RESUMO
Heterobifunctional PROTAC degraders are gaining attention as a differentiated therapeutic modality with the potential for oral dosing in the clinic. Belonging to the beyond Rule of Five domain of physicochemical property space, we have sought to understand the determinants of oral absorption for this class of molecules for the rapid development of novel oral agents. We have collected a large data set from PROTAC molecules that have been dosed orally and intravenously in rats to estimate the fraction absorbed from oral dosing. Through this estimation, effects from differential hepatic clearance are normalized, allowing for a better assessment of the absorption. We demonstrate that rats are less permissive to PROTAC absorption than mice. The physicochemical properties of the molecules are then evaluated once compounds are rank-ordered by the fraction absorbed. We derive suggested design constraints on physicochemical properties for PROTAC molecules that are associated with higher probability of being orally absorbed.
Assuntos
Fígado , Ratos , Camundongos , Animais , ProteóliseRESUMO
Heterobifunctional protein degraders, such as PROteolysis TArgeting Chimera (PROTAC) protein degraders, constitute a novel therapeutic modality that harnesses the cell's natural protein-degradation machinery - that is, the ubiquitin-proteasome system - to selectively target proteins involved in disease pathogenesis for elimination. Protein degraders have several potential advantages over small-molecule inhibitors that have traditionally been used for cancer treatment, including their event-driven (rather than occupancy-driven) pharmacology, which permits sub-stoichiometric drug concentrations for activity, their capacity to act iteratively and target multiple copies of a protein of interest, and their potential to target nonenzymatic proteins that were previously considered 'undruggable'. Following numerous innovations in protein degrader design and rigorous evaluation in preclinical models, protein degraders entered clinical testing in 2019. Currently, 18 protein degraders are in phase I or phase I/II clinical trials that involve patients with various tumour types, with a phase III trial of one initiated in 2022. The first safety, efficacy and pharmacokinetic data from these studies are now materializing and, although considerably more evidence is needed, protein degraders are showing promising activity as cancer therapies. Herein, we review advances in protein degrader development, the preclinical research that supported their entry into clinical studies, the available data for protein degraders in patients and future directions for this new class of drugs.
Assuntos
Neoplasias , Humanos , Neoplasias/tratamento farmacológico , Proteínas/metabolismo , ProteóliseRESUMO
6,7-Dihydro-5H-2,1-benzisoxazol-4-one analogs are potent inhibitors of aldosterone synthase (CYP11B2) with selectivity over the highly homologous enzyme cortisol synthase (CYP11B1). These compounds are unique among inhibitors of CYP11B2 in their lack of a strong-heme binding group such as a pyridine or imidazole. Poor metabolic stability in hepatocyte incubations was found to proceed via a reduction of the isoxazole ring. While the enzyme responsible for the reductive metabolism remains unknown, the rate of metabolism could be attenuated by the addition of polar functionality. The in vitro CYP11B2 potency and selectivity were confirmed in vivo in a cynomolgus monkey model by the inhibition of ACTH stimulated aldosterone production without impacting plasma cortisol concentrations.
Assuntos
Citocromo P-450 CYP11B2/antagonistas & inibidores , Inibidores Enzimáticos/farmacologia , Isoxazóis/farmacologia , Citocromo P-450 CYP11B2/metabolismo , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Humanos , Isoxazóis/síntese química , Isoxazóis/química , Estrutura Molecular , Relação Estrutura-AtividadeRESUMO
The kinase selectivity and pharmacokinetic optimization of a series of 7-aminofuro[2,3-c]pyridine inhibitors of TAK1 is described. The intersection of insights from molecular modeling, computational prediction of metabolic sites, and in vitro metabolite identification studies resulted in a simple and unique solution to both of these problems. These efforts culminated in the discovery of compound 13a, a potent, relatively selective inhibitor of TAK1 with good pharmacokinetic properties in mice, which was active in an in vivo model of ovarian cancer.
