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1.
Blood ; 121(9): 1524-33, 2013 Feb 28.
Artigo em Inglês | MEDLINE | ID: mdl-23264589

RESUMO

We report the safety and tolerability of 87 infusions of lentiviral vector­modified autologous CD4 T cells (VRX496-T; trade name, Lexgenleucel-T) in 17 HIV patients with well-controlled viremia. Antiviral effects were studied during analytic treatment interruption in a subset of 13 patients. VRX496-T was associated with a decrease in viral load set points in 6 of 8 subjects (P = .08). In addition, A → G transitions were enriched in HIV sequences after infusion, which is consistent with a model in which transduced CD4 T cells exert antisense-mediated genetic pressure on HIV during infection. Engraftment of vector-modified CD4 T cells was measured in gut-associated lymphoid tissue and was correlated with engraftment in blood. The engraftment half-life in the blood was approximately 5 weeks, with stable persistence in some patients for up to 5 years. Conditional replication of VRX496 was detected periodically through 1 year after infusion. No evidence of clonal selection of lentiviral vector­transduced T cells or integration enrichment near oncogenes was detected. This is the first demonstration that gene-modified cells can exert genetic pressure on HIV. We conclude that gene-modified T cells have the potential to decrease the fitness of HIV-1 and conditionally replicative lentiviral vectors have a promising safety profile in T cells.


Assuntos
Linfócitos T CD4-Positivos/transplante , Terapia Genética/métodos , Infecções por HIV/terapia , HIV-1/genética , Lentivirus/genética , Oligonucleotídeos Antissenso/farmacologia , Transferência Adotiva/métodos , Adulto , Antivirais/efeitos adversos , Antivirais/metabolismo , Antivirais/farmacologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD4-Positivos/fisiologia , Feminino , Terapia Genética/efeitos adversos , Vetores Genéticos/efeitos adversos , Vetores Genéticos/metabolismo , Vetores Genéticos/farmacologia , Infecções por HIV/genética , Infecções por HIV/imunologia , HIV-1/imunologia , Humanos , Lentivirus/metabolismo , Lentivirus/fisiologia , Masculino , Pessoa de Meia-Idade , Oligonucleotídeos/administração & dosagem , Oligonucleotídeos/genética , Oligonucleotídeos Antissenso/administração & dosagem , Oligonucleotídeos Antissenso/efeitos adversos , Oligonucleotídeos Antissenso/genética , Transdução Genética/métodos , Carga Viral/efeitos dos fármacos , Replicação Viral/genética
2.
Blood ; 117(3): 788-97, 2011 Jan 20.
Artigo em Inglês | MEDLINE | ID: mdl-21030558

RESUMO

In a phase 1/2 two-arm trial, 54 patients with myeloma received autografts followed by ex vivo anti-CD3/anti-CD28 costimulated autologous T cells at day 2 after transplantation. Study patients positive for human leukocyte antigen A2 (arm A, n = 28) also received pneumococcal conjugate vaccine immunizations before and after transplantation and a multipeptide tumor antigen vaccine derived from the human telomerase reverse transcriptase and the antiapoptotic protein survivin. Patients negative for human leukocyte antigen A2 (arm B, n = 26) received the pneumococcal conjugate vaccine only. Patients exhibited robust T-cell recoveries by day 14 with supraphysiologic T-cell counts accompanied by a sustained reduction in regulatory T cells. The median event-free survival (EFS) for all patients is 20 months (95% confidence interval, 14.6-24.7 months); the projected 3-year overall survival is 83%. A subset of patients in arm A (36%) developed immune responses to the tumor antigen vaccine by tetramer assays, but this cohort did not exhibit better EFS. Higher posttransplantation CD4(+) T-cell counts and a lower percentage of FOXP3(+) T cells were associated with improved EFS. Patients exhibited accelerated polyclonal immunoglobulin recovery compared with patients without T-cell transfers. Adoptive transfer of tumor antigen vaccine-primed and costimulated T cells leads to augmented and accelerated cellular and humoral immune reconstitution, including antitumor immunity, after autologous stem cell transplantation for myeloma. This study was registered at www.clinicaltrials.gov as NCT00499577.


