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BACKGROUND AND AIMS: Intestinal epithelial cell (IEC) damage is a hallmark of celiac disease (CeD); however, its role in gluten-dependent T-cell activation is unknown. We investigated IEC-gluten-T cell interactions in organoid monolayers expressing human MHC class II (HLA-DQ2.5), which facilitates gluten antigen recognition by CD4+ T cells in CeD. METHODS: Epithelial MHC class II (MHCII) was determined in active and treated CeD, and in non-immunized and gluten-immunized DR3-DQ2.5 transgenic mice, lacking mouse MHCII molecules. Organoid monolayers from DR3-DQ2.5 mice were treated with or without IFN-γ, and MHCII expression was evaluated by flow cytometry. Organoid monolayers and CD4+ T cell co-cultures were incubated with gluten, pre-digested, or not by elastase-producing Pseudomonas aeruginosa or its lasB mutant. T cell function was assessed based on proliferation, expression of activation markers, and cytokine release in the co-culture supernatants. RESULTS: Active CeD patients and gluten-immunized DR3-DQ2.5 mice demonstrated epithelial MHCII expression. Organoid monolayers derived from gluten-immunized DR3-DQ2.5 mice expressed MHCII, which was upregulated by IFN-γ. In organoid monolayer-T cell co-cultures, gluten increased the proliferation of CD4+ T cells, expression of T cell activation markers, and the release of IL-2, IFN-γ, and IL-15 in co-culture supernatants. Gluten metabolized by P. aeruginosa, but not the lasB mutant, enhanced CD4+ T cell proliferation and activation. CONCLUSIONS: Gluten antigens are efficiently presented by MHCII-expressing IECs, resulting in the activation of gluten-specific CD4+ T cells, which is enhanced by gluten pre-digestion with microbial elastase. Therapeutics directed at IECs may offer a novel approach for modulating both adaptive and innate immunity in CeD patients.
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Chronic infection with human CMV may contribute to poor vaccine efficacy in older adults. We assessed the effects of CMV serostatus on Ab quantity and quality, as well as cellular memory recall responses, after two and three SARS-CoV-2 mRNA vaccine doses, in older adults in assisted living facilities. CMV serostatus did not affect anti-Spike and anti-receptor-binding domain IgG Ab levels, nor neutralization capacity against wild-type or ß variants of SARS-CoV-2 several months after vaccination. CMV seropositivity altered T cell expression of senescence-associated markers and increased effector memory re-expressing CD45RA T cell numbers, as has been previously reported; however, this did not impact Spike-specific CD4+ T cell memory recall responses. CMV-seropositive individuals did not have a higher incidence of COVID-19, although prior infection influenced humoral immunity. Therefore, CMV seropositivity may alter T cell composition but does not impede the durability of humoral protection or cellular memory responses after SARS-CoV-2 mRNA vaccination in older adults.
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COVID-19 , Infecções por Citomegalovirus , Humanos , Idoso , Vacinas contra COVID-19 , Citomegalovirus , SARS-CoV-2 , COVID-19/prevenção & controle , Anticorpos , Vacinação , Vacinas de mRNARESUMO
Immunogenicity of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) bivalent mRNA-1273.214 vaccine (Original/Omicron B.1.1.529 [BA.1]) is underreported in vulnerable older adults in congregate care settings. In residents of 26 long-term care and retirement homes in Ontario, Canada, humoral (i.e., serum anti-spike and anti-receptor binding domain [anti-RBD]) IgG and IgA antibodies and live SARS-CoV-2 neutralization) and cellular (i.e., CD4+ and CD8+ activation-induced marker spike-specific T cell memory) responses were assessed 7-120 days postvaccination with four monovalent mRNA vaccines (n = 494) or subsequent bivalent mRNA-1273.214 vaccination (fifth vaccine) (n = 557). Within 4 months, anti-spike and anti-RBD antibody levels were similar after monovalent and bivalent vaccination in infection-naïve individuals. Hybrid immunity (i.e., vaccination and natural infection) generally increased humoral responses. After bivalent vaccination, compared to monovalent vaccination, residents with hybrid immunity had elevated anti-spike and anti-RBD IgG and IgA antibodies. Omicron BA.1 antibody-mediated neutralization, and CD8+ T cell memory responses to the Omicron BA.1 spike protein, were also higher after bivalent vaccination. Humoral and cellular responses were, therefore, noninferior within 4 months of bivalent mRNA-1273.214 vaccination compared to monovalent mRNA vaccination. Waning of humoral but not cellular immunity was particularly evident in individuals without hybrid immunity. Continued monitoring of vaccine-associated and hybrid immunity against emerging Omicron variants of concern is necessary to assess longevity of protection.
