RESUMO
Inhibition of soluble epoxide hydrolase (sEH) has recently emerged as a new approach to treat cardiovascular disease and respiratory disease. Inhibitors based on 1,3,5-triazine chemotype were discovered through affinity selection against two triazine-based DNA-encoded libraries. The structure and activity relationship study led to the expansion of the original 1,4-cycloalkyl series to related aniline, piperidine, quinoline, aryl-ether and benzylic series. The 1,3-cycloalkyl chemotype led to the discovery of a clinical candidate (GSK2256294) for COPD.
Assuntos
Cicloexilaminas/farmacologia , Epóxido Hidrolases/antagonistas & inibidores , Triazinas/farmacologia , Cicloexilaminas/química , Descoberta de Drogas , Humanos , Estrutura Molecular , Doença Pulmonar Obstrutiva Crônica/tratamento farmacológico , Bibliotecas de Moléculas Pequenas , Triazinas/químicaRESUMO
Libraries of DNA-Encoded small molecules created using combinatorial chemistry and synthetic oligonucleotides are being applied to drug discovery projects across the pharmaceutical industry. The majority of reported projects describe the discovery of reversible, i.e. non-covalent, target modulators. We synthesized multiple DNA-encoded chemical libraries terminated in electrophiles and then used them to discover covalent irreversible inhibitors and report the successful discovery of acrylamide- and epoxide-terminated Bruton's Tyrosine Kinase (BTK) inhibitors. We also demonstrate their selectivity, potency and covalent cysteine engagement using a range of techniques including X-ray crystallography, thermal transition shift assay, reporter displacement assay and intact protein complex mass spectrometry. The epoxide BTK inhibitors described here are the first ever reported to utilize this electrophile for this target.
Assuntos
Tirosina Quinase da Agamaglobulinemia/antagonistas & inibidores , DNA/química , Descoberta de Drogas , Inibidores de Proteínas Quinases/farmacologia , Bibliotecas de Moléculas Pequenas/farmacologia , Tirosina Quinase da Agamaglobulinemia/metabolismo , Cristalografia por Raios X , Relação Dose-Resposta a Droga , Humanos , Estrutura Molecular , Inibidores de Proteínas Quinases/química , Bibliotecas de Moléculas Pequenas/química , Relação Estrutura-AtividadeRESUMO
We have identified and characterized novel potent inhibitors of Bruton's tyrosine kinase (BTK) from a single DNA-encoded library of over 110 million compounds by using multiple parallel selection conditions, including variation in target concentration and addition of known binders to provide competition information. Distinct binding profiles were observed by comparing enrichments of library building block combinations under these conditions; one enriched only at high concentrations of BTK and was competitive with ATP, and another enriched at both high and low concentrations of BTK and was not competitive with ATP. A compound representing the latter profile showed low nanomolar potency in biochemical and cellular BTK assays. Results from kinetic mechanism of action studies were consistent with the selection profiles. Analysis of the co-crystal structure of the most potent compound demonstrated a novel binding mode that revealed a new pocket in BTK. Our results demonstrate that profile-based selection strategies using DNA-encoded libraries form the basis of a new methodology to rapidly identify small molecule inhibitors with novel binding modes to clinically relevant targets.
Assuntos
DNA/química , Inibidores de Proteínas Quinases/química , Proteínas Tirosina Quinases/antagonistas & inibidores , Bibliotecas de Moléculas Pequenas/química , Tirosina Quinase da Agamaglobulinemia , Sítios de Ligação , Linhagem Celular , Citocromo P-450 CYP3A/química , Citocromo P-450 CYP3A/metabolismo , DNA/metabolismo , Humanos , Cinética , Simulação de Acoplamento Molecular , Ligação Proteica , Inibidores de Proteínas Quinases/metabolismo , Estrutura Terciária de Proteína , Proteínas Tirosina Quinases/genética , Proteínas Tirosina Quinases/metabolismo , Bibliotecas de Moléculas Pequenas/metabolismoRESUMO
DNA-encoded chemical library (DEL) technology provides a time- and cost-efficient method to simultaneously screen billions of compounds for their affinity to a protein target of interest. Here we report its use to identify a novel chemical series of inhibitors of the thioesterase activity of polyketide synthase 13 (Pks13) from Mycobacterium tuberculosis (Mtb). We present three chemically distinct series of inhibitors along with their enzymatic and Mtb whole cell potency, the measure of on-target activity in cells, and the crystal structures of inhibitor-enzyme complexes illuminating their interactions with the active site of the enzyme. One of these inhibitors showed a favorable pharmacokinetic profile and demonstrated efficacy in an acute mouse model of tuberculosis (TB) infection. These findings and assay developments will aid in the advancement of TB drug discovery.
Assuntos
Antituberculosos , Inibidores Enzimáticos , Mycobacterium tuberculosis , Policetídeo Sintases , Bibliotecas de Moléculas Pequenas , Tioléster Hidrolases , Animais , Humanos , Camundongos , Antituberculosos/química , Antituberculosos/farmacologia , Antituberculosos/uso terapêutico , Proteínas de Bactérias/antagonistas & inibidores , Proteínas de Bactérias/química , Cristalografia por Raios X , Modelos Animais de Doenças , Descoberta de Drogas , Avaliação Pré-Clínica de Medicamentos , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/química , Mycobacterium tuberculosis/enzimologia , Mycobacterium tuberculosis/efeitos dos fármacos , Policetídeo Sintases/metabolismo , Policetídeo Sintases/química , Policetídeo Sintases/genética , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Tioléster Hidrolases/antagonistas & inibidores , Tioléster Hidrolases/metabolismo , Tioléster Hidrolases/química , Tioléster Hidrolases/genética , Tuberculose/tratamento farmacológico , Tuberculose/microbiologiaRESUMO
Evasion of apoptosis is critical for the development and growth of tumors. The pro-survival protein myeloid cell leukemia 1 (Mcl-1) is an antiapoptotic member of the Bcl-2 family, associated with tumor aggressiveness, poor survival, and drug resistance. Development of Mcl-1 inhibitors implies blocking of protein-protein interactions, generally requiring a lengthy optimization process of large, complex molecules. Herein, we describe the use of DNA-encoded chemical library synthesis and screening to directly generate complex, yet conformationally privileged macrocyclic hits that serve as Mcl-1 inhibitors. By applying a conceptual combination of conformational analysis and structure-based design in combination with a robust synthetic platform allowing rapid analoging, we optimized in vitro potency of a lead series into the low nanomolar regime. Additionally, we demonstrate fine-tuning of the physicochemical properties of the macrocyclic compounds, resulting in the identification of lead candidates 57/59 with a balanced profile, which are suitable for future development toward therapeutic use.
Assuntos
Antineoplásicos , Neoplasias , Humanos , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Apoptose , Conformação Molecular , DNA , Linhagem Celular Tumoral , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Antineoplásicos/farmacologia , Antineoplásicos/químicaRESUMO
Bfl-1 is overexpressed in both hematological and solid tumors; therefore, inhibitors of Bfl-1 are highly desirable. A DNA-encoded chemical library (DEL) screen against Bfl-1 identified the first known reversible covalent small-molecule ligand for Bfl-1. The binding was validated through biophysical and biochemical techniques, which confirmed the reversible covalent mechanism of action and pointed to binding through Cys55. This represented the first identification of a cyano-acrylamide reversible covalent compound from a DEL screen and highlights further opportunities for covalent drug discovery through DEL screening. A 10-fold improvement in potency was achieved through a systematic SAR exploration of the hit. The more potent analogue compound 13 was successfully cocrystallized in Bfl-1, revealing the binding mode and providing further evidence of a covalent interaction with Cys55.
RESUMO
Dysregulation of IL17A drives numerous inflammatory and autoimmune disorders with inhibition of IL17A using antibodies proven as an effective treatment. Oral anti-IL17 therapies are an attractive alternative option, and several preclinical small molecule IL17 inhibitors have previously been described. Herein, we report the discovery of a novel class of small molecule IL17A inhibitors, identified via a DNA-encoded chemical library screen, and their subsequent optimization to provide in vivo efficacious inhibitors. These new protein-protein interaction (PPI) inhibitors bind in a previously undescribed mode in the IL17A protein with two copies binding symmetrically to the central cavities of the IL17A homodimer.
Assuntos
DNA , Descoberta de Drogas , Interleucina-17 , Bibliotecas de Moléculas Pequenas , Interleucina-17/metabolismo , Interleucina-17/antagonistas & inibidores , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , DNA/metabolismo , DNA/química , Humanos , Animais , Relação Estrutura-Atividade , Ligação Proteica , CamundongosRESUMO
The natural product fumagillin exhibits potent antiproliferative and antiangiogenic properties. The semisynthetic analog PPI-2458, [(3R,4S,5S,6R)-5-methoxy-4-[(2R,3R)-2-methyl-3-(3-methylbut-2-enyl)oxiran-2-yl]-1-oxaspiro[2.5]octan-6-yl] N-[(2R)-1-amino-3-methyl-1-oxobutan-2-yl]carbamate, demonstrates rapid inactivation of its molecular target, methionine aminopeptidase-2 (MetAP2), and good efficacy in several rodent models of cancer and inflammation with oral dosing despite low apparent oral bioavailability. To probe the basis of its in vivo efficacy, the metabolism of PPI-2458 was studied in detail. Reaction phenotyping identified CYP3A4/5 as the major source of metabolism in humans. Six metabolites were isolated from liver microsomes and characterized by mass spectrometry and nuclear resonance spectroscopy, and their structures were confirmed by chemical synthesis. The synthetic metabolites showed correlated inhibition of MetAP2 enzymatic activity and vascular endothelial cell growth. In an ex vivo experiment, MetAP2 inhibition in white blood cells, thymus, and lymph nodes in rats after single dosing with PPI-2458 and the isolated metabolites was found to correlate with the in vitro activity of the individual species. In a phase 1 clinical study, PPI-2458 was administered to patients with non-Hodgkin lymphoma. At 15 mg administered orally every other day, MetAP2 in whole blood was 80% inactivated for up to 48 hours, although the exposure of the parent compound was only â¼10% that of the summed cytochrome P450 metabolites. Taken together, the data confirm the participation of active metabolites in the in vivo efficacy of PPI-2458. The structures define a metabolic pathway for PPI-2458 that is distinct from that of TNP-470 ([(3R,4S,5S,6R)-5-methoxy-4-[(2R,3R)-2-methyl-3-(3-methylbut-2-enyl)oxiran-2-yl]-1-oxaspiro[2.5]octan-6-yl] N-(2-chloroacetyl)carbamate). The high level of MetAP2 inhibition achieved in vivo supports the value of fumagillin-derived therapeutics for angiogenic diseases.
Assuntos
Aminopeptidases/antagonistas & inibidores , Inibidores da Angiogênese/farmacologia , Inibidores da Angiogênese/farmacocinética , Compostos de Epóxi/farmacocinética , Metaloendopeptidases/antagonistas & inibidores , Valina/análogos & derivados , Aminopeptidases/sangue , Animais , Esquema de Medicação , Compostos de Epóxi/administração & dosagem , Compostos de Epóxi/análise , Compostos de Epóxi/farmacologia , Humanos , Leucócitos/efeitos dos fármacos , Leucócitos/metabolismo , Linfonodos/efeitos dos fármacos , Linfonodos/metabolismo , Linfoma não Hodgkin/sangue , Linfoma não Hodgkin/tratamento farmacológico , Metaloendopeptidases/sangue , Microssomos Hepáticos/química , Microssomos Hepáticos/metabolismo , Ratos , Relação Estrutura-Atividade , Timo/efeitos dos fármacos , Timo/metabolismo , Valina/administração & dosagem , Valina/análise , Valina/farmacocinética , Valina/farmacologiaRESUMO
WD40 repeat-containing protein 91 (WDR91) regulates early-to-late endosome conversion and plays vital roles in endosome fusion, recycling, and transport. WDR91 was recently identified as a potential host factor for viral infection. We employed DNA-encoded chemical library (DEL) selection against the WDR domain of WDR91, followed by machine learning to predict ligands from the synthetically accessible Enamine REAL database. Screening of predicted compounds identified a WDR91 selective compound 1, with a KD of 6 ± 2 µM by surface plasmon resonance. The co-crystal structure confirmed the binding of 1 to the WDR91 side pocket, in proximity to cysteine 487, which led to the discovery of covalent analogues 18 and 19. The covalent adduct formation for 18 and 19 was confirmed by intact mass liquid chromatography-mass spectrometry. The discovery of 1, 18, and 19, accompanying structure-activity relationship, and the co-crystal structures provide valuable insights for designing potent and selective chemical tools against WDR91 to evaluate its therapeutic potential.
Assuntos
DNA , Bibliotecas de Moléculas Pequenas , DNA/química , Biblioteca Gênica , Ligantes , Aprendizado de Máquina , Bibliotecas de Moléculas Pequenas/farmacologia , Bibliotecas de Moléculas Pequenas/químicaRESUMO
Biochemical combinatorial techniques such as phage display, RNA display and oligonucleotide aptamers have proven to be reliable methods for generation of ligands to protein targets. Adapting these techniques to small synthetic molecules has been a long-sought goal. We report the synthesis and interrogation of an 800-million-member DNA-encoded library in which small molecules are covalently attached to an encoding oligonucleotide. The library was assembled by a combination of chemical and enzymatic synthesis, and interrogated by affinity selection. We describe methods for the selection and deconvolution of the chemical display library, and the discovery of inhibitors for two enzymes: Aurora A kinase and p38 MAP kinase.
Assuntos
DNA/química , Desenho de Fármacos , Inibidores de Proteínas Quinases/síntese química , Bibliotecas de Moléculas Pequenas/síntese química , Animais , Aurora Quinases , Técnicas de Química Combinatória , DNA/genética , Modelos Moleculares , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/farmacologia , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Bibliotecas de Moléculas Pequenas/química , Bibliotecas de Moléculas Pequenas/farmacologia , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidoresRESUMO
This study describes a novel series of UDP-N-acetylglucosamine acyltransferase (LpxA) inhibitors that was identified through affinity-mediated selection from a DNA-encoded compound library. The original hit was a selective inhibitor of Pseudomonas aeruginosa LpxA with no activity against Escherichia coli LpxA. The biochemical potency of the series was optimized through an X-ray crystallography-supported medicinal chemistry program, resulting in compounds with nanomolar activity against P. aeruginosa LpxA (best half-maximal inhibitory concentration (IC50) <5 nM) and cellular activity against P. aeruginosa (best minimal inhibitory concentration (MIC) of 4 µg/mL). Lack of activity against E. coli was maintained (IC50 > 20 µM and MIC > 128 µg/mL). The mode of action of analogues was confirmed through genetic analyses. As expected, compounds were active against multidrug-resistant isolates. Further optimization of pharmacokinetics is needed before efficacy studies in mouse infection models can be attempted. To our knowledge, this is the first reported LpxA inhibitor series with selective activity against P. aeruginosa.
Assuntos
Aciltransferases/antagonistas & inibidores , Antibacterianos/farmacologia , Descoberta de Drogas , Inibidores Enzimáticos/farmacologia , Pseudomonas aeruginosa/efeitos dos fármacos , Antibacterianos/química , Cristalografia por Raios X , Farmacorresistência Bacteriana/efeitos dos fármacos , Inibidores Enzimáticos/química , Escherichia coli/enzimologia , Testes de Sensibilidade Microbiana , Estrutura Molecular , Relação Estrutura-AtividadeRESUMO
Fumagillin, an irreversible inhibitor of MetAP2, has been shown to potently inhibit growth of malaria parasites in vitro. Here, we demonstrate activity of fumagillin analogs with an improved pharmacokinetic profile against malaria parasites, trypanosomes, and amoebas. A subset of the compounds showed efficacy in a murine malaria model. The observed SAR forms a basis for further optimization of fumagillin based inhibitors against parasitic targets by inhibition of MetAP2.
Assuntos
Aminopeptidases/antagonistas & inibidores , Antimaláricos/química , Cicloexanos/química , Ácidos Graxos Insaturados/química , Metaloendopeptidases/antagonistas & inibidores , Administração Oral , Aminopeptidases/metabolismo , Animais , Antimaláricos/síntese química , Antimaláricos/farmacologia , Cicloexanos/síntese química , Cicloexanos/farmacologia , Ácidos Graxos Insaturados/síntese química , Ácidos Graxos Insaturados/farmacologia , Metaloendopeptidases/metabolismo , Camundongos , Testes de Sensibilidade Parasitária , Sesquiterpenos/síntese química , Sesquiterpenos/química , Sesquiterpenos/farmacologiaRESUMO
Encoded library technology (ELT) is an effective approach to the discovery of novel small-molecule ligands for biological targets. A key factor for the success of the technology is the chemical diversity of the libraries. Here we report the development of DNA-conjugated benzimidazoles. Using 4-fluoro-3-nitrobenzoic acid as a key synthon, we synthesized a 320 million-member DNA-encoded benzimidazole library using Fmoc-protected amino acids, amines and aldehydes as diversity elements. Affinity selection of the library led to the discovery of a novel, potent and specific antagonist of the NK3 receptor.
Assuntos
Benzimidazóis/síntese química , DNA/química , Bibliotecas de Moléculas Pequenas/síntese química , Aminoácidos/química , Benzimidazóis/química , Concentração de Íons de Hidrogênio , Nitrobenzoatos/química , Bibliotecas de Moléculas Pequenas/químicaRESUMO
This corrects the article DOI: 10.1038/ncomms16081.
RESUMO
Mcl-1 is a pro-apoptotic BH3 protein family member similar to Bcl-2 and Bcl-xL. Overexpression of Mcl-1 is often seen in various tumors and allows cancer cells to evade apoptosis. Here we report the discovery and optimization of a series of non-natural peptide Mcl-1 inhibitors. Screening of DNA-encoded libraries resulted in hit compound 1, a 1.5 µM Mcl-1 inhibitor. A subsequent crystal structure demonstrated that compound 1 bound to Mcl-1 in a ß-turn conformation, such that the two ends of the peptide were close together. This proximity allowed for the linking of the two ends of the peptide to form a macrocycle. Macrocyclization resulted in an approximately 10-fold improvement in binding potency. Further exploration of a key hydrophobic interaction with Mcl-1 protein and also with the moiety that engages Arg256 led to additional potency improvements. The use of protein-ligand crystal structures and binding kinetics contributed to the design and understanding of the potency gains. Optimized compound 26 is a <3 nM Mcl-1 inhibitor, while inhibiting Bcl-2 at only 5 µM and Bcl-xL at >99 µM, and induces cleaved caspase-3 in MV4-11 cells with an IC50 of 3 µM after 6 h.
RESUMO
ATAD2 (ANCCA) is an epigenetic regulator and transcriptional cofactor, whose overexpression has been linked to the progress of various cancer types. Here, we report a DNA-encoded library screen leading to the discovery of BAY-850, a potent and isoform selective inhibitor that specifically induces ATAD2 bromodomain dimerization and prevents interactions with acetylated histones in vitro, as well as with chromatin in cells. These features qualify BAY-850 as a chemical probe to explore ATAD2 biology.
Assuntos
ATPases Associadas a Diversas Atividades Celulares/antagonistas & inibidores , ATPases Associadas a Diversas Atividades Celulares/metabolismo , Proteínas de Ligação a DNA/antagonistas & inibidores , Proteínas de Ligação a DNA/metabolismo , Sondas Moleculares/química , Sondas Moleculares/farmacologia , Mapas de Interação de Proteínas/efeitos dos fármacos , Multimerização Proteica/efeitos dos fármacos , ATPases Associadas a Diversas Atividades Celulares/química , Linhagem Celular Tumoral , Cromatina/metabolismo , Proteínas de Ligação a DNA/química , Descoberta de Drogas , Histonas/metabolismo , Humanos , Ligantes , Modelos Moleculares , Isoformas de Proteínas/antagonistas & inibidores , Isoformas de Proteínas/química , Isoformas de Proteínas/metabolismoRESUMO
[This corrects the article DOI: 10.1021/acsmedchemlett.6b00464.].
RESUMO
The identification and prioritization of chemically tractable therapeutic targets is a significant challenge in the discovery of new medicines. We have developed a novel method that rapidly screens multiple proteins in parallel using DNA-encoded library technology (ELT). Initial efforts were focused on the efficient discovery of antibacterial leads against 119 targets from Acinetobacter baumannii and Staphylococcus aureus. The success of this effort led to the hypothesis that the relative number of ELT binders alone could be used to assess the ligandability of large sets of proteins. This concept was further explored by screening 42 targets from Mycobacterium tuberculosis. Active chemical series for six targets from our initial effort as well as three chemotypes for DHFR from M. tuberculosis are reported. The findings demonstrate that parallel ELT selections can be used to assess ligandability and highlight opportunities for successful lead and tool discovery.
Assuntos
Acinetobacter baumannii/efeitos dos fármacos , Antibacterianos/farmacologia , Descoberta de Drogas/métodos , Biblioteca Gênica , Mycobacterium tuberculosis/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas , Staphylococcus aureus/efeitos dos fármacos , Acinetobacter baumannii/metabolismo , Avaliação Pré-Clínica de Medicamentos , Terapia de Alvo Molecular , Mycobacterium tuberculosis/metabolismo , Staphylococcus aureus/metabolismoRESUMO
DNA-encoded library technology (ELT) is a powerful tool for the discovery of new small-molecule ligands to various protein targets. Here we report the design and synthesis of biaryl DNA-encoded libraries based on the scaffold of 5-formyl 3-iodobenzoic acid. Three reactions on DNA template, acylation, Suzuki-Miyaura coupling and reductive amination, were applied in the library synthesis. The three cycle library of 3.5 million diversity has delivered potent hits for phosphoinositide 3-kinase α (PI3Kα).