RÉSUMÉ
Mycobacterium tuberculosis drug resistance is emerging and new drug targets are needed. Tryptophan biosynthesis is necessary for M. tuberculosis replication and virulence. Indole-3-glycerol phosphate synthase (IGPS) catalyzes a step in M. tuberculosis tryptophan biosynthesis and has been suggested as a potential anti-infective target, but our understanding of this enzyme is limited. To aid in inhibitor design and gain a greater mechanistic picture of this enzyme, there is a need to understand the roles of active site amino acids in ligand binding and catalysis. In this work, we explored the roles of conserved active site amino acids Glu57, Lys59, Lys119, Glu168, and Glu219. Mutation of each to Ala results in loss of all detectable activity. The Glu57Gln, Lys59Arg, Lys119Arg, Glu168Gln, and Glu219Asp mutations result in large activity losses, while Glu219Gln has enhanced activity. Analysis of the enzymatic data yields the following main conclusions: (A) Lys119 is the likely catalytic acid in the CdRP ring closure step. (B) Glu168 stabilizes a charged reaction intermediate and may also be the catalytic base. (C) Glu57, Glu219, and Lys119 form a closely arranged triad in which Glu57 and Glu219 modulate the pKa of Lys119, and thus overall activity. This increased understanding of inter- and intramolecular interactions and demonstration of the highly coordinated nature of the M. tuberculosis IGPS active site provide new mechanistic information and guidance for future work with this potential new drug target.
RÉSUMÉ
Lymphatic filariasis is a debilitating illness with an estimated 50 million cases as of 2018. The majority of cases are caused by the parasitic worm W. bancrofti and additional cases by the worms B. malayi and B. timori. Dihydrofolate reductase (DHFR) is an established target in the treatment of cancer, bacterial, and protozoal infections and may be a potential target for drugs targeting parasitic worm infections, including filariasis. Recent studies have shown that known antifolate compounds, including methotrexate, inhibit the activity of W. bancrofti DHFR (WbDHFR). However, the absence of structural information for filarial DHFRs has limited the study of more in-depth structure-function relationships. We report the structure of WbDHFR complexed with NADPH and folate using X-ray diffraction data measured to 2.47 Å resolution. The structure of WbDHFR reveals the usual DHFR fold and is currently only the second nematode DHFR structure in the Protein Data Bank. The equilibrium dissociation constants for NADPH (90 ± 29 nM) and folate (23 ± 4 nM) were determined by equilibrium titrations. The interactions of known antifolates with WbDHFR were analyzed using molecular docking programs and molecular dynamics simulations. Antifolates with a hydrophobic core and extended linker formed favorable interactions with WbDHFR. These combined data should now facilitate the rational design of filarial DHFR inhibitors, which in turn can be used to determine whether DHFR is a viable drug target for filariasis and whether existing antifolates may be repurposed for its treatment.
Sujet(s)
Filariose lymphatique , Antifoliques , Animaux , Antifoliques/pharmacologie , Antifoliques/métabolisme , Wuchereria bancrofti , Acide folique , Dihydrofolate reductase/métabolisme , NADP , Simulation de docking moléculaireRÉSUMÉ
Many antibacterial and antiparasitic drugs work by competitively inhibiting dihydrofolate reductase (DHFR), a vital enzyme in folate metabolism. The interactions between inhibitors and DHFR active site residues are known in many homologs but the contributions from distal residues are less understood. Identifying distal residues that aid in inhibitor binding can improve targeted drug development programs by accounting for distant influences that may be less conserved and subject to frequent resistance causing mutations. Previously, a novel, homology-based, computational approach that mines ligand inhibition data was used to predict residues involved in inhibitor selectivity in the DHFR family. Expectedly, some inhibitor selectivity determining residue positions were predicted to lie in the active site and coincide with experimentally known inhibitor selectivity determining positions. However, other residues that group spatially in clusters distal to the active site have not been previously investigated. In this study, the effect of introducing amino acid substitutions at one of these predicted clusters (His38-Ala39-Ile40) on the inhibitor selectivity profile in Bacillus stearothermophilus dihydrofolate reductase (Bs DHFR) was investigated. Mutations were introduced into these cluster positions to change sidechain chemistry and size. We determined kcat and KM values and measured KD values at equilibrium for two competitive DHFR inhibitors, trimethoprim (TMP) and pyrimethamine (PYR). Mutations in the His38-Ala39-Ile40 cluster significantly impacted inhibitor binding and TMP/PYR selectivity - seven out of nine mutations resulted in tighter binding to PYR when compared to TMP. These data suggest that the His38-Ala39-Ile40 cluster is a distal inhibitor selectivity determining region that favors PYR binding in Bs DHFR and, possibly, throughout the DHFR family.
Sujet(s)
Protéines bactériennes/antagonistes et inhibiteurs , Protéines bactériennes/composition chimique , Antifoliques/composition chimique , Geobacillus stearothermophilus/enzymologie , Mutation faux-sens , Dihydrofolate reductase/composition chimique , Substitution d'acide aminé , Protéines bactériennes/génétique , Geobacillus stearothermophilus/génétique , Dihydrofolate reductase/génétiqueRÉSUMÉ
Filariasis is a tropical disease caused by the parasitic nematodes Wuchereria bancrofti and Brugia malayi. Known inhibitors of dihydrofolate reductase (DHFR) have been previously shown to kill Brugia malayi nematodes and to inhibit Brugia malayi DHFR (BmDHFR) at nanomolar concentrations. These data suggest that BmDHFR is a potential target for the treatment of filariasis. Here, protocols for cloning, expression and purification of Wuchereria bancrofti DHFR (WbDHFR) were developed. The Uniprot entry J9F199-1 predicts a 172 amino acid protein for WbDHFR but alignment of this sequence to the previously described BmDHFR shows that this WbDHFR sequence lacks a crucial, conserved 13 amino acid loop. The presence of the loop in WbDHFR is supported by a noncanonical splicing event and the loop sequence was therefore included in the gene design. Subsequently, the KM for dihydrofolate (3.7 ± 2 µM), kcat (7.4 ± 0.6 s-1), and pH dependence of activity were determined. IC50 values of methotrexate, trimethoprim, pyrimethamine, raltitrexed, aminopterin, (-)-epicatechin gallate, (-)-epicatechin, and vitexin were measured for WbDHFR and BmDHFR. Methotrexate and structurally related aminopterin were found to be effective inhibitors of WbDHFR, with an KI of 1.2 ± 0.2 nM and 2.1 ± 0.5 nM, respectively, suggesting that repurposing of known antifolate compound may be an effective strategy to treating filariasis. Most compounds showed similar inhibition profiles toward both enzymes, suggesting that the two enzymes have important similarities in their active site environments and can be targeted with the same compound, once a successful inhibitor is identified.
Sujet(s)
Dihydrofolate reductase/composition chimique , Dihydrofolate reductase/métabolisme , Wuchereria bancrofti/enzymologie , Séquence d'acides aminés , Animaux , Brugia malayi/enzymologie , Brugia malayi/génétique , Clonage moléculaire , Antienzymes/pharmacologie , Flavonoïdes/pharmacologie , Régulation de l'expression des gènes codant pour des enzymes/effets des médicaments et des substances chimiques , Alignement de séquences , Dihydrofolate reductase/génétique , Dihydrofolate reductase/isolement et purification , Wuchereria bancrofti/génétiqueRÉSUMÉ
Tankyrase activity has been linked to the regulation of intracellular axin levels, which have been shown to be crucial for the Wnt pathway. Deregulated Wnt signaling is important for the genesis of many diseases including cancer. We describe herein the discovery and development of a new series of tankyrase inhibitors. These pyranopyridones are highly active in various cell-based assays. A fragment/structure based optimization strategy led to a compound with good pharmacokinetic properties that is suitable for in vivo studies and further development.
RÉSUMÉ
The field of small-molecule inhibitors of protein-protein interactions is rapidly advancing and the specific area of inhibitors of the p53/MDM2 interaction is a prime example. Several groups have published on this topic and multiple compounds are in various stages of clinical development. Building on the strength of the discovery of RG7112, a Nutlin imidazoline-based compound, and RG7388, a pyrrolidine-based compound, we have developed additional scaffolds that provide opportunities for future development. Here, we report the discovery and optimization of a highly potent and selective series of spiroindolinone small-molecule MDM2 inhibitors, culminating in RO8994.
Sujet(s)
Indoles/composition chimique , Indolizidines/composition chimique , Protéines proto-oncogènes c-mdm2/antagonistes et inhibiteurs , Spiranes/composition chimique , Apoptose/effets des médicaments et des substances chimiques , Sites de fixation , Lignée cellulaire tumorale , Prolifération cellulaire/effets des médicaments et des substances chimiques , Évaluation préclinique de médicament , Humains , Imidazolines/composition chimique , Indoles/usage thérapeutique , Indoles/toxicité , Indolizidines/usage thérapeutique , Indolizidines/toxicité , Simulation de dynamique moléculaire , Tumeurs/traitement médicamenteux , Liaison aux protéines , Structure tertiaire des protéines , Protéines proto-oncogènes c-mdm2/métabolisme , Pyrrolidines/composition chimique , Spiranes/usage thérapeutique , Spiranes/toxicité , Protéine p53 suppresseur de tumeur/antagonistes et inhibiteurs , Protéine p53 suppresseur de tumeur/métabolisme , para-Aminobenzoates/composition chimiqueRÉSUMÉ
DYRK1B is a kinase over-expressed in certain cancer cells (including colon, ovarian, pancreatic, etc.). Recent publications have demonstrated inhibition of DYRK1B could be an attractive target for cancer therapy. From a data-mining effort, the team has discovered analogues of pyrido[2,3-d]pyrimidines as potent enantio-selective inhibitors of DYRK1B. Cells treated with a tool compound from this series showed the same cellular effects as down regulation of DYRK1B with siRNA. Such effects are consistent with the proposed mechanism of action. Progress of the SAR study is presented.
Sujet(s)
Inhibiteurs de protéines kinases/composition chimique , Inhibiteurs de protéines kinases/pharmacologie , Protein-Serine-Threonine Kinases/antagonistes et inhibiteurs , Protein-tyrosine kinases/antagonistes et inhibiteurs , Pyrimidines/composition chimique , Animaux , Sites de fixation , Cristallographie aux rayons X , Activation enzymatique/effets des médicaments et des substances chimiques , Période , Humains , Simulation de dynamique moléculaire , Inhibiteurs de protéines kinases/pharmacocinétique , Protein-Serine-Threonine Kinases/métabolisme , Structure tertiaire des protéines , Protein-tyrosine kinases/métabolisme , Pyrimidines/pharmacocinétique , Rats , Relation structure-activité ,RÉSUMÉ
XIAP is a key regulator of apoptosis, and its overexpression in cancer cells may contribute to their survival. The antiapoptotic function of XIAP derives from its BIR domains, which bind to and inhibit pro-apoptotic caspases. Most known IAP inhibitors are selective for the BIR3 domain and bind to cIAP1 and cIAP2 as well as XIAP. Pathways activated upon cIAP binding contribute to the function of these compounds. Inhibitors selective for XIAP should exert pro-apoptotic effects through competition with the terminal caspases. This paper details our synthetic explorations of a novel XIAP BIR2-selective benzazepinone screening hit with a focus on increasing BIR2 potency and overcoming high in vivo clearance. These efforts led to the discovery of benzoxazepinone 40, a potent BIR2-selective inhibitor with good in vivo pharmacokinetic properties which potentiates apoptotic signaling in a manner mechanistically distinct from that of known pan-IAP inhibitors.
Sujet(s)
Composés hétérocycliques/synthèse chimique , Composés hétérocycliques/pharmacologie , Protéines IAP/antagonistes et inhibiteurs , Protéine inhibitrice de l'apoptose liée au chromosome X/antagonistes et inhibiteurs , Alanine/analogues et dérivés , Alanine/synthèse chimique , Alanine/pharmacocinétique , Alanine/pharmacologie , Animaux , Anticorps monoclonaux/pharmacologie , Apoptose/effets des médicaments et des substances chimiques , Technique de Western , Caspase-3/métabolisme , Caspase-7/métabolisme , Lignée cellulaire tumorale , Survie cellulaire/effets des médicaments et des substances chimiques , Cristallographie aux rayons X , Femelle , Composés hétérocycliques/pharmacocinétique , Humains , Protéines IAP/composition chimique , Protéines IAP/métabolisme , Souris , Souris nude , Modèles chimiques , Modèles moléculaires , Structure moléculaire , Oxazépines/synthèse chimique , Oxazépines/pharmacocinétique , Oxazépines/pharmacologie , Structure tertiaire des protéines , Rats , Ubiquitin-protein ligases , Protéine inhibitrice de l'apoptose liée au chromosome X/composition chimique , Protéine inhibitrice de l'apoptose liée au chromosome X/métabolisme , Tests d'activité antitumorale sur modèle de xénogreffeRÉSUMÉ
The IAPs are key regulators of the apoptotic pathways and are commonly overexpressed in many cancer cells. IAPs contain one to three BIR domains that are crucial for their inhibitory function. The pro-survival properties of XIAP come from binding of the BIR domains to the pro-apoptotic caspases. The BIR3 domain of XIAP binds and inhibits caspase 9, while the BIR2 domain binds and inhibits the terminal caspases 3 and 7. While XIAP BIR3 inhibitors have previously been reported, they also inhibit cIAP1/2 and promote the release of TNFα, potentially limiting their therapeutic utility. This paper will focus on the optimization of selective XIAP BIR2 inhibitors leading to the discovery of highly potent benzodiazepinone 36 (IC50 = 45 nM), which has high levels of selectivity over XIAP BIR3 and cIAP1 BIR2/3 and shows efficacy in a xenograft pharmacodynamic model monitoring caspase activity while not promoting the release of TNFα in vitro.
Sujet(s)
Composés hétérocycliques/synthèse chimique , Composés hétérocycliques/pharmacologie , Protéines IAP/antagonistes et inhibiteurs , Protéine inhibitrice de l'apoptose liée au chromosome X/antagonistes et inhibiteurs , Alanine/analogues et dérivés , Alanine/synthèse chimique , Alanine/pharmacocinétique , Alanine/pharmacologie , Animaux , Apoptose/effets des médicaments et des substances chimiques , Benzodiazépinones/synthèse chimique , Benzodiazépinones/pharmacocinétique , Benzodiazépinones/pharmacologie , Technique de Western , Caspase-3/métabolisme , Caspase-7/métabolisme , Lignée cellulaire tumorale , Survie cellulaire/effets des médicaments et des substances chimiques , Cristallographie aux rayons X , Femelle , Composés hétérocycliques/pharmacocinétique , Humains , Protéines IAP/composition chimique , Protéines IAP/métabolisme , Souris , Souris nude , Modèles chimiques , Modèles moléculaires , Structure moléculaire , Structure tertiaire des protéines , Ubiquitin-protein ligases , Protéine inhibitrice de l'apoptose liée au chromosome X/composition chimique , Protéine inhibitrice de l'apoptose liée au chromosome X/métabolisme , Tests d'activité antitumorale sur modèle de xénogreffeRÉSUMÉ
XIAP, a member of the inhibitor of apoptosis family of proteins, is a critical regulator of apoptosis. Inhibition of the BIR domain-caspase interaction is a promising approach towards treating cancer. Previous work has been directed towards inhibiting the BIR3-caspase-9 interaction, which blocks the intrinsic apoptotic pathway; selectively inhibiting the BIR2-caspase-3 interaction would also block the extrinsic pathway. The BIR2 domain of XIAP has successfully been crystallized; peptides and small-molecule inhibitors can be soaked into these crystals, which diffract to high resolution. Here, the BIR2 apo crystal structure and the structures of five BIR2-tetrapeptide complexes are described. The structural flexibility observed on comparing these structures, along with a comparison with XIAP BIR3, affords an understanding of the structural elements that drive selectivity between BIR2 and BIR3 and which can be used to design BIR2-selective inhibitors.
Sujet(s)
Caspase-3/composition chimique , Caspase-3/métabolisme , Inhibiteurs des caspases/composition chimique , Protéines IAP/composition chimique , Nucleopolyhedrovirus/composition chimique , Protéines virales/composition chimique , Protéine inhibitrice de l'apoptose liée au chromosome X/composition chimique , Séquence d'acides aminés , Apoprotéines/composition chimique , Apoprotéines/génétique , Apoptose/génétique , Cristallographie aux rayons X , Humains , Protéines IAP/génétique , Données de séquences moléculaires , Famille multigénique/génétique , Nucleopolyhedrovirus/génétique , Oligopeptides/composition chimique , Oligopeptides/génétique , Cartographie d'interactions entre protéines , Structure tertiaire des protéines/génétique , Protéines virales/génétique , Protéine inhibitrice de l'apoptose liée au chromosome X/génétiqueRÉSUMÉ
Restoration of p53 activity by inhibition of the p53-MDM2 interaction has been considered an attractive approach for cancer treatment. However, the hydrophobic protein-protein interaction surface represents a significant challenge for the development of small-molecule inhibitors with desirable pharmacological profiles. RG7112 was the first small-molecule p53-MDM2 inhibitor in clinical development. Here, we report the discovery and characterization of a second generation clinical MDM2 inhibitor, RG7388, with superior potency and selectivity.
Sujet(s)
Antinéoplasiques/synthèse chimique , Protéines proto-oncogènes c-mdm2/antagonistes et inhibiteurs , Pyrrolidines/synthèse chimique , Protéine p53 suppresseur de tumeur/antagonistes et inhibiteurs , para-Aminobenzoates/synthèse chimique , Animaux , Antinéoplasiques/pharmacologie , Antinéoplasiques/usage thérapeutique , Découverte de médicament , Humains , Mâle , Souris , Souris de lignée C57BL , Pyrrolidines/pharmacologie , Pyrrolidines/usage thérapeutique , para-Aminobenzoates/pharmacologie , para-Aminobenzoates/usage thérapeutiqueRÉSUMÉ
A series of amino-pyrimidines was developed based upon an initial kinase cross-screening hit from a CDK2 program. Kinase profiling and structure-based drug design guided the optimization from the initial 1,2,3-benzotriazole hit to a potent and selective JNK inhibitor, compound 24f (JNK1 and 2 IC(50)=16 and 66 nM, respectively), with bioavailability in rats and suitable for further in vivo pharmacological evaluation.
Sujet(s)
JNK Mitogen-Activated Protein Kinases/antagonistes et inhibiteurs , Inhibiteurs de protéines kinases/composition chimique , Inhibiteurs de protéines kinases/pharmacologie , Pyrimidines/composition chimique , Pyrimidines/pharmacologie , Triazoles/composition chimique , Triazoles/pharmacologie , Animaux , Cristallographie aux rayons X , Conception de médicament , Humains , JNK Mitogen-Activated Protein Kinases/métabolisme , Modèles moléculaires , Inhibiteurs de protéines kinases/synthèse chimique , Pyrimidines/synthèse chimique , Rats , Relation structure-activité , Triazoles/synthèse chimiqueRÉSUMÉ
Protein-protein interaction (PPI) systems represent a rich potential source of targets for drug discovery, but historically have proven to be difficult, particularly in the lead identification stage. Application of the fragment-based approach may help toward success with this target class. To provide an example toward understanding the potential issues associated with such an application, we have deconstructed one of the best established protein-protein inhibitors, the Nutlin series that inhibits the interaction between MDM2 and p53, into fragments, and surveyed the resulting binding properties using heteronuclear single quantum coherence nuclear magnetic resonance (HSQC NMR), surface plasmon resonance (SPR), and X-ray crystallography. We report the relative contributions toward binding affinity for each of the key substituents of the Nutlin molecule and show that this series could hypothetically have been discovered via a fragment approach. We find that the smallest fragment of Nutlin that retains binding accesses two subpockets of MDM2 and has a molecular weight at the high end of the range that normally defines fragments.
RÉSUMÉ
Activation of p53 tumor suppressor by antagonizing its negative regulator murine double minute (MDM)2 has been considered an attractive strategy for cancer therapy and several classes of p53-MDM2 binding inhibitors have been developed. However, these compounds do not inhibit the p53-MDMX interaction, and their effectiveness can be compromised in tumors overexpressing MDMX. Here, we identify small molecules that potently block p53 binding with both MDM2 and MDMX by inhibitor-driven homo- and/or heterodimerization of MDM2 and MDMX proteins. Structural studies revealed that the inhibitors bind into and occlude the p53 pockets of MDM2 and MDMX by inducing the formation of dimeric protein complexes kept together by a dimeric small-molecule core. This mode of action effectively stabilized p53 and activated p53 signaling in cancer cells, leading to cell cycle arrest and apoptosis. Dual MDM2/MDMX antagonists restored p53 apoptotic activity in the presence of high levels of MDMX and may offer a more effective therapeutic modality for MDMX-overexpressing cancers.
Sujet(s)
Apoptose/physiologie , Hydantoïnes/pharmacologie , Modèles moléculaires , Protéines nucléaires/antagonistes et inhibiteurs , Protéines proto-oncogènes c-mdm2/antagonistes et inhibiteurs , Protéines proto-oncogènes/antagonistes et inhibiteurs , Protéine p53 suppresseur de tumeur/métabolisme , Apoptose/effets des médicaments et des substances chimiques , Technique de Western , Protéines du cycle cellulaire , Lignée cellulaire tumorale , Cristallisation , Dimérisation , Transfert d'énergie par résonance de fluorescence , Humains , Résonance magnétique nucléaire biomoléculaire , Protéines nucléaires/composition chimique , Protéines proto-oncogènes/composition chimique , Protéines proto-oncogènes c-mdm2/composition chimique , Transduction du signal/effets des médicaments et des substances chimiques , Transduction du signal/physiologie , Sels de tétrazolium , ThiazolesRÉSUMÉ
3-[4-((1S,2S,3R,5S,7S)-5-Hydroxyadamantan-2-ylcarbamoyl)benzyl]-4-oxo-1-phenyl-1,4-dihydro-[1,8]naphthyridine-2-carboxylic acid methyl ester (4) was identified as a novel, druglike and selective quinolone pan JNK inhibitor. In this communication, some of the structure-activity relationship of the azaquinolone analogues leading to 4 is discussed. The focus is on how changes at the amide functionality affected the biochemical potency, cellular potency, metabolