RESUMO
Tumor infiltration by T cells profoundly affects cancer progression and responses to immunotherapy. However, the tumor immunosuppressive microenvironment can impair the induction, trafficking, and local activity of antitumor T cells. Here, we investigated whether intratumoral injection of virus-derived peptide epitopes could activate preexisting antiviral T cell responses locally and promote antitumor responses or antigen spreading. We focused on a mouse model of cytomegalovirus (CMV), a highly prevalent human infection that induces vigorous and durable T cell responses. Mice persistently infected with murine CMV (MCMV) were challenged with lung (TC-1), colon (MC-38), or melanoma (B16-F10) tumor cells. Intratumoral injection of MCMV-derived T cell epitopes triggered in situ and systemic expansion of their cognate, MCMV-specific CD4+ or CD8+ T cells. The MCMV CD8+ T cell epitopes injected alone provoked arrest of tumor growth and some durable remissions. Intratumoral injection of MCMV CD4+ T cell epitopes with polyinosinic acid:polycytidylic acid (pI:C) preferentially elicited tumor antigen-specific CD8+ T cells, promoted tumor clearance, and conferred long-term protection against tumor rechallenge. Notably, secondary proliferation of MCMV-specific CD8+ T cells correlated with better tumor control. Importantly, intratumoral injection of MCMV-derived CD8+ T cell-peptide epitopes alone or CD4+ T cell-peptide epitopes with pI:C induced potent adaptive and innate immune activation of the tumor microenvironment. Thus, CMV-derived peptide epitopes, delivered intratumorally, act as cytotoxic and immunotherapeutic agents to promote immediate tumor control and long-term antitumor immunity that could be used as a stand-alone therapy. The tumor antigen-agnostic nature of this approach makes it applicable across a broad range of solid tumors regardless of their origin.
Assuntos
Linfócitos T CD8-Positivos , Infecções por Citomegalovirus , Citomegalovirus , Epitopos de Linfócito T , Neoplasias , Animais , Antígenos de Neoplasias/imunologia , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/virologia , Citomegalovirus/imunologia , Infecções por Citomegalovirus/imunologia , Epitopos de Linfócito T/administração & dosagem , Epitopos de Linfócito T/imunologia , Imunoterapia , Camundongos , Neoplasias/imunologia , Neoplasias/terapia , Poli I-C/administração & dosagem , Poli I-C/imunologia , Microambiente TumoralRESUMO
African swine fever (ASF) is a highly fatal swine disease threatening the global pig industry. Currently, vaccine is not commercially available for ASF. Hence, it is desirable to develop effective subunit vaccines against ASF. Here, we expressed and purified two recombinant fusion proteins comprising ASFV proteins p30 and p54 fused to a novel cell-penetrating peptide Z12, which were labeled as ZPM (Z12-p30-modified p54) and ZPMT (Z12-p30-modified p54-T cell epitope). Purified recombinant p30 and modified p54 expressed alone or fused served as controls. The transduction capacity of these recombinant proteins was assessed in RAW264.7 cells. Both ZPM and ZPMT exhibited higher transduction efficiency than the other proteins. Subsequently, humoral and cellular immune responses elicited by these proteins were evaluated in mice. ZPMT elicited the highest levels of antigen-specific IgG responses, cytokines (interleukin-2, interferon-γ, and tumor necrosis factor-α) and lymphocyte proliferation. Importantly, sera from mice immunized with ZPM or ZPMT neutralized greater than 85% of ASFV in vitro. Our results indicate that ZPMT induces potent neutralizing antibody responses and cellular immunity in mice. Therefore, ZPMT may be a suitable candidate to elicit immune responses in swine, providing valuable information for the development of subunit vaccines against ASF.
Assuntos
Vírus da Febre Suína Africana/imunologia , Febre Suína Africana/imunologia , Vacinas Virais/imunologia , Febre Suína Africana/genética , Animais , Anticorpos Neutralizantes/sangue , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Peptídeos Penetradores de Células/administração & dosagem , Peptídeos Penetradores de Células/genética , Peptídeos Penetradores de Células/imunologia , Epitopos de Linfócito T/administração & dosagem , Epitopos de Linfócito T/genética , Epitopos de Linfócito T/imunologia , Feminino , Imunidade Celular/imunologia , Camundongos , Fosfoproteínas/administração & dosagem , Fosfoproteínas/genética , Fosfoproteínas/imunologia , Proteínas Recombinantes de Fusão/administração & dosagem , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/imunologia , Suínos , Desenvolvimento de Vacinas , Vacinas de Subunidades Antigênicas/administração & dosagem , Vacinas de Subunidades Antigênicas/genética , Vacinas de Subunidades Antigênicas/imunologia , Proteínas Virais/administração & dosagem , Proteínas Virais/genética , Proteínas Virais/imunologia , Proteínas Estruturais Virais/administração & dosagem , Proteínas Estruturais Virais/genética , Proteínas Estruturais Virais/imunologia , Vacinas Virais/administração & dosagem , Vacinas Virais/genéticaRESUMO
BACKGROUND: Based on the interaction between cytotoxic T lymphocyte (CTL) dominant epitopes and dendritic cells (DCs), CD8+T cells are specifically activated into CTL cells. Targeted killing is a type of tumor vaccine for immunotherapy with great development potential. However, because of the disadvantages of poor stability in vivo and low uptake rate of DCs caused by single use of dominant epitope peptide drugs, its use is limited. Here, we investigated the antitumor potential of M-YL/LA-Lipo, a novel liposome drug delivery system. METHODS: We assembled mannose on the surface of liposome, which has a highly targeted effect on the mannose receptor on the surface of DCs. The dominant epitope peptide drugs were encapsulated into the liposome using membrane hydration method, and the encapsulation rate, release rate, in vitro stability, and microstructure were characterized using ultrafiltration method, dialysis method, and negative staining transmission electron microscopy. In addition, its targeting ability was verified by in vitro interaction with DCs, and its anticancer effect was verified by animal experiments. RESULTS: We have successfully prepared a liposome drug delivery system with stable physical and chemical properties. Moreover, we demonstrated that it was highly uptaken by DCs and promoted DC maturation in vitro. Furthermore, in vivo animal experiments indicated that M-YL/LA-Lipo specific CTL significantly inhibited the hematogenous spread of lung metastasis of triple negative breast cancer. CONCLUSIONS: we successfully constructed a new polypeptide liposome drug delivery system by avoiding the disadvantages of single use of dominant epitope peptide drugs and accurate targeted therapy for tumors.
Assuntos
Vacinas Anticâncer/administração & dosagem , Epitopos de Linfócito T/administração & dosagem , Manose/química , Neoplasias/terapia , Animais , Vacinas Anticâncer/imunologia , Linhagem Celular Tumoral , Células Dendríticas/imunologia , Epitopos de Linfócito T/imunologia , Humanos , Imunogenicidade da Vacina , Lipossomos , Receptor de Manose , Camundongos Transgênicos , Neoplasias/imunologia , Cultura Primária de Células , Linfócitos T Citotóxicos/imunologia , Vacinas de Subunidades Antigênicas/administração & dosagem , Vacinas de Subunidades Antigênicas/imunologiaRESUMO
AIMS: Human papillomavirus (HPV) L1, L2 and E7 proteins were used as target antigens for development of preventive and therapeutic vaccines. Moreover, linkage of antigens to heat shock proteins (HSPs) could enhance the potency of vaccines. Curcumin and nanocurcumin compounds were suggested as the chemopreventive and chemotherapeutic agents against cancer. In this study, two multiepitope DNA and peptide-based vaccine constructs (L1-L2-E7 and HSP70-L1-L2-E7) were used along with curcumin and nanocurcumin to evaluate immune responses, and protective/therapeutic effects in tumor mouse model. MAIN METHODS: At first, the multiepitope L1-L2-E7 and HSP70-L1-L2-E7 fusion genes were subcloned in eukaryotic and prokaryotic expression vectors. The recombinant multiepitope peptides were generated in E. coli strain. Then, the cytotoxic effects of curcumin and nanocurcumin were evaluated on HEK-293 T non-cancerous and C3 cancerous cells. Finally, mice vaccination was performed using different regimens. Curcumin and nanocurcumin compounds were administered alone or along with different vaccine constructs. KEY FINDINGS: Our data indicated that the use of nanocurcumin along with the multiepitope HSP70-L1-L2-E7 vaccine construct could completely protect mice against HPV-related C3 tumor cells, and eradicate tumors in a therapeutic test. Furthermore, nanocurcumin showed higher protection than curcumin alone. Generally, curcumin and nanocurcumin compounds could reduce tumor growth synergistically with the multiepitope vaccine constructs, but they did not influence the immune responses in different regimens. SIGNIFICANCE: These data demonstrated that the designed multiepitope vaccine constructs along with curcumin and nanocurcumin can be used as a promising method for HPV vaccine development.
Assuntos
Antineoplásicos/farmacologia , Vacinas Anticâncer/imunologia , Proteínas do Capsídeo/imunologia , Curcumina/farmacologia , Proteínas Oncogênicas Virais/imunologia , Proteínas E7 de Papillomavirus/imunologia , Vacinas contra Papillomavirus/imunologia , Neoplasias do Colo do Útero/imunologia , Animais , Antineoplásicos/administração & dosagem , Vacinas Anticâncer/genética , Proteínas do Capsídeo/administração & dosagem , Proteínas do Capsídeo/genética , Clonagem Molecular , Curcumina/administração & dosagem , Citocinas/metabolismo , Epitopos de Linfócito T/administração & dosagem , Epitopos de Linfócito T/genética , Epitopos de Linfócito T/imunologia , Escherichia coli , Feminino , Vetores Genéticos , Células HEK293 , Proteínas de Choque Térmico HSP70/administração & dosagem , Proteínas de Choque Térmico HSP70/genética , Proteínas de Choque Térmico HSP70/imunologia , Humanos , Camundongos Endogâmicos C57BL , Neoplasias Experimentais/imunologia , Neoplasias Experimentais/terapia , Proteínas Oncogênicas Virais/administração & dosagem , Proteínas Oncogênicas Virais/genética , Proteínas E7 de Papillomavirus/administração & dosagem , Proteínas E7 de Papillomavirus/genética , Vacinas contra Papillomavirus/administração & dosagem , Vacinas contra Papillomavirus/genética , Proteínas Recombinantes de Fusão/administração & dosagem , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/imunologia , Neoplasias do Colo do Útero/terapia , Vacinas de Subunidades Antigênicas/administração & dosagem , Vacinas de Subunidades Antigênicas/genética , Vacinas de Subunidades Antigênicas/imunologiaRESUMO
Ebola virus (EBOV) glycoprotein (GP) can be recognized by neutralizing antibodies (NAbs) and is the main target for vaccine design. Here, we first investigate the contribution of the stalk and heptad repeat 1-C (HR1C) regions to GP metastability. Specific stalk and HR1C modifications in a mucin-deleted form (GPΔmuc) increase trimer yield, whereas alterations of HR1C exert a more complex effect on thermostability. Crystal structures are determined to validate two rationally designed GPΔmuc trimers in their unliganded state. We then display a modified GPΔmuc trimer on reengineered protein nanoparticles that encapsulate a layer of locking domains (LD) and a cluster of helper T-cell epitopes. In mice and rabbits, GP trimers and nanoparticles elicit cross-ebolavirus NAbs, as well as non-NAbs that enhance pseudovirus infection. Repertoire sequencing reveals quantitative profiles of vaccine-induced B-cell responses. This study demonstrates a promising vaccine strategy for filoviruses, such as EBOV, based on GP stabilization and nanoparticle display.
Assuntos
Vacinas contra Ebola/administração & dosagem , Glicoproteínas/administração & dosagem , Doença pelo Vírus Ebola/terapia , Proteínas Virais/administração & dosagem , Animais , Anticorpos Neutralizantes/sangue , Anticorpos Neutralizantes/imunologia , Antígenos Virais/administração & dosagem , Antígenos Virais/genética , Antígenos Virais/imunologia , Antígenos Virais/ultraestrutura , Linfócitos B/imunologia , Cristalografia por Raios X , Modelos Animais de Doenças , Vacinas contra Ebola/genética , Vacinas contra Ebola/imunologia , Ebolavirus/genética , Ebolavirus/imunologia , Epitopos de Linfócito T/administração & dosagem , Epitopos de Linfócito T/genética , Epitopos de Linfócito T/imunologia , Epitopos de Linfócito T/ultraestrutura , Feminino , Glicoproteínas/genética , Glicoproteínas/imunologia , Glicoproteínas/ultraestrutura , Doença pelo Vírus Ebola/sangue , Doença pelo Vírus Ebola/imunologia , Doença pelo Vírus Ebola/virologia , Humanos , Camundongos , Nanopartículas/química , Domínios Proteicos/genética , Domínios Proteicos/imunologia , Engenharia de Proteínas , Multimerização Proteica/genética , Multimerização Proteica/imunologia , Estabilidade Proteica , Coelhos , Linfócitos T Auxiliares-Indutores/imunologia , Vacinas de Subunidades Antigênicas/administração & dosagem , Vacinas de Subunidades Antigênicas/genética , Vacinas de Subunidades Antigênicas/imunologia , Proteínas Virais/genética , Proteínas Virais/imunologia , Proteínas Virais/ultraestruturaRESUMO
T-cell receptor mimic (TCRm) antibodies have expanded the repertoire of antigens targetable by monoclonal antibodies, to include peptides derived from intracellular proteins that are presented by major histocompatibility complex class I (MHC-I) molecules on the cell surface. We have previously used this approach to target p53, which represents a valuable target for cancer immunotherapy because of the high frequency of its deregulation by mutation or other mechanisms. The T1-116C TCRm antibody targets the wild type p5365-73 peptide (RMPEAAPPV) presented by HLA-A*0201 (HLA-A2) and exhibited in vivo efficacy against triple receptor negative breast cancer xenografts. Here we report a comprehensive mutational analysis of the p53 RMPEAAPPV peptide to assess the T1-116C epitope and its peptide specificity. Antibody binding absolutely required the N-terminal arginine residue, while amino acids in the center of the peptide contributed little to specificity. Data mining the immune epitope database with the T1-116C binding consensus and validation of peptide recognition using the T2 stabilization assay identified additional tumor antigens targeted by T1-116C, including WT1, gp100, Tyrosinase and NY-ESO-1. Most peptides recognized by T1-116C were conserved in mice and human HLA-A2 transgenic mice showed no toxicity when treated with T1-116C in vivo. We conclude that comprehensive validation of TCRm antibody target specificity is critical for assessing their safety profile.
Assuntos
Antígeno HLA-A2/genética , Peptídeos/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Proteína Supressora de Tumor p53/química , Sequência de Aminoácidos , Animais , Especificidade de Anticorpos , Linhagem Celular Tumoral , Epitopos de Linfócito T/administração & dosagem , Epitopos de Linfócito T/imunologia , Feminino , Antígeno HLA-A2/metabolismo , Humanos , Rim/patologia , Fígado/patologia , Camundongos , Camundongos Transgênicos , Mutagênese Sítio-Dirigida , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Peptídeos/química , Peptídeos/metabolismo , Receptores de Antígenos de Linfócitos T/química , Proteína Supressora de Tumor p53/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismoRESUMO
Respiratory syncytial virus (RSV) is the leading cause of lower respiratory tract infection in young children. The T cell response plays a critical role in facilitating clearance of an acute RSV infection, and memory T cell responses are vital for protection against secondary RSV exposures. Tissue-resident memory (TRM) T cells have been identified as a subset of memory T cells that reside in nonlymphoid tissues and are critical for providing long-term immunity. There is currently limited information regarding the establishment and longevity of TRM T cell responses elicited following an acute RSV infection as well as their role in protection against repeated RSV infections. In this study, we examined the magnitude, phenotype, and protective capacity of TRM CD4 and CD8 T cells in the lungs of BALB/c mice following an acute RSV infection. TRM CD4 and CD8 T cells were established within the lungs and waned by 149 d following RSV infection. To determine the protective capacity of TRMs, FTY720 administration was used to prevent trafficking of peripheral memory T cells into the lungs prior to challenge of RSV-immune mice, with a recombinant influenza virus expressing either an RSV-derived CD4 or CD8 T cell epitope. We observed enhanced viral clearance in RSV-immune mice, suggesting that TRM CD8 T cells can contribute to protection against a secondary RSV infection. Given the protective capacity of TRMs, future RSV vaccine candidates should focus on the generation of these cell populations within the lung to induce effective immunity against RSV infection.
