RESUMO
Small-molecule inhibitors of MDM2 that block the MDM2-p53 protein-protein interaction have been considered as potential therapeutic agents for the treatment of cancer. Here, we identify five highly potent inhibitors of MDM2 (termed as WY 1-5) that display significant inhibitory effects on MDM2-p53 interaction by using a combined strategy of pharmacophore modeling, virtual screening, and molecular docking studies. Among them, WY-5 is the most active MDM2 inhibitor with an IC50 value of 14.1±2.8â nM. Moreover, WY-5 significantly up-regulate the protein level of p53 in SK-Hep-1 cells harboring wild-type p53. In vitro anticancer study reveals that WY-5 markedly inhibits the survival of SK-Hep-1 cells. In vivo anticancer study suggests that WY-5 significantly inhibits the growth of SK-Hep-1 cells-derived xenograft in nude mice, with no observable toxicity. Our results demonstrate that WY-5 may be a promising candidate for the treatment of cancer harboring wild-type p53.
Assuntos
Antineoplásicos/farmacologia , Descoberta de Drogas , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Avaliação Pré-Clínica de Medicamentos , Feminino , Humanos , Neoplasias Hepáticas Experimentais/tratamento farmacológico , Neoplasias Hepáticas Experimentais/metabolismo , Neoplasias Hepáticas Experimentais/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Modelos Moleculares , Estrutura Molecular , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Relação Estrutura-AtividadeRESUMO
PURPOSE: Glioblastoma (GBM) is a malignant brain tumor with a poor long-term prognosis due to recurrence from highly resistant GBM cancer stem cells (CSCs), for which the current standard of treatment with temozolomide (TMZ) alone will unlikely produce a viable cure. In addition, CSCs regenerate rapidly and overexpress methyl transferase which overrides the DNA-alkylating mechanism of TMZ, leading to resistance. The objective of this research was to apply the concepts of nanotechnology to develop a multi-drug therapy, TMZ and idasanutlin (RG7388, a potent mouse double minute 2 (MDM2) antagonist), loaded in functionalized nanoparticles (NPs) that target the GBM CSC subpopulation, reduce the cell viability and provide possibility of in vivo preclinical imaging. METHODS: Polymer-micellar NPs composed of poly(styrene-b-ethylene oxide) (PS-b-PEO) and poly(lactic-co-glycolic) acid (PLGA) were developed by a double emulsion technique loading TMZ and/or RG7388. The NPs were covalently bound to a 15-nucleotide base-pair CD133 aptamer to target the CD133 antigen expressed on the surfaces of GBM CSCs. For diagnostic functionality, the NPs were labelled with radiotracer Zirconium-89 (89Zr). RESULTS: NPs maintained size range less than 100 nm, a low negative charge and exhibited the ability to target and kill the CSC subpopulation when TMZ and RG7388 were used in combination. The targeting function of CD133 aptamer promoted killing in GBM CSCs providing impetus for further development of targeted nanosystems for localized therapy in future in vivo models. CONCLUSIONS: This work has provided a potential clinical application for targeting GBM CSCs with simultaneous diagnostic imaging.
Assuntos
Antígeno AC133/metabolismo , Neoplasias Encefálicas/metabolismo , Sistemas de Liberação de Medicamentos/métodos , Glioblastoma/metabolismo , Nanopartículas/metabolismo , Células-Tronco Neoplásicas/metabolismo , Animais , Neoplasias Encefálicas/tratamento farmacológico , Linhagem Celular Tumoral , Relação Dose-Resposta a Droga , Desenvolvimento de Medicamentos/métodos , Avaliação Pré-Clínica de Medicamentos/métodos , Glioblastoma/tratamento farmacológico , Humanos , Camundongos , Micelas , Nanopartículas/administração & dosagem , Células-Tronco Neoplásicas/efeitos dos fármacos , Polímeros/administração & dosagem , Polímeros/metabolismo , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Pirrolidinas/administração & dosagem , Pirrolidinas/metabolismo , Temozolomida/administração & dosagem , Temozolomida/metabolismo , para-Aminobenzoatos/administração & dosagem , para-Aminobenzoatos/metabolismoRESUMO
BACKGROUND: Lung cancer is a leading fatal malignancy due to the high incidence of treatment failure. Dysfunction of the tumor suppressor p53 contributes to cancer initiation, progression, and therapeutic resistance. Targeting MDM2, a negative regulator of p53, has recently attracted interest in cancer drug research as it may restore tumor suppressive function. PURPOSE: The present study aimed to investigate the effect of 3,4-dihydroxy-5,4'-dimethoxybibenzyl (DS-1) on targeting MDM2 and restoring p53 function in lung cancer cells. METHODS: The efficacy of DS-1 alone or in combination with cisplatin in lung cancer cells was determined by MTT, nuclear staining, and annexin V/PI assay. The expression of apoptosis-related proteins was determined by western blot analysis. To evaluate the role of DS-1 on the stabilization and degradation of p53, cycloheximide chasing assay and immunoprecipitation were conducted, and the active form of p53 was investigated by immunofluorescent staining assay. To confirm and demonstrate the site interaction between DS-1 and the MDM2 protein, in silico computational analysis was performed. RESULTS: DS-1 exhibited a cytotoxic effect and sensitized lung cancer cells to cisplatin-induced apoptosis. DS-1 caused a significant increase in the cellular level of p53 protein, while the active form of p53 (phosphorylation at Ser15) was unaltered. DS-1 treatment in combination with cisplatin could enhance activated p-p53 (Ser15) and p53 downstream signaling (Bax, Bcl-2, and Akt), leading to a higher level of apoptosis. Immunoprecipitation analysis revealed that DS-1 decreased the p53-ubiquitin complex, a prerequisite step in p53 proteasomal degradation. Molecular docking simulation further evidenced that DS-1 interacts with MDM2 within the p53-binding domain by carbon-hydrogen bond interaction at Lys27, π-alkyl interactions at Ile37 and Leu30, and van der Waals interactions at Ile75, Val51, Val69, Phe67, Met38, Tyr43, Gly34, and Phe31. Treatment by DS-1 and cisplatin in patient-derivated primary lung cancer cells showed consistent effects by increasing cisplatin sensitivity. CONCLUSIONS: Our findings provide evidence that DS-1 is an MDM2 inhibitor and its underlying mechanism involves MDM2 binding and p53 induction, which may benefit the development of this compound for lung cancer treatment.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Bibenzilas/farmacologia , Neoplasias Pulmonares/patologia , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Proteína Supressora de Tumor p53/metabolismo , Adulto , Idoso , Linhagem Celular Tumoral , Cisplatino/farmacologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Simulação de Acoplamento Molecular , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Transdução de Sinais/efeitos dos fármacosRESUMO
Modification-dependent and -independent biomolecular interactions, including protein-protein, protein-DNA/RNA, protein-sugar, and protein-lipid interactions, play crucial roles in all cellular processes. Dysregulation of these biomolecular interactions or malfunction of the associated enzymes results in various diseases; therefore, these interactions and enzymes are attractive targets for therapies. High-throughput screening can greatly facilitate the discovery of drugs for these targets. Here, we describe a biomolecular interaction detection method, called phase-separated condensate-aided enrichment of biomolecular interactions in test tubes (CEBIT). The readout of CEBIT is the selective recruitment of biomolecules into phase-separated condensates harboring their cognate binding partners. We tailored CEBIT to detect various biomolecular interactions and activities of biomolecule-modifying enzymes. Using CEBIT-based high-throughput screening assays, we identified known inhibitors of the p53/MDM2 (MDM2) interaction and of the histone methyltransferase, suppressor of variegation 3-9 homolog 1 (SUV39H1), from a compound library. CEBIT is simple and versatile, and is likely to become a powerful tool for drug discovery and basic biomedical research.
Assuntos
Avaliação Pré-Clínica de Medicamentos/métodos , Ensaios de Triagem em Larga Escala/métodos , Mapeamento de Interação de Proteínas/métodos , Descoberta de Drogas/métodos , Humanos , Metiltransferases/antagonistas & inibidores , Metiltransferases/metabolismo , Transição de Fase , Mapas de Interação de Proteínas/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Proteínas Repressoras/antagonistas & inibidores , Proteínas Repressoras/metabolismo , Proteína Supressora de Tumor p53/antagonistas & inibidores , Proteína Supressora de Tumor p53/metabolismoRESUMO
AIMS: Inhibition of P53-MDM2/X interaction is known as an effective cancer therapy strategy. In this regard, pDI peptide was introduced previously with the potential of targeting MDM2. In this research, the large-scale peptide mutation screening was used to achieve the best sequence of pDI with the highest affinity for inhibition activity against MDM2/X. MAIN METHODS: Three mutant peptides of pDI as dual inhibitor peptides including single mutations of pDIm/4W, pDIm/11M and double mutations of pDIdm/4W11M were presented with the high affinities to inhibit both MDM2/X. The selected mutants were then evaluated comprehensively to confirm their ability as potent MDM2/X inhibitors, using a theoretical simulation approach. KEY FINDINGS: MD simulations analyses confirmed their dual inhibition potential against both MDM2/X interactions with p53 protein. The developed pDIm and mainly pDIdm peptides showed stable conformations over the simulation time with conserved secondary structure and effective interaction with MDM2/X by physical binding such as hydrogen bonding. Besides, umbrella sampling free energy calculation indicated higher binding energy, ΔGbinding, of pDIm-MDM2/X and pDIdm-MDM2/X compared to pDI-MDM2/X. SIGNIFICANCE: The optimized and improved mutant pDI, pDIdm, with more effective ΔGbinding values of -30 and -25 kcal/mol to MDMX and MDM2, respectively, is recommended as a promising anticancer agent and suitable candidate for experimental evaluations.
Assuntos
Antineoplásicos/farmacologia , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Avaliação Pré-Clínica de Medicamentos , Humanos , Ligação Proteica/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Relação Estrutura-Atividade , Proteína Supressora de Tumor p53/antagonistas & inibidoresRESUMO
INTRODUCTION: Uveal melanoma (UM) is a rare and malignant intraocular tumour with a dismal prognosis. Despite a good control of the primary tumour by radiation or surgery, up to 50% of patients subsequently develop metastasis for which no efficient treatment is yet available. METHODOLOGY: To identify therapeutic opportunities, we performed an in vitro screen of 30 combinations of different inhibitors of pathways that are dysregulated in UM. Effects of drug combinations on viability, cell cycle and apoptosis were assessed in eight UM cell lines. The best synergistic combinations were further evaluated in six UM patient-derived xenografts (PDXs). RESULTS: We demonstrated that the Bcl-2/XL/W inhibitor (ABT263) sensitised the UM cell lines to other inhibitors, mainly to mammalian target of rapamycin (mTOR), mitogen-activated protein kinase kinase (MEK) and murine double minute 2 (MDM2) inhibitors. mTOR (RAD001) and MEK1/2 (trametinib) inhibitors were efficient as single agents, but their combinations with ABT263 displayed no synergism in UM PDXs. In contrast, the combination of ABT263 with MDM2 inhibitor (HDM201) showed a trend for a synergistic effect. CONCLUSION: We showed that inhibition of Bcl-2/XL/W sensitised the UM cell lines to other treatments encouraging investigation of the underlying mechanisms. Furthermore, our findings highlighted Bcl-2/XL/W and MDM2 co-inhibition as a promising strategy in UM.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Avaliação Pré-Clínica de Medicamentos/métodos , Melanoma/tratamento farmacológico , Neoplasias Uveais/tratamento farmacológico , Compostos de Anilina/administração & dosagem , Animais , Apoptose/efeitos dos fármacos , Proteínas Reguladoras de Apoptose/antagonistas & inibidores , Proteínas Reguladoras de Apoptose/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Combinação de Medicamentos , Everolimo/administração & dosagem , Humanos , Imidazóis/administração & dosagem , Melanoma/metabolismo , Melanoma/patologia , Camundongos , Quinases de Proteína Quinase Ativadas por Mitógeno/antagonistas & inibidores , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Piridonas/administração & dosagem , Pirimidinas/administração & dosagem , Pirimidinonas/administração & dosagem , Pirróis/administração & dosagem , Sulfonamidas/administração & dosagem , Serina-Treonina Quinases TOR/antagonistas & inibidores , Serina-Treonina Quinases TOR/metabolismo , Neoplasias Uveais/metabolismo , Neoplasias Uveais/patologia , Ensaios Antitumorais Modelo de Xenoenxerto/métodos , Proteína bcl-X/antagonistas & inibidores , Proteína bcl-X/metabolismoRESUMO
This study explored the probable in vivo cardiac and renal toxicities together with in silico approaches for predicting the apoptogenic potential of Euphorbia peplus methanolic extract (EPME) in rats. Cardiac and renal injury biomarkers were estimated with histopathological and immunohistochemical evaluations of both kidney and heart. The probable underlying mechanism of E. peplus compounds to potentiate p53 activity is examined using Molecular Operating Environment (MOE) docking software and validated experimentally by immunohistochemical localization of p53 protein in the kidney and heart tissues. The gas chromatography/mass spectrometry analysis of E. peplus revealed the presence of nine different compounds dominated by di-(2-ethylhexyl) phthalate (DEHP). Significant elevations of troponin, creatine phosphokinase, creatine kinase-myocardium bound, lactate dehydrogenase, aspartate transaminase, alkaline phosphatase, urea, creatinine, and uric acid were evident in the EPME treated rats. The EPME treated rats showed strong renal and cardiac p53 expression and moderate cardiac TNF-α expression. Further, our in silico results predicted the higher affinity and good inhibition of DEHP, glyceryl linolenate, and lucenin 2 to the MDM2-p53 interface compared to the standard reference 15 a compound. Conclusively, EPME long-term exposure could adversely affect the cardiac and renal tissues probably due to their inflammatory and apoptotic activity. Moreover, the in silico study hypothesizes that EPME inhibits MDM2-mediated degradation of p53 suggesting possible anticancer potentials which confirmed experimental by strong p53 expression in renal and cardiac tissues.
Assuntos
Apoptose/efeitos dos fármacos , Euphorbia/química , Coração/efeitos dos fármacos , Rim/efeitos dos fármacos , Extratos Vegetais/toxicidade , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Proteína Supressora de Tumor p53/antagonistas & inibidores , Animais , Biomarcadores/sangue , Sobrevivência Celular/efeitos dos fármacos , Simulação por Computador , Rim/metabolismo , Rim/patologia , Masculino , Simulação de Acoplamento Molecular , Miocárdio/metabolismo , Miocárdio/patologia , Extratos Vegetais/isolamento & purificação , Ratos , Ratos WistarRESUMO
BACKGROUND: Neural tube defects (NTDs) are common congenital malformations resulting in failure of the neural tube closure during early embryonic development. Although it is known that maternal folate deficiency increases the risk of NTDs, the mechanism remains elusive. RESULTS: Herein, we report that histone H2A monoubiquitination (H2AK119ub1) plays a role in neural tube closure. We found that the folate antagonist methotrexate induced H2AK119ub1 in mouse embryonic stem cells. We demonstrated that an increase in H2AK119ub1 downregulated expression of the neural tube closure-related genes Cdx2, Nes, Pax6, and Gata4 in mouse embryonic stem cells under folate deficiency conditions. We also determined that the E3 ligase Mdm2 was responsible for the methotrexate-induced increase in H2AK119ub1 and downregulation of neural tube closure-related genes. Surprisingly, we found that Mdm2 is required for MTX-induced H2A ubiquitination and is recruited to the sites of DSB, which is dependent on DNA damage signaling kinase ATM. Furthermore, folic acid supplementation restored H2AK119ub1 binding to neural tube closure-related genes. Downregulation of these genes was also observed in both brain tissue of mouse and human NTD cases, and high levels of H2AK119ub1 were found in the corresponding NTDs samples with their maternal serum folate under low levels. Pearson correlation analysis showed a significant negative correlation between expression of the neural precursor genes and H2AK119ub1. CONCLUSION: Our results indicate that folate deficiency contributes to the onset of NTDs by altering H2AK119ub1 and subsequently affecting expression of neural tube closure-related genes. This may be a potential risk factor for NTDs in response to folate deficiency.
Assuntos
Regulação para Baixo , Histonas/metabolismo , Defeitos do Tubo Neural/patologia , Animais , Proteínas Mutadas de Ataxia Telangiectasia/antagonistas & inibidores , Proteínas Mutadas de Ataxia Telangiectasia/genética , Proteínas Mutadas de Ataxia Telangiectasia/metabolismo , Fator de Transcrição CDX2/genética , Fator de Transcrição CDX2/metabolismo , Dano ao DNA , Regulação para Baixo/efeitos dos fármacos , Desenvolvimento Embrionário/efeitos dos fármacos , Células-Tronco Embrionárias/citologia , Células-Tronco Embrionárias/metabolismo , Ácido Fólico/metabolismo , Ácido Fólico/farmacologia , Ácido Fólico/uso terapêutico , Metotrexato/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Defeitos do Tubo Neural/metabolismo , Defeitos do Tubo Neural/prevenção & controle , Fator de Transcrição PAX6/genética , Fator de Transcrição PAX6/metabolismo , Ligação Proteica , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-mdm2/genética , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Interferência de RNA , RNA Interferente Pequeno/metabolismo , UbiquitinaçãoRESUMO
BACKGROUND: Mouse Double Minute 2 protein (MDM2) is a cellular regulator of p53 tumor suppressor (p53). Inhibition of the interaction between MDM2 and p53 proteins is a promising anticancer therapy. OBJECTIVE: This updated patent review is an attempt to compile the research and achievements of the various researchers working on small molecule MDM2 inhibitors from 2010 to date. We provide an outlook into the future for therapy based on MDM2 inhibition by presenting an overview of the most relevant patents which have recently appeared in the literature. METHODS: Literature and recent patents focusing on the anticancer potential of MDM2-p53 interaction inhibitors and its applications have been analyzed. We put the main emphasis on the most perspective compounds which are or were examined in clinical trials. RESULTS: Literature data indicated that MDM2 inhibitors are therapeutically effective in specific types of cancer or non-cancer diseases. A great number of patents and research work around new MDM2- p53 interaction inhibitors, possible combinations, new indications, clinical regimens in previous years prove that this targeted therapy is in the scope of interest for many business and academic research groups. CONCLUSION: Novel MDM2 inhibitors thanks to higher potency and better ADME properties have shown effectiveness in preclinical and clinical development however the final improvement of therapeutic potential for MDM2 inhibitors might depend on the useful combination therapy and exploring new cancer and non-cancer indications.
Assuntos
Antineoplásicos/uso terapêutico , Neoplasias/tratamento farmacológico , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Proteína Supressora de Tumor p53/antagonistas & inibidores , Animais , Antineoplásicos/farmacologia , Ensaios Clínicos como Assunto , Avaliação Pré-Clínica de Medicamentos , Humanos , Terapia de Alvo Molecular , Neoplasias/metabolismo , Neoplasias/patologia , Patentes como Assunto , Domínios e Motivos de Interação entre ProteínasRESUMO
INTRODUCTION: PI3K pathway signaling has received attention as a molecular target in clear cell ovarian carcinoma (CCOC). MDM2 is one of the AKT effectors in the PI3K pathway, which binds to and degrades p53. In this study, we aimed to clarify the prognostic significance of PIK3CA and MDM2 expression, and potential therapeutic effect of a dual inhibition of the PI3K pathway and MDM2. MATERIALS AND METHODS: cDNA expression was evaluated by using microarray data using 75 samples of CCOC. DS-7423 (dual inhibitor of pan-PI3K and mTOR) and RG7112 (MDM2 inhibitor) were used on CCOC cell lines to evaluate cell proliferation, expression level of MDM2 related proteins, and apoptosis by MTT assay, western blotting, and flow cytometry. DS-7423 (3â¯mg/kg) and/or RG7112 (50â¯mg/kg) were orally administrated every day for three weeks, and the anti-tumor effect was evaluated using tumor xenografts, along with immunohistochemistry. RESULTS: Tumors with high expression of both PIK3CA and MDM2 showed significantly worse prognosis in expression array of 71 CCOCs (Pâ¯=â¯0.013). Dual inhibition of the PI3K pathway by DS-7423 and MDM2 by RG7112 showed synergistic anti-proliferative effect in 4 CCOC cell lines without TP53 mutations. The combination therapy more robustly induced pro-apoptotic proteins (PUMA and cleaved PARP) with increase of sub G1 population and apoptotic cells, compared with either single agent alone. The combination therapy significantly reduced tumor volume in mice (Pâ¯<â¯0.001 in OVISE, and Pâ¯=â¯0.038 in RMG-I) without severe body weight loss. Immunohistochemistry from the xenograft tumors showed that the combination treatment significantly reduced vascularity and cell proliferation, with an increase of apoptotic cell death. CONCLUSION: A combination therapy targeting the PI3K pathway and MDM2 might be a promising therapeutic strategy in CCOC.
Assuntos
Neoplasias Ovarianas/tratamento farmacológico , Inibidores de Fosfoinositídeo-3 Quinase , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Adenina/análogos & derivados , Adenina/farmacologia , Adenocarcinoma de Células Claras , Animais , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Classe I de Fosfatidilinositol 3-Quinases , DNA Complementar/metabolismo , Feminino , Xenoenxertos , Imidazolinas/farmacologia , Camundongos Nus , Transplante de Neoplasias/fisiologia , Neoplasias Ovarianas/metabolismo , Piperazinas/farmacologia , Inibidores de Proteínas Quinases/farmacologia , RNA Mensageiro/metabolismo , RNA Neoplásico/metabolismo , Distribuição AleatóriaRESUMO
The p53 protein, also called guardian of the genome, plays a critical role in the cell cycle regulation and apoptosis. This protein is frequently inactivated in several types of human cancer by abnormally high levels of its negative regulator, mouse double minute 2 (MDM2). As a result, restoration of p53 function by inhibiting p53-MDM2 protein-protein interaction has been pursued as a compelling strategy for cancer therapy. To date, a limited number of small-molecules have been reported as effective p53-MDM2 inhibitors. X-ray structures of MDM2 in complex with some ligands are available in Protein Data Bank and herein, these data have been exploited to efficiently identify new p53-MDM2 interaction antagonists through a hierarchical virtual screening strategy. For this purpose, the first step was aimed at compiling a focused library of 686,630 structurally suitable compounds, from PubChem database, similar to two known effective inhibitors, Nutlin-3a and DP222669. These compounds were subjected to the subsequent structure-based approaches (quantum polarized ligand docking and molecular dynamics simulation) to select potential compounds with highest binding affinity for MDM2 protein. Additionally, ligand binding energy, ADMET properties and PAINS analysis were also considered as filtering criteria for selecting the most promising drug-like molecules. On the basis of these analyses, three top-ranked hit molecules, CID_118439641, CID_60452010 and CID_3106907, were found to have acceptable pharmacokinetics properties along with superior in silico inhibitory ability towards the p53-MDM2 interaction compared to known inhibitors. Molecular docking and molecular dynamics results well confirmed the interactions of the final selected compounds with critical residues within p53 binding site on the MDM2 hydrophobic clefts with satisfactory thermodynamics stability. Consequently, the new final scaffolds identified by the presented computational approach could offer a set of guidelines for designing promising anti-cancer agents targeting p53-MDM2 interaction.
Assuntos
Simulação por Computador , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Teoria Quântica , Bibliotecas de Moléculas Pequenas/farmacologia , Proteína Supressora de Tumor p53/antagonistas & inibidores , Animais , Cristalografia por Raios X , Avaliação Pré-Clínica de Medicamentos , Ligantes , Camundongos , Modelos Moleculares , Estrutura Molecular , Ligação Proteica/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-mdm2/química , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Bibliotecas de Moléculas Pequenas/química , Proteína Supressora de Tumor p53/química , Proteína Supressora de Tumor p53/metabolismoRESUMO
Mutations at the TP53 gene are readily detected (approximately 50-75%) in pancreatic ductal adenocarcinoma (PDAC) patients. TP53 was previously thought to be a difficult target as it is often mutated, deleted or inactivated on both chromosomes in certain cancers. In the following study, the effects of restoration of wild-type (WT) TP53 activity on the sensitivities of MIA-PaCa-2 pancreatic cancer cells to the MDM2 inhibitor nutlin-3a in combination with chemotherapy, targeted therapy, as well as, nutraceuticals were examined. Upon introduction of the WT-TP53 gene into MIA-PaCa-2â¯cells, which contain a TP53 gain of function (GOF) mutation, the sensitivity to the MDM2 inhibitor increased. However, effects of nutlin-3a were also observed in MIA-PaCa-2â¯cells lacking WT-TP53, as upon co-treatment with nutlin-3a, the sensitivity to certain inhibitors, chemotherapeutic drugs and nutraceuticals increased. Interestingly, co-treatment with nutlin-3a and certain chemotherapeutic drug such as irinotecan and oxaliplatin resulted in antagonistic effects in cells both lacking and containing WT-TP53 activity. These studies indicate the sensitizing abilities that WT-TP53 activity can have in PDAC cells which normally lack WT-TP53, as well as, the effects that the MDM2 inhibitor nutlin-3a can have in both cells containing and lacking WT-TP53 to various therapeutic agents.
Assuntos
Antineoplásicos/farmacologia , Carcinoma Ductal Pancreático/metabolismo , Imidazóis/farmacologia , Neoplasias Pancreáticas/metabolismo , Piperazinas/farmacologia , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Proteína Supressora de Tumor p53/metabolismo , Carcinoma Ductal Pancreático/tratamento farmacológico , Carcinoma Ductal Pancreático/genética , Linhagem Celular Tumoral , Suplementos Nutricionais/análise , Humanos , Irinotecano/farmacologia , Oxaliplatina/farmacologia , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/genética , Proteínas Proto-Oncogênicas c-mdm2/genética , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Transdução de Sinais/efeitos dos fármacos , Proteína Supressora de Tumor p53/genéticaRESUMO
The ubiquitin/proteasome system is a primary conduit for selective intracellular protein degradation. Since its discovery over 30 years ago, this highly regulated system continues to be an active research area for drug discovery that is exemplified by several approved drugs. Here we review compounds in preclinical testing, clinical trials, and approved drugs, with the aim of highlighting innovative discoveries and breakthrough therapies that target the ubiquitin system.
Assuntos
Inibidores Enzimáticos/química , Complexo de Endopeptidases do Proteassoma/metabolismo , Ubiquitina/metabolismo , Avaliação Pré-Clínica de Medicamentos , Inibidores Enzimáticos/metabolismo , Humanos , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Ubiquitina/química , Enzimas Ativadoras de Ubiquitina/química , Enzimas Ativadoras de Ubiquitina/metabolismo , Enzimas de Conjugação de Ubiquitina/química , Enzimas de Conjugação de Ubiquitina/metabolismo , Ubiquitina-Proteína Ligases/química , Ubiquitina-Proteína Ligases/metabolismo , UbiquitinaçãoRESUMO
BACKGROUND: The tumor suppressor protein p53 plays an important role in preventing tumor formation and progression through its involvement in cell division control and initiation of apoptosis. Mdm2 protein controls the activity of p53 protein through working as ubiquitin E3 ligase promoting p53 degradation through the proteasome degradation pathway. Inhibitors for Mdm2-p53 interaction have restored the activity of p53 protein and induced cancer fighting properties in the cell. PURPOSE: The objective of this study is to use computer-aided drug discovery techniques to search for new Mdm2-p53 interaction inhibitors. METHODS: A set of pharmacophoric features were created based on a standard Mdm2 inhibitor and this was used to screen a commercial drug-like ligand library; then potential inhibitors were docked and ranked in a multi-step protocol using GLIDE. Top ranked ligands from docking were evaluated for their inhibition activity of Mdm2-p53 interaction using ELISA testing. RESULTS: Several compounds showed inhibition activity at the submicromolar level, which is comparable to the standard inhibitor Nutlin-3a. Furthermore, the discovered inhibitors were evaluated for their anticancer activities against different breast cancer cell lines, and they showed an interesting inhibition pattern. CONCLUSION: The reported inhibitors can represent a starting point for further SAR studies in the future and can help in the discovery of new anticancer agents.
Assuntos
Antineoplásicos/farmacologia , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Pirimidinas/farmacologia , Triazóis/farmacologia , Proteína Supressora de Tumor p53/antagonistas & inibidores , Antineoplásicos/síntese química , Antineoplásicos/química , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Desenho Assistido por Computador , Descoberta de Drogas , Avaliação Pré-Clínica de Medicamentos , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Modelos Moleculares , Estrutura Molecular , Proteínas Proto-Oncogênicas c-mdm2/química , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Pirimidinas/síntese química , Pirimidinas/química , Triazóis/síntese química , Triazóis/química , Proteína Supressora de Tumor p53/química , Proteína Supressora de Tumor p53/metabolismoRESUMO
T- and NK-cell lymphomas (TCL) are a heterogenous group of lymphoid malignancies with poor prognosis. In contrast to B-cell and myeloid malignancies, there are few preclinical models of TCLs, which has hampered the development of effective therapeutics. Here we establish and characterize preclinical models of TCL. We identify multiple vulnerabilities that are targetable with currently available agents (e.g., inhibitors of JAK2 or IKZF1) and demonstrate proof-of-principle for biomarker-driven therapies using patient-derived xenografts (PDXs). We show that MDM2 and MDMX are targetable vulnerabilities within TP53-wild-type TCLs. ALRN-6924, a stapled peptide that blocks interactions between p53 and both MDM2 and MDMX has potent in vitro activity and superior in vivo activity across 8 different PDX models compared to the standard-of-care agent romidepsin. ALRN-6924 induced a complete remission in a patient with TP53-wild-type angioimmunoblastic T-cell lymphoma, demonstrating the potential for rapid translation of discoveries from subtype-specific preclinical models.
Assuntos
Antineoplásicos/farmacologia , Regulação Neoplásica da Expressão Gênica , Linfoma Extranodal de Células T-NK/tratamento farmacológico , Linfoma de Células T/tratamento farmacológico , Proteínas Nucleares/genética , Peptídeos/farmacologia , Proteínas Proto-Oncogênicas c-mdm2/genética , Proteínas Proto-Oncogênicas/genética , Proteína Supressora de Tumor p53/genética , Animais , Proteínas de Ciclo Celular , Depsipeptídeos/farmacologia , Avaliação Pré-Clínica de Medicamentos , Humanos , Fator de Transcrição Ikaros/antagonistas & inibidores , Fator de Transcrição Ikaros/genética , Fator de Transcrição Ikaros/metabolismo , Imidazolinas/farmacologia , Janus Quinase 2/antagonistas & inibidores , Janus Quinase 2/genética , Janus Quinase 2/metabolismo , Linfoma Extranodal de Células T-NK/genética , Linfoma Extranodal de Células T-NK/metabolismo , Linfoma Extranodal de Células T-NK/patologia , Linfoma de Células T/genética , Linfoma de Células T/metabolismo , Linfoma de Células T/patologia , Camundongos , Proteínas Nucleares/antagonistas & inibidores , Proteínas Nucleares/metabolismo , Ligação Proteica/efeitos dos fármacos , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Indução de Remissão , Transdução de Sinais , Proteína Supressora de Tumor p53/antagonistas & inibidores , Proteína Supressora de Tumor p53/metabolismo , Sequenciamento do Exoma , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The biological activity of curcumin (CUR), a promising naturally occurring dietary compound for the treatment of hepatocellular carcinoma (HCC), was closely associated with its metabolite. Octahydrocurcumin (OHC) is the final hydrogenated metabolite of CUR and has been reported to have potential biological activities. However, difficulties in access have hampered its biological studies. In the current investigation, we designed an efficient synthesis method to produce OHC, and comparatively explored the anti-cancer effect and potential mechanism of OHC and CUR in an H22 ascites tumor-bearing mice model. The results indicated that OHC had a relatively wide margin of safety, and exhibited superior effects to CUR in suppressing the tumor growth, including ascending weight, abdominal circumference, ascites volume and cancer cell viability. OHC significantly induced H22 cell apoptosis by upregulating the p53 expression and downregulating the MDM2 expression. OHC also remarkably decreased the Bcl-2 and Bcl-xl protein expressions, and increased the Bax and Bad expressions in ascitic cells. Furthermore, THC substantially induced the release of cytochrome C, caspase-3, caspase-9 and the cleavage of PARP to induce H22 cell apoptosis. Taken together, OHC was more effective than CUR in suppressing H22-induced HCC through the activation of the mitochondrial apoptosis pathway. OHC may thus be a promising anti-HCC agent.
Assuntos
Antineoplásicos Fitogênicos/uso terapêutico , Apoptose , Carcinoma Hepatocelular/dietoterapia , Curcumina/análogos & derivados , Neoplasias Hepáticas Experimentais/dietoterapia , Animais , Animais não Endogâmicos , Antineoplásicos Fitogênicos/síntese química , Antineoplásicos Fitogênicos/metabolismo , Proteínas Reguladoras de Apoptose/agonistas , Proteínas Reguladoras de Apoptose/antagonistas & inibidores , Proteínas Reguladoras de Apoptose/genética , Proteínas Reguladoras de Apoptose/metabolismo , Biomarcadores/metabolismo , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Sobrevivência Celular , Curcumina/síntese química , Curcumina/metabolismo , Curcumina/uso terapêutico , Regulação Neoplásica da Expressão Gênica , Hidrogenação , Neoplasias Hepáticas Experimentais/metabolismo , Neoplasias Hepáticas Experimentais/patologia , Masculino , Camundongos , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-mdm2/genética , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Distribuição Aleatória , Análise de Sobrevida , Carga Tumoral , Proteína Supressora de Tumor p53/agonistas , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismoRESUMO
In most types of cancer, overexpression of murine double minute 2 (MDM2) often leads to inactivation of p53. The crystal structure of MDM2, with a 109-residue amino-terminal domain, reveals that MDM2 has a core hydrophobic region to which p53 binds as an amphipathic α helix. The interface depends on the steric complementarity between MDM2 and the hydrophobic region of p53. Especially, on p53's triad, amino acids Phe19, Trp23 and Leu26 bind to the MDM2 core. Results from studies suggest that the structural motif of both p53 and MDM2 can be attributed to similarities in the amphipathic α helix. Thus, in the current investigation it is hypothesized that the similarity in the structural motif might be the cause of p53 inactivation by MDM2. Hence, molecular docking and phytochemical screening approaches are appraised to inhibit the hydrophobic cleft of MDM2 and to stop p53-MDM2 interaction, resulting in reactivation of p53 activity. For this purpose, a library of 2295 phytochemicals were screened against p53-MDM2 to find potential candidates. Of these, four phytochemicals including epigallocatechin gallate, alvaradoin M, alvaradoin E and nordihydroguaiaretic acid were found to be potential inhibitors of p53-MDM2 interaction. The screened phytochemicals, derived from natural extracts, may have negligible side effects and can be explored as potent antagonists of p53-MDM2 interactions, resulting in reactivation of the normal transcription of p53.
Assuntos
Antineoplásicos/farmacologia , Extratos Vegetais/farmacologia , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-mdm2/química , Proteína Supressora de Tumor p53/antagonistas & inibidores , Proteína Supressora de Tumor p53/química , Antineoplásicos/química , Ensaios de Seleção de Medicamentos Antitumorais , Interações Hidrofóbicas e Hidrofílicas , Simulação de Acoplamento Molecular , Extratos Vegetais/química , Conformação Proteica , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Proteína Supressora de Tumor p53/metabolismoRESUMO
Inflammation is recently recognized as one of the hallmarks of human cancer. Chronic inflammatory response plays a critical role in cancer development, progression, metastasis, and resistance to chemotherapy. Conversely, the oncogenic aberrations also generate an inflammatory microenvironment, enabling the development and progression of cancer. The molecular mechanisms of action that are responsible for inflammatory cancer and cancer-associated inflammation are not fully understood due to the complex crosstalk between oncogenic and pro-inflammatory genes. However, molecular mediators that regulate both inflammation and cancer, such as NF-κB and STAT have been considered as promising targets for preventing and treating these diseases. Recent works have further demonstrated an important role of oncogenes (e.g., NFAT1, MDM2) and tumor suppressor genes (e.g., p53) in cancer-related inflammation. Natural products that target these molecular mediators have shown anticancer and anti-inflammatory activities in preclinical and clinical studies. Sesquiterpenoids (STs), a class of novel plant-derived secondary metabolites have attracted great interest in recent years because of their diversity in chemical structures and pharmacological activities. At present, we and other investigators have found that dimeric sesquiterpenoids (DSTs) may exert enhanced activity and binding affinity to molecular targets due to the increased number of alkylating centers and improved conformational flexibility and lipophilicity. Here, we focus our discussion on the activities and mechanisms of action of STs and DSTs in treating inflammation and cancer as well as their structure-activity relationships.
Assuntos
Anti-Inflamatórios/farmacologia , Antineoplásicos Fitogênicos/farmacologia , Sesquiterpenos/farmacologia , Animais , Humanos , Inflamação/tratamento farmacológico , Inflamação/etiologia , NF-kappa B/antagonistas & inibidores , Fatores de Transcrição NFATC/antagonistas & inibidores , Neoplasias/tratamento farmacológico , Neoplasias/etiologia , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-mdm2/fisiologia , Sesquiterpenos/química , Relação Estrutura-AtividadeRESUMO
Antigen-activated lymphocytes undergo extraordinarily rapid cell division in the course of immune responses. We hypothesized that this unique aspect of lymphocyte biology leads to unusual genomic stress in recently antigen-activated lymphocytes and that targeted manipulation of DNA damage-response (DDR) signaling pathways would allow for selective therapeutic targeting of pathological T cells in disease contexts. Consistent with these hypotheses, we found that activated mouse and human T cells display a pronounced DDR in vitro and in vivo. Upon screening a variety of small-molecule compounds, we found that potentiation of p53 (via inhibition of MDM2) or impairment of cell cycle checkpoints (via inhibition of CHK1/2 or WEE1) led to the selective elimination of activated, pathological T cells in vivo. The combination of these strategies [which we termed "p53 potentiation with checkpoint abrogation" (PPCA)] displayed therapeutic benefits in preclinical disease models of hemophagocytic lymphohistiocytosis and multiple sclerosis, which are driven by foreign antigens or self-antigens, respectively. PPCA therapy targeted pathological T cells but did not compromise naive, regulatory, or quiescent memory T-cell pools, and had a modest nonimmune toxicity profile. Thus, PPCA is a therapeutic modality for selective, antigen-specific immune modulation with significant translational potential for diverse immune-mediated diseases.
Assuntos
Dano ao DNA/imunologia , Doenças do Sistema Imunitário/terapia , Animais , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/imunologia , Modelos Animais de Doenças , Avaliação Pré-Clínica de Medicamentos , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/terapia , Etoposídeo/administração & dosagem , Humanos , Doenças do Sistema Imunitário/imunologia , Ativação Linfocitária , Linfo-Histiocitose Hemofagocítica/imunologia , Linfo-Histiocitose Hemofagocítica/terapia , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , Esclerose Múltipla/imunologia , Esclerose Múltipla/terapia , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-mdm2/imunologia , Transdução de Sinais/imunologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Linfócitos T/patologia , Proteína Supressora de Tumor p53/imunologiaRESUMO
Uridine-cytidine kinase 2 is an enzyme that is overexpressed in abnormal cell growth and its implication is considered a hallmark of cancer. Due to the selective expression of UCK2 in cancer cells, a selective inhibition of this key enzyme necessitates the discovery of its potential inhibitors for cancer chemotherapy. The present study was carried out to demonstrate the potentials of natural phytochemicals from the rhizome of Alpinia mutica to inhibit UCK2 useful for colorectal cancer. Here, we employed the used of in vitro to investigate the effectiveness of natural UCK2 inhibitors to cause HT-29 cell death. Extracts, flavokawain B, and alpinetin compound from the rhizome of Alpinia mutica was used in the study. The study demonstrated that the expression of UCK2 mRNA were substantially reduced in treated HT-29 cells. In addition, downregulation in expression of 18S ribosomal RNA was also observed in all treated HT-29 cells. This was confirmed by fluorescence imaging to measure the level of expression of 18S ribosomal RNA in live cell images. The study suggests the possibility of MDM2 protein was downregulated and its suppression subsequently activates the expression of p53 during inhibition of UCK2 enzyme. The expression of p53 is directly linked to a blockage of cell cycle progression at G0/G1 phase and upregulates Bax, cytochrome c, and caspase 3 while Bcl2 was deregulated. In this respect, apoptosis induction and DNA fragmentation were observed in treated HT-29 cells. Initial results from in vitro studies have shown the ability of the bioactive compounds of flavokawain B and alpinetin to target UCK2 enzyme specifically, inducing cell cycle arrest and subsequently leading to cancer cell death, possibly through interfering the MDM2-p53 signalling pathway. These phenomena have proven that the bioactive compounds could be useful for future therapeutic use in colon cancer.