RESUMEN
CD74 is a cell-surface receptor for the cytokine macrophage migration inhibitory factor. Macrophage migration inhibitory factor binding to CD74 induces its intramembrane cleavage and the release of its cytosolic intracellular domain (CD74-ICD), which regulates cell survival. In the present study, we characterized the transcriptional activity of CD74-ICD in chronic lymphocytic B cells. We show that following CD74 activation, CD74-ICD interacts with the transcription factors RUNX (Runt related transcription factor) and NF-κB and binds to proximal and distal regulatory sites enriched for genes involved in apoptosis, immune response, and cell migration. This process leads to regulation of expression of these genes. Our results suggest that identifying targets of CD74 will help in understanding of essential pathways regulating B-cell survival in health and disease.
RESUMEN
The control of lymphoid homeostasis is the result of a very fine balance between lymphocyte production, proliferation, and apoptosis. In this study, we focused on the role of T cells in the maintenance/survival of the mature naive peripheral B cell population. We show that naive B and T cells interact via the signaling lymphocyte activation molecule (SLAM) family receptor, SLAMF6. This interaction induces cell type-specific signals in both cell types, mediated by the SLAM-associated protein (SAP) family of adaptors. This signaling results in an upregulation of the expression of the cytokine migration inhibitory factor in the T cells and augmented expression of its receptor CD74 on the B cell counterparts, consequently enhancing B cell survival. Furthermore, in X-linked lymphoproliferative disease patients, SAP deficiency reduces CD74 expression, resulting in the perturbation of B cell maintenance from the naive stage. Thus, naive T cells regulate B cell survival in a SLAMF6- and SAP-dependent manner.
Asunto(s)
Subgrupos de Linfocitos B/fisiología , Linfocitos B/fisiología , Células Sanguíneas/fisiología , Trastornos Linfoproliferativos/inmunología , Proteína Asociada a la Molécula de Señalización de la Activación Linfocitaria/metabolismo , Familia de Moléculas Señalizadoras de la Activación Linfocitaria/metabolismo , Linfocitos T/fisiología , Animales , Anticuerpos Bloqueadores/administración & dosificación , Comunicación Celular , Diferenciación Celular , Proliferación Celular , Supervivencia Celular , Células Cultivadas , Homeostasis , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , ARN Interferente Pequeño/genética , Proteína Asociada a la Molécula de Señalización de la Activación Linfocitaria/genética , Familia de Moléculas Señalizadoras de la Activación Linfocitaria/genéticaRESUMEN
Chemokines and chemokine receptors establish a complex network modulating immune cell migration and localization. These molecules were also suggested to mediate the differentiation of leukocytes; however, their intrinsic, direct regulation of lymphocyte fate remained unclear. CCR2 is the main chemokine receptor inducing macrophage and monocyte recruitment to sites of inflammation, and it is also expressed on T cells. To assess whether CCR2 directly regulates T cell responses, we followed the fates of CCR2-/- T cells in T cell-specific inflammatory models. Our in vitro and in vivo results show that CCR2 intrinsically mediates the expression of inflammatory T cell cytokines, and its absence on T cells results in attenuated colitis progression. Moreover, CCR2 deficiency in T cells promoted a program inducing the accumulation of Foxp3+ regulatory T cells, while decreasing the levels of Th17 cells in vivo, indicating that CCR2 regulates the immune response by modulating the effector/regulatory T ratio.
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Inmunidad Celular , Receptores CCR2/metabolismo , Linfocitos T Reguladores/inmunología , Animales , Movimiento Celular , Colitis/inmunología , Citocinas/genética , Citocinas/inmunología , Factores de Transcripción Forkhead/genética , Factores de Transcripción Forkhead/metabolismo , Regulación de la Expresión Génica , Macrófagos/inmunología , Ratones , Receptores CCR2/deficiencia , Receptores CCR2/genética , Receptores CCR2/inmunología , Receptores CCR5/inmunología , Receptores CCR5/metabolismo , Células Th17/inmunología , Células Th17/fisiologíaRESUMEN
BACKGROUND: Peroxisome proliferator-activated receptor gamma (PPARγ) agonists are clinically used to counteract hyperglycemia. However, so far experienced unwanted side effects, such as weight gain, promote the search for new PPARγ activators. METHODS: We used a combination of in silico, in vitro, cell-based and in vivo models to identify and validate natural products as promising leads for partial novel PPARγ agonists. RESULTS: The natural product honokiol from the traditional Chinese herbal drug Magnolia bark was in silico predicted to bind into the PPARγ ligand binding pocket as dimer. Honokiol indeed directly bound to purified PPARγ ligand-binding domain (LBD) and acted as partial agonist in a PPARγ-mediated luciferase reporter assay. Honokiol was then directly compared to the clinically used full agonist pioglitazone with regard to stimulation of glucose uptake in adipocytes as well as adipogenic differentiation in 3T3-L1 pre-adipocytes and mouse embryonic fibroblasts. While honokiol stimulated basal glucose uptake to a similar extent as pioglitazone, it did not induce adipogenesis in contrast to pioglitazone. In diabetic KKAy mice oral application of honokiol prevented hyperglycemia and suppressed weight gain. CONCLUSION: We identified honokiol as a partial non-adipogenic PPARγ agonist in vitro which prevented hyperglycemia and weight gain in vivo. GENERAL SIGNIFICANCE: This observed activity profile suggests honokiol as promising new pharmaceutical lead or dietary supplement to combat metabolic disease, and provides a molecular explanation for the use of Magnolia in traditional medicine.
Asunto(s)
Productos Biológicos/farmacología , Compuestos de Bifenilo/farmacología , Lignanos/farmacología , PPAR gamma/agonistas , Células 3T3-L1 , Tejido Adiposo/citología , Tejido Adiposo/efectos de los fármacos , Animales , Productos Biológicos/aislamiento & purificación , Compuestos de Bifenilo/aislamiento & purificación , Diferenciación Celular/efectos de los fármacos , Diabetes Mellitus Experimental/fisiopatología , Células HEK293 , Humanos , Lignanos/aislamiento & purificación , Ratones , Simulación del Acoplamiento MolecularRESUMEN
Myocardial injury may ultimately lead to adverse ventricular remodeling and development of heart failure (HF), which is a major cause of morbidity and mortality worldwide. Given the slow pace and substantial costs of developing new therapeutics, drug repurposing is an attractive alternative. Studies of many organs, including the heart, highlight the importance of the immune system in modulating injury and repair outcomes. Glatiramer acetate (GA) is an immunomodulatory drug prescribed for patients with multiple sclerosis. Here, we report that short-term GA treatment improves cardiac function and reduces scar area in a mouse model of acute myocardial infarction and a rat model of ischemic HF. We provide mechanistic evidence indicating that, in addition to its immunomodulatory functions, GA exerts beneficial pleiotropic effects, including cardiomyocyte protection and enhanced angiogenesis. Overall, these findings highlight the potential repurposing of GA as a future therapy for a myriad of heart diseases.
RESUMEN
Post-translational modification (PTM) of antigens provides an additional source of specificities targeted by immune responses to tumors or pathogens, but identifying antigen PTMs and assessing their role in shaping the immunopeptidome is challenging. Here we describe the Protein Modification Integrated Search Engine (PROMISE), an antigen discovery pipeline that enables the analysis of 29 different PTM combinations from multiple clinical cohorts and cell lines. We expanded the antigen landscape, uncovering human leukocyte antigen class I binding motifs defined by specific PTMs with haplotype-specific binding preferences and revealing disease-specific modified targets, including thousands of new cancer-specific antigens that can be shared between patients and across cancer types. Furthermore, we uncovered a subset of modified peptides that are specific to cancer tissue and driven by post-translational changes that occurred in the tumor proteome. Our findings highlight principles of PTM-driven antigenicity, which may have broad implications for T cell-mediated therapies in cancer and beyond.
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Neoplasias , Procesamiento Proteico-Postraduccional , Humanos , Procesamiento Proteico-Postraduccional/genética , Péptidos/genética , Antígenos , Antígenos de Histocompatibilidad Clase I/genética , Antígenos de Histocompatibilidad Clase I/metabolismo , Neoplasias/genéticaRESUMEN
Immunotherapy revolutionized treatment options in cancer, yet the mechanisms underlying resistance in many patients remain poorly understood. Cellular proteasomes have been implicated in modulating antitumor immunity by regulating antigen processing, antigen presentation, inflammatory signaling and immune cell activation. However, whether and how proteasome complex heterogeneity may affect tumor progression and the response to immunotherapy has not been systematically examined. Here, we show that proteasome complex composition varies substantially across cancers and impacts tumor-immune interactions and the tumor microenvironment. Through profiling of the degradation landscape of patient-derived non-small-cell lung carcinoma samples, we find that the proteasome regulator PSME4 is upregulated in tumors, alters proteasome activity, attenuates presented antigenic diversity and associates with lack of response to immunotherapy. Collectively, our approach affords a paradigm by which proteasome composition heterogeneity and function should be examined across cancer types and targeted in the context of precision oncology.
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Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Humanos , Presentación de Antígeno , Neoplasias Pulmonares/patología , Medicina de Precisión , Complejo de la Endopetidasa Proteasomal/metabolismo , Microambiente TumoralRESUMEN
B cells have essential functions in multiple sclerosis and in its mouse model, experimental autoimmune encephalomyelitis, both as drivers and suppressors of the disease. The suppressive effects are driven by a regulatory B cell (Breg) population that functions, primarily but not exclusively, via the production of IL-10. However, the mechanisms modulating IL-10-producing Breg abundance are poorly understood. Here we identify SLAMF5 for controlling IL-10+ Breg maintenance and function. In EAE, the deficiency of SLAMF5 in B cells causes accumulation of IL10+ Bregs in the central nervous system and periphery. Blocking SLAMF5 in vitro induces both human and mouse IL-10-producing Breg cells and increases their survival with a concomitant increase of a transcription factor, c-Maf. Finally, in vivo SLAMF5 blocking in EAE elevates IL-10+ Breg levels and ameliorates disease severity. Our results suggest that SLAMF5 is a negative moderator of IL-10+ Breg cells, and may serve as a therapeutic target in MS and other autoimmune diseases.
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Linfocitos B Reguladores/inmunología , Encefalomielitis Autoinmune Experimental/patología , Interleucina-10/inmunología , Familia de Moléculas Señalizadoras de la Activación Linfocitaria/metabolismo , Animales , Supervivencia Celular/inmunología , Células Cultivadas , Modelos Animales de Enfermedad , Encefalomielitis Autoinmune Experimental/inmunología , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Esclerosis Múltiple/inmunología , Familia de Moléculas Señalizadoras de la Activación Linfocitaria/antagonistas & inhibidores , Familia de Moléculas Señalizadoras de la Activación Linfocitaria/genéticaRESUMEN
Multiple myeloma (MM) is characterized by an accumulation of malignant plasma cells (PCs) within the BM. The BM microenvironment supports survival of the malignant cells and is composed of cellular fractions that foster myeloma development and progression by suppression of the immune response. Despite major progress in understanding the biology and pathophysiology of MM, this disease is still incurable and requires aggressive treatment with significant side effects. CD84 is a self-binding immunoreceptor belonging to the signaling lymphocyte activation molecule (SLAM) family. Previously, we showed that CD84 bridges between chronic lymphocytic leukemia cells and their microenvironment, and it regulates T cell function. In the current study, we investigated the role of CD84 in MM. Our results show that MM cells express low levels of CD84. However, these cells secrete the cytokine macrophage migration inhibitory factor (MIF), which induces CD84 expression on cells in their microenvironment. Its activation leads to an elevation of expression of genes regulating differentiation to monocytic/granulocytic-myeloid-derived suppressor cells (M-MDSCs and G-MDSCs, respectively) and upregulation of PD-L1 expression on MDSCs, which together suppress T cell function. Downregulation of CD84 or its blocking reduce MDSC accumulation, resulting in elevated T cell activity and reduced tumor load. Our data suggest that CD84 might serve as a novel therapeutic target in MM.
Asunto(s)
Mieloma Múltiple/inmunología , Familia de Moléculas Señalizadoras de la Activación Linfocitaria/genética , Familia de Moléculas Señalizadoras de la Activación Linfocitaria/inmunología , Familia de Moléculas Señalizadoras de la Activación Linfocitaria/metabolismo , Microambiente Tumoral/inmunología , Animales , Antígeno B7-H1 , Línea Celular Tumoral , Humanos , Inmunoterapia , Oxidorreductasas Intramoleculares/metabolismo , Leucemia Linfocítica Crónica de Células B/inmunología , Activación de Linfocitos , Factores Inhibidores de la Migración de Macrófagos/metabolismo , Ratones , Mieloma Múltiple/terapia , Células Supresoras de Origen Mieloide/inmunología , Linfocitos T/inmunologíaRESUMEN
The Golgi is a dynamic organelle whose correct assembly is crucial for cellular homeostasis. Perturbations in Golgi structure are associated with numerous disorders from neurodegeneration to cancer. However, whether and how dispersal of the Golgi apparatus is actively regulated under stress, and the consequences of Golgi dispersal, remain unknown. Here we demonstrate that 26S proteasomes are associated with the cytosolic surface of Golgi membranes to facilitate Golgi Apparatus-Related Degradation (GARD) and degradation of GM130 in response to Golgi stress. The degradation of GM130 is dependent on p97/VCP and 26S proteasomes, and required for Golgi dispersal. Finally, we show that perturbation of Golgi homeostasis induces cell death of multiple myeloma in vitro and in vivo, offering a therapeutic strategy for this malignancy. Taken together, this work reveals a mechanism of Golgi-localized proteasomal degradation, providing a functional link between proteostasis control and Golgi architecture, which may be critical in various secretion-related pathologies.
Asunto(s)
Aparato de Golgi/metabolismo , Ionóforos/uso terapéutico , Mieloma Múltiple/tratamiento farmacológico , Complejo de la Endopetidasa Proteasomal/metabolismo , Proteostasis/fisiología , Animales , Apoptosis/efectos de los fármacos , Autoantígenos/metabolismo , Línea Celular Tumoral/trasplante , Modelos Animales de Enfermedad , Aparato de Golgi/efectos de los fármacos , Células HEK293 , Humanos , Membranas Intracelulares/metabolismo , Ionóforos/farmacología , Proteínas de la Membrana/metabolismo , Ratones , Monensina/farmacología , Monensina/uso terapéutico , Mieloma Múltiple/patología , Proteolisis/efectos de los fármacos , Proteostasis/efectos de los fármacos , Ubiquitinación/efectos de los fármacos , Proteína que Contiene Valosina/metabolismoRESUMEN
Data on the biological effects of some overexpressed oncogenes and their cooperation with cellular factors are, at least partially, contradictory. There are reports on the strong pro-apoptotic action of temperature-sensitive (ts) p53(135val) in transformed cells at permissive temperature. However, in our experience very high levels of p53(135val) induce in transformed rat cells at permissive temperature cell cycle arrest but not apoptosis. Comparison of the experimental protocols reveals that cells used for transfection strongly differ. Therefore, we decided to explore the impact of primary cells used for generation of cell clones on the biological effects evoked by p53 and c-Ha-Ras. In the present study, we used primary rat cells (RECs) isolated from rat embryos of different age: at 13.5 gd (y) and 15.5 gd (o). We immortalized rat cells using ts p53(135val) mutant and additionally generated transformed cells after co-transfection with oncogenic Ras. The RECs were transfected with a constitutively activated Ha-Ras protein, a mutation that is found in a wide variety of human tumors. The ts p53(135Val) mutant, switching between wild-type (wt) and mutant conformation, offers the possibility to study the escape from p53-mediated cell cycle control in a model of malignant transformation in cells with the same genetic background. Surprisingly, the kinetics of cell proliferation at non-permissive temperature and that of cell cycle arrest at 32 degrees C strongly differed between cell clones established from yRECs and oRECs, thereby indicating that overexpression of genes such as ts p53(135Val) mutant and oncogenic-Ha-Ras does not fully override the intrinsic cellular program.
Asunto(s)
Ciclo Celular/fisiología , Ambiente , Proteínas Proto-Oncogénicas p21(ras)/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Animales , Línea Celular Transformada , Proliferación Celular , Células Cultivadas , Embrión de Mamíferos/citología , Embrión de Mamíferos/fisiología , Femenino , Edad Gestacional , Humanos , Ratones , Embarazo , Proteínas Proto-Oncogénicas p21(ras)/genética , Ratas , Temperatura , Proteína p53 Supresora de Tumor/genéticaRESUMEN
Antigen presentation on HLA molecules is a major mechanism by which the immune system monitors self and non-self-recognition. Importantly, HLA-I presentation has gained much attention through its role in eliciting anti-tumor immunity. Several determinants controlling the peptides presented on HLA have been uncovered, mainly through the study of model substrates and large-scale immunopeptidome analyses. These determinants include the relative abundances of proteins in the cell, the stability or turnover rate of these proteins and the binding affinities of a given peptide to the HLA haplotypes found in a cell. However, the regulatory principles involved in selection and regulation of specific antigens in response to tumor pro-inflammatory signals remain largely unknown. Here, we chose to examine the effect that TNFα and IFNγ stimulation may exert on the immunopeptidome landscape of lung cancer cells. We show that the expression of many of the proteins involved in the class I antigen presentation pathway are changed by pro-inflammatory cytokines. Further, we could show that increased expression of the HLA-B allomorph drives a significant change in HLA-bound antigens, independently of the significant changes observed in the cellular proteome. Finally, we observed increased HLA-B levels in correlation with tumor infiltration across the TCGA lung cancer cohorts. Taken together, our results suggest that the immunopeptidome landscape should be examined in the context of anti-tumor immunity whereby signals in the microenvironment may be critical in shaping and modulating this important aspect of host-tumor interactions.
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Antígenos HLA-B/inmunología , Interferón gamma/farmacología , Neoplasias Pulmonares/inmunología , Péptidos/inmunología , Factor de Necrosis Tumoral alfa/farmacología , Células A549 , Humanos , ProteomaRESUMEN
Activating mutations of Ras that frequently occur during malignant transformation, enhance growth-promoting signal transduction, allowing cells to bypass stringent control of cell cycle progression, thereby rendering them highly proliferative. Abundantly expressed c-Ha-ras protein in human cervical HeLa cells is farnesylated and attached to the plasma membrane, inducing enhanced signal transduction. Exposure of HeLa cells to cisplatin very efficiently inhibits cell proliferation and induces apoptosis. Unfortunately, high doses of cisplatin are strongly cytotoxic, therefore, an alternative therapeutic strategy allowing dose reduction of cisplatin by inhibition of farnesylation could increase the curative effects of cisplatin, thereby benefiting cancer patients. We used two inhibitors of farnesyl protein transferase (FPTase), FTI, and L-744,832, to sensitize HeLa cells to the action of cisplatin. The combined administration of cisplatin and inhibitors of FPTase increased the cytostatic potency of cisplatin. L-744,832 exhibited a stronger synergistic effect in combination with cisplatin than FTI. Moreover, the efficiency of the combined therapy strongly depended on the treatment regimen: The highest efficiency was achieved after combined treatment for 24 h and post-incubation with an inhibitor of FPTase for 48 h. Following this optimized treatment, apoptosis was induced in approximately 50% of HeLa cells treated with 1 microM cisplatin, representing approximately a threefold increase as compared to cisplatin monotherapy. Combined treatment of HeLa cells with cisplatin and inhibitors of FPTase significantly increases the efficacy of the therapy and allows to reduce the dose of cisplatin. Importantly, best therapeutic effects can be achieved by post-treatment with inhibitors of FPTase.
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Transferasas Alquil y Aril/antagonistas & inhibidores , Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Cisplatino/farmacología , Metionina/análogos & derivados , Apoptosis/efectos de los fármacos , Sinergismo Farmacológico , Quimioterapia Combinada , Femenino , Células HeLa , Humanos , Metionina/farmacología , Proteínas Proto-Oncogénicas p21(ras) , Neoplasias del Cuello Uterino/tratamiento farmacológico , Neoplasias del Cuello Uterino/patologíaRESUMEN
It is generally accepted that exposure of cells to a variety of DNA-damaging agents leads to up-regulation and activation of wild-type (wt) p53 protein. We investigated the (re)-activation of p53 protein in two human cancer cell lines in which the gene for this tumor suppressor is not mutated: HeLaS(3) cervix carcinoma and MCF-7 breast cancer cells, by induction via different genotoxic and cytotoxic stimuli. Treatment of human cells with the alkylating agent N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) or different anti-cancer drugs resulted in a strong DNA damage as evidenced by Comet assay and a marked increase in site-specific phosphorylation of H2AX. Unlike in MCF-7 cells, in HeLaS(3) cells the expression of p53 protein did not increase after MNNG treatment despite a strong DNA damage. However, other agents for example doxorubicin markedly induced p53 response in HeLaS(3) cells. After exposure of these cells to MNNG, the ATM-dependent effector proteins Chk2 and NBS1 were phosphorylated, thereby evidencing that MNNG-induced DNA breakage was recognized and properly signaled. In HeLaS(3) cells wt p53 protein is not functional due to E6-mediated targeting for accelerated ubiquitylation and degradation. Therefore, the activation of a p53 response to genotoxic stress in HeLaS(3) cells seems to depend on the status of E6 oncoprotein. Indeed, the induction of p53 protein in HeLaS(3) cells in response to distinct agents inversely correlates with the cellular level of E6 oncoprotein. This implicates that the capability of different agents to activate p53 in HeLaS(3) cells primarily depends on their inhibitory effect on expression of E6 oncoprotein.
Asunto(s)
Daño del ADN , Proteínas Oncogénicas Virales/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Antibióticos Antineoplásicos/farmacología , Proteínas de Ciclo Celular/metabolismo , Quinasa de Punto de Control 2 , Daño del ADN/efectos de los fármacos , Doxorrubicina/farmacología , Células HeLa , Humanos , Metilnitronitrosoguanidina/farmacología , Proteínas Nucleares/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Transducción de Señal/efectos de los fármacos , Ubiquitinación/efectos de los fármacos , Regulación hacia Arriba/efectos de los fármacosRESUMEN
Exposure of human HeLaS(3) cervix carcinoma cells to high doses of conventional cytostatic drugs, e.g. cisplatin (CP) strongly inhibits their proliferation. However, most cytostatic agents are genotoxic and may generate a secondary malignancy. Therefore, therapeutic strategy using alternative, not cytotoxic drugs would be beneficial. Inhibition of cyclin-dependent kinases (CDKs) by pharmacological inhibitors became recently a promising therapeutic option. Roscovitine (ROSC), a selective CDK inhibitor, efficiently targets human malignant cells. ROSC induces cell cycle arrest and apoptosis in human MCF-7 breast cancer cells. ROSC also activates p53 protein. Activation of p53 tumor suppressor protein is essential for induction of apoptosis in MCF-7 cells. Considering the fact that in HeLaS(3) cells wt p53 is inactivated by the action of HPV-encoded E6 oncoprotein, we addressed the question whether ROSC would be able to reactivate p53 protein in them. Their exposure to ROSC for 24 h induced cell cycle arrest at G(2)/M and reduced the number of viable cells. Unlike CP, ROSC in the used doses did not induce DNA damage and was not directly cytotoxic. Despite lack of detectable DNA lesions, ROSC activated wt p53 protein. The increase of p53 levels was attributable to the ROSC-mediated protein stabilization. Further analyses revealed that ROSC induced site-specific phosphorylation of p53 protein at Ser46. After longer exposure, ROSC induced apoptosis in HeLaS(3) cells. These results indicate that therapy of HeLaS(3) cells by ROSC could offer an advantage over that by CP due to its increased selectivity and markedly reduced risk of generation of a secondary cancer.
Asunto(s)
Antineoplásicos/farmacología , Apoptosis , Carcinoma/tratamiento farmacológico , Purinas/farmacología , Proteína p53 Supresora de Tumor/genética , Neoplasias del Cuello Uterino/tratamiento farmacológico , Antineoplásicos/uso terapéutico , Carcinoma/metabolismo , Ciclo Celular , Línea Celular Tumoral , Supervivencia Celular , Femenino , Células HeLa , Humanos , Roscovitina , Proteína p53 Supresora de Tumor/metabolismo , Neoplasias del Cuello Uterino/metabolismoRESUMEN
Human MCF-7 breast cancer cells are relatively resistant to anti-cancer drugs. Recently, we reported that roscovitine (ROSC), a selective cyclin-dependent kinase (CDK) inhibitor, arrested human MCF-7 breast cancer cells in G2 phase of the cell cycle and concomitantly induced apoptosis. Moreover, we observed that the effect of the CDK inhibitor was dependent on the content of the culture medium. The cell cycle inhibiting action of ROSC was markedly diminished in human MCF-7 cells cultivated in medium supplemented with phenol red. These observations indicated that the therapeutic effects of ROSC can be affected by the components of the tissue medium. Recently, a number of epidemiological and experimental studies indicated that polyphenols (e.g. resveratrol, epicatechins etc.), abundant micronutrients in food, are anti-oxidant agents and could have strong anti-mitotic as well as pro-apoptotic activities. In the present contribution we raised the question whether the ROSC-mediated cell cycle arrest could be additionally modulated by compounds of natural origin, especially by polyphenols. Considering the potential benefits of the dietary components during the post-chemotherapy period, we focused our attention on the effects of resveratrol administration after treatment with ROSC. We analyzed whether the combined treatment with resveratrol would exert any additional effect on the cell cycle status of ROSC-treated human cancer cells. Resveratrol exhibited low direct cytotoxicity. The combined treatment with ROSC enhanced the ROSC-mediated inhibition of cell proliferation and cell cycle arrest. These results indicate that targeted combination of anti-cancer drugs with distinct naturally occurring compounds could increase the efficacy of the therapy and concomitantly reduce the undesired side effects exerted by cytostatic drugs.
Asunto(s)
Antineoplásicos/farmacología , Antioxidantes/farmacología , Ciclo Celular/efectos de los fármacos , Purinas/farmacología , Estilbenos/farmacología , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/patología , Recuento de Células , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , ADN/biosíntesis , ADN/genética , Femenino , Citometría de Flujo , Humanos , Resveratrol , RoscovitinaRESUMEN
Chronic lymphocytic leukemia (CLL) is characterized by clonal proliferation and progressive accumulation of mature B lymphocytes in the peripheral blood, lymphoid tissues, and bone marrow. CLL is characterized by profound immune defects leading to severe infectious complications. T cells are numerically, phenotypically, and functionally highly abnormal in CLL, with only limited ability to exert antitumor immune responses. Exhaustion of T cells has also been suggested to play an important role in antitumor responses. CLL-mediated T cell exhaustion is achieved by the aberrant expression of several inhibitory molecules on CLL cells and their microenvironment, prominently the programmed cell death ligand 1/programmed cell death 1 (PD-L1/PD-1) receptors. Previously, we showed that CD84, a member of the SLAM family of receptors, bridges between CLL cells and their microenvironment. In the current study, we followed CD84 regulation of T cell function. We showed that cell-cell interaction mediated through human and mouse CD84 upregulates PD-L1 expression on CLL cells and in their microenvironment and PD-1 expression on T cells. This resulted in suppression of T cell responses and activity in vitro and in vivo. Thus, our results demonstrate a role for CD84 in the regulation of immune checkpoints by leukemia cells and identify CD84 blockade as a therapeutic strategy to reverse tumor-induced immune suppression.
Asunto(s)
Antígeno B7-H1/inmunología , Regulación Leucémica de la Expresión Génica/inmunología , Leucemia Linfocítica Crónica de Células B/inmunología , Proteínas de Neoplasias/inmunología , Receptor de Muerte Celular Programada 1/inmunología , Familia de Moléculas Señalizadoras de la Activación Linfocitaria/inmunología , Animales , Antígeno B7-H1/genética , Regulación Leucémica de la Expresión Génica/genética , Humanos , Leucemia Linfocítica Crónica de Células B/genética , Leucemia Linfocítica Crónica de Células B/patología , Ratones , Ratones Noqueados , Proteínas de Neoplasias/genética , Receptor de Muerte Celular Programada 1/genética , Familia de Moléculas Señalizadoras de la Activación Linfocitaria/genéticaRESUMEN
Cellular function is critically regulated through degradation of substrates by the proteasome. To enable direct analysis of naturally cleaved proteasomal peptides under physiological conditions, we developed mass spectrometry analysis of proteolytic peptides (MAPP), a method for proteasomal footprinting that allows for capture, isolation and analysis of proteasome-cleaved peptides. Application of MAPP to cancer cell lines as well as primary immune cells revealed dynamic modulation of the cellular degradome in response to various stimuli, such as proinflammatory signals. Further, we performed analysis of minute amounts of clinical samples by studying cells from the peripheral blood of patients with systemic lupus erythematosus (SLE). We found increased degradation of histones in patient immune cells, thereby suggesting a role of aberrant proteasomal degradation in the pathophysiology of SLE. Thus, MAPP offers a broadly applicable method to facilitate the study of the cellular-degradation landscape in various cellular conditions and diseases involving changes in proteasomal degradation, including protein aggregation diseases, autoimmunity and cancer.
RESUMEN
The naturally occurring triterpenoid betulinic acid (BA) shows pronounced polypharmacology ranging from anti-inflammatory to anti-lipogenic activities. Recent evidence suggests that rather diverse cellular signaling events may be attributed to the same common upstream switch in cellular metabolism. In this study we therefore examined the metabolic changes induced by BA (10 µM) administration, with focus on cellular glucose metabolism. We demonstrate that BA elevates the rates of cellular glucose uptake and aerobic glycolysis in mouse embryonic fibroblasts with concomitant reduction of glucose oxidation. Without eliciting signs of obvious cell death BA leads to compromised mitochondrial function, increased expression of mitochondrial uncoupling proteins (UCP) 1 and 2, and liver kinase B1 (LKB1)-dependent activation AMP-activated protein kinase. AMPK activation accounts for the increased glucose uptake and glycolysis which in turn are indispensable for cell viability upon BA treatment. Overall, we show for the first time a significant impact of BA on cellular bioenergetics which may be a central mediator of the pleiotropic actions of BA.