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1.
Nat Commun ; 11(1): 4659, 2020 09 16.
Artigo em Inglês | MEDLINE | ID: mdl-32938936

RESUMO

The αvß6 integrin plays a key role in the activation of transforming growth factor-ß (TGFß), a pro-fibrotic mediator that is pivotal to the development of idiopathic pulmonary fibrosis (IPF). We identified a selective small molecule αvß6 RGD-mimetic, GSK3008348, and profiled it in a range of disease relevant pre-clinical systems. To understand the relationship between target engagement and inhibition of fibrosis, we measured pharmacodynamic and disease-related end points. Here, we report, GSK3008348 binds to αvß6 with high affinity in human IPF lung and reduces downstream pro-fibrotic TGFß signaling to normal levels. In human lung epithelial cells, GSK3008348 induces rapid internalization and lysosomal degradation of the αvß6 integrin. In the murine bleomycin-induced lung fibrosis model, GSK3008348 engages αvß6, induces prolonged inhibition of TGFß signaling and reduces lung collagen deposition and serum C3M, a marker of IPF disease progression. These studies highlight the potential of inhaled GSK3008348 as an anti-fibrotic therapy.


Assuntos
Butiratos/farmacologia , Fibrose Pulmonar Idiopática/tratamento farmacológico , Integrinas/antagonistas & inibidores , Naftiridinas/farmacologia , Pirazóis/farmacologia , Pirrolidinas/farmacologia , Administração por Inalação , Animais , Antígenos de Neoplasias/metabolismo , Bleomicina/toxicidade , Butiratos/administração & dosagem , Butiratos/metabolismo , Butiratos/farmacocinética , Colágeno/metabolismo , Modelos Animais de Doenças , Células Epiteliais/efeitos dos fármacos , Humanos , Fibrose Pulmonar Idiopática/induzido quimicamente , Fibrose Pulmonar Idiopática/patologia , Integrinas/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Simulação de Acoplamento Molecular , Naftiridinas/administração & dosagem , Naftiridinas/metabolismo , Naftiridinas/farmacocinética , Pirazóis/administração & dosagem , Pirazóis/metabolismo , Pirazóis/farmacocinética , Pirrolidinas/administração & dosagem , Pirrolidinas/metabolismo , Pirrolidinas/farmacocinética , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Tomografia Computadorizada de Emissão de Fóton Único , Fator de Crescimento Transformador beta/metabolismo , Pesquisa Médica Translacional
2.
Anticancer Res ; 40(9): 4929-4935, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32878781

RESUMO

BACKGROUND: Mutations in the isocitrate dehydrogenase 1 (IDH1) gene are frequently found in various cancer types. IDH1 mutants produce 2-hydroxyglutarate (2-HG), an oncometabolite, from alpha-ketoglutarate (α-KG). This 2-HG plays a key role in tumorigenesis via inhibition of α-KG dependent enzymes. For this reason, IDH1 mutant could be an ideal target for the treatment of cancer. MATERIALS AND METHODS: To find a new IDH1 inhibitor, 8,364 compounds were obtained from Korea Chemical Bank. Using high-throughput screening (HTS) of a chemical library, we unveiled a compound that could inhibit the IDH1 mutant. RESULTS: According to the enzyme assay, our compound (KRC-09) effectively inhibited the activity of IDH1 R132H mutant. In addition, KRC-09 decreased the concentration of intracellular 2-HG in the U-87 MG cell line harboring IDH1 R132H. CONCLUSION: In this article, we present a novel chemical scaffold that suppresses the activity of an IDH1 mutant.


Assuntos
Antineoplásicos/farmacologia , Isocitrato Desidrogenase/antagonistas & inibidores , Isocitrato Desidrogenase/genética , Antineoplásicos/química , Linhagem Celular Tumoral , Descoberta de Drogas , Glutaratos/metabolismo , Humanos , Isocitrato Desidrogenase/metabolismo , Estrutura Molecular , Mutação , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia
3.
PLoS One ; 15(7): e0236710, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32735606

RESUMO

Argonaute (AGO) proteins are the key component of the RNA interference machinery that suppresses gene expression by forming an RNA-induced silencing complex (RISC) with microRNAs (miRNAs). Each miRNA is involved in various cellular processes, such as development, differentiation, tumorigenesis, and viral infection. Thus, molecules that regulate miRNA function are expected to have therapeutic potential. In addition, the biogenesis of miRNA is a multistep process involving various proteins, although the complete pathway remains to be elucidated. Therefore, identification of molecules that can specifically modulate each step will help understand the mechanism of gene suppression. To date, several AGO2 inhibitors have been identified. However, these molecules were identified through a single screening method, and no studies have specifically evaluated a combinatorial strategy. Here, we demonstrated a combinatorial screening (SCR) approach comprising an in silico molecular docking study, surface plasmon resonance (SPR) analysis, and nuclear magnetic resonance (NMR) analysis, focusing on the strong binding between the 5'-terminal phosphate of RNA and the AGO2 middle (MID) domain. By combining SPR and NMR, we identified binding modes of amino acid residues binding to AGO2. First, using a large chemical library (over 6,000,000 compounds), 171 compounds with acidic functional groups were screened using in silico SCR. Next, we constructed an SPR inhibition system that could analyze only the 5'-terminal binding site of RNA, and nine molecules that strongly bound to the AGO2 MID domain were selected. Finally, using NMR, three molecules that bound to the desired site were identified. The RISC inhibitory ability of the "hit" compounds was analyzed in human cell lysate, and all three hit compounds strongly inhibited the binding between double-stranded RNA and AGO2.


Assuntos
Proteínas Argonauta/antagonistas & inibidores , Avaliação Pré-Clínica de Medicamentos/métodos , Espectroscopia de Ressonância Magnética , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Ressonância de Plasmônio de Superfície , Proteínas Argonauta/química , Proteínas Argonauta/metabolismo , Simulação por Computador , Células HeLa , Humanos , Simulação de Acoplamento Molecular , Conformação Proteica , Bibliotecas de Moléculas Pequenas/metabolismo
4.
J Mol Graph Model ; 100: 107697, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32739642

RESUMO

Angiotensin-converting enzyme 2 (ACE2) is a membrane-bound zinc metallopeptidase that generates the vasodilatory peptide angiotensin 1-7 and thus performs a protective role in heart disease. It is considered an important therapeutic target in controlling the COVID-19 outbreak, since SARS-CoV-2 enters permissive cells via an ACE2-mediated mechanism. The present in silico study attempted to repurpose existing drugs for use as prospective viral-entry inhibitors targeting human ACE2. Initially, a clinically approved drug library of 7,173 ligands was screened against the receptor using molecular docking, followed by energy minimization and rescoring of docked ligands. Finally, potential binders were inspected to ensure molecules with different scaffolds were engaged in favorable contacts with both the metal cofactor and the critical residues lining the receptor's active site. The results of the calculations suggest that lividomycin, burixafor, quisinostat, fluprofylline, pemetrexed, spirofylline, edotecarin, and diniprofylline emerge as promising repositionable drug candidates for stabilizing the closed (substrate/inhibitor-bound) conformation of ACE2, thereby shifting the relative positions of the receptor's critical exterior residues recognized by SARS-CoV-2. This study is among the rare ones in the relevant scientific literature to search for potential ACE2 inhibitors. In practical terms, the drugs, unmodified as they are, may be introduced into the therapeutic armamentarium of the ongoing fight against COVID-19 now, or their scaffolds may serve as rich skeletons for designing novel ACE2 inhibitors in the near future.


Assuntos
Inibidores da Enzima Conversora de Angiotensina/química , Antivirais/química , Betacoronavirus/química , Peptidil Dipeptidase A/química , Bibliotecas de Moléculas Pequenas/química , Motivos de Aminoácidos , Betacoronavirus/enzimologia , Carbazóis/química , Domínio Catalítico , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/virologia , Reposicionamento de Medicamentos , Difilina/análogos & derivados , Difilina/química , Interações Hospedeiro-Patógeno , Humanos , Ácidos Hidroxâmicos/química , Ligantes , Simulação de Acoplamento Molecular , Pandemias , Paromomicina/análogos & derivados , Paromomicina/química , Pemetrexede/química , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/virologia , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Estrutura Secundária de Proteína , Relação Estrutura-Atividade , Termodinâmica
5.
J Mol Graph Model ; 100: 107710, 2020 11.
Artigo em Inglês | MEDLINE | ID: mdl-32829149

RESUMO

The emergence of SARS-CoV-2 has prompted a worldwide health emergency. There is an urgent need for therapeutics, both through the repurposing of approved drugs and the development of new treatments. In addition to the viral drug targets, a number of human drug targets have been suggested. In theory, targeting human proteins should provide an advantage over targeting viral proteins in terms of drug resistance, which is commonly a problem in treating RNA viruses. This paper focuses on the human protein TMPRSS2, which supports coronavirus life cycles by cleaving viral spike proteins. The three-dimensional structure of TMPRSS2 is not known and so we have generated models of the TMPRSS2 in the apo state as well as in complex with a peptide substrate and putative inhibitors to aid future work. Importantly, many related human proteases have 80% or higher identity with TMPRSS2 in the S1-S1' subsites, with plasminogen and urokinase-type plasminogen activator (uPA) having 95% identity. We highlight 376 approved, investigational or experimental drugs targeting S1A serine proteases that may also inhibit TMPRSS2. Whilst the presence of a relatively uncommon lysine residue in the S2/S3 subsites means that some serine protease inhibitors will not inhibit TMPRSS2, this residue is likely to provide a handle for selective targeting in a focused drug discovery project. We discuss how experimental drugs targeting related serine proteases might be repurposed as TMPRSS2 inhibitors to treat coronaviruses.


Assuntos
Antivirais/química , Betacoronavirus/química , Inibidores de Proteases/química , Serina Endopeptidases/química , Bibliotecas de Moléculas Pequenas/química , Sequência de Aminoácidos , Betacoronavirus/enzimologia , Domínio Catalítico , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/virologia , Reposicionamento de Medicamentos , Interações Hospedeiro-Patógeno , Humanos , Ligantes , Simulação de Dinâmica Molecular , Pandemias , Plasminogênio/antagonistas & inibidores , Plasminogênio/química , Plasminogênio/metabolismo , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/virologia , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Estrutura Secundária de Proteína , Alinhamento de Sequência , Serina Endopeptidases/metabolismo , Homologia Estrutural de Proteína , Relação Estrutura-Atividade , Termodinâmica , Ativador de Plasminogênio Tipo Uroquinase/antagonistas & inibidores , Ativador de Plasminogênio Tipo Uroquinase/química , Ativador de Plasminogênio Tipo Uroquinase/metabolismo
6.
J Chem Inf Model ; 60(8): 3910-3934, 2020 08 24.
Artigo em Inglês | MEDLINE | ID: mdl-32786511

RESUMO

Protein-protein interactions (PPIs) are attractive targets for drug design because of their essential role in numerous cellular processes and disease pathways. However, in general, PPIs display exposed binding pockets at the interface, and as such, have been largely unexploited for therapeutic interventions with low-molecular weight compounds. Here, we used docking and various rescoring strategies in an attempt to recover PPI inhibitors from a set of active and inactive molecules for 11 targets collected in ChEMBL and PubChem. Our focus is on the screening power of the various developed protocols and on using fast approaches so as to be able to apply such a strategy to the screening of ultralarge libraries in the future. First, we docked compounds into each target using the fast "pscreen" mode of the structure-based virtual screening (VS) package Surflex. Subsequently, the docking poses were postprocessed to derive a set of 3D topological descriptors: (i) shape similarity and (ii) interaction fingerprint similarity with a co-crystallized inhibitor, (iii) solvent-accessible surface area, and (iv) extent of deviation from the geometric center of a reference inhibitor. The derivatized descriptors, together with descriptor-scaled scoring functions, were utilized to investigate possible impacts on VS performance metrics. Moreover, four standalone scoring functions, RF-Score-VS (machine-learning), DLIGAND2 (knowledge-based), Vinardo (empirical), and X-SCORE (empirical), were employed to rescore the PPI compounds. Collectively, the results indicate that the topological scoring algorithms could be valuable both at a global level, with up to 79% increase in areas under the receiver operating characteristic curve for some targets, and in early stages, with up to a 4-fold increase in enrichment factors at 1% of the screened collections. Outstandingly, DLIGAND2 emerged as the best scoring function on this data set, outperforming all rescoring techniques in terms of VS metrics. The described methodology could help in the rational design of small-molecule PPI inhibitors and has direct applications in many therapeutic areas, including cancer, CNS, and infectious diseases such as COVID-19.


Assuntos
Desenho de Fármacos , Descoberta de Drogas , Mapas de Interação de Proteínas/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/farmacologia , Algoritmos , Betacoronavirus/efeitos dos fármacos , Betacoronavirus/metabolismo , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/metabolismo , Bases de Dados de Proteínas , Humanos , Ligantes , Aprendizado de Máquina , Simulação de Acoplamento Molecular , Terapia de Alvo Molecular , Pandemias , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/metabolismo , Proteínas/química , Proteínas/metabolismo , Bibliotecas de Moléculas Pequenas/química
7.
Sci Rep ; 10(1): 13093, 2020 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-32753646

RESUMO

A novel coronavirus, named SARS-CoV-2, emerged in 2019 in China and rapidly spread worldwide. As no approved therapeutics exists to treat COVID-19, the disease associated to SARS-Cov-2, there is an urgent need to propose molecules that could quickly enter into clinics. Repurposing of approved drugs is a strategy that can bypass the time-consuming stages of drug development. In this study, we screened the PRESTWICK CHEMICAL LIBRARY composed of 1,520 approved drugs in an infected cell-based assay. The robustness of the screen was assessed by the identification of drugs that already demonstrated in vitro antiviral effect against SARS-CoV-2. Thereby, 90 compounds were identified as positive hits from the screen and were grouped according to their chemical composition and their known therapeutic effect. Then EC50 and CC50 were determined for a subset of 15 compounds from a panel of 23 selected drugs covering the different groups. Eleven compounds such as macrolides antibiotics, proton pump inhibitors, antiarrhythmic agents or CNS drugs emerged showing antiviral potency with 2 < EC50 ≤ 20 µM. By providing new information on molecules inhibiting SARS-CoV-2 replication in vitro, this study provides information for the selection of drugs to be further validated in vivo. Disclaimer: This study corresponds to the early stages of antiviral development and the results do not support by themselves the use of the selected drugs to treat SARS-CoV-2 infection.


Assuntos
Betacoronavirus/fisiologia , Bibliotecas de Moléculas Pequenas/química , Animais , Antivirais/química , Antivirais/metabolismo , Antivirais/farmacologia , Betacoronavirus/isolamento & purificação , Células CACO-2 , Sobrevivência Celular/efeitos dos fármacos , Chlorocebus aethiops , Infecções por Coronavirus/patologia , Infecções por Coronavirus/virologia , Aprovação de Drogas , Avaliação Pré-Clínica de Medicamentos , Reposicionamento de Medicamentos , Humanos , Pandemias , Pneumonia Viral/patologia , Pneumonia Viral/virologia , Bibliotecas de Moléculas Pequenas/metabolismo , Bibliotecas de Moléculas Pequenas/farmacologia , Células Vero , Replicação Viral/efeitos dos fármacos
8.
Int J Mol Sci ; 21(15)2020 Jul 23.
Artigo em Inglês | MEDLINE | ID: mdl-32718020

RESUMO

The ongoing pandemic of coronavirus disease-2019 (COVID-19) is being caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). The disease continues to present significant challenges to the health care systems around the world. This is primarily because of the lack of vaccines to protect against the infection and the lack of highly effective therapeutics to prevent and/or treat the illness. Nevertheless, researchers have swiftly responded to the pandemic by advancing old and new potential therapeutics into clinical trials. In this review, we summarize potential anti-COVID-19 therapeutics that block the early stage of the viral life cycle. The review presents the structures, mechanisms, and reported results of clinical trials of potential therapeutics that have been listed in clinicaltrials.gov. Given the fact that some of these therapeutics are multi-acting molecules, other relevant mechanisms will also be described. The reviewed therapeutics include small molecules and macromolecules of sulfated polysaccharides, polypeptides, and monoclonal antibodies. The potential therapeutics target viral and/or host proteins or processes that facilitate the early stage of the viral infection. Frequent targets are the viral spike protein, the host angiotensin converting enzyme 2, the host transmembrane protease serine 2, and clathrin-mediated endocytosis process. Overall, the review aims at presenting update-to-date details, so as to enhance awareness of potential therapeutics, and thus, to catalyze their appropriate use in combating the pandemic.


Assuntos
Antivirais/uso terapêutico , Betacoronavirus/fisiologia , Infecções por Coronavirus/tratamento farmacológico , Pneumonia Viral/tratamento farmacológico , Anticorpos Monoclonais/química , Anticorpos Monoclonais/farmacologia , Anticorpos Monoclonais/uso terapêutico , Antivirais/química , Antivirais/farmacologia , Betacoronavirus/efeitos dos fármacos , Ensaios Clínicos como Assunto , Humanos , Pandemias , Peptídeos/química , Peptídeos/farmacologia , Peptídeos/uso terapêutico , Polissacarídeos/química , Polissacarídeos/farmacologia , Polissacarídeos/uso terapêutico , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Bibliotecas de Moléculas Pequenas/uso terapêutico , Ligação Viral/efeitos dos fármacos , Internalização do Vírus/efeitos dos fármacos
9.
Proc Natl Acad Sci U S A ; 117(31): 18431-18438, 2020 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-32690700

RESUMO

Influenza hemagglutinin (HA) glycoprotein is the primary surface antigen targeted by the host immune response and a focus for development of novel vaccines, broadly neutralizing antibodies (bnAbs), and therapeutics. HA enables viral entry into host cells via receptor binding and membrane fusion and is a validated target for drug discovery. However, to date, only a very few bona fide small molecules have been reported against the HA. To identity new antiviral lead candidates against the highly conserved fusion machinery in the HA stem, we synthesized a fluorescence-polarization probe based on a recently described neutralizing cyclic peptide P7 derived from the complementarity-determining region loops of human bnAbs FI6v3 and CR9114 against the HA stem. We then designed a robust binding assay compatible with high-throughput screening to identify molecules with low micromolar to nanomolar affinity to influenza A group 1 HAs. Our simple, low-cost, and efficient in vitro assay was used to screen H1/Puerto Rico/8/1934 (H1/PR8) HA trimer against ∼72,000 compounds. The crystal structure of H1/PR8 HA in complex with our best hit compound F0045(S) confirmed that it binds to pockets in the HA stem similar to bnAbs FI6v3 and CR9114, cyclic peptide P7, and small-molecule inhibitor JNJ4796. F0045 is enantioselective against a panel of group 1 HAs and F0045(S) exhibits in vitro neutralization activity against multiple H1N1 and H5N1 strains. Our assay, compound characterization, and small-molecule candidate should further stimulate the discovery and development of new compounds with unique chemical scaffolds and enhanced influenza antiviral capabilities.


Assuntos
Antivirais/farmacologia , Avaliação Pré-Clínica de Medicamentos/métodos , Polarização de Fluorescência/métodos , Vírus da Influenza A Subtipo H1N1/efeitos dos fármacos , Virus da Influenza A Subtipo H5N1/efeitos dos fármacos , Influenza Humana/virologia , Bibliotecas de Moléculas Pequenas/farmacologia , Antivirais/química , Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Glicoproteínas de Hemaglutininação de Vírus da Influenza/metabolismo , Humanos , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H1N1/metabolismo , Virus da Influenza A Subtipo H5N1/genética , Virus da Influenza A Subtipo H5N1/metabolismo , Bibliotecas de Moléculas Pequenas/química
10.
Proc Natl Acad Sci U S A ; 117(28): 16292-16301, 2020 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-32601208

RESUMO

Notch pathway signaling is implicated in several human cancers. Aberrant activation and mutations of Notch signaling components are linked to tumor initiation, maintenance, and resistance to cancer therapy. Several strategies, such as monoclonal antibodies against Notch ligands and receptors, as well as small-molecule γ-secretase inhibitors (GSIs), have been developed to interfere with Notch receptor activation at proximal points in the pathway. However, the use of drug-like small molecules to target the downstream mediators of Notch signaling, the Notch transcription activation complex, remains largely unexplored. Here, we report the discovery of an orally active small-molecule inhibitor (termed CB-103) of the Notch transcription activation complex. We show that CB-103 inhibits Notch signaling in primary human T cell acute lymphoblastic leukemia and other Notch-dependent human tumor cell lines, and concomitantly induces cell cycle arrest and apoptosis, thereby impairing proliferation, including in GSI-resistant human tumor cell lines with chromosomal translocations and rearrangements in Notch genes. CB-103 produces Notch loss-of-function phenotypes in flies and mice and inhibits the growth of human breast cancer and leukemia xenografts, notably without causing the dose-limiting intestinal toxicity associated with other Notch inhibitors. Thus, we describe a pharmacological strategy that interferes with Notch signaling by disrupting the Notch transcription complex and shows therapeutic potential for treating Notch-driven cancers.


Assuntos
Receptores Notch/metabolismo , Bibliotecas de Moléculas Pequenas/farmacologia , Ativação Transcricional/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Sítios de Ligação , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Drosophila , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Células HeLa , Humanos , Proteína de Ligação a Sequências Sinal de Recombinação J de Imunoglobina/química , Proteína de Ligação a Sequências Sinal de Recombinação J de Imunoglobina/genética , Proteína de Ligação a Sequências Sinal de Recombinação J de Imunoglobina/metabolismo , Intestino Delgado/efeitos dos fármacos , Intestino Delgado/metabolismo , Camundongos , Mutação , Fenótipo , Multimerização Proteica , Transdução de Sinais/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/uso terapêutico
11.
Int J Mol Sci ; 21(13)2020 Jun 30.
Artigo em Inglês | MEDLINE | ID: mdl-32630064

RESUMO

Vimentin is an intermediate filament protein that plays key roles in integration of cytoskeletal functions, and therefore in basic cellular processes such as cell division and migration. Consequently, vimentin has complex implications in pathophysiology. Vimentin is required for a proper immune response, but it can also act as an autoantigen in autoimmune diseases or as a damage signal. Although vimentin is a predominantly cytoplasmic protein, it can also appear at extracellular locations, either in a secreted form or at the surface of numerous cell types, often in relation to cell activation, inflammation, injury or senescence. Cell surface targeting of vimentin appears to associate with the occurrence of certain posttranslational modifications, such as phosphorylation and/or oxidative damage. At the cell surface, vimentin can act as a receptor for bacterial and viral pathogens. Indeed, vimentin has been shown to play important roles in virus attachment and entry of severe acute respiratory syndrome-related coronavirus (SARS-CoV), dengue and encephalitis viruses, among others. Moreover, the presence of vimentin in specific virus-targeted cells and its induction by proinflammatory cytokines and tissue damage contribute to its implication in viral infection. Here, we recapitulate some of the pathophysiological implications of vimentin, including the involvement of cell surface vimentin in interaction with pathogens, with a special focus on its role as a cellular receptor or co-receptor for viruses. In addition, we provide a perspective on approaches to target vimentin, including antibodies or chemical agents that could modulate these interactions to potentially interfere with viral pathogenesis, which could be useful when multi-target antiviral strategies are needed.


Assuntos
Vírus da SARS/fisiologia , Vimentina/metabolismo , Viroses/patologia , Anticorpos/imunologia , Anticorpos/metabolismo , Anticorpos/uso terapêutico , Betacoronavirus/isolamento & purificação , Infecções por Coronavirus/tratamento farmacológico , Interações Hospedeiro-Patógeno , Humanos , Pandemias , Pneumonia Viral/tratamento farmacológico , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Bibliotecas de Moléculas Pequenas/uso terapêutico , Vimentina/química , Vimentina/imunologia , Viroses/tratamento farmacológico , Viroses/metabolismo , Replicação Viral/efeitos dos fármacos
12.
Mol Cell ; 79(3): 390-405.e7, 2020 08 06.
Artigo em Inglês | MEDLINE | ID: mdl-32619402

RESUMO

Despite their apparent lack of catalytic activity, pseudokinases are essential signaling molecules. Here, we describe the structural and dynamic properties of pseudokinase domains from the Wnt-binding receptor tyrosine kinases (PTK7, ROR1, ROR2, and RYK), which play important roles in development. We determined structures of all pseudokinase domains in this family and found that they share a conserved inactive conformation in their activation loop that resembles the autoinhibited insulin receptor kinase (IRK). They also have inaccessible ATP-binding pockets, occluded by aromatic residues that mimic a cofactor-bound state. Structural comparisons revealed significant domain plasticity and alternative interactions that substitute for absent conserved motifs. The pseudokinases also showed dynamic properties that were strikingly similar to those of IRK. Despite the inaccessible ATP site, screening identified ATP-competitive type-II inhibitors for ROR1. Our results set the stage for an emerging therapeutic modality of "conformational disruptors" to inhibit or modulate non-catalytic functions of pseudokinases deregulated in disease.


Assuntos
Moléculas de Adesão Celular/química , Inibidores de Proteínas Quinases/farmacologia , Receptores Proteína Tirosina Quinases/química , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/química , Sequência de Aminoácidos , Animais , Baculoviridae/genética , Baculoviridae/metabolismo , Sítios de Ligação , Moléculas de Adesão Celular/antagonistas & inibidores , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismo , Linhagem Celular , Clonagem Molecular , Cristalografia por Raios X , Expressão Gênica , Humanos , Camundongos , Modelos Moleculares , Células Precursoras de Linfócitos B/citologia , Células Precursoras de Linfócitos B/metabolismo , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Inibidores de Proteínas Quinases/química , Receptores Proteína Tirosina Quinases/antagonistas & inibidores , Receptores Proteína Tirosina Quinases/genética , Receptores Proteína Tirosina Quinases/metabolismo , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/antagonistas & inibidores , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/genética , Receptores Órfãos Semelhantes a Receptor Tirosina Quinase/metabolismo , Receptores da Família Eph/antagonistas & inibidores , Receptores da Família Eph/química , Receptores da Família Eph/genética , Receptores da Família Eph/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Células Sf9 , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Spodoptera , Homologia Estrutural de Proteína , Especificidade por Substrato
13.
J Comput Aided Mol Des ; 34(10): 1091-1103, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32601839

RESUMO

Herein, the LASSBio Chemical Library is presented as a valuable source of compounds for screening to identify hits suitable for subsequent hit-to-lead optimization stages. A feature of the LASSBio Chemical Library worth highlighting is the fact that it is a smart library designed by medicinal chemists with pharmacological activity as the main priority. The great majority of the compounds part of this library have shown in vivo activity in animal models, which is an indication that they possess overall favorable bioavailability properties and, hence, adequate pharmacokinetic profiles. This, in turn, is supported by the fact that approximately 85% of the compounds are compliant with Lipinski's rule of five and ca. 95% are compliant with Veber's rules, two important guidelines for oral bioavailability. In this work it is presented a virtual screening methodology combining a pharmacophore-based model and an empirical Gibbs free energy-based model for the ligand-protein interaction to explore the LASSBio Chemical Library as a source of new hits for the inhibition of the phosphatidylinositol 4-kinase IIIß (PI4KIIIß) enzyme, which is related to the development of viral infections (including enteroviruses, SARS coronavirus, and hepatitis C virus), cancers and neurological diseases. The approach resulted in the identification of two hits, LASSBio-1799 (7) and LASSBio-1814 (10), which inhibited the target enzyme with IC50 values of 3.66 µM and IC50 and 6.09 µM, respectively. This study also enabled the determination of the structural requirements for interactions with the active site of PI4KIIIß, demonstrating the importance of both acceptor and donor hydrogen bonding groups for forming interactions with binding site residues Val598 and Lys549.


Assuntos
Avaliação Pré-Clínica de Medicamentos/métodos , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Fosfotransferases (Aceptor do Grupo Álcool)/antagonistas & inibidores , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Sítios de Ligação , Domínio Catalítico , Ligação de Hidrogênio , Ligantes , Modelos Moleculares , Fosfotransferases (Aceptor do Grupo Álcool)/química , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia
14.
Biomolecules ; 10(7)2020 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-32679775

RESUMO

Therapeutic strategies for the treatment of any severe disease are based on the discovery and validation of druggable targets. The human genome encodes only 600-1500 targets for small-molecule drugs, but posttranslational modifications lead to a considerably larger druggable proteome. The spontaneous conversion of asparagine (Asn) residues to aspartic acid or isoaspartic acid is a frequent modification in proteins as part of the process called deamidation. Triosephosphate isomerase (TIM) is a glycolytic enzyme whose deamidation has been thoroughly studied, but the prospects of exploiting this phenomenon for drug design remain poorly understood. The purpose of this study is to demonstrate the properties of deamidated human TIM (HsTIM) as a selective molecular target. Using in silico prediction, in vitro analyses, and a bacterial model lacking the tim gene, this study analyzed the structural and functional differences between deamidated and nondeamidated HsTIM, which account for the efficacy of this protein as a druggable target. The highly increased permeability and loss of noncovalent interactions of deamidated TIM were found to play a central role in the process of selective enzyme inactivation and methylglyoxal production. This study elucidates the properties of deamidated HsTIM regarding its selective inhibition by thiol-reactive drugs and how these drugs can contribute to the development of cell-specific therapeutic strategies for a variety of diseases, such as COVID-19 and cancer.


Assuntos
Infecções por Coronavirus/tratamento farmacológico , Inibidores Enzimáticos/farmacologia , Neoplasias/tratamento farmacológico , Pneumonia Viral/tratamento farmacológico , Bibliotecas de Moléculas Pequenas/farmacologia , Triose-Fosfato Isomerase/antagonistas & inibidores , Amidas/antagonistas & inibidores , Amidas/metabolismo , Cristalografia por Raios X , Inibidores Enzimáticos/química , Humanos , Modelos Moleculares , Mutação , Pandemias , Proteoma/antagonistas & inibidores , Proteoma/genética , Proteoma/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/isolamento & purificação , Proteínas Recombinantes/metabolismo , Bibliotecas de Moléculas Pequenas/química , Triose-Fosfato Isomerase/química , Triose-Fosfato Isomerase/metabolismo
15.
Proc Natl Acad Sci U S A ; 117(29): 17195-17203, 2020 07 21.
Artigo em Inglês | MEDLINE | ID: mdl-32606248

RESUMO

The vast majority of intracellular protein targets are refractory toward small-molecule therapeutic engagement, and additional therapeutic modalities are needed to overcome this deficiency. Here, the identification and characterization of a natural product, WDB002, reveals a therapeutic modality that dramatically expands the currently accepted limits of druggability. WDB002, in complex with the FK506-binding protein (FKBP12), potently and selectively binds the human centrosomal protein 250 (CEP250), resulting in disruption of CEP250 function in cells. The recognition mode is unprecedented in that the targeted domain of CEP250 is a coiled coil and is topologically featureless, embodying both a structural motif and surface topology previously considered on the extreme limits of "undruggability" for an intracellular target. Structural studies reveal extensive protein-WDB002 and protein-protein contacts, with the latter being distinct from those seen in FKBP12 ternary complexes formed by FK506 and rapamycin. Outward-facing structural changes in a bound small molecule can thus reprogram FKBP12 to engage diverse, otherwise "undruggable" targets. The flat-targeting modality demonstrated here has the potential to expand the druggable target range of small-molecule therapeutics. As CEP250 was recently found to be an interaction partner with the Nsp13 protein of the SARS-CoV-2 virus that causes COVID-19 disease, it is possible that WDB002 or an analog may exert useful antiviral activity through its ability to form high-affinity ternary complexes containing CEP250 and FKBP12.


Assuntos
Actinobacteria/genética , Genoma Bacteriano , Domínios e Motivos de Interação entre Proteínas/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/farmacologia , Proteína 1A de Ligação a Tacrolimo/química , Proteína 1A de Ligação a Tacrolimo/metabolismo , Actinobacteria/metabolismo , Sequência de Aminoácidos , Antibacterianos/química , Antibacterianos/metabolismo , Autoantígenos/genética , Autoantígenos/metabolismo , Calcineurina/genética , Calcineurina/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Evolução Molecular , Células HEK293 , Humanos , Modelos Moleculares , Conformação Proteica , Homologia de Sequência , Sirolimo/química , Sirolimo/metabolismo , Bibliotecas de Moléculas Pequenas/química , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo
16.
Proc Natl Acad Sci U S A ; 117(29): 16782-16789, 2020 07 21.
Artigo em Inglês | MEDLINE | ID: mdl-32641511

RESUMO

DNA-encoded chemical libraries are collections of compounds individually coupled to unique DNA tags serving as amplifiable identification barcodes. By bridging split-and-pool combinatorial synthesis with the ligation of unique encoding DNA oligomers, million- to billion-member libraries can be synthesized for use in hundreds of healthcare target screens. Although structural diversity and desirable molecular property ranges generally guide DNA-encoded chemical library design, recent reports have highlighted the utility of focused DNA-encoded chemical libraries that are structurally biased for a class of protein targets. Herein, a protease-focused DNA-encoded chemical library was designed that utilizes chemotypes known to engage conserved catalytic protease residues. The three-cycle library features functional moieties such as guanidine, which interacts strongly with aspartate of the protease catalytic triad, as well as mild electrophiles such as sulfonamide, urea, and carbamate. We developed a DNA-compatible method for guanidinylation of amines and reduction of nitriles. Employing these optimized reactions, we constructed a 9.8-million-membered DNA-encoded chemical library. Affinity selection of the library with thrombin, a common protease, revealed a number of enriched features which ultimately led to the discovery of a 1 nM inhibitor of thrombin. Thus, structurally focused DNA-encoded chemical libraries have tremendous potential to find clinically useful high-affinity hits for the rapid discovery of drugs for targets (e.g., proteases) with essential functions in infectious diseases (e.g., severe acute respiratory syndrome coronavirus 2) and relevant healthcare conditions (e.g., male contraception).


Assuntos
DNA/química , DNA/metabolismo , Descoberta de Drogas , Biblioteca Gênica , Inibidores de Proteases/farmacologia , Bibliotecas de Moléculas Pequenas/farmacologia , Trombina/antagonistas & inibidores , Técnicas de Química Combinatória , Humanos , Inibidores de Proteases/química , Bibliotecas de Moléculas Pequenas/química
17.
Nat Chem Biol ; 16(7): 716-724, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32572259

RESUMO

Largely non-overlapping sets of cyclin-dependent kinases (CDKs) regulate cell division and RNA polymerase II (Pol II)-dependent transcription. Here we review the molecular mechanisms by which specific CDKs are thought to act at discrete steps in the transcription cycle and describe the recent emergence of transcriptional CDKs as promising drug targets in cancer. We emphasize recent advances in understanding the transcriptional CDK network that were facilitated by development and deployment of small-molecule inhibitors with increased selectivity for individual CDKs. Unexpectedly, several of these compounds have also shown selectivity in killing cancer cells, despite the seemingly universal involvement of their target CDKs during transcription in all cells. Finally, we describe remaining and emerging challenges in defining functions of individual CDKs in transcription and co-transcriptional processes and in leveraging CDK inhibition for therapeutic purposes.


Assuntos
Antineoplásicos/farmacologia , Quinases Ciclina-Dependentes/antagonistas & inibidores , Regulação Neoplásica da Expressão Gênica , Neoplasias/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , RNA Polimerase II/genética , Bibliotecas de Moléculas Pequenas/farmacologia , Animais , Antineoplásicos/química , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/genética , Quinases Ciclina-Dependentes/genética , Quinases Ciclina-Dependentes/metabolismo , Modelos Animais de Doenças , Humanos , Neoplasias/enzimologia , Neoplasias/genética , Neoplasias/patologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Inibidores de Proteínas Quinases/química , Proteólise , RNA Polimerase II/antagonistas & inibidores , RNA Polimerase II/metabolismo , Transdução de Sinais , Bibliotecas de Moléculas Pequenas/química , Transcrição Genética
18.
Nature ; 585(7824): 293-297, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32494016

RESUMO

Molecular glue compounds induce protein-protein interactions that, in the context of a ubiquitin ligase, lead to protein degradation1. Unlike traditional enzyme inhibitors, these molecular glue degraders act substoichiometrically to catalyse the rapid depletion of previously inaccessible targets2. They are clinically effective and highly sought-after, but have thus far only been discovered serendipitously. Here, through systematically mining databases for correlations between the cytotoxicity of 4,518 clinical and preclinical small molecules and the expression levels of E3 ligase components across hundreds of human cancer cell lines3-5, we identify CR8-a cyclin-dependent kinase (CDK) inhibitor6-as a compound that acts as a molecular glue degrader. The CDK-bound form of CR8 has a solvent-exposed pyridyl moiety that induces the formation of a complex between CDK12-cyclin K and the CUL4 adaptor protein DDB1, bypassing the requirement for a substrate receptor and presenting cyclin K for ubiquitination and degradation. Our studies demonstrate that chemical alteration of surface-exposed moieties can confer gain-of-function glue properties to an inhibitor, and we propose this as a broader strategy through which target-binding molecules could be converted into molecular glues.


Assuntos
Ciclinas/deficiência , Ciclinas/metabolismo , Proteólise/efeitos dos fármacos , Purinas/química , Purinas/farmacologia , Piridinas/química , Piridinas/farmacologia , Linhagem Celular Tumoral , Quinases Ciclina-Dependentes/antagonistas & inibidores , Quinases Ciclina-Dependentes/química , Quinases Ciclina-Dependentes/metabolismo , Ciclinas/química , Proteínas de Ligação a DNA/metabolismo , Humanos , Modelos Moleculares , Complexo de Endopeptidases do Proteassoma/metabolismo , Ligação Proteica/efeitos dos fármacos , Purinas/toxicidade , Piridinas/toxicidade , Bibliotecas de Moléculas Pequenas/análise , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Ubiquitinação/efeitos dos fármacos
19.
Virus Res ; 286: 198068, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32565126

RESUMO

The emergence of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has led to a renewed interest in studying the role of the spike S glycoprotein in regulating coronavirus infections in the natural host. Taking advantage of the cryo-electron microscopy structure of SARS-CoV-2 S trimer in the prefusion conformation, we performed a virtual screening simulation with the aim to identify novel molecules that could be used as fusion inhibitors. The spike glycoprotein structure has been completed using modeling techniques and its inner cavity, needful for the postfusion transition of the trimer, has been scanned for the identification of strongly interacting available drugs. Finally, the stability of the protein-drug top complexes has been tested using classical molecular dynamics simulations. The free energy of interaction of the molecules to the spike protein has been evaluated through the MM/GBSA method and per-residue decomposition analysis. Results have been critically discussed considering previous scientific knowledge concerning the selected compounds and sequence alignments have been carried out to evaluate the spike glycoprotein similarity among the betacoronavirus family members. Finally, a cocktail of drugs that may be used as SARS-CoV-2 fusion inhibitors has been suggested.


Assuntos
Antivirais/química , Betacoronavirus/química , Compostos Heterocíclicos de 4 ou mais Anéis/química , Indóis/química , Perileno/análogos & derivados , Glicoproteína da Espícula de Coronavírus/química , Sulfonamidas/química , Antivirais/farmacologia , Betacoronavirus/patogenicidade , Sítios de Ligação , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/virologia , Expressão Gênica , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Ensaios de Triagem em Larga Escala , Humanos , Indóis/farmacologia , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , Pandemias , Perileno/química , Perileno/farmacologia , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/virologia , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Multimerização Proteica , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Glicoproteína da Espícula de Coronavírus/antagonistas & inibidores , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/metabolismo , Sulfonamidas/farmacologia , Termodinâmica , Interface Usuário-Computador , Internalização do Vírus/efeitos dos fármacos
20.
Nat Commun ; 11(1): 3183, 2020 06 23.
Artigo em Inglês | MEDLINE | ID: mdl-32576815

RESUMO

High-diversity genetically-encoded combinatorial libraries (108-1013 members) are a rich source of peptide-based binding molecules, identified by affinity selection. Synthetic libraries can access broader chemical space, but typically examine only ~ 106 compounds by screening. Here we show that in-solution affinity selection can be interfaced with nano-liquid chromatography-tandem mass spectrometry peptide sequencing to identify binders from fully randomized synthetic libraries of 108 members-a 100-fold gain in diversity over standard practice. To validate this approach, we show that binders to a monoclonal antibody are identified in proportion to library diversity, as diversity is increased from 106-108. These results are then applied to the discovery of p53-like binders to MDM2, and to a family of 3-19 nM-affinity, α/ß-peptide-based binders to 14-3-3. An X-ray structure of one of these binders in complex with 14-3-3σ is determined, illustrating the role of ß-amino acids in facilitating a key binding contact.


Assuntos
Biblioteca de Peptídeos , Peptídeos/química , Bibliotecas de Moléculas Pequenas/química , Sequência de Aminoácidos , Aminoácidos , Anticorpos Monoclonais/química , Afinidade de Anticorpos , Proteínas de Transporte/química , Cromatografia Líquida , Cristalografia por Raios X , Desenho de Fármacos , Descoberta de Drogas , Modelos Moleculares , Proteínas Proto-Oncogênicas c-mdm2/química , Proteínas Proto-Oncogênicas c-mdm2/metabolismo
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