaccine group, 56% of the volunteers developed neutralizing antibodies to one or both viruses. These results demonstrate that the HTNV and PUUV DNA vaccines are safe and can be immunogenic in humans when delivered by PMED.
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Vírus Hantaan/patogenicidade , Febre Hemorrágica com Síndrome Renal/prevenção & controle , Virus Puumala/patogenicidade , Vacinas de DNA/imunologia , Vacinas Virais/imunologia , Adulto , Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , Biolística , Feminino , Vírus Hantaan/imunologia , Febre Hemorrágica com Síndrome Renal/imunologia , Humanos , Masculino , Pessoa de Meia-Idade , Testes de Neutralização , Virus Puumala/imunologia , Vacinas de DNA/administração & dosagem , Vacinas de DNA/efeitos adversos , Vacinas Virais/administração & dosagem , Vacinas Virais/efeitos adversos , Adulto JovemRESUMO
The genetic diversity of HIV-1 represents a major challenge in vaccine development. In this study, we establish a rationale for eliminating HIV-1-infected cells by targeting cellular immune responses against stable human endogenous retroviral (HERV) antigens. HERV DNA sequences in the human genome represent the remnants of ancient infectious retroviruses. We show that the infection of CD4+ T cells with HIV-1 resulted in transcription of the HML-2 lineage of HERV type K [HERV-K(HML-2)] and the expression of Gag and Env proteins. HERV-K(HML-2)-specific CD8+ T cells obtained from HIV-1-infected human subjects responded to HIV-1-infected cells in a Vif-dependent manner in vitro. Consistent with the proposed mode of action, a HERV-K(HML-2)-specific CD8+ T cell clone exhibited comprehensive elimination of cells infected with a panel of globally diverse HIV-1, HIV-2, and SIV isolates in vitro. We identified a second T cell response that exhibited cross-reactivity between homologous HIV-1-Pol and HERV-K(HML-2)-Pol determinants, raising the possibility that homology between HIV-1 and HERVs plays a role in shaping, and perhaps enhancing, the T cell response to HIV-1. This justifies the consideration of HERV-K(HML-2)-specific and cross-reactive T cell responses in the natural control of HIV-1 infection and for exploring HERV-K(HML-2)-targeted HIV-1 vaccines and immunotherapeutics.
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Linfócitos T CD4-Positivos/virologia , Retrovirus Endógenos/fisiologia , HIV-1/fisiologia , HIV-2/fisiologia , Imunidade Celular , Vírus da Imunodeficiência Símia/fisiologia , Sequência de Aminoácidos , Animais , Antígenos Virais/genética , Antígenos Virais/imunologia , Antígenos Virais/metabolismo , Linfócitos T CD4-Positivos/imunologia , Células Cultivadas , Retrovirus Endógenos/imunologia , Retrovirus Endógenos/metabolismo , Regulação Viral da Expressão Gênica , Produtos do Gene gag/genética , Produtos do Gene gag/imunologia , Produtos do Gene gag/metabolismo , Infecções por HIV/imunologia , Infecções por HIV/virologia , HIV-1/imunologia , HIV-1/isolamento & purificação , HIV-2/imunologia , HIV-2/isolamento & purificação , Interações Hospedeiro-Patógeno , Humanos , Dados de Sequência Molecular , Vírus da Imunodeficiência Símia/imunologia , Vírus da Imunodeficiência Símia/isolamento & purificação , Ativação Transcricional , Proteínas do Envelope Viral/genética , Proteínas do Envelope Viral/imunologia , Proteínas do Envelope Viral/metabolismo , Integração Viral , Internalização do Vírus , Produtos do Gene vif do Vírus da Imunodeficiência Humana/fisiologiaRESUMO
Despite several attempts to develop an effective prophylactic vaccine for HSV-2, all have failed to show efficacy in the clinic. The most recent of these failures was the GlaxoSmithKline (GSK) subunit vaccine based on the glycoprotein gD with the adjuvant monophosphoryl lipid A (MPL). In a phase 3 clinical trial, this vaccine failed to protect from HSV-2 disease, even though good neutralizing antibody responses were elicited. We aimed to develop a superior, novel HSV-2 vaccine containing either gD or gB alone or in combination, together with the potent adjuvant CpG oligodeoxynucleotides (CPG). The immunogenic properties of these vaccines were compared in mice. We show that gB/CPG/alum elicited a neutralizing antibody response similar to that elicited by gD/CPG/alum vaccine but a significantly greater gamma interferon (IFN-γ) T cell response. Furthermore, the combined gB-gD/CPG/alum vaccine elicited significantly greater neutralizing antibody and T cell responses than gD/MPL/alum. The efficacies of these candidate vaccines were compared in the mouse and guinea pig disease models, including a novel male guinea pig genital disease model. These studies demonstrated that increased immune response did not correlate to improved protection. First, despite a lower IFN-γ T cell response, the gD/CPG/alum vaccine was more effective than gB/CPG/alum in mice. Furthermore, the gB-gD/CPG/alum vaccine was no more effective than gD/MPL/alum in mice or male guinea pigs. We conclude that difficulties in correlating immune responses to efficacy in animal models will act as a deterrent to researchers attempting to develop effective HSV vaccines.