Assuntos
Epitopos de Linfócito T/imunologia , Memória Imunológica , Vacinas contra Influenza/imunologia , Células T de Memória/imunologia , Infecções por Vírus Respiratório Sincicial/prevenção & controle , Animais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Epitopos de Linfócito T/administração & dosagem , Epitopos de Linfócito T/genética , Feminino , Cloridrato de Fingolimode/farmacologia , Vacinas contra Influenza/administração & dosagem , Vacinas contra Influenza/genética , Pulmão/imunologia , Pulmão/virologia , Células T de Memória/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Infecções por Vírus Respiratório Sincicial/imunologia , Vírus Sinciciais Respiratórios/imunologia , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/imunologiaRESUMO
Respiratory syncytial virus (RSV) can cause recurrent infection in people because it does not stimulate a long-lived immunological memory. There is an urgent need to develop a safe and efficacious vaccine against RSV that would induce immunological memory without causing immunopathology following natural RSV infection. We have previously generated two recombinant live attenuated influenza vaccine (LAIV) viruses that encode immunodominant T-cell epitopes of RSV M2 protein in the neuraminidase or NS1 genes. These chimeric vaccines afforded protection against influenza and RSV infection in mice, without causing pulmonary eosinophilia or inflammatory RSV disease. The current study assessed the formation of influenza-specific and RSV-specific CD4 and CD8 T-cell responses in the lungs of mice, with special attention to the lung tissue-resident memory T cell subsets (TRM). The RSV epitopes did not affect influenza-specific CD4 effector memory T cell (Tem) levels in the lungs. The majority of these cells formed by LAIV or LAIV-RSV viruses had CD69+CD103- phenotype. Both LAIV+NA/RSV and LAIV+NS/RSV recombinant viruses induced significant levels of RSV M282 epitope-specific lung-localized CD8 Tem cells expressing both CD69 and CD103 TRM markers. Surprisingly, the CD69+CD103+ influenza-specific CD8 Tem responses were augmented by the addition of RSV epitopes, possibly as a result of the local microenvironment formed by the RSV-specific memory T cells differentiating to TRM in the lungs of mice immunized with LAIV-RSV chimeric viruses. This study provides evidence that LAIV vector-based vaccination can induce robust lung-localized T-cell immunity to the inserted T-cell epitope of a foreign pathogen, without altering the immunogenicity of the viral vector itself.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Epitopos de Linfócito T/imunologia , Memória Imunológica , Vacinas contra Influenza/imunologia , Infecções por Orthomyxoviridae/prevenção & controle , Animais , Anticorpos Neutralizantes/sangue , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Epitopos de Linfócito T/administração & dosagem , Epitopos de Linfócito T/genética , Feminino , Humanos , Subtipo H7N9 do Vírus da Influenza A/genética , Subtipo H7N9 do Vírus da Influenza A/imunologia , Vacinas contra Influenza/administração & dosagem , Vacinas contra Influenza/genética , Pulmão/imunologia , Pulmão/virologia , Camundongos , Camundongos Endogâmicos BALB C , Infecções por Orthomyxoviridae/imunologia , Vírus Sincicial Respiratório Humano/imunologia , Vacinas Atenuadas/administração & dosagem , Vacinas Atenuadas/imunologiaRESUMO
Trichuris trichiura is a parasite that infects 500 million people worldwide, leading to colitis, growth retardation and Trichuris dysentery syndrome. There are no licensed vaccines available to prevent Trichuris infection and current treatments are of limited efficacy. Trichuris infections are linked to poverty, reducing children's educational performance and the economic productivity of adults. We employed a systematic, multi-stage process to identify a candidate vaccine against trichuriasis based on the incorporation of selected T-cell epitopes into virus-like particles. We conducted a systematic review to identify the most appropriate in silico prediction tools to predict histocompatibility complex class II (MHC-II) molecule T-cell epitopes. These tools were used to identify candidate MHC-II epitopes from predicted ORFs in the Trichuris genome, selected using inclusion and exclusion criteria. Selected epitopes were incorporated into Hepatitis B core antigen virus-like particles (VLPs). Bone marrow-derived dendritic cells and bone marrow-derived macrophages responded in vitro to VLPs irrespective of whether the VLP also included T-cell epitopes. The VLPs were internalized and co-localized in the antigen presenting cell lysosomes. Upon challenge infection, mice vaccinated with the VLPs+T-cell epitopes showed a significantly reduced worm burden, and mounted Trichuris-specific IgM and IgG2c antibody responses. The protection of mice by VLPs+T-cell epitopes was characterised by the production of mesenteric lymph node (MLN)-derived Th2 cytokines and goblet cell hyperplasia. Collectively our data establishes that a combination of in silico genome-based CD4+ T-cell epitope prediction, combined with VLP delivery, offers a promising pipeline for the development of an effective, safe and affordable helminth vaccine.
Assuntos
Epitopos de Linfócito T/imunologia , Tricuríase/prevenção & controle , Trichuris/imunologia , Vacinas/imunologia , Animais , Anticorpos Anti-Helmínticos/imunologia , Simulação por Computador , Células Dendríticas/imunologia , Epitopos de Linfócito T/administração & dosagem , Epitopos de Linfócito T/genética , Antígenos de Histocompatibilidade Classe II/genética , Antígenos de Histocompatibilidade Classe II/imunologia , Humanos , Imunogenicidade da Vacina , Macrófagos/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Tricuríase/imunologia , Tricuríase/parasitologia , Trichuris/genética , Vacinas/administração & dosagem , Vacinas/genéticaRESUMO
Type 1 Diabetes (T1D) is an autoimmune disease that is associated with effector T cell (Teff) destruction of insulin-producing pancreatic beta-islet cells. Among the therapies being evaluated for T1D is the restoration of regulatory T cell (Treg) activity, specifically directed toward down-modulation of beta-islet antigen-specific T effector cells. This is also known as antigen-specific adaptive tolerance induction for T1D (T1D ASATI). Tregitopes (T regulatory cell epitopes) are natural T cell epitopes derived from immunoglobulin G (IgG) that were identified in 2008 and have been evaluated in several autoimmune disease models. In the T1D ASATI studies presented here, Tregitope peptides were administered to non-obese diabetic (NOD) mice at the onset of diabetes within two clinically-relevant delivery systems (liposomes and in human serum albumin [HSA]-fusion products) in combination with preproinsulin (PPI) target antigen peptides. The combination of Tregitope-albumin fusions and PPI peptides reduced the incidence of severe diabetes and reversed mild diabetes, over 49 days of treatment and observation. Combining HSA-Tregitope fusions with PPI peptides is a promising ASATI approach for therapy of T1D.
Assuntos
Diabetes Mellitus Tipo 1/tratamento farmacológico , Epitopos de Linfócito T/administração & dosagem , Tolerância Imunológica , Insulina/administração & dosagem , Peptídeos/administração & dosagem , Precursores de Proteínas/administração & dosagem , Albumina Sérica Humana/administração & dosagem , Animais , Diabetes Mellitus Tipo 1/imunologia , Epitopos de Linfócito T/genética , Feminino , Humanos , Insulina/genética , Camundongos Endogâmicos NOD , Peptídeos/genética , Precursores de Proteínas/genética , Proteínas Recombinantes de Fusão/administração & dosagem , Proteínas Recombinantes de Fusão/genética , Albumina Sérica Humana/genética , Linfócitos T Reguladores/imunologiaRESUMO
BACKGROUND: An effective vaccine against human immunodeficiency virus 1 (HIV-1) is an important global health priority. Despite many efforts in the development of the HIV-1 vaccine, no effective vaccine has been approved yet. Recently, polyepitope vaccines including several immunogenic and conserved epitopes of HIV-1 proteins have received special attention. METHODS: In this study, HIV-1 Nef, Tat, Gp160 and P24 proteins were considered for selection of immunodominant and conserved epitopes due to their critical roles in the viral life cycle and pathogenesis. At first, the Nef60-84-Nef126-144-Tat29-49-Gp16030-53-Gp160308-323-P248-151 DNA construct was designed using in silico studies. Then, the DNA construct was subcloned in pEGFP-N1 and pET- 24a (+) expression vectors and the rNef-Tat-Gp160-P24 polyepitope peptide was generated in E.coli expression system for in vitro delivery using novel cell-penetrating peptides (CPPs), LDP-NLS and CyLoP-1, in a non-covalent manner. Also, the HR9 and MPG CPPs were used to transfer the DNA construct. RESULTS: Our results showed that the recombinant polyepitope peptide generated in Rosetta strain migrated as a clear band of ~31 kDa in SDS-PAGE. The SEM data confirmed the formation of stable nanoparticles with a size below 250 nm. MTT assay revealed that the complexes did not represent any considerable cytotoxic effect compared to untreated cells. The results of fluorescence microscopy, flow cytometry and western blotting indicated that these CPPs successfully delivered polyepitope constructs into HEK-293T cell line. CONCLUSION: These data suggested that these CPPs can be used as a promising approach for the development of the HIV-1 vaccine.
Assuntos
Peptídeos Penetradores de Células/administração & dosagem , Epitopos/administração & dosagem , HIV-1 , Proteínas Virais , Sequência de Aminoácidos , Animais , Linhagem Celular , Peptídeos Penetradores de Células/química , Peptídeos Penetradores de Células/genética , Peptídeos Penetradores de Células/imunologia , Mapeamento de Epitopos , Epitopos/química , Epitopos/genética , Epitopos/imunologia , Epitopos de Linfócito T/administração & dosagem , Epitopos de Linfócito T/química , Epitopos de Linfócito T/genética , Epitopos de Linfócito T/imunologia , Expressão Gênica , Vetores Genéticos/genética , HIV-1/genética , HIV-1/imunologia , HIV-1/metabolismo , Humanos , Modelos Moleculares , Nanopartículas , Proteínas Recombinantes , Relação Estrutura-Atividade , Transfecção , Proteínas Virais/química , Proteínas Virais/genética , Proteínas Virais/imunologiaRESUMO
Cytomegalovirus (CMV) is a ubiquitous ß-herpesvirus that establishes life-long latent infection in a high percentage of the population worldwide. CMV induces the strongest and most durable CD8+ T cell response known in human clinical medicine. Due to its unique properties, the virus represents a promising candidate vaccine vector for the induction of persistent cellular immunity. To take advantage of this, we constructed a recombinant murine CMV (MCMV) expressing an MHC-I restricted epitope from influenza A virus (IAV) H1N1 within the immediate early 2 (ie2) gene. Only mice that were immunized intranasally (i.n.) were capable of controlling IAV infection, despite the greater potency of the intraperitoneally (i.p.) vaccination in inducing a systemic IAV-specific CD8+ T cell response. The protective capacity of the i.n. immunization was associated with its ability to induce IAV-specific tissue-resident memory CD8+ T (CD8TRM) cells in the lungs. Our data demonstrate that the protective effect exerted by the i.n. immunization was critically mediated by antigen-specific CD8+ T cells. CD8TRM cells promoted the induction of IFNγ and chemokines that facilitate the recruitment of antigen-specific CD8+ T cells to the lungs. Overall, our results showed that locally applied MCMV vectors could induce mucosal immunity at sites of entry, providing superior immune protection against respiratory infections.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Imunidade nas Mucosas , Vacinas contra Influenza/imunologia , Muromegalovirus/imunologia , Administração Intranasal , Sequência de Aminoácidos , Animais , Linhagem Celular , Quimiocinas/biossíntese , Epitopos de Linfócito T/administração & dosagem , Epitopos de Linfócito T/genética , Epitopos de Linfócito T/imunologia , Feminino , Produtos do Gene env/administração & dosagem , Produtos do Gene env/genética , Produtos do Gene env/imunologia , Vetores Genéticos , Humanos , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H1N1/imunologia , Vacinas contra Influenza/administração & dosagem , Vacinas contra Influenza/genética , Influenza Humana/imunologia , Influenza Humana/prevenção & controle , Pulmão/imunologia , Pulmão/virologia , Camundongos , Camundongos Endogâmicos BALB C , Muromegalovirus/genética , Células NIH 3T3 , Fragmentos de Peptídeos/administração & dosagem , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/imunologia , Mucosa Respiratória/imunologia , Mucosa Respiratória/virologia , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/genética , Vacinas Sintéticas/imunologiaRESUMO
Anti-tumor immunity is limited by a number of factors including the lack of fully activated T-cells, insufficient antigenic stimulation and the immune-suppressive tumor microenvironment. We addressed these hurdles by developing a novel class of immunoconjugates, Antibody-Targeted Pathogen-derived Peptides (ATPPs), which were designed to efficiently deliver viral T-cell epitopes to tumors with the aim of redirecting virus-specific memory T-cells against the tumor. ATPPs were generated through covalent binding of mature MHC class I peptides to antibodies specific for cell surface-expressed tumor antigens that mediate immunoconjugate internalization. By means of a cleavable linker, the peptides are released in the endosomal compartment, from which they are loaded into MHC class I without the need for further processing. Pulsing of tumor cells with ATPPs was found to sensitize these for recognition by virus-specific CD8+ T-cells with much greater efficiency than exogenous loading with free peptides. Systemic injection of ATPPs into tumor-bearing mice enhanced the recruitment of virus-specific T-cells into the tumor and, when combined with immune checkpoint blockade, suppressed tumor growth. Our data thereby demonstrate the potential of ATPPs as a means of kick-starting the immune response against "cold" tumors and increasing the efficacy of checkpoint inhibitors.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Epitopos de Linfócito T/administração & dosagem , Imunoconjugados/administração & dosagem , Neoplasias/terapia , Peptídeos/administração & dosagem , Animais , Linhagem Celular Tumoral , Feminino , Herpesvirus Humano 4 , Humanos , Imunoterapia , CamundongosRESUMO
Bacillus Calmette-Guerin (BCG) is the only vaccine against TB and has limited protection efficacy, which wanes past adolescence. Multifunctional CD8+ T cells (IFN-γ+/TNF-α+/IL-2+) are associated with lower reactivation risk and enhanced control of active Mtb infection. Since boosting with BCG is contraindicated, booster vaccines that augment T cell immunity in the lungs of BCG-vaccinated individuals are urgently needed. We developed a vaccination strategy based on self-assembling peptide nanofibers presenting Mtb-specific CD8+ or CD4+ T cell epitopes that induce high frequency and antigen-specific effector memory T cells producing IFN-γ and IL-2. Intranasal immunization with peptide nanofibers was well tolerated in mice leading to increased antigen-specific CD8+ T cell population in the lungs. Co-assembled nanofibers of CD8+ T cell epitopes and toll-like receptor 2 (TLR2) agonists induced a 8-fold expansion in multifunctional CD8+ T cell populations in the lungs of vaccinated mice. Aerosol challenge with Mtb in BCG-primed and nanofiber-boosted mice provided an additional 0.5-log CFU reduction in lung bacterial load and indicating enhanced protection compared to BCG alone. Together, these data suggest that heterologous prime-boost with BCG and peptide nanofiber vaccines induces cell mediated immunity in the lung, reduces bacterial burden, and is a potentially safer alternative for boosting BCG-primed immunity.
Assuntos
Antígenos de Bactérias/química , Epitopos de Linfócito B/administração & dosagem , Epitopos de Linfócito T/administração & dosagem , Mycobacterium tuberculosis/imunologia , Peptídeos/administração & dosagem , Linfócitos T/imunologia , Administração Intranasal , Animais , Vacina BCG/administração & dosagem , Vacina BCG/imunologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Epitopos de Linfócito B/imunologia , Epitopos de Linfócito T/imunologia , Imunização Secundária , Infusões Parenterais , Interferon gama/metabolismo , Interleucina-2/metabolismo , Camundongos , Nanofibras , Peptídeos/síntese química , Peptídeos/imunologiaRESUMO
Dendritic cell (DC)-based cytotoxic T lymphocyte (CTL) epitope vaccines are effective to induce CTL responses but require complex ex vivo DC preparation and epitope-loading. To take advantage of DC-based epitope vaccines without involving the ex vivo procedures, we aimed to develop carriers to directly load CTL epitopes onto DCs in vivo. Here, we first engineered a carrier consisting of a hydrophilic polypeptide, immune-tolerant elastin-like polypeptide (iTEP) and a substrate peptide of matrix metalloproteinases-9 (sMMP). The iTEP was able to solubilize CTL epitopes. CTL epitopes were connected to the carrier, iTEP-sMMP, through sMMP so that the epitopes can be cleaved from the carrier by MMP-9. iTEP-sMMP was found to release its epitope payloads in the DC culture media, which contained MMP-9 released from DCs. iTEP-sMMP allowed for the direct loading of CTL epitopes onto the surface MHC class I complexes of DCs. Importantly, iTEP-sMMP resulted in greater epitope presentation by DCs both in vitro and in vivo than a control carrier that cannot directly load epitopes. iTEP-sMMP also induced 2-fold stronger immune responses than the control carrier. To further enhance the direct epitope-loading strategy, we furnished iTEP-sMMP with an albumin-binding domain (ABD) and found the new carrier, ABD-iTEP-sMMP, had greater lymph node (LN) accumulation than iTEP-sMMP. ABD-iTEP-sMMP also resulted in greater immune responses than iTEP-sMMP by 1.5-fold. Importantly, ABD-iTEP-sMMP-delivered CTL epitope vaccine induced stronger immune responses than free CTL epitope vaccine. Taken together, these carriers utilized two physiological features of DCs to realize direct epitope-loading in vivo: the accumulation of DCs in LNs and MMP-9 released from DCs. These carriers are a potential substitute for DC-based CTL epitope vaccines.
Assuntos
Vacinas Anticâncer/administração & dosagem , Vacinas Anticâncer/imunologia , Células Dendríticas/imunologia , Epitopos de Linfócito T/administração & dosagem , Epitopos de Linfócito T/imunologia , Linfócitos T Citotóxicos/imunologia , Animais , Células Dendríticas/química , Portadores de Fármacos/química , Feminino , Antígenos de Histocompatibilidade Classe I/química , Antígenos de Histocompatibilidade Classe I/imunologia , Metaloproteinase 9 da Matriz/imunologia , Camundongos Endogâmicos C57BL , Neoplasias/imunologia , Neoplasias/prevenção & controle , Peptídeos/químicaRESUMO
Influenza vaccines that can be administered intranasally or by other needle-free delivery routes have potential advantages over injected formulations in terms of patient compliance, cost, and ease of global distribution. Supramolecular peptide nanofibers have been investigated previously as platforms for vaccines and immunotherapies and have been shown to raise immune responses in the absence of exogenous adjuvants and without measurable inflammation. However, at present it has not been tested whether the immunogenicity of these materials extends to the intranasal route. Here we investigated the extent to which self-assembled peptide nanofibers bearing an influenza peptide epitope elicit antigen-specific CD8+ T cell responses when delivered intranasally, and we compared these responses with those elicited by subcutaneous immunization. Peptides containing an epitope from influenza acid polymerase (PA) and the Q11 self-assembly domain formed nanofibers that were avidly taken up by dendritic cells in lung-draining mediastinal lymph nodes after intranasal immunization. Intranasally delivered nanofibers generated greater antigen-specific CD8+ T cell responses in the lung-draining lymph nodes than subcutaneous immunizations while retaining the non-inflammatory character of the materials observed in other delivery sites. The CD8+ T cells elicited systemically were functional as assessed by their ability to produce IFN-γ ex vivo, lyse epitope-pulsed target cells in vivo, and diminish viral loads in infected mice. Compared to subcutaneously delivered nanofibers, intranasally delivered peptide nanofibers significantly increased the number of persisting antigen-specific tissue resident memory CD8+ T cells in the lung, allowing for a more rapid response to infection at 6â¯weeks post-vaccination. These results indicate that intranasally delivered self-assembled peptide nanofibers are immunogenic when delivering CD8+ epitopes without adjuvant or CD4+ epitopes, are non-inflammatory, and promote more lung-resident memory CD8+ T cells compared to subcutaneous immunization.
Assuntos
Alphainfluenzavirus/imunologia , Linfócitos T CD8-Positivos/imunologia , Vacinas contra Influenza/administração & dosagem , Nanofibras/administração & dosagem , Infecções por Orthomyxoviridae/prevenção & controle , Peptídeos/administração & dosagem , Administração Intranasal , Animais , Epitopos de Linfócito T/administração & dosagem , Epitopos de Linfócito T/imunologia , Epitopos de Linfócito T/uso terapêutico , Vacinas contra Influenza/imunologia , Vacinas contra Influenza/uso terapêutico , Camundongos Endogâmicos C57BL , Nanofibras/uso terapêutico , Nanofibras/ultraestrutura , Infecções por Orthomyxoviridae/imunologia , Peptídeos/imunologia , Peptídeos/uso terapêuticoRESUMO
Peptides have been studied as an important class of components in medicine to control many major diseases with vaccination. Polymers as adjuvants are capable of enhancing the vaccine potential against various diseases by improving the delivery of antigens, and they reduce the booster doses of vaccines. In brief, polymers are promising candidates for peptide-based vaccine delivery platforms. The purpose of the present study was to create a possible alternative approach in the treatment of malignant melanoma and/or to prevent metastasis of melanoma. The study was designed as both an experimental and an in vivo study. We prepared a complex and covalent conjugate of MAGE-3 121-134 (L-L-K-Y-R-A-R-E-P-V-T-K-A-E) T-cell epitope as a vaccine candidate for melanoma. These conjugates were able to generate an immune response in mice after a single immunization, without the help of any external adjuvant. The peptide-polymer complexes activated the immune system in the best way and formed the highest antigen specific immune response. These results indicate the adjuvant activity of Poly(N-vinyl-2- pyrrolidone-co-acrylic acid) [P(VP-co-AA)] and the potential use of P(VP-coAA)-peptide based vaccine prototypes for future melanoma cancer vaccine formulations.
Assuntos
Resinas Acrílicas/administração & dosagem , Anticorpos Antineoplásicos/sangue , Antígenos de Neoplasias/imunologia , Vacinas Anticâncer/administração & dosagem , Imunidade Humoral/efeitos dos fármacos , Melanoma/prevenção & controle , Proteínas de Neoplasias/imunologia , Peptídeos/administração & dosagem , Povidona/análogos & derivados , Neoplasias Cutâneas/prevenção & controle , Resinas Acrílicas/química , Adjuvantes Imunológicos/administração & dosagem , Adjuvantes Imunológicos/química , Sequência de Aminoácidos , Animais , Antígenos de Neoplasias/química , Vacinas Anticâncer/química , Vacinas Anticâncer/imunologia , Epitopos de Linfócito T/administração & dosagem , Epitopos de Linfócito T/química , Epitopos de Linfócito T/imunologia , Humanos , Imunização Secundária , Imunoconjugados/administração & dosagem , Imunoconjugados/química , Masculino , Melanoma/sangue , Melanoma/imunologia , Melanoma/patologia , Camundongos , Camundongos Endogâmicos BALB C , Proteínas de Neoplasias/química , Peptídeos/química , Peptídeos/imunologia , Povidona/administração & dosagem , Povidona/química , Soroalbumina Bovina/administração & dosagem , Soroalbumina Bovina/química , Soroalbumina Bovina/imunologia , Neoplasias Cutâneas/sangue , Neoplasias Cutâneas/imunologia , Neoplasias Cutâneas/patologia , Linfócitos T/química , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Vacinação , Vacinas de Subunidades Antigênicas , Melanoma Maligno CutâneoRESUMO
Cytotoxic T lymphocyte (CTL) can have remarkable abilities to kill tumor cells. However, the establishment of successful CTL-based anticancer therapy has met with many challenges. Within tumor cells, there exist subpopulations with low or no expression of the targeted antigen (termed as antigen-loss variants). In addition, tumor cells can downregulate the levels of major histocompatibility complex class I (MHC-I) molecules on cell surface due to immune pressure. As a result, some tumor cells can escape the immune pressure bestowed by CTLs, resulting in treatment failure. To address these difficulties, a new approach is developed to deliver foreign high-affinity CTL epitopes to tumor tissues utilizing pH-responsive "smart" microparticles (MPs). These MPs could encapsulate CTL peptide epitope, release the peptide under acidic condition encountered in tumor tissues and enhance CTL activation. Mice bearing pre-established tumor as "antigen-loss variant" solid tumor models were administered intratumorally with MPs containing the CTL peptide, which showed 100% survival following the treatment. In contrast, all control mice died from tumor. Significant protection from tumor-induced death was also observed with systemic administration of CTL peptide-MPs. The therapeutic efficacy can be attributed to enhanced delivery of the epitope to tumor tissues, presentation of the epitope by tumor cells as well as tumor stromal cells and/or generation of epitope-specific CTLs by the peptide-containing MPs. These findings offer a promising new direction for treating established solid tumor using CTL therapy.
Assuntos
Adenocarcinoma/terapia , Epitopos de Linfócito T/administração & dosagem , Imunoterapia/métodos , Lipossomos/administração & dosagem , Linfoma/terapia , Peptídeos/administração & dosagem , Linfócitos T Citotóxicos/imunologia , Adenocarcinoma/imunologia , Animais , Linhagem Celular Tumoral , Epitopos de Linfócito T/química , Feminino , Humanos , Concentração de Íons de Hidrogênio , Lipossomos/química , Ativação Linfocitária , Linfoma/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Transplante de Neoplasias , Neoplasias Experimentais , Peptídeos/química , Evasão TumoralRESUMO
Plasmodium falciparum malaria is one of the leading infectious causes of childhood mortality in Africa. EP-1300 is a polyepitope plasmid DNA vaccine expressing 38 cytotoxic T cell epitopes and 16 helper T cell epitopes derived from P. falciparum antigens expressed predominantly in the liver phase of the parasite's life cycle. We performed a phase 1 randomized, placebo-controlled, dose escalation clinical trial of the EP-1300 DNA vaccine administered via electroporation using the TriGrid Delivery System device (Ichor Medical Systems). Although the delivery of the EP-1300 DNA vaccine via electroporation was safe, tolerability was less than that usually observed with standard needle and syringe intramuscular administration. This was primarily due to acute local discomfort at the administration site during electroporation. Despite the use of electroporation, the vaccine was poorly immunogenic. The reasons for the poor immunogenicity of this polyepitope DNA vaccine remain uncertain. CLINICAL TRIALS REGISTRATION: ClinicalTrials.gov NCT01169077.
Assuntos
Imunogenicidade da Vacina , Vacinas Antimaláricas/administração & dosagem , Vacinas Antimaláricas/imunologia , Malária Falciparum/prevenção & controle , Vacinas de DNA/administração & dosagem , Vacinas de DNA/imunologia , Adolescente , Adulto , África , Anticorpos Antiprotozoários/biossíntese , Anticorpos Antiprotozoários/sangue , Relação Dose-Resposta Imunológica , Eletroporação , Epitopos de Linfócito T/administração & dosagem , Epitopos de Linfócito T/imunologia , Feminino , Humanos , Vacinas Antimaláricas/efeitos adversos , Malária Falciparum/epidemiologia , Masculino , Plasmodium falciparum/imunologia , Proteínas de Protozoários/imunologia , Vacinas de DNA/efeitos adversos , Vacinas de DNA/química , Adulto JovemRESUMO
AIM: Peptide-based vaccines are designed to carry the minimum required antigen to trigger the desired immune responses; however, they are usually poorly immunogenic and require appropriate delivery system. RESULTS: Peptides, B-cell epitope (J14) derived from group A streptococcus M-protein and universal T-helper (PADRE) epitope, were conjugated to a variety of linear and branched polyacrylates. All produced conjugates formed submicron-sized particles and induced a high level of IgG titres in mice after subcutaneous immunization. These polymer-peptide conjugates demonstrated high opsonization capacity against group A streptococcus clinical isolates. CONCLUSION: We have successfully demonstrated that submicron-sized polymer-peptide conjugates were capable of inducing strong humoral immune responses after single immunization.