RESUMO
Tumor growth and its chemotherapeutic regimens manifest myelosuppression, which is one of the possible causes underlying the limited success of immunotherapeutic anticancer strategies. Hence, approaches are being designed to develop safer therapeutic regimens that may have minimal damaging action on the process of myelopoiesis. 3-Bromopyruvate (3-BP) is a highly potent antimetabolic agent displaying a broad spectrum antineoplastic activity. However, 3-BP has not been investigated for its effect on the process of myelopoiesis in a tumor-bearing host. Hence, in this investigation, we studied the myelopoietic effect of in vivo administration of 3-BP to a murine host bearing a progressively growing ascitic thymoma designated as Dalton's lymphoma (DL). 3-BP administration to the DL-bearing mice resulted in a myelopotentiating action, reflected by an elevated count of bone marrow cells (BMC) accompanied by augmented proliferative ability and a declined induction of apoptosis. The BMC of 3-BP-administered mice displayed enhanced responsiveness to macrophage colony-stimulating factor for colony-forming ability of myeloid lineage along with an enhanced differentiation of F4/80+ bone marrow-derived macrophages (BMDM). BMDM differentiated from the BMC of 3-BP-administered DL-bearing mice showed an augmented response to lipopolysaccharide and interferon-γ for activation, displaying an augmented tumor cytotoxicity, expression of cytokines, reactive oxygen species, nitric oxide, CD11c, TLR-4, and HSP70. These features are indicative of the differentiation of M1 subtype of macrophages. Thus, this study demonstrates the myelopotentiating action of 3-BP, indicating its hematopoietic safety and potential for reinforcing the differentiation of macrophages in a tumor-bearing host.
Assuntos
Antimetabólitos Antineoplásicos/uso terapêutico , Piruvatos/uso terapêutico , Timoma/tratamento farmacológico , Neoplasias do Timo/tratamento farmacológico , Animais , Ascite , Células da Medula Óssea , Diferenciação Celular , Proliferação de Células , Macrófagos , Camundongos Endogâmicos BALB C , Mielopoese/efeitos dos fármacos , Complexo Piruvato Desidrogenase , Timoma/sangue , Neoplasias do Timo/sangueRESUMO
Diclofenac is a potent NSAID of clinical choice, which is widely used for containing inflammation. Moreover, recent experimental evidences overwhelmingly substantiate its antineoplastic potential. However, the precise molecular mechanisms of diclofenac's anticancer activity remain poorly understood. Neoplastic cells display reprogrammed metabolic features, which are manifested and regulated by a complex networking of molecular pathways. However, the effect of diclofenac on tumor cell metabolism are not yet clearly deciphered. Hence, the present investigation was carried out to identify and characterize key diclofenac targets of tumor metabolism, cell survival and chemoresistance. The interactions of diclofenac with such targets was analysed by PatchDock and YASARA (Yet Another Scientific Artificial Reality Application). The docking ability of diclofenac with its targets was based on analysis of dissociation constant (Kd), geometric shape complementarity score (GSC score), approximate interface area (AI area) and binding energy. The findings of this investigation reveal that diclofenac is capable of interacting with all of the selected molecular targets. Prominent interactions were observed with GLUT1, MCT4, LDH A, COX1, COX2, BCRP/ABCG2, HDM2/MDM2 and MRP1 compared to other targets. Interactions were of noncovalent nature involving ionic, hydrophobic interactions, Van der Waals forces and H-bonds, which varied depending on targets. This study for the first time, characterizes the nature of molecular interactions of diclofenac with selected targets involved in cancer cell metabolism, pH homeostasis, chemosensitivity, cell signalling and inflammation. Hence, these findings will be highly beneficial in optimizing the utility of diclofenac in development of novel cancer therapeutics.
Assuntos
Diclofenaco/metabolismo , Simulação de Acoplamento Molecular , Proteínas de Neoplasias/metabolismo , Neoplasias/metabolismo , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/química , Membro 2 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/metabolismo , Anti-Inflamatórios não Esteroides/química , Anti-Inflamatórios não Esteroides/metabolismo , Anti-Inflamatórios não Esteroides/farmacologia , Ciclo-Oxigenase 1/química , Ciclo-Oxigenase 1/metabolismo , Diclofenaco/química , Diclofenaco/farmacologia , Transportador de Glucose Tipo 1/química , Transportador de Glucose Tipo 1/metabolismo , Humanos , Cinética , Estrutura Molecular , Transportadores de Ácidos Monocarboxílicos/química , Transportadores de Ácidos Monocarboxílicos/metabolismo , Proteínas Musculares/química , Proteínas Musculares/metabolismo , Proteínas de Neoplasias/química , Neoplasias/tratamento farmacológico , Ligação Proteica , Conformação Proteica , Proteínas Proto-Oncogênicas c-mdm2/química , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Transdução de Sinais/efeitos dos fármacosRESUMO
Evidences demonstrate that metabolic inhibitor 3-bromopyruvate (3-BP) exerts a potent antitumor action against a wide range of malignancies. However, the effect of 3-BP on progression of the tumors of thymic origin remains unexplored. Although, constituents of tumor microenvironment (TME) plays a pivotal role in regulation of tumor progression, it remains unclear if 3-BP can alter the composition of the crucial tumor growth regulatory components of the external surrounding of tumor cells. Thus, the present investigation attempts to understand the effect of 3-BP administration to a host bearing a progressively growing tumor of thymic origin on tumor growth regulatory soluble, cellular and biophysical components of tumor milieu vis-à-vis understanding its association with tumor progression, accompanying cell cycle events and mode of cell death. Further, the expression of cell survival regulatory molecules and hemodynamic characteristics of the tumor milieu were analysed to decipher mechanisms underlying the antitumor action of 3-BP. Administration of 3-BP to tumor-bearing hosts retarded tumor progression accompanied by induction of tumor cell death, cell cycle arrest, declined metabolism, inhibited mitochondrial membrane potential, elevated release of cytochrome c and altered hemodynamics. Moreover, 3-BP reconstituted the external milieu, in concurrence with deregulated glucose and pH homeostasis and increased tumor infiltration by NK cells, macrophages, and T lymphocytes. Further, 3-BP administration altered the expression of key regulatory molecules involved in glucose uptake, intracellular pH and tumor cell survival. The outcomes of this study will help in optimizing the therapeutic application of 3-BP by targeting crucial tumor growth regulatory components of tumor milieu.
Assuntos
Antineoplásicos/farmacologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Piruvatos/farmacologia , Carga Tumoral/efeitos dos fármacos , Microambiente Tumoral/efeitos dos fármacos , Animais , Pontos de Checagem do Ciclo Celular/fisiologia , Morte Celular/efeitos dos fármacos , Morte Celular/fisiologia , Inibidores Enzimáticos/farmacologia , Camundongos , Camundongos Endogâmicos BALB C , Mitocôndrias/metabolismo , Mitocôndrias/patologia , Carga Tumoral/fisiologia , Microambiente Tumoral/fisiologiaRESUMO
Tuberculosis, one of the leading cause of death from infectious diseases, is caused by Mycobacterium tuberculosis. The genome of M. tuberculosis has been sequenced and nearly 40% of the whole genome sequence was categorized as hypothetical. Rv0774c was annotated as membrane exported hypothetical protein in TB database. In silico analysis revealed that Rv0774c is a paralog of PE-PGRS multi gene family with 100 aa N-terminal domain similar to PE domain of PE-PGRS proteins. Its C-terminal domain is quite different from PGRS domain, having characteristic lipase signature GXSXG & HG and catalytic residues predicted for lipolytic activity. Therefore, DNA coding for Rv0774c (303 aa), its N-terminal (1-100 aa) and C- terminal domain (100-303 aa) were separately cloned from M. tuberculosis and were over expressed in E. coli. Rv0774c gene and its C-terminal lipolytic domain preferably hydrolyzed short chain esters. Though no enzyme activity was observed in N-terminus PE like domain, it was demonstrated to enhance the thermostability of full length Rv0774c. Tetrahydrolipstatin inhibited the enzyme activity and predicted catalytic residues (Ser-185, Asp-255 and His-281) were confirmed by site directed mutagenesis. Rv0774c was secreted out in culture media by M. tuberculosis and was up-regulated in iron limiting conditions. Treatment of THP-1 cells with rRv0774c resulted in a decline in the LPS induced production of NO and expression of iNOS. rRv0774c treated THP-1 cells also showed an enhanced expression of IL-10 and TLR2. On contrary, it suppressed the LPS induced production of IL-12, chemokines MCP-1 and IL-8. Rv0774c inhibited the LPS induced phosphorylation of p38. These observations suggested that Rv0774c could modulate the pro-inflammatory immune response to support intracellular survival of the mycobacterium.
Assuntos
Proteínas de Bactérias/metabolismo , Citocinas/antagonistas & inibidores , Esterases/metabolismo , Terapia de Imunossupressão , Mycobacterium tuberculosis/enzimologia , Mycobacterium tuberculosis/fisiologia , Receptor 2 Toll-Like/antagonistas & inibidores , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Domínio Catalítico , Linhagem Celular , Clonagem Molecular , Análise Mutacional de DNA , Estabilidade Enzimática , Escherichia coli/genética , Escherichia coli/metabolismo , Esterases/química , Esterases/genética , Expressão Gênica , Interações Hospedeiro-Patógeno , Humanos , Lipopolissacarídeos/metabolismo , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Mutagênese Sítio-Dirigida , Especificidade por Substrato , TemperaturaRESUMO
Etiological role of viral proteins E6 and E7 of high-risk HPV in cervical carcinogenesis is well established. However, their contribution in chemoresistance and epithelial-mesenchymal transition (EMT) that leads to advanced metastatic lesions and chemoresistance is poorly defined. In the present study, contribution of viral oncoproteins in acquisition of EMT character during onset of chemoresistance was assessed. A chemoresistant cell line (SiHaCR) was developed from an established HPV16-positive cervical cancer cell line, SiHa, by escalating selection pressure of 5-fluorouracil (5-FU). Expression of Survivin, ABCG2, Snail, Slug, Twist, and Vimentin was examined in SiHa and SiHaCR cells by reverse transcriptase-PCR (RT-PCR) and immunoblotting assays. Mesenchymal phenotype in SiHaCR cells was confirmed by assessment of migration and invasion potentials. SiHaCR cells displayed elevated level of functional and molecular markers associated with chemoresistance (Survivin, ABCG2) and EMT (Snail, Slug, Twist, Vimentin) and reduced E-cadherin. SiHaCR also showed increased levels of HPV16 E6 and E7 transcripts. Specific silencing of HPV16 E6, but not E7 using corresponding siRNA, demonstrated a differential involvement of HPV oncogenes in manifestation of EMT. HPV16 E6 silencing resulted in reduction of Slug and Twist expression. However, the expression of Snail and Vimentin was only marginally affected. In contrast, there was an increase in the expression of E-cadherin. A reduced migration and invasion capabilities were observed only in E6-silenced SiHaCR cells, which further confirmed functional contribution of HPV16 E6 in manifestation of EMT. Taken together, our study demonstrated an active involvement of HPV16 E6 in regulation of EMT, which promotes chemoresistance in cervical cancer.
Assuntos
Resistencia a Medicamentos Antineoplásicos , Transição Epitelial-Mesenquimal , Fluoruracila/farmacologia , Regulação Neoplásica da Expressão Gênica , Proteínas Oncogênicas Virais/metabolismo , Proteínas E7 de Papillomavirus/metabolismo , Proteínas Repressoras/metabolismo , Neoplasias do Colo do Útero/patologia , Antimetabólitos Antineoplásicos/farmacologia , Apoptose , Western Blotting , Movimento Celular , Proliferação de Células , Feminino , Humanos , Técnicas Imunoenzimáticas , Proteínas Oncogênicas Virais/antagonistas & inibidores , Proteínas Oncogênicas Virais/genética , Proteínas E7 de Papillomavirus/antagonistas & inibidores , Proteínas E7 de Papillomavirus/genética , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Proteínas Repressoras/antagonistas & inibidores , Proteínas Repressoras/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células Tumorais Cultivadas , Neoplasias do Colo do Útero/genética , Neoplasias do Colo do Útero/virologia , CicatrizaçãoRESUMO
BACKGROUND: Orlistat, a fatty acid synthase (FASN) inhibitor, has been demonstrated to inhibit tumor cell survival. However, the mechanism(s) of its tumor growth retarding action against malignancies of hematological origin remains unclear. It is also not understood if the antitumor action of orlistat implicates modulated susceptibility of tumor cell to anticancer drugs. Therefore, the present investigation focuses to study the antitumor and chemosensitizing action of orlistat in a murine host bearing a progressively growing T cell lymphoma. METHODS: Tumor-bearing mice were administered with vehicle alone or containing orlistat followed by administration of PBS with or without cisplatin. Tumor progression and survival of tumor-bearing host were monitored along with analysis of tumor cell survival and apoptosis. Tumor ascitic fluid was examined for pH, NO and cytokines. Expression of genes and proteins was investigated by RT-PCR and western blot respectively. ROS was analyzed by DCFDA staining and FASN activity by spectrophotometry. RESULTS: Orlistat administration to tumor-bearing mice resulted in tumor growth retardation, prolonged life span, declined tumor cell survival and chemosensitization to cisplatin. It was accompanied by increased osmotic fragility, modulated acidosis, expression of ROS, NO, cytokines, MCT-1 and VH(+) ATPase, Bcl2, Caspase-3, P53, inhibited FASN activity and declined expression of MDR and MRP-1 proteins. CONCLUSION: Orlistat manifests antitumor and chemosensitizing action implicating modulated regulation of cell survival, reconstituted-tumor microenvironment and altered MDR phenotype. GENERAL SIGNIFICANCE: These observations indicate that orlistat could be utilized as an adjunct regimen for improving antitumor efficacy of cisplatin.
Assuntos
Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Ácido Graxo Sintases/antagonistas & inibidores , Lactonas/farmacologia , Linfoma de Células T/prevenção & controle , Microambiente Tumoral/efeitos dos fármacos , Animais , Fármacos Antiobesidade/farmacologia , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Western Blotting , Proliferação de Células/efeitos dos fármacos , Cisplatino/farmacologia , Modelos Animais de Doenças , Quimioterapia Combinada , Ensaio de Imunoadsorção Enzimática , Ácido Graxo Sintases/genética , Ácido Graxo Sintases/metabolismo , Citometria de Fluxo , Linfoma de Células T/metabolismo , Linfoma de Células T/patologia , Camundongos , Camundongos Endogâmicos BALB C , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/metabolismo , Nitritos/metabolismo , Orlistate , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Bicarbonate transporter (BCT) plays a crucial role in maintaining pH homeostasis of tumor cells by import of HCO3(-). This helps the tumor cells in manifesting extracellular tumor acidosis, accompanied by a relative intracellular alkalinization, which in turn promotes tumor progression. Therefore, blocking BCT-mediated HCO3(-) transport is envisaged as a promising anticancer therapeutic approach. Thus, using a murine model of a T cell lymphoma, designated as Dalton's lymphoma (DL), in the present in vitro investigation the antitumor consequences of blocking BCT function by its inhibitor 4-acetamido-4-isothiocyanostilbene-2,2-disulfonate (SITS) were explored. Treatment of DL cells with SITS resulted in an increase in the extracellular pH, associated with a decline in DL cell survival and augmented induction of apoptosis. BCT inhibition also elevated the expression of cytochrome c, caspase-9, caspase-3, Bax, reactive oxygen species, and nitric oxide along with inhibition of HSP-70 and Bcl2, which regulate tumor cell survival and apoptosis. SITS-treated DL cells displayed upregulated production of IFN-γ and IL-6 along with a decline of IL-10. Treatment of DL cells with SITS also inhibited the expression of fatty acid synthase, which is crucial for membrane biogenesis in neoplastic cells. The expression of lactate transporter MCT-1 and multidrug resistance regulating protein MRP-1 got inhibited along with hampered uptake of glucose and lactate production in SITS-treated DL cells. Thus, the declined tumor cell survival following inhibition of BCT could be the consequence of interplay of several inter-connected regulatory molecular events. The outcome of this study indicates the potential of BCT inhibition as a novel therapeutic approach for treatment of hematological malignancies.
Assuntos
Ácido 4-Acetamido-4'-isotiocianatostilbeno-2,2'-dissulfônico/farmacologia , Proteínas de Transporte de Ânions/antagonistas & inibidores , Apoptose/efeitos dos fármacos , Homeostase/efeitos dos fármacos , Linfoma de Células T/tratamento farmacológico , Proteínas de Neoplasias/metabolismo , Neoplasias Experimentais/tratamento farmacológico , Simportadores/antagonistas & inibidores , Animais , Proteínas de Transporte de Ânions/genética , Proteínas de Transporte de Ânions/metabolismo , Apoptose/genética , Proteínas Reguladoras de Apoptose/genética , Proteínas Reguladoras de Apoptose/metabolismo , Bicarbonatos/metabolismo , Citocinas/genética , Citocinas/metabolismo , Homeostase/genética , Concentração de Íons de Hidrogênio , Linfoma de Células T/genética , Linfoma de Células T/metabolismo , Linfoma de Células T/patologia , Camundongos , Camundongos Endogâmicos BALB C , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/genética , Neoplasias Experimentais/genética , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Simportadores/genética , Simportadores/metabolismoRESUMO
Transcription Termination Factor 1 (TTF1) is a multifunctional mammalian protein with vital roles in various cellular processes, including Pol I-mediated transcription initiation and termination, pre-rRNA processing, chromatin remodelling, DNA damage repair, and polar replication fork arrest. It comprises of two distinct functional regions; the N-terminal regulatory region (1-445 aa), and the C-terminal catalytic region (445-859 aa). The Myb domain located at the C-terminal region is a conserved DNA binding domain spanning from 550 to 732 aa (183 residues). Despite its critical role in various cellular processes, the physical structure of TTF1 remains unsolved. Attempts to purify the functional TTF1 protein have been unsuccessful till date. Therefore, we focused on characterizing the Myb domain of this essential protein. We started with predicting a 3-D model of the Myb domain using homology modelling, and ab-initio method. We then determined its stability through MD simulation in an explicit solvent. The model predicted is highly stable, which stabilizes at 200ns. To experimentally validate the computational model, we cloned and expressed the codon optimized Myb domain into a bacterial expression vector and purified the protein to homogeneity. Further, characterization of the protein shows that, Myb domain is predominantly helical (65%) and is alone sufficient to bind the Sal Box DNA. This is the first-ever study to report a complete in-silico model of the Myb domain, which is physically characterized. The above study will pave the way towards solving the atomic structure of this essential mammalian protein.
RESUMO
BACKGROUND: De novo fatty acid synthesis catalyzed by fatty acid synthase (FASN) is crucial for tumor cell survival. Thus therapeutic targeting of FASN is considered as a novel antineoplastic strategy. However, little is understood in this respect regarding malignancies of hematological origin. The present investigation was therefore, undertaken to study the molecular mechanisms of the antitumor action of FASN inhibitor orlistat (tetrahydrolipstatin) using a murine model of a T cell lymphoma. METHODS: The antitumor efficacy of orlistat was investigated in vitro by estimating cell survival by MTT assay and apoptosis by Wright Giemsa, TUNEL, Annexin-V/PI staining and % DNA fragmentation. Generation of reactive oxygen species (ROS) in tumor cells was studied using fluorescence microscopy. Expression of genes and proteins was carried out by RT-PCR and western blot analyses respectively. FASN and CPT-1 activity was estimated by spectrophotometer. Cytokines expression was analyzed by ELISA. RESULTS: We report that inhibition of FASN with its specific inhibitor orlistat manifests tumor-specific inhibition of cell survival, accompanied by induction of apoptosis. Orlistat-treated tumor cells showed an altered ROS generation, shift in cytokine balance and modulated expression of cell survival regulatory molecules like HSP70, Bcl2, p53, PUMA, Caspase-3 and CAD. It was observed that IFN-γ mediates orlistat-dependent modulation of FASN expression. CONCLUSION AND GENERAL SIGNIFICANCE: In this study, we report some of the so far unexplored novel aspects underlying the molecular mechanisms associated with orlistat-dependent modulation of tumor cell survival. These observations will help in designing antineoplastic therapeutic protocols using orlistat against malignancies of hematological origin.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Ácido Graxo Sintases/antagonistas & inibidores , Lactonas/farmacologia , Linfoma de Células T/tratamento farmacológico , Animais , Sobrevivência Celular/efeitos dos fármacos , Citocinas/biossíntese , Relação Dose-Resposta a Droga , Proteínas de Choque Térmico HSP70/fisiologia , Interferon gama/fisiologia , Linfoma de Células T/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Orlistate , Espécies Reativas de Oxigênio/metabolismoRESUMO
The present investigation was undertaken to study the effect of in vitro exposure of Colo205, colonadenocarcinoma cells, to monocarboxylate transporter inhibitor α-cyano-4-hydroxycinnamate (αCHC) on cell survival and evolution of resistance to chemotherapeutic drug cisplatin. αCHC-treated Colo205 cells showed inhibition of survival accompanied by an augmented induction of apoptosis. Changes in cell survival properties were associated with alterations in lactate efflux, pH homeostasis, expression of glucose transporters, glucose uptake, HIF-1α, generation of nitric oxide, expression pattern of some key cell survival regulatory molecules: Bcl2, Bax, active caspase-3 and p53. Pretreatment of Colo205 cells with αCHC also altered their susceptibility to the cytotoxicity of cisplatin accompanied by altered expression of multidrug resistance regulating MDR1 and MRP1 genes. This study for the first time deciphers some of the key molecular events underlying modulation of cell survival of cancer cells of colorectal origin by αCHC and its contribution to chemosensitization against cisplatin. Thus these findings will be of immense help in further research for optimizing the use of αCHC for improving the chemotherapeutic efficacy of anticancer drugs like cisplatin.
Assuntos
Adenocarcinoma/fisiopatologia , Apoptose/efeitos dos fármacos , Cisplatino/farmacologia , Neoplasias do Colo/fisiopatologia , Ácidos Cumáricos/farmacologia , Resistencia a Medicamentos Antineoplásicos , Transportadores de Ácidos Monocarboxílicos/antagonistas & inibidores , Proteínas Musculares/antagonistas & inibidores , Adenocarcinoma/tratamento farmacológico , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Caspase 3/genética , Caspase 3/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Neoplasias do Colo/tratamento farmacológico , Neoplasias do Colo/genética , Neoplasias do Colo/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Transportadores de Ácidos Monocarboxílicos/genética , Transportadores de Ácidos Monocarboxílicos/metabolismo , Proteínas Musculares/genética , Proteínas Musculares/metabolismoRESUMO
Using a murine tumor model, we demonstrate that tumor cells display a tumor stage-dependent differential glucose utilization associated with an altered GLUT-1 expression. Hyperglycemic tumor microenvironment modulates the tumorigenic ability, survival, apoptosis, and glucose utilization of tumor cells in late tumor-bearing stage accompanied by an altered tumor acidosis and expression of cell survival regulatory molecules: HIF-1α, p53, Bcl2, caspase-activated DNase, IL-4, IL-6, IL-10, IFN-γ, TGF-ß, and VEGF. Glucose-exposed tumor cells of late tumor-bearing stage also show a declined susceptibility to the cytotoxic action of chemotherapeutic drugs: cisplatin and methotrexate, accompanied by an increased expression of MDR-1 gene. Taken together the results show that hyperglycemic tumor microenvironment differentially alters tumor growth depending on the stage of tumor progression.
Assuntos
Resistencia a Medicamentos Antineoplásicos , Glucose/metabolismo , Hiperglicemia/patologia , Linfoma de Células T/metabolismo , Linfoma de Células T/patologia , Microambiente Tumoral , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/genética , Animais , Transporte Biológico , Sobrevivência Celular , Cisplatino/farmacologia , Citocinas/metabolismo , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Glucose/farmacologia , Transportador de Glucose Tipo 1/genética , Transportador de Glucose Tipo 1/metabolismo , Hiperglicemia/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Linfoma de Células T/tratamento farmacológico , Masculino , Camundongos , Camundongos Endogâmicos BALB CRESUMO
Targeting of tumor metabolism is emerging as a novel therapeutic strategy against cancer. Dichloroacetate (DCA), an inhibitor of pyruvate dehydrogenase kinase (PDK), has been shown to exert a potent tumoricidal action against a variety of tumor cells. The main mode of its antineoplastic action implicates a shift of glycolysis to oxidative metabolism of glucose, leading to generation of cytotoxic reactive oxygen intermediates. However, the effect of DCA on tumor microenvironment, which in turn regulates tumor cell survival; remains speculative to a large extent. It is also unclear if DCA can exert any modulatory effect on the process of hematopoiesis, which is in a compromised state in tumor-bearing hosts undergoing chemotherapy. In view of these lacunas, the present study was undertaken to investigate the so far unexplored aspects with respect to the molecular mechanisms of DCA-dependent tumor growth retardation and chemosensitization. BALB/c mice were transplanted with Dalton's lymphoma (DL) cells, a T cell lymphoma of spontaneous origin, followed by administration of DCA with or without cisplatin. DCA-dependent tumor regression and chemosensitization to cisplatin was found to be associated with altered repertoire of key cell survival regulatory molecules, modulated glucose metabolism, accompanying reconstituted tumor microenvironment with respect to pH homeostasis, cytokine balance and alternatively activated TAM. Moreover, DCA administration also led to an alteration in the MDR phenotype of tumor cells and myelopoietic differentiation of macrophages. The findings of this study shed a new light with respect to some of the novel mechanisms underlying the antitumor action of DCA and thus may have immense clinical applications.
Assuntos
Antineoplásicos/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Ácido Dicloroacético/farmacologia , Glucose/metabolismo , Macrófagos/efeitos dos fármacos , Microambiente Tumoral/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Diferenciação Celular/efeitos dos fármacos , Cisplatino/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Glicólise , Marcação In Situ das Extremidades Cortadas , Linfoma de Células T/patologia , Macrófagos/citologia , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/metabolismo , Piruvato Desidrogenase Quinase de Transferência de AcetilRESUMO
In the present investigation, we explored the molecular mechanisms of the tumoricidal action of α-cyano-4-hydroxycinnamate (αCHC) on the cells of Dalton's lymphoma (DL), which is a murine T cell lymphoma. In-vitro treatment of the DL cells with αCHC resulted in the modulation of the biophysical parameters of the tumor cell culture medium with respect to pH, nitric oxide, glucose, and lactate, accompanied by an alteration in the expression of cytokines (IL-10, IL-6, and IFN-γ) and cell survival-regulatory proteins such as Bcl2, p53, caspase-3, caspase-activated DNase, HSP70, and IL2R. The expression of the pH-regulatory proteins vacuolar ATPase and MCT1 was also found to be altered. The study discusses the possible role of the aforementioned alterations triggered by αCHC in the induction of tumor cell apoptosis and decreased cell survival. These findings will provide a new insight into the novel molecular mechanisms of the antitumor action of αCHC.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Ácidos Cumáricos/farmacologia , Linfoma de Células T/tratamento farmacológico , Animais , Proteínas de Ciclo Celular/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Glucose/metabolismo , Interferon gama/metabolismo , Interleucina-10/metabolismo , Interleucina-6/metabolismo , Ácido Láctico/metabolismo , Linfoma de Células T/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Óxido Nítrico/metabolismo , Proteínas Oncogênicas/metabolismo , Timócitos/efeitos dos fármacos , Timócitos/metabolismo , Células Tumorais CultivadasRESUMO
Recent experimental evidence from our and other laboratories has strongly indicated that glutor, a piperazine-2-one derivative, which is a pan-GLUT inhibitor, displays a promising antineoplastic action by hampering glucose uptake owing to its ability to inhibit GLUT1 and GLUT3, which are overexpressed in neoplastic cells. However, the molecular mechanism(s) of the inhibiting action of glutor has remained elusive. Thus, for optimal utilization of the antineoplastic potential of glutor, it is essential to decipher the precise mechanism(s) of its interaction with GLUTs. Therefore, the present investigation was carried out to understand the molecular mechanism(s) of the binding of glutor to GLUT1 and GLUT3 in silico. This study suggests that glutor can effectively bind to GLUTs at the reported binding site. Moreover, the docking of glutor to GLUT was stabilised by several contacts between these two partners as shown by the 200 ns long molecular dynamic simulation carried out using Gromacs, indicating the formation of a stable complex. Moreover, glutor was found to possess all characteristics conducive to its drug-likeness. Hence, these observations suggest that glutor has the potential to be used in antineoplastic therapeutic applications.Communicated by Ramaswamy H. Sarma.
Assuntos
Antineoplásicos , Transportador de Glucose Tipo 1 , Transportador de Glucose Tipo 3 , Antineoplásicos/farmacologia , Sítios de Ligação , Transporte Biológico , Simulação de Dinâmica Molecular , Simulação de Acoplamento MolecularRESUMO
The present investigation studied the effect of aspirin administration in tumor-bearing hosts on bone marrow cellularity and myelopoiesis. Aspirin administration to mice bearing a transplantable T-cell lymphoma, designated as Dalton's lymphoma (DL), augmented proliferation of bone marrow cells (BMC). BMC of aspirin-administered tumor-bearing mice were found to be predominantly in the S phase of cell cycle releasing them from G0/G1 arrest. Aspirin-exposed BMC also showed an altered expression of survival and cell cycle regulatory proteins p53, bcl2, caspase-activated deoxyribonuclease (CAD), cyclin B1 and cyclin D. Moreover, the BMC of aspirin-administered tumor-bearing mice showed an augmented colony-forming ability and differentiation in the macrophage lineage with an activated phenotype of bone marrow-derived macrophages (BMDM) with respect to macrophage-mediated tumoricidal activity and production of nitric oxide, IL-1ß, TNFα and VEGF. On the other hand aspirin administration to normal mice showed little effect on bone marrow cellularity and myeloid differentiation. In this model, aspirin had a myelopoetic action in tumor-bearing host.
Assuntos
Aspirina/farmacologia , Linfoma de Células T/metabolismo , Mielopoese/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Aspirina/administração & dosagem , Células da Medula Óssea/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Proteínas de Ciclo Celular/metabolismo , Proliferação de Células , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB CRESUMO
Aspirin is an anti-inflammatory drug demonstrated to possess a tremendous anticancer potential. As progression of some tumors is influenced by sex hormones, we investigated if the antineoplastic action of aspirin shows gender dependence. Using a murine model of T-cell lymphoma, the present investigation was undertaken to study if the antitumor actions of aspirin against lymphoma cells display gender dimorphism. The findings of the present investigation indicate that aspirin administration to male and female tumor-bearing hosts resulted in gender dependent differential tumor growth retardation. Such gender dichotomy of aspirin's antitumor action was associated with a differential impact on cell cycle progression and expression of cell survival regulatory molecules. Aspirin administration was also found to modulate crucial parameters of tumor microenvironment, including contents of glucose, lactate and cell growth regulatory cytokines, in a gender specific manner. Aspirin was found to reverse estrogen-dependent augmentation of tumor cell survival in vitro. Taken together the results of the present study suggest that the antineoplastic action of aspirin is gender-dependent and should be considered in designing of gender-specific therapeutic applications of aspirin.
Assuntos
Antineoplásicos/uso terapêutico , Aspirina/uso terapêutico , Linfoma de Células T/tratamento farmacológico , Androgênios/farmacologia , Animais , Antineoplásicos/farmacologia , Aspirina/farmacologia , Ciclo Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Modelos Animais de Doenças , Estrogênios/farmacologia , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Linfoma de Células T/genética , Linfoma de Células T/mortalidade , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Fatores Sexuais , Microambiente Tumoral/efeitos dos fármacosRESUMO
The present investigation was undertaken to study if curcumin, which is recognized for its potential as an antineoplastic and immunopotentiating agent, can also influence the process of myelopoiesis in a tumor-bearing host. Administration of curcumin to tumor-bearing host augmented count of bone marrow cell (BMC) accompanied by an up-regulated BMC survival and a declined induction of apoptosis. Curcumin administration modulated expression of cell survival regulatory molecules: Bcl2, p53, caspase-activated DNase (CAD) and p53-upregulated modulator of apoptosis (PUMA) along with enhanced expression of genes of receptors for M-CSF and GM-CSF in BMC. The BMC harvested from curcumin-administered hosts showed an up-regulated colony forming ability with predominant differentiation into bone marrow-derived macrophages (BMDM), responsive for activation to tumoricidal state. The number of F4/80 positive bone marrow resident macrophages (BMM), showing an augmented expression of M-CSF, was also augmented in the bone marrow of curcumin-administered host. In vitro reconstitution experiments indicated that only BMM of curcumin-administered hosts, but not in vitro curcumin-exposed BMM, augmented BMC survival. It suggests that curcumin-dependent modulation of BMM is of indirect nature. Such prosurvival action of curcumin is associated with altered T(H1)/T(H2) cytokine balance in serum. Augmented level of serum-borne IFN-γ was found to mediate modulation of BMM to produce enhanced amount of monokines (IL-1, IL-6, TNF-α), which are suggested to augment the BMC survival. Taken together the present investigation indicates that curcumin can potentiate myelopoiesis in a tumor-bearing host, which may have implications in its therapeutic utility.
Assuntos
Curcumina/farmacologia , Macrófagos/efeitos dos fármacos , Mielopoese/efeitos dos fármacos , Neoplasias Experimentais/fisiopatologia , Animais , Apoptose/efeitos dos fármacos , Western Blotting , Citocinas/fisiologia , Relação Dose-Resposta a Droga , Ensaio de Imunoadsorção Enzimática , Marcação In Situ das Extremidades Cortadas , Macrófagos/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Mielopoese/fisiologia , Transplante de NeoplasiasRESUMO
The present study was conducted to investigate if anti-inflammatory drug aspirin could alter the cytotoxic action of cisplatin on tumor cells. Using a transplantable T cell lymphoma in a murine model, we demonstrate that exposure to aspirin exerts a priming action on tumor cells, rendering them susceptible to induction of cell death by cisplatin with consequences on retardation of tumor progression. The priming action of aspirin on tumor cells was found to be dependent on an altered constitution of tumor microenvironment with respect to decline of acidosis and modulation in the expression of cell cycle and survival regulatory molecules like cyclin B1, cyclin D, bcl-2, bcl-xL, p53, and cytokines: IL-4, IL-10, IFN- γ & VEGF. The study also discusses possible mechanisms underlying augmentary action of aspirin on cisplatin-mediated tumor cells killing. This is the first report showing that pre-exposure of tumor cells to aspirin lowers the concentration of cisplatin to exert its cytotoxic action. The finding of this study will help in designing novel antitumor protocols with reduced dose of cisplatin.
Assuntos
Antineoplásicos/farmacologia , Apoptose , Aspirina/farmacologia , Ciclo Celular/efeitos dos fármacos , Cisplatino/farmacologia , Animais , Morte Celular , Sobrevivência Celular , Ciclinas/metabolismo , Citocinas/metabolismo , Sinergismo Farmacológico , Interleucina-10/metabolismo , Interleucina-4/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Células Tumorais Cultivadas , Microambiente TumoralRESUMO
Neoplastic cells displayed altered metabolism with accelerated glycolysis. Therefore, these cells need a mammoth supply of glucose for which they display an upregulated expression of various glucose transporters (GLUT). Thus, novel antineoplastic strategies focus on inhibiting GLUT to intersect the glycolytic lifeline of cancer cells. This review focuses on the current status of various GLUT inhibition scenarios. The GLUT inhibitors belong to both natural and synthetic small inhibitory molecules category. As neoplastic cells express multiple GLUT isoforms, it is necessary to use pan-GLUT inhibitors. Nevertheless, it is also necessary that such pan-GLUT inhibitors exert their action at a low concentration so that normal healthy cells are left unharmed and minimal injury is caused to the other vital organs and systems of the body. Moreover, approaches are also emerging from combining GLUT inhibitors with other chemotherapeutic agents to potentiate the antineoplastic action. A new pan-GLUT inhibitor named glutor, a piperazine-one derivative, has shown a potent antineoplastic action owing to its inhibitory action exerted at nanomolar concentrations. The review discusses the merits and limitations of the existing GLUT inhibitory approach with possible future outcomes.
RESUMO
Neoplastic cells overexpress glucose transporters (GLUT), particularly GLUT1 and GLUT3, to support altered metabolism. Hence, novel strategies are being explored to effectively inhibit GLUTs for a daunting interference of glucose uptake. Glutor, a piperazine-2-one derivative, is a newly reported pan-GLUT inhibitor with a promising antineoplastic potential. However, several aspects of the underlying mechanisms remain obscure. To understand this better, tumor cells of thymic origin designated as Dalton's lymphoma (DL) were treated with glutor and analyzed for survival and metabolism regulatory molecular events. Treatment of tumor cells with glutor caused a decrease in cell survival with augmented induction of apoptosis. It also caused a decrease in glucose uptake associated with altered expression of GLUT1 and GLUT3. HIF-1α, HK-2, LDH-A, and MCT1 also decreased with diminished lactate production and deregulated pH homeostasis. Moreover, glutor treatment modulated the expression of cell survival regulatory molecules p53, Hsp70, IL-2 receptor CD25, and C-myc along with mitochondrial membrane depolarization, increased intracellular ROS expression, and altered Bcl-2/BAX ratio. Glutor also enhanced the chemosensitivity of tumor cells to cisplatin, accompanied by decreased MDR1 expression. Adding fructose to the culture medium containing glutor reversed the latter's inhibitory action on tumor cell survival. These results demonstrate that in addition to inhibited glucose uptake, modulated tumor growth regulatory molecular pathways are also implicated in the manifestation of the antineoplastic action of glutor. Thus, the novel findings of this study will have a long-lasting clinical significance in evaluating and optimizing the use of glutor in anticancer therapeutic strategies.