Assuntos
Adjuvantes Imunológicos/administração & dosagem , Herpes Genital/prevenção & controle , Herpesvirus Humano 2/imunologia , Vacinas contra Herpesvirus/imunologia , Proteínas do Envelope Viral/imunologia , Compostos de Alúmen/administração & dosagem , Animais , Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , Modelos Animais de Doenças , Cobaias , Herpes Genital/imunologia , Herpes Genital/patologia , Vacinas contra Herpesvirus/administração & dosagem , Interferon gama/metabolismo , Lipídeo A/administração & dosagem , Lipídeo A/análogos & derivados , Camundongos , Oligodesoxirribonucleotídeos/administração & dosagem , Índice de Gravidade de Doença , Linfócitos T/imunologia , Proteínas do Envelope Viral/administração & dosagemRESUMO
BACKGROUND: The recent H5N1 avian and H1N1 swine-origin influenza virus outbreaks reaffirm that the threat of a world-wide influenza pandemic is both real and ever-present. Vaccination is still considered the best strategy for protection against influenza virus infection but a significant challenge is to identify new vaccine approaches that offer accelerated production, broader protection against drifted and shifted strains, and the capacity to elicit anti-viral immune responses in the respiratory tract at the site of viral entry. As a safe alternative to live attenuated vaccines, the mucosal and systemic immunogenicity of an H1N1 influenza (A/New Caledonia/20/99) HA DNA vaccine administered by particle-mediated epidermal delivery (PMED or gene gun) was analyzed in rhesus macaques. METHODOLOGY/PRINCIPAL FINDINGS: Macaques were immunized at weeks 0, 8, and 16 using a disposable single-shot particle-mediated delivery device designed for clinical use that delivers plasmid DNA directly into cells of the epidermis. Significant levels of hemagglutination inhibiting (HI) antibodies and cytokine-secreting HA-specific T cells were observed in the periphery of macaques following 1-3 doses of the PMED HA DNA vaccine. In addition, HA DNA vaccination induced detectable levels of HA-specific mucosal antibodies and T cells in the lung and gut-associated lymphoid tissues of vaccinated macaques. Importantly, co-delivery of a DNA encoding the rhesus macaque GM-CSF gene was found to significantly enhance both the systemic and mucosal immunogenicity of the HA DNA vaccine. CONCLUSIONS/SIGNIFICANCE: These results provide strong support for the development of a particle-mediated epidermal DNA vaccine for protection against respiratory pathogens such as influenza and demonstrate, for the first time, the ability of skin-delivered GM-CSF to serve as an effective mucosal adjuvant for vaccine induction of immune responses in the gut and respiratory tract.
Assuntos
Adjuvantes Imunológicos/farmacologia , Anticorpos Antivirais/biossíntese , Epiderme/imunologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/farmacologia , Vírus da Influenza A Subtipo H1N1/imunologia , Mucosa/efeitos dos fármacos , Vacinas de DNA/administração & dosagem , Animais , Anticorpos Antivirais/sangue , Imunidade Celular/efeitos dos fármacos , Macaca mulatta , Linfócitos T/imunologia , Vacinas de DNA/imunologiaRESUMO
BACKGROUND: We have developed a Trivalent DNA vaccine for influenza consisting of three plasmids expressing haemagglutinin from different seasonal influenza virus strains delivered using PMED (particle mediated epidermal delivery). We set out to determine whether this vaccine (with and without a molecular adjuvant DNA Encoded Immunostimulator-Labile Toxin (DEI-LT)) could protect subjects from a controlled influenza virus challenge. METHODS: Healthy adult subjects were screened for susceptibility to infection with influenza A/H3 Panama/2007/99 then vaccinated with 4microg Trivalent influenza DNA vaccine, 2microg Trivalent influenza DNA vaccine plus DEI-LT or placebo. Safety and serological responses to vaccination were assessed and on Day 56 subjects were challenged with A/H3 Panama/2007/99 virus. RESULTS: Vaccination with 4microg Trivalent or 2microg Trivalent/DEI-LT was well tolerated and induced antibody responses to two of the three influenza virus vaccine strains. Post challenge, subjects in the 4microg Trivalent group (N=27) showed reductions in disease symptoms and viral shedding compared to placebo (N=27), with an overall vaccine efficacy of 41% (95% confidence interval (CI)=?1.5, 67.7) for 'Any illness with or without fever' and 53% for 'Upper respiratory tract infection' (95% CI=8.0, 77.7). CONCLUSION: It was concluded that PMED vaccination with 4microg Trivalent influenza DNA vaccine was safe and elicited immunological responses that protected human subjects from influenza; this is the first report of protection of human subjects from disease by DNA vaccination.
Assuntos
Vacinas contra Influenza/imunologia , Vacinas de DNA/imunologia , Adjuvantes Imunológicos/administração & dosagem , Adulto , Anticorpos Antivirais/sangue , Toxinas Bacterianas/administração & dosagem , Método Duplo-Cego , Enterotoxinas/administração & dosagem , Proteínas de Escherichia coli/administração & dosagem , Feminino , Humanos , Vacinas contra Influenza/efeitos adversos , Masculino , Pessoa de Meia-Idade , Vacinação , Vacinas de DNA/efeitos adversosRESUMO
Mice were vaccinated with a DNA plasmid encoding the haemagglutinin (HA) antigen of H5N1 influenza A/Vietnam/1194/2004 by particle mediated epidermal delivery (PMED). Vaccination led to potent anti-HA serological responses that were significantly enhanced by the inclusion of a plasmid expressing the A and B subunits of Escherichia coli heat labile enterotoxin (designated DEI-LT). Mice were vaccinated with H5 or H5/DEI-LT and challenged with 100LD50 H5N1 A/Vietnam/1194/2004 virus. Vaccination provided considerable protection, and mice that received two doses (prime-boost) of H5/DEI-LT showed no symptoms of disease post vaccination, did not shed detectable virus and did not show any rise in anti-H5N1 HI titre post challenge, indicating that they were fully protected. These results demonstrate that the PMED technology may hold promise for the prophylaxis of pandemic influenza.
Assuntos
Glicoproteínas de Hemaglutininação de Vírus da Influenza/imunologia , Virus da Influenza A Subtipo H5N1/imunologia , Vacinas contra Influenza/administração & dosagem , Vacinas de DNA/administração & dosagem , Animais , Anticorpos Antivirais/sangue , Sistemas de Liberação de Medicamentos , Epiderme , Feminino , Ouro , Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Vacinas contra Influenza/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Plasmídeos , Vacinação , Vacinas de DNA/imunologiaRESUMO
This review provides an overview of studies employing particle-mediated epidermal delivery (PMED) or the gene gun to administer DNA vaccines for infectious diseases in preclinical studies employing large animal models and in human clinical trials. It reviews the immunogenicity and protective efficacy of PMED DNA vaccines in nonhuman primates and swine and studies that have directly compared the effectiveness of PMED in these large animal models to existing licensed vaccines and intramuscular or intradermal delivery of DNA vaccines with a needle. Various clinical trials employing PMED have been completed and an overview of the immunogenicity, safety, and tolerability of this approach in humans is described. Finally, efforts currently in progress for commercial development of particle-mediated DNA vaccines are discussed.