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1.
Mol Ther ; 22(1): 42-51, 2014 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-24077034

RESUMO

Self-complementary adeno-associated viral (AAV) vectors expressing human factor IX (hF.IX) have achieved transient or sustained correction of hemophilia B in human volunteers. High doses of AAV2 or AAV8 vectors delivered to the liver caused in several patients an increase in transaminases accompanied by a rise in AAV capsid-specific T cells and a decrease in circulating hF.IX levels suggesting immune-mediated destruction of vector-transduced cells. Kinetics of these adverse events differed in patients receiving AAV2 or AAV8 vectors causing rise in transaminases at 3 versus 8 weeks after vector injection, respectively. To test if CD8+ T cells to AAV8 vectors, which are similar to AAV2 vectors are fully-gutted vectors and thereby fail to encode structural viral proteins, could cause damage at this late time point, we tested in a series of mouse studies how long major histocompatibility (MHC) class I epitopes within AAV8 capsid can be presented to CD8+ T cells. Our results clearly show that depending on the vectors' genome, CD8+ T cells can detect such epitopes on AAV8's capsid for up to 6 months indicating that the capsid of AAV8 degrades slowly in mice.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Capsídeo/imunologia , Dependovirus/imunologia , Vetores Genéticos/genética , Vetores Genéticos/imunologia , Genoma , Animais , Proteínas do Capsídeo/imunologia , Dependovirus/genética , Epitopos de Linfócito T , Vetores Genéticos/normas , Humanos , Memória Imunológica , Ativação Linfocitária/imunologia , Masculino , Camundongos , Controle de Qualidade , Especificidade do Receptor de Antígeno de Linfócitos T , Transdução Genética
2.
Mol Ther ; 21(3): 696-706, 2013 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-23229092

RESUMO

To determine if an ordered and repetitive display of an epitope promoted induction of superior antibody responses, we compared B-cell responses to an influenza A virus epitope that was either encoded as a transgene by an adenovirus (Ad) vector or expressed on the vector's surface. To this end, we constructed a panel of influenza A virus vaccines based on chimpanzee-derived replication-defective adenovirus (AdC) vectors of serotype SAd-V25 also called AdC68. AdC68 vectors were modified to express a linear B-cell epitope of the ectodomain of matrix 2 (M2e) within variable regions 1 (VR1) or 4 (VR4) of the adenovirus hexon. Additional vectors with wild-type or M2e-modified hexon encoded M2e fused to the influenza A virus nucleoprotein (NP) as a transgene product. Hexon-modified vectors were tested for immunogenicity and efficacy in mice in comparison to vectors with native hexon expressing the M2e-NP fusion protein. Upon priming, vectors expressing M2e within VR1 of hexon induced M2e-specific antibody responses of higher magnitude and avidity than those carrying M2e within VR4 or vectors expressing the M2e as part of a transgene product. CD8(+) T-cell responses to the transgenic NP were comparable between vectors. M2e-specific antibody responses could be boosted by a second dose of the VR1 hexon-modified vector but not by repeated immunization with the VR4 hexon-modified vector.


Assuntos
Adenoviridae/genética , Vetores Genéticos , Vírus da Influenza A/imunologia , Vacinas contra Influenza/genética , Animais , Linfócitos T CD8-Positivos/imunologia , Epitopos de Linfócito B/imunologia , Terapia Genética , Vacinas contra Influenza/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos ICR , Proteínas do Nucleocapsídeo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/imunologia , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/imunologia , Sensibilidade e Especificidade , Proteínas do Core Viral/genética , Proteínas do Core Viral/imunologia , Proteínas da Matriz Viral/genética , Proteínas da Matriz Viral/imunologia
3.
Mol Ther ; 19(3): 536-46, 2011 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-21157435

RESUMO

Using adoptive transfer models we determined that an adeno-associated viral vector of serotype 2 (AAV2) induces in mice proliferation of CD8(+) T cells that recognize an epitope within the viral capsid. Proliferation to an endogenous epitope within viral protein (VP)3 could be observed for at least 3 weeks while a foreign epitope placed at multiple copies within VP2 elicited CD8(+) T cell expansion for at least 10 weeks. These data show that capsid antigens of AAV2 degrade slowly over a period of weeks and during this period provide targets to CD8(+) T cells.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Proteínas do Capsídeo/imunologia , Dependovirus/genética , Vetores Genéticos/genética , Ativação Linfocitária/imunologia , Animais , Apresentação de Antígeno/imunologia , Células Cultivadas , Células Dendríticas/imunologia , Epitopos de Linfócito T/química , Epitopos de Linfócito T/genética , Epitopos de Linfócito T/metabolismo , Fator IX/genética , Fator IX/metabolismo , Feminino , Ordem dos Genes , Técnicas de Transferência de Genes , Hepatócitos/imunologia , Humanos , Imunidade Celular , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL
4.
Mol Ther ; 19(2): 417-26, 2011 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-21081905

RESUMO

Despite enormous efforts by the scientific community, an effective HIV vaccine remains elusive. To further address to what degree T cells in absence of antibodies may protect against simian immunodeficiency virus (SIV) disease progression, rhesus macaques were vaccinated intramuscularly with a chimpanzee-derived Ad vector (AdC) serotype 6 and then boosted intramuscularly with a serologically distinct AdC vector of serotype 7 both expressing Gag of SIVmac239. Animals were subsequently boosted intramuscularly with a modified vaccinia Ankara (MVA) virus expressing Gag and Tat of the homologous SIV before mucosal challenge with a high dose of SIVmac239 given rectally. Whereas vaccinated animals showed only a modest reduction of viral loads, their overall survival was improved, in association with a substantial protection from the loss of CD4(+) T cells. In addition, the two vaccinated Mamu-A*01(+) macaques controlled viral loads to levels below detection within weeks after challenge. These data strongly suggest that T cells, while unable to affect SIV acquisition upon high-dose rectal infection, can reduce disease progression. Induction of potent T-cell responses should thus remain a component of our efforts to develop an efficacious vaccine to HIV-1.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Macaca mulatta/imunologia , Macaca mulatta/virologia , Vacinas contra a SAIDS/imunologia , Vírus da Imunodeficiência Símia/imunologia , Vírus da Imunodeficiência Símia/patogenicidade , Animais , Feminino , Masculino
5.
Mol Ther ; 18(12): 2182-9, 2010 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-20877342

RESUMO

A universal influenza vaccine, designed to induce broadly cross-reactive immunity against current and future influenza A virus strains, is in critical demand to reduce the need for annual vaccinations with vaccines chosen upon predicting the predominant circulating viral strains, and to ameliorate the threat of cyclically occurring pandemics that have, in the past, killed tens of millions. Here, we describe a vaccine regimen based on sequential immunization with two serologically distinct chimpanzee-derived replication-defective adenovirus (Ad) vectors expressing the matrix-2 protein ectodomain (M2e) from three divergent strains of influenza A virus fused to the influenza virus nucleoprotein (NP) for induction of antibodies to M2e and virus-specific CD8(+) T cells to NP. In preclinical mouse models, the Ad vaccines expressing M2e and NP elicit robust NP-specific CD8(+) T-cell responses and moderate antibody responses to all three M2e sequences. Most importantly, vaccinated mice are protected against morbidity and mortality following challenge with high doses of different influenza virus strains. Protection requires both antibodies to M2e and cellular immune responses to NP.


