RESUMO
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
Millions of individuals are infected with and die from tuberculosis (TB) each year, and multidrug-resistant (MDR) strains of TB are increasingly prevalent. As such, there is an urgent need to identify novel drugs to treat TB infections. Current frontline therapies include the drug isoniazid, which inhibits the essential NADH-dependent enoyl-acyl-carrier protein (ACP) reductase, InhA. To inhibit InhA, isoniazid must be activated by the catalase-peroxidase KatG. Isoniazid resistance is linked primarily to mutations in the katG gene. Discovery of InhA inhibitors that do not require KatG activation is crucial to combat MDR TB. Multiple discovery efforts have been made against InhA in recent years. Until recently, despite achieving high potency against the enzyme, these efforts have been thwarted by lack of cellular activity. We describe here the use of DNA-encoded X-Chem (DEX) screening, combined with selection of appropriate physical properties, to identify multiple classes of InhA inhibitors with cell-based activity. The utilization of DEX screening allowed the interrogation of very large compound libraries (1011 unique small molecules) against multiple forms of the InhA enzyme in a multiplexed format. Comparison of the enriched library members across various screening conditions allowed the identification of cofactor-specific inhibitors of InhA that do not require activation by KatG, many of which had bactericidal activity in cell-based assays.
Assuntos
Proteínas de Bactérias/antagonistas & inibidores , Mycobacterium tuberculosis/enzimologia , Oxirredutases/antagonistas & inibidores , Testes de Sensibilidade Microbiana , Bibliotecas de Moléculas PequenasRESUMO
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
The inhibition of protein-protein interactions remains a challenge for traditional small molecule drug discovery. Here we describe the use of DNA-encoded library technology for the discovery of small molecules that are potent inhibitors of the interaction between lymphocyte function-associated antigen 1 and its ligand intercellular adhesion molecule 1. A DNA-encoded library with a potential complexity of 4.1 billion compounds was exposed to the I-domain of the target protein and the bound ligands were affinity selected, yielding an enriched small-molecule hit family. Compounds representing this family were synthesized without their DNA encoding moiety and found to inhibit the lymphocyte function-associated antigen 1/intercellular adhesion molecule-1 interaction with submicromolar potency in both ELISA and cell adhesion assays. Re-synthesized compounds conjugated to DNA or a fluorophore were demonstrated to bind to cells expressing the target protein.
Assuntos
Descoberta de Drogas , Antígeno-1 Associado à Função Linfocitária/metabolismo , Bibliotecas de Moléculas Pequenas/farmacologia , Adesão Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Humanos , Células Jurkat , Ligantes , Estrutura Molecular , Ligação Proteica/efeitos dos fármacos , Bibliotecas de Moléculas Pequenas/síntese química , Bibliotecas de Moléculas Pequenas/química , Relação Estrutura-AtividadeRESUMO
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
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
DCAF1 is a substrate receptor of two distinct E3 ligases (CRL4DCAF1 and EDVP), plays a critical physiological role in protein degradation, and is considered a drug target for various cancers. Antagonists of DCAF1 could be used toward the development of therapeutics for cancers and viral treatments. We used the WDR domain of DCAF1 to screen a 114-billion-compound DNA encoded library (DEL) and identified candidate compounds using similarity search and machine learning. This led to the discovery of a compound (Z1391232269) with an SPR KD of 11 µM. Structure-guided hit optimization led to the discovery of OICR-8268 (26e) with an SPR KD of 38 nM and cellular target engagement with EC50 of 10 µM as measured by cellular thermal shift assay (CETSA). OICR-8268 is an excellent tool compound to enable the development of next-generation DCAF1 ligands toward cancer therapeutics, further investigation of DCAF1 functions in cells, and the development of DCAF1-based PROTACs.
Assuntos
Neoplasias , Ubiquitina-Proteína Ligases , Humanos , Ligantes , Ubiquitina-Proteína Ligases/metabolismo , Proteínas de Transporte/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
Herein we report the discovery of 2,4-1H-imidazole carboxamides as novel, biochemically potent, and kinome selective inhibitors of transforming growth factor ß-activated kinase 1 (TAK1). The target was subjected to a DNA-encoded chemical library (DECL) screen. After hit analysis a cluster of compounds was identified, which was based on a central pyrrole-2,4-1H-dicarboxamide scaffold, showing remarkable kinome selectivity. A scaffold-hop to the corresponding imidazole resulted in increased biochemical potency. Next, X-ray crystallography revealed a distinct binding mode compared to other TAK1 inhibitors. A benzylamide was found in a perpendicular orientation with respect to the core hinge-binding imidazole. Additionally, an unusual amide flip was observed in the kinase hinge region. Using structure-based drug design (SBDD), key substitutions at the pyrrolidine amide and the glycine resulted in a significant increase in biochemical potency.
RESUMO
Inhibition of hydroxy acid oxidase 1 (HAO1) is a strategy to mitigate the accumulation of toxic oxalate that results from reduced activity of alanine-glyoxylate aminotransferase (AGXT) in primary hyperoxaluria 1 (PH1) patients. DNA-Encoded Chemical Library (DECL) screening provided two novel chemical series of potent HAO1 inhibitors, represented by compounds 3-6. Compound 5 was further optimized via various structure-activity relationship (SAR) exploration methods to 29, a compound with improved potency and absorption, distribution, metabolism, and excretion (ADME)/pharmacokinetic (PK) properties. Since carboxylic acid-containing compounds are often poorly permeable and have potential active glucuronide metabolites, we undertook a brief, initial exploration of acid replacements with the aim of identifying non-acid-containing HAO1 inhibitors. Structure-based drug design initiated with Compound 5 led to the identification of a nonacid inhibitor of HAO1, 31, which has weaker potency and increased permeability.
Assuntos
Oxirredutases do Álcool/antagonistas & inibidores , DNA/química , Bibliotecas de Moléculas Pequenas/química , Oxirredutases do Álcool/metabolismo , Animais , Sítios de Ligação , Cristalografia por Raios X , DNA/metabolismo , Desenho de Fármacos , Meia-Vida , Humanos , Hiperoxalúria Primária/metabolismo , Hiperoxalúria Primária/patologia , Indóis/química , Indóis/metabolismo , Masculino , Camundongos , Simulação de Acoplamento Molecular , Bibliotecas de Moléculas Pequenas/metabolismo , Relação Estrutura-Atividade , Tiazóis/química , Tiazóis/metabolismo , Transaminases/genética , Transaminases/metabolismoRESUMO
Bispecific degraders (PROTACs) of ERα are expected to be advantageous over current inhibitors of ERα signaling (aromatase inhibitors/SERMs/SERDs) used to treat ER+ breast cancer. Information from DNA-encoded chemical library (DECL) screening provides a method to identify novel PROTAC binding features as the linker positioning, and binding elements are determined directly from the screen. After screening â¼120 billion DNA-encoded molecules with ERα WT and 3 gain-of-function (GOF) mutants, with and without estradiol to identify features that enrich ERα competitively, the off-DNA synthesized small molecule exemplar 7 exhibited nanomolar ERα binding, antagonism, and degradation. Click chemistry synthesis on an alkyne E3 ligase engagers panel and an azide variant of 7 rapidly generated bispecific nanomolar degraders of ERα, with PROTACs 18 and 21 inhibiting ER+ MCF7 tumor growth in a mouse xenograft model of breast cancer. This study validates this approach toward identifying novel bispecific degrader leads from DECL screening with minimal optimization.
Assuntos
DNA/química , Receptor alfa de Estrogênio/metabolismo , Bibliotecas de Moléculas Pequenas/química , Animais , Neoplasias da Mama/tratamento farmacológico , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Química Click , DNA/metabolismo , Antagonistas de Estrogênios/química , Antagonistas de Estrogênios/metabolismo , Antagonistas de Estrogênios/farmacologia , Antagonistas de Estrogênios/uso terapêutico , Receptor alfa de Estrogênio/química , Receptor alfa de Estrogênio/genética , Feminino , Meia-Vida , Humanos , Indóis/química , Indóis/metabolismo , Cinética , Camundongos , Bibliotecas de Moléculas Pequenas/metabolismo , Bibliotecas de Moléculas Pequenas/farmacologia , Bibliotecas de Moléculas Pequenas/uso terapêutico , Relação Estrutura-Atividade , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
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
The activity of the secreted phosphodiesterase autotaxin produces the inflammatory signaling molecule LPA and has been associated with a number of human diseases including idiopathic pulmonary fibrosis (IPF). We screened a single DNA-encoded chemical library (DECL) of 225 million compounds and identified a series of potent inhibitors. Optimization of this series led to the discovery of compound 1 (X-165), a highly potent, selective, and bioavailable small molecule. Cocrystallization of compound 1 with human autotaxin demonstrated that it has a novel binding mode occupying both the hydrophobic pocket and a channel near the autotaxin active site. Compound 1 inhibited the production of LPA in human and mouse plasma at nanomolar levels and showed efficacy in a mouse model of human lung fibrosis. After successfully completing IND-enabling studies, compound 1 was approved by the FDA for a Phase I clinical trial. These results demonstrate that DECL hits can be readily optimized into clinical candidates.
Assuntos
Hidantoínas/uso terapêutico , Fibrose Pulmonar Idiopática/tratamento farmacológico , Inibidores de Fosfodiesterase/uso terapêutico , Diester Fosfórico Hidrolases/metabolismo , Piperidinas/uso terapêutico , Compostos de Espiro/uso terapêutico , Animais , Bleomicina , Cristalografia por Raios X , DNA/química , Cães , Humanos , Hidantoínas/síntese química , Hidantoínas/metabolismo , Fibrose Pulmonar Idiopática/induzido quimicamente , Fibrose Pulmonar Idiopática/patologia , Pulmão/patologia , Masculino , Camundongos Endogâmicos C57BL , Inibidores de Fosfodiesterase/síntese química , Inibidores de Fosfodiesterase/metabolismo , Piperidinas/síntese química , Piperidinas/metabolismo , Ligação Proteica , Ratos , Compostos de Espiro/síntese química , Compostos de Espiro/metabolismoRESUMO
DNA-encoded small molecule libraries (DELs) have enabled discovery of novel inhibitors for many distinct protein targets of therapeutic value. We demonstrate a new approach applying machine learning to DEL selection data by identifying active molecules from large libraries of commercial and easily synthesizable compounds. We train models using only DEL selection data and apply automated or automatable filters to the predictions. We perform a large prospective study (â¼2000 compounds) across three diverse protein targets: sEH (a hydrolase), ERα (a nuclear receptor), and c-KIT (a kinase). The approach is effective, with an overall hit rate of â¼30% at 30 µM and discovery of potent compounds (IC50 < 10 nM) for every target. The system makes useful predictions even for molecules dissimilar to the original DEL, and the compounds identified are diverse, predominantly drug-like, and different from known ligands. This work demonstrates a powerful new approach to hit-finding.
Assuntos
DNA/química , Descoberta de Drogas/métodos , Redes Neurais de Computação , Bibliotecas de Moléculas Pequenas/química , Epóxido Hidrolases/antagonistas & inibidores , Receptor alfa de Estrogênio/antagonistas & inibidores , Ligantes , Inibidores de Proteínas Quinases/química , Proteínas Proto-Oncogênicas c-kit/antagonistas & inibidoresRESUMO
The discovery of ligands via affinity-mediated selection of DNA-encoded chemical libraries is driven by the quality and concentration of the protein target. G-protein-coupled receptors (GPCRs) and other membrane-bound targets can be difficult to isolate in their functional state and at high concentrations, and therefore have been challenging for affinity-mediated selection. Here, we report a successful selection campaign against protease-activated receptor 2 (PAR2). Using a thermo-stabilized mutant of PAR2, we conducted affinity selection using our >100-billion-compound DNA-encoded library. We observed a number of putative ligands enriched upon selection, and subsequent cellular profiling revealed these ligands to comprise both agonists and antagonists. The agonist series shared structural similarity with known agonists. The antagonists were shown to bind in a novel allosteric binding site on the PAR2 protein. This report serves to demonstrate that cell-free affinity selection against GPCRs can be achieved with mutant stabilized protein targets.
Assuntos
DNA/genética , Mutação/efeitos dos fármacos , Receptores Acoplados a Proteínas G/agonistas , Receptores Acoplados a Proteínas G/antagonistas & inibidores , Bibliotecas de Moléculas Pequenas/farmacologia , Sítio Alostérico/efeitos dos fármacos , Linhagem Celular , Células HEK293 , Humanos , Ligantes , Proteínas/genética , Receptor PAR-2 , Receptores Acoplados a Proteínas G/genéticaRESUMO
Tumor progression loci-2 (Tpl2) (Cot/MAP3K8) is a serine/threonine kinase in the MAP3K family directly upstream of MEK. Recent studies using Tpl2 knockout mice have indicated an important role for Tpl2 in the lipopolysaccharide (LPS) induced production of tumor necrosis factor alpha (TNF-alpha) and other proinflammatory cytokines involved in diseases such as rheumatoid arthritis. Initial 4-anilino-6-aminoquinoline-3-carbonitrile leads showed poor selectivity for Tpl2 over epidermal growth factor receptor (EGFR) kinase. Using molecular modeling and crystallographic data of the EGFR kinase domain with and without an EGFR kinase-specific 4-anilinoquinazoline inhibitor (erlotinib, Tarceva), we hypothesized that we could diminish the inhibition of EGFR kinase by substitution at the C-8 position of our 4-anilino-6-aminoquinoline-3-carbonitrile leads. The 8-substituted-4-anilino-6-aminoquinoline-3-carbonitriles were prepared from the appropriate 2-substituted 4-nitroanilines. Modifications to the C-6 and C-8 positions led to the identification of compounds with increased inhibition of TNF-alpha release from LPS-stimulated rat and human blood, and these analogues were also highly selective for Tpl2 kinase over EGFR kinase. Further structure-activity based modifications led to the identification of 8-bromo-4-(3-chloro-4-fluorophenylamino)-6-[(1-methyl-1H-imidazol-4-yl)methylamino]quinoline-3-carbonitrile, which demonstrated in vitro as well as in vivo efficacy in inhibition of LPS-induced TNF-alpha production.
Assuntos
Aminoquinolinas/síntese química , Anti-Inflamatórios não Esteroides/síntese química , Imidazóis/síntese química , MAP Quinase Quinase Quinases/antagonistas & inibidores , Modelos Moleculares , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Aminoquinolinas/farmacocinética , Aminoquinolinas/farmacologia , Animais , Anti-Inflamatórios não Esteroides/farmacocinética , Anti-Inflamatórios não Esteroides/farmacologia , Cristalografia por Raios X , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/química , Cloridrato de Erlotinib , Feminino , Humanos , Imidazóis/farmacocinética , Imidazóis/farmacologia , Técnicas In Vitro , MAP Quinase Quinase Quinases/biossíntese , MAP Quinase Quinase Quinases/química , Microssomos Hepáticos/metabolismo , Estrutura Terciária de Proteína , Proteínas Proto-Oncogênicas/biossíntese , Proteínas Proto-Oncogênicas/química , Quinazolinas/química , Ratos , Ratos Sprague-Dawley , Relação Estrutura-Atividade , Fator de Necrose Tumoral alfa/biossíntese , Fator de Necrose Tumoral alfa/químicaRESUMO
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.