Assuntos
Imunoterapia/métodos , Mieloma Múltiplo/terapia , Fragmentos de Peptídeos/imunologia , Vacinação/métodos , Adulto , Idoso , Sequência de Aminoácidos , Antígenos de Neoplasias/imunologia , Terapia Combinada , Exantema/etiologia , Feminino , Transplante de Células-Tronco Hematopoéticas/métodos , Humanos , Imunoterapia/efeitos adversos , Imunoterapia Adotiva , Proteínas Inibidoras de Apoptose , Estimativa de Kaplan-Meier , Masculino , Proteínas Associadas aos Microtúbulos/química , Proteínas Associadas aos Microtúbulos/imunologia , Pessoa de Meia-Idade , Mieloma Múltiplo/imunologia , Mieloma Múltiplo/patologia , Náusea/etiologia , Survivina , Linfócitos T/imunologia , Linfócitos T/transplante , Telomerase/química , Telomerase/imunologia , Transplante Autólogo , Resultado do Tratamento , Vacinação/efeitos adversos , Vômito/etiologia
3.
Nat Med ; 11(11): 1230-7, 2005 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-16227990

RESUMO

Immunodeficiency is a barrier to successful vaccination in individuals with cancer and chronic infection. We performed a randomized phase 1/2 study in lymphopenic individuals after high-dose chemotherapy and autologous hematopoietic stem cell transplantation for myeloma. Combination immunotherapy consisting of a single early post-transplant infusion of in vivo vaccine-primed and ex vivo costimulated autologous T cells followed by post-transplant booster immunizations improved the severe immunodeficiency associated with high-dose chemotherapy and led to the induction of clinically relevant immunity in adults within a month after transplantation. Immune assays showed accelerated restoration of CD4 T-cell numbers and function. Early T-cell infusions also resulted in significantly improved T-cell proliferation in response to antigens that were not contained in the vaccine, as assessed by responses to staphylococcal enterotoxin B and cytomegalovirus antigens (P < 0.05). In the setting of lymphopenia, combined vaccine therapy and adoptive T-cell transfer fosters the development of enhanced memory T-cell responses.


Assuntos
Imunoterapia Adotiva , Linfócitos T/imunologia , Transferência Adotiva , Adulto , Idoso , Feminino , Humanos , Linfopenia/terapia , Masculino , Pessoa de Meia-Idade , Mieloma Múltiplo/terapia , Vacinas Pneumocócicas/uso terapêutico , Resultado do Tratamento , Vacinação
4.
Nat Med ; 8(1): 47-53, 2002 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-11786906

RESUMO

To study the safety and feasibility of T-cell reconstitution in HIV-infected individuals, we adoptively transferred activated autologous CD4+ T cells. Polyclonal peripheral blood CD4+ cells were costimulated ex vivo and subjects were given infusions of up to 3 x 1010 activated CD4+ cells. Dose-dependent increases in CD4+ cell counts and in the CD4:CD8 ratio were observed. Sustained increases in the fraction of cytokine-secreting T cells and decreases in the percentage of CD4+CCR5+ cells were noted in vivo, suggesting enhanced function and resistance to HIV infection. The frequency of CD4+Ki-67+ cells increased whereas CD4+ T cells containing T cell-receptor rearrangement excision circles (TRECs) decreased. These findings indicate that expansion of the peripheral T-cell pool mediated the increase in CD4 counts and suggest that approaches to reconstitute CD4 helper cell activity and decrease CCR5 expression may augment natural immunity to HIV infection.


Assuntos
Transferência Adotiva , Linfócitos T CD4-Positivos/transplante , Infecções por HIV/imunologia , Receptores CCR5/biossíntese , Linfócitos T/imunologia , Adulto , Remoção de Componentes Sanguíneos , Relação CD4-CD8 , Estudos de Viabilidade , Feminino , Infecções por HIV/terapia , Humanos , Ativação Linfocitária , Masculino , Pessoa de Meia-Idade , Transplante Autólogo
5.
Cytotherapy ; 11(7): 923-35, 2009.
Artigo em Inglês | MEDLINE | ID: mdl-19903104

RESUMO

BACKGROUND AIMS: Clinical-scale lymphocyte enrichment from a leukapheresis product has been performed most routinely using costly magnetic bead separation systems that deplete monocytes, but this procedure may leave behind residual beads or antibodies in the enriched cell product. Counterflow centrifugal elutriation has been demonstrated previously to enrich monocytes efficiently for generation of dendritic cells. This study describes a modified elutriation procedure for efficient bead-free economical enrichment of lymphocytes from leukapheresis products from healthy donors and study subjects with human immunodeficiency virus (HIV) infection or malignancy. METHODS: Modified program settings and conditions for the CaridianBCT Elutra device were investigated to optimize lymphocyte enrichment and recovery. Lymphocyte enrichment was measured using a novel approach utilizing cell sizing analysis on a Beckman Coulter Multisizer and confirmed by flow cytometry phenotypic analysis. RESULTS: Efficient enrichment and recovery of lymphocytes from leukapheresis cell products was achieved using modified elutriation settings for flow rate and fraction volume. Elutriation allowed for enrichment of larger numbers of lymphocytes compared with depletion of monocytes by bead adherence, with a trend toward increased lymphocyte purity and yield via elutriation, resulting in a substantial reduction in the cost of enrichment per cell. Importantly, significant lymphocyte enrichment could be accomplished using leukapheresis samples from healthy donors (n=12) or from study subjects with HIV infection (n=15) or malignancy (n=12). CONCLUSIONS: Clinical-scale closed-system elutriation can be performed efficiently for the selective enrichment of lymphocytes for immunotherapy protocols. This represents an improvement in cost, yield and purity over current methods that require the addition of monocyte-depleting beads.


Assuntos
Separação Celular/métodos , Imunoterapia Adotiva/métodos , Linfócitos/metabolismo , Neoplasias/terapia , Separação Celular/instrumentação , Células Cultivadas , Centrifugação , Humanos , Imunoterapia Adotiva/instrumentação , Leucaférese , Linfócitos/citologia , Neoplasias/imunologia
6.
Hum Gene Ther ; 24(3): 245-58, 2013 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-23360514

RESUMO

Since HIV requires CD4 and a co-receptor, most commonly C-C chemokine receptor 5 (CCR5), for cellular entry, targeting CCR5 expression is an attractive approach for therapy of HIV infection. Treatment of CD4(+) T cells with zinc-finger protein nucleases (ZFNs) specifically disrupting chemokine receptor CCR5 coding sequences induces resistance to HIV infection in vitro and in vivo. A chimeric Ad5/F35 adenoviral vector encoding CCR5-ZFNs permitted efficient delivery and transient expression following anti-CD3/anti-CD28 costimulation of T lymphocytes. We present data showing CD3/CD28 costimulation substantially improved transduction efficiency over reported methods for Ad5/F35 transduction of T lymphocytes. Modifications to the laboratory scale process, incorporating clinically compatible reagents and methods, resulted in a robust ex vivo manufacturing process capable of generating >10(10) CCR5 gene-edited CD4+ T cells from healthy and HIV+ donors. CD4+ T-cell phenotype, cytokine production, and repertoire were comparable between ZFN-modified and control cells. Following consultation with regulatory authorities, we conducted in vivo toxicity studies that showed no detectable ZFN-specific toxicity or T-cell transformation. Based on these findings, we initiated a clinical trial testing the safety and feasibility of CCR5 gene-edited CD4+ T-cell transfer in study subjects with HIV-1 infection.


Assuntos
Enzimas de Restrição do DNA/genética , Vetores Genéticos/normas , Infecções por HIV/genética , Infecções por HIV/imunologia , Receptores CCR5/genética , Dedos de Zinco/genética , Adenovírus Humanos/genética , Transferência Adotiva , Animais , Antígenos CD28/imunologia , Complexo CD3/imunologia , Enzimas de Restrição do DNA/metabolismo , Feminino , Vetores Genéticos/administração & dosagem , Vetores Genéticos/efeitos adversos , Vetores Genéticos/genética , Infecções por HIV/terapia , Humanos , Ativação Linfocitária/imunologia , Masculino , Camundongos , Fenótipo , Receptores CCR5/imunologia , Linfócitos T/imunologia , Linfócitos T/metabolismo , Transdução Genética/métodos , Transdução Genética/normas , Transplante Heterólogo
7.
Clin Cancer Res ; 15(13): 4499-507, 2009 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-19509133

RESUMO

PURPOSE: Previously, we showed that adoptive transfer of in vivo vaccine-primed and ex vivo (anti-CD3/anti-CD28) costimulated autologous T cells (ex-T) at day +12 after transplant increased CD4 and CD8 T-cell counts at day +42 and augmented vaccine-specific immune responses in patients with myeloma. Here, we investigated the safety and kinetics of T-cell recovery after infusing ex-T at day +2 after transplant. EXPERIMENTAL DESIGN: In this phase I/II two-arm clinical trial, 50 patients with myeloma received autografts after high-dose melphalan followed by infusions of ex-T at day +2 after transplant. Patients also received pretransplant and posttransplant immunizations using a pneumococcal conjugate vaccine only (arm B; n = 24) or the pneumococcal conjugate vaccine plus an HLA-A2-restricted microltipeptide vaccine for HLA-A2(+) patients (arm A; n = 26). RESULTS: The mean number of T cells infused was 4.26 x 10(10) (range, 1.59-5.0). At day 14 after transplant, the median CD3, CD4, and CD8 counts were 4,198, 1,545, and 2,858 cells/microL, respectively. Interleukin (IL)-6 and IL-15 levels increased early after transplant and IL-15 levels correlated significantly to day 14 T-cell counts. Robust vaccine-specific B- and T-cell responses were generated. T-cell infusions were well tolerated with no effect on hematopoietic recovery. Eight patients (16%) developed a T-cell "engraftment syndrome" characterized by diarrhea and fever that was clinically and histopathologically indistinguishable from grade 1 to 3 acute graft-versus-host disease (GVHD) of the gastrointestinal tract (seven patients) and/or grade 1 to 2 cutaneous GVHD (four patients). CONCLUSIONS: Adoptive T-cell transfers achieve robust T-cell recovery early after transplant and induce moderate-to-severe autologous GVHD in a subset of patients.


Assuntos
Doença Enxerto-Hospedeiro/reabilitação , Imunoterapia Adotiva , Mieloma Múltiplo/terapia , Recuperação de Função Fisiológica/imunologia , Linfócitos T/transplante , Adulto , Idoso , Algoritmos , Células Cultivadas , Feminino , Doença Enxerto-Hospedeiro/imunologia , Antígeno HLA-A2/metabolismo , Humanos , Imunoterapia Adotiva/efeitos adversos , Imunoterapia Adotiva/métodos , Ativação Linfocitária/imunologia , Ativação Linfocitária/fisiologia , Masculino , Melfalan/uso terapêutico , Pessoa de Meia-Idade , Mieloma Múltiplo/imunologia , Agonistas Mieloablativos/uso terapêutico , Síndrome , Transplante Autólogo
8.
Clin Immunol ; 111(3): 262-74, 2004 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-15183147

RESUMO

We have previously shown that adoptive transfer of in vitro CD3/CD28 activated autologous CD4(+) T cells results in increased CD4 counts and CD4/CD8 ratios in HIV+ subjects. In this report, analysis of variable beta (Vbeta) chain T cell receptor (TCR) repertoire showed that CD3/CD28 stimulation was able to increase polyclonality within skewed spectra types in vitro. In vivo, two of eight subjects showed increase in TCR diversity and importantly, in no subject did a highly skewed in vivo repertoire emerge. Measurement of proliferative response to alloantigen showed increases following infusions. Response to pharmacological stimulus and lectin via Interferon-gamma ELISpot assay showed increases in a subset of subjects following infusions. However, interferon-gamma response to HIV antigens and peptides declined concurrent with stable or diminishing latent infectious viral load in CD4(+) T cells. These data provide further evidence that adoptive transfer of activated autologous CD4(+) T cells can augment the immune system.


Assuntos
Antígenos CD28/imunologia , Complexo CD3/imunologia , Linfócitos T CD4-Positivos/imunologia , Infecções por HIV/terapia , Imunoterapia Adotiva , Adulto , Feminino , Genes Codificadores da Cadeia beta de Receptores de Linfócitos T/imunologia , Infecções por HIV/imunologia , Humanos , Interferon gama/metabolismo , Teste de Cultura Mista de Linfócitos , Masculino , Pessoa de Meia-Idade
9.
Blood ; 102(6): 2004-13, 2003 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-12763934

RESUMO

We explored the feasibility and toxicity of administering escalating doses of anti-CD3/CD28 ex vivo costimulated T cells as a therapeutic adjunct for patients with relapsed, refractory, or chemotherapy-resistant, aggressive non-Hodgkin lymphoma (NHL) following high-dose chemotherapy and CD34+-selected hematopoietic cell transplantation (HCT). Sixteen patients had infusions on day 14 after HCT of autologous T cells that had been stimulated using beads coated with anti-CD3 and anti-CD28 monoclonal antibodies. At baseline, the subjects had severe quantitative and functional T-cell impairments. The culture procedure partially reversed impaired cytokine responsiveness in T cells in vitro and in vivo. Transient dose-dependent infusion toxicities were observed. There was a rapid reconstitution of lymphocytes; however, there were persistent defects in CD4 T cells. Most interestingly, 5 patients had a delayed lymphocytosis between day 30 and day 120 after HCT. Maximal clinical responses included 5 patients with a complete response (CR), 7 patients with a partial response (PR), and 4 patients with stable disease. At a median follow-up of 33 months (range, 26-60 months), 5 patients are alive with stable or relapsed disease and 3 patients remain in CR. In conclusion, this phase 1 trial demonstrates that adoptive transfer of autologous costimulated T cells (1) is feasible in heavily pretreated patients with advanced NHL, (2) is associated with a rapid recovery of lymphocyte counts, (3) reverses cytokine activation deficits in vitro, and (4) is associated with delayed lymphocytosis in a subset of patients.


Assuntos
Transferência Adotiva , Transplante de Células-Tronco Hematopoéticas , Linfocitose/imunologia , Linfoma não Hodgkin/terapia , Linfócitos T/transplante , Adolescente , Transferência Adotiva/efeitos adversos , Adulto , Idoso , Antígenos CD34/análise , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Carmustina/uso terapêutico , Terapia Combinada , Ciclofosfamida/uso terapêutico , Citarabina/uso terapêutico , Etoposídeo/uso terapêutico , Feminino , Seguimentos , Células-Tronco Hematopoéticas/química , Humanos , Linfoma não Hodgkin/tratamento farmacológico , Linfoma não Hodgkin/imunologia , Masculino , Pessoa de Meia-Idade , Recidiva , Indução de Remissão
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