Assuntos
Adenoviridae , Vírus da Influenza A , Vacinas contra Influenza , Influenza Humana/prevenção & controle , Nucleoproteínas/metabolismo , Proteínas da Matriz Viral/metabolismo , Adenoviridae/genética , Sequência de Aminoácidos , Animais , Humanos , Vírus da Influenza A/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Dados de Sequência Molecular , Proteínas Recombinantes de Fusão/genética
6.
Trends Mol Med ; 15(1): 32-9, 2009 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-19101205

RESUMO

Immune responses to viral vectors pose one of the main obstacles to successful human gene replacement therapy, unless gene transfer vectors are applied to immune-privileged sites. Both innate and adaptive immunity work in concert against sustained gene transfer, but the functions of patients' regulatory T cells (Tregs) and tolerogenic dendritic cells (DCs) could potentially be harnessed to reduce these immune responses. Over the last few years, immunologists have gained an ever-increasing knowledge of immunoregulatory pathways, especially those that prevent or dampen adaptive immune responses. The gene therapy community is now in a position to use this expanding knowledge in basic immunology to overcome the so far nearly unsurpassable obstacles posed by the immune system to the long-term replacement of missing or faulty genes by the use of viral vectors. Here, we discuss the current challenges in overcoming immune barriers to gene therapy. In addition, we point out potential strategies that might allow circumvention of cellular or humoral immune responses against the vector or the transgene product.


Assuntos
Terapia Genética , Vetores Genéticos/imunologia , Imunidade Inata , Vírus , Linfócitos T CD8-Positivos/imunologia , Terapia Genética/tendências , Vetores Genéticos/genética , Humanos , Linfócitos T Reguladores/imunologia , Vírus/genética
7.
Hum Gene Ther ; 24(4): 431-42, 2013 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-23461589

RESUMO

In humans adeno-associated virus (AAV)-mediated gene transfer is followed by expansion of AAV capsid-specific T cells, evidence of cell damage, and loss of transgene product expression, implicating immunological rejection of vector-transduced cells, which may be prevented by immunosuppressive drugs. We undertook this study to assess the effect of immunosuppression (IS) used for organ transplantation on immune responses to AAV capsid antigens. Recipients of liver or kidney transplants were tested before and 4 weeks after induction of IS in comparison with matched samples from healthy human adults and an additional cohort with comorbid conditions similar to those of the transplant patients. Our data show that transplant patients and comorbid control subjects have markedly higher frequencies of circulating AAV capsid-specific T cells compared with healthy adults. On average, IS resulted in a reduction of AAV-specific CD4⁺ T cells, whereas numbers of circulating CD8⁺ effector and central memory T cells tended to increase. Independent of the type of transplant or the IS regimens, the trend of AAV capsid-specific T cell responses after drug treatment varied; in some patients responses were unaffected whereas others showed decreases or even pronounced increases, casting doubt on the usefulness of prophylactic IS for AAV vector recipients.


Assuntos
Dependovirus/genética , Terapia de Imunossupressão , Adulto , Idoso , Antígenos Virais/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD4-Positivos/patologia , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/patologia , Capsídeo/imunologia , Estudos de Casos e Controles , Dependovirus/imunologia , Feminino , Vetores Genéticos , Humanos , Tolerância Imunológica , Transplante de Rim , Fígado/imunologia , Fígado/metabolismo , Transplante de Fígado , Masculino , Pessoa de Meia-Idade , Imunologia de Transplantes
8.
Adv Drug Deliv Rev ; 63(8): 671-7, 2011 Jul 18.
Artigo em Inglês | MEDLINE | ID: mdl-21616108

RESUMO

Viral delivery for cancer gene therapy is a promising approach, where traditional radiotherapy or chemotherapy to limit proliferation and movement of cancer cells has met resistance. Based on the new understanding of the biology of the viral vectors, therapeutic viral vectors for cancer gene therapy have been improved for greater safety and efficacy as well as transitioned from being non-replicating to replication-competent. Traditional oncolytic vectors have focused on eliminating tumor growth, while novel vectors simultaneously target epithelial-to-mesenchymal transition (EMT) in cancer cells, which could further prevent and reverse the aggressive tumor progression. In this review, we highlight the illustrative examples of cancer gene therapy in clinical trials as well as preclinical data and include proposals on methods to further enhance the safety and efficacy of oncolytic viral vectors in cancer gene therapy.