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COVID-19 , Assistência de Longa Duração , Humanos , Idoso , Ontário , Aposentadoria , SARS-CoV-2/genética , COVID-19/prevenção & controle , Vacinas de mRNA , Vacinação , Estudos de Coortes , Imunoglobulina A , Imunoglobulina G , Anticorpos Antivirais , Anticorpos NeutralizantesRESUMO
Immunomodulatory cytokines can alter the tumor microenvironment and promote tumor eradication. Interleukin (IL)-27 is a pleiotropic cytokine that has potential to augment anti-tumor immunity while also facilitating anti-myeloma activity. We engineered human T cells to express a recombinant single-chain (sc)IL-27 and a synthetic antigen receptor targeting the myeloma antigen, B-cell maturation antigen, and evaluated the anti-tumor function of T cells bearing scIL-27 in vitro and in vivo. We discovered that T cells bearing scIL-27 sustained anti-tumor immunity and cytotoxicity yet manifested a profound reduction in pro-inflammatory cytokines granulocyte-macrophage colony-stimulating factor and tumor necrosis factor alpha. IL-27-expressing T cells therefore present a potential avenue to avert treatment-related toxicities commonly associated with engineered T-cell therapy due to the reduced pro-inflammatory cytokine profile.
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Interleucina-27 , Neoplasias , Humanos , Linfócitos T , Citocinas/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Neoplasias/terapia , Interleucinas , Interleucina-2 , Microambiente TumoralRESUMO
B-cell maturation antigen (BCMA) is a clinically validated target for multiple myeloma. T-cell engineered with chimeric antigen receptors (CARs) directed against BCMA have demonstrated robust therapeutic activity in clinical trials, but toxicities remain a significant concern for a subset of patients, supporting continued investigation of other engineered T-cell platforms that may offer equal efficacy with an improved toxicity profile. The authors recently described a BCMA-specific, T-cell-centric synthetic antigen receptor, the T-cell antigen coupler (TAC) receptor, that can be used to engineer T-cell with robust anti-myeloma activity. Here the authors describe the creation of a fully humanized BCMA-specific TAC receptor. Single-chain variable fragments (scFvs) were developed from BCMA-specific F(ab)s that were identified in a fully human phage display library. Twenty-four configurations of the F(ab)s were evaluated in a medium-throughput screening using primary T-cell, and a single F(ab), TRAC 3625, emerged as the most robust following in vitro and in vivo evaluation. An optimized BCMA-specific TAC receptor was developed through iterations of the BCMA-TAC design that evaluated a next-generation TAC scaffold sequence, different domains connecting the TAC to the 3625 scFv and different orientations of the TRAC 3625 heavy and light variable regions.
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Mieloma Múltiplo , Linfócitos T , Humanos , Mieloma Múltiplo/terapia , Antígeno de Maturação de Linfócitos B , Imunoterapia Adotiva , Receptores de Antígenos de Linfócitos TRESUMO
BACKGROUND AIMS: T cells engineered with synthetic receptors have delivered powerful therapeutic results for patients with relapsed/refractory hematologic malignancies. The authors have recently described the T-cell antigen coupler (TAC) receptor, which co-opts the endogenous T-cell receptor (TCR) and activates engineered T cells in an HLA-independent manner. Here the authors describe the evolution of a next-generation TAC receptor with a focus on developing a TAC-engineered T cell for multiple myeloma. METHODS: To optimize the TAC scaffold, the authors employed a bona fide antigen-binding domain derived from the B-cell maturation antigen-specific monoclonal antibody C11D5.3, which has been used successfully in the clinic. The authors first tested humanized versions of the UCHT1 domain, which is used by the TAC to co-opt the TCR. The authors further discovered that the signal peptide affected surface expression of the TAC receptor. Higher density of the TAC receptor enhanced target binding in vitro, which translated into higher levels of Lck at the immunological synapse and stronger proliferation when only receptor-ligand interactions were present. RESULTS: The authors observed that the humanized UCHT1 improved surface expression and in vivo efficacy. Using TAC T cells derived from both healthy donors and multiple myeloma patients, the authors determined that despite the influence of receptor density on early activation events and effector function, receptor density did not impact late effector functions in vitro, nor did the receptor density affect in vivo efficacy. CONCLUSIONS: The modifications to the TAC scaffold described herein represent an important step in the evolution of this technology, which tolerates a range of expression levels without impacting therapeutic efficacy.
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Antígeno de Maturação de Linfócitos B , Mieloma Múltiplo , Humanos , Imunoterapia Adotiva , Mieloma Múltiplo/terapia , Recidiva Local de Neoplasia , Receptores de Antígenos de Linfócitos T/genética , Linfócitos TRESUMO
Clinical and experimental studies have shown that estradiol (E2) confers protection against HIV and other sexually transmitted infections. Here, we investigated the underlying mechanism. Better protection in E2-treated mice, immunized against genital HSV-2, coincided with earlier recruitment and higher proportions of Th1 and Th17 effector cells in the vagina post-challenge, compared to placebo-treated controls. Vaginal APCs isolated from E2-treated mice induced 10-fold higher Th17 and Th1 responses, compared to APCs from progesterone-treated, placebo-treated, and estradiol-receptor knockout mice in APC-T cell co-cultures. CD11c+ DCs in the vagina were the predominant APC population responsible for priming these Th17 responses, and a potent source of IL-6 and IL-1ß, important factors for Th17 differentiation. Th17 responses were abrogated in APC-T cell co-cultures containing IL-1ß KO, but not IL-6 KO vaginal DCs, showing that IL-1ß is a critical factor for Th17 induction in the genital tract. E2 treatment in vivo directly induced high expression of IL-1ß in vaginal DCs, and addition of IL-1ß restored Th17 induction by IL-1ß KO APCs in co-cultures. Finally, we examined the role of IL-17 in anti-HSV-2 memory T cell responses. IL-17 KO mice were more susceptible to intravaginal HSV-2 challenge, compared to WT controls, and vaginal DCs from these mice were defective at priming efficient Th1 responses in vitro, indicating that IL-17 is important for the generation of efficient anti-viral memory responses. We conclude that the genital mucosa has a unique microenvironment whereby E2 enhances CD4+ T cell anti-viral immunity by priming vaginal DCs to induce Th17 responses through an IL-1-dependent pathway.
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Linfócitos T CD4-Positivos/imunologia , Células Dendríticas/imunologia , Estradiol/imunologia , Células Th17/imunologia , Vagina/imunologia , Animais , Técnicas de Cocultura , Citocinas/biossíntese , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática , Estradiol/farmacologia , Feminino , Citometria de Fluxo , Herpes Genital/imunologia , Herpesvirus Humano 2/imunologia , Interleucina-1/imunologia , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Vagina/virologiaRESUMO
[This corrects the article DOI: 10.1371/journal.ppat.1005589.].
RESUMO
Effector T cells (TEFF) are a barrier to booster vaccination because they can rapidly kill Ag-bearing APCs before memory T cells are engaged. We report in this study that i.v. delivery of rhabdoviral vectors leads to direct infection of follicular B cells in the spleen, where the earliest evidence of secondary T cell responses was observed. This allows booster immunizations to rapidly expand CD8(+) central memory T cells (TCM) during the acute phase of the primary response that is dominated by TEFF Interestingly, although the ablation of B cells before boosting with rhabdoviral vectors diminishes the expansion of memory T cells, B cells do not present Ags directly. Instead, depletion of CD11c(+) dendritic cells abrogates secondary T cell expansion, suggesting that virus-infected follicular B cells may function as an Ag source for local DCs to subsequently capture and present the Ag. Because TCM are located within B cell follicles in the spleen whereas TEFF cannot traffic through follicular regions, Ag production and presentation by follicular APCs represent a unique mechanism to secure engagement of TCM during an ongoing effector response. Our data offer insights into novel strategies for rapid expansion of CD8(+) T cells using prime-boost vaccines by targeting privileged sites for Ag presentation.
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Apresentação de Antígeno/imunologia , Linfócitos B/imunologia , Linfócitos T CD8-Positivos/imunologia , Células Dendríticas Foliculares/imunologia , Baço/citologia , Baço/imunologia , Vírus da Estomatite Vesicular Indiana/imunologia , Vacinas Virais/imunologia , Animais , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos KnockoutRESUMO
CD4(+) T cell help is critical for optimal CD8(+) T cell memory differentiation and maintenance in many experimental systems. In addition, many reports have identified reduced primary CD8(+) T cell responses in the absence of CD4(+) T cell help, which often coincides with reduced Ag or pathogen clearance. In this study, we demonstrate that absence of CD4(+) T cells at the time of adenovirus vector immunization of mice led to immediate impairments in early CD8(+) T cell functionality and differentiation. Unhelped CD8(+) T cells exhibited a reduced effector phenotype, decreased ex vivo cytotoxicity, and decreased capacity to produce cytokines. This dysfunctional state was imprinted within 3 d of immunization. Unhelped CD8(+) T cells expressed elevated levels of inhibitory receptors and exhibited transcriptomic exhaustion and anergy profiles by gene set enrichment analysis. Dysfunctional, impaired effector differentiation also occurred following immunization of CD4(+) T cell-deficient mice with a poxvirus vector. This study demonstrates that following priming with viral vectors, CD4(+) T cell help is required to promote both the expansion and acquisition of effector functions by CD8(+) T cells, which is accomplished by preventing immediate dysfunction.
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Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Vacinas/imunologia , Adenoviridae/genética , Animais , Linfócitos T CD8-Positivos/fisiologia , Citocinas/imunologia , Citotoxicidade Imunológica , Feminino , Regulação da Expressão Gênica , Vetores Genéticos , Imunização , Memória Imunológica , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BLRESUMO
Background: Elderly long-term care residents often exhibit a myriad of risk factors for immune dysfunction, including chronic inflammation and multiple comorbid conditions, which undoubtedly contribute to their enhanced susceptibility to infection. Hence, understanding the factors required for optimal vaccine responsiveness is critical. Methods: We examined 187 elderly nursing home residents (aged 80-102 years) and 50 community-dwelling seniors (aged 60-75 years) immunized with the live-attenuated varicella-zoster virus (VZV) vaccine. Specifically, we examined whether vaccine responsiveness was associated with serum C-reactive protein (CRP), tumor necrosis factor, interleukin 1ß, 6, and 10, leukocyte telomere length, chronic disease status, and frailty. Results: Elderly participants had significantly higher levels of CRP, tumor necrosis factor, and interleukin 6 and shorter leukocyte telomere length. Vaccine responsiveness was inversely related to the CRP level in elderly participants, but not seniors, and those with congestive heart failure were less likely to achieve a 2-fold response (odds ratio, 0.08). The latter relationship is probably due to immunosenescence, because heart failure was associated with increased senescent CD4+ T cells, and reduced naive and effector and central memory CD8+ T cells. Conclusions: In summary, these data improve our understanding of vaccine responsiveness for those in long-term care, suggesting that certain risk factors are associated with a greater likelihood of vaccine failure.
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Proteína C-Reativa/análise , Insuficiência Cardíaca/epidemiologia , Vacina contra Herpes Zoster/imunologia , Linfócitos T/imunologia , Idoso , Idoso de 80 Anos ou mais , Canadá , Citocinas/sangue , Feminino , Herpes Zoster/prevenção & controle , Herpesvirus Humano 3 , Instituição de Longa Permanência para Idosos , Humanos , Imunidade Celular , Imunossenescência , Modelos Lineares , Modelos Logísticos , Assistência de Longa Duração , Masculino , Pessoa de Meia-Idade , Casas de Saúde , Telômero/ultraestruturaRESUMO
Immunosuppressive mechanisms within the tumor microenvironment have emerged as a major impediment to cancer immunotherapy. While a broad range of secreted factors, receptors/ligands, and cell populations have been described that contribute to the immunosuppression, the involvement of these processes in immune evasion by tumors is typically considered to be an intrinsic property of the tumor. Evidence is now emerging that the processes underlying immune suppression within the tumor are, in fact, triggered by immune attack and reflect a dynamic interplay between the tumor and the host's immune system. The term adaptive resistance has been coined to describe the induction of immune suppressive pathways in the tumor following active attack on the tumor. Adaptive resistance is a scalable process where the magnitude of immune suppression matches the magnitude of the immune attack; the net balance between suppression and attack determines the durability of the anti-tumor response and tumor outcome. In this chapter, we will examine the data supporting adaptive resistance and the opposing roles of T cells in simultaneously promoting both anti-tumor immunity and immune suppression within the tumor microenvironment. The clinical implications of adaptive resistance in the design and application of immunotherapeutic strategies is also discussed.
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Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Imunidade Celular , Imunoterapia/métodos , Neoplasias/imunologia , Neoplasias/terapia , Animais , Linfócitos T CD4-Positivos/patologia , Linfócitos T CD8-Positivos/patologia , Humanos , Neoplasias/patologiaRESUMO
BACKGROUND: Whether a candidate tuberculosis vaccine induces clinically relevant protective T-cell repertoires in humans will not be known until the completion of costly efficacy clinical trials. METHODS: We have developed an integrated immunologic approach to investigate the clinical relevance of T cells induced by a novel tuberculosis vaccine in a phase 1 trial. This approach consists of screening for likely dominant T-cell epitopes, establishing antigen-specific memory T-cell lines for identifying CD8+ and CD4+ T-cell epitopes, determining the ability of vaccine-induced T cells to inhibit mycobacterial growth in infected cells, and examining the genetic diversity of HLA recognition and the clinical relevance of identified T-cell epitopes. RESULTS: A single-dose immunization in BCG-primed adults with an adenovirus-based tuberculosis vaccine elicits a repertoire of memory T cells capable of recognizing multiple Ag85A epitopes. These T cells are polyfunctional and cytotoxic and can inhibit mycobacterial growth in infected target cells. Some identified T-cell epitopes are promiscuous and recognizable by the common HLA alleles. These epitopes are clinically relevant to the epitopes identified in people with latent Mycobacterium tuberculosis infection and treated patients with tuberculosis. CONCLUSIONS: These data support further clinical development of this candidate vaccine. Our approach helps fill the gap in clinical tuberculosis vaccine development.
Assuntos
Adenoviridae/genética , Portadores de Fármacos , Mycobacterium tuberculosis/imunologia , Linfócitos T/imunologia , Vacinas contra a Tuberculose/imunologia , Tuberculose/prevenção & controle , Aciltransferases/genética , Aciltransferases/imunologia , Adulto , Antígenos de Bactérias/genética , Antígenos de Bactérias/imunologia , Epitopos de Linfócito T/genética , Epitopos de Linfócito T/imunologia , Humanos , Mycobacterium tuberculosis/genética , Tuberculose/imunologia , Vacinas contra a Tuberculose/administração & dosagem , Vacinas contra a Tuberculose/genética , Vacinas de Subunidades Antigênicas/administração & dosagem , Vacinas de Subunidades Antigênicas/genética , Vacinas de Subunidades Antigênicas/imunologia , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/genética , Vacinas Sintéticas/imunologiaRESUMO
BACKGROUND: Little is known about the immunogenicity of live-attenuated Oka/Merck varicella zoster virus (VZV)-containing vaccine (hereafter, "varicella vaccine") in frail nursing homes residents nor about immune phenotypes associated with a response. METHODS: A cohort of 190 frail nursing home residents aged 80-102 years and a cohort of 50 community-dwelling seniors aged 60-75 years (a comparison group) received varicella vaccine. Interferon γ (IFN-γ) enzyme-linked immunospot assays were performed before and 6 weeks after vaccination. Cellular markers of immunosenescence were measured in the nursing home elderly. RESULTS: The average number of IFN-γ spot-forming cells at baseline was significantly lower in the elderly nursing home residents than in the community-dwelling seniors. However, following vaccination, the VZV immune response increased in both cohorts, and no difference was noted in the fold difference of the response between the 2 cohorts. Upon further examination of the elderly nursing home residents, we found that higher frequencies of regulatory T cells and cytomegalovirus-specific CD4+ T cells correlated negatively with the magnitude of VZV-specific responses. CONCLUSIONS: The Oka/Merck varicella vaccine induces VZV immunity in elderly nursing home residents that is similar to that produced in community-dwelling seniors. CLINICAL TRIALS REGISTRATION: NCT01328548.
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Vacina contra Varicela/imunologia , Interferon gama/metabolismo , Leucócitos Mononucleares/imunologia , Casas de Saúde , Idoso de 80 Anos ou mais , Vacina contra Varicela/administração & dosagem , Estudos de Coortes , ELISPOT , Feminino , Humanos , MasculinoRESUMO
Ligands for the NKG2D receptor are overexpressed on tumors, making them interesting immunotherapy targets. To assess the tumoricidal properties of T cells directed to attack NKG2D ligands, we engineered murine T cells with two distinct NKG2D-based chimeric antigen receptors (CARs): (i) a fusion between the NKG2D receptor and the CD3ζ chain and (ii) a conventional second-generation CAR, where the extracellular domain of NKG2D was fused to CD28 and CD3ζ. To enhance the CAR surface expression, we also engineered T cells to coexpress DAP10. In vitro functionality and surface expression levels of all three CARs was greater in BALB/c T cells than C57BL/6 T cells, indicating strain-specific differences. Upon adoptive transfer of NKG2D-CAR-T cells into syngeneic animals, we observed significant clinical toxicity resulting in morbidity and mortality. The severity of these toxicities varied between the CAR configurations and paralleled their in vitro NKG2D surface expression. BALB/c mice were more sensitive to these toxicities than C57BL/6 mice, consistent with the higher in vitro functionality of BALB/c T cells. Treatment with cyclophosphamide prior to adoptive transfer exacerbated the toxicity. We conclude that while NKG2D ligands may be useful targets for immunotherapy, the pursuit of NKG2D-based CAR-T cell therapies should be undertaken with caution.
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Citotoxicidade Imunológica , Proteínas Recombinantes de Fusão , Transferência Adotiva , Animais , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Ciclofosfamida/farmacologia , Citocinas/metabolismo , Modelos Animais de Doenças , Feminino , Expressão Gênica , Vetores Genéticos/genética , Imunoterapia Adotiva , Ligantes , Camundongos , Subfamília K de Receptores Semelhantes a Lectina de Células NK/metabolismo , Neoplasias/genética , Neoplasias/imunologia , Neoplasias/patologia , Neoplasias/terapia , Pneumonia/imunologia , Pneumonia/metabolismo , Pneumonia/patologia , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/metabolismo , Receptores Imunológicos/genética , Receptores Imunológicos/metabolismo , Retroviridae/genética , Linfócitos T/imunologia , Linfócitos T/metabolismo , Condicionamento Pré-TransplanteRESUMO
Rapid boosting of memory CD8(+) T cells (TM) is essential in cancer immunotherapy and the control of certain infectious diseases. However, effector T cells (TE) are a barrier to booster vaccination because they can rapidly kill antigen-bearing antigen-presenting cells (APCs) before TM are engaged. We demonstrate that viral-vectored vaccines delivered by B cells elicit robust TM expansion in the presence of TE, enabling booster immunizations to bypass TE-mediated negative feedback regulation. Our data indicate that viral vector-loaded B cells home to the follicular regions in secondary lymphoid organs, which are anatomically separated from TE and in close proximity to TM. The B cells, however, do not serve as APCs in this area. Rather, classic CD11c(+) dendritic cells serve to stimulate the secondary CD8(+) T-cell response. Our data reveal that B cells represent a novel and readily accessible delivery system that can effectively engage secondary CD8(+) T-cell activation for prime-boost strategies.
Assuntos
Adenoviridae , Linfócitos B/transplante , Linfócitos T CD8-Positivos/imunologia , Vetores Genéticos/administração & dosagem , Memória Imunológica , Imunoterapia Adotiva/métodos , Aceleração , Adenoviridae/genética , Animais , Linfócitos B/metabolismo , Células Cultivadas , Técnicas de Transferência de Genes , Imunização Secundária/métodos , Memória Imunológica/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Vacinação/métodosRESUMO
The rhabdovirus Maraba has recently been characterized as a potent oncolytic virus. In the present study, we engineered an attenuated Maraba strain, defined as MG1, to express a melanoma-associated tumor antigen. Its ability to mount an antitumor immunity was evaluated in tumor-free and melanoma tumor-bearing mice. Alone, the MG1 vaccine appeared insufficient to prime detectable adaptive immunity against the tumor antigen. However, when used as a boosting vector in a heterologous prime-boost regimen, MG1 vaccine rapidly generated strong antigen-specific T-cell immune responses. Once applied for treating syngeneic murine melanoma tumors, our oncolytic prime-boost vaccination protocol involving Maraba MG1 dramatically extended median survival and allowed complete remission in more than 20% of the animals treated. This work describes Maraba virus MG1 as a potent vaccine vector for cancer immunotherapy displaying both oncolytic activity and a remarkable ability to boost adaptive antitumor immunity.
Assuntos
Vetores Genéticos/genética , Vírus Oncolíticos/genética , Rhabdoviridae/genética , Animais , Linfócitos T CD8-Positivos/imunologia , Vacinas Anticâncer/imunologia , Efeito Citopatogênico Viral , Feminino , Expressão Gênica , Vetores Genéticos/imunologia , Imunização Secundária/métodos , Oxirredutases Intramoleculares/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/mortalidade , Neoplasias Pulmonares/secundário , Neoplasias Pulmonares/terapia , Melanoma Experimental , Camundongos , Neoplasias/genética , Neoplasias/imunologia , Neoplasias/mortalidade , Neoplasias/patologia , Neoplasias/terapia , Vírus Oncolíticos/imunologia , Rhabdoviridae/imunologia , Resultado do Tratamento , Vesiculovirus/genética , Vesiculovirus/imunologia , Tropismo ViralRESUMO
Despite clear evidence of immunogenicity, cancer vaccines only provide a modest clinical benefit. To evaluate the mechanisms that limit tumor regression following vaccination, we have investigated the weak efficacy of a highly immunogenic experimental vaccine using a murine melanoma model. We discovered that the tumor adapts rapidly to the immune attack instigated by tumor-specific CD8+ T cells in the first few days following vaccination, resulting in the upregulation of a complex set of biological networks, including multiple immunosuppressive processes. This rapid adaptation acts to prevent sustained local immune attack, despite continued infiltration by increasing numbers of tumor-specific T cells. Combining vaccination with adoptive transfer of tumor-specific T cells produced complete regression of the treated tumors but did not prevent the adaptive immunosuppression. In fact, the adaptive immunosuppressive pathways were more highly induced in regressing tumors, commensurate with the enhanced level of immune attack. Examination of tumor infiltrating T-cell functionality revealed that the adaptive immunosuppression leads to a progressive loss in T-cell function, even in tumors that are regressing. These novel observations that T cells produced by therapeutic intervention can instigate a rapid adaptive immunosuppressive response within the tumor have important implications for clinical implementation of immunotherapies.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Imunoterapia , Neoplasias/imunologia , Neoplasias/terapia , Imunidade Adaptativa/genética , Adenovírus Humanos/genética , Animais , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/imunologia , Linfócitos T CD8-Positivos/metabolismo , Vacinas Anticâncer/imunologia , Feminino , Vetores Genéticos/genética , Vetores Genéticos/imunologia , Imunomodulação/genética , Imunomodulação/imunologia , Imunoterapia Adotiva , Interferon gama/imunologia , Interferon gama/metabolismo , Oxirredutases Intramoleculares/genética , Oxirredutases Intramoleculares/imunologia , Linfócitos do Interstício Tumoral/imunologia , Camundongos , Recidiva Local de Neoplasia , Neoplasias/genética , Neoplasias/patologia , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Carga Tumoral/genética , Carga Tumoral/imunologia , Vacinação , Vacinas SintéticasRESUMO
We used SILAC (stable isotope labelling of amino acids in cell culture) and high-throughput quantitative MS mass spectrometry to analyse the protein composition of highly purified WT wild type adenoviruses, mutant adenoviruses lacking an internal protein component (protein V) and recombinant adenoviruses of the type commonly used in gene therapy, including one virus that had been used in a clinical trial. We found that the viral protein abundance and composition were consistent across all types of virus examined except for the virus lacking protein V, which also had reduced amounts of another viral core protein, protein VII. In all the samples analysed we found no evidence of consistent packaging or contamination with cellular proteins. We believe this technique is a powerful method to analyse the protein composition of this important gene therapy vector and genetically engineered or synthetic virus-like particles. The raw data have been deposited at proteomexchange, identifer PXD001120.
Assuntos
Adenovírus Humanos/química , Adenovírus Humanos/genética , Adenovírus Humanos/isolamento & purificação , Aminoácidos/análise , Linhagem Celular , Células HEK293 , Células HeLa , Humanos , Marcação por Isótopo , Mutação , Proteômica , Espectrometria de Massas em Tandem , Proteínas Virais/química , Proteínas Virais/genéticaRESUMO
As humans age, they experience a progressive loss of thymic function and a corresponding shift in the makeup of the circulating CD8+ T cell population from naïve to memory phenotype. These alterations are believed to result in impaired CD8+ T cell responses in older individuals; however, evidence that these global changes impact virus-specific CD8+ T cell immunity in the elderly is lacking. To gain further insight into the functionality of virus-specific CD8+ T cells in older individuals, we interrogated a cohort of individuals who were acutely infected with West Nile virus (WNV) and chronically infected with Epstein Barr virus (EBV) and Cytomegalovirus (CMV). The cohort was stratified into young (<40 yrs), middle-aged (41-59 yrs) and aged (>60 yrs) groups. In the aged cohort, the CD8+ T cell compartment displayed a marked reduction in the frequency of naïve CD8+ T cells and increased frequencies of CD8+ T cells that expressed CD57 and lacked CD28, as previously described. However, we did not observe an influence of age on either the frequency of virus-specific CD8+ T cells within the circulating pool nor their functionality (based on the production of IFNγ, TNFα, IL2, Granzyme B, Perforin and mobilization of CD107a). We did note that CD8+ T cells specific for WNV, CMV or EBV displayed distinct functional profiles, but these differences were unrelated to age. Collectively, these data fail to support the hypothesis that immunosenescence leads to defective CD8+ T cell immunity and suggest that it should be possible to develop CD8+ T cell vaccines to protect aged individuals from infections with novel emerging viruses.