RESUMEN

RATIONALE OF THE TRIAL: Although the use of engineered T cells in cancer immunotherapy has greatly advanced the treatment of hematological malignancies, reaching meaningful clinical responses in the treatment of solid tumors is still challenging. We investigated the safety and tolerability of IMA202 in a first-in-human, dose escalation basket trial in human leucocyte antigen A*02:01 positive patients with melanoma-associated antigen A1 (MAGEA1)-positive advanced solid tumors. TRIAL DESIGN: The 2+2 trial design was an algorithmic design based on a maximally acceptable dose-limiting toxicity (DLT) rate of 25% and the sample size was driven by the algorithmic design with a maximum of 16 patients. IMA202 consists of autologous genetically modified cytotoxic CD8+ T cells expressing a T cell receptor (TCR), which is specific for a nine amino acid peptide derived from MAGEA1. Eligible patients underwent leukapheresis, T cells were isolated, transduced with lentiviral vector carrying MAGEA1-specific TCR and following lymphodepletion (fludarabine/cyclophosphamide), infused with a median of 1.4×109 specific T cells (range, 0.086×109-2.57×109) followed by interleukin 2. SAFETY OF IMA202: No DLT was observed. The most common grade 3-4 adverse events were cytopenias, that is, neutropenia (81.3%), lymphopenia (75.0%), anemia (50.0%), thrombocytopenia (50.0%) and leukopenia (25.0%). 13 patients experienced cytokine release syndrome, including one grade 3 event. Immune effector cell-associated neurotoxicity syndrome was observed in two patients and was grade 1 in both. EFFICACY OF IMA202: Of the 16 patients dosed, 11 (68.8%) patients had stable disease (SD) as their best overall response (Response Evaluation Criteria in Solid Tumors V.1.1). Five patients had initial tumor shrinkage in target lesions and one patient with SD experienced continued shrinkage in target lesions for 3 months in total but had to be classified as progressive disease due to progressive non-target lesions. IMA202 T cells were persistent in peripheral blood for several weeks to months and were also detectable in tumor tissue. Peak persistence was higher in patients who received higher doses. CONCLUSION: In conclusion, IMA202 had a manageable safety profile, and it was associated with biological and potential clinical activity of MAGEA1-targeting genetically engineered TCR-T cells in a poor prognosis, multi-indication solid tumor cohort. TRIAL REGISTRATION NUMBERS: NCT04639245, NCT05430555.

Asunto(s)

Antígenos de Neoplasias , Inmunoterapia Adoptiva , Neoplasias , Humanos , Femenino , Masculino , Antígenos de Neoplasias/inmunología , Persona de Mediana Edad , Anciano , Neoplasias/terapia , Neoplasias/inmunología , Adulto , Inmunoterapia Adoptiva/métodos , Inmunoterapia Adoptiva/efectos adversos , Receptores de Antígenos de Linfocitos T/inmunología , Receptores de Antígenos de Linfocitos T/genética , Proteínas de Neoplasias/inmunología , Proteínas de Neoplasias/genética

RESUMEN

IMA101 is an actively personalized, multi-targeted adoptive cell therapy (ACT), whereby autologous T cells are directed against multiple novel defined peptide-HLA (pHLA) cancer targets. HLA-A*02:01-positive patients with relapsed/refractory solid tumors expressing ≥1 of 8 predefined targets underwent leukapheresis. Endogenous T cells specific for up to 4 targets were primed and expanded in vitro. Patients received lymphodepletion (fludarabine, cyclophosphamide), followed by T-cell infusion and low-dose IL2 (Cohort 1). Patients in Cohort 2 received atezolizumab for up to 1 year (NCT02876510). Overall, 214 patients were screened, 15 received lymphodepletion (13 women, 2 men; median age, 44 years), and 14 were treated with T-cell products. IMA101 treatment was feasible and well tolerated. The most common adverse events were cytokine release syndrome (Grade 1, n = 6; Grade 2, n = 4) and expected cytopenias. No patient died during the first 100 days after T-cell therapy. No neurotoxicity was observed. No objective responses were noted. Prolonged disease stabilization was noted in three patients lasting for 13.7, 12.9, and 7.3 months. High frequencies of target-specific T cells (up to 78.7% of CD8+ cells) were detected in the blood of treated patients, persisted for >1 year, and were detectable in posttreatment tumor tissue. Individual T-cell receptors (TCR) contained in T-cell products exhibited broad variation in TCR avidity, with the majority being low avidity. High-avidity TCRs were identified in some patients' products. This study demonstrates the feasibility and tolerability of an actively personalized ACT directed to multiple defined pHLA cancer targets. Results warrant further evaluation of multi-target ACT approaches using potent high-avidity TCRs. See related Spotlight by Uslu and June, p. 865.

