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1.
Anticancer Res ; 39(11): 5903-5910, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31704814

RESUMO

BACKGROUND: Gastric cancer (GC) exhibits heterogeneous clinical and molecular features, requiring the development of new biomarkers to further understand this disease. Our transcriptomic analysis detected overexpression of melanoma-associated antigen A6 (MAGEA6) in metastatic GC, leading us to determine the clinical significance of MAGEA6 in GC. MATERIALS AND METHODS: Fourteen GC cell lines and 230 pairs of surgically resected gastric tissues were subjected to mRNA expression analysis. Polymerase chain reaction array analysis was performed to identify coordinately expressed cancer-related genes, and immunohistochemistry (IHC) was used to detected MAGEA6 expression in situ. RESULTS: MAGEA6 mRNA levels were positively correlated with the expression of matrix metallopeptidase 9 mRNA. MAGEA6 mRNA levels were higher in GC tissues compared with those in normal adjacent tissues. Patients with high MAGEA6 expression had significantly worse prognosis. MAGEA6 protein levels in primary lesions predicted the likelihood of recurrence. CONCLUSION: Overexpression of MAGEA6 in GC tissues represents a promising biomarker for assessing the malignant phenotype of GC.


Assuntos
Antígenos de Neoplasias/metabolismo , Biomarcadores Tumorais/metabolismo , Gastrectomia/mortalidade , Neoplasias Hepáticas/metabolismo , Proteínas de Neoplasias/metabolismo , Recidiva Local de Neoplasia/metabolismo , Neoplasias Gástricas/metabolismo , Adenocarcinoma/metabolismo , Adenocarcinoma/secundário , Adenocarcinoma/cirurgia , Adenocarcinoma Mucinoso/metabolismo , Adenocarcinoma Mucinoso/secundário , Adenocarcinoma Mucinoso/cirurgia , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígenos de Neoplasias/genética , Biomarcadores Tumorais/genética , Carcinoma de Células em Anel de Sinete/metabolismo , Carcinoma de Células em Anel de Sinete/secundário , Carcinoma de Células em Anel de Sinete/cirurgia , Estudos de Casos e Controles , Feminino , Seguimentos , Humanos , Neoplasias Hepáticas/secundário , Neoplasias Hepáticas/cirurgia , Metástase Linfática , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica , Proteínas de Neoplasias/genética , Recidiva Local de Neoplasia/secundário , Recidiva Local de Neoplasia/cirurgia , Prognóstico , Neoplasias Gástricas/patologia , Neoplasias Gástricas/cirurgia , Taxa de Sobrevida
2.
Anticancer Res ; 39(11): 5943-5951, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31704819

RESUMO

BACKGROUND/AIM: To investigate the function of preferentially expressed antigen of melanoma (PRAME) in esophageal squamous cell carcinoma (ESCC). MATERIALS AND METHODS: mRNA expression levels of PRAME were analyzed in resected esophageal tissues of 150 ESCC patients and correlated with clinicopathological parameters. We also investigated the potential function of PRAME by analyzing coordinately expressed genes in 13 ESCC cell lines. RESULTS: RT-qPCR analysis of clinical samples revealed aberrantly high PRAME expression in tumors compared with normal esophageal tissues. High PRAME expression was significantly associated with shorter disease-specific survival and hematogenous recurrence, but not with overall recurrence. The cumulative incidence of hematogenous recurrence was significantly greater for patients with high compared to those with low PRAME expression. In vitro, PCR array analysis revealed that PRAME was coordinately expressed with EGFR, ITGB, and TCF3. CONCLUSION: PRAME is overexpressed in ESCC tissues and may serve as a novel biomarker for predicting hematogenous recurrence.


Assuntos
Antígenos de Neoplasias/metabolismo , Biomarcadores Tumorais/metabolismo , Neoplasias Esofágicas/patologia , Carcinoma de Células Escamosas do Esôfago/secundário , Recidiva Local de Neoplasia/patologia , RNA Mensageiro/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígenos de Neoplasias/genética , Biomarcadores Tumorais/genética , Estudos de Casos e Controles , Estudos de Coortes , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/metabolismo , Carcinoma de Células Escamosas do Esôfago/genética , Carcinoma de Células Escamosas do Esôfago/metabolismo , Feminino , Seguimentos , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica , Metástase Neoplásica , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/metabolismo , Prognóstico , RNA Mensageiro/genética , Taxa de Sobrevida
3.
Anticancer Res ; 39(11): 6015-6023, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31704827

RESUMO

BACKGROUND/AIM: We previously reported that expression of melanoma-associated antigen (MAGE)-D4 mRNA was a prognostic factor for esophageal squamous cell carcinoma (ESCC). The aim of this study was to validate the expression of MAGE-D4 in two additional patient cohorts, and to investigate its biological functions. MATERIALS AND METHODS: The role of MAGE-D4 in cell proliferation, adhesion, and migration was determined by gene knockdown experiments in the KYSE590 cell line. MAGE-D4 protein expression was analyzed in ESCC tissues by immunohistochemistry. A second validation cohort consisted of an ESCC mRNA dataset from The Cancer Genome Atlas. RESULTS: Knockdown of MAGE-D4 significantly decreased cell proliferation and migration. Expression of MAGE-D4 protein was significantly associated with disease-free survival. In the second validation cohort, high MAGE-D4 mRNA expression was associated with significantly shorter overall survival and disease-free survival. CONCLUSION: MAGE-D4 plays an important role in the malignant behavior of ESCC. MAGE-D4 was validated as a prognostic indicator in two independent ESCC patient cohorts.


Assuntos
Antígenos de Neoplasias/metabolismo , Biomarcadores Tumorais/metabolismo , Neoplasias Esofágicas/patologia , Carcinoma de Células Escamosas do Esôfago/secundário , Proteínas de Neoplasias/metabolismo , Idoso , Antígenos de Neoplasias/genética , Apoptose , Biomarcadores Tumorais/genética , Adesão Celular , Proliferação de Células , Estudos de Coortes , Neoplasias Esofágicas/metabolismo , Neoplasias Esofágicas/cirurgia , Carcinoma de Células Escamosas do Esôfago/metabolismo , Carcinoma de Células Escamosas do Esôfago/cirurgia , Feminino , Seguimentos , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Invasividade Neoplásica , Metástase Neoplásica , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/genética , Prognóstico , RNA Interferente Pequeno/genética , Taxa de Sobrevida , Células Tumorais Cultivadas
4.
Cancer Immunol Immunother ; 68(11): 1891-1899, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31628525

RESUMO

Acute myeloid leukemia (AML) is the most common acute leukemia in adults and overall survival remains poor. Chemotherapy is the standard of care for intensive induction therapy. Patients who achieve a complete remission require post-remission therapies to prevent relapse. There is no standard of care for patients with minimal residual disease (MRD), and stem cell transplantation is a salvage therapy. Considering the AML genetic heterogeneity and the leukemia immune-suppressive properties, novel cellular immune therapies to effectively harness immunological responses to prevent relapse are needed. We developed a novel modality of immune therapy consisting of monocytes reprogrammed with lentiviral vectors expressing GM-CSF, IFN-α and antigens. Preclinical studies in humanized mice showed that the reprogrammed monocytes self-differentiated into highly viable induced dendritic cells (iDCs) in vivo which migrated effectively to lymph nodes, producing remarkable effects in the de novo regeneration of T and B cell responses. For the first-in-man clinical trial, the patient's monocytes will be transduced with an integrase-defective tricistronic lentiviral vector expressing GM-CSF, IFN-α and a truncated WT1 antigen. For transplanted patients, pre-clinical development of iDCs co-expressing cytomegalovirus antigens is ongoing. To simplify the product chain for a de-centralized supply model, we are currently exploring a closed automated system for a short two-day manufacturing of iDCs. A phase I clinical trial study is in preparation for immune therapy of AML patients with MRD. The proposed cell therapy can fill an important gap in the current and foreseeable future immunotherapies of AML.


Assuntos
Antígenos de Neoplasias/metabolismo , Vetores Genéticos/administração & dosagem , Fator Estimulador de Colônias de Granulócitos e Macrófagos/metabolismo , Imunoterapia , Interferon-alfa/metabolismo , Leucemia Mieloide Aguda/terapia , Monócitos/imunologia , Células Dendríticas/imunologia , Humanos , Lentivirus/genética , Leucemia Mieloide Aguda/imunologia , Leucemia Mieloide Aguda/metabolismo , Medicina de Precisão , Transplante de Células-Tronco
5.
Anticancer Res ; 39(9): 5071-5076, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31519617

RESUMO

BACKGROUND/AIM: Approximately 20% of pleural effusions are associated with cancer; about 50% require invasive procedures to perform diagnosis. Determination of the concentration of soluble cytokeratin 19-fragments (CYFRA21-1) may help identify patients with malignant effusions. However, pathologies other than cancer can increase its concentration. The identification of these possible false positives with routine tests CRP, ADA, % polymorphonuclear cells (PN) may improve diagnostic accuracy. This study aimed to determine the diagnostic accuracy of CYFRA21-1 in the detection of malignant pleural effusions and the possible false positives. MATERIALS AND METHODS: Analysis of CYFRA21-1, adenosine deaminase (ADA), C-reactive protein (CRP), and the percentage of polymorphonuclear leukocytes (PN%) in the fluid from 643 consecutive undiagnosed pleural effusions was performed. RESULTS: CYFRA21-1 showed 38.7% sensitivity and 97.3% specificity at 175 ng/ml cut-off. Effusions not suspicious of a false-positive showed 39.0% sensitivity and 98.2% specificity, while effusions suspicious of false positive showed lower sensitivity (36.4%) and specificity (95.0%). CONCLUSION: The diagnostic accuracy of CYFRA21-1 in pleural effusions can be improved by classification according to the possibility of false positives.


Assuntos
Antígenos de Neoplasias/metabolismo , Biomarcadores Tumorais , Queratina-19/metabolismo , Derrame Pleural/diagnóstico , Derrame Pleural/metabolismo , Biópsia , Feminino , Humanos , Masculino , Derrame Pleural/etiologia , Curva ROC , Valores de Referência , Reprodutibilidade dos Testes , Sensibilidade e Especificidade
6.
Eur J Med Chem ; 181: 111586, 2019 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-31401537

RESUMO

We have synthetized a novel series of ß-hydroxy tellurides bearing the benzenesulfonamide group as potent inhibitors of carbonic anhydrase enzymes. In a one pot procedure, we discovered both the ring opening reaction of the three-membered ring and the cleavage of the sulfonamide protecting moiety at the same time. Moreover, the first X-ray co-crystallographic structure of a ß-hydroxy telluride derivative with hCA II is reported. The potent effects of these compounds against the tumor-associated hCA IX with low nanomolar constant inhibition values give the possibility to evaluate their activity in vitro using a breast cancer cell line (MDA-MB-231). Compounds 7e and 7g induced significant toxic effects against tumor cells after 48 h incubation in normoxic conditions killing over 50% of tumor cells at 3 µM, but their efficacy decreased in hypoxic conditions reaching the 50% of the tumor cell viability only at 30 µM. These unusual features make them interesting lead compounds to act as antitumor agents, not only as Carbonic Anhydrase IX inhibitors, but reasonably in different pathways, where hCA IX is not overexpressed.