Assuntos
Inibidores Enzimáticos , MAP Quinase Quinase Quinases/antagonistas & inibidores , Piridinas , Aminas/síntese química , Aminas/química , Aminas/farmacologia , Animais , Cristalografia por Raios X , Ativação Enzimática/efeitos dos fármacos , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/farmacocinética , Inibidores Enzimáticos/farmacologia , Furanos/síntese química , Furanos/química , Furanos/farmacologia , Humanos , Concentração Inibidora 50 , MAP Quinase Quinase Quinases/metabolismo , Camundongos , Estrutura Molecular , Neoplasias/tratamento farmacológico , Fosfotransferases/química , Fosfotransferases/metabolismo , Piridinas/síntese química , Piridinas/farmacocinética , Piridinas/farmacologia , Relação Estrutura-Atividade , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The discovery and potency optimization of a series of 7-aminofuro[2,3-c]pyridine inhibitors of TAK1 is described. Micromolar hits taken from high-throughput screening were optimized for biochemical and cellular mechanistic potency to ~10nM, as exemplified by compound 12az. Application of structure-based drug design aided by co-crystal structures of TAK1 with inhibitors significantly shortened the number of iterations required for the optimization.
Assuntos
MAP Quinase Quinase Quinases/antagonistas & inibidores , Piridinas , Aminas/síntese química , Aminas/química , Aminas/farmacologia , Animais , Cristalografia por Raios X , Desenho de Fármacos , Ativação Enzimática/efeitos dos fármacos , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/farmacocinética , Inibidores Enzimáticos/farmacologia , Furanos/síntese química , Furanos/química , Furanos/farmacologia , Humanos , Concentração Inibidora 50 , Camundongos , Estrutura Molecular , Neoplasias/tratamento farmacológico , Piridinas/síntese química , Piridinas/farmacocinética , Piridinas/farmacologia , Relação Estrutura-Atividade , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Hyperactive signaling of the MAP kinase pathway resulting from the constitutively active B-Raf(V600E) mutated enzyme has been observed in a number of human tumors, including melanomas. Herein we report the discovery and biological evaluation of GSK2118436, a selective inhibitor of Raf kinases with potent in vitro activity in oncogenic B-Raf-driven melanoma and colorectal carcinoma cells and robust in vivo antitumor and pharmacodynamic activity in mouse models of B-Raf(V600E) human melanoma. GSK2118436 was identified as a development candidate, and early clinical results have shown significant activity in patients with B-Raf mutant melanoma.
RESUMO
In this study, the development of an image-based high-content screening (HCS) binding assay for the seven-transmembrane (7TM) receptor Smoothened (Smo) is described. Using BacMam-based gene delivery of Smo, BODIPY-cyclopamine as a fluorescent probe, and a confocal imaging system, a robust 384-well assay that could be used for high-throughput compound profiling activities was developed. The statistically robust HCS binding assay was developed through optimization of multiple parameters, including cell transduction conditions, Smo expression levels, the image analysis algorithm, and staining procedures. Evaluation of structurally diverse compounds, including functional Smo activators, inhibitors, and related analogs, demonstrated good compound potency correlations between high-content imaging binding, membrane fluorescence polarization binding, and gene reporter assays. Statistical analysis of data from a screening test set of compounds at a single 10-µM concentration suggested that the high-content imaging Smo binding assay is amenable for use in hit identification. The 384-well HCS assay was rapidly developed and met statistical assay performance targets, thus demonstrating its utility as a fluorescent whole-cell binding assay suitable for compound screening and profiling.
Assuntos
Benzamidas/metabolismo , Benzimidazóis/metabolismo , Cicloexilaminas/metabolismo , Avaliação Pré-Clínica de Medicamentos/métodos , Ensaios de Triagem em Larga Escala/métodos , Receptores Acoplados a Proteínas G/antagonistas & inibidores , Receptores Acoplados a Proteínas G/metabolismo , Tiofenos/metabolismo , Algoritmos , Baculoviridae/genética , Benzamidas/química , Benzamidas/farmacologia , Benzimidazóis/química , Benzimidazóis/farmacologia , Linhagem Celular , Cicloexilaminas/química , Cicloexilaminas/farmacologia , Corantes Fluorescentes , Genes Reporter , Células HEK293 , Humanos , Morfolinas/química , Morfolinas/metabolismo , Morfolinas/farmacologia , Piperazinas/química , Piperazinas/metabolismo , Piperazinas/farmacologia , Ligação Proteica , Purinas/química , Purinas/metabolismo , Purinas/farmacologia , Pirazóis/química , Pirazóis/metabolismo , Pirazóis/farmacologia , Receptor Smoothened , Tiofenos/química , Tiofenos/farmacologiaRESUMO
A potent series of inhibitors against the B-Raf(V600E) kinase have been developed that show excellent activity in cellular assays and good oral bioavailability in rats. The key structural features of the series are an arylsulfonamide headgroup, a thiazole core, and a fluorine ortho to the sulfonamide nitrogen.
Assuntos
Inibidores de Proteínas Quinases/química , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Sulfonamidas/química , Administração Oral , Substituição de Aminoácidos , Animais , Sítios de Ligação , Domínio Catalítico , Flúor/química , Microssomos Hepáticos/metabolismo , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/farmacocinética , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas B-raf/metabolismo , Ratos , Relação Estrutura-AtividadeRESUMO
The scope of Pd-catalyzed synthesis of N-Boc-protected anilines from aryl bromides and commercially available tert-butyl carbamate is described. For the first time, this process can be conducted at room temperature (17-22 degrees C) using a combination of Pd(2)dba(3).CHCl(3) and a monodentate ligand, tert-butyl X-Phos. Use of sodium tert-butoxide is crucial to the success of the reaction, which proceeds in 43-83% yield.
Assuntos
Compostos de Anilina/síntese química , Bromobenzenos/química , Carbamatos/química , Catálise , Compostos Organometálicos/química , Paládio/química , TemperaturaRESUMO
The Smoothened receptor (Smo) mediates hedgehog (Hh) signaling critical for development, cell growth, and migration, as well as stem cell maintenance. Aberrant Hh signaling pathway activation has been implicated in a variety of cancers, and small-molecule antagonists of Smo have entered human clinical trials for the treatment of cancer. Here, we report the biochemical characterization of allosteric interactions of agonists and antagonists for Smo. Binding of two radioligands, [(3)H]3-chloro-N-[trans-4-(methylamino)cyclohexyl]-N-{[3-(4-pyridinyl)-phenyl]methyl}-1-benzothiophene-2-carboxamide (SAG-1.3) (agonist) and [(3)H]cyclopamine (antagonist), was characterized using human Smo expressed in human embryonic kidney 293F membranes. We observed full displacement of [(3)H]cyclopamine by all Smo agonist and antagonist ligands examined. N-[(1E)-(3,5-Dimethyl-1-phenyl-1H-pyrazol-4-yl)methylidene]-4-(phenylmethyl)-1-piperazinamine (SANT-1), an antagonist, did not fully inhibit the binding of [(3)H]SAG-1.3. In a functional cell-based beta-lactamase reporter gene assay, SANT-1 and N-[3-(1H-benzimidazol-2-yl)-4-chlorophenyl]-3,4,5-tris(ethyloxy)-benzamide (SANT-2) fully inhibited 3-chloro-4,7-difluoro-N-[trans-4-(methylamino)cyclohexyl]-N-{[3-(4-pyridinyl)phenyl]methyl}-1-benzothiophene-2-carboxamide (SAG-1.5)-induced Hh pathway activation. Detailed "Schild-type" radioligand binding analysis with [(3)H]SAG-1.3 revealed that two structurally distinct Smoothened receptor antagonists, SANT-1 and SANT-2, bound in a manner consistent with that of allosteric modulation. Our mechanism of action characterization of radioligand binding to Smo combined with functional data provides a better understanding of small-molecule interactions with Smo and their influence on the Hh pathway.
Assuntos
Receptores Acoplados a Proteínas G/antagonistas & inibidores , Receptores Acoplados a Proteínas G/metabolismo , Anilidas , Animais , Benzamidas/química , Benzamidas/metabolismo , Benzimidazóis/química , Benzimidazóis/metabolismo , Sítios de Ligação , Ligação Competitiva , Linhagem Celular , Membrana Celular/metabolismo , Cicloexilaminas/química , Cicloexilaminas/metabolismo , Genes Reporter/genética , Humanos , Cinética , Camundongos , Estrutura Molecular , Morfolinas/química , Morfolinas/metabolismo , Células NIH 3T3 , Piperazinas/química , Piperazinas/metabolismo , Purinas/química , Purinas/metabolismo , Pirazóis/química , Pirazóis/metabolismo , Piridinas , Ensaio Radioligante , Receptores Acoplados a Proteínas G/agonistas , Receptores Acoplados a Proteínas G/química , Proteínas Recombinantes/agonistas , Proteínas Recombinantes/antagonistas & inibidores , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Receptor Smoothened , Tiofenos/química , Tiofenos/metabolismo , Tomatina/análogos & derivados , Tomatina/química , Tomatina/metabolismo , Transfecção , Alcaloides de Veratrum/química , Alcaloides de Veratrum/metabolismo , beta-Lactamases/metabolismoRESUMO
A series of thiophene PLK1 inhibitors was optimized for increased solubility and reduced protein binding through the appendage of basic amine functionality. Interesting selectivity between PLK1 and PLK3 was also obtained through these modifications.
Assuntos
Proteínas de Ciclo Celular/antagonistas & inibidores , Química Farmacêutica/métodos , Inibidores de Proteínas Quinases/síntese química , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Tiofenos/química , Ciclo Celular , Proteínas de Ciclo Celular/química , Linhagem Celular Tumoral , Desenho de Fármacos , Humanos , Concentração Inibidora 50 , Mitose , Modelos Químicos , Conformação Molecular , Ligação Proteica , Inibidores de Proteínas Quinases/farmacologia , Proteínas Serina-Treonina Quinases/química , Proteínas Proto-Oncogênicas/química , Solubilidade , Proteínas Supressoras de Tumor , Quinase 1 Polo-LikeRESUMO
Aniline 'headgroups' were synthesized and incorporated into an alkynyl thienopyrimidine series of EGFR and ErbB-2 inhibitors. Potent inhibition of enzyme activity and cellular proliferation was observed. In certain instances, protein binding was reduced and oral exposure was found to be somewhat improved relative to compounds containing the reference aniline.
Assuntos
Compostos de Anilina/síntese química , Receptores ErbB/antagonistas & inibidores , Inibidores de Proteínas Quinases/síntese química , Pirimidinas/síntese química , Receptor ErbB-2/antagonistas & inibidores , Administração Oral , Compostos de Anilina/administração & dosagem , Compostos de Anilina/farmacologia , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos/métodos , Receptores ErbB/metabolismo , Inibidores do Crescimento/administração & dosagem , Inibidores do Crescimento/síntese química , Inibidores do Crescimento/farmacologia , Humanos , Camundongos , Inibidores de Proteínas Quinases/administração & dosagem , Inibidores de Proteínas Quinases/farmacologia , Pirimidinas/administração & dosagem , Pirimidinas/farmacologia , Receptor ErbB-2/metabolismoRESUMO
A novel class of substituted pyrrolidinyl-acetylenic thieno[3,2-d]pyrimidines has been identified that are potent and selective inhibitors of both EGFR/ErbB-2 receptor tyrosine kinases. The inhibitors are found to display a range of enzyme and cellular potency and also to display a varying level of covalent modification of the kinase targets. Selected molecules, including compound 15h, were found to be potent in enzymatic and cellular assays while also demonstrating exposure in the mouse from an oral dose.
Assuntos
Receptores ErbB/antagonistas & inibidores , Pirimidinas/farmacologia , Receptor ErbB-2/antagonistas & inibidores , Animais , Linhagem Celular , Camundongos , Ligação Proteica , Pirimidinas/química , Relação Estrutura-AtividadeRESUMO
Yeast hexokinase (EC 2.7.1.1) catalyzes the phosphorylation of pyranose and furanose analogs of glucose at 0.01-125% of the rate of glucose. The enzyme is highly tolerant of structural changes at C-2 and C-3 of glucopyranose and less tolerant of changes at C-1 and C-4. Preparative phosphorylations were performed on compounds having 0.01-100% of the activity of glucose, using phosphoenolpyruvate and pyruvate kinase to regenerate ATP. The effects of inhibition of hexokinase by phosphoenolpyruvate and acetyl phosphate on cofactor regeneration are discussed.