Asunto(s)

Inmunoterapia Adoptiva , Neoplasias , Adulto , Femenino , Humanos , Masculino , Linfocitos T CD8-positivos , Estudios de Factibilidad , Inmunoterapia Adoptiva/efectos adversos , Inmunoterapia Adoptiva/métodos , Neoplasias/terapia , Neoplasias/etiología , Receptores de Antígenos de Linfocitos T/genética

RESUMEN

BACKGROUND: Despite advances in care, the 5 year overall survival for patients with relapsed and or metastatic sarcoma remains as low as < 35%. Currently, there are no biomarkers available to assess disease status in patients with sarcomas and as such, disease surveillance remains reliant on serial imaging which increases the risk of secondary malignancies and heightens patient anxiety. METHODS: Here, for the first time reported in the literature, we have enumerated the cell surface vimentin (CSV+) CTCs in the blood of 92 sarcoma pediatric and adolescent and young adult (AYA) patients as a possible marker of disease. RESULTS: We constructed a ROC with an AUC of 0.831 resulting in a sensitivity of 85.3% and a specificity of 75%. Additionally, patients who were deemed to be CSV+ CTC positive were found to have a worse overall survival compared to those who were CSV+ CTC negative. We additionally found the use of available molecular testing increased the accuracy of our diagnostic and prognostic tests. CONCLUSIONS: Our findings indicate that CSV+ CTCs have prognostic value and can possibly serve as a measure of disease burden.

RESUMEN

Neuroblastoma (NB) is a deadly childhood disease that carries a 50% chance of relapse for anyone in remission and similar level of 5-year survival. We investigated the value of our proprietary approach-cell surface vimentin (CSV) positive circulating tumor cells (CTC) to monitor treatment response and predict relapse in NB patients under remission in a Phase II long-term preventative clinical trial. We longitudinally analyzed peripheral blood samples from 93 patients for 27 cycles (~25 months) and discovered that the presence of CSV+ CTCs in the first two sequential samples (baseline, cycle 4 [month 3-4]) was a significant indicator of earlier relapse. We observed strong correlation between relapse-free survival (RFS) and lack of CSV+ CTCs in first 4 cycles of therapy (95%). There was sensitivity reaching 100% in predicting RFS in patients who had neither CSV+ CTCs nor MycN amplification. Of note, the low number of CSV+ CTCs seems equivalent to low tumor load because the prevention therapy difluoromethylornithine yields faster reduction of relapse risk when none or only 1-2 CSV+ CTCs (every 6 mL) are present in the blood samples compared to >3 CSV+ CTCs. To the best of our knowledge, this is the first study that directly observes CTCs in under remission NB patients for relapse prediction and the first to gather sequential CSV+ CTC data in any study in a long-term longitudinal manner.

Asunto(s)

Recurrencia Local de Neoplasia/diagnóstico , Células Neoplásicas Circulantes/metabolismo , Neuroblastoma/diagnóstico , Vimentina/metabolismo , Biomarcadores de Tumor/metabolismo , Línea Celular Tumoral , Ensayos Clínicos Fase II como Asunto , Detección Precoz del Cáncer , Eflornitina/uso terapéutico , Transición Epitelial-Mesenquimal , Femenino , Humanos , Estudios Longitudinales , Masculino , Recurrencia Local de Neoplasia/metabolismo , Neuroblastoma/metabolismo , Sensibilidad y Especificidad , Análisis de Supervivencia

RESUMEN

Immunotherapy is revolutionizing cancer treatment and has shown success in particular for tumors with a high mutational load. These effects have been linked to neoantigens derived from patient-specific mutations. To expand efficacious immunotherapy approaches to the vast majority of tumor types and patient populations carrying only a few mutations and maybe not a single presented neoepitope, it is necessary to expand the target space to non-mutated cancer-associated antigens. Mass spectrometry enables the direct and unbiased discovery and selection of tumor-specific human leukocyte antigen (HLA) peptides that can be used to define targets for immunotherapy. Combining these targets into a warehouse allows for multi-target therapy and accelerated clinical application. For precise personalization aimed at optimally ensuring treatment efficacy and safety, it is necessary to assess the presence of the target on each individual patient's tumor. Here we show how LC-MS paired with gene expression data was used to define mRNA biomarkers currently being used as diagnostic test IMADETECT™ for patient inclusion and personalized target selection within two clinical trials (NCT02876510, NCT03247309). Thus, we present a way how to translate HLA peptide presentation into gene expression thresholds for companion diagnostics in immunotherapy considering the peptide-specific correlation to its encoding mRNA.

Asunto(s)

Antígenos de Neoplasias/metabolismo , Antígenos HLA/metabolismo , Inmunoterapia , Neoplasias/metabolismo , Fragmentos de Péptidos/metabolismo , Medicina de Precisión , Proteogenómica/métodos , Presentación de Antígeno/inmunología , Antígenos de Neoplasias/análisis , Antígenos de Neoplasias/inmunología , Toma de Decisiones , Epítopos/inmunología , Epítopos/metabolismo , Antígenos HLA/análisis , Antígenos HLA/inmunología , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Espectrometría de Masas/métodos , Neoplasias/inmunología , Fragmentos de Péptidos/análisis , Fragmentos de Péptidos/inmunología , ARN Mensajero/análisis , ARN Mensajero/genética , ARN Mensajero/inmunología

RESUMEN

Recent advances in the field of circulating tumor cells (CTC) have shown promise in this liquid biopsy-based prognosis of patient outcome. However, not all of the circulating cells are tumor cells, as evidenced by a lack of tumor-specific markers. The current FDA standard for capturing CTCs (CellSearch) relies on an epithelial marker and cells captured via CellSearch cannot be considered to have undergone EMT. Therefore, it is difficult to ascertain the presence and relevance of any mesenchymal or EMT-like CTCs. To address this gap in technology, we recently discovered the utility of cell-surface vimentin (CSV) as a marker for detecting mesenchymal CTCs from sarcoma, breast, and colon cancer. Here we studied peripheral blood samples of 48 prostate cancer (PCA) patients including hormone sensitive and castration resistant sub-groups. Blood samples were analyzed for three different properties including our own CSV-based CTC enumeration (using 84-1 mAb against CSV), CellSearch-based epithelial CTC counts, and serum prostate-specific antigen (PSA) quantification. Our data demonstrated that in comparison with CellSearch, the CSV-based method had greater sensitivity and specificity. Further, we observed significantly greater numbers of CTCs in castration resistant patients as measured by our CSV method but not CellSearch. Our data suggests CSV-guided CTC enumeration may hold prognostic value and should be further validated as a possible measurement of PCA progression towards the deadly, androgen-independent form.

Asunto(s)

Membrana Celular/metabolismo , Transición Epitelial-Mesenquimal , Células Neoplásicas Circulantes/metabolismo , Neoplasias de la Próstata/metabolismo , Neoplasias de la Próstata/patología , Vimentina/metabolismo , Anciano , Biomarcadores de Tumor , Estudios de Casos y Controles , Línea Celular Tumoral , Progresión de la Enfermedad , Transición Epitelial-Mesenquimal/genética , Humanos , Masculino , Persona de Mediana Edad , Clasificación del Tumor , Metástasis de la Neoplasia , Neoplasias de la Próstata/terapia , Curva ROC , Vimentina/genética

RESUMEN

Cancer cell signaling, growth, morphology, proliferation and tumorigenic potential are largely depending on the signaling molecules present naturally in the tumor microenvironment and the identification of key molecules that drive the tumor progression is critical for the development of new modalities for the prevention of tumor progression. High concentrations of vimentin in the blood of cancer patients have been reported, however the function of blood circulating vimentin remains unknown. Here, we investigated the functional role of exogenously supplemented vimentin on colon cancer cells and examined the Wnt Signaling activation and cancer cell invasion. Vimentin when supplemented to the cancer cells remained bound to the surface of the cancer cells. Furthermore, bound vimentin activates Wnt signaling pathway as detectable by increased ß-catenin accumulation in the nucleus with concomitant activation of ß-catenin-dependent transcription of Wnt signaling downstream targets. Functionally, there was an increase in the rate of cellular invasion in these cancer cells upon binding with vimentin. Our results thus suggest that free vimentin in the tumor microenvironment acts as a positive regulator of the ß-catenin signaling pathway, thus providing a basis for cancer invasive properties.

RESUMEN

Although circulating tumor cells (CTCs) have potential as diagnostic biomarkers for cancer, determining their prognostic role in cancer patients undergoing treatment is a challenge. We evaluated the prognostic value of programmed death-ligand 1 (PD-L1) expression in CTCs in colorectal and prostate cancer patients undergoing treatment. Peripheral blood samples were collected from 62 metastatic colorectal cancer patients and 30 metastatic prostate cancer patients. CTCs were isolated from the samples using magnetic separation with the cell-surface vimentin(CSV)-specific 84-1 monoclonal antibody that detects epithelial-mesenchymal transitioned (EMT) CTCs. CTCs were enumerated and analyzed for PD-L1 expression using confocal microscopy. PD-L1 expression was detectable in CTCs and was localized in the membrane and/or cytoplasm and nucleus. CTC detection alone was not associated with poor progression-free or overall survival in colorectal cancer or prostate cancer patients, but nuclear PD-L1 (nPD-L1) expression in these patients was significantly associated with short survival durations. These results demonstrated that nPD-L1 has potential as a clinically relevant prognostic biomarker for colorectal and prostate cancer. Our data thus suggested that use of CTC-based models of cancer for risk assessment can improve the standard cancer staging criteria and supported the incorporation of nPD-L1 expression detection in CTCs detection in such models.

Asunto(s)

Antígeno B7-H1/metabolismo , Biomarcadores de Tumor/metabolismo , Neoplasias/metabolismo , Células Neoplásicas Circulantes/metabolismo , Vimentina/metabolismo , Antígeno B7-H1/genética , Núcleo Celular/metabolismo , Neoplasias Colorrectales/sangre , Neoplasias Colorrectales/diagnóstico , Neoplasias Colorrectales/metabolismo , Supervivencia sin Enfermedad , Células HCT116 , Células HEK293 , Humanos , Masculino , Neoplasias/sangre , Neoplasias/diagnóstico , Pronóstico , Neoplasias de la Próstata/sangre , Neoplasias de la Próstata/diagnóstico , Neoplasias de la Próstata/metabolismo , Estudios Retrospectivos

RESUMEN

BACKGROUND: Expression of programmed cell death ligand 1 (PD-L1) is an important process by which tumor cells suppress antitumor immunity in the tumor microenvironment. Bone marrow (BM)-derived immune cells are an important component of the tumor microenvironment. However, the link between PD-L1 induction on tumor cells and communication with BM cells is unknown. RESULTS: This study demonstrates that BM cells have a direct effect in inducing PD-L1 expression on tumor cells, which contributes to the tumor cells' drug resistance. This novel discovery was revealed using a co-incubation system with BM cells and tumor cells. BM cells from wild-type C57BL6 mice and the immune-deficient mouse strains B-cell(-/-), CD28(-/-), perforin(-/-), and Rag2(-/-) but not CD11b(-/-) dramatically increased the expression of tumor cell surface PD-L1. This PD-L1 induction was dependent on CD11b-positive BM cells through direct contact with tumor cells. Furthermore, p38 signaling was activated in tumor cells after co-incubation with BM cells, whereas the expression of PD-L1 was remarkably decreased after co-culture of cells treated with a p38 inhibitor. The increase in PD-L1 induced by BM cell co-culture protected tumor cells from drug-induced apoptosis. CONCLUSIONS: PD-L1 expression is increased on tumor cells by direct contact with BM-derived CD11b-positive cells through the p38 signaling pathway. PD-L1 may play an important role in drug resistance, which often causes failure of the antitumor response.

Asunto(s)

Antígeno B7-H1/biosíntesis , Células de la Médula Ósea/metabolismo , Antígeno CD11b/metabolismo , Comunicación Celular , Sistema de Señalización de MAP Quinasas , Proteínas de Neoplasias/metabolismo , Neoplasias/metabolismo , Animales , Apoptosis/genética , Antígeno B7-H1/genética , Células de la Médula Ósea/patología , Antígeno CD11b/genética , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica , Ratones , Ratones Noqueados , Proteínas de Neoplasias/genética , Neoplasias/genética , Neoplasias/patología , Células del Estroma/metabolismo , Células del Estroma/patología , Microambiente Tumoral/genética , Proteínas Quinasas p38 Activadas por Mitógenos/genética , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo

RESUMEN

PURPOSE: This study aimed to detect cell-surface vimentin (CSV) on the surface of epithelial-mesenchymal transitioned (EMT) circulating tumor cells (CTC) from blood of patients with epithelial cancers. EXPERIMENTAL DESIGN: In this study, 101 patients undergoing postsurgery adjuvant chemotherapy for metastatic colon cancer were recruited. EMT CTCs were detected from blood of patients using the 84-1 monoclonal antibody against CSV as a marker. EMT CTCs isolated were characterized further using EMT-specific markers, fluorescent in situ hybridization, and single-cell mutation analysis. RESULTS: Using the 84-1 antibody, we detected CSV exclusively on EMT CTCs from a variety of tumor types but not in the surrounding normal cells in the blood. The antibody exhibited very high specificity and sensitivity toward different epithelial cancer cells. With this antibody, we detected and enumerated EMT CTCs from patients. From our observations, we defined a cutoff of <5 or ≥5 EMT CTCs as the optimal threshold with respect to therapeutic response using ROC curves. Using this defined threshold, the presence of ≥5 EMT CTCs was associated with progressive disease, whereas patients with <5 EMT CTCs showed therapeutic response. CONCLUSION: Taken together, the number of EMT CTCs detected correlated with the therapeutic outcome of the disease. These results establish CSV as a universal marker for EMT CTCs from a wide variety of tumor types and thus provide the foundation for emerging CTC detection technologies and for studying the molecular regulation of these EMT CTCs.

Asunto(s)

Biomarcadores de Tumor/análisis , Neoplasias del Colon/diagnóstico , Transición Epitelial-Mesenquimal , Células Neoplásicas Circulantes , Vimentina/análisis , Adulto , Anciano , Anticuerpos Monoclonales , Progresión de la Enfermedad , Femenino , Citometría de Flujo , Humanos , Hibridación Fluorescente in Situ , Masculino , Persona de Mediana Edad , Sensibilidad y Especificidad , Análisis de la Célula Individual , Vimentina/biosíntesis

RESUMEN

Recent advances in cancer stem cell biology have shown that cancer stem-like cells with epithelial-mesenchymal transition (EMT) phenotypes are more aggressive and cause relapse; however, absence of a specific marker to isolate these EMT stem-like cells hampers research in this direction. Cell surface markers have been identified for isolating cancer stem-like cells, but none has been identified for isolating cancer stem-like cells with EMT phenotype. Recently, we discovered that Vimentin, an intracellular EMT tumor cell marker, is present on the surface of colon metastatic tumor nodules in the liver. In our study, we examined the potential of targeting cell surface Vimentin (CSV) to isolate stem-like cancer cells with EMT phenotype, by using a specific CSV-binding antibody, 84-1. Using this antibody, we purified the CSV-positive, CD133-negative (csVim(+) CD133(-) ) cell population from primary liver tumor cell suspensions and characterized for stem cell properties. The results of sphere assays and staining for the stem cell markers Sox2 and Oct4A demonstrated that csVim(+) CD133(-) cells have stem-like properties similar to csVim(-) CD133(+) population. Our investigation further revealed that the csVim(+) CD133(-) cells had EMT phenotypes, as evidenced by the presence of Twist and Slug in the nucleus, the absence of EpCAM on the cell surface and basal level of expression of epithelial marker E-cadherin. The csVimentin-negative CD133-positive stem cells do not have any EMT phenotypes. csVim(+) CD133(-) cells exhibited more aggressively metastatic in livers than csVim(-) CD133(+) cells. Our findings indicate that csVim(+) CD133(-) cells are promising targets for treatment and prevention of metastatic hepatocellular carcinoma.

Asunto(s)

Antígenos CD/metabolismo , Biomarcadores de Tumor/metabolismo , Transición Epitelial-Mesenquimal , Glicoproteínas/metabolismo , Células Madre Neoplásicas/metabolismo , Péptidos/metabolismo , Vimentina/metabolismo , Antígeno AC133 , Animales , Anticuerpos/inmunología , Anticuerpos/metabolismo , Western Blotting , Carcinoma Hepatocelular/metabolismo , Membrana Celular/metabolismo , Humanos , Neoplasias Hepáticas/metabolismo , Ratones Endogámicos NOD , Ratones Noqueados , Ratones SCID , Microscopía Confocal , Neoplasias Experimentales/metabolismo , Factor 3 de Transcripción de Unión a Octámeros/metabolismo , Unión Proteica/inmunología , Factores de Transcripción SOXB1/metabolismo , Células Tumorales Cultivadas , Vimentina/inmunología

RESUMEN

BACKGROUND: Detection, isolation, and enumeration of circulating tumor cells (CTCs) from cancer patients has become an important modality in clinical management of patients with breast cancer. Although CellSearch, an epithelial cell adhesion molecule (EpCAM)-based method that is used to isolate epithelial CTCs, has gained prominence, its inability to detect mesenchymal CTCs from breast cancer patients raises concerns regarding its utility in clinical management. METHODS: To address this gap in technology, we recently discovered the utility of cell-surface vimentin (CSV) as a marker for detecting mesenchymal CTCs from sarcoma tumors. In the present study, we tested the sensitivity and specificity of detecting CTCs from blood collected at a random time during therapy from each of 58 patients with metastatic breast cancer by use of 84-1 (a monoclonal antibody against CSV to detect epithelial/mesenchymal-transition CTCs) and CellSearch methods. Additionally, we tested the possibility of improving the sensitivity and specificity of detection by use of additional parameters including nuclear EpCAM localization and epithelial mesenchymal ratios. RESULTS: CTC counts with CSV were significant (P = 0.0053) in differentiating populations responsive and nonresponsive to treatment compared with CTC counts with CellSearch (P = 0.0564). The specificity of CTC detection was found to be highest when the sum of CTC counts from the 2 methods was above a threshold of 8 CTCs/7.5 mL. CONCLUSIONS: The sum of CTC counts from the CellSearch and CSV methods appears to provide new insights for assessment of therapeutic response and thus provides a new approach to personalized medicine in breast cancer patients.

Asunto(s)

Antígenos de Neoplasias/sangre , Biomarcadores de Tumor/sangre , Neoplasias de la Mama/sangre , Moléculas de Adhesión Celular/sangre , Membrana Celular/metabolismo , Transición Epitelial-Mesenquimal , Células Neoplásicas Circulantes/patología , Vimentina/metabolismo , Anticuerpos Monoclonales/inmunología , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Recuento de Células , Molécula de Adhesión Celular Epitelial , Transición Epitelial-Mesenquimal/efectos de los fármacos , Femenino , Humanos , Metástasis de la Neoplasia , Células Neoplásicas Circulantes/efectos de los fármacos , Proyectos Piloto , Estudios Retrospectivos , Resultado del Tratamiento , Vimentina/inmunología

RESUMEN

UNLABELLED: Chronic hepatic diseases, such as cirrhosis, hepatocellular carcinoma, and virus-mediated immunopathogenic infections, affect billions of people worldwide. These diseases commonly initiate with fibrosis. Owing to the various side effects of antifibrotic therapy and the difficulty of diagnosing asymptomatic patients, suitable medication remains a major concern. To overcome this drawback, the use of cytokine-based sustained therapy might be a suitable alternative with minimal side effects. Here, we studied the therapeutic efficacy and potential mechanisms of interleukin (IL)-30 as antifibrosis therapy in murine liver fibrosis models. CCl4 or 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) 0.1% (wt/wt) Purina 5015 Chow (LabDiet, St. Louis, MO) was fed for 3 weeks to induce liver fibrosis. Either control vector (pCtr) or pIL30 was injected hydrodynamically once per week. A significant decrease in collagen deposition and reduced expression of alpha-smooth muscle actin (α-SMA) protein indicated that IL-30-based gene therapy dramatically reduced bridging fibrosis that was induced by CCl4 or DDC. Immunophenotyping and knockout studies showed that IL-30 recruits natural-killer-like T (NKT) cells to the liver to remove activated hepatic stellate cells (HSCs) significantly and ameliorate liver fibrosis. Both flow cytometric and antibody-mediated neutralization studies showed that liver NKT cells up-regulate the natural killer group 2, member D (NKG2D) ligand and bind with the NKG2D ligand, retinoic acid early inducible 1 (Rae1), and positively activated HSCs to ameliorate liver fibrosis. Furthermore, adoptive transfer of liver NKT cells in T-cell-deficient mice showed reduction of fibrosis upon IL-30 administration. CONCLUSIONS: Highly target-specific liver NKT cells selectively remove activated HSCs through an NKG2D-Rae1 interaction to ameliorate liver fibrosis after IL-30 treatment.

Asunto(s)

Células Estrelladas Hepáticas/efectos de los fármacos , Interleucinas/uso terapéutico , Cirrosis Hepática/tratamiento farmacológico , Subfamilia K de Receptores Similares a Lectina de Células NK/metabolismo , Células T Asesinas Naturales/efectos de los fármacos , Proteínas Asociadas a Matriz Nuclear/metabolismo , Proteínas de Transporte Nucleocitoplasmático/metabolismo , Animales , Tetracloruro de Carbono , Evaluación Preclínica de Medicamentos , Femenino , Células Estrelladas Hepáticas/metabolismo , Interleucinas/farmacología , Cirrosis Hepática/inducido químicamente , Cirrosis Hepática/inmunología , Ratones Endogámicos C57BL , Células T Asesinas Naturales/metabolismo , Piridinas

RESUMEN

To date, no specific marker exists for the detection of circulating tumor cells (CTC) from different types of sarcomas, though tools are available for detection of CTCs in peripheral blood of patients with cancer for epithelial cancers. Here, we report cell-surface vimentin (CSV) as an exclusive marker on sarcoma CTC regardless of the tissue origin of the sarcoma as detected by a novel monoclonal antibody. Utilizing CSV as a probe, we isolated and enumerated sarcoma CTCs with high sensitivity and specificity from the blood of patients bearing different types of sarcoma, validating their phenotype by single cell genomic amplification, mutation detection, and FISH. Our results establish the first universal and specific CTC marker described for enumerating CTCs from different types of sarcoma, thereby providing a key prognosis tool to monitor cancer metastasis and relapse.

Asunto(s)

Biomarcadores de Tumor/metabolismo , Células Neoplásicas Circulantes/metabolismo , Sarcoma/patología , Vimentina/metabolismo , Anticuerpos Monoclonales/química , Secuencia de Bases , Línea Celular Tumoral , Análisis Mutacional de ADN , Citometría de Flujo , Técnica del Anticuerpo Fluorescente Indirecta , Amplificación de Genes , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Humanos , Microscopía Fluorescente , Pronóstico , Sarcoma/diagnóstico , Sarcoma/metabolismo , Sensibilidad y Especificidad , Análisis de la Célula Individual , Proteína p53 Supresora de Tumor/genética , Vimentina/inmunología

RESUMEN

The safest and most effective cytokine therapies require the favorable accumulation of the cytokine in the tumor environment. While direct treatment into the neoplasm is ideal, systemic tumor-targeted therapies will be more feasible. Electroporation-mediated transfection of cytokine plasmid DNA including a tumor-targeting peptide-encoding sequence is one method for obtaining a tumor-targeted cytokine produced by the tumor-bearing patient's tissues. Here, the impact on efficacy of the location of targeting peptide, choice of targeting peptide, tumor histotype, and cytokine utilization are studied in multiple syngeneic murine tumor models. Within the same tumor model, the location of the targeting peptide could either improve or reduce the antitumor effect of interleukin (IL)12 gene treatments, yet in other tumor models the tumor-targeted IL12 plasmid DNAs were equally effective regardless of the peptide location. Similarly, the same targeting peptide that enhances IL12 therapies in one model fails to improve the effect of either IL15 or PF4 for inhibiting tumor growth in the same model. These interesting and sometimes contrasting results highlight both the efficacy and personalization of tumor-targeted cytokine gene therapies while exposing important aspects of these same therapies which must be considered before progressing into approved treatment options.

Asunto(s)

Citocinas/metabolismo , Regulación Neoplásica de la Expresión Génica , Terapia Genética/métodos , Neoplasias/terapia , Procesamiento Proteico-Postraduccional , Animales , Línea Celular Tumoral , ADN/metabolismo , Modelos Animales de Enfermedad , Electroporación , Femenino , Interleucina-12/genética , Interleucina-15/genética , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C3H , Ratones Endogámicos C57BL , Neoplasias/genética , Péptidos/química , Plásmidos/metabolismo , Transfección

RESUMEN

Bioflavonoids are of considerable interest to human health as these serve as antioxidant and anticancer agents. Although epidemiological and experimental studies suggest that luteolin, a natural bioflavonoid, exhibits chemopreventive properties, its effectiveness as an antiproliferative agent against multidrug resistant (MDR) cancers is unclear. Thus, we assessed the antiproliferative effects of luteolin and associated molecular mechanisms using two MDR cancer cell lines that express high levels of P-glycoprotein and ABCG2. In this article, we demonstrate that luteolin induces apoptosis in P-glycoprotein- and ABCG2-expressing MDR cancer cells without affecting the transport functions of these drug transporters. Analysis of various proliferative signaling pathways indicated that luteolin-induced apoptosis involves reactive oxygen species generation, DNA damage, activation of ATR → Chk2 → p53 signaling pathway, inhibition of NF-kB signaling pathway, activation of p38 pathway and depletion of antiapoptotic proteins. Importantly, use of luteolin in these analyses also identified specific molecular characteristics of NCI-ADR/RES and MCF-7/Mito(R) cells that highlight their different tissue origins. These results suggest that luteolin possesses therapeutic potential to control the proliferation of MDR cancers without affecting the physiological function of drug transporters in the body tissues.

Asunto(s)

Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Luteolina/farmacología , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/fisiología , Transportador de Casetes de Unión a ATP, Subfamilia G, Miembro 2 , Transportadoras de Casetes de Unión a ATP/fisiología , Caspasa 7/metabolismo , Puntos de Control del Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Resistencia a Múltiples Medicamentos , Resistencia a Antineoplásicos , Humanos , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Proteínas de Neoplasias/fisiología , Especies Reactivas de Oxígeno/metabolismo

RESUMEN

Vimentin, a major constituent of the intermediate filament family of proteins, is ubiquitously expressed in normal mesenchymal cells and is known to maintain cellular integrity and provide resistance against stress. Vimentin is overexpressed in various epithelial cancers, including prostate cancer, gastrointestinal tumors, tumors of the central nervous system, breast cancer, malignant melanoma, and lung cancer. Vimentin's overexpression in cancer correlates well with accelerated tumor growth, invasion, and poor prognosis; however, the role of vimentin in cancer progression remains obscure. In recent years, vimentin has been recognized as a marker for epithelial-mesenchymal transition (EMT). Although EMT is associated with several tumorigenic events, vimentin's role in the underlying events mediating these processes remains unknown. By virtue of its overexpression in cancer and its association with tumor growth and metastasis, vimentin serves as an attractive potential target for cancer therapy; however, more research would be crucial to evaluate its specific role in cancer. Our recent discovery of a vimentin-binding mini-peptide has generated further impetus for vimentin-targeted tumor-specific therapy. Furthermore, research directed toward elucidating the role of vimentin in various signaling pathways would reveal new approaches for the development of therapeutic agents. This review summarizes the expression and functions of vimentin in various types of cancer and suggests some directions toward future cancer therapy utilizing vimentin as a potential molecular target.

Asunto(s)

Sistemas de Liberación de Medicamentos/métodos , Transición Epitelial-Mesenquimal/fisiología , Regulación Neoplásica de la Expresión Génica/fisiología , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Transducción de Señal/fisiología , Vimentina/metabolismo , Transición Epitelial-Mesenquimal/genética , Regulación Neoplásica de la Expresión Génica/genética , Humanos , Modelos Biológicos , Transducción de Señal/genética , Vimentina/genética

RESUMEN

Galectin-1 (gal-1) is an important molecule secreted by many tumors, which induces apoptosis in activated T-cells and promotes tumor angiogenesis, both of which phenomena facilitate successful establishment of tumor in the body. However, little is known about the function of intracellular gal-1 or its transcriptional regulation in colorectal cancer (CRC). Here, we demonstrate that gal-1 expression is epigenetically regulated in CRC through promoter hypermethylation. Intracellular gal-1 induces cell cycle arrest and apoptosis in CRC cells with concomitant down-regulation of Wnt and NF-κB signaling pathways. Together, these data suggested that gal-1 silencing imparts CRC with the ability to proliferate and escape apoptosis.

Asunto(s)

Apoptosis , Neoplasias Colorrectales/etiología , Metilación de ADN , Galectina 1/fisiología , Silenciador del Gen , Regiones Promotoras Genéticas , Apoptosis/efectos de los fármacos , Butiratos/farmacología , Caspasas/fisiología , Ciclo Celular , Línea Celular Tumoral , Proliferación Celular , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/patología , Galectina 1/antagonistas & inhibidores , Galectina 1/genética , Humanos , Potencial de la Membrana Mitocondrial

RESUMEN

Development of colorectal cancer (CRC) involves a series of genetic alterations with altered expression of proteins and cell signaling pathways. Here, we identified that galectin-4 (gal-4), a marker of differentiation, was down-regulated in CRC. The goal of this work was to determine the function of gal-4 in CRC. Toward this goal, the human colon biopsies and tissue microarrays containing a gradient of pathology were analyzed for gal-4 expression by immunohistochemistry. Cell proliferation, migration, motility, forced expression, knockdown, cell cycle and apoptosis assays were used to characterize gal-4 function. Immunohistochemistry identified that gal-4 expression was significantly down-regulated in adenomas and was essentially absent in invasive carcinomas. Forced expression of gal-4 in gal-4 -ve cells induced cell cycle arrest and retarded cell migration and motility. Further, gal-4 sensitized the cells to camptothecin-induced apoptosis. Gal-4 knockdown resulted in increased cell proliferation, migration and motility. Gal-4 was found to be associated with Wnt signaling proteins. Finally, gal-4 expression led to down-regulation of Wnt signaling target genes. This study demonstrates that loss of gal-4 is a common and specific event in CRC. This study also shows that gal-4 exhibits tumor suppressive effects in CRC cells in vitro. Through its ability to interact with and down-regulate the functions of Wnt signaling pathway, gal-4 reveals a new dimension in the control of the Wnt signaling pathway. Thus, gal-4 may prove to be an important molecule in understanding the biology of CRC.

Asunto(s)

Adenocarcinoma/metabolismo , Adenoma/metabolismo , Neoplasias Colorrectales/metabolismo , Galectina 4/metabolismo , Genes Supresores de Tumor , Adenocarcinoma/genética , Adenocarcinoma/patología , Adenoma/genética , Adenoma/patología , Apoptosis , Western Blotting , Adhesión Celular , Ciclo Celular , Movimiento Celular , Proliferación Celular , Colon/metabolismo , Colon/patología , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/patología , Regulación hacia Abajo , Galectina 4/antagonistas & inhibidores , Galectina 4/genética , Humanos , Técnicas para Inmunoenzimas , Invasividad Neoplásica , ARN Interferente Pequeño/genética , Transducción de Señal , Células Tumorales Cultivadas , Proteínas Wnt/metabolismo , Cicatrización de Heridas , beta Catenina/metabolismo

RESUMEN

RNF2, a member of polycomb group (PcG) proteins, is involved in chromatin remodeling. However, mechanisms that regulate RNF2 function are unknown. To identify such mechanisms, RNF2 was expressed in HEK-293 cells and analyzed by 2-D electrophoresis. RNF2 was resolved into at least seven protein spots, migrating toward the lower pI from its expected pI of 6.38, suggesting that RNF2 undergoes post-translational modifications. Western blotting indicated that majority of these RNF2 spots contained phosphoserine(s), which were completely dephosphorylated upon treatment with a phosphatase. SB203580, a specific inhibitor of p38 MAPK, inhibited RNF2 phosphorylation at one site. On the other hand, PD98059, an inhibitor of MEK1/2, inhibited majority of the phosphorylation events in RNF2. Mass spectrometry analysis identified that RNF2 expressed in Sf9 insect cells undergoes co-translational excision of (1)Met coupled to N-acetylation of (2)Ser, and phosphorylation of (41)Ser. Interestingly, (41)Ser is a predicted p38/MAPK phosphorylation site, consistent with the loss of phosphorylation induced by SB203580. Further analysis indicated that RNF2 phosphorylation differentially modulates the expression of transcription factors and histone 2B acetylation. These results provide first evidence for phosphorylation of RNF2, and suggest that the mitogen activated protein kinases including p38 MAPK and ERK1/2 regulate growth, stress response, differentiation and other cellular processes, through phosphorylation of RNF2.

Asunto(s)

Proteínas de Unión al ADN/metabolismo , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Sistema de Señalización de MAP Quinasas , Proteínas Represoras/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Línea Celular , Proteínas de Unión al ADN/genética , Electroforesis en Gel Bidimensional , Regulación de la Expresión Génica , Humanos , Fosforilación , Fosfoserina/metabolismo , Complejo Represivo Polycomb 1 , Proteínas del Grupo Polycomb , Procesamiento Proteico-Postraduccional , Proteómica , Proteínas Represoras/genética , Espectrometría de Masas en Tándem , Ubiquitina-Proteína Ligasas/genética