Assuntos
Antineoplásicos/farmacologia , Inibidores da Anidrase Carbônica/farmacologia , Sulfonamidas/farmacologia , Telúrio/farmacologia , Antígenos de Neoplasias/química , Antígenos de Neoplasias/metabolismo , Antineoplásicos/síntese química , Antineoplásicos/química , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/metabolismo , Anidrase Carbônica II/antagonistas & inibidores , Anidrase Carbônica II/química , Anidrase Carbônica II/metabolismo , Anidrase Carbônica IX/antagonistas & inibidores , Anidrase Carbônica IX/química , Anidrase Carbônica IX/metabolismo , Inibidores da Anidrase Carbônica/síntese química , Inibidores da Anidrase Carbônica/química , Linhagem Celular Tumoral , Cristalografia por Raios X , Feminino , Humanos , Simulação de Acoplamento Molecular , Sulfonamidas/síntese química , Sulfonamidas/química , Telúrio/química
7.
Nat Biotechnol ; 37(9): 1049-1058, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31332324

RESUMO

Chimeric antigen receptor (CAR)-T-cell therapy for solid tumors is limited due to heterogeneous target antigen expression and outgrowth of tumors lacking the antigen targeted by CAR-T cells directed against single antigens. Here, we developed a bicistronic construct to drive expression of a CAR specific for EGFRvIII, a glioblastoma-specific tumor antigen, and a bispecific T-cell engager (BiTE) against EGFR, an antigen frequently overexpressed in glioblastoma but also expressed in normal tissues. CART.BiTE cells secreted EGFR-specific BiTEs that redirect CAR-T cells and recruit untransduced bystander T cells against wild-type EGFR. EGFRvIII-specific CAR-T cells were unable to completely treat tumors with heterogenous EGFRvIII expression, leading to outgrowth of EGFRvIII-negative, EGFR-positive glioblastoma. However, CART.BiTE cells eliminated heterogenous tumors in mouse models of glioblastoma. BiTE-EGFR was locally effective but was not detected systemically after intracranial delivery of CART.BiTE cells. Unlike EGFR-specific CAR-T cells, CART.BiTE cells did not result in toxicity against human skin grafts in vivo.


Assuntos
Anticorpos Biespecíficos/uso terapêutico , Antígenos de Neoplasias/imunologia , Glioblastoma/terapia , Receptores de Antígenos Quiméricos , Animais , Antígenos de Neoplasias/metabolismo , Diferenciação Celular , Receptores ErbB , Glioblastoma/imunologia , Glioblastoma/metabolismo , Humanos , Camundongos , Neoplasias Experimentais , Linfócitos T/fisiologia
8.
J Dermatol ; 46(7): 610-614, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31166031

RESUMO

Primary cicatricial alopecia (PCA) is a group of poorly understood mechanisms in which the destruction of hair follicles leads to permanent hair loss. Lichen planopilaris (LPP) is a type of lymphocytic PCA and it has been known for epidermal Langerhans cells (LC) to disappear in the scar of LPP. We also found that epidermal LC also disappeared in the scar of folliculitis decalvans (FD), a type of neutrophilic PCA. Of note was that epidermal LC did not disappear in the scar of discoid lupus erythematosus, another type of lymphocytic PCA, suggesting that LC disappearance in the scar was not always a common feature of PCA. We found that the expression of integrin (ITG)-αvß6 in scar epidermis was significantly diminished in LPP and FD, but not in other PCA and disorders accompanied with scar formation. We also found that exogenous interleukin-1ß and α-interferon downregulated ITG-αvß6 expression in normal human epidermal keratinocytes. These data suggest that downregulation of ITG-αvß6 may be one of the causes of LC disappearance in the scar of LPP and FD.


Assuntos
Alopecia/patologia , Antígenos de Neoplasias/metabolismo , Cicatriz/patologia , Foliculite/patologia , Integrinas/metabolismo , Células de Langerhans/imunologia , Líquen Plano/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Alopecia/imunologia , Antígenos de Neoplasias/imunologia , Cicatriz/imunologia , Regulação para Baixo , Células Epidérmicas/imunologia , Epiderme/imunologia , Epiderme/patologia , Feminino , Foliculite/imunologia , Folículo Piloso/imunologia , Folículo Piloso/patologia , Humanos , Integrinas/imunologia , Queratinócitos , Líquen Plano/imunologia , Masculino , Pessoa de Meia-Idade
9.
Cancer Immunol Immunother ; 68(8): 1245-1261, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31222486

RESUMO

The efficacy of cancer immunotherapy, including treatment with immune-checkpoint inhibitors, often is limited by ineffective presentation of antigenic peptides that elicit T-cell-mediated anti-tumor cytotoxic responses. Manipulation of antigen presentation pathways is an emerging approach for enhancing the immunogenicity of tumors in immunotherapy settings. ER aminopeptidase 1 (ERAP1) is an intracellular enzyme that trims peptides as part of the system that generates peptides for binding to MHC class I molecules (MHC-I). We hypothesized that pharmacological inhibition of ERAP1 in cells could regulate the cellular immunopeptidome. To test this hypothesis, we treated A375 melanoma cells with a recently developed potent ERAP1 inhibitor and analyzed the presented MHC-I peptide repertoire by isolating MHC-I, eluting bound peptides, and identifying them using capillary chromatography and tandem mass spectrometry (LC-MS/MS). Although the inhibitor did not reduce cell-surface MHC-I expression, it induced qualitative and quantitative changes in the presented peptidomes. Specifically, inhibitor treatment altered presentation of about half of the total 3204 identified peptides, including about one third of the peptides predicted to bind tightly to MHC-I. Inhibitor treatment altered the length distribution of eluted peptides without change in the basic binding motifs. Surprisingly, inhibitor treatment enhanced the average predicted MHC-I binding affinity, by reducing presentation of sub-optimal long peptides and increasing presentation of many high-affinity 9-12mers, suggesting that baseline ERAP1 activity in this cell line is destructive for many potential epitopes. Our results suggest that chemical inhibition of ERAP1 may be a viable approach for manipulating the immunopeptidome of cancer.


