RESUMO
The inhibition of kynurenine production is considered a promising target for cancer immunotherapy. In this study, an amino acid derivative, compound 1 was discovered using a cell-based assay with our screening library. Compound 1 suppressed kynurenine production without inhibiting indoleamine 2,3-dioxygenase 1 (IDO1) activity. The activity of 1 was derived from the inhibition of IDO1 by a metabolite of 1, O-benzylhydroxylamine (OBHA, 2a). A series of N-substituted 2a derivatives that exhibit potent activity in cell-based assays may represent effective prodrugs. Therefore, we synthesized and evaluated novel N,O-substituted hydroxylamine derivatives. The structure-activity relationships revealed that N,O-substituted hydroxylamine 2c inhibits kynurenine production in a cell-based assay. We conducted an in vivo experiment with 2c, although the effectiveness of O-substituted hydroxylamine derivatives in vivo has not been previously reported. The results indicate that N,O-substituted hydroxylamine derivatives are promising IDO1 inhibitors.
Assuntos
Hidroxilamina , Indolamina-Pirrol 2,3,-Dioxigenase , Cinurenina , Cinurenina/metabolismo , Indolamina-Pirrol 2,3,-Dioxigenase/antagonistas & inibidores , Indolamina-Pirrol 2,3,-Dioxigenase/metabolismo , Relação Estrutura-Atividade , Humanos , Hidroxilamina/química , Hidroxilamina/farmacologia , Hidroxilaminas/química , Hidroxilaminas/farmacologia , Estrutura Molecular , Animais , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Camundongos , Relação Dose-Resposta a DrogaRESUMO
Large regions in tumor tissues, particularly pancreatic cancer, are hypoxic and nutrient-deprived because of unregulated cell growth and insufficient vascular supply. Certain cancer cells, such as those inside a tumor, can tolerate these severe conditions and survive for prolonged periods. We hypothesized that small molecular agents, which can preferentially reduce cancer cell survival under nutrient-deprived conditions, could function as anticancer drugs. In this study, we constructed a high-throughput screening system to identify such small molecules and screened chemical libraries and microbial culture extracts. We were able to determine that some small molecular compounds, such as penicillic acid, papyracillic acid, and auranofin, exhibit preferential cytotoxicity to human pancreatic cancer cells under nutrient-deprived compared with nutrient-sufficient conditions. Further analysis revealed that these compounds target to redox systems such as GSH and thioredoxin and induce accumulation of reactive oxygen species in nutrient-deprived cancer cells, potentially contributing to apoptosis under nutrient-deprived conditions. Nutrient-deficient cancer cells are often deficient in GSH; thus, they are susceptible to redox system inhibitors. Targeting redox systems might be an attractive therapeutic strategy under nutrient-deprived conditions of the tumor microenvironment.
Assuntos
Alcenos/química , Auranofina/química , Glutationa/química , Ácido Penicílico/química , Compostos de Espiro/química , Tiorredoxinas/química , Alcenos/farmacologia , Animais , Antineoplásicos/química , Antineoplásicos/uso terapêutico , Auranofina/farmacologia , Auranofina/uso terapêutico , Caspase 3/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Feminino , Glutationa/metabolismo , Humanos , Metaboloma/efeitos dos fármacos , Camundongos , Camundongos Nus , Nutrientes/química , Nutrientes/deficiência , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Ácido Penicílico/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Compostos de Espiro/farmacologia , Tiorredoxinas/metabolismo , Regulação para Cima/efeitos dos fármacosRESUMO
BACKGROUND: Toxoplasmosis, a parasitic disease caused by Toxoplasma gondii, is an important cause of miscarriage or adverse fetal effects, including neurological and ocular manifestations in humans. Current anti-Toxoplasma drugs have limited efficacy against toxoplasmosis and also have severe side effects. Therefore, novel efficacious drugs are urgently needed. Here, we identified metacytofilin (MCF) from a fungal Metarhizium species as a potential anti-Toxoplasma compound. METHODS: Anti-Toxoplasma activities of MCF and its derivatives were evaluated in vitro and in vivo using nonpregnant and pregnant mice. To understand the mode of action of MCF, the RNA expression of host and parasite genes was investigated by RNAseq. RESULTS: In vitro, MCF inhibited the viability of intracellular and extracellular T. gondii. Administering MCF intraperitoneally or orally to mice after infection with T. gondii tachyzoites increased mouse survival compared with the untreated animals. Remarkably, oral administration of MCF to pregnant mice prevented vertical transmission of the parasite. Interestingly, RNA sequencing of T. gondii-infected cells treated with MCF showed that MCF inhibited DNA replication and enhanced RNA degradation in the parasites. CONCLUSIONS: With its potent anti-T. gondii activity, MCF is a strong candidate for future drug development against toxoplasmosis.
Assuntos
Antiparasitários/uso terapêutico , Transmissão Vertical de Doenças Infecciosas/prevenção & controle , Oxazinas/uso terapêutico , Toxoplasma/efeitos dos fármacos , Toxoplasmose/tratamento farmacológico , Toxoplasmose/mortalidade , Administração Intravenosa , Administração Oral , Animais , Antiparasitários/administração & dosagem , Antiparasitários/farmacologia , Replicação do DNA/efeitos dos fármacos , DNA de Protozoário , Modelos Animais de Doenças , Feminino , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Oxazinas/administração & dosagem , Oxazinas/farmacologia , Gravidez , Taxa de Sobrevida , Toxoplasma/genética , Toxoplasmose/parasitologia , Toxoplasmose/transmissão , Resultado do TratamentoRESUMO
Modulation of prostate stromal cells (PrSCs) within tumor tissues is gaining attention for the treatment of solid tumors. Using our original in vitro coculture system, we previously reported that leucinostatin (LCS)-A, a peptide mycotoxin, inhibited prostate cancer DU-145 cell growth through reduction of insulin-like growth factor 1 (IGF-I) expression in PrSCs. To further obtain additional bioactive compounds from LCS-A, we designed and synthesized a series of LCS-A derivatives as compounds that target PrSCs. Among the synthesized LCS-A derivatives, LCS-7 reduced IGF-I expression in PrSCs with lower toxicity to PrSCs and mice than LCS-A. As LCS-A has been suggested to interact with mitochondrial adenosine triphosphate (ATP) synthase, a docking study was performed to elucidate the mechanism of reduced IGF-I expression in the PrSCs. As expected, LCS-A and LCS-7 directly interacted with mitochondrial ATP synthase, and like LCS-A and LCS-7, other mitochondrial ATP synthase inhibitors also reduced the expression of IGF-I by PrSCs. Furthermore, LCS-A and LCS-7 significantly decreased the growth of mouse xenograft tumors. Based on these data, we propose that the mitochondrial ATP synthases-IGF-I axis of PrSCs plays a critical role on cancer cell growth and inhibition could be a potential anticancer target for prostate cancer.
Assuntos
Peptídeos Catiônicos Antimicrobianos/farmacologia , Fator de Crescimento Insulin-Like I/metabolismo , ATPases Mitocondriais Próton-Translocadoras/antagonistas & inibidores , Neoplasias da Próstata/tratamento farmacológico , Células Estromais/efeitos dos fármacos , Animais , Peptídeos Catiônicos Antimicrobianos/uso terapêutico , Linhagem Celular Tumoral , Técnicas de Cocultura , Feminino , Humanos , Masculino , Camundongos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/enzimologia , ATPases Mitocondriais Próton-Translocadoras/metabolismo , Simulação de Acoplamento Molecular , Próstata/citologia , Próstata/efeitos dos fármacos , Próstata/patologia , Neoplasias da Próstata/patologia , Células Estromais/metabolismo , Células Estromais/patologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The now clinically-used anti-epidermal growth factor receptor (EGFR) monoclonal antibodies have demonstrated significant efficacy only in patients with metastatic colorectal cancer (mCRC), with wild-type Kirsten rat sarcoma viral oncogene homolog (KRAS). However, no effective treatments for patients with mCRC with KRAS mutated tumors have been approved yet. Therefore, a new strategy for targeting mCRC with KRAS mutated tumors is desired. In the present study, we examined the anti-tumor activities of a novel anti-EGFR monoclonal antibody, EMab-17 (mouse IgG2a, kappa), in colorectal cancer (CRC) cells with the KRAS p.G13D mutation. This antibody recognized endogenous EGRF in CRC cells with or without KRAS mutations, and showed a high sensitivity for CRC cells in flow cytometry, indicating that EMab-17 possesses a high binding affinity to the endogenous EGFR. In vitro experiments showed that EMab-17 exhibited antibody-dependent cellular cytotoxicity and complement-dependent cytotoxicity activities against CRC cells. In vivo analysis revealed that EMab-17 inhibited the metastases of HCT-15 and HCT-116 cells in the livers of nude mouse metastatic models, unlike the anti-EGFR monoclonal antibody EMab-51 of subtype mouse IgG1. In conclusion, EMab-17 may be useful in an antibody-based therapy against mCRC with the KRAS p.G13D mutation.
Assuntos
Anticorpos Monoclonais/farmacologia , Antineoplásicos Imunológicos/farmacologia , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Hepáticas/tratamento farmacológico , Mutação , Proteínas Proto-Oncogênicas p21(ras)/genética , Animais , Citotoxicidade Celular Dependente de Anticorpos , Células CHO , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Neoplasias Colorretais/genética , Neoplasias Colorretais/imunologia , Neoplasias Colorretais/patologia , Proteínas do Sistema Complemento/genética , Cricetulus , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/genética , Receptores ErbB/imunologia , Feminino , Regulação Neoplásica da Expressão Gênica , Células HCT116 , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/imunologia , Neoplasias Hepáticas/secundário , Camundongos , Camundongos Nus , Proteínas Proto-Oncogênicas p21(ras)/imunologia , Resultado do Tratamento , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Small cell lung cancer (SCLC) is a severe malignancy associated with early and widespread metastasis. To study SCLC metastasis, we previously developed an orthotopic transplantation model using the human SCLC cell line DMS273. In the model, metastatic foci were found in distant tissues such as bone and the adrenal gland, similarly as observed in patients with SCLC. In this study, we evaluated the differentially expressed genes between orthotopic and metastatic tumors in the model. We isolated tumor cells from orthotopic and metastatic sites, and the tumor cell RNA was analyzed using DNA microarray analysis. We found that 19 genes in metastatic tumors were upregulated by more than 4-fold compared with their expression in orthotopic tumors. One of these genes encodes a transmembrane protein, interferon (IFN)-induced transmembrane protein 1 (IFITM1), and immunohistochemical analysis confirmed the higher expression of the protein in metastatic sites than in orthotopic sites. IFITM1 was also detected in some SCLC cell lines and lung tumors from patients with SCLC. The overexpression of IFITM1 in DMS273 cells increased their metastatic formation in the orthotopic model and in an experimental metastasis model. Conversely, the silencing of IFITM1 suppressed metastatic formation by DMS273 cells. We also found that IFITM1 overexpression promoted the metastatic formation of NCI-H69 human SCLC cells. These results demonstrate that IFITM1 promotes distant metastasis in xenograft models of human SCLC.
Assuntos
Antígenos de Diferenciação/fisiologia , Carcinoma de Células Pequenas/secundário , Neoplasias Pulmonares/patologia , Metástase Neoplásica/fisiopatologia , Proteínas de Neoplasias/fisiologia , Neoplasias das Glândulas Suprarrenais/secundário , Animais , Antígenos de Diferenciação/genética , Neoplasias Ósseas/secundário , Carcinoma de Células Pequenas/genética , Carcinoma de Células Pequenas/metabolismo , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Xenoenxertos , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Camundongos , Camundongos Nus , Invasividade Neoplásica , Metástase Neoplásica/genética , Proteínas de Neoplasias/genética , Transplante de Neoplasias , Especificidade de Órgãos , Interferência de RNA , RNA Neoplásico/genética , RNA Interferente Pequeno/genéticaRESUMO
Coxsackievirus and adenovirus receptor (CAR) is a single-pass transmembrane protein that is associated with adenoviral infection. CAR is involved in the formation of epithelial tight junctions and promotes tumor growth in some cancers. Previously, we developed mouse monoclonal antibodies against human CAR and found that one, mu6G10A, significantly inhibited tumor growth in xenografts of human cancer cells. Herein, we generated and characterized a mouse-human chimeric anti-CAR antibody (ch6G10A) from mu6G10A. ch6G10A had binding activity, inducing antibody-dependent cellular cytotoxicity and complement-dependent cytotoxicity, and in vivo anti-tumor activity against CAR-expressing prostate cancer DU-145 cells. In addition, cancer tissue array analysis confirmed that CAR is highly expressed in neuroendocrine lung cancers including small cell lung cancer, and treatment with ch6G10A effectively inhibited in vivo subcutaneous tumor growth of NCI-H69 small cell lung cancer cells in nude mice. Moreover, treatment with mu6G10A effectively inhibited both in vivo orthotopic tumor growth and distant metastatic formation in mouse xenograft models of a highly metastatic subline of human small cell lung cancer DMS273 cells. These results suggest that targeting therapy to CAR with a therapeutic antibody might be effective against several cancer types including small cell lung cancer.
Assuntos
Anticorpos Monoclonais/uso terapêutico , Proteína de Membrana Semelhante a Receptor de Coxsackie e Adenovirus/antagonistas & inibidores , Neoplasias Pulmonares/terapia , Neoplasias da Próstata/terapia , Carcinoma de Pequenas Células do Pulmão/terapia , Animais , Citotoxicidade Celular Dependente de Anticorpos , Proteínas do Sistema Complemento/imunologia , Proteína de Membrana Semelhante a Receptor de Coxsackie e Adenovirus/imunologia , Proteína de Membrana Semelhante a Receptor de Coxsackie e Adenovirus/metabolismo , Neoplasias das Glândulas Endócrinas/metabolismo , Neoplasias das Glândulas Endócrinas/terapia , Feminino , Humanos , Neoplasias Pulmonares/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Terapia de Alvo Molecular , Neoplasias da Próstata/metabolismo , Carcinoma de Pequenas Células do Pulmão/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: Helicobacter pylori (H. pylori) infection causes various gastrointestinal diseases including gastric cancer. Hence, eradication of this infection could prevent these diseases. The most popular first-line treatment protocol to eradicate H. pylori is termed "triple therapy" and consists of a proton pump inhibitor (PPI), clarithromycin, and amoxicillin or metronidazole. However, the antibiotics used to treat H. pylori infection are hindered by the antibiotics-resistant bacteria and by their antimicrobial activity against intestinal bacteria, leading to side effects. Therefore, an alternative treatment with fewer adverse side effects is urgently required to improve the overall eradication rate of H. pylori. OBJECTIVE: The aim of this study was to assess the effectiveness and mechanism of action of an antitumor agent, intervenolin, and its derivatives as an agent for the treatment of H. pylori infection. RESULTS: We demonstrate that intervenolin, and its derivatives showed selective anti-H. pylori activity, including antibiotic-resistant strains, without any effect on intestinal bacteria. We showed that dihydroorotate dehydrogenase, a key enzyme for de novo pyrimidine biosynthesis, is a target and treatment with intervenolin or its derivatives decreased the protein and mRNA levels of H. pylori urease, which protects H. pylori against acidic conditions in the stomach. Using a mouse model of H. pylori infection, oral monotherapy with the intervenolin derivative AS-1934 had a stronger anti-H. pylori effect than the triple therapy commonly used worldwide to eradicate H. pylori. CONCLUSION: AS-1934 has potential advantages over current treatment options for H. pylori infection.
Assuntos
Infecções por Helicobacter/tratamento farmacológico , Quinolonas/uso terapêutico , Antibacterianos/uso terapêutico , Helicobacter pylori/efeitos dos fármacos , Helicobacter pylori/patogenicidade , Humanos , Resultado do TratamentoRESUMO
BACKGROUND: EGFRvIII is a mutant form of the epidermal growth factor receptor gene (EGFR) that lacks exons 2-7. The resulting protein does not bind to ligands and is constitutively activated. The expression of EGFRvIII is likely confined to various types of cancer, particularly glioblastomas. Although an anti-EGFRvIII vaccine is of great interest, low-molecular-weight substances are needed to obtain better therapeutic efficacy. Thus, the purpose of this study is to identify low molecular weight substances that can suppress EGFRvIII-dependent transformation. METHODS: We constructed a new throughput screening system and searched for substances that decreased cell survival of NIH3T3/EGFRvIII spheres under 3-dimensional (3D)-culture conditions, but retained normal NIH3T3 cell growth under 2D-culture conditions. In vivo activity was examined using a mouse transplantation model, and derivatives were chemically synthesized. Functional characterization of the candidate molecules was investigated using an EGFR kinase assay, immunoprecipitation, western blotting, microarray analysis, quantitative polymerase chain reaction analysis, and measurement of lactate and ATP synthesis. RESULTS: In the course of screening 30,000 substances, a reagent, "Ertredin" was found to inhibit anchorage-independent 3D growth of sphere-forming cells transfected with EGFRvIII cDNA. Ertredin also inhibited sphere formation in cells expressing wild-type EGFR in the presence of EGF. However, it did not affect anchorage-dependent 2D growth of parental NIH3T3 cells. The 3D-growth-inhibitory activity of some derivatives, including those with new structures, was similar to Ertredin. Furthermore, we demonstrated that Ertredin suppressed tumor growth in an allograft transplantation mouse model injected with EGFRvIII- or wild-type EGFR-expressing cells; a clear toxicity to host animals was not observed. Functional characterization of Ertredin in cells expressing EGFRvIII indicated that it stimulated EGFRvIII ubiquitination, suppressed both oxidative phosphorylation and glycolysis under 3D conditions, and promoted cell apoptosis. CONCLUSION: We developed a high throughput screening method based on anchorage-independent sphere formation induced by EGFRvIII-dependent transformation. In the course of screening, we identified Ertredin, which inhibited anchorage-independent 3D growth and tumor formation in nude mice. Functional analysis suggests that Ertredin suppresses both mitochondrial oxidative phosphorylation and cytosolic glycolysis in addition to promoting EGFRvIII degradation, and stimulates apoptosis in sphere-forming, EGFRvIII-overexpressing cells.
Assuntos
Antineoplásicos/farmacologia , Receptores ErbB/metabolismo , Glicólise/efeitos dos fármacos , Fosforilação Oxidativa/efeitos dos fármacos , Quinoxalinas/farmacologia , Esferoides Celulares/efeitos dos fármacos , Carga Tumoral/efeitos dos fármacos , Animais , Antineoplásicos/química , Apoptose/efeitos dos fármacos , Apoptose/genética , Western Blotting , Técnicas de Cultura de Células , Transformação Celular Neoplásica/efeitos dos fármacos , Transformação Celular Neoplásica/genética , Receptores ErbB/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Estrutura Molecular , Células NIH 3T3 , Quinoxalinas/química , Interferência de RNA , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Esferoides Celulares/metabolismo , Transplante Homólogo , Carga Tumoral/genéticaRESUMO
The proteasome degrades numerous regulatory proteins that are critical for tumor growth. Thus, proteasome inhibitors are promising antitumor agents. New proteasome inhibitors, such as tyropeptins and tyropeptin-boronic acid derivatives, have a potent inhibitory activity. Here we report the antitumor effects of two new tyropeptin-boronic acid derivatives, AS-06 and AS-29. AS-06 and AS-29 significantly suppress the degradation of the proteasome-sensitive fluorescent proteins in HEK293PS cells, and induce the accumulation of ubiquitinated proteins in human multiple myeloma cells. We show that these derivatives also suppress the degradation of the NF-κB inhibitor IκB-α and the nuclear translocation of NF-κB p65 in multiple myeloma cells, resulting in the inhibition of NF-κB activation. Furthermore, we demonstrate that AS-06 and AS-29 induce apoptosis through the caspase-8 and caspase-9 cascades. In a xenograft mouse model, i.v. administration of tyropeptin-boronic acid derivatives inhibits proteasome in tumors and clearly suppresses tumor growth in mice bearing human multiple myeloma. Our results indicate that tyropeptin-boronic acid derivatives could be lead therapeutic agents against human multiple myeloma.
Assuntos
Antineoplásicos/farmacologia , Ácidos Borônicos/farmacologia , Dipeptídeos/farmacologia , Mieloma Múltiplo/tratamento farmacológico , Inibidores de Proteassoma/farmacologia , Animais , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Células HEK293 , Humanos , Camundongos , Mieloma Múltiplo/patologia , Neoplasias Experimentais , Ubiquitinação/efeitos dos fármacos , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Dihydroorotate dehydrogenase (DHODH) is a central enzyme of the de novo pyrimidine biosynthesis pathway and is a promising drug target for the treatment of cancer and autoimmune diseases. This study presents the identification of a potent DHODH inhibitor by proteomic profiling. Cell-based screening revealed that NPD723, which is reduced to H-006 in cells, strongly induces myeloid differentiation and inhibits cell growth in HL-60 cells. H-006 also suppressed the growth of various cancer cells. Proteomic profiling of NPD723-treated cells in ChemProteoBase showed that NPD723 was clustered with DHODH inhibitors. H-006 potently inhibited human DHODH activity in vitro, whereas NPD723 was approximately 400 times less active than H-006. H-006-induced cell death was rescued by the addition of the DHODH product orotic acid. Moreover, metabolome analysis revealed that H-006 treatment promotes marked accumulation of the DHODH substrate dihydroorotic acid. These results suggest that NPD723 is reduced in cells to its active metabolite H-006, which then targets DHODH and suppresses cancer cell growth. Thus, H-006-related drugs represent a potentially powerful treatment for cancer and other diseases.
Assuntos
Di-Hidro-Orotato Desidrogenase , Proteômica , Humanos , Transformação Celular Neoplásica , Ciclo Celular , Morte CelularRESUMO
Sickle cell disease (SCD) is a heritable disorder caused by ß-globin gene mutations. Induction of fetal γ-globin is an established therapeutic strategy. Recently, epigenetic modulators, including G9a inhibitors, have been proposed as therapeutic agents. However, the molecular mechanisms whereby these small molecules reactivate γ-globin remain unclear. Here we report the development of a highly selective and non-genotoxic G9a inhibitor, RK-701. RK-701 treatment induces fetal globin expression both in human erythroid cells and in mice. Using RK-701, we find that BGLT3 long non-coding RNA plays an essential role in γ-globin induction. RK-701 selectively upregulates BGLT3 by inhibiting the recruitment of two major γ-globin repressors in complex with G9a onto the BGLT3 gene locus through CHD4, a component of the NuRD complex. Remarkably, BGLT3 is indispensable for γ-globin induction by not only RK-701 but also hydroxyurea and other inducers. The universal role of BGLT3 in γ-globin induction suggests its importance in SCD treatment.
Assuntos
Anemia Falciforme , RNA Longo não Codificante , Camundongos , Humanos , Animais , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , gama-Globinas/genética , Células Eritroides/metabolismo , Anemia Falciforme/tratamento farmacológico , Anemia Falciforme/genética , Anemia Falciforme/metabolismo , Expressão Gênica , Hemoglobina Fetal/genética , Hemoglobina Fetal/metabolismoRESUMO
A proteasome degrades numerous regulatory proteins that are critical for tumor growth and is therefore recognized as a promising anticancer target. Determining proteasome activity in the tumors of mice bearing xenografts is essential for the development of novel proteasome inhibitors. We developed a system for in vivo imaging of proteasome inhibition in the tumors of living mice, using a proteasome-sensitive fluorescent reporter, ZsProSensor-1. This reporter consists of a green fluorescent protein, ZsGreen, fused to mouse ornithine decarboxylase, which is degraded by the proteasome without being ubiquitinated. In stably transfected cells expressing ZsProSensor-1, the fluorescent reporter was rapidly degraded under steady-state conditions, whereas it was stabilized in the presence of proteasome inhibitors. Subcutaneous inoculation of the transfected cells into nude mice resulted in tumor formation. When the proteasome inhibitor bortezomib was intravenously administered to mice bearing these tumors, the ZsProSensor-1 protein accumulated in the tumors and emitted a fluorescent signal in a dose-dependent manner. Robust fluorescence was sustained for 3 days and then gradually decreased to baseline levels within 15 days. Intravenous administration of bortezomib also showed potent antitumor activity. In contrast, oral administration of bortezomib did not result in fluorescent protein accumulation in tumors or exhibit any antitumor activity. These results indicate that in vivo imaging using the ZsProSensor-1 fluorescent protein can be used as an indicator of antitumor activity and will be a powerful tool for the development of novel proteasome inhibitors.
Assuntos
Antineoplásicos/farmacologia , Ácidos Borônicos/farmacologia , Genes Reporter , Proteínas de Fluorescência Verde/genética , Imagem Molecular , Complexo de Endopeptidases do Proteassoma/genética , Inibidores de Proteassoma , Pirazinas/farmacologia , Animais , Antineoplásicos/administração & dosagem , Ácidos Borônicos/administração & dosagem , Bortezomib , Linhagem Celular , Feminino , Proteínas de Fluorescência Verde/metabolismo , Células HEK293 , Humanos , Camundongos , Camundongos Nus , Neoplasias/diagnóstico , Neoplasias/genética , Neoplasias/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Pirazinas/administração & dosagem , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
A bisabolane sesquiterpene endoperoxide compound, 3,6-epidioxy-1,10-bisaboladiene (EDBD), was isolated from edible wild plants grown in the northern area of Japan, Cacalia delphiniifolia and Cacalia hastata, using a mutant yeast (cdc2-1 rad9Δ). It showed cytotoxicity at IC(50) = 3.4 µM and induced apoptosis against the human promyelocytic leukemia cell line HL60 through a new stable rearrangement product (1) when in the presence of FeSO(4). This conversion mechanism is different from another sesquiterpene endoperoxide lactone compound, dihydroartemisinin (DHA), which is an anti-malarial drug. The cytotoxicity of EDBD decreased in the presence of the ferrous ion chelating drug deferoxamine mesylate (DFOM), and this suggested that the structural change of the drug caused by Fe(2+) may be responsible for its biological activities. EDBD induced apoptosis via phosphorylation of p38 mitogen-activated protein kinase (MAPK) in HL60 cells, and was detected by Western blot. EDBD resulted in an immediate increase in DCF fluorescence intensity in HL60 cells using DCFH-DA (2',7'-dichlorofluorescin diacetate) assay. The in vitro reaction of EDBD with FeSO(4) also increased DCF fluorescence intensity in a dose dependent manner. These results showed that the biological activity of EDBD involves an unstable carbon-centered radical intermediate. Furthermore, there was no similarity between the JFCR39 fingerprints of EDBD and DHA (correlation coefficient on COMPARE Analysis γ = 0.158). EDBD showed anti-tumor effects against a xenograft of Lox-IMVI cells in vivo.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Asteraceae/química , Neoplasias Experimentais/tratamento farmacológico , Plantas Comestíveis/química , Sesquiterpenos/farmacologia , Animais , Antineoplásicos Fitogênicos/química , Antineoplásicos Fitogênicos/isolamento & purificação , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Células HL-60 , Humanos , Camundongos , Camundongos Nus , Neoplasias Experimentais/patologia , Sesquiterpenos/química , Sesquiterpenos/isolamento & purificação , Relação Estrutura-AtividadeRESUMO
Nonreducing disaccharide trehalose is used as a stabilizer and humectant in various products and is a potential medicinal drug, showing curative effects on the animal models of various diseases. However, its use is limited as it is hydrolyzed by trehalase, a widely expressed enzyme in multiple organisms. Several trehalose analogs are prepared, including a microbial metabolite 4-trehalosamine, and their high biological stability is confirmed. For further analysis, 4-trehalosamine is selected as it shows high producibility. Compared with trehalose, 4-trehalosamine exhibits better or comparable protective activities and a high buffer capacity around the neutral pH. Another advantage of 4-trehalosamine is its chemical modifiability: simple reactions produce its various derivatives. Labeled probes and detergents are synthesized in one-pot reactions to exemplify the feasibility of their production, and their utility is confirmed for their respective applications. The labeled probes are used for mycobacterial staining. Although the derivative detergents can be effectively used in membrane protein research, long-chain detergents show 1000-3000-fold stronger autophagy-inducing activity in cultured cells than trehalose and are expected to become a drug lead and research reagent. These results indicate that 4-trehalosamine is a useful trehalose substitute for various purposes and a material to produce new useful derivative substances.
Assuntos
Detergentes , Trealose , Animais , Dissacarídeos , Trealase/metabolismo , Trealose/análogos & derivados , Trealose/farmacologiaRESUMO
The androgen receptor (AR) is an important therapeutic target for all clinical states of prostate cancer. We screened cultured broths of microorganisms for their ability to suppress androgen-dependent growth of human prostate cancer LNCaP and VCaP cells without cytotoxicity. We have already identified androprostamine A (APA) from a Streptomyces culture broth as a functional inhibitor of AR. APA repressed R1881 (the synthetic androgen methyltrienolone)-induced androgen-regulated gene expression and dramatically inhibited R1881-induced prostate-specific antigen levels. However, APA did not act as an AR antagonist and did not inhibit AR transcriptional activity. Moreover, AS2405, an APA derivative, significantly inhibited the growth of VCaP cells in SCID mice upon oral administration.
Assuntos
Antineoplásicos/uso terapêutico , Cinamatos/uso terapêutico , Oligopeptídeos/uso terapêutico , Neoplasias da Próstata/tratamento farmacológico , Antineoplásicos/química , Antineoplásicos/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cinamatos/química , Cinamatos/metabolismo , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Masculino , Estrutura Molecular , Oligopeptídeos/química , Oligopeptídeos/metabolismo , Streptomyces/metabolismoRESUMO
Cancer cells reprogram their metabolism to survive and grow. Small-molecule inhibitors targeting cancer are useful for studying its metabolic pathways and functions and for developing anticancer drugs. Here, we discovered that glutipyran and its derivatives inhibit glycolytic activity and cell growth in human pancreatic cancer cells. According to proteomic profiling of glutipyran-treated cells using our ChemProteoBase, glutipyran was clustered within the group of endoplasmic reticulum (ER) stress inducers that included glycolysis inhibitors. Glutipyran inhibited glucose uptake and suppressed the growth of various cancer cells, including A431 cells that express glucose transporter class I (GLUT1) and DLD-1 GLUT1 knockout cells. When cotreated with the mitochondrial respiration inhibitor metformin, glutipyran exhibited a synergistic antiproliferative effect. Metabolome analysis revealed that glutipyran markedly decreased most metabolites of the glycolytic pathway and the pentose phosphate pathway. Glutipyran significantly suppressed tumor growth in a xenograft mouse model of pancreatic cancer. These results suggest that glutipyran acts as a broad-spectrum GLUT inhibitor and reduces cancer cell growth.
Assuntos
Antineoplásicos/uso terapêutico , Proteínas Facilitadoras de Transporte de Glucose/antagonistas & inibidores , Neoplasias/tratamento farmacológico , Piranos/uso terapêutico , Animais , Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Sinergismo Farmacológico , Feminino , Glucose/metabolismo , Glicólise/efeitos dos fármacos , Humanos , Metabolômica , Metformina/uso terapêutico , Camundongos Endogâmicos BALB C , Camundongos Nus , Proteômica , Piranos/síntese química , Piranos/farmacologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Dihydroorotate dehydrogenase (DHODH) catalyzes the rate-limiting step in de novo pyrimidine biosynthesis and is a promising cancer treatment target. This study reports the identification of indoluidin D and its derivatives as inhibitors of DHODH. Cell-based phenotypic screening revealed that indoluidin D promoted myeloid differentiation and inhibited the proliferation of acute promyelocytic leukemia HL-60 cells. Indoluidin D also suppressed cell growth in various other types of cancer cells. Cancer cell sensitivity profiling with JFCR39 and proteomic profiling with ChemProteoBase revealed that indoluidin D is a DHODH inhibitor. Indoluidin D inhibited human DHODH activity in vitro; the DHODH reaction product orotic acid rescued indoluidin D-induced cell differentiation. We synthesized several indoluidin D diastereomer derivatives and demonstrated that stereochemistry was vital to their molecular activity. The indoluidin D derivative indoluidin E showed similar activity to its parent compound and suppressed tumor growth in a murine lung cancer xenograft model. Hence, indoluidin D and its derivatives selectively inhibit DHODH and suppress cancer cell growth.
Assuntos
Antineoplásicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Di-Hidro-Orotato Desidrogenase/antagonistas & inibidores , Inibidores Enzimáticos/farmacologia , Animais , Antineoplásicos/química , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Bases de Dados de Proteínas , Inibidores Enzimáticos/química , Humanos , Camundongos , Proteômica , Estereoisomerismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The disruption of the tumor microenvironment (TME) is a promising anti-cancer strategy, but its effective targeting for solid tumors remains unknown. Here, we investigated the anti-cancer activity of the mitochondrial complex I inhibitor intervenolin (ITV), which modulates the TME independent of energy depletion. By modulating lactate metabolism, ITV induced the concomitant acidification of the intra- and extracellular environment, which synergistically suppressed S6K1 activity in cancer cells through protein phosphatase-2A-mediated dephosphorylation via G-protein-coupled receptor(s). Other complex I inhibitors including metformin and rotenone were also found to exert the same effect through an energy depletion-independent manner as ITV. In mouse and patient-derived xenograft models, ITV was found to suppress tumor growth and its mode of action was further confirmed. The TME is usually acidic owing to glycolytic cancer cell metabolism, and this condition is more susceptible to complex I inhibitors. Thus, we have demonstrated a potential treatment strategy for solid tumors.
RESUMO
Targeting stroma in tumor tissues is an attractive new strategy for cancer treatment. We developed in vitro coculture system, in which the growth of human prostate cancer DU-145 cells is stimulated by prostate stromal cells (PrSC) through insulin-like growth factor I (IGF-I). Using this system, we have been searching for small molecules that inhibit tumor growth through modulation of tumor-stromal cell interactions. As a result, we have found that leucinostatins and atpenins, natural antifungal antibiotics, inhibit the growth of DU-145 cells cocultured with PrSC more strongly than that of DU-145 cells alone. In this study we examined the antitumor effects of these small molecules in vitro and in vivo. When DU-145 cells were coinoculated with PrSC subcutaneously in nude mice, leucinostatin A was found to significantly suppress the tumor growth more than atpenin B. The antitumor effect of leucinostatin A in vivo was not obtained against the tumors of DU-145 cells alone. RT-PCR experiments revealed that leucinostatin A specifically inhibited IGF-I expression in PrSC without effect on expressions of other IGF axis molecules. Leucinostatins and atpenins are known to abrogate mitochondrial functions. However, when we used mitochondrial DNA-depleted, pseudo-rho(0) cells, we found that one of leucinostain A actions certainly depended on mitochondrial function, but it actually inhibited the growth of DU-145 cells more strongly in coculture with pseudo-rho(0) PrSC and reduced IGF-I expression in pseudo-rho(0) PrSC. Taken together, our results suggested that leucinostatin A inhibited prostate cancer cell growth through reduction of IGF-I expression in PrSC.