Assuntos
Terapia Genética/métodos , Neoplasias/terapia , Vírus Oncolíticos/genética , Animais , Movimento Celular/genética , Técnicas de Transferência de Genes , Terapia Genética/efeitos adversos , Vetores Genéticos , Humanos , Neoplasias/patologia , Replicação Viral , Vírus/genética
9.
Virology ; 379(1): 78-86, 2008 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-18649912

RESUMO

Many studies report the level of total viral DNA in HIV-infected patients, but few studies report the level of integrated DNA. It is important to measure integrated DNA in HIV-infected patients because the information could shed light on the effectiveness of antiretroviral therapy, especially intensified therapy, when viral loads may remain undetectable. In order to develop an integration assay for patient samples, we enhanced the sensitivity of our prior integration assay. To do this, we exploited a technique that we developed, called repetitive sampling, and optimized reaction conditions for rare event detection, rather than large dynamic range. We also designed our primers to match more conserved regions of HIV. The result is a new, sensitive, quantitative assay that allows us to measure integrated DNA in HIV-infected patients. When we applied our integration assay to patient PBMCs, we found that the use of HAART is associated with reduced levels of integrated DNA.


Assuntos
Fármacos Anti-HIV/uso terapêutico , Terapia Antirretroviral de Alta Atividade , Infecções por HIV/tratamento farmacológico , Infecções por HIV/virologia , HIV/efeitos dos fármacos , Integração Viral/efeitos dos fármacos , Primers do DNA/genética , DNA Viral/genética , Humanos , Leucócitos Mononucleares/virologia , Técnicas de Amplificação de Ácido Nucleico/métodos
10.
Heart Vessels ; 20(5): 217-23, 2005 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-16160904

RESUMO

Information of the effect of statin on lipoproteins such as apolipoprotein (apo) A-I, lipoprotein (a) [Lp (a)], or apolipoprotein B levels is limited. This investigation was a crossover study designed to evaluate the efficacy and safety of atorvastatin and simvastatin in patients with hyperlipidemia. Sixty-six patients were involved in the study. Group I consisted of 32 patients, who were first treated with atorvastatin (10 mg) then switched to simvastatin (10 mg). Group II consisted of 34 patients, who were first treated with simvastatin then switched to atorvastatin. Each regimen was used for 3 months (phase I), stopped for 2 months, and then restarted for another 3 months (phase II). Both statins effectively reduced total cholesterol, low-density lipoprotein cholesterol (LDL-C), apo B, and Lp (a) (P < 0.001 in all comparisons). A significant increase in the high-density lipoprotein cholesterol (HDL-C) was noted after both statin treatments (P < 0.05 in all comparisons). Both statins caused an increase in the apo A-I levels, and the extent of changes in apo A-I revealed no difference between the two drugs. Compared to the simvastatin group, there were more patients in the atorvastatin group achieving the National Cholesterol Education Program ATP-III LDL-C goal (P < 0.05) and European LDL-C goal (P < 0.001). Both treatments were well tolerated; no patient was withdrawn from the study. This study demonstrates that both statins can effectively improve lipid profiles in patients with hyperlipidemia. Atorvastatin is more effective in helping patients reach the ATP-III and European LDL-C goals than simvastatin at the same dosage.


Assuntos
Anticolesterolemiantes/uso terapêutico , Ácidos Heptanoicos/uso terapêutico , Hipercolesterolemia/tratamento farmacológico , Lipoproteínas/efeitos dos fármacos , Pirróis/uso terapêutico , Sinvastatina/uso terapêutico , Adulto , Atorvastatina , Estudos Cross-Over , Feminino , Humanos , Masculino , Estatísticas não Paramétricas , Resultado do Tratamento
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