Assuntos
Aminopeptidases/metabolismo , Antígenos de Neoplasias/metabolismo , Antineoplásicos/farmacologia , Vacinas Anticâncer/imunologia , Epitopos de Linfócito T/metabolismo , Imunoterapia/métodos , Melanoma/tratamento farmacológico , Antígenos de Histocompatibilidade Menor/metabolismo , Peptídeos/metabolismo , Inibidores de Proteases/farmacologia , Linfócitos T Citotóxicos/imunologia , Aminopeptidases/antagonistas & inibidores , Apresentação do Antígeno , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/imunologia , Linhagem Celular Tumoral , Citotoxicidade Imunológica , Epitopos de Linfócito T/genética , Epitopos de Linfócito T/imunologia , Antígenos HLA/metabolismo , Antígenos de Histocompatibilidade Classe I/metabolismo , Humanos , Imunogenicidade da Vacina , Ativação Linfocitária , Terapia de Alvo Molecular , Peptídeos/genética , Peptídeos/imunologia , Ligação Proteica
10.
Biochim Biophys Acta Proteins Proteom ; 1867(9): 854-865, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31247344

RESUMO

Falcipain-2(FP2), a cysteine protease from Plasmodium falciparum, cleaves host erythrocyte hemoglobin and specific membrane skeleton components during the parasite life cycle. Therefore its inhibition has been considered as an attractive approach to combat the disease. SerpinB3 (SPB3) belongs to the ovalbumin-serpin family and is a potent cross-class inhibitor of cysteine cathepsins L, K, S and papain. This study explored the possibility of inhibition of FP2 by SPB3. It turned out that general proteolytic activities as well as specific hemoglobinolytic activity of FP2 have been inhibited by SPB3. Furthermore, studies have been designed to investigate and characterize the mechanism of inhibition in comparison with proteases Cathepsin L (CTSL) and papain. The Ki value of inhibition for FP2, measured against its specific substrate (VLK-pNA), is 338.11 nM and stoichiometry (I/E ratio) of inhibition is 1. These values are comparable to CTSL and papain. Analytical gel filtration profile and CD spectroscopy data confirm FP2-SPB3 complex formation. Our studies revealed that interaction of SPB3 with FP2 is non-covalent type like that of CTSL and papain but unlike other serine protease-inhibiting serpins. An in-silico docking and simulation study have been performed with FP2 as well as CTSL and results suggest different binding mode for FP2 and CTSL, though both the complexes are stable with significant contribution from electrostatic energy of interaction. We further showed a disease state mutant SPB3-Gly351Ala performed better anti-protease activity against FP2. This study, for the first time, has shown a serpin family inhibitor from human could efficiently inhibit activity of FP2.


Assuntos
Antígenos de Neoplasias/química , Cisteína Endopeptidases/química , Simulação de Acoplamento Molecular , Plasmodium falciparum/enzimologia , Serpinas/química , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/metabolismo , Catepsina L/antagonistas & inibidores , Catepsina L/química , Catepsina L/genética , Catepsina L/metabolismo , Cisteína Endopeptidases/genética , Cisteína Endopeptidases/metabolismo , Humanos , Papaína/antagonistas & inibidores , Papaína/química , Papaína/genética , Papaína/metabolismo , Plasmodium falciparum/genética , Serpinas/genética , Serpinas/metabolismo
11.
Cancer Immunol Immunother ; 68(7): 1039-1058, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31165204

RESUMO

The emergence of immunotherapy has revolutionized medical oncology with unprecedented advances in cancer treatment over the past two decades. However, a major obstacle in cancer immunotherapy is identifying appropriate tumor-specific antigens to make targeted therapy achievable with fewer normal cells being impaired. The similarity between placentation and tumor development and growth has inspired many investigators to discover antigens for effective immunotherapy of cancers. Placenta-specific 1 (PLAC1) is one of the recently discovered placental antigens with limited normal tissue expression and fundamental roles in placental function and development. There is a growing body of evidence showing that PLAC1 is frequently activated in a wide variety of cancer types and promotes cancer progression. Based on the restricted expression of PLAC1 in testis, placenta and a wide variety of cancers, we have designated this molecule with new terminology, cancer-testis-placenta (CTP) antigen, a feature that PLAC1 shares with many other cancer testis antigens. Recent reports from our lab provide compelling evidence on the preferential expression of PLAC1 in prostate cancer and its potential utility in prostate cancer immunotherapy. PLAC1 may be regarded as a potential CTP antigen for targeted cancer immunotherapy based on the available data on its promoting function in cancer development and also its expression in cancers of different histological origin. In this review, we will summarize current data on PLAC1 with emphasis on its association with cancer development and immunotherapy.


Assuntos
Antígenos de Neoplasias/imunologia , Antineoplásicos Imunológicos/uso terapêutico , Biomarcadores Tumorais/antagonistas & inibidores , Neoplasias/terapia , Proteínas da Gravidez/antagonistas & inibidores , Antígenos de Neoplasias/metabolismo , Antineoplásicos Imunológicos/farmacologia , Biomarcadores Tumorais/imunologia , Biomarcadores Tumorais/metabolismo , Progressão da Doença , Feminino , Humanos , Imunoterapia/métodos , Masculino , Terapia de Alvo Molecular/métodos , Neoplasias/imunologia , Neoplasias/patologia , Placenta/patologia , Gravidez , Proteínas da Gravidez/imunologia , Proteínas da Gravidez/metabolismo , Testículo/patologia
12.
Cancer Immunol Immunother ; 68(7): 1195-1209, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31177329

RESUMO

The anti-tumor efficacy of TCR-engineered T cells in vivo depends largely on less-differentiated subsets such as T cells with naïve-like T cell (TN) phenotypes with greater expansion and long-term persistence. To increase these subsets, we compared the generation of New York esophageal squamous cell carcinoma-1 (NY-ESO-1)-specific T cells under supplementation with either IL-2 or IL-7/IL-15. PBMCs were transduced with MS3II-NY-ESO-1-siTCR retroviral vector. T cell generation was adapted from a CD19-specific CART cell production protocol. Comparable results in viability, expansion and transduction efficiency of T cells under stimulation with either IL-2 or IL-7/IL-15 were observed. IL-7/IL-15 led to an increase of CD4+ T cells and a decrease of CD8+ T cells, enriched the amount of TN among CD4+ T cells but not among CD8+ T cells. In a 51Cr release assay, similar specific lysis of NY-ESO-1-positive SW982 sarcoma cells was achieved. However, intracellular cytokine staining revealed a significantly increased production of IFN-γ and TNF-α in T cells generated by IL-2 stimulation. To validate these unexpected findings, NY-ESO-1-specific T cell production was evaluated in another protocol originally established for TCR-engineered T cells. IL-7/IL-15 increased the proportion of TN. However, the absolute number of TN did not increase due to a significantly slower expansion of T cells with IL-7/IL-15. In conclusion, IL-7/IL-15 does not seem to be superior to IL-2 for the generation of NY-ESO-1-specific T cells. This is in sharp contrast to the observations in CD19-specific CART cells. Changes of cytokine cocktails should be carefully evaluated for individual vector systems.


Assuntos
Antígenos de Neoplasias/metabolismo , Engenharia Celular/métodos , Imunoterapia Adotiva/métodos , Proteínas de Membrana/metabolismo , Neoplasias/terapia , Receptores de Antígenos Quiméricos/imunologia , Antígenos CD19/metabolismo , Antígenos de Neoplasias/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD4-Positivos/transplante , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/transplante , Técnicas de Cultura de Células/métodos , Linhagem Celular Tumoral , Meios de Cultura , Humanos , Interleucina-15/imunologia , Interleucina-2/imunologia , Interleucina-7/imunologia , Proteínas de Membrana/imunologia , Neoplasias/imunologia , Receptores de Antígenos Quiméricos/genética
13.
Nat Commun ; 10(1): 2484, 2019 06 06.
Artigo em Inglês | MEDLINE | ID: mdl-31171773

RESUMO

Tumor-specific antibody drugs can serve as cancer therapy with minimal side effects. A humanized antibody, PRL3-zumab, specifically binds to an intracellular oncogenic phosphatase PRL3, which is frequently expressed in several cancers. Here we show that PRL3-zumab specifically inhibits PRL3+ cancer cells in vivo, but not in vitro. PRL3 antigens are detected on the cell surface and outer exosomal membranes, implying an 'inside-out' externalization of PRL3. PRL3-zumab binds to surface PRL3 in a manner consistent with that in classical antibody-dependent cell-mediated cytotoxicity or antibody-dependent cellular phagocytosis tumor elimination pathways, as PRL3-zumab requires an intact Fc region and host FcγII/III receptor engagement to recruit B cells, NK cells and macrophages to PRL3+ tumor microenvironments. PRL3 is overexpressed in 80.6% of 151 fresh-frozen tumor samples across 11 common cancers examined, but not in patient-matched normal tissues, thereby implicating PRL3 as a tumor-associated antigen. Targeting externalized PRL3 antigens with PRL3-zumab may represent a feasible approach for anti-tumor immunotherapy.


Assuntos
Citotoxicidade Celular Dependente de Anticorpos/efeitos dos fármacos , Antineoplásicos Imunológicos/farmacologia , Carcinoma Hepatocelular/metabolismo , Citofagocitose/efeitos dos fármacos , Hepatócitos/efeitos dos fármacos , Neoplasias Hepáticas/metabolismo , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas Tirosina Fosfatases/antagonistas & inibidores , Microambiente Tumoral/efeitos dos fármacos , Animais , Anticorpos Monoclonais Humanizados , Anticorpos Monoclonais Murinos , Antígenos de Neoplasias/metabolismo , Linfócitos B , Linhagem Celular Tumoral , Células Hep G2 , Hepatócitos/metabolismo , Humanos , Imunoterapia , Células Matadoras Naturais , Macrófagos , Camundongos , Terapia de Alvo Molecular , Proteínas de Neoplasias/metabolismo , Transplante de Neoplasias , Neoplasias/metabolismo , Proteínas Oncogênicas/metabolismo , Proteínas Tirosina Fosfatases/metabolismo , Receptores de IgG , Microambiente Tumoral/imunologia , Ensaios Antitumorais Modelo de Xenoenxerto
14.
Gastroenterology ; 157(3): 823-837, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31078621

RESUMO

BACKGROUND & AIMS: Most pancreatic ductal adenocarcinomas (PDACs) express an activated form of KRAS, become hypoxic and dysplastic, and are refractory to chemo and radiation therapies. To survive in the hypoxic environment, PDAC cells upregulate enzymes and transporters involved in pH regulation, including the extracellular facing carbonic anhydrase 9 (CA9). We evaluated the effect of blocking CA9, in combination with administration of gemcitabine, in mouse models of pancreatic cancer. METHODS: We knocked down expression of KRAS in human (PK-8 and PK-1) PDAC cells with small hairpin RNAs. Human and mouse (KrasG12D/Pdx1-Cre/Tp53/RosaYFP) PDAC cells were incubated with inhibitors of MEK (trametinib) or extracellular signal-regulated kinase (ERK), and some cells were cultured under hypoxic conditions. We measured levels and stability of the hypoxia-inducible factor 1 subunit alpha (HIF1A), endothelial PAS domain 1 protein (EPAS1, also called HIF2A), CA9, solute carrier family 16 member 4 (SLC16A4, also called MCT4), and SLC2A1 (also called GLUT1) by immunoblot analyses. We analyzed intracellular pH (pHi) and extracellular metabolic flux. We knocked down expression of CA9 in PDAC cells, or inhibited CA9 with SLC-0111, incubated them with gemcitabine, and assessed pHi, metabolic flux, and cytotoxicity under normoxic and hypoxic conditions. Cells were also injected into either immune-compromised or immune-competent mice and growth of xenograft tumors was assessed. Tumor fragments derived from patients with PDAC were surgically ligated to the pancreas of mice and the growth of tumors was assessed. We performed tissue microarray analyses of 205 human PDAC samples to measure levels of CA9 and associated expression of genes that regulate hypoxia with outcomes of patients using the Cancer Genome Atlas database. RESULTS: Under hypoxic conditions, PDAC cells had increased levels of HIF1A and HIF2A, upregulated expression of CA9, and activated glycolysis. Knockdown of KRAS in PDAC cells, or incubation with trametinib, reduced the posttranscriptional stabilization of HIF1A and HIF2A, upregulation of CA9, pHi, and glycolysis in response to hypoxia. CA9 was expressed by 66% of PDAC samples analyzed; high expression of genes associated with metabolic adaptation to hypoxia, including CA9, correlated with significantly reduced survival times of patients. Knockdown or pharmacologic inhibition of CA9 in PDAC cells significantly reduced pHi in cells under hypoxic conditions, decreased gemcitabine-induced glycolysis, and increased their sensitivity to gemcitabine. PDAC cells with knockdown of CA9 formed smaller xenograft tumors in mice, and injection of gemcitabine inhibited tumor growth and significantly increased survival times of mice. In mice with xenograft tumors grown from human PDAC cells, oral administration of SLC-0111 and injection of gemcitabine increased intratumor acidosis and increased cell death. These tumors, and tumors grown from PDAC patient-derived tumor fragments, grew more slowly than xenograft tumors in mice given control agents, resulting in longer survival times. In KrasG12D/Pdx1-Cre/Tp53/RosaYFP genetically modified mice, oral administration of SLC-0111 and injection of gemcitabine reduced numbers of B cells in tumors. CONCLUSIONS: In response to hypoxia, PDAC cells that express activated KRAS increase expression of CA9, via stabilization of HIF1A and HIF2A, to regulate pH and glycolysis. Disruption of this pathway slows growth of PDAC xenograft tumors in mice and might be developed for treatment of pancreatic cancer.


Assuntos
Antígenos de Neoplasias/metabolismo , Anidrase Carbônica IX/metabolismo , Carcinoma Ductal Pancreático/enzimologia , Neoplasias Pancreáticas/enzimologia , Proteínas Proto-Oncogênicas p21(ras)/genética , Microambiente Tumoral , Animais , Antígenos de Neoplasias/genética , Antimetabólitos Antineoplásicos/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Anidrase Carbônica IX/antagonistas & inibidores , Anidrase Carbônica IX/genética , Inibidores da Anidrase Carbônica/farmacologia , Carcinoma Ductal Pancreático/tratamento farmacológico , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/patologia , Hipóxia Celular , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Desoxicitidina/análogos & derivados , Desoxicitidina/farmacologia , Feminino , Regulação Neoplásica da Expressão Gênica , Predisposição Genética para Doença , Glicólise/efeitos dos fármacos , Humanos , Concentração de Íons de Hidrogênio , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Camundongos SCID , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Fenótipo , Compostos de Fenilureia/farmacologia , Transdução de Sinais , Sulfonamidas/farmacologia , Carga Tumoral/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
15.
Med Sci Monit ; 25: 3495-3502, 2019 05 11.
Artigo em Inglês | MEDLINE | ID: mdl-31077581

RESUMO

BACKGROUND The use of adoptive T cell therapy has proven to be effective in some advanced malignancies. This study aimed to investigate the effects of lymphocyte-activation gene 3 (LAG-3) immune checkpoint receptor in the enrichment of tumor antigen-specific CD8⁺ T lymphocytes derived from peripheral blood mononuclear cells (PBMCs) in patients with colorectal cancer. MATERIAL AND METHODS Peripheral blood samples were obtained from 20 patients with colorectal cancer and apheresis was performed with enrichment and cell sorting to obtain CD8⁺LAG-3⁺ T cells, which were expanded using high-dose interleukin-2 (IL-2). T cell subsets were detected using fluorescence-activated cell sorting (FACS), and T cell receptor (TCR) sequencing was used to determine specific clone types. Interferon-γ (IFN-γ) enzyme-linked immunospot (ELISpot) and cell counting kit-8 (CCK-8) assays were used to measure cell avidity and cytotoxicity. RESULTS The cultured cells increased in number over time and had the greatest proliferative activity at 15 days, at which time the percentage of CD3⁺, CD3⁺CD8⁺, and CD8⁺CD28⁺ reached maximal levels. High purity CD8⁺LAG-3⁺ T cells were isolated by FACS and at 15 days. TCR sequencing showed that CD8⁺LAG-3⁺ T cells were oligoclonal, ELISpot identified increased production of tumor-specific IFN-γ, and the CCK-8 assay showed increased cytotoxicity when compared with pre-cultured CD8⁺LAG-3⁻ T cells. CONCLUSIONS In patients with colorectal cancer, CD8⁺LAG-3⁺ T cells showed more specific anti-tumor activity following cell culture in vitro, which supported the potential role for the LAG-3 immune checkpoint receptor in enriching tumor-specific T cells in patients with cancer.


Assuntos
Antígenos CD/genética , Linfócitos T CD8-Positivos/imunologia , Neoplasias Colorretais/genética , Adulto , Idoso , Antígenos CD/metabolismo , Antígenos de Neoplasias/metabolismo , Antígenos CD28/imunologia , Complexo CD3/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Linhagem Celular Tumoral , China , Neoplasias Colorretais/metabolismo , Feminino , Citometria de Fluxo , Humanos , Interferon gama/metabolismo , Leucócitos Mononucleares/metabolismo , Ativação Linfocitária/imunologia , Masculino , Pessoa de Meia-Idade , Receptores de Antígenos de Linfócitos T , Receptores Imunológicos/metabolismo
16.
J Nanobiotechnology ; 17(1): 59, 2019 May 04.
Artigo em Inglês | MEDLINE | ID: mdl-31054582

RESUMO

BACKGROUND: Technology enabling the separation of rare circulating tumor cells (CTCs) provides the potential to enhance our knowledge of cancer metastasis and improve the care of cancer patients. Modern detection approaches commonly depend on tumor antigens or the physical size of CTCs. However, few studies report the detection of CTCs by the electrical differences between cancer cells and normal cells. RESULTS: In this study, we report a procedure for capturing CTCs from blood samples using electrically charged superparamagnetic nanoparticles (NPs). We found that only positively charged NPs attached to cancer cells, while negatively charged NPs did not. The capture method with positively charged NPs offered a sensitivity of down to 4 CTCs in 1 mL blood samples and achieved a superior capture yield (> 70%) for a high number of CTCs in blood samples (103-106 cells/mL). Following an in vitro evaluation, S180-bearing mice were employed as an in vivo model to assess the specificity and sensitivity of the capture procedure. The number of CTCs in blood from tumor-bearing mice was significantly higher than that in blood from healthy controls (on average, 75.8 ± 16.4 vs. zero CTCs/100 µL of blood, p < 0.0001), suggesting the high sensitivity and specificity of our method. CONCLUSIONS: Positively charged NPs combined with an in vivo tumor model demonstrated that CTCs can be distinguished and isolated from other blood cells based on their electrical properties.


Assuntos
Neoplasias da Mama/diagnóstico , Óxido Ferroso-Férrico/química , Nanopartículas de Magnetita/química , Células Neoplásicas Circulantes/metabolismo , Adulto , Idoso , Animais , Antígenos de Neoplasias/metabolismo , Técnicas Biossensoriais/métodos , Sangue , Contagem de Células/métodos , Linhagem Celular Tumoral , Separação Celular/métodos , Feminino , Humanos , Magnetismo , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Modelos Animais , Tamanho da Partícula , Sensibilidade e Especificidade , Propriedades de Superfície
17.
BMC Cancer ; 19(1): 510, 2019 May 29.
Artigo em Inglês | MEDLINE | ID: mdl-31142270

RESUMO

BACKGROUND: Hypoxia is an important factor that contributes to tumour aggressiveness and correlates with poor prognosis and resistance to conventional therapy. Therefore, identifying hypoxic environments within tumours is extremely useful for understanding cancer biology and developing novel therapeutic strategies. Several studies have suggested that carbonic anhydrase 9 (CA9) is a reliable biomarker of hypoxia and a potential therapeutic target, while pimonidazole has been identified as an exogenous hypoxia marker. However, other studies have suggested that CA9 expression is not directly induced by hypoxia and it is not expressed in all types of tumours. Thus, in this study, we focused on endoplasmic reticulum disulphide oxidase 1α (ERO1α), a protein that localises in the endoplasmic reticulum and is involved in the formation of disulphide bonds in proteins, to determine whether it could serve as a potential tumour-hypoxia biomarker. METHODS: Using quantitative real-time polymerase chain reaction, we analysed the mRNA expression of ERO1α and CA9 in different normal and cancer cell lines. We also determined the protein expression levels of ERO1α and CA9 in these cell lines by western blotting. We then investigated the hypoxia-inducible ERO1α and CA9 expression and localisation in HCT116 and HeLa cells, which express low (CA9-low) and high (CA9-high) levels of CA9, respectively. A comparative analysis was performed using pimonidazole, an exogenous hypoxic marker, as a positive control. The expression and localisation of ERO1α and CA9 in tumour spheres during hypoxia were analysed by a tumour sphere formation assay. Finally, we used a mouse model to investigate the localisation of ERO1α and CA9 in tumour xenografts using several cell lines. RESULTS: We found that ERO1α expression increased under chronic hypoxia. Our results show that ERO1α was hypoxia-induced in all the tested cancer cell lines. Furthermore, in the comparative analysis using CA9 and pimonidazole, ERO1α had a similar localisation to pimonidazole in both CA9-low and CA9-high cell lines. CONCLUSION: ERO1α can serve as a novel endogenous chronic hypoxia marker that is more reliable than CA9 and can be used as a diagnostic biomarker and therapeutic target for cancer.


Assuntos
Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Neoplasias/metabolismo , Oxirredutases/genética , Oxirredutases/metabolismo , Animais , Antígenos de Neoplasias/genética , Antígenos de Neoplasias/metabolismo , Anidrase Carbônica IX/genética , Anidrase Carbônica IX/metabolismo , Hipóxia Celular , Linhagem Celular Tumoral , Células HCT116 , Células HT29 , Células HeLa , Humanos , Camundongos , Transplante de Neoplasias , Neoplasias/genética , Nitroimidazóis/metabolismo
18.
Biomed Res Int ; 2019: 2807123, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31080815

RESUMO

Cervical cancer is a life-threatening complication, appearing as the uncontrolled growth of abnormal cells in the lining of the cervix. Every year, increasing numbers of cervical cancer cases are reported worldwide. Different identification strategies were proposed to detect cervical cancer at the earlier stages using various biomarkers. Squamous cell carcinoma antigen (SCC-Ag) is one of the potential biomarkers for this diagnosis. Nanomaterial-based detection systems were shown to be efficient with different clinical biomarkers. In this study, we have demonstrated strontium oxide-modified interdigitated electrode (IDE) fabrication by the sol-gel method and characterized by scanning electron microscopy and high-power microscopy. Analysis of the bare devices indicated the reproducibility with the fabrication, and further pH scouting on the device revealed that the reliability of the working pH ranges from 3 to 9. The sensing surface was tested to detect SCC-Ag against its specific antibody; the detection limit was found to be 10 pM, and the sensitivity was in the range between 1 and 10 pM as calculated by 3σ. The specificity experiment was carried out using major proteins from human serum, such as albumin and globulin. SCC-Ag was shown to be selectively detected on the strontium oxide-modified IDE surface.


Assuntos
Antígenos de Neoplasias/administração & dosagem , Biomarcadores Tumorais/metabolismo , Carcinoma de Células Escamosas/diagnóstico , Serpinas/administração & dosagem , Estrôncio/administração & dosagem , Neoplasias do Colo do Útero/diagnóstico , Antígenos de Neoplasias/metabolismo , Carcinoma de Células Escamosas/metabolismo , Colo do Útero/metabolismo , Eletrodos , Feminino , Humanos , Recidiva Local de Neoplasia/diagnóstico , Recidiva Local de Neoplasia/metabolismo , Óxidos , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Serpinas/metabolismo , Neoplasias do Colo do Útero/metabolismo
19.
Comput Biol Chem ; 80: 234-243, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31009872

RESUMO

Nowadays, different approaches have been pursued with the intent to develop sulfonamide-like carbonic anhydrase inhibitors that possess better selectivity profiles toward the different human isoforms of the enzyme. Here, we used conventional 3D-QSAR methods, including comparative molecular field analysis (CoMFA), comparative molecular similarity indices analysis (CoMSIA), and Topomer CoMFA, to construct three-dimensional quantitative structure-activity relationship (3D-QSAR) models for benzenesulfonamide derivatives as human carbonic anhydrase (hCA) II/IX inhibitors. The theoretical models had good reliability (R2>0.75) and predictability (Q2>0.55), and the contour maps could graphically present the contributions of the force fields for activity and identify the structural divergence between human carbonic anhydrase II inhibitors and human carbonic anhydrase IX inhibitors. Consequently, we explored the selectivity of inhibitor for human carbonic anhydrase II and IX through molecular docking, and the difference of activity coincides with the potential binding mode well. According to the results of the predicted values and the molecule docking, we found that the inhibitors published in the literature had stronger inhibition on the hCA IX; based on the theoretical models, we designed seven new compounds with good potential activity and reasonably good ADMET profile, which could selectively inhibit hCA IX. Molecular Dynamics Simulation showed that newly-designed compound D7 had good selectivity on hCA IX. The findings from 3D-QSAR and docking studies maybe helpful in the rational drug design of isoform-selective inhibitors.


Assuntos
Antígenos de Neoplasias/química , Anidrase Carbônica II/química , Anidrase Carbônica IX/química , Inibidores da Anidrase Carbônica/química , Sulfonamidas/química , Antígenos de Neoplasias/metabolismo , Anidrase Carbônica II/metabolismo , Anidrase Carbônica IX/metabolismo , Inibidores da Anidrase Carbônica/metabolismo , Inibidores da Anidrase Carbônica/farmacocinética , Conjuntos de Dados como Assunto , Desenho de Drogas , Humanos , Análise dos Mínimos Quadrados , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Estrutura Molecular , Ligação Proteica , Relação Quantitativa Estrutura-Atividade , Sulfonamidas/metabolismo , Sulfonamidas/farmacocinética
20.
Nagoya J Med Sci ; 81(1): 1-18, 2019 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30962651

RESUMO

CD4+ regulatory T cells (Tregs) expressing the transcription factor forkhead box P3 (FoxP3) play an important role in self-tolerance and immune homeostasis. Tregs have evolved to protect the host from aberrant immune responses against self-components and collateral damages occurring in the process of defense against invading pathogens by softening immune responses. However, they turned to be a scourge in malignant tumors by not only allowing and promoting tumor growth but also suppressing effective antitumor actions, both inherent (host's immune surveillance) and extrinsic (anticancer therapy). An increase in the number of Tregs infiltrating into tumor sites and a concomitant decrease in the number of CD8+ cytotoxic T lymphocytes are associated with a poor prognosis for various types of cancers, marking Tregs as notorious meddlers with an effective antitumor response. Various cancer immunotherapy approaches are often dampened by meddling Tregs, making them one of the major targets in the treatment of cancer. The recent success of immune checkpoint inhibitors (ICIs) that target immune checkpoint molecules expressed by Tregs or effector T cells implies, that "meddling with meddlers" represents an effective strategy in cancer immunotherapy. However, clinical responses to ICIs are effective and durable only in some patients with cancer, whereas more than half of them do not show significant clinical improvement. This implies that a therapeutic approach based on the use of a single ICI, or targeting Tregs alone, is insufficient, highlighting the need for combinatorial approaches. With regard to antitumor immune stimulation, several approaches, such as vaccination with peptides (or the corresponding DNA) to stimulate antigen-presenting CD8+ T cells with tumor-specific neoantigens, cancer/testis antigens, or cancer stem cell antigens, that eventually boost effective cytotoxic antitumor responses are being tested. This review describes the immunosuppressive physiology of Tregs and their meddling with the host's antitumor immunity; current and prospective approaches to curb Tregs; and approaches to augment antitumor immunity.


Assuntos
Linfócitos T Reguladores/metabolismo , Animais , Antígenos de Neoplasias/imunologia , Antígenos de Neoplasias/metabolismo , Fatores de Transcrição Forkhead/metabolismo , Receptor Celular 2 do Vírus da Hepatite A/metabolismo , Humanos , Tolerância Imunológica/imunologia , Tolerância Imunológica/fisiologia , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Modelos Biológicos , Receptor A2A de Adenosina/metabolismo , Receptores CCR4/metabolismo , Receptores de Interleucina-2/metabolismo , Linfócitos T Reguladores/imunologia
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