Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 40
Filtrar
1.
ChemMedChem ; 18(6): e202200686, 2023 03 14.
Artículo en Inglés | MEDLINE | ID: mdl-36649575

RESUMEN

The bromodomain and PHD-finger containing transcription factor (BPTF) is part of the nucleosome remodeling factor (NURF) complex and has been implicated in multiple cancer types. Here, we report the discovery of a potent and selective chemical probe targeting the bromodomain of BPTF with an attractive pharmacokinetic profile enabling cellular and in vivo experiments in mice. Microarray-based transcriptomics in presence of the probe in two lung cancer cell lines revealed only minor effects on the transcriptome. Profiling against a panel of cancer cell lines revealed that the antiproliferative effect does not correlate with BPTF dependency score in depletion screens. Both observations and the multi-domain architecture of BPTF suggest that depleting the protein by proteolysis targeting chimeras (PROTACs) could be a promising strategy to target cancer cell proliferation. We envision that the presented chemical probe and the related negative control will enable the research community to further explore scientific hypotheses with respect to BPTF bromodomain inhibition.


Asunto(s)
Neoplasias Pulmonares , Factores de Transcripción , Animales , Ratones , Proliferación Celular , Regulación de la Expresión Génica , Proteínas Nucleares/metabolismo , Factores de Transcripción/metabolismo
2.
Nat Commun ; 13(1): 5969, 2022 10 10.
Artículo en Inglés | MEDLINE | ID: mdl-36216795

RESUMEN

Targeted protein degradation offers an alternative modality to classical inhibition and holds the promise of addressing previously undruggable targets to provide novel therapeutic options for patients. Heterobifunctional molecules co-recruit a target protein and an E3 ligase, resulting in ubiquitylation and proteosome-dependent degradation of the target. In the clinic, the oral route of administration is the option of choice but has only been achieved so far by CRBN- recruiting bifunctional degrader molecules. We aimed to achieve orally bioavailable molecules that selectively degrade the BAF Chromatin Remodelling complex ATPase SMARCA2 over its closely related paralogue SMARCA4, to allow in vivo evaluation of the synthetic lethality concept of SMARCA2 dependency in SMARCA4-deficient cancers. Here we outline structure- and property-guided approaches that led to orally bioavailable VHL-recruiting degraders. Our tool compound, ACBI2, shows selective degradation of SMARCA2 over SMARCA4 in ex vivo human whole blood assays and in vivo efficacy in SMARCA4-deficient cancer models. This study demonstrates the feasibility for broadening the E3 ligase and physicochemical space that can be utilised for achieving oral efficacy with bifunctional molecules.


Asunto(s)
Adenosina Trifosfatasas , Factores de Transcripción , Adenosina Trifosfatasas/genética , Adenosina Trifosfatasas/metabolismo , ADN Helicasas/genética , ADN Helicasas/metabolismo , Humanos , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteolisis , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Ubiquitina-Proteína Ligasas/genética , Ubiquitina-Proteína Ligasas/metabolismo , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/genética , Proteína Supresora de Tumores del Síndrome de Von Hippel-Lindau/metabolismo
3.
Nat Cancer ; 3(7): 821-836, 2022 07.
Artículo en Inglés | MEDLINE | ID: mdl-35883003

RESUMEN

Oncogenic alterations in human epidermal growth factor receptor 2 (HER2) occur in approximately 2% of patients with non-small cell lung cancer and predominantly affect the tyrosine kinase domain and cluster in exon 20 of the ERBB2 gene. Most clinical-grade tyrosine kinase inhibitors are limited by either insufficient selectivity against wild-type (WT) epidermal growth factor receptor (EGFR), which is a major cause of dose-limiting toxicity or by potency against HER2 exon 20 mutant variants. Here we report the discovery of covalent tyrosine kinase inhibitors that potently inhibit HER2 exon 20 mutants while sparing WT EGFR, which reduce tumor cell survival and proliferation in vitro and result in regressions in preclinical xenograft models of HER2 exon 20 mutant non-small cell lung cancer, concomitant with inhibition of downstream HER2 signaling. Our results suggest that HER2 exon 20 insertion-driven tumors can be effectively treated by a potent and highly selective HER2 inhibitor while sparing WT EGFR, paving the way for clinical translation.


Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Receptores ErbB/genética , Exones/genética , Genes erbB-2 , Humanos , Neoplasias Pulmonares/tratamiento farmacológico , Inhibidores de Proteínas Quinasas/farmacología , Receptor ErbB-2/genética
4.
ChemMedChem ; 16(23): 3576-3587, 2021 12 06.
Artículo en Inglés | MEDLINE | ID: mdl-34524728

RESUMEN

The NRF2 transcription factor is a key regulator in cellular oxidative stress response, and acts as a tumor suppressor. Aberrant activation of NRF2 has been implicated in promoting chemo-resistance, tumor growth, and metastasis by activating its downstream target genes. Hence, inhibition of NRF2 promises to be an attractive therapeutic strategy to suppress cell proliferation and enhance cell apoptosis in cancer. Direct targeting of NRF2 with small-molecules to discover protein-DNA interaction inhibitors is challenging as it is a largely intrinsically disordered protein. To discover molecules that bind to NRF2 at the DNA binding interface, we performed an NMR-based fragment screen against its DNA-binding domain. We discovered several weakly binding fragment hits that bind to a region overlapping with the DNA binding site. Using SAR by catalogue we developed an initial structure-activity relationship for the most interesting initial hit series. By combining NMR chemical shift perturbations and data-driven docking, binding poses which agreed with NMR information and the observed SAR were elucidated. The herein discovered NRF2 hits and proposed binding modes form the basis for future structure-based optimization campaigns on this important but to date 'undrugged' cancer driver.


Asunto(s)
ADN/antagonistas & inhibidores , Factor 2 Relacionado con NF-E2/antagonistas & inhibidores , Unión Proteica/efectos de los fármacos , Bibliotecas de Moléculas Pequeñas/química , Sitios de Unión , ADN/metabolismo , Humanos , Simulación del Acoplamiento Molecular , Estructura Molecular , Factor 2 Relacionado con NF-E2/química , Factor 2 Relacionado con NF-E2/metabolismo , Resonancia Magnética Nuclear Biomolecular , Dominios Proteicos , Relación Estructura-Actividad
5.
ChemMedChem ; 16(9): 1420-1424, 2021 05 06.
Artículo en Inglés | MEDLINE | ID: mdl-33275320

RESUMEN

Aberrant WNT pathway activation, leading to nuclear accumulation of ß-catenin, is a key oncogenic driver event. Mutations in the tumor suppressor gene APC lead to impaired proteasomal degradation of ß-catenin and subsequent nuclear translocation. Restoring cellular degradation of ß-catenin represents a potential therapeutic strategy. Here, we report the fragment-based discovery of a small molecule binder to ß-catenin, including the structural elucidation of the binding mode by X-ray crystallography. The difficulty in drugging ß-catenin was confirmed as the primary screening campaigns identified only few and very weak hits. Iterative virtual and NMR screening techniques were required to discover a compound with sufficient potency to be able to obtain an X-ray co-crystal structure. The binding site is located between armadillo repeats two and three, adjacent to the BCL9 and TCF4 binding sites. Genetic studies show that it is unlikely to be useful for the development of protein-protein interaction inhibitors but structural information and established assays provide a solid basis for a prospective optimization towards ß-catenin proteolysis targeting chimeras (PROTACs) as alternative modality.


Asunto(s)
Bibliotecas de Moléculas Pequeñas/química , beta Catenina/antagonistas & inhibidores , Sitios de Unión , Cristalografía por Rayos X , Humanos , Simulación de Dinámica Molecular , Mapas de Interacción de Proteínas/efectos de los fármacos , Bibliotecas de Moléculas Pequeñas/metabolismo , Bibliotecas de Moléculas Pequeñas/farmacología , Relación Estructura-Actividad , beta Catenina/metabolismo
6.
Antioxidants (Basel) ; 9(2)2020 Feb 21.
Artículo en Inglés | MEDLINE | ID: mdl-32098277

RESUMEN

The flavonoid kaempferol is almost ubiquitously contained in edible and medicinal plants and exerts a broad range of interesting pharmacological activities. Interactions with central inflammatory processes can be exploited to treat or attenuate symptoms of disorders associated with chronic immune activation during infections, malignancies, and neurodegenerative or cardiovascular disorders. Many drugs, phytochemicals, and nutritional components target the catabolism of the essential amino acid tryptophan by indoleamine 2,3-dioxygenase 1 (IDO-1) for immunomodulation. We studied the effects of kaempferol by in vitro models with human peripheral blood mononuclear cells (PBMC) and THP-1 derived human myelomonocytic cell lines. Kaempferol suppressed interferon-γ dependent immunometabolic pathways: Formation of the oxidative stress biomarker neopterin and catabolism of tryptophan were inhibited dose-dependently in stimulated cells. In-silico docking studies revealed a potential interaction of kaempferol with the catalytic domain of IDO-1. Kaempferol stimulated nuclear factor kappa B (NF-κB) signaling in lipopolysaccharide (LPS)-treated THP-1 cells, thereby increasing the mRNA expression of interleukin (IL) 1 beta, tumor necrosis factor, and nuclear factor kappa B subunit 1, while IL6 was downregulated. Data suggest that concerted effects of kaempferol on multiple immunologically relevant targets are responsible for its immunomodulatory activity. However, the immunosuppressive effects may be more relevant in a T-cell dominated context.

7.
J Med Chem ; 62(22): 10272-10293, 2019 11 27.
Artículo en Inglés | MEDLINE | ID: mdl-31689114

RESUMEN

The epidermal growth factor receptor (EGFR), when carrying an activating mutation like del19 or L858R, acts as an oncogenic driver in a subset of lung tumors. While tumor responses to tyrosine kinase inhibitors (TKIs) are accompanied by marked tumor shrinkage, the response is usually not durable. Most patients relapse within two years of therapy often due to acquisition of an additional mutation in EGFR kinase domain that confers resistance to TKIs. Crucially, oncogenic EGFR harboring both resistance mutations, T790M and C797S, can no longer be inhibited by currently approved EGFR TKIs. Here, we describe the discovery of BI-4020, which is a noncovalent, wild-type EGFR sparing, macrocyclic TKI. BI-4020 potently inhibits the above-described EGFR variants and induces tumor regressions in a cross-resistant EGFRdel19 T790M C797S xenograft model. Key was the identification of a highly selective but moderately potent benzimidazole followed by complete rigidification of the molecule through macrocyclization.


Asunto(s)
Antineoplásicos/química , Antineoplásicos/farmacología , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Animales , Antineoplásicos/farmacocinética , Bencimidazoles/química , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Pulmón de Células no Pequeñas/patología , Línea Celular Tumoral , Cristalografía por Rayos X , Ciclización , Entropía , Receptores ErbB/antagonistas & inhibidores , Receptores ErbB/química , Receptores ErbB/genética , Femenino , Hepatocitos , Humanos , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Ratones , Ratones Transgénicos , Mutación , Conformación Proteica , Inhibidores de Proteínas Quinasas/farmacocinética , Relación Estructura-Actividad , Ensayos Antitumor por Modelo de Xenoinjerto
8.
Nat Chem Biol ; 15(8): 822-829, 2019 08.
Artículo en Inglés | MEDLINE | ID: mdl-31285596

RESUMEN

Here, we report the fragment-based discovery of BI-9321, a potent, selective and cellular active antagonist of the NSD3-PWWP1 domain. The human NSD3 protein is encoded by the WHSC1L1 gene located in the 8p11-p12 amplicon, frequently amplified in breast and squamous lung cancer. Recently, it was demonstrated that the PWWP1 domain of NSD3 is required for the viability of acute myeloid leukemia cells. To further elucidate the relevance of NSD3 in cancer biology, we developed a chemical probe, BI-9321, targeting the methyl-lysine binding site of the PWWP1 domain with sub-micromolar in vitro activity and cellular target engagement at 1 µM. As a single agent, BI-9321 downregulates Myc messenger RNA expression and reduces proliferation in MOLM-13 cells. This first-in-class chemical probe BI-9321, together with the negative control BI-9466, will greatly facilitate the elucidation of the underexplored biological function of PWWP domains.


Asunto(s)
N-Metiltransferasa de Histona-Lisina/antagonistas & inhibidores , Proteínas Nucleares/antagonistas & inhibidores , Sistemas CRISPR-Cas , Línea Celular , Proliferación Celular/efectos de los fármacos , Supervivencia Celular , Regulación de la Expresión Génica/efectos de los fármacos , N-Metiltransferasa de Histona-Lisina/genética , N-Metiltransferasa de Histona-Lisina/metabolismo , Humanos , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Dominios Proteicos , Proteínas Proto-Oncogénicas c-myc/genética , Proteínas Proto-Oncogénicas c-myc/metabolismo
9.
ChemMedChem ; 14(1): 88-93, 2019 01 08.
Artículo en Inglés | MEDLINE | ID: mdl-30458062

RESUMEN

Mouse double minute 2 (MDM2) is a main and direct inhibitor of the crucial tumor suppressor p53. Reports from initial clinical trials showed that blocking this interaction with a small-molecule inhibitor can have great value in the treatment of cancer for patients with p53 wild-type tumors; however, it also revealed dose-limiting hematological toxicities and drug-induced resistance as main issues. To overcome the former, an inhibitor with superior potency and pharmacokinetic properties to ultimately achieve full efficacy with less-frequent dosing schedules is required. Toward this aim, we optimized our recently reported spiro-oxindole inhibitors by focusing on the crucial interaction with the amino acid side chain of His96MDM2 . The designed molecules required the targeted synthesis of structurally complex spiro[indole-3,2'-pyrrolo[2,3-c]pyrrole]-2,4'-diones for which we developed an unprecedented intramolecular azomethine ylide cycloaddition and investigated the results by computational methods. One of the new compounds showed superior cellular potency over previously reported BI-0252. This finding is a significant step toward an inhibitor suitable to potentially mitigate hematological on-target adverse effects.


Asunto(s)
Compuestos Azo/farmacología , Indoles/farmacología , Pirrolidinonas/farmacología , Compuestos de Espiro/farmacología , Tiosemicarbazonas/farmacología , Animales , Compuestos Azo/química , Línea Celular Tumoral , Cristalografía por Rayos X , Ciclización , Teoría Funcional de la Densidad , Relación Dosis-Respuesta a Droga , Humanos , Indoles/síntesis química , Indoles/química , Ratones , Modelos Moleculares , Estructura Molecular , Proteínas Proto-Oncogénicas c-mdm2/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-mdm2/química , Proteínas Proto-Oncogénicas c-mdm2/metabolismo , Pirrolidinonas/síntesis química , Pirrolidinonas/química , Compuestos de Espiro/síntesis química , Compuestos de Espiro/química , Relación Estructura-Actividad , Tiosemicarbazonas/química , Proteína p53 Supresora de Tumor/antagonistas & inhibidores , Proteína p53 Supresora de Tumor/química , Proteína p53 Supresora de Tumor/metabolismo
10.
J Chem Inf Model ; 59(1): 137-148, 2019 01 28.
Artículo en Inglés | MEDLINE | ID: mdl-30532974

RESUMEN

The protozoan cysteine proteases cruzain in Trypanosoma cruzi and rhodesain in Trypanosoma brucei are therapeutic targets for Chagas disease and Human African Trypanosomiasis (HAT), respectively. A benzimidazole series was previously characterized as potent noncovalent competitive cruzain and rhodesain inhibitors with activity against trypanosomes. Common structure-activity relationships (SAR) trends and structural modifications leading to selectivity against each enzyme were described. However, some of these trends could not be understood based on the reported binding mode of lead compound 1. Therefore, we employed microsecond molecular dynamics simulations and free energy calculations to understand qualitative SAR trends and to quantitatively recapitulate them. Simulations revealed the most stable protein-ligand interactions and provided insights concerning enzyme selectivity. Calculated relative binding free energies of compound 1 analogs exhibited deviations of 1.1 and 2.2 kcal/mol from the experimental values for cruzain and rhodesain, respectively. These data encourage prospective thermodynamic integration (TI) studies to optimize this series and facilitate the prioritization of compounds for synthesis.


Asunto(s)
Bencimidazoles/química , Inhibidores de Cisteína Proteinasa/química , Simulación del Acoplamiento Molecular , Trypanosoma brucei brucei/enzimología , Animales , Cisteína Endopeptidasas/metabolismo , Interacciones Hidrofóbicas e Hidrofílicas , Proteínas Protozoarias/química , Proteínas Protozoarias/metabolismo , Relación Estructura-Actividad
11.
Proc Natl Acad Sci U S A ; 115(44): E10505-E10514, 2018 10 30.
Artículo en Inglés | MEDLINE | ID: mdl-30309962

RESUMEN

Human BCL-2-associated death promoter (hBAD) is an apoptosis-regulatory protein mediating survival signals to carcinoma cells upon phosphorylation of Ser99, among other residues. Herein, we screened multiple small-molecule databases queried in a Laplacian-modified naive Bayesian-based cheminformatics platform and identified a Petasis reaction product as a site-specific inhibitor for hBAD phosphorylation. Based on apoptotic efficacy against mammary carcinoma cells, N-cyclopentyl-3-((4-(2,3-dichlorophenyl) piperazin-1-yl) (2-hydroxyphenyl) methyl) benzamide (NPB) was identified as a potential lead compound. In vitro biochemical analyses demonstrated that NPB inhibited the phosphorylation of hBAD specifically on Ser99. NPB was observed to exert this effect independently of AKT and other kinase activities despite the demonstration of AKT-mediated BAD-Ser99 phosphorylation. Using a structure-based bioinformatics platform, we observed that NPB exhibited predicted interactions with hBAD in silico and verified the same by direct binding kinetics. NPB reduced phosphorylation of BAD-Ser99 and enhanced caspase 3/7 activity with associated loss of cell viability in various human cancer cell lines derived from mammary, endometrial, ovarian, hepatocellular, colon, prostatic, and pancreatic carcinoma. Furthermore, by use of a xenograft model, it was observed that NPB, as a single agent, markedly diminished BAD phosphorylation in tumor tissue and significantly inhibited tumor growth. Similar doses of NPB utilized in acute toxicity studies in mice did not exhibit significant effects. Hence, we report a site-specific inhibitor of BAD phosphorylation with efficacy in tumor models.


Asunto(s)
Antineoplásicos/farmacología , Benzamidas/farmacología , Supervivencia Celular/efectos de los fármacos , Piperazinas/farmacología , Serina/química , Proteína Letal Asociada a bcl/antagonistas & inhibidores , Antineoplásicos/química , Apoptosis , Benzamidas/química , Proliferación Celular , Bases de Datos Factuales , Sistemas de Liberación de Medicamentos , Descubrimiento de Drogas , Humanos , Células MCF-7 , Fosforilación , Piperazinas/química , Interferencia de ARN , Bibliotecas de Moléculas Pequeñas , Resonancia por Plasmón de Superficie
12.
J Chem Inf Model ; 58(5): 982-992, 2018 05 29.
Artículo en Inglés | MEDLINE | ID: mdl-29652495

RESUMEN

Macrocycles are of considerable interest as highly specific drug candidates, yet they challenge standard conformer generators with their large number of rotatable bonds and conformational restrictions. Here, we present a molecular dynamics-based routine that bypasses current limitations in conformational sampling and extensively profiles the free energy landscape of peptidic macrocycles in solution. We perform accelerated molecular dynamics simulations to capture a diverse conformational ensemble. By applying an energetic cutoff, followed by geometric clustering, we demonstrate the striking robustness and efficiency of the approach in identifying highly populated conformational states of cyclic peptides. The resulting structural and thermodynamic information is benchmarked against interproton distances from NMR experiments and conformational states identified by X-ray crystallography. Using three different model systems of varying size and flexibility, we show that the method reliably reproduces experimentally determined structural ensembles and is capable of identifying key conformational states that include the bioactive conformation. Thus, the described approach is a robust method to generate conformations of peptidic macrocycles and holds promise for structure-based drug design.


Asunto(s)
Compuestos Macrocíclicos/química , Simulación de Dinámica Molecular , Péptidos/química , Conformación Proteica , Termodinámica
13.
J Biomol Struct Dyn ; 36(15): 4072-4084, 2018 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-29210603

RESUMEN

A ten microsecond molecular dynamics simulation of a kallikrein-related peptidase 7 peptide complex revealed an unexpected change in binding mode. After more than two microseconds unrestrained sampling we observe a spontaneous transition of the binding pose including a 180° rotation around the P1 residue. Subsequently, the substrate peptide occupies the prime side region rather than the cognate non-prime side in a stable conformation. We characterize the unexpected binding mode in terms of contacts, solvent-accessible surface area, molecular interactions and energetic properties. We compare the new pose to inhibitor-bound structures of kallikreins with occupied prime side and find that a similar orientation is adopted. Finally, we apply in silico mutagenesis based on the alternative peptide binding position to explore the prime side specificity of kallikrein-related peptidase 7 and compare it to available experimental data. Our study provides the first microsecond time scale simulation data on a kallikrein protease and shows previously unexplored prime side interactions. Therefore, we expect our study to advance the rational design of inhibitors targeting kallikrein-related peptidase 7, an emerging drug target involved in several skin diseases as well as cancer.


Asunto(s)
Clorometilcetonas de Aminoácidos/química , Calicreínas/química , Simulación de Dinámica Molecular , Oligopéptidos/química , Inhibidores de Proteasas/química , Dominio Catalítico , Humanos , Calicreínas/antagonistas & inhibidores , Cinética , Ligandos , Unión Proteica , Conformación Proteica en Hélice alfa , Conformación Proteica en Lámina beta , Dominios y Motivos de Interacción de Proteínas , Especificidad por Sustrato , Termodinámica
14.
Biophys J ; 113(7): 1485-1495, 2017 Oct 03.
Artículo en Inglés | MEDLINE | ID: mdl-28978442

RESUMEN

Mutations of positively charged amino acids in the S4 transmembrane segment of a voltage-gated ion channel form ion-conducting pathways through the voltage-sensing domain, named ω-current. Here, we used structure modeling and MD simulations to predict pathogenic ω-currents in CaV1.1 and CaV1.3 Ca2+ channels bearing several S4 charge mutations. Our modeling predicts that mutations of CaV1.1-R1 (R528H/G, R897S) or CaV1.1-R2 (R900S, R1239H) linked to hypokalemic periodic paralysis type 1 and of CaV1.3-R3 (R990H) identified in aldosterone-producing adenomas conducts ω-currents in resting state, but not during voltage-sensing domain activation. The mechanism responsible for the ω-current and its amplitude depend on the number of charges in S4, the position of the mutated S4 charge and countercharges, and the nature of the replacing amino acid. Functional characterization validates the modeling prediction showing that CaV1.3-R990H channels conduct ω-currents at hyperpolarizing potentials, but not upon membrane depolarization compared with wild-type channels.


Asunto(s)
Canales de Calcio Tipo L/metabolismo , Adenoma/genética , Adenoma/metabolismo , Animales , Canales de Calcio Tipo L/química , Canales de Calcio Tipo L/genética , Simulación por Computador , Células HEK293 , Humanos , Parálisis Periódica Hipopotasémica/genética , Parálisis Periódica Hipopotasémica/metabolismo , Potenciales de la Membrana/fisiología , Modelos Moleculares , Mutación , Dominios Proteicos , Estructura Secundaria de Proteína , Conejos , Homología Estructural de Proteína , Agua/química , Agua/metabolismo
15.
Front Pharmacol ; 8: 552, 2017.
Artículo en Inglés | MEDLINE | ID: mdl-28894419

RESUMEN

Dark chemical matter compounds are small molecules that have been recently identified as highly potent and selective hits. For this reason, they constitute a promising class of possible candidates in the process of drug discovery and raise the interest of the scientific community. To this purpose, Wassermann et al. (2015) have described the application of 2D descriptors to characterize dark chemical matter. However, their definition was based on the number of reported positive assays rather than the number of known targets. As there might be multiple assays for one single target, the number of assays does not fully describe target selectivity. Here, we propose an alternative classification of active molecules that is based on the number of known targets. We cluster molecules in four classes: black, gray, and white compounds are active on one, two to four, and more than four targets respectively, whilst inactive compounds are found to be inactive in the considered assays. In this study, black and inactive compounds are found to have not only higher solubility, but also a higher number of chiral centers, sp3 carbon atoms and aliphatic rings. On the contrary, white compounds contain a higher number of double bonds and fused aromatic rings. Therefore, the design of a screening compound library should consider these molecular properties in order to achieve target selectivity or polypharmacology. Furthermore, analysis of four main target classes (GPCRs, kinases, proteases, and ion channels) shows that GPCR ligands are more selective than the other classes, as the number of black compounds is higher in this target superfamily. On the other side, ligands that hit kinases, proteases, and ion channels bind to GPCRs more likely than to other target classes. Consequently, depending on the target protein family, appropriate screening libraries can be designed in order to minimize the likelihood of unwanted side effects early in the drug discovery process. Additionally, synergistic effects may be obtained by library design toward polypharmacology.

16.
Hum Mol Genet ; 26(18): 3531-3544, 2017 09 15.
Artículo en Inglés | MEDLINE | ID: mdl-28911204

RESUMEN

Human proteins are vulnerable towards disease-associated single amino acid replacements affecting protein stability and function. Interestingly, a few studies have shown that consensus amino acids from mammals or vertebrates can enhance protein stability when incorporated into human proteins. Here, we investigate yet unexplored relationships between the high vulnerability of human proteins towards disease-associated inactivation and recent evolutionary site-specific divergence of stabilizing amino acids. Using phylogenetic, structural and experimental analyses, we show that divergence from the consensus amino acids at several sites during mammalian evolution has caused local protein destabilization in two human proteins linked to disease: cancer-associated NQO1 and alanine:glyoxylate aminotransferase, mutated in primary hyperoxaluria type I. We demonstrate that a single consensus mutation (H80R) acts as a disease suppressor on the most common cancer-associated polymorphism in NQO1 (P187S). The H80R mutation reactivates P187S by enhancing FAD binding affinity through local and dynamic stabilization of its binding site. Furthermore, we show how a second suppressor mutation (E247Q) cooperates with H80R in protecting the P187S polymorphism towards inactivation through long-range allosteric communication within the structural ensemble of the protein. Our results support that recent divergence of consensus amino acids may have occurred with neutral effects on many functional and regulatory traits of wild-type human proteins. However, divergence at certain sites may have increased the propensity of some human proteins towards inactivation due to disease-associated mutations and polymorphisms. Consensus mutations also emerge as a potential strategy to identify structural hot-spots in proteins as targets for pharmacological rescue in loss-of-function genetic diseases.


Asunto(s)
Angiotensinógeno/genética , Proteínas/genética , Alanina/genética , Alanina Transaminasa/genética , Alanina Transaminasa/metabolismo , Aminoácidos/genética , Angiotensinógeno/metabolismo , Animales , Sitios de Unión , Secuencia de Consenso/genética , Evolución Molecular , Humanos , Mutación , NAD(P)H Deshidrogenasa (Quinona)/genética , NAD(P)H Deshidrogenasa (Quinona)/metabolismo , Filogenia , Polimorfismo Genético , Unión Proteica , Estabilidad Proteica , Proteínas/metabolismo , Transaminasas/genética , Transaminasas/metabolismo
17.
BMC Cancer ; 17(1): 235, 2017 03 31.
Artículo en Inglés | MEDLINE | ID: mdl-28359266

RESUMEN

BACKGROUND: Expression and activity of heparanase, an endoglycosidase that cleaves heparan sulfate (HS) side chains of proteoglycans, is associated with progression and poor prognosis of many cancers which makes it an attractive drug target in cancer therapeutics. METHODS: In the present work, we report the in vitro screening of a library of 150 small molecules with the scaffold bearing quinolones, oxazines, benzoxazines, isoxazoli(di)nes, pyrimidinones, quinolines, benzoxazines, and 4-thiazolidinones, thiadiazolo[3,2-a]pyrimidin-5-one, 1,2,4-triazolo-1,3,4-thiadiazoles, and azaspiranes against the enzymatic activity of human heparanase. The identified lead compounds were evaluated for their heparanase-inhibiting activity using sulfate [35S] labeled extracellular matrix (ECM) deposited by cultured endothelial cells. Further, anti-invasive efficacy of lead compound was evaluated against hepatocellular carcinoma (HepG2) and Lewis lung carcinoma (LLC) cells. RESULTS: Among the 150 compounds screened, we identified 1,2,4-triazolo-1,3,4-thiadiazoles bearing compounds to possess human heparanase inhibitory activity. Further analysis revealed 2,4-Diiodo-6-(3-phenyl-[1, 2, 4]triazolo[3,4-b][1, 3, 4]thiadiazol-6yl)phenol (DTP) as the most potent inhibitor of heparanase enzymatic activity among the tested compounds. The inhibitory efficacy was demonstrated by a colorimetric assay and further validated by measuring the release of radioactive heparan sulfate degradation fragments from [35S] labeled extracellular matrix. Additionally, lead compound significantly suppressed migration and invasion of LLC and HepG2 cells with IC50 value of ~5 µM. Furthermore, molecular docking analysis revealed a favourable interaction of triazolo-thiadiazole backbone with Asn-224 and Asp-62 of the enzyme. CONCLUSIONS: Overall, we identified biologically active heparanase inhibitor which could serve as a lead structure in developing compounds that target heparanase in cancer.


Asunto(s)
Antineoplásicos/farmacología , Inhibidores Enzimáticos/farmacología , Glucuronidasa/antagonistas & inhibidores , Neoplasias/enzimología , Neoplasias/patología , Tiadiazoles/química , Triazoles/química , Antineoplásicos/química , Apoptosis/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Inhibidores Enzimáticos/química , Humanos , Neoplasias/tratamiento farmacológico , Células Tumorales Cultivadas
18.
Sci Rep ; 7: 44532, 2017 03 14.
Artículo en Inglés | MEDLINE | ID: mdl-28291250

RESUMEN

Disease associated genetic variations often cause intracellular enzyme inactivation, dysregulation and instability. However, allosteric communication of mutational effects to distant functional sites leading to loss-of-function remains poorly understood. We characterize here interdomain site-to-site communication by which a common cancer-associated single nucleotide polymorphism (c.C609T/p.P187S) reduces the activity and stability in vivo of NAD(P)H:quinone oxidoreductase 1 (NQO1). NQO1 is a FAD-dependent, two-domain multifunctional stress protein acting as a Phase II enzyme, activating cancer pro-drugs and stabilizing p53 and p73α oncosuppressors. We show that p.P187S causes structural and dynamic changes communicated to functional sites far from the mutated site, affecting the FAD binding site located at the N-terminal domain (NTD) and accelerating proteasomal degradation through dynamic effects on the C-terminal domain (CTD). Structural protein:protein interaction studies reveal that the cancer-associated polymorphism does not abolish the interaction with p73α, indicating that oncosuppressor destabilization largely mirrors the low intracellular stability of p.P187S. In conclusion, we show how a single disease associated amino acid change may allosterically perturb several functional sites in an oligomeric and multidomain protein. These results have important implications for the understanding of loss-of-function genetic diseases and the identification of novel structural hot spots as targets for pharmacological intervention.


Asunto(s)
NAD(P)H Deshidrogenasa (Quinona)/genética , Neoplasias/genética , Conformación Proteica , Proteína Tumoral p73/genética , Regulación Alostérica/genética , Flavina-Adenina Dinucleótido/química , Flavina-Adenina Dinucleótido/genética , Predisposición Genética a la Enfermedad , Humanos , Mutación , NAD(P)H Deshidrogenasa (Quinona)/química , Neoplasias/patología , Polimorfismo de Nucleótido Simple , Complejo de la Endopetidasa Proteasomal/química , Complejo de la Endopetidasa Proteasomal/genética , Unión Proteica , Dominios Proteicos/genética , Dominios y Motivos de Interacción de Proteínas/genética , Proteína Tumoral p73/química , Proteína p53 Supresora de Tumor/química , Proteína p53 Supresora de Tumor/genética
19.
Curr Protein Pept Sci ; 18(9): 905-913, 2017.
Artículo en Inglés | MEDLINE | ID: mdl-27455965

RESUMEN

BACKGROUND: Recent advances in proteomics methodologies allow for high throughput profiling of proteolytic cleavage events. The resulting substrate peptide distributions provide deep insights in the underlying macromolecular recognition events, as determinants of biomolecular specificity identified by proteomics approaches may be compared to structure-based analysis of corresponding protein-protein interfaces. METHOD: Here, we present an overview of experimental and computational methodologies and tools applied in the area and provide an outlook beyond the protein class of proteases. RESULTS AND CONCLUSION: We discuss here future potential, synergies and needs of the emerging overlap disciplines of proteomics and structure-based modelling.


Asunto(s)
Péptidos/química , Animales , Sitios de Unión , Humanos , Modelos Moleculares , Péptido Hidrolasas/química , Péptido Hidrolasas/metabolismo , Péptidos/metabolismo , Unión Proteica , Conformación Proteica , Proteolisis , Proteoma/química , Proteoma/metabolismo , Proteómica , Especificidad por Sustrato
20.
J Chem Inf Model ; 56(6): 1228-35, 2016 06 27.
Artículo en Inglés | MEDLINE | ID: mdl-27247997

RESUMEN

Protease substrate profiling has nowadays almost become a routine task for experimentalists, and the knowledge on protease peptide substrates is easily accessible via the MEROPS database. We present a shape-based virtual screening workflow using vROCS that applies the information about the specificity of the proteases to find new small-molecule inhibitors. Peptide substrate sequences for three to four substrate positions of each substrate from the MEROPS database were used to build the training set. Two-dimensional substrate sequences were converted to three-dimensional conformations through mutation of a template peptide substrate. The vROCS query was built from single amino acid queries for each substrate position considering the relative frequencies of the amino acids. The peptide-substrate-based shape-based virtual screening approach gives good performance for the four proteases thrombin, factor Xa, factor VIIa, and caspase-3 with the DUD-E data set. The results show that the method works for protease targets with different specificity profiles as well as for targets with different active-site mechanisms. As no structure of the target and no information on small-molecule inhibitors are required to use our approach, the method has significant advantages in comparison with conventional structure- and ligand-based methods.


Asunto(s)
Bases de Datos Farmacéuticas , Evaluación Preclínica de Medicamentos/métodos , Aprendizaje Automático , Péptido Hidrolasas/metabolismo , Péptidos/metabolismo , Inhibidores de Proteasas/metabolismo , Inhibidores de Proteasas/farmacología , Ligandos , Modelos Moleculares , Péptido Hidrolasas/química , Conformación Proteica , Bibliotecas de Moléculas Pequeñas/metabolismo , Bibliotecas de Moléculas Pequeñas/farmacología , Especificidad por Sustrato , Interfaz Usuario-Computador
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA