RESUMO
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.
Assuntos
Vacinas Anticâncer , Neoplasias Renais , Neoplasias Ovarianas , Humanos , Imunidade Humoral , Recidiva Local de Neoplasia , Neoplasias Ovarianas/terapia , Peptídeos , Linfócitos T Citotóxicos , Vacinas de Subunidades Antigênicas , Proteínas WT1RESUMO
The cell wall skeleton of Bacillus Calmette-Guérin (BCG-CWS) is a bioactive component that is a strong immune adjuvant for cancer immunotherapy. BCG-CWS activates the innate immune system through various pattern recognition receptors and is expected to elicit antigen-specific cellular immune responses when co-administered with tumor antigens. To determine the recommended dose (RD) of BCG-CWS based on its safety profile, we conducted a phase I dose-escalation study of BCG-CWS in combination with WT1 peptide for patients with advanced cancer.The primary endpoint was the proportion of treatment-related adverse events (AEs) at each BCG-CWS dose. The secondary endpoints were immune responses and clinical effects. A BCG-CWS dose of 50, 100, or 200âµg/body was administered intradermally on days 0, 7, 21, and 42, followed by 2âmg of WT1 peptide on the next day. For the escalation of a dose level, 3â+â3 design was used.Study subjects were 18 patients with advanced WT1-expressing cancers refractory to standard anti-cancer therapies (7 melanoma, 5 colorectal, 4 hepatobiliary, 1 ovarian, and 1 lung). Dose-limiting toxicity occurred in the form of local skin reactions in 2 patients at a dose of 200âµg although no serious treatment-related systemic AEs were observed. Neutrophils and monocytes transiently increased in response to BCG-CWS. Some patients demonstrated the induction of the CD4 T cell subset and its differentiation from the naïve to memory phenotype, resulting in a tumor response.The RD of BCG-CWS was determined to be 100âµg/body. This dose was well tolerated and showed promising clinical effects with the induction of an appropriate immune response.
Assuntos
Adjuvantes Imunológicos/uso terapêutico , Vacina BCG/uso terapêutico , Esqueleto da Parede Celular/uso terapêutico , Mycobacterium bovis , Adjuvantes Imunológicos/administração & dosagem , Adulto , Idoso , Vacina BCG/administração & dosagem , Contagem de Linfócito CD4 , Esqueleto da Parede Celular/administração & dosagem , Neoplasias Colorretais/tratamento farmacológico , Feminino , Humanos , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Pulmonares/tratamento farmacológico , Masculino , Melanoma/tratamento farmacológico , Pessoa de Meia-Idade , Neoplasias Ovarianas/tratamento farmacológico , Neoplasias Cutâneas/tratamento farmacológico , Resultado do TratamentoRESUMO
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
OBJECTIVE: The aim of the present study was to analyze the long-term survival effects of WT1 peptide vaccine, in addition to its anti-tumor effects and toxicity. METHODS: A phase II clinical trial was conducted during the period of 2004-2010 at Osaka University Hospital, Osaka, Japan. The patients who had gynecologic malignancies progressing against previous treatments received WT1 peptide vaccine intradermally at 1-week intervals for 12 weeks. The vaccination was allowed to further continue, unless the patient's condition became significantly worse due to the disease progression. RESULTS: Forty out of 42 patients, who met all the inclusion criteria, underwent WT1 peptide vaccine. Among these 40 patients, stable disease was observed in 16 cases (40 %). Skin toxicity of a grade 1, 2 and 3 occurred in 25 cases (63 %), 9 cases (23 %) and a single case (3 %), respectively, and liver toxicity of grade 1 in a single case (3 %). The overall survival period was significantly longer in cases positive for the WT1 peptide-specific delayed-type hypersensitivity (DTH) reaction after the vaccination, compared to those negative for the DTH reaction (p = 0.023). Multivariate Cox proportional hazards analysis demonstrated that the adjusted hazard ratio for the negative DTH reaction was 2.73 (95 % CI 1.04-7.19, p = 0.043). CONCLUSION: WT1 peptide vaccine may be a potential treatment, with limited toxicity, for gynecologic malignancies that have become resistant to conventional therapies. Larger scale of clinical studies is required to establish the efficacy of the WT1 peptide vaccine for gynecologic malignancies.
Assuntos
Carcinoma/terapia , Neoplasias dos Genitais Femininos/terapia , Oligopeptídeos/uso terapêutico , Proteínas WT1/uso terapêutico , Adulto , Idoso , Carcinoma/mortalidade , Carcinoma/patologia , Progressão da Doença , Feminino , Neoplasias dos Genitais Femininos/mortalidade , Neoplasias dos Genitais Femininos/patologia , Humanos , Imunoterapia/efeitos adversos , Imunoterapia/métodos , Pessoa de Meia-Idade , Oligopeptídeos/efeitos adversos , Terapia de Salvação , Análise de Sobrevida , Falha de Tratamento , Resultado do Tratamento , Proteínas WT1/efeitos adversosRESUMO
Wilms' tumor (WT1) protein is one of the most promising target antigens for cancer immunotherapy. In fact, clinical responses, such as growth stabilization or shrinkage of tumor with immunological responses, have been reported in patients vaccinated with WT1 peptide. Here, we performed WT1 peptide-based immunotherapy for a patient with chemotherapy-resistant salivary gland cancer, whose histologic type was carcinoma ex pleomorphic adenoma. The patient with its pulmonary metastasis, refractory to chemotherapy, was intradermally injected with 3 mg of WT1 peptide emulsified with Montanide ISA51 adjuvant at one-week intervals for 12 weeks. The considerably rapid growth of tumor was inhibited after WT1 vaccination, and stable disease, lasting three months, was achieved. Concomitantly, immunological responses, i.e. an increase in frequencies of WT1 tetramer(+) CD8(+)T cells and delayed type hypersensitivity response, were detected after the vaccination. These results indicate the potential of WT1 peptide-based immunotherapy for the treatment of chemotherapy-resistant salivery gland cancer.
Assuntos
Antineoplásicos/uso terapêutico , Neoplasias das Glândulas Salivares/tratamento farmacológico , Proteínas WT1/uso terapêutico , Humanos , Masculino , Pessoa de Meia-Idade , Proteínas WT1/químicaRESUMO
Aurora kinase A (AURKA) is overexpressed in leukemias. Previously, we demonstrated that AURKA-specific CD8(+) T cells specifically and selectively lysed leukemia cells, indicating that AURKA is an excellent target for immunotherapy. In this study, we examined the feasibility of adoptive therapy using redirected T cells expressing an HLA-A*0201-restricted AURKA(207-215)-specific T-cell receptor (TCR). Retrovirally transduced T cells recognized relevant peptide-pulsed but not control target cells. Furthermore, TCR-redirected CD8(+) T cells lysed AURKA-overexpressing human leukemic cells in an HLA-A*0201-restricted manner, but did not kill HLA-A*0201(+) normal cells, including hematopoietic progenitors. In addition, AURKA(207-215)-specific TCR-transduced CD4(+) T cells displayed target-responsive Th1 cytokine production. Finally, AURKA(207-215)-specific TCR-transduced CD8(+) T cells displayed antileukemia efficacy in a xenograft mouse model. Collectively, these data demonstrate the feasibility of redirected T cell-based AURKA-specific immunotherapy for the treatment of human leukemia.
Assuntos
Genes Codificadores dos Receptores de Linfócitos T/genética , Terapia Genética , Imunoterapia , Leucemia/terapia , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/imunologia , Linfócitos T/imunologia , Animais , Aurora Quinase A , Aurora Quinases , Western Blotting , Estudos de Casos e Controles , Estudos de Viabilidade , Citometria de Fluxo , Antígeno HLA-A2/imunologia , Humanos , Leucemia/genética , Leucemia/imunologia , Luciferases/metabolismo , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Receptores de Interleucina-2/fisiologia , Especificidade do Receptor de Antígeno de Linfócitos T , Linfócitos T Citotóxicos/imunologia , Transdução Genética , Células Tumorais CultivadasRESUMO
To induce and activate tumor-associated antigen-specific cytotoxic T lymphocytes (CTLs) for cancer immunity, it is important not only to select potent CTL epitopes but also to combine them with appropriate immunopotentiating agents. Here we investigated whether tumor immunity induced by WT1 peptide vaccination could be enhanced by IFN-ß. For the experimental group, C57BL/6 mice were twice pre-treated with WT1 peptide vaccine, implanted with WT1-expressing C1498 cells, and treated four times with WT1 peptide vaccine at one-week intervals. During the vaccination period, IFN-ß was injected three times a week. Mice in control groups were treated with WT1 peptide alone, IFN-ß alone, or PBS alone. The mice in the experimental group rejected tumor cells and survived significantly longer than mice in the control groups. The overall survival on day 75 was 40% for the mice treated with WT1 peptide+IFN-ß, while it was 7, 7, and 0% for those treated with WT1 peptide alone, IFN-ß alone or PBS alone, respectively. Induction of WT1-specific CTLs and enhancement of NK activity were detected in splenocytes from mice in the experimental group. Furthermore, administration of IFN-ß enhanced expression of MHC class I molecules on the implanted tumor cells. In conclusion, our results showed that co-administration of WT1 peptide+IFN-ß enhanced tumor immunity mainly through the induction of WT1-specific CTLs, enhancement of NK activity, and promotion of MHC class I expression on the tumor cells. WT1 peptide vaccination combined with IFN-ß administration can thus be expected to enhance the clinical efficacy of WT1 immunotherapy.
Assuntos
Adjuvantes Imunológicos/administração & dosagem , Vacinas Anticâncer/imunologia , Interferon beta/administração & dosagem , Neoplasias Renais/prevenção & controle , Proteínas WT1/imunologia , Tumor de Wilms/prevenção & controle , Animais , Vacinas Anticâncer/administração & dosagem , Neoplasias Renais/imunologia , Células Matadoras Naturais/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Baço/imunologia , Análise de Sobrevida , Linfócitos T Citotóxicos/imunologia , Proteínas WT1/administração & dosagem , Tumor de Wilms/imunologiaRESUMO
Adoptive T-cell therapy for malignancies using redirected T cells genetically engineered by tumor antigen-specific T-cell receptor (TCR) gene transfer is associated with mispairing between introduced and endogenous TCR chains with unknown specificity. Therefore, deterioration of antitumor reactivity and serious autoimmune reactivity are major concerns. To address this problem, we have recently established a novel retroviral vector system encoding siRNAs for endogenous TCR genes (siTCR vector). In this study, to test the clinical application of siTCR gene therapy for human leukemia, we examined in detail the efficacy and safety of WT1-siTCR-transduced T cells. Compared with conventional WT1-TCR (WT1-coTCR) gene-transduced T cells, these cells showed significant enhancement of antileukemia reactivity resulting from stronger expression of the introduced WT1-specific TCR with inhibition of endogenous TCRs. Notably, WT1-siTCR gene-transduced T cells were remarkably expandable after repetitive stimulation with WT1 peptide in vitro, without any deterioration of antigen specificity. WT1-siTCR gene-transduced T cells from leukemia patients successfully lysed autologous leukemia cells, but not normal hematopoietic progenitor cells. In a mouse xenograft model, adoptively transferred WT1-siTCR gene-transduced T cells exerted distinct antileukemia efficacy but did not inhibit human hematopoiesis. Our results suggest that gene-immunotherapy for leukemia using this WT1-siTCR system holds considerable promise.
Assuntos
Imunoterapia Adotiva/métodos , Leucemia/terapia , RNA Interferente Pequeno/genética , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Proteínas WT1/genética , Animais , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Células Cultivadas , Citotoxicidade Imunológica/imunologia , Feminino , Terapia Genética/métodos , Humanos , Subunidade gama Comum de Receptores de Interleucina/deficiência , Subunidade gama Comum de Receptores de Interleucina/genética , Células Jurkat , Células K562 , Leucemia/genética , Leucemia/patologia , Camundongos , Camundongos Endogâmicos NOD , Camundongos Knockout , Camundongos SCID , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/imunologia , Fragmentos de Peptídeos/metabolismo , RNA Interferente Pequeno/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/imunologia , Receptores de Antígenos de Linfócitos T alfa-beta/metabolismo , Linfócitos T Citotóxicos/imunologia , Linfócitos T Citotóxicos/metabolismo , Transfecção , Células Tumorais Cultivadas , Proteínas WT1/química , Proteínas WT1/imunologia , Proteínas WT1/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
How to treat CML patients who are resistant to inhibitors of BCR-ABL tyrosine kinase such as Imatinib is a very important and urgent issue in clinical hematology. Here, we report a case of Imatinib-treated CML in which intradermally administered WT1 peptide vaccine elicited WT1-specific immune responses and the resultant reduction in the persistent residual disease in co-administration of Imatinib. BCR-ABL mRNA levels were being maintained under the detection limit for 8 months since week 77 of vaccination. No adverse effects except local erythema at the injection sites were observed. The tetramer assay revealed that the decrease in BCR-ABL mRNA levels was associated with the increase in frequency of WT1-specific cytotoxic T lymphocytes, notably effector-memory type of that, in the patient's peripheral blood. The case presented here indicates that WT1 peptide vaccine may become a safe and cure-oriented therapy for CML patients who have residual disease regardless of the treatment with Imatinib.
Assuntos
Vacinas Anticâncer/uso terapêutico , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/terapia , Proteínas WT1/imunologia , Idoso , Antineoplásicos/uso terapêutico , Benzamidas , Vacinas Anticâncer/química , Vacinas Anticâncer/imunologia , Terapia Combinada , Resistencia a Medicamentos Antineoplásicos , Feminino , Proteínas de Fusão bcr-abl/antagonistas & inibidores , Humanos , Mesilato de Imatinib , Leucemia Mielogênica Crônica BCR-ABL Positiva/imunologia , Neoplasia Residual , Piperazinas/uso terapêutico , Proteínas Tirosina Quinases/antagonistas & inibidores , Pirimidinas/uso terapêutico , RNA Mensageiro , Linfócitos T Citotóxicos/imunologia , Vacinas de Subunidades Antigênicas/imunologia , Vacinas de Subunidades Antigênicas/uso terapêutico , Proteínas WT1/químicaRESUMO
Clinical studies of WT1-targeted cancer vaccine are being performed. However, WT1-specific Th response in cancer patients remains unclear. Using quantitative real-time RT-PCR, we investigated IFN-gamma and IL-10 mRNA expression from Th cells by stimulation with helper peptide WT1(332). Seventeen patients, of whom 10 had achieved stable disease and the remaining 7 had progressive disease, were weekly vaccinated with WT1 CTL epitope (modified WT1(235)) and examined for WT1(332)-specific Th response. A clear correlation between WT1(332)-specific Th response and clinical response was observed at 4 weeks post-vaccination. In patients who responded, a clear inverse correlation between IL-10-type and IFN-gamma-type WT1(332)-specific Th response was detected at pre- and 4 weeks post-vaccination, and the shift of the Th response from IL-10-type dominancy at early phase to IFN-gamma-type dominancy at late phase was observed. From this study we concluded that occurrence of WT1(332)-specific Th response could predict good clinical response of WT1 CTL epitope vaccination.
Assuntos
Vacinas Anticâncer/imunologia , Epitopos de Linfócito T , Linfócitos T Citotóxicos/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Vacinação , Proteínas WT1/imunologia , Adulto , Idoso , Feminino , Humanos , Interferon gama/genética , Interleucina-10/genética , Masculino , Pessoa de Meia-Idade , RNA Mensageiro/análiseRESUMO
WT1 (Wilms' tumor gene 1) protein is a potent pan-tumor-associated antigen (TAA) and WT1-specific cytotoxic T lymphocytes (WT1 tetramer(+) CD8(+) T cells) are spontaneously induced in patients with acute myeloid leukemia (AML) or myelodysplastic syndrome (MDS). We conducted a single-cell level comparative analysis of T-cell receptor beta-chain variable region (TCR-BV) gene families of a total of 1242 spontaneously induced WT1 tetramer(+) CD8(+) T cells in HLA-A*2402(+) patients with AML or MDS and those in healthy donors (HDs). This is the first report of direct usage analysis of TCR-BV gene families of individual TAA-specific CD8(+) T cells at single-cell level. Usage analysis using single-cell RT-PCR of TCR-BV gene families of individual FACS-sorted WT1 tetramer(+) CD8(+) T cells showed for the first time (i) that BVs 5, 6, 20, and 27 were commonly biased in both HDs and patients; (ii) that BV4 was commonly biased in HDs and MDS patients; (iii) that BV19 was commonly biased in the patients; and (iv) that BVs 7 and 28, BVs 9 and 15, and BVs 12 and 29 were specifically biased in HDs, AML, and MDS patients, respectively. However, statistical analysis of similarity among HD, AML, and MDS of individual usage frequencies of 24 kinds of TCR-BV gene families indicated that the usage frequencies of TCR-BV gene families in AML and MDS patients reflect those in HDs. These findings represent a novel insight for a better understanding of WT1-specific immune response.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Genes Codificadores da Cadeia beta de Receptores de Linfócitos T , Leucemia Mieloide Aguda/imunologia , Síndromes Mielodisplásicas/imunologia , Proteínas WT1/imunologia , Adulto , Idoso , Feminino , Humanos , Antígenos Comuns de Leucócito/análise , Masculino , Pessoa de Meia-Idade , Receptores CCR7/análiseRESUMO
In tumor-bearing patients, tumor-associated antigen (TAA)-specific CTLs are spontaneously induced as a result of immune response to TAAs and play an important role in anti-tumor immunity. Wilms' tumor gene 1 (WT1) is overexpressed in various types of tumor and WT1 protein is a promising pan-TAA because of its high immunogenicity. In this study, to clarify the immune response to the WT1 antigen, WT1-specific CD8(+) T cells that were spontaneously induced in patients with solid tumor were comparatively analyzed in both bone marrow (BM) and peripheral blood (PB). WT1-specific CD8(+) T cells more frequently existed in BM than in PB, whereas frequencies of naïve (CCR7(+) CD45RA(+)), central memory (CCR7(+) CD45RA-), effector-memory (CCR7- CD45RA(-)), and effector (CCR7- CD45RA(+)) subsets were not significantly different between BM and PB. However, analysis of these subsets for the expression of CD57 and CD28, which were associated with differentiation, revealed that effector-memory and effector subsets of the WT1-specific CD8(+) T cells in BM had less differentiated phenotypes and more proliferative potential than those in PB. Furthermore, CD107a/b functional assay for WT1 peptide-specific cytotoxic potential and carboxyfluorescein diacetate succinimidyl ester dilution assay for WT1 peptide-specific proliferation also showed that WT1-specific CD8(+) T cells in BM were less cytotoxic and more proliferative in response to WT1 peptide than those in PB. These results implied that BM played an important role as a secondary lymphoid organ in tumor-bearing patients. Preferential residence of WT1-specific CD8(+) T cells in BM could be, at least in part, explained by higher expression of chemokine receptor CCR5, whose ligand was expressed on BM fibroblasts on the WT1-specific CD8(+) T cells in BM, compared to those in PB. These results should provide us with an insight into WT1-specific immune response in tumor-bearing patients and give us an idea of enhancement of clinical response in WT1 protein-targeted immunotherapy.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Neoplasias/imunologia , Proteínas WT1/fisiologia , Adolescente , Idoso , Medula Óssea/química , Medula Óssea/imunologia , Medula Óssea/metabolismo , Linfócitos T CD8-Positivos/patologia , Diferenciação Celular/imunologia , Proliferação de Células , Feminino , Humanos , Memória Imunológica , Antígenos Comuns de Leucócito/análise , Antígenos Comuns de Leucócito/imunologia , Contagem de Linfócitos , Subpopulações de Linfócitos/imunologia , Masculino , Pessoa de Meia-IdadeRESUMO
Wilms' tumor gene (WT1) possesses oncogenic functions and is expressed in various kinds of malignancies, which suggests that the gene's product, the WT1 protein, should be one of the most promising cancer antigens. In fact, the WT1 protein was shown to be highly immunogenic in cancer patients. WT1 peptides that could induce WT1-specific CTLs (WT1 CTL peptides) were identified, and vaccination of cancer patients with these WT1 CTL peptides induced immunological responses, which were assessed by ex vivo immuno-monitoring, such as the tetramer assay, and in vivo immuno-monitoring, such as the peptide-specific delayed type hypersensitivity reaction. The induced immunological responses then led to clinical responses such as solid tumor shrinkage, a decrease in leukemia cells, and reduction of M-protein (multiple myeloma). Long-term stabilization of disease with good quality of life, which might be characteristic of cancer vaccine therapy, was also reported. It is noteworthy that injection with a "single" kind of WT1 peptide elicited an immunological response strong enough to induce a clinical response, indicating that the WT1 peptide vaccine has therapeutic potential. The number of reports of the successful treatment of cancer patients (not only adult but also childhood malignancies) with WT1 vaccination is increasing. Strategies for further improvement in the efficacy of therapy, including combined use of chemotherapy drugs, molecular-target-based drugs, or WT1 helper peptides, are being proposed. WT1 peptide vaccination in an "adjuvant setting" should be considered a promising treatment to protect against progression or relapse of malignancies in cases with minimal residual disease.
Assuntos
Antígenos de Neoplasias/imunologia , Vacinas Anticâncer/uso terapêutico , Imunoterapia , Neoplasias/terapia , Peptídeos/uso terapêutico , Proteínas WT1/química , Sequência de Aminoácidos , Animais , Vacinas Anticâncer/administração & dosagem , Vacinas Anticâncer/imunologia , Ensaios Clínicos como Assunto , Modelos Animais de Doenças , Humanos , Camundongos , Neoplasias/imunologia , Peptídeos/administração & dosagem , Peptídeos/química , Peptídeos/imunologia , Linfócitos T Citotóxicos/imunologiaRESUMO
A high level protein synthesis is one of the characteristics of cancer cells. The aim of this study is to show the contribution of eukaryotic elongation factor 2 (eEF2), which plays an essential role in the polypeptide chain elongation step, in the tumorigenesis of gastrointestinal cancers. In the present study, we demonstrated by using immunohistochemistry that eEF2 protein was overexpressed in 92.9% (13 of 14) of gastric and 91.7% (22 of 24) of colorectal cancers. No mutations were found in any of the exons of the eEF2 gene in six gastric and six colorectal cancers. Knockdown of eEF2 by eEF2-specific short-hairpin RNA (shEF2) inhibited cancer cell growth in two gastric cancer cell lines, AZ-521 and MKN28, and one colon cancer cell line, SW620. Flow cytometric analysis showed that knockdown of eEF2 induced G2/M arrest and resulted in inactivation of Akt and cdc2 (a G2/M regulator) and activation of eEF2 kinase (a negative regulator of eEF2) in these cancer cells. Conversely, forced expression of eEF2 in AZ-521 cells significantly enhanced the cell growth through promotion of G2/M progression in cell cycle, activated Akt and cdc2, and inactivated eEF2 kinase. Furthermore, forced expression of eEF2 in these cancer cells enhanced in vivo tumorigenicity in a mouse xenograft model. These results showed that overexpressed eEF2 in gastrointestinal cancers promoted G2/M progression and enhanced their cell growth in vitro and in vivo. These results also suggested a novel linkage between translational elongation and cell cycle mechanisms, implying that the linkage might play an important role to orchestrate the deregulated translation and cell cycle mechanisms for promotion of the development of gastrointestinal cancers.
Assuntos
Adenocarcinoma/genética , Divisão Celular/genética , Fase G2/genética , Neoplasias Gastrointestinais/genética , Fator 2 de Elongação de Peptídeos/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Animais , Ciclo Celular/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Modelos Biológicos , Células Tumorais Cultivadas , Regulação para Cima , Adulto JovemRESUMO
There are urgent needs to develop methods for early detection of nonsmall cell lung cancer (NSCLC) because of its increasing incidence and poor prognosis. Here, we analyzed the production of IgG antibody (WT1 Ab) against WT1 (Wilms' tumor gene) protein that was overexpressed in the majority of NSCLC. Enzyme-linked immuno-sorbent assay showed that WT1 Ab was produced in all of 91 NSCLC patients and 70 healthy individuals and that WT1 Ab titers were significantly higher in NSCLC patients compared with healthy individuals. When the cut-off level of WT1 Ab titers were fixed at mean + 3SD of those in healthy individuals, 26.4% of NSCLC patients had WT1 Ab titers over the cut-off level, and positive rates of WT1 Ab at each clinical stage were 25.0, 30.8 and 38.4% in stage I, II and III NSCLC, respectively. When WT1 Ab was combined with CEA or CYFRA for detection of NSCLC, positive detection rates increased from 25.0 to 34.1 and 31.8%, respectively, in stage I and from 38.4 to 69.2 and 46.1%, respectively, in stage III, but not changed in stage II. Western blot analysis showed that dominant subclass of WT1 Ab was Th1-type IgG2. Interestingly, elevation of WT1 Ab titers was significantly associated with longer disease-free survival in patients with stages I-III NSCLC. These results showed that WT1 Ab could be a useful marker for early detection of NSCLC and its prognostic prediction. These results also suggested that WT1-specific immune responses played an important role in anti-cancer immunity in NSCLC.
Assuntos
Biomarcadores Tumorais/sangue , Carcinoma Pulmonar de Células não Pequenas/diagnóstico , Imunoglobulina G/sangue , Neoplasias Pulmonares/diagnóstico , Proteínas WT1/genética , Western Blotting , Carcinoma Pulmonar de Células não Pequenas/imunologia , Diagnóstico Precoce , Ensaio de Imunoadsorção Enzimática , Humanos , Imuno-Histoquímica , Neoplasias Pulmonares/imunologia , PrognósticoRESUMO
The Wilms' tumor gene WT1 is overexpressed in various kinds of hematopoietic malignancies as well as solid cancers, and this protein has been demonstrated to be an attractive target antigen for cancer immunotherapy. WT1-specific CTL epitopes with a restriction of HLA-A 2402 or HLA-A 0201 have been already identified. In the present study it has been demonstrated that a 9-mer WT1-derived WT1(187) peptide, which had already been shown to elicit a WT1-specific CTL response with a restriction of HLA-A 0201, can also elicit a CTL response with a restriction of HLA-A 0206. In all three different HLA-A 0206(+) healthy donors examined, WT1(187) peptide-specific CTL could be generated from peripheral blood mononuclear cells, and the CTL showed cytotoxic activity that depended on dual expression of WT1 and HLA-A 0206 molecules. The present study describes the first identification of a HLA-A 0206-restricted, WT1-specific CTL epitope. The present results should help to broaden the application of WT1 peptide-based immunotherapy from only HLA-A 0201-positive to HLA-A 0206-positive cancer patients as well.
Assuntos
Epitopos de Linfócito T/imunologia , Antígenos HLA-A/imunologia , Linfócitos T Citotóxicos/imunologia , Proteínas WT1/imunologia , Linfócitos T CD8-Positivos/imunologia , Linhagem Celular Tumoral , Testes Imunológicos de Citotoxicidade , Mapeamento de Epitopos , Genes do Tumor de Wilms , Antígeno HLA-A2 , Humanos , Oligopeptídeos/imunologiaRESUMO
Cancer immunotherapy targeting tumor-associated antigens is now being developed. Wilms' tumor gene WT1-encoding protein is one of the promising target antigens for cancer immunotherapy, because the gene has an oncogenic function and is expressed in many kinds of malignancies. Furthermore, a series of investigations indicated that WT1 protein was highly immunogenic in cancer patients. Based on the analysis of anchor residues that were important for the interaction between peptides and HLA class I molecules, WT1 cytotoxic T lymphocyte (CTL) epitopes with the restriction of HLA-A 0201 and HLA-A 2402 were identified, and clinical trials of WT1 peptide vaccination for cancer patients with these HLA class I types were started. The vaccination-driven immunological and/or clinical responses were reported in patients with myeloid malignancies, multiple myeloma, and several solid cancers. Pediatric malignancies also may be target diseases for WT1 peptide vaccination in the future. Addition of HLA class II-restricted WT1 helper epitope peptide, chemotherapy, or molecular-target-based drug to WT1 CTL epitope peptide-based vaccination may enhance the power and usefulness of WT1 peptide vaccine. Other modalities, including gene therapy using genes encoding WT1-specific T cell receptor or DNA vaccination, are also expected to be developed.
Assuntos
Antígenos de Neoplasias/química , Antineoplásicos/farmacologia , Proteínas WT1/química , Animais , Antígenos de Neoplasias/efeitos dos fármacos , Antineoplásicos/química , Ensaios Clínicos como Assunto , Humanos , Imunoterapia , Peptídeos/química , Peptídeos/farmacologia , Proteínas WT1/efeitos dos fármacosRESUMO
OBJECT: The object of this study was to investigate the safety and clinical responses of immunotherapy targeting the WT1 (Wilms tumor 1) gene product in patients with recurrent glioblastoma multiforme (GBM). METHODS: Twenty-one patients with WT1/HLA-A*2402-positive recurrent GBM were included in a Phase II clinical study of WT1 vaccine therapy. In all patients, the tumors were resistant to standard therapy. Patients received intra-dermal injections of an HLA-A*2402-restricted, modified 9-mer WT1 peptide every week for 12 weeks. Tumor size, which was obtained by measuring the contrast-enhanced area on magnetic resonance images, was determined every 4 weeks. The responses were analyzed according to Response Evaluation Criteria in Solid Tumors (RECIST) 12 weeks after the initial vaccination. Patients who achieved an effective response continued to be vaccinated until tumor progression occurred. Progression-free survival and overall survival after initial WT1 treatment were estimated. RESULTS: The protocol was well tolerated; only local erythema occurred at the WT1 vaccine injection site. The clinical responses were as follows: partial response in 2 patients, stable disease in 10 patients, and progressive disease in 9 patients. No patient had a complete response. The overall response rate (cases with complete or partial response) was 9.5%, and the disease control rate (cases with complete or partial response as well as those in which disease was stable) was 57.1%. The median progression-free survival (PFS) period was 20.0 weeks, and the 6-month (26-week) PFS rate was 33.3%. CONCLUSIONS: Although a small uncontrolled nonrandomized trial, this study showed that WT1 vaccine therapy for patients with WT1/HLA-A*2402-positive recurrent GBM was safe and produced a clinical response. Based on these results, further clinical studies of WT1 vaccine therapy in patients with malignant glioma are warranted.
Assuntos
Vacinas Anticâncer/uso terapêutico , Glioblastoma/terapia , Proteínas Nucleares/imunologia , Adulto , Idoso , Neoplasias Encefálicas/mortalidade , Neoplasias Encefálicas/patologia , Neoplasias Encefálicas/terapia , Proteínas de Ciclo Celular , Feminino , Glioblastoma/mortalidade , Glioblastoma/patologia , Humanos , Injeções Intradérmicas , Masculino , Pessoa de Meia-Idade , Fatores de Processamento de RNA , Resultado do Tratamento , VacinaçãoRESUMO
Wilms' tumor gene WT1 is overexpressed in leukemia and various types of solid tumors and plays an important role in leukemogenesis and tumorigenesis. We tested apoptosis-inducing ability of short hairpin RNAs targeting exon 5 (shWTE5), exon10 (shWTE10) and 3'UTR (shWT3U) of the WT1 gene. Among the three WT1-shRNAs, since shWTE5 most effectively induced apoptosis, its ability as an apoptosis-inducing agent was intensively examined. shWTE5 induced mitochondrial damage and resultant apoptosis in five WT1-expressing solid cancer cells originated from gastric (AZ-521), lung (LU99B), ovarian (TYKnuCPr) cancers, fibrosarcoma (HT-1080) and glioblastoma (A172). Moreover, shWTE5 significantly enhanced apoptosis induced by chemotherapeutic agents, doxorubicin (DOX) and etoposide (ETP), or by death ligand TRAIL in all of the four solid tumor cells examined (HT-1080, LU99B, TYK and A172). Transduction of one each of WT1 isoforms with exon 5 [17AA(+)KTS(+) and 17AA(+)KTS(-)] prevented mitochondrial damage induced by ETP or TRAIL and inhibited apoptosis. These results showed that shWTE5 induced apoptosis through the suppression of the WT1 isoform with exon 5. Furthermore, shWTE5 increased expression of proapoptotic Bak and Bax proteins and decreased antiapoptotic Bcl-xL and Bcl-2 proteins in WT1-expressing HT-1080 cells, indicating that WT1 isoforms with exon 5 might play an antiapoptotic role through regulation of Bcl-2 family genes in solid tumor cells. The results presented here demonstrated that WT1-shRNA targeting exon 5 should serve as a potent anti-cancer agent for various types of solid tumors.