RESUMO
Wilms' tumor 1 (WT1) is a promising tumor-associated antigen for cancer immunotherapy. We developed an oral protein vaccine platform composed of WT1-anchored, genetically engineered Bifidobacterium longum (B. longum) and conducted an in vivo study in mice to examine its anticancer activity. Mice were orally treated with phosphate-buffered saline, wild-type B. longum105-A, B. longum 2012 displaying only galacto-N-biose/lacto-N-biose I-binding protein (GLBP), and WT1 protein- and GLBP-expressing B. longum 420. Tumor size reduced significantly in the B. longum 420 group than in the B. longum 105-A and 2012 groups (P < 0.00 l each), indicating B. longum 420's antitumor activity via WT1-specific immune responses. CD8+ T cells played a major role in the antitumor activity of B. longum 420. The proportion of CD103+CD11b+CD11c+ dendritic cells (DCs) increased in the Peyer's patches (PPs) from mice in the B. longum 420 group, indicating the definite activation of DCs. In the PPs, the number and proportion of CD8+ T cells capable of producing interferon-gamma were significantly greater in the B. longum 420 group than in the B. longum 2012 group (P < 0.05 or < 0.01). The production of WT1-specific IgG antibody was significantly higher in the B. longum 420 group than in the 2012 group (P < 0.05). The B. longum 420 group showed the most intense intratumoral infiltration of CD4+ and CD8+ T cells primed by activated DCs in the PPs of mice in the B. longum 420 group. Our findings provide insights into a novel, intestinal bacterium-based, cancer immunotherapy through intestinal immunity.
Assuntos
Bifidobacterium longum , Vacinas Anticâncer , Leucemia Mieloide Aguda , Camundongos , Animais , Proteínas WT1 , Linfócitos T CD8-PositivosRESUMO
CD4+ T cells that recognize antigenic peptides presented on HLA class II are essential for inducing an optimal anti-tumor immune response, and adoptive transfer of tumor antigen-specific TCR-transduced CD4+ T cells with high responsiveness against tumor is a promising strategy for cancer treatment. Whereas a precise evaluation method of functional avidity, an indicator of T cell responsiveness against tumors, has been established for HLA class I-restricted TCRs, it remains unestablished for HLA class II-restricted TCRs. In this study, we generated a novel platform cell line, CD4-2D3, in which GFP reporter was expressed by NFAT activation via TCR signaling, for correctly evaluating functional avidity of HLA class II-restricted TCRs. Furthermore, using this platform cell line, we succeeded in maturating functional avidity of an HLA class II-restricted TCR specific for a WT1-derived helper peptide by substituting amino acids in complementarity determining region 3 (CDR3) of the TCR. Importantly, we demonstrated that transduction of an avidity-maturated TCR conferred strong cytotoxicity against WT1-expressing leukemia cells on CD4+ T cells, compared to that of its original TCR. Thus, CD4-2D3 cell line should be useful not only to evaluate TCR functional avidity in HLA class II-restricted TCRs but also to screen appropriate TCRs for clinical applications such as cancer immunotherapy.
Assuntos
Imunoterapia Adotiva , Neoplasias , Humanos , Imunoterapia Adotiva/métodos , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/metabolismo , Linfócitos T CD4-Positivos , Antígenos de NeoplasiasRESUMO
BACKGROUND: A Wilms' tumor 1 (WT1) oral vaccine, Bifidobacterium longum (B. longum) 420, in which the bacterium is used as a vector for WT1 protein, triggers immune responses through cellular immunity consisting of cytotoxic T lymphocytes (CTLs) and other immunocompetent cells (e.g., helper T cells). We developed a novel, oral, helper epitope-containing WT1 protein vaccine (B. longum 2656) to examine whether or not B. longum 420/2656 combination further accelerates the CD4+ T cell help-enhanced antitumor activity in a model of murine leukemia. METHODS: C1498-murine WT1-a genetically-engineered, murine leukemia cell line to express murine WT1-was used as tumor cell. Female C57BL/6 J mice were allocated to the B. longum 420, 2656, and 420/2656 combination groups. The day of subcutaneous inoculation of tumor cells was considered as day 0, and successful engraftment was verified on day 7. The oral administration of the vaccine by gavage was initiated on day 8. Tumor volume, the frequency and phenotypes of WT1-specific CTLs in CD8+ T cells in peripheral blood (PB) and tumor-infiltrating lymphocytes (TILs), as well as the proportion of interferon-gamma (INF-γ)-producing CD3+CD4+ T cells pulsed with WT135-52 peptide in splenocytes and TILs were determined. RESULTS: Tumor volume was significantly smaller (p < 0.01) in the B. longum 420/2656 combination group than in the B. longum 420 group on day 24. WT1-specific CTL frequency in CD8+ T cells in PB was significantly greater in the B. longum 420/2656 combination group than in the B. longum 420 group at weeks 4 (p < 0.05) and 6 (p < 0.01). The proportion of WT1-specific, effector memory CTLs in PB increased significantly in the B. longum 420/2656 combination group than in the B. longum 420 group at weeks 4 and 6 (p < 0.05 each). WT1-specific CTL frequency in intratumoral CD8+ T cells and the proportion of IFN-γ-producing CD3+CD4+ T cells in intratumoral CD4+ T cells increased significantly (p < 0.05 each) in the B. longum 420/2656 combination group than in the 420 group. CONCLUSIONS: B. longum 420/2656 combination further accelerated antitumor activity that relies on WT1-specific CTLs in the tumor compared with B. longum 420.
Assuntos
Vacinas Anticâncer , Neoplasias Renais , Leucemia , Tumor de Wilms , Feminino , Animais , Camundongos , Proteínas WT1 , Linfócitos T CD8-Positivos , Epitopos , Camundongos Endogâmicos C57BL , Linfócitos T Citotóxicos , Interferon gamaRESUMO
We have previously revealed the overexpression of Wilms' tumor gene 1 (WT1) in malignant glioma and developed WT1 peptide vaccine cancer immunotherapy. A phase II clinical trial indicated the clinical efficacy of the WT1 peptide vaccine for recurrent malignant glioma. Here, we aimed to investigate the immunological microenvironment in glioma tissues before and after WT1 peptide vaccine treatment. Paired tissue samples were obtained from 20 malignant glioma patients who had received the WT1 peptide vaccine for > 3 months and experienced tumor progression, confirmed radiographically and/or clinically, during vaccination. We discovered that the expression of WT1 and HLA class I antigens in the tumor cells significantly decreased after vaccination. Maintenance of WT1 expression, which is the target molecule of immunotherapy, in tumor cells during the vaccination period was significantly associated with a longer progression-free and overall survival. A high expression of HLA class I antigens and low CD4+/CD8+ tumor-infiltrating lymphocytes (TIL) ratio in pre-vaccination specimens, were also associated with a good prognosis. No statistically significant difference existed in the number of infiltrating CD3+ or CD8+ T cells between the pre- and post-vaccination specimens, whereas the number of infiltrating CD4+ T cells significantly decreased in the post-vaccination specimens. This study provides insight into the mechanisms of intra-tumoral immune reaction/escape during WT1 peptide vaccine treatment and suggests potential clinical strategies for cancer immunotherapy.
Assuntos
Neoplasias Encefálicas/diagnóstico , Neoplasias Encefálicas/metabolismo , Regulação Neoplásica da Expressão Gênica , Glioma/diagnóstico , Glioma/metabolismo , Imunoterapia/métodos , Proteínas WT1/biossíntese , Adulto , Biomarcadores Tumorais/biossíntese , Complexo CD3/biossíntese , Linfócitos T CD4-Positivos/citologia , Vacinas Anticâncer , Ensaios Clínicos Fase I como Assunto , Ensaios Clínicos Fase II como Assunto , Feminino , Perfilação da Expressão Gênica , Antígenos de Histocompatibilidade Classe I/imunologia , Humanos , Masculino , Pessoa de Meia-Idade , Peptídeos/química , Prognóstico , Modelos de Riscos ProporcionaisRESUMO
Simultaneous induction of tumor antigen-specific cytotoxic T lymphocytes (CTLs) and helper T lymphocytes (HTLs) is required for an optimal anti-tumor immune response. WT1332, a 16-mer WT1-derived helper peptide, induce HTLs in an HLA class II-restricted manner and enhance the induction of WT1-specific CTLs in vitro. However, in vivo immune reaction to WT1332 vaccination in tumor-bearing patients remained unclear. Here, a striking difference in WT1-specific T cell responses was shown between WT1 CTL + WT1 helper peptide and WT1 CTL peptide vaccines in patients with recurrent glioma. WT1-specific CTLs were more strongly induced in the patients who were immunized with WT1 CTL + WT1 helper peptide vaccine, compared to those who were immunized with WT1 CTL vaccine alone. Importantly, a clear correlation was demonstrated between WT1-specific CTL and WT1332-specific HTL responses. Interestingly, two novel distinct populations of WT1-tetramerlow WT1-TCRlow CD5low and WT1-tetramerhigh WT1-TCRhigh CD5high CTLs were dominantly detected in WT1 CTL + WT1 helper peptide vaccine. Although natural WT1 peptide-reactive CTLs in the latter population were evidently less than those in the former population, the latter population showed natural WT1 peptide-specific proliferation capacity comparable to the former population, suggesting that the latter population highly expressing CD5, a marker of resistance to activation-induced cell death, should strongly expand and persist for a long time in patients. These results demonstrated the advantage of WT1 helper peptide vaccine for the enhancement of WT1-specific CTL induction by WT1 CTL peptide vaccine.
Assuntos
Antígenos de Neoplasias/imunologia , Vacinas Anticâncer/imunologia , Peptídeos/imunologia , Linfócitos T Citotóxicos/imunologia , Vacinas de Subunidades Antigênicas/imunologia , Proteínas WT1/imunologia , Antígenos CD5/imunologia , Morte Celular/imunologia , Proliferação de Células/fisiologia , Células Cultivadas , Humanos , Leucócitos Mononucleares/imunologia , Recidiva Local de Neoplasia/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Vacinação/métodosRESUMO
Helper T lymphocytes (HTLs) play a central role in cancer immunity because they can not only help the induction and proliferation of cytotoxic T lymphocytes (CTLs) but also their differentiation into cytotoxic CD4+ T cells and directly kill the target cells.This study describes the identification of three novel mouse Th epitope peptides, WT135-52, WT186-102 and WT1294-312, derived from WT1 protein, which is the most potent tumor-associated antigen. Compared to immunization with WT1 CTL peptide alone, immunization with the addition of these WT1-specific Th peptides strongly induced WT1-specific CTLs, continued to maintain them, and efficiently rejected the challenge of WT1-expressing tumor cells. Importantly, the majority of WT1-specific CTLs induced by the co-immunization with WT1 CTL and the WT1-specific Th peptides were CD44+CD62L- effector memory CD8+ T cells, which played a central role in tumor rejection. Establishment of mouse models suitable for the analysis of the detailed mechanism of these functions of HTLs is very important. These results clearly showed that WT1-specific HTLs perform an essential function in WT1-specific tumor immunity. Therefore, the WT1-specific Th peptides identified here should make a major contribution to elucidation of the mutual roles of WT1-specific CTLs and HTLs in cancer immunity in in vivo mouse models.
Assuntos
Antígenos de Neoplasias/imunologia , Epitopos de Linfócito T/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Proteínas WT1/imunologia , Animais , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Linfócitos T Citotóxicos/imunologiaRESUMO
PURPOSE: The safety and clinical efficacy of WT1 human leukocyte antigen (HLA) class I peptide vaccine have been established, but the safety of a cocktail vaccine of WT1 HLA class I and II peptides has not. To verify its safety, we performed a phase I clinical trial for patients with recurrent malignant gliomas and assessed the immunological responses and survival data. PATIENTS AND METHODS: Fourteen HLA-A*24:02-positive patients with recurrent malignant glioma (2 with grade 3, 12 with grade 4) were enrolled. Every week, the patients received alternately a vaccine containing 3 mg of WT1 HLA-A*24:02-restricted (HLA class I) peptide and a cocktail vaccine of the HLA class I peptide and one of 0.75, 1.5 or 3 mg of the WT1 HLA class II peptide. For patients who showed no significant adverse effects within 6 weeks, the WT1 vaccine was continued at 2-4-week intervals. RESULTS: Eleven of the 14 patients completed WT1 vaccination for 6 weeks, while 3 patients dropped out earlier due to disease progression. All patients showed grade I level of skin disorders at the injection sites. No grade III/IV toxicity or dose-limiting toxicity was observed for any dose of WT1 HLA class II peptide. Six of the 14 patients had stable disease at 6 weeks. Median OS and 1-year OS rates were 24.7 weeks and 36%, respectively. CONCLUSION: The safety of a cocktail vaccine of WT1 HLA class I and II peptides for malignant gliomas was verified. This vaccine is, therefore, considered promising for patients with recurrent malignant glioma.
Assuntos
Neoplasias Encefálicas/tratamento farmacológico , Vacinas Anticâncer/uso terapêutico , Glioma/tratamento farmacológico , Vacinas de Subunidades Antigênicas/uso terapêutico , Adulto , Neoplasias Encefálicas/imunologia , Neoplasias Encefálicas/patologia , Vacinas Anticâncer/imunologia , Feminino , Glioma/imunologia , Glioma/patologia , Antígeno HLA-A24/imunologia , Antígenos de Histocompatibilidade Classe II/imunologia , Humanos , Masculino , Pessoa de Meia-Idade , Recidiva Local de Neoplasia , Análise de Sobrevida , Resultado do Tratamento , Vacinação/métodos , Vacinas de Subunidades Antigênicas/imunologia , Proteínas WT1/imunologiaRESUMO
Thymic epithelial tumors are rare malignancies, and no optimal therapeutic regimen has been defined for patients with advanced disease. Patients with advanced thymic epithelial tumors, which were resistant or intolerable to prior therapies, were eligible for this study. Patients received 9 mer-WT1-derived peptide emulsified with Montanide ISA51 adjuvant via intradermal administration once a week as a monotherapy. After the 3-month-protocol treatment, the treatment was continued mostly at intervals of 2-4 weeks until disease progression or intolerable adverse events occurred. Of the 15 patients enrolled, 11 had thymic carcinoma (TC) and 4 had invasive thymoma (IT). Median period from diagnosis to the start of treatment was 13.3 and 65.5 months for TC and IT, respectively. No patients achieved a complete or partial response. Of the 8 evaluable TC patients, 6 (75.0%) had stable disease (SD) and 2 had progressive disease (PD). Of the 4 evaluable IT patients, 3 (75.0%) had SD and 1 (25.0%) had PD. Median period of monotherapy treatment was 133 and 683 days in TC and IT patients, respectively. No severe adverse events occurred during the 3-month-protocol treatment. As adverse events in long responders, thymoma-related autoimmune complications, pure red cell aplasia and myasthenia gravis occurred in two IT patients. Cerebellar hemorrhage developed in a TC patient complicated with Von Willebrand disease. Induction of WT1-specific immune responses was observed in the majority of the patients. WT1 peptide vaccine immunotherapy may have antitumor potential against thymic malignancies.
Assuntos
Imunoterapia , Neoplasias Epiteliais e Glandulares/imunologia , Neoplasias Epiteliais e Glandulares/patologia , Peptídeos/imunologia , Neoplasias do Timo/imunologia , Neoplasias do Timo/patologia , Proteínas WT1/imunologia , Adulto , Idoso , Vacinas Anticâncer/imunologia , Vacinas Anticâncer/uso terapêutico , Terapia Combinada , Feminino , Expressão Gênica , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Neoplasias Epiteliais e Glandulares/diagnóstico por imagem , Neoplasias Epiteliais e Glandulares/tratamento farmacológico , Neoplasias do Timo/diagnóstico por imagem , Neoplasias do Timo/tratamento farmacológico , Tomografia Computadorizada por Raios X , Resultado do Tratamento , Proteínas WT1/química , Proteínas WT1/metabolismoRESUMO
We previously evaluated Wilms' tumor gene 1 (WT1) peptide vaccination in a large number of patients with leukemia or solid tumors and have reported that HLA-A*24:02 restricted, 9-mer WT1-235 peptide (CYTWNQMNL) vaccine induces cellular immune responses and elicits WT1-235-specific cytotoxic T lymphocytes (CTLs). However, whether this vaccine induces humoral immune responses to produce WT1 antibody remains unknown. Thus, we measured IgG antibody levels against the WT1-235 peptide (WT1-235 IgG antibody) in patients with glioblastoma multiforme (GBM) receiving the WT1 peptide vaccine. The WT1-235 IgG antibody, which was undetectable before vaccination, became detectable in 30 (50.8%) of a total of 59 patients during 3 months of WT1 peptide vaccination. The dominant WT1-235 IgG antibody subclass was Th1-type, IgG1 and IgG3 . WT1-235 IgG antibody production was significantly and positively correlated with both progression-free survival (PFS) and overall survival (OS). Importantly, the combination of WT1-235 IgG antibody production and positive delayed type-hypersensitivity (DTH) to the WT1-235 peptide was a better prognostic marker for long-term OS than either parameter alone. These results suggested that WT1-235 peptide vaccination induces not only WT1-235-specific CTLs as previously described but also WT1-235-specific humoral immune responses associated with antitumor cellular immune response. Our results indicate that the WT1 IgG antibody against the WT1 peptide may be a useful predictive marker, with better predictive performance in combination with DTH to WT1 peptide, and provide a new insight into the antitumor immune response induction in WT1 peptide vaccine-treated patients.
Assuntos
Vacinas Anticâncer/imunologia , Glioblastoma/imunologia , Glioblastoma/mortalidade , Imunoglobulina G/imunologia , Peptídeos/imunologia , Proteínas WT1/imunologia , Adulto , Idoso , Biomarcadores , Vacinas Anticâncer/administração & dosagem , Linhagem Celular Tumoral , Terapia Combinada , Ensaio de Imunoadsorção Enzimática , Feminino , Glioblastoma/terapia , Antígeno HLA-A24/imunologia , Humanos , Imunoglobulina G/sangue , Imunoterapia , Masculino , Pessoa de Meia-Idade , Peptídeos/administração & dosagem , Prognóstico , Linfócitos T Citotóxicos/imunologia , Linfócitos T Citotóxicos/metabolismo , Células Th1/imunologia , Células Th1/metabolismo , Resultado do Tratamento , Vacinação , Proteínas WT1/química , Adulto JovemRESUMO
In our previous study, we showed that miR-125a directly targeted a WT1 oncogene, which was overexpressed in leukemia and various kinds of solid tumors including lung, breast, gastric, and colon cancers, and brain tumors and was deeply involved in leukemogenesis and tumorigenesis and that miR-125a knockout mice overexpressed WT1 and developed myeloproliferative disease. It had been also reported that miR-125a is downregulated in leukemia and various types of solid tumors such as lung cancers, suggesting its tumor suppressor function. Therefore, it is important to elucidate what is target(s) of miR-125a for understandings of such functions although few target genes for it are known. In the present study, Zbtb7a oncogene was identified as a potential target for miR-125a by gene expression profiling in miR-125a knockout mice combined with bioinformatics target prediction. EGFP-3'UTR reporter assay showed that miR-125a suppressed Zbtb7a expression through its direct binding to the Zbtb7a-3'UTR. Zbtb7a knockdown by siRNA suppressed cell proliferation and induced G1 cell cycle arrest and apoptosis in lung cancer cells. Furthermore, miR-125a expression showed a negative correlation with Zbtb7a expression in non-small cell lung cancer tissues. The present study showed for the first time that Zbtb7a was a direct target for miR-125a and was involved in cell cycle progression and apoptosis of lung cancer cells. These results also demonstrated that deregulation of miR-125a-Zbtb7a signaling was associated with the development and progression of lung cancer. © 2015 Wiley Periodicals, Inc.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Proteínas de Ligação a DNA/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/genética , MicroRNAs/genética , Fatores de Transcrição/genética , Animais , Apoptose , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Proliferação de Células , Humanos , Pulmão/metabolismo , Pulmão/patologia , Neoplasias Pulmonares/patologia , Camundongos Knockout , Proto-Oncogene MasRESUMO
BACKGROUND: Advances in cancer immunotherapy in the pediatric field are needed in order to improve the prognosis of children with malignancies. We conducted a prospective phase I/II study of WT1 peptide vaccination for children with relapsed or refractory malignancies. METHODS: The main eligibility criteria were affected tissues or leukemic cells expressing the WT1 gene, and patients (and donors for allogeneic hematopoietic stem cell transplantation) having HLA-A*24:02. Vaccination using the WT1 peptide (CYTWNQMNL), which was modified for higher affinity to this HLA-type molecule with the adjuvant Montanide ISA51, was performed weekly 12 times. RESULTS: Twenty-six patients were enrolled and 13 (50.0%) completed the vaccination 12 times. Evidence for the induction of WT1-specific cytotoxic T-lymphocyte (CTL) responses without severe systemic side effects was obtained. Two out of 12 patients with bulky disease exhibited a transient clinical effect (one mixed response and one stable disease), three out of six patients with minimal residual disease achieved transient molecular remission, and five out of eight patients without a detectable level of the molecular marker, but with a high risk of relapse, had the best outcome of long-term continuous complete remission. CONCLUSIONS: WT1 vaccination is a safe immunotherapy and induced WT1-specific CTL responses in children; however, as a single agent, vaccination only provided patients in remission, but with a high risk of relapse, with "long-term benefits" in the context of its use for relapse prevention. WT1 peptide-based treatments in combination with other modalities, such as anti-tumor drugs or immunomodulating agents, need to be planned.
Assuntos
Vacinas Anticâncer/uso terapêutico , Neoplasias/terapia , Peptídeos/uso terapêutico , Proteínas WT1/imunologia , Proteínas WT1/uso terapêutico , Adjuvantes Imunológicos , Adolescente , Antineoplásicos/uso terapêutico , Criança , Pré-Escolar , Feminino , Citometria de Fluxo , Humanos , Imuno-Histoquímica , Lactente , Recém-Nascido , Masculino , Manitol/administração & dosagem , Manitol/análogos & derivados , Neoplasias/genética , Ácidos Oleicos/administração & dosagem , Peptídeos/imunologia , Reação em Cadeia da Polimerase , Linfócitos T Citotóxicos/imunologiaRESUMO
Wilms' tumor gene 1 (WT1) protein is a promising tumor-associated antigen for cancer immunotherapy. We have been performing WT1 peptide vaccination with good clinical responses in over 750 patients with leukemia or solid cancers. In this study, we generated single-cell gene-expression profiles of the effector memory (EM) subset of WT1-specific cytotoxic T lymphocytes (CTLs) in peripheral blood of nine acute myeloid leukemia patients treated with WT1 peptide vaccine, in order to discriminate responders (WT1 mRNA levels in peripheral blood decreased to undetectable levels, decreased but stayed at abnormal levels, were stable at undetectable levels, or remained unchanged from the initial abnormal levels more than 6 months after WT1 vaccination) from non-responders (leukemic blast cells and/or WT1 mRNA levels increased relative to the initial state within 6 months of WT1 vaccination) prior to WT1 vaccination. Cluster and principal component analyses performed using 83 genes did not discriminate between responders and non-responders prior to WT1 vaccination. However, these analyses revealed that EM subset of WT1-specific CTLs could be divided into two groups: the "activated" and "quiescent" states; in responders, EM subset of the CTLs shifted to the "quiescent" state, whereas in non-responders, those shifted to the "activated" state following WT1 vaccination. These results demonstrate for the first time the existence of two distinct EM states, each of which was characteristic of responders or non-responders, of WT1-specific CTLs in AML patients, and raises the possibility of using advanced gene-expression profile analysis to clearly discriminate between responders and non-responders prior to WT1 vaccination.
Assuntos
Antígenos de Neoplasias/imunologia , Memória Imunológica/imunologia , Leucemia Mieloide Aguda/imunologia , Linfócitos T Citotóxicos/imunologia , Proteínas WT1/imunologia , Adulto , Idoso , Antígenos de Neoplasias/genética , Vacinas Anticâncer/imunologia , Vacinas Anticâncer/uso terapêutico , Feminino , Perfilação da Expressão Gênica , Humanos , Imunoterapia/métodos , Leucemia Mieloide Aguda/terapia , Masculino , Pessoa de Meia-Idade , Análise de Componente Principal , RNA Mensageiro/sangue , RNA Mensageiro/genética , Linfócitos T Citotóxicos/citologia , Vacinas de Subunidades Antigênicas/imunologia , Vacinas de Subunidades Antigênicas/uso terapêutico , Proteínas WT1/genéticaRESUMO
To investigate the safety of combined Wilms tumor 1 peptide vaccination and temozolomide treatment of glioblastoma, a phase I clinical trial was designed. Seven patients with histological diagnosis of glioblastoma underwent concurrent radiotherapy and temozolomide therapy. Patients first received Wilms tumor 1 peptide vaccination 1 week after the end of combined concurrent radio/temozolomide therapy, and administration was continued once per week for 7 weeks. Temozolomide maintenance was started and performed for up to 24 cycles, and the observation period for safety encompassed 6 weeks from the first administration of maintenance temozolomide. All patients showed good tolerability during the observation period. Skin disorders, such as grade 1/2 injection-site reactions, were observed in all seven patients. Although grade 3 lymphocytopenia potentially due to concurrent radio/temozolomide therapy was observed in five patients (71.4 %), no other grade 3/4 hematological or neurological toxicities were observed. No autoimmune reactions were observed. All patients are still alive, and six are on Wilms tumor 1 peptide vaccination without progression, yielding a progression-free survival from histological diagnosis of 5.2-49.1 months. Wilms tumor 1 peptide vaccination was stopped in one patient after 12 injections by the patient's request. The safety profile of the combined Wilms tumor 1 peptide vaccination and temozolomide therapy approach for treating glioblastoma was confirmed.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Neoplasias Encefálicas/terapia , Vacinas Anticâncer/administração & dosagem , Dacarbazina/análogos & derivados , Glioblastoma/terapia , Proteínas WT1/administração & dosagem , Proteínas WT1/imunologia , Adulto , Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/imunologia , Estudos de Coortes , Terapia Combinada , Dacarbazina/administração & dosagem , Dacarbazina/efeitos adversos , Progressão da Doença , Feminino , Glioblastoma/tratamento farmacológico , Glioblastoma/imunologia , Humanos , Masculino , Pessoa de Meia-Idade , Temozolomida , Proteínas WT1/efeitos adversosAssuntos
Antineoplásicos Imunológicos/uso terapêutico , Vacinas Anticâncer/uso terapêutico , Leucemia Mieloide Aguda/tratamento farmacológico , Proteínas WT1/imunologia , Adulto , Idoso , Intervalo Livre de Doença , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Fatores de Risco , Prevenção Secundária , Resultado do TratamentoRESUMO
Th17 plays important roles in the pathogenesis of various inflammatory and autoimmune diseases. Although the importance of Th17 in tumor immunity has also been suggested, precise roles of tumor-associated antigen-specific Th17 still remain poorly understood, especially in humans. We previously identified WT1332, a 16-mer helper epitope derived from tumor-associated antigen Wilms' tumor gene 1 (WT1) product, and WT1332-specific Th1 clones were established. In the present study, WT1-specific Th17 clones were established by the stimulation of peripheral blood mononuclear cells with the WT1332 helper peptide under human Th17-polarizing conditions. The WT1-specific Th17 clone exhibited the helper function for proliferation of conventional CD4(+) T cells in the antigenic stimulation-specific manner. This is the first report of establishment of functional Th17 clones with both antigen (WT1332) specificity and antigen-specific helper activity. Th17 clones established here and the method to establish antigen-specific Th17 clones should be a useful tool to further analyze the roles of human Th17 in tumor immunity.
Assuntos
Epitopos de Linfócito T/imunologia , Células Th17/imunologia , Proteínas WT1/imunologia , Células Clonais , Antígenos HLA-DP/imunologia , Cadeias beta de HLA-DQ/imunologia , Antígenos HLA-DR/imunologia , Humanos , Interleucina-17/biossíntese , Interleucina-17/imunologia , Células Th17/citologiaRESUMO
No standard treatment has been established for most rare cancers. Here, we report a clinical trial of a biweekly WT1 tri-peptide-based vaccine for recurrent or advanced rare cancers. Due to the insufficient number of patients available for a traditional clinical trial, the trial was designed for rare cancers expressing shared target molecule WT1. The recruitment criteria included WT1-expressing tumors as well as HLA-A*24:02 or 02:01. The primary endpoints were immunoglobulin G (IgG) antibody (Ab) production against the WT1-235 cytotoxic T lymphocyte (CTL) epitope and delayed-type hypersensitivity (DTH) skin reactions to targeted WT1 CTL epitopes. The secondary endpoints were safety and clinical efficacy. Forty-five patients received WT1 Trio, and 25 (55.6%) completed the 3-month protocol treatment. WT1-235 IgG Ab was positive in 88.0% of patients treated with WT1 Trio at 3 months, significantly higher than 62.5% of the weekly WT1-235 CTL peptide vaccine. The DTH positivity rate in WT1 Trio was 62.9%, which was not significantly different from 60.7% in the WT1-235 CTL peptide vaccine. The WT1 Trio safety was confirmed without severe treatment-related adverse events, except grade 3 myasthenia gravis-like symptoms observed in a patient with thymic cancer. Fifteen (33.3%) patients achieved stable disease after 3 months of treatment. In conclusion, the biweekly WT1 Trio vaccine containing the WT1-332 helper T lymphocyte peptide induced more robust immune responses targeting WT1 than the weekly WT1-235 CTL peptide vaccine. Therefore, WT1-targeted immunotherapy may be a potential therapeutic strategy for rare cancers.
RESUMO
Wilms' tumor gene 1 (WT1) protein is a promising tumor-associated antigen. In patients with WT1-expressing malignancies, WT1-specific CTLs are spontaneously induced as a result of an immune response to the WT1 protein. In the present study, we performed single cell-level comparative analysis of T cell receptor ß-chain variable region (TCR-BV) gene families of a total of 750 spontaneously induced WT1(126) peptide (amino acids 126-134, WT1(126))-specific CTLs in both HLA-A*0201(+) patients with solid tumors and healthy donors (HDs). This is the first report of direct usage analysis of 24 kinds of TCR-BV gene families of WT1(126)-specific CTLs at the single cell level. Usage analysis with single-cell RT-PCR of TCR-BV gene families of individual FACS-sorted WT1(126) tetramer(+) CD8(+) T cells showed, for the first time, that: (i) BVs 3, 6, 7, 20, 27, and 28 were commonly biased in patients and HDs; (ii) BVs 2, 11, and 15 were biased only in patients; and (iii) BVs 4, 5, 9, and 19 were biased only in HDs. However, statistical analysis of similarity of individual usage frequencies of 24 kinds of TCR-BV gene families between patients and HDs indicated that the usage frequencies of TCR-BV gene families in patients reflected those in HDs. These results should provide us with a novel insight for a better understanding of WT1-specific immune responses.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Neoplasias/genética , Neoplasias/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Adolescente , Adulto , Separação Celular , Epitopos de Linfócito T/genética , Epitopos de Linfócito T/imunologia , Feminino , Citometria de Fluxo , Genes do Tumor de Wilms , Antígeno HLA-A2/genética , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Receptores de Antígenos de Linfócitos T alfa-beta/imunologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Adulto JovemRESUMO
Monoclonal antibody (mAb) drugs are desirable for the improvement of multiple myeloma (MM) treatment. In this study, we found for the first time that CD48 was highly expressed on MM plasma cells. In 22 out of 24 MM patients, CD48 was expressed on more than 90% of MM plasma cells at significantly higher levels than it was on normal lymphocytes and monocytes. CD48 was only weakly expressed on some CD34(+) haematopoietic stem/progenitor cells, and not expressed on erythrocytes or platelets. We next examined whether CD48 could serve as a target antigen for mAb therapy against MM. A newly generated in-house anti-CD48 mAb induced mild antibody-dependent cell-mediated cytotoxicity and marked complement-dependent cytotoxicity against not only MM cell lines but also primary MM plasma cells in vitro. Administration of the anti-CD48 mAb significantly inhibited tumour growth in severe combined immunodeficient mice inoculated subcutaneously with MM cells. Furthermore, anti-CD48 mAb treatment inhibited growth of MM cells transplanted directly into murine bone marrow. Finally and importantly, we demonstrated that the anti-CD48 mAb did not damage normal CD34(+) haematopoietic stem/progenitor cells. These results suggest that the anti-CD48 mAb has the potential to become an effective therapeutic mAb against MM.
Assuntos
Anticorpos Monoclonais/farmacologia , Antígenos CD/imunologia , Mieloma Múltiplo/imunologia , Mieloma Múltiplo/terapia , Animais , Anticorpos Monoclonais/imunologia , Citotoxicidade Celular Dependente de Anticorpos/efeitos dos fármacos , Citotoxicidade Celular Dependente de Anticorpos/imunologia , Antígenos CD/biossíntese , Antígeno CD48 , Linhagem Celular Tumoral , Feminino , Humanos , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Camundongos Transgênicos , Terapia de Alvo Molecular/métodos , Mieloma Múltiplo/patologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Memory T cells play an essential role in infectious and tumor immunity. Vitamin A metabolites such as retinoic acid are immune modulators, but the role of vitamin A metabolism in memory T-cell differentiation is unclear. In this study, we identified retinol dehydrogenase 10 (Rdh10), which metabolizes vitamin A to retinal (RAL), as a key molecule for regulating T cell differentiation. T cell-specific Rdh10 deficiency enhanced memory T-cell formation through blocking RAL production in infection model. Epigenetic profiling revealed that retinoic acid receptor (RAR) signaling activated by vitamin A metabolites induced comprehensive epigenetic repression of memory T cell-associated genes, including TCF7, thereby promoting effector T-cell differentiation. Importantly, memory T cells generated by Rdh deficiency and blocking RAR signaling elicited potent anti-tumor responses in adoptive T-cell transfer setting. Thus, T cell differentiation is regulated by vitamin A metabolism and its signaling, which should be novel targets for memory T cell-based cancer immunotherapy.
Assuntos
Neoplasias , Vitamina A , Oxirredutases do Álcool/genética , Oxirredutases do Álcool/metabolismo , Imunoterapia , Células T de Memória , Neoplasias/terapia , Tretinoína/farmacologia , Vitamina A/metabolismoRESUMO
The HLA-A*24:02-restricted peptide vaccine targeting Wilms' tumor 1 (WT1) (WT1 vaccine) is a promising therapeutic strategy for ovarian cancer; however, its efficacy varies among patients. In this study, we analyzed WT1-specific immune responses in patients with advanced or recurrent ovarian cancer that was refractory to standard chemotherapies and their associations with clinical outcomes. In 25 patients, the WT1 vaccine was administered subcutaneously weekly for 3 months and biweekly thereafter until disease progression or severe adverse events. We assessed Wilms' tumor 1-specific cytotoxic T lymphocytes (WT1-CTLs) and Wilms' tumor 1 peptide-specific immunoglobulin G (WT1235-IgG). After vaccination, the percentage of tetramer high-avidity population of WT1-CTLs among CD8+ T lymphocytes (%tet-hi WT1-CTL) and the WT1235-IgG titer increased significantly, although the values were extremely low or below the limit of detection before vaccination (%tet-hi WT1-CTL: 0.003%-0.103%.; WT1235-IgG: <0.05-0.077 U/mL). Patients who had %tet-hi WT1-CTL of ≥0.25% (n=6) or WT1235-IgG of ≥0.10 U/mL (n=12) had a significantly longer progression-free survival than those of patients in the other groups. In addition, an increase in WT1235-IgG corresponded to a significantly longer progression-free survival (P=0.0496). In patients with systemic inflammation, as evidenced by elevated C-reactive protein levels, the induction of tet-hi WT1-CTL or WT1235-IgG was insufficient. Decreased serum albumin levels, multiple tumor lesions, poor performance status, and excess ascites negatively influenced the clinical effectiveness of the WT1 vaccine. In conclusion, the WT1 vaccine induced antigen-specific cellular and humoral immunity in patients with refractory ovarian cancer. Both %tet-hi WT1-CTL and WT1235-IgG levels are prognostic markers for the WT1 vaccine.