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1.
J Mol Model ; 29(2): 55, 2023 Jan 26.
Artigo em Inglês | MEDLINE | ID: mdl-36700982

RESUMO

CONTEXT: The mutations in the TP53 gene are the most frequent (50-60% of human cancer) genetic alterations in cancer cells, indicating the critical role of wild-type p53 in the regulation of cell proliferation and apoptosis upon oncogenic stress. Most missense mutations are clustered in the DNA-binding core domain, disrupting DNA binding ability. However, some mutations like Y220C occur outside the DNA binding domain and are associated with p53 structure destabilization. Overall, the results of these mutations are single amino acid substitutions in p53 and the production of dysfunctional p53 protein in large amounts, consequently allowing the escape of apoptosis and rapid progression of tumor growth. Thus, therapeutic targeting of mutant p53 in tumors to restore its wild-type tumor suppression activity has immense potential for translational cancer research. Various molecules have been discovered with modern scientific techniques to reactivate mutant p53 by reverting structural changes and/or DNA binding ability. These compounds include small molecules, various peptides, and phytochemicals. TP53 protein is long thought of as a potential target; however, its translation for therapeutic purposes is still in its infancy. The study comprehensively analyzed the therapeutic potential of small phytochemicals from Foeniculum vulgare (Fennel) with drug-likeness and capability to reactivate mutant p53 (Y220C) through molecular docking simulation. The docking study and the stable molecular dynamic simulations revealed juglalin (- 8.6 kcal/mol), retinol (- 9.14 kcal/mol), and 3-nitrofluoranthene (- 8.43 kcal/mol) significantly bind to the mutated site suggesting the possibility of drug designing against the Y220C mutp53. The study supports these compounds for further animal based in vivo and in vitro research to validate their efficacy. METHODS: For the purposes of drug repurposing, recently in-silico methods have presented with opportunity to rule out many compounds which have less probability to act as a drug based on their structural moiety and interaction with the target macromolecule. The study here utilizes molecular docking via Autodock 4.2.6 and molecular dynamics using Schrodinger 2021 to find potential therapeutic options which are capable to reactive the mutated TP53 protein.


Assuntos
Foeniculum , Neoplasias , Animais , Humanos , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/química , Proteína Supressora de Tumor p53/metabolismo , Foeniculum/genética , Foeniculum/metabolismo , Genes p53 , Simulação de Acoplamento Molecular , Neoplasias/tratamento farmacológico , Neoplasias/genética , Neoplasias/patologia , Mutação , DNA
2.
J Biomol Struct Dyn ; 41(12): 5817-5826, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-35822492

RESUMO

Cancer is a major global health issue that has a high mortality rate. p53, which functions as a tumor suppressor, is critical in preventing tumor development by regulating the cell cycle and inducing apoptosis in damaged cells. However, the tumor suppressor function of p53 is effectively inhibited by its direct interaction with the hydrophobic cleft of MDM2 protein via multiple mechanisms As a result, restoring p53 activity by blocking the p53-MDM2 protein-protein interaction has been proposed as a compelling therapeutic strategy for cancer treatment. The use of molecular docking and phytochemical screening procedures are appraised to inhibit MDM2's hydrophobic cleft and disrupt the p53-MDM2 interaction. For this purpose, a library of 51 bioactive compounds from 10 medicinal plants was compiled and subjected to structure-based virtual screening. Out of these, only 3 compounds (Atalantoflavone, Cudraxanthone 1, and Ursolic acid) emerged as promising inhibitors of MDM2-p53 based on their binding affinities (-9.1 kcal/mol, -8.8 kcal/mol, and -8.8 kcal/mol respectively) when compared to the standard (-8.8 kcal/mol). Moreover, these compounds showed better pharmacokinetic and drug-like profiling than the standard inhibitor (Chromonotriazolopyrimidine 1). Finally, the 100 ns MD simulation analysis confirmed no significant perturbation in the conformational dynamics of the simulated binary complexes when compared to the standard. In particular, Ursolic acid was found to satisfy the molecular enumeration the most compared to the other inhibitors. Our overall molecular modeling finding shows why these compounds may emerge as potent arsenals for cancer therapeutics. Nonetheless, extensive experimental and clinical research is needed to augment their use in clinics.Communicated by Ramaswamy H. Sarma.


Assuntos
Neoplasias , Plantas Medicinais , Humanos , Simulação de Acoplamento Molecular , Proteína Supressora de Tumor p53/química , Proteínas Proto-Oncogênicas c-mdm2/química , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Dimerização , Neoplasias/tratamento farmacológico , Ligação Proteica , Ácido Ursólico
3.
Biochem Biophys Res Commun ; 578: 15-20, 2021 11 12.
Artigo em Inglês | MEDLINE | ID: mdl-34534740

RESUMO

Interaction between human positive coactivator 4 (PC4), an abundant nuclear protein, and the tumor suppressor protein p53 plays a crucial role in initiating apoptosis. In certain neurodegenerative diseases PC4 assisted-p53-dependent apoptosis may play a central role. Thus, disruption of p53-PC4 interaction may be a good drug target for certain disease pathologies. A p53-derived short peptide (AcPep) that binds the C-terminal domain of PC4 (C-PC4) is known to disrupt PC4-p53 interaction. To fully characterize its binding mode and binding site on PC4, we co-crystallized C-PC4 with the peptide and determined its structure. The crystal, despite exhibiting mass spectrometric signature of the peptide, lacked peptide electron density and showed a novel crystal lattice, when compared to C-PC4 crystals without the peptide. Using peptide-docked models of crystal lattices, corresponding to our structure and the peptide-devoid structure we show the origin of the novel crystal lattice to be dynamically bound peptide at the previously identified putative binding site. The weak binding is proposed to be due to the lack of the N-terminal domain of PC4 (N-PC4), which we experimentally show to be disordered with no effect on PC4 stability. Taking cue from the structure, virtual screening of ∼18.6 million small molecules from the ZINC15 database was performed, followed by toxicity and binding free energy filtering. The novel crystal lattice of C-PC4 in presence of the peptide, the role of the disordered N-PC4 and the high throughput identification of potent small molecules will allow a better understanding and control of p53-PC4 interaction.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Doenças Neurodegenerativas/patologia , Peptídeos/farmacologia , Bibliotecas de Moléculas Pequenas/farmacologia , Fatores de Transcrição/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Apoptose/fisiologia , Sítios de Ligação , Biologia Computacional/métodos , Cristalografia por Raios X , Proteínas de Ligação a DNA/química , Avaliação Pré-Clínica de Medicamentos/métodos , Humanos , Doenças Neurodegenerativas/tratamento farmacológico , Doenças Neurodegenerativas/metabolismo , Peptídeos/química , Domínios e Motivos de Interação entre Proteínas , Fatores de Transcrição/química , Proteína Supressora de Tumor p53/química
4.
Nat Commun ; 12(1): 986, 2021 02 12.
Artigo em Inglês | MEDLINE | ID: mdl-33579943

RESUMO

Epigallocatechin gallate (EGCG) from green tea can induce apoptosis in cancerous cells, but the underlying molecular mechanisms remain poorly understood. Using SPR and NMR, here we report a direct, µM interaction between EGCG and the tumor suppressor p53 (KD = 1.6 ± 1.4 µM), with the disordered N-terminal domain (NTD) identified as the major binding site (KD = 4 ± 2 µM). Large scale atomistic simulations (>100 µs), SAXS and AUC demonstrate that EGCG-NTD interaction is dynamic and EGCG causes the emergence of a subpopulation of compact bound conformations. The EGCG-p53 interaction disrupts p53 interaction with its regulatory E3 ligase MDM2 and inhibits ubiquitination of p53 by MDM2 in an in vitro ubiquitination assay, likely stabilizing p53 for anti-tumor activity. Our work provides insights into the mechanisms for EGCG's anticancer activity and identifies p53 NTD as a target for cancer drug discovery through dynamic interactions with small molecules.


Assuntos
Apoptose/efeitos dos fármacos , Catequina/análogos & derivados , Catequina/farmacologia , Proteínas Proto-Oncogênicas c-mdm2/química , Proteína Supressora de Tumor p53/química , Sítios de Ligação , Linhagem Celular Tumoral , Epitopos , Humanos , Ligação Proteica , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Espalhamento a Baixo Ângulo , Chá , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Ubiquitinação , Difração de Raios X
5.
Int J Cancer ; 148(1): 8-17, 2021 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-32638380

RESUMO

RAS, TP53 (p53) and MYC are among the most frequently altered driver genes in cancer. Thus, RAS is the most frequently mutated oncogene, MYC the most frequently amplified gene and TP53 the most frequently mutated tumor suppressor gene and overall the most frequently mutated gene in cancer. Theoretically, therefore, these genes are highly attractive targets for cancer treatment. However, as the protein products of each of these genes lack an accessible hydrophobic pocket into which low molecular weight compounds might bind with high affinity, they have proved difficult to target and have traditionally been referred to as "undruggable." Despite this branding, several low molecular weight compounds targeting each of these proteins have recently been reported to have anticancer activity in preclinical models. Indeed, several drugs inhibiting mutant KRAS, MYC overexpression or reactivating mutant p53 have undergone or are currently undergoing clinical trials. For targeting mutant KRAS and reactivating mutant p53, trials have progressed to a Phase III stage, that is, the mutant-p53 reactivating drug, APR-246 is currently being investigated in patients with myelodysplastic syndrome (MDS) and the RAS inhibitor, rigosertib is also undergoing evaluation in patients with MDS. Although there appears to be no directly acting MYC inhibitor currently being tested in a clinical trial, an anti-MYC compound, known as OmoMYC has been extensively validated in multiple preclinical models and is being developed for clinical evaluation. Based on current evidence, the traditional perception of RAS, p53 and MYC as being "undruggable" would appear to be coming to an end.


Assuntos
Antineoplásicos/farmacologia , Neoplasias/tratamento farmacológico , Proteínas Proto-Oncogênicas c-myc/antagonistas & inibidores , Proteínas Proto-Oncogênicas p21(ras)/antagonistas & inibidores , Proteína Supressora de Tumor p53/agonistas , Animais , Antineoplásicos/uso terapêutico , Sítios de Ligação , Carcinogênese/efeitos dos fármacos , Carcinogênese/genética , Ensaios Clínicos como Assunto , Avaliação Pré-Clínica de Medicamentos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Interações Hidrofóbicas e Hidrofílicas , Camundongos , Mutação , Neoplasias/genética , Proteínas Proto-Oncogênicas c-myc/química , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas p21(ras)/química , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteína Supressora de Tumor p53/química , Proteína Supressora de Tumor p53/genética
6.
Sci Rep ; 10(1): 11448, 2020 07 10.
Artigo em Inglês | MEDLINE | ID: mdl-32651427

RESUMO

Yinzhihuang granules (YZHG) is a patented Chinese medicine for the treatment of hepatitis B. This study aimed to investigate the intrinsic mechanisms of YZHG in the treatment of hepatitis B and to provide new evidence and insights for its clinical application. The chemical compounds of YZHG were searched in the CNKI and PUBMED databases, and their putative targets were then predicted through a search of the SuperPred and Swiss Target Prediction databases. In addition, the targets of hepatitis B were obtained from TTD, PharmGKB and DisGeNET. The abovementioned data were visualized using Cytoscape 3.7.1, and network construction identified a total of 13 potential targets of YZHG in the treatment of hepatitis B. Molecular docking verification showed that CDK6, CDK2, TP53 and BRCA1 might be strongly correlated with hepatitis B treatment. Furthermore, GO and KEGG analyses indicated that the treatment of hepatitis B by YZHG might be related to positive regulation of transcription, positive regulation of gene expression, the hepatitis B pathway and the viral carcinogenesis pathway. Network pharmacology intuitively shows the multicomponent, multitarget and multichannel pharmacological effects of YZHG in the treatment of hepatitis B and provides a scientific basis for its mechanism of action.


Assuntos
Biologia Computacional , Medicamentos de Ervas Chinesas/química , Hepatite B/tratamento farmacológico , Medicina Tradicional Chinesa , Proteína BRCA1/química , Proteína BRCA1/genética , Quinase 2 Dependente de Ciclina/química , Quinase 2 Dependente de Ciclina/genética , Quinase 6 Dependente de Ciclina/química , Quinase 6 Dependente de Ciclina/genética , Bases de Dados Factuais , Medicamentos de Ervas Chinesas/uso terapêutico , Hepatite B/virologia , Humanos , Simulação de Acoplamento Molecular , Transdução de Sinais/efeitos dos fármacos , Proteína Supressora de Tumor p53/química , Proteína Supressora de Tumor p53/genética
7.
Biochem Biophys Res Commun ; 524(3): 730-735, 2020 04 09.
Artigo em Inglês | MEDLINE | ID: mdl-32035620

RESUMO

Post-translational modifications (PTMs) play pivotal roles in controlling the stability and activity of the tumor suppressor p53 in response to distinct stressors. Here we report an unexpected finding of a short chain fatty acid modification of p53 in human cells. Crotonic acid (CA) treatment induces p53 crotonylation, but surprisingly reduces its protein, but not mRNA level, leading to inhibition of p53 activity in a dose dependent fashion. Surprisingly this crotonylation targets serine 46, instead of any predicted lysine residues, of p53, as detected in TCEP-probe labeled crotonylation and anti-crotonylated peptide antibody reaction assays. This is further confirmed by substitution of serine 46 with alanine, which abolishes p53 crotonylation in vitro and in cells. CA increases p53-dependent glycolytic activity, and augments cancer cell proliferation in response to metabolic or DNA damage stress. Since serine 46 is only found in human p53, our studies unveil an unconventional PTM unique for human p53, impairing its activity in response to CA. Because CA is likely produced by the gut microbiome, our results also predict that this type of PTM might play a role in early human colorectal neoplasia development by negating p53 activity without mutation of this tumor suppressor gene.


Assuntos
Crotonatos/metabolismo , Processamento de Proteína Pós-Traducional , Serina/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Crotonatos/química , Glucose/deficiência , Glicólise , Humanos , Lisina/metabolismo , Mitocôndrias/metabolismo , Proteína Supressora de Tumor p53/química
8.
Comput Biol Chem ; 83: 107105, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31473433

RESUMO

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/metabolismo
9.
J Phys Chem Lett ; 10(15): 4382-4400, 2019 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-31304749

RESUMO

It has been demonstrated that MMP13 enzyme is related to most cancer cell tumors. The world's largest traditional Chinese medicine database was applied to screen for structure-based drug design and ligand-based drug design. To predict drug activity, machine learning models (Random Forest (RF), AdaBoost Regressor (ABR), Gradient Boosting Regressor (GBR)), and Deep Learning models were utilized to validate the Docking results, and we obtained an R2 of 0.922 on the training set and 0.804 on the test set in the RF algorithm. For the Deep Learning algorithm, R2 of the training set is 0.90, and R2 of the test set is 0.810. However, these TCM compounds fly away during the molecular dynamics (MD) simulation. We seek another method: peptide design. All peptide database were screened by the Docking process. Modification peptides were optimized the interaction modes, and the affinities were assessed with ZDOCK protocol and Refine Docked protein protocol. The 300 ns MD simulation evaluated the stability of receptor-peptide complexes. The double-site effect appeared on S2, a designed peptide based on a known inhibitor, when complexed with BCL2. S3, a designed peptide referred from endogenous inhibitor P16, competed against cyclin when binding with CDK6. The MDM2 inhibitors S5 and S6 were derived from the P53 structure and stable binding with MDM2. A flexible region of peptides S5 and S6 may enhance the binding ability by changing its own conformation, which was unforeseen. These peptides (S2, S3, S5, and S6) are potentially interesting to treat cancer; however, these findings need to be affirmed by biological testing, which will be conducted in the near future.


Assuntos
Antineoplásicos/química , Aprendizado Profundo , Aprendizado de Máquina , Modelos Moleculares , Peptídeos/química , Proteínas/química , Algoritmos , Sítios de Ligação , Quinase 6 Dependente de Ciclina/química , Inibidor p16 de Quinase Dependente de Ciclina/química , Bases de Dados de Produtos Farmacêuticos , Bases de Dados de Proteínas , Desenho de Fármacos , Ligantes , Metaloproteinase 13 da Matriz/química , Mutação , Proteínas Proto-Oncogênicas c-bcl-2/química , Proteínas Proto-Oncogênicas c-mdm2/química , Proteína Supressora de Tumor p53/química , Proteína Supressora de Tumor p53/genética
10.
J Exp Clin Cancer Res ; 38(1): 103, 2019 Feb 26.
Artigo em Inglês | MEDLINE | ID: mdl-30808373

RESUMO

BACKGROUND: Tumor suppressor p53 protein is frequently mutated in a large majority of cancers. These mutations induce local or global changes in protein structure thereby affecting its binding to DNA. The structural differences between the wild type and mutant p53 thus provide an opportunity to selectively target mutated p53 harboring cancer cells. Restoration of wild type p53 activity in mutants using small molecules that can revert the structural changes have been considered for cancer therapeutics. METHODS: We used bioinformatics and molecular docking tools to investigate the structural changes between the wild type and mutant p53 proteins (p53V143A, p53R249S, p53R273H and p53Y220C) and explored the therapeutic potential of Withaferin A and Withanone for restoration of wild type p53 function in cancer cells. Cancer cells harboring the specific mutant p53 proteins were used for molecular assays to determine the mutant or wild type p53 functions. RESULTS: We found that p53V143A mutation does not show any significant structural changes and was also refractory to the binding of withanolides. p53R249S mutation critically disturbed the H-bond network and destabilized the DNA binding site. However, withanolides did not show any selective binding to either this mutant or other similar variants. p53Y220C mutation created a cavity near the site of mutation with local loss of hydrophobicity and water network, leading to functionally inactive conformation. Mutated structure could accommodate withanolides suggesting their conformational selectivity to target p53Y220C mutant. Using human cell lines containing specific p53 mutant proteins, we demonstrated that Withaferin A, Withanone and the extract rich in these withanolides caused restoration of wild type p53 function in mutant p53Y220C cells. This was associated with induction of p21WAF-1-mediated growth arrest/apoptosis. CONCLUSION: The study suggested that withanolides may serve as highly potent anticancer compounds for treatment of cancers harboring a p53Y220C mutation.


Assuntos
Antineoplásicos/farmacologia , Extratos Vegetais/farmacologia , Proteína Supressora de Tumor p53/química , Proteína Supressora de Tumor p53/genética , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Biologia Computacional , Humanos , Conformação Molecular , Simulação de Acoplamento Molecular , Proteína Supressora de Tumor p53/efeitos dos fármacos , Vitanolídeos/farmacologia
11.
Nucleic Acids Res ; 47(4): 1637-1652, 2019 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-30649466

RESUMO

The DNA binding domain (DBD) of the tumor suppressor p53 is the site of several oncogenic mutations. A subset of these mutations lowers the unfolding temperature of the DBD. Unfolding leads to the exposure of a hydrophobic ß-strand and nucleates aggregation which results in pathologies through loss of function and dominant negative/gain of function effects. Inspired by the hypothesis that structural changes that are associated with events initiating unfolding in DBD are likely to present opportunities for inhibition, we investigate the dynamics of the wild type (WT) and some aggregating mutants through extensive all atom explicit solvent MD simulations. Simulations reveal differential conformational sampling between the WT and the mutants of a turn region (S6-S7) that is contiguous to a known aggregation-prone region (APR). The conformational properties of the S6-S7 turn appear to be modulated by a network of interacting residues. We speculate that changes that take place in this network as a result of the mutational stress result in the events that destabilize the DBD and initiate unfolding. These perturbations also result in the emergence of a novel pocket that appears to have druggable characteristics. FDA approved drugs are computationally screened against this pocket.


Assuntos
Proteínas de Ligação a DNA/química , Proteínas Mutantes/química , Bibliotecas de Moléculas Pequenas/química , Proteína Supressora de Tumor p53/química , Proteínas de Ligação a DNA/genética , Avaliação Pré-Clínica de Medicamentos/métodos , Humanos , Interações Hidrofóbicas e Hidrofílicas/efeitos dos fármacos , Modelos Moleculares , Simulação de Dinâmica Molecular , Proteínas Mutantes/genética , Mutação/genética , Conformação Proteica/efeitos dos fármacos , Domínios Proteicos/efeitos dos fármacos , Domínios Proteicos/genética , Desdobramento de Proteína/efeitos dos fármacos , Proteína Supressora de Tumor p53/genética
12.
Drug Des Devel Ther ; 12: 3741-3752, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30464405

RESUMO

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/metabolismo
13.
J Recept Signal Transduct Res ; 38(3): 179-190, 2018 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-29734849

RESUMO

The aim of this study was to find the efficacy of 5-hydroxy 3',4',7-trimethoxyflavone (HTMF), a flavonoid compound isolated from the medicinal plant Lippia nodiflora, in inhibiting the proliferation and inducing apoptosis in human breast cancer cell line MCF-7. The anti-proliferative effect of the compound HTMF was confirmed using MTT cytotoxicity assay. Increased apoptotic induction by HTMF was demonstrated by acridine orange/ethidium bromide (AO/EtBr) and Hoechst 33258 staining studies. The phosphatidylserine translocation, an early feature of apoptosis and DNA damage were revealed through AnnexinV-Cy3 staining and comet assay. Moreover, the significant elevation of cellular ROS was observed in the treated cells, as measured by 2,7-diacetyl dichlorofluorescein (DCFH-DA). The mRNA expression studies also supported the effectiveness of HTMF by shifting the Bax:Bcl-2 ratio. The treatment of MCF-7 cells with HTMF encouraged apoptosis through the modulation of apoptotic markers, such as p53, Bcl-2, Bax, and cleaved PARP. In silico molecular docking and dynamics studies with MDM2-p53 protein revealed that HTMF was more potent compound that could inhibit the binding of MDM2 with p53 and, therefore, could trigger apoptosis in cancer cell. Overall, this study brings up scientific evidence for the efficacy of HTMF against MCF-7 breast cancer cells.


Assuntos
Apoptose/efeitos dos fármacos , Neoplasias da Mama/tratamento farmacológico , Flavonas/farmacologia , Proteínas Proto-Oncogênicas c-mdm2/genética , Proteína Supressora de Tumor p53/genética , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Proliferação de Células/efeitos dos fármacos , Simulação por Computador , Feminino , Flavonas/química , Regulação Neoplásica da Expressão Gênica , Humanos , Células MCF-7 , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Poli(ADP-Ribose) Polimerases/genética , Ligação Proteica/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-mdm2/química , Proteína Supressora de Tumor p53/química
14.
Anticancer Drugs ; 28(9): 1032-1038, 2017 10.
Artigo em Inglês | MEDLINE | ID: mdl-28723868

RESUMO

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/metabolismo
15.
Stem Cells ; 33(5): 1657-69, 2015 May.
Artigo em Inglês | MEDLINE | ID: mdl-25694388

RESUMO

Retinoblastoma tumor suppressor protein (RB) is inactivated more frequently during tumor progression than during tumor initiation. However, its exact role in controlling the malignant features associated with tumor progression is poorly understood. We established in vivo and in vitro models to investigate the undifferentiated state induced by Rb inactivation. Rb heterozygous mice develop well-differentiated thyroid medullary carcinoma. We found that additional deletion of Trp53, without change in lineage, converted these Rb-deficient tumors to a poorly differentiated type associated with higher self-renewal activity. Freshly prepared mouse embryonic fibroblasts (MEFs) of Rb(-/-) ; Trp53(-/-) background formed stem cell-like spheres that expressed significant levels of embryonic genes despite of lacking the ability to form colonies on soft agar or tumors in immune-deficient mice. This suggested that Rb-p53 double inactivation resulted in an undifferentiated status but without carcinogenic conversion. We next established Rb(-/-) ; N-ras(-/-) MEFs that harbored a spontaneous carcinogenic mutation in Trp53. These cells (RN6), in an Rb-dependent manner, efficiently generated spheres that expressed very high levels of embryonic genes, and appeared to be carcinogenic. We then screened an FDA-approved drug library to search for agents that suppressed the spherogenic activity of RN6 cells. Data revealed that RN6 cells were sensitive to specific agents including ones those are effective against cancer stem cells. Taken together, all these findings suggest that the genetic interaction between Rb and p53 is a critical determinant of the undifferentiated state in normal and tumor cells.


Assuntos
Diferenciação Celular , Embrião de Mamíferos/citologia , Fibroblastos/citologia , Células Neuroendócrinas/citologia , Proteína do Retinoblastoma/metabolismo , Glândula Tireoide/citologia , Proteína Supressora de Tumor p53/metabolismo , Sequência de Aminoácidos , Animais , Comportamento Animal , Linhagem Celular , Avaliação Pré-Clínica de Medicamentos , Fibroblastos/metabolismo , Heterozigoto , Camundongos Knockout , Dados de Sequência Molecular , Mutação/genética , Fenótipo , Proteína do Retinoblastoma/deficiência , Esferoides Celulares/metabolismo , Proteína Supressora de Tumor p53/química , Proteína Supressora de Tumor p53/deficiência , Proteína Supressora de Tumor p53/genética , Proteínas ras/metabolismo
16.
Chem Biol Drug Des ; 86(2): 180-9, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-25388787

RESUMO

Two libraries of substituted benzimidazoles were designed using a 'scaffold-hopping' approach based on reported MDM2-p53 inhibitors. Substituents were chosen following library enumeration and docking into an MDM2 X-ray structure. Benzimidazole libraries were prepared using an efficient solution-phase approach and screened for inhibition of the MDM2-p53 and MDMX-p53 protein-protein interactions. Key examples showed inhibitory activity against both targets.


Assuntos
Benzimidazóis/farmacologia , Proteínas Nucleares/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-mdm2/antagonistas & inibidores , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Proteína Supressora de Tumor p53/antagonistas & inibidores , Benzimidazóis/química , Proteínas de Ciclo Celular , Avaliação Pré-Clínica de Medicamentos , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Ensaio de Imunoadsorção Enzimática , Humanos , Modelos Moleculares , Simulação de Acoplamento Molecular , Proteínas Nucleares/química , Proteínas Nucleares/metabolismo , Ligação Proteica/efeitos dos fármacos , Proteínas Proto-Oncogênicas/química , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Proto-Oncogênicas c-mdm2/química , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Proteína Supressora de Tumor p53/química , Proteína Supressora de Tumor p53/metabolismo
17.
Chem Biol Drug Des ; 86(2): 163-72, 2015 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-25407396

RESUMO

The guardian of the genome, p53, is the most mutated protein found in all cancer cells. Restoration of wild-type activity to mutant p53 offers promise to eradicate cancer cells using novel pharmacological agents. Several molecules have already been found to activate mutant p53. While the exact mechanism of action of these compounds has not been fully understood, a transiently open pocket has been identified in some mutants. In our study, we docked twelve known activators to p53 into the open pocket to further understand their mechanism of action and rank the best binders. In addition, we predicted the absorption, distribution, metabolism, excretion and toxicity properties of these compounds to assess their pharmaceutical usefulness. Our studies showed that alkylating ligands do not all bind at the same position, probably due to their varying sizes. In addition, we found that non-alkylating ligands are capable of binding at the same pocket and directly interacting with Cys124. The comparison of the different ligands demonstrates that stictic acid has a great potential as a p53 activator in terms of less adverse effects although it has poorer pharmacokinetic properties.


Assuntos
Proteína Supressora de Tumor p53/química , Proteína Supressora de Tumor p53/metabolismo , Alquilação , Amifostina/química , Amifostina/farmacocinética , Amifostina/toxicidade , Compostos Aza/química , Compostos Aza/farmacocinética , Compostos Aza/toxicidade , Compostos Bicíclicos Heterocíclicos com Pontes/química , Compostos Bicíclicos Heterocíclicos com Pontes/farmacocinética , Compostos Bicíclicos Heterocíclicos com Pontes/toxicidade , Avaliação Pré-Clínica de Medicamentos , Elipticinas/química , Elipticinas/farmacocinética , Elipticinas/farmacologia , Elipticinas/toxicidade , Compostos Heterocíclicos de 4 ou mais Anéis/química , Compostos Heterocíclicos de 4 ou mais Anéis/farmacocinética , Compostos Heterocíclicos de 4 ou mais Anéis/toxicidade , Humanos , Cinética , Ligantes , Mercaptoetilaminas/química , Mercaptoetilaminas/farmacocinética , Mercaptoetilaminas/toxicidade , Simulação de Dinâmica Molecular , Mutagênese Sítio-Dirigida , Mutação , Oxepinas/química , Oxepinas/farmacocinética , Oxepinas/toxicidade , Ligação Proteica , Pirimidinas/química , Pirimidinas/farmacologia , Pirimidinas/toxicidade , Quinuclidinas/química , Quinuclidinas/farmacocinética , Quinuclidinas/toxicidade , Proteína Supressora de Tumor p53/genética
18.
Clin Cancer Res ; 19(20): 5777-87, 2013 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-23983256

RESUMO

PURPOSE: The TP53 tumor suppressor is frequently mutated in colon cancer, but the influence of such mutations on survival remains controversial. We investigated whether mutations in the DNA-binding domain of TP53 are associated with survival in stage III colon cancer. EXPERIMENTAL DESIGN: The impact of TP53 genotype was prospectively evaluated in Cancer and Leukemia Group B 89803, a trial that randomized stage III colon cancer patients to receive adjuvant 5-fluorouracil/leucovorin (5FU/LV) or 5FU/LV with irinotecan (IFL). RESULTS: TP53 mutations were identified in 274 of 607 cases. The presence of any TP53 mutation did not predict disease-free survival (DFS) or overall survival with either adjuvant regimen when men and women were considered together or as separate groups. However, outcome differences among women became apparent when tumor TP53 genotype was stratified as wild-type versus zinc- or non-zinc-binding mutations in the TP53 DNA-binding domain. DFS at 5 years was 0.59, 0.52, and 0.78 for women with TP53 wild-type tumors, and tumors with zinc- or non-zinc-binding mutations, respectively. Survival at 5 years for these same women was 0.72, 0.59, and 0.90, respectively. No differences in survival by TP53 genotype were observed in men. CONCLUSIONS: The presence of any TP53 mutation within the DNA-binding domain did not predict survival in stage III colon cancer. However, TP53 genotype was predictive of survival in women following adjuvant therapy. Future colon cancer therapeutic trials, with inclusion of correlative molecular markers, should be designed to permit evaluation of survival and/or response to treatment in women separately from men.


Assuntos
Neoplasias do Colo/genética , Neoplasias do Colo/patologia , Mutação , Proteína Supressora de Tumor p53/genética , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Quimioterapia Adjuvante , Neoplasias do Colo/mortalidade , Neoplasias do Colo/terapia , DNA/química , DNA/metabolismo , Feminino , Genótipo , Humanos , Masculino , Instabilidade de Microssatélites , Modelos Moleculares , Conformação Molecular , Estadiamento de Neoplasias , Ligação Proteica , Fatores Sexuais , Proteína Supressora de Tumor p53/química , Proteína Supressora de Tumor p53/metabolismo , Zinco/química , Zinco/metabolismo
19.
Cell Death Dis ; 4: e484, 2013 Feb 07.
Artigo em Inglês | MEDLINE | ID: mdl-23392172

RESUMO

The role of p53 in neurodegenerative diseases is essentially associated with neuronal death. Recently an alternative point of view is emerging, as altered p53 conformation and impaired protein function have been found in fibroblasts and blood cells derived from Alzheimer's disease patients. Here, using stable transfected SH-SY5Y cells overexpressing APP751wt (SY5Y-APP) we demonstrated that the expression of an unfolded p53 conformation compromised neuronal functionality. In particular, these cells showed (i) augmented expression of amyloid precursor protein (APP) and its metabolites, including the C-terminal fragments C99 and C83 and ß-amyloid peptide (ii) high levels of oxidative markers, such as 4-hydroxy-2-nonenal Michael-adducts and 3-nitro-tyrosine and (iii) altered p53 conformation, mainly due to nitration of its tyrosine residues. The consequences of high-unfolded p53 expression resulted in loss of p53 pro-apoptotic activity, and reduction of growth-associated protein 43 (GAP-43) mRNA and protein levels. The role of unfolded p53 in cell death resistance and lack of GAP-43 transcription was demonstrated by ZnCl(2) treatment. Zinc supplementation reverted p53 wild-type tertiary structure, increased cells sensitivity to acute cytotoxic injury and GAP-43 levels in SY5Y-APP clone.


Assuntos
Proteína GAP-43/metabolismo , Neurônios/metabolismo , Proteína Supressora de Tumor p53/química , Precursor de Proteína beta-Amiloide/genética , Precursor de Proteína beta-Amiloide/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cloretos/farmacologia , Proteína GAP-43/genética , Humanos , Peróxido de Hidrogênio/toxicidade , Estresse Oxidativo , Estrutura Terciária de Proteína , Desdobramento de Proteína , RNA Mensageiro/metabolismo , Transcrição Gênica/efeitos dos fármacos , Proteína Supressora de Tumor p53/metabolismo , Tirosina/análogos & derivados , Tirosina/química , Compostos de Zinco/farmacologia
20.
Toxicol Sci ; 126(1): 60-71, 2012 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-22143989

RESUMO

The multifactorial origin of gastric cancer encompasses environmental factors mainly associated with diet. Pteridium aquilinum-bracken fern-is the only higher plant known to cause cancer in animals. Its carcinogenic toxin, ptaquiloside, has been identified in milk of cows and groundwater. Humans can be directly exposed by consumption of the plant, contaminated water or milk, and spore inhalation. Epidemiological studies have shown an association between bracken exposure and gastric cancer. In the present work, the genotoxicity of P. aquilinum and ptaquiloside, including DNA damaging effects and DNA damage response, was characterized in human gastric epithelial cells and in a mouse model. In vitro, the highest doses of P. aquilinum extracts (40 mg/ml) and ptaquiloside (60 µg/ml) decreased cell viability and induced apoptosis. γH2AX and P53-binding protein 1 analysis indicated induction of DNA strand breaks in treated cells. P53 level also increased after exposure, associated with ATR-Chk1 signaling pathway activation. The involvement of ptaquiloside in the DNA damage activity of P. aquilinum was confirmed by deregulation of the expression of a panel of genes related to DNA damage signaling pathways and DNA repair, in response to purified ptaquiloside. Oral administration of P. aquilinum extracts to mice increased gastric cell proliferation and led to frameshift events in intron 2 of the P53 gene. Our data demonstrate the direct DNA damaging and mutagenic effects of P. aquilinum. These results are in agreement with the carcinogenic properties attributed to this fern and its ptaquiloside toxin and support their role in promoting gastric carcinogenesis.


Assuntos
Dano ao DNA , Mucosa Gástrica/efeitos dos fármacos , Indanos/toxicidade , Mutagênicos/toxicidade , Extratos Vegetais/toxicidade , Pteridium/química , Sesquiterpenos/toxicidade , Neoplasias Gástricas/induzido quimicamente , Animais , Apoptose/efeitos dos fármacos , Carcinoma/induzido quimicamente , Linhagem Celular Tumoral , Mucosa Gástrica/metabolismo , Mucosa Gástrica/patologia , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Indanos/administração & dosagem , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mutagênicos/administração & dosagem , Proteínas Mutantes/química , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Proteínas de Neoplasias/química , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Extratos Vegetais/administração & dosagem , Folhas de Planta/efeitos adversos , Folhas de Planta/química , Pteridium/efeitos adversos , RNA Mensageiro/metabolismo , Sesquiterpenos/administração & dosagem , Organismos Livres de Patógenos Específicos , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologia , Proteína Supressora de Tumor p53/química , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo
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