RESUMEN
The regulatory role of protein tyrosine kinases in ß1- and ß2-integrin activation and in the survival of chronic lymphocytic leukemia (CLL) cells is well established. In contrast, the involvement of protein tyrosine phosphatases in CLL biology was less investigated. We show that selective activation of the protein tyrosine phosphatase receptor type γ (PTPRG) strongly suppresses integrin activation and survival in leukemic B cells isolated from patients with CLL. Activation of PTPRG specifically inhibits CXCR4- as well as BCR-induced triggering of LFA-1 and VLA-4 integrins and mediated rapid adhesion. Triggering of LFA-1 affinity is also prevented by PTPRG activity. Analysis of signaling mechanisms shows that activation of PTPRG blocks chemokine-induced triggering of JAK2 and Bruton's tyrosine kinase protein tyrosine kinases and of the small GTP-binding protein RhoA. Furthermore, activated PTPRG triggers rapid and robust caspase-3/7-mediated apoptosis in CLL cells in a manner quantitatively comparable to the Bruton's tyrosine kinase inhibitor ibrutinib. However, in contrast to ibrutinib, PTPRG-triggered apoptosis is insensitive to prosurvival signals generated by CXCR4 and BCR signaling. Importantly, PTPRG activation does not trigger apoptosis in healthy B lymphocytes. The data show that activated PTPRG inhibits, at once, the signaling pathways controlling adhesion and survival of CLL cells, thus emerging as a negative regulator of CLL pathogenesis. These findings suggest that pharmacological potentiation of PTPRG tyrosine-phosphatase enzymatic activity could represent a novel approach to CLL treatment.
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Agammaglobulinemia Tirosina Quinasa/metabolismo , Adhesión Celular/fisiología , Supervivencia Celular/fisiología , Leucemia Linfocítica Crónica de Células B/metabolismo , Proteínas Tirosina Fosfatasas Clase 5 Similares a Receptores/metabolismo , Adenina/análogos & derivados , Adenina/farmacología , Apoptosis/efectos de los fármacos , Apoptosis/fisiología , Linfocitos B/efectos de los fármacos , Linfocitos B/metabolismo , Adhesión Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Humanos , Integrina alfa4beta1/metabolismo , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Antígeno-1 Asociado a Función de Linfocito/metabolismo , Piperidinas/farmacología , Inhibidores de Proteínas Quinasas/farmacología , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiologíaRESUMEN
Mitochondria are critical organelles in the regulation of intrinsic apoptosis. As a general feature of blood cancers, different antiapoptotic members of the BCL-2 protein family localize at the outer mitochondrial membrane to sequester variable amounts of proapoptotic activators, and hence protect cancer cells from death induction. However, the impact of distinct anti-apoptotic members on apoptosis prevention, a concept termed anti-apoptotic dependence, differs remarkably across disease entities. Over the last two decades, several genetic and functional methodologies have been established to uncover the anti-apoptotic dependencies of the majority of blood cancers, inspiring the development of a new class of small molecules called BH3 mimetics. In this review, we highlight the rationale of targeting mitochondrial apoptosis in hematology, and provide a comprehensive map of the anti-apoptotic dependencies that are currently guiding novel therapeutic strategies. Cell-extrinsic and -intrinsic mechanisms conferring resistance to BH3 mimetics are also examined, with insights on potential strategies to overcome them. Finally, we discuss how the field of mitochondrial apoptosis might be complemented with other dimensions of precision medicine for more successful treatment of 'highly complex' hematologic malignancies.
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Neoplasias Hematológicas , Medicina de Precisión , Apoptosis , Biología , Neoplasias Hematológicas/tratamiento farmacológico , Neoplasias Hematológicas/genética , Neoplasias Hematológicas/metabolismo , Humanos , Mitocondrias/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismoRESUMEN
Tumor dormancy is a poorly understood stage in cancer progression characterized by mitotic cycle arrest in G0/G1 phase and low metabolism. The cells survive in a quiescent state and wait for appropriate environmental conditions to begin proliferation again giving rise to metastasis. Despite their key role in cancer development and metastasis, the knowledge about their biology and origin is still very limited due to the poorness of established in vitro models that faithfully recapitulated tumor dormancy. Using at least three cycles of 1% O2 hypoxia and reoxygenation, we establish and characterize the hypoxia-resistant human breast cancer cell line chMDA-MB-231 that can stably survive under 1% O2 condition by entering into dormant state characterized by arrest in G0/G1 phase and low metabolism. This dormant state is reversible since once replaced in normoxia the cells recover the proliferation rate in 2 weeks. We show that chronic hypoxia induces autophagy that may be the survival mechanism of chMDA-MB-231 cells. Furthermore, the data in this work demonstrate that cycling hypoxic/reoxygenation stress selects MDA-MB-231 population that presents the cancer stem-like phenotype characterized by CD24- /CD44+ /ESA+ expression and spheroid forming capacity. We believe that our study presents a promising approach to select dormant breast cancer cells with stem-like phenotype using the hypoxia/reoxygenation regimen that may represent an area with profound implications for therapeutic developments in oncology. J. Cell. Biochem. 118: 3237-3248, 2017. © 2017 Wiley Periodicals, Inc.
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Neoplasias de la Mama/metabolismo , Fase G1 , Células Madre Neoplásicas/metabolismo , Fase de Descanso del Ciclo Celular , Neoplasias de la Mama/patología , Hipoxia de la Célula , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Células MCF-7 , Metástasis de la Neoplasia , Células Madre Neoplásicas/patologíaRESUMEN
New effective treatments are needed to improve outcomes for multiple myeloma (MM) patients. Receptors with restricted expression on plasma cells (PCs) represent attractive new therapeutic targets. The endothelin-1 (EDN1) axis, consisting of EDN1 acting through EDN-receptor A (EDNRA) and B (EDNRB), was previously shown to be overexpressed in several tumours, including MM. However, there is incomplete understanding of how EDN1 axis regulates MM growth and response to therapy. Besides EDNRA, the majority of MM cell lines and primary malignant PCs express high levels of EDNRB and release EDN1. Similarly, bone-marrow microenvironment cells also secrete EDN1. Investigating the extent of epigenetic dysregulation of EDNRB gene in MM, we found that hypermethylation of EDNRB promoter and subsequent down-regulation of EDNRB gene was observed in PCs or B lymphocytes from healthy donors compared to EDNRB-expressing malignant PCs. Pharmacological blockade with the dual EDN1 receptor antagonist bosentan decreased cell viability and MAPK activation of U266 and RPMI-8226 cells. Interestingly, the combination of bosentan and the proteasome inhibitor bortezomib, currently approved for MM treatment, resulted in synergistic cytotoxic effects. Overall, our data has uncovered EDN1-mediated autocrine and paracrine mechanisms that regulate malignant PCs growth and drug response, and support EDN1 receptors as new therapeutic targets in MM.
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Antagonistas de los Receptores de la Endotelina A/farmacología , Mieloma Múltiple/sangre , Receptor de Endotelina A/sangre , Adulto , Anciano , Anciano de 80 o más Años , Comunicación Autocrina/fisiología , Bortezomib/farmacología , Bosentán , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Metilación de ADN , ADN de Neoplasias/genética , Sinergismo Farmacológico , Endotelina-1/sangre , Endotelina-1/fisiología , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Persona de Mediana Edad , Terapia Molecular Dirigida/métodos , Mieloma Múltiple/genética , Mieloma Múltiple/patología , Células Plasmáticas/metabolismo , Regiones Promotoras Genéticas , Receptor de Endotelina A/genética , Sulfonamidas/farmacología , Células Tumorales Cultivadas/efectos de los fármacos , Células Tumorales Cultivadas/patologíaRESUMEN
A new immunological investigation was carried out to study the association between non-Hodgkin lymphoma and Simian virus 40 (SV40). To this end, a new indirect ELISA was employed with two mimotopes from SV40 large T antigen (Tag), the viral oncoprotein, to analyse for specific reactions to antibodies in sera from non-Hodgkin lymphoma patients and controls, represented by healthy subjects (HS) and breast carcinoma (BC) patients. This study allowed us to assay a new sera collection from non-Hodgkin lymphoma patients (NHL, n = 254). To verify the association between NHL and SV40 Tag, two totally independent cohorts were analysed: NHL1 n = 150 and NHL2 n = 104. The epidemiological survey included sera from HS1, n = 150; HS2, n = 104 and BC, n = 78. This new indirect ELISA revealed that antibodies against SV40 Tag mimotopes are detectable in NHL1 and NHL2 sera with a prevalence of 37 and 36%, respectively. The prevalence of SV40-antibodies detected in both NHL1 and NHL2 cohorts differs statistically from controls, at 19% for HS1 (p < 0.01), HS2 (p < 0.05) and BC patients (p < 0.05). This study, carried out with an immunological assay with specific Tag oncoprotein mimotopes of Simian virus 40, reports the presence of IgG antibodies against the large Tumour antigen in non-Hodgkin lymphomas for the first time. Our immunological data with two independent NHL cohorts show a statistically significant association between Simian virus 40 Tag and non-Hodgkin lymphoma. These results suggest that SV40-positive non-Hodgkin lymphomas could be treated differently from those tested SV40-negative.
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Anticuerpos Antivirales/inmunología , Antígenos Virales de Tumores/inmunología , Linfoma no Hodgkin/inmunología , Proteínas Oncogénicas/metabolismo , Virus 40 de los Simios/inmunología , Adulto , Animales , Femenino , Humanos , Linfoma no Hodgkin/patología , Ratones , Ratones Transgénicos , Persona de Mediana Edad , PrevalenciaRESUMEN
The sesquiterpene α-bisabolol (α-BSB) has been shown to be an effective cytotoxic agent for a variety of human cancer cells in culture and animal models. However, much of its intracellular action remains elusive. We evaluated the cytotoxic action of α-BSB against CML-T1, Jurkat and HeLa cell lines, as preclinical models for myeloid, lymphoid and epithelial neoplasias. The approach included single cell analysis (flow cytometry, immunocytology) combined with cytotoxicity and proliferation assays to characterize organelle damage, autophagy, cytostatic effect, and apoptosis. The study focuses on the relevant steps in the cytotoxic cascade triggered by α-BSB: (1) the lipid rafts through which α-BSB enters the cells, (2) the opening of pores in the mitochondria and lysosomes, (3) the activation of both caspase-dependent and caspase-independent cell death pathways, (4) the induction of autophagy and (5) apoptosis. The effectiveness of α-BSB as an agent against tumor cells is grounded on its capability to act on different layers of cell regulation to elicit different concurrent death signals, thereby neutralizing a variety of aberrant survival mechanisms leading to treatment resistance in neoplastic cell.
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Antineoplásicos/farmacología , Muerte Celular/efectos de los fármacos , Lisosomas/efectos de los fármacos , Mitocondrias/efectos de los fármacos , Sesquiterpenos/farmacología , Apoptosis/efectos de los fármacos , Caspasas/metabolismo , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Células HeLa , Humanos , Células Jurkat , Lisosomas/metabolismo , Mitocondrias/metabolismo , Sesquiterpenos MonocíclicosRESUMEN
BACKGROUND: Non-Hodgkin lymphoma (NHL), the most common cancer of the lymphatic system, is of unknown etiology. The identification of etiologic factors in the onset of NHL is a key event that could facilitate the prevention and cure of this malignancy. Simian virus 40 (SV40) has been considered an oncogenic agent in the onset/progression of NHL. METHODS: In this study, an indirect enzyme-linked immunosorbent assay with 2 synthetic peptides that mimic SV40 antigens of viral capsid proteins 1 to 3 was employed to detect specific antibodies against SV40. Serum samples were taken from 2 distinct cohorts of NHL-affected patients (NHL1 [n = 89] and NHL2 [n = 61]) along with controls represented by oncologic patients affected by breast cancer (BC; n = 78) and undifferentiated nasopharyngeal carcinoma (UNPC; n = 64) and 3 different cohorts of healthy subjects (HSs; HS1 [n = 130], HS2 [n = 83], and HS3 [n = 87]). RESULTS: Immunologic data indicated that in serum samples from NHL patients, antibodies against SV40 mimotopes were detectable with a prevalence of 40% in NHL1 patients and with a prevalence of 43% in NHL2 patients. In HSs of the same median age as NHL patients, the prevalence was 16% for the HS1 group (57 years) and 14% for the HS2 group (65 years). The difference was statistically significant (P < .0001 and P < .001). Interestingly, the difference between NHL1/NHL2 patients and BC patients (40%/43% vs 15%, P < .001) and between NHL1/NHL2 patients and UNPC patients (40%/43% vs 25%, P < .05) was significant. CONCLUSIONS: Our data indicate a strong association between NHL and SV40 and thus a need for innovative therapeutic approaches for this hematologic malignancy.
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Anticuerpos Antivirales/sangre , Linfoma no Hodgkin/virología , Infecciones por Polyomavirus/complicaciones , Virus 40 de los Simios/aislamiento & purificación , Infecciones Tumorales por Virus/virología , Adulto , Anciano , Proteínas de la Cápside/inmunología , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , Linfoma no Hodgkin/clasificación , Linfoma no Hodgkin/inmunología , Masculino , Persona de Mediana Edad , Estudios Seroepidemiológicos , Infecciones Tumorales por Virus/clasificaciónAsunto(s)
Endotelina-1 , Mieloma Múltiple , Humanos , Pirimidinas , Receptor de Endotelina A , SulfonamidasRESUMEN
Richter's syndrome (RS) is the transformation of chronic lymphocytic leukemia (CLL) into a high-grade B-cell malignancy. Molecular and functional studies have pointed out that CLL cells are close to the apoptotic threshold and dependent on BCL-2 for survival. However, it remains undefined how evasion from apoptosis evolves during disease transformation. Here, we employed functional and static approaches to compare the regulation of mitochondrial apoptosis in CLL and RS. BH3 profiling of 17 CLL and 9 RS samples demonstrated that RS cells had reduced apoptotic priming and lower BCL-2 dependence than CLL cells. While a subset of RS was dependent on alternative anti-apoptotic proteins and was sensitive to specific BH3 mimetics, other RS cases harbored no specific anti-apoptotic addiction. Transcriptomics of paired CLL/RS samples revealed downregulation of pro-apoptotic sensitizers during disease transformation. Albeit expressed, effector and activator members were less likely to colocalize with mitochondria in RS compared to CLL. Electron microscopy highlighted reduced cristae width in RS mitochondria, a condition further promoting apoptosis resistance. Collectively, our data suggest that RS cells evolve multiple mechanisms that lower the apoptotic priming and shift the anti-apoptotic dependencies away from BCL-2, making direct targeting of mitochondrial apoptosis more challenging after disease transformation.
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Apoptosis , Leucemia Linfocítica Crónica de Células B , Mitocondrias , Proteínas Proto-Oncogénicas c-bcl-2 , Humanos , Leucemia Linfocítica Crónica de Células B/patología , Leucemia Linfocítica Crónica de Células B/genética , Leucemia Linfocítica Crónica de Células B/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/genética , Mitocondrias/metabolismo , Masculino , Femenino , Persona de Mediana EdadRESUMEN
Patients with chronic lymphocytic leukemia (CLL) relapsing on ibrutinib are often treated with the Bcl-2 inhibitor venetoclax. However, the transition from one agent to another poses some clinical challenges due to disease flares sometimes occurring right after ibrutinib interruption. Here, we describe three clinical vignettes highlighting two distinct patterns of ibrutinib-to-venetoclax transition. While patients following the favorable pattern transited to venetoclax without experiencing disease flare, the one patient who took the unfavorable path showed rapid disease rebound, with large-cell transformation occurring one week after ibrutinib interruption. A high burden of BTK and PLCG2 mutations was found only in patients with the favorable transition pattern, suggesting that removing BTK inhibition might be particularly harmful if CLL cells are progressing through mechanisms external to the BTK axis.
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Leucemia Linfocítica Crónica de Células B , Adenina/análogos & derivados , Compuestos Bicíclicos Heterocíclicos con Puentes/uso terapéutico , Humanos , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Leucemia Linfocítica Crónica de Células B/genética , Piperidinas/uso terapéutico , SulfonamidasRESUMEN
BACKGROUND: We previously demonstrated that the plant-derived agent α-bisabolol enters cells via lipid rafts, binds to the pro-apoptotic Bcl-2 family protein BID, and may induce apoptosis. Here we studied the activity of α-bisabolol in acute leukemia cells. METHODS: We tested ex vivo blasts from 42 acute leukemias (14 Philadelphia-negative and 14 Philadelphia-positive B acute lymphoid leukemias, Ph-/Ph+B-ALL; 14 acute myeloid leukemias, AML) for their sensitivity to α-bisabolol in 24-hour dose-response assays. Concentrations and time were chosen based on CD34+, CD33+my and normal peripheral blood cell sensitivity to increasing α-bisabolol concentrations for up to 120 hours. RESULTS: A clustering analysis of the sensitivity over 24 hours identified three clusters. Cluster 1 (14 ± 5 µM α-bisabolol IC50) included mainly Ph-B-ALL cells. AML cells were split into cluster 2 and 3 (45 ± 7 and 65 ± 5 µM IC50). Ph+B-ALL cells were scattered, but mainly grouped into cluster 2. All leukemias, including 3 imatinib-resistant cases, were eventually responsive, but a subset of B-ALL cells was fairly sensitive to low α-bisabolol concentrations. α-bisabolol acted as a pro-apoptotic agent via a direct damage to mitochondrial integrity, which was responsible for the decrease in NADH-supported state 3 respiration and the disruption of the mitochondrial membrane potential. CONCLUSION: Our study provides the first evidence that α-bisabolol is a pro-apoptotic agent for primary human acute leukemia cells.
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Apoptosis/efectos de los fármacos , Leucemia/patología , Modelos Biológicos , Sesquiterpenos/farmacología , Adolescente , Adulto , Proteína Proapoptótica que Interacciona Mediante Dominios BH3/metabolismo , Benzamidas , Crisis Blástica/metabolismo , Crisis Blástica/patología , Células Sanguíneas/efectos de los fármacos , Respiración de la Célula/efectos de los fármacos , Análisis por Conglomerados , Medios de Cultivo , Ensayos de Selección de Medicamentos Antitumorales , Femenino , Proteínas de Fusión bcr-abl/genética , Humanos , Mesilato de Imatinib , Leucemia/diagnóstico , Leucemia/metabolismo , Masculino , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Persona de Mediana Edad , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Sesquiterpenos Monocíclicos , Mutación/genética , Piperazinas/farmacología , Pirimidinas/farmacología , Sesquiterpenos/química , Solubilidad , Factores de Tiempo , Células Tumorales Cultivadas , Adulto JovenRESUMEN
BACKGROUND: Increased numbers of tumour-associated macrophages correlate with shortened survival in some cancers. The molecular bases of this correlation are not thoroughly understood. Events triggered by CXCL12 may play a part, as CXCL12 drives the migration of both CXCR4-positive cancer cells and macrophages and may promote a molecular crosstalk between them. RESULTS: Samples of HER1-positive colon cancer metastases in liver, a tissue with high expression of CXCL12, were analysed by immunohistochemistry. In all of the patient biopsies, CD68-positive tumour-associated macrophages presented a mixed CXCL10 (M1)/CD163 (M2) pattern, expressed CXCR4, GM-CSF and HB-EGF, and some stained positive for CXCL12. Cancer cells stained positive for CXCR4, CXCL12, HER1, HER4 and GM-CSF. Regulatory interactions among these proteins were validated via experiments in vitro involving crosstalk between human mononuclear phagocytes and the cell lines DLD-1 (human colon adenocarcinoma) and HeLa (human cervical carcinoma), which express the above-mentioned ligand/receptor repertoire. CXCL12 induced mononuclear phagocytes to release HB-EGF, which activated HER1 and triggered anti-apoptotic and proliferative signals in cancer cells. The cancer cells then proliferated and released GM-CSF, which in turn activated mononuclear phagocytes and induced them to release more HB-EGF. Blockade of GM-CSF with neutralising antibodies or siRNA suppressed this loop. CONCLUSIONS: CXCL12-driven stimulation of cancer cells and macrophages may elicit and reinforce a GM-CSF/HB-EGF paracrine loop, whereby macrophages contribute to cancer survival and expansion. The involvement of mixed M1/M2 GM-CSF-stimulated macrophages in a tumour-promoting loop may challenge the paradigm of tumour-favouring macrophages as polarized M2 mononuclear phagocytes.
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Quimiocina CXCL12/farmacología , Neoplasias del Colon/metabolismo , Factor Estimulante de Colonias de Granulocitos y Macrófagos/metabolismo , Péptidos y Proteínas de Señalización Intercelular/metabolismo , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Anciano , Animales , Apoptosis/efectos de los fármacos , Northern Blotting , Línea Celular , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Ensayo de Inmunoadsorción Enzimática , Receptores ErbB/metabolismo , Citometría de Flujo , Factor Estimulante de Colonias de Granulocitos y Macrófagos/farmacología , Células HeLa , Factor de Crecimiento Similar a EGF de Unión a Heparina , Humanos , Inmunoquímica , Técnicas In Vitro , Péptidos y Proteínas de Señalización Intercelular/farmacología , Espectrometría de Masas , Ratones , Fosforilación , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
The objectives of the study are to evaluate DNase I serum levels and their correlation with soluble Fas (sFas) and soluble Fas ligand (sFasL) and with cell surface Fas expression in patients with systemic lupus erythematosus (SLE), thus contributing to the dysregulated apoptosis typical of the disease. The methods include the following: Serum DNase I levels in patients and in controls were detected using the dot blot method and quantified by densitometry; sFas and sFasL were quantified using an ELISA system. Cell surface Fas expression was evaluated by FACS analysis. Apoptosis was studied by means of internucleosomal DNA degradation using a commercially available kit. The results are as follows: We found a significant difference in DNase I, sFas and sFasL serum levels between patients and controls. Levels of DNase I <7.79 ng ml(-1) are more represented in patients with SLE. Active SLE is strongly associated with high sFas levels and detectable sFasL. DNase I does not correlate with sFas or sFasL, whereas it correlates with T cell surface Fas expression that is higher in patients with active SLE than in healthy controls. Finally, administration of exogenous human recombinant DNase (hrDNase) I to freshly isolated T cells up-regulates cell surface Fas expression and induces increased susceptibility to Fas-mediated apoptosis. In conclusion, our findings confirm that DNase I is low in SLE and suggest that it may play a role in apoptosis in SLE by regulating the surface expression of the cell death molecule Fas. This role may contribute to explain the inefficacy of hrDNase I in SLE, a treatment proposed for the ability of DNase I to remove DNA from auto-antigenic nucleoprotein complexes.
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Desoxirribonucleasa I/inmunología , Proteína Ligando Fas/inmunología , Lupus Eritematoso Sistémico/inmunología , Proteínas Recombinantes/inmunología , Receptor fas/sangre , Adulto , Apoptosis/efectos de los fármacos , Apoptosis/inmunología , Separación Celular , Células Cultivadas , Desoxirribonucleasa I/sangre , Desoxirribonucleasa I/farmacología , Desoxirribonucleasa I/uso terapéutico , Proteína Ligando Fas/sangre , Femenino , Citometría de Flujo , Humanos , Lupus Eritematoso Sistémico/sangre , Lupus Eritematoso Sistémico/tratamiento farmacológico , Lupus Eritematoso Sistémico/fisiopatología , Masculino , Persona de Mediana Edad , Proteínas Recombinantes/sangre , Proteínas Recombinantes/farmacología , Proteínas Recombinantes/uso terapéutico , Resultado del Tratamiento , Receptor fas/genética , Receptor fas/inmunologíaRESUMEN
Classic Hodgkin lymphoma (cHL) is a unique lymphoid neoplasm characterized by extensive immune infiltrates surrounding rare malignant Hodgkin Reed-Sternberg (HRS) cells. Different subsets of T and NK cells have long been recognized in the cHL microenvironment, yet their distinct contribution to disease pathogenesis has remained enigmatic. Very recently, novel platforms for high dimensional analysis of immune cells, such as single-cell RNA sequencing and mass cytometry, have revealed unanticipated insights into the composition of T- and NK-cell compartments in cHL. Advances in imaging techniques have better defined specific T-helper subpopulations physically interacting with neoplastic cells. In addition, the identification of novel cytotoxic subsets with an exhausted phenotype, typically enriched in cHL milieu, is shedding light on previously unrecognized immune evasion mechanisms. This review examines the immunological features and the functional properties of T and NK subsets recently identified in the cHL microenvironment, highlighting their pathological interplay with HRS cells. We also discuss how this knowledge can be exploited to predict response to immunotherapy and to design novel strategies to improve PD-1 blockade efficacy.
RESUMEN
In classical Hodgkin lymphoma (cHL), the significance of the interplay between Hodgkin and Reed-Sternberg cells (HRS) and reactive T cells remains poorly defined. By immunohistochemistry on bioptic cHL specimens, we found that HRS and surrounding T lymphocytes stained positive for IL-17 in 40% of cases. IL-17 was detectable in a similar proportion of patients' sera and correlated with disease burden. Supernatants of KM-H2 and HDLM-2 cHL cell lines guided preferential chemotaxis of CCR6+ T lymphocytes. Coculture of cHL cell lines with PBMC promoted the enrichment of Th17 lymphocytes and Foxp3+/IL-17+ cells, whereas T regulatory cells slightly decreased. Soluble CD30 downmodulated membrane CD30 expression on T cells and contributed to their polarization shift by stimulating IL-17 production and reducing IFN-γ synthesis. Thus, HRS and a number of reactive CD4+ T cells, attracted by tumor-secreted chemokines, produce an IL-17 tumor-shaped inflammatory milieu in a cHL subset.
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Ligando CD30/inmunología , Linfocitos T CD4-Positivos/inmunología , Enfermedad de Hodgkin/inmunología , Interleucina-17/inmunología , Antígeno Ki-1/inmunología , Linfocitos T Reguladores/inmunología , Microambiente Tumoral/inmunología , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Apoptosis , Ligando CD30/metabolismo , Linfocitos T CD4-Positivos/metabolismo , Movimiento Celular , Proliferación Celular , Femenino , Estudios de Seguimiento , Enfermedad de Hodgkin/metabolismo , Enfermedad de Hodgkin/patología , Humanos , Interleucina-17/metabolismo , Antígeno Ki-1/metabolismo , Leucocitos Mononucleares/inmunología , Leucocitos Mononucleares/metabolismo , Leucocitos Mononucleares/patología , Masculino , Persona de Mediana Edad , Pronóstico , Células de Reed-Sternberg , Linfocitos T Reguladores/metabolismo , Adulto JovenRESUMEN
West Nile virus is a zoonotic agent causing life-threatening encephalitis in a proportion of infected patients. Older age, immunosuppression, and mutations in specific host genes (e.g., CCR5 delta-32 mutation) predispose to neuroinvasive infection. We report on two cases of severe West Nile encephalitis in recently-treated, different-aged, chronic lymphocytic leukemia patients. Both patients developed high-grade fever associated with severe neurological impairment. The younger one harboured germ-line CCR5 delta-32 mutation, which might have played a role in the pathogenesis of its neuroinvasive manifestations.
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Ibrutinib is increasingly adopted for treating lymphoid malignancies. While growing amounts of data pile up about Ibrutinib mechanism of action on neoplastic B cells, little is known about its impact on other immune cells. Here we investigated the effect of Ibrutinib on monocyte/macrophage functions. (1) Ibrutinib treatment of purified human monocytes affected both chemoattractant-triggered inside-out as well as integrin-mediated outside-in signaling events, thus provoking defective adhesion and spreading on purified integrin ligands, respectively. (2) In in vitro cell-culture experiments, Ibrutinib promoted a differentiation shift of monocytes to fibrocyte-like cells, characterized by the acquisition of a typical elongated cell morphology. Importantly, this clear-cut shape transition also occurred upon culturing monocytes with sera derived from Ibrutinib-treated patients, thus clearly suggesting that the drug concentrations achievable in vivo can generate the phenotypic shift. (3) Ibrutinib-induced fibrocyte-like cells showed adhesion deficiency, altered phagocytic properties, and, with respect to macrophages, they acquired the capability of generating larger amounts of reactive oxygen species, possibly displaying different metabolic activities. Taken together, our results indicate that Ibrutinib has profound effects on the monocyte/macrophage immunobiology. They may finally shed some light about the biological ground of several Ibrutinib-related toxicities.
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α-Bisabolol (BSB) is a plant-derived sesquiterpene alcohol able to trigger regulated cell death in transformed cells, while deprived of the general toxicity in several mouse models. Here, we investigated the involvement of lysosomal and mitochondrial compartments in the cytotoxic effects of BSB, with a specific focus on the BH3-only activator protein BID. We found that BSB particularly accumulated in cancer cell lines, displaying a higher amount of lipid rafts as compared to normal blood cells. By means of western blotting and microscopy techniques, we documented rapid BSB-induced BID translocation to lysosomes and mitochondria, both of them becoming dysfunctional. Lysosomal membranes were permeabilized, thus blocking the cytoprotective autophagic flux and provoking cathepsin B leakage into the cytosol. Multiple flow cytometry-based experiments demonstrated the loss of mitochondrial membrane potential due to pore formation across the lipid bilayer. These parallel events converged on neoplastic cell death, an outcome significantly prevented by BID knockdown. Therefore, BSB promoted BID redistribution to the cell death executioner organelles, which in turn activated anti-autophagic and proapoptotic mechanisms. This is an example of how xenohormesis can be exploited to modulate basic cellular programs in cancer.
Asunto(s)
Apoptosis/efectos de los fármacos , Proteína Proapoptótica que Interacciona Mediante Dominios BH3/metabolismo , Lisosomas/metabolismo , Mitocondrias/metabolismo , Sesquiterpenos Monocíclicos/farmacología , Autofagia/efectos de los fármacos , Bencimidazoles/farmacología , Carbocianinas/farmacología , Ciclo Celular/efectos de los fármacos , Línea Celular , Gangliósido G(M1)/metabolismo , Técnicas de Silenciamiento del Gen , Humanos , Lisosomas/efectos de los fármacos , Microdominios de Membrana/metabolismo , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Proteínas de la Membrana/metabolismo , Mitocondrias/efectos de los fármacos , Transporte de Proteínas/efectos de los fármacosRESUMEN
CXCL12/CXCR4 axis relies on both heterotrimeric Gi protein and ß-arrestin coupling to trigger downstream responses. G protein activation allows for calcium flux, chemotaxis and early extracellular-signal regulated kinases 1/2 (ERK1/2) phosphorylation, whereas ß-arrestin recruitment leads to late signaling, receptor desensitization and internalization. Together they may regulate the balance between transactivation and transinhibition of epithelial growth factor receptor 1 (HER1). Since we have previously noted significant differences between CXCL12 and its structural variant [N33A]CXCL12 in CXCR4 signaling, we sought to better characterize them by performing cAMP inhibition and ß-arrestin recruitment assays, as well as functional tests that separately investigate G protein and ß-arrestin-induced responses. [N33A]CXCL12 showed reduced potency both in Gαi coupling and ß-arrestin recruitment as compared to the wild type chemokine, acting as an unbiased ligand. While these findings translated into reduced potency within Gαi-dependent functions, ß-arrestin-dependent modules were affected in a more peculiar way. Unlike CXCL12, the mutant analogue did not restore HB-EGF-stimulated HER1 from CXCR4-induced transinhibition, and did not trigger the late wave of ERK1/2 phosphorylation. Instead, CXCR4 internalization was not impaired upon [N33A]CXCL12 stimulation. These differences highlight the novel opportunity to dissect CXCL12 signaling within the ß-arrestin layer, in which the mutant chemokine clearly favors the internalization module over the other pathways. Such functional selectivity has an impact on HER1 activation status and may play a relevant part in the crosstalk between tyrosine kinase and seven transmembrane receptors.
RESUMEN
Bruton's tyrosine kinase (BTK) regulates the B-cell receptor (BCR) signaling pathway, which, in turn, plays a critical role in B-cell chronic lymphocytic leukemia (B-CLL) pathogenesis. The BTK-specific inhibitor Ibrutinib blocks BCR signaling and is now approved as effective B-CLL therapy. Chemokines, such as the homeostatic chemokine CXCL12, play a central role in B-CLL pathogenesis and progression, by regulating CLL cell interaction with the stromal microenvironment, leading to cells survival and proliferation. In this study, we investigated, in normal versus CLL B-lymphocytes, the role of BTK in signal transduction activated by the CXCL12-CXCR4 signaling axis and its involvement in rapid integrin activation. We show that BTK is rapidly activated by CXCL12 in healthy as well as CLL B-lymphocytes, with a kinetic of tyr-phosphorylation coherent with rapid adhesion triggering. BTK inhibition prevents CXCL12-induced triggering of lymphocyte function-associated antigen-1 (LFA-1) and very late antigen-4 (VLA-4) integrins. Furthermore, BTK inhibition blocks the activation of the small GTP-binding protein RhoA, controlling integrin affinity. Very importantly, we show that BTK tyr-phosphorylation and activation by CXCL12 depends on upstream activation of JAK2 tyrosine kinase. A comparative analysis of 36 B-CLL patients demonstrates that JAK2-dependent BTK regulatory role on integrin activation by CXCL12 is fully conserved in CLL cells. Finally, we show that the JAK2-BTK axis also regulates signaling to integrin activation by BCR. Thus, BTK and JAK protein tyrosine kinases (PTKs) manifest a hierarchical activity both in chemokine- as well as BCR-mediated integrin activation and dependent adhesion, potentially suggesting the possibility of combined therapeutic approaches to B-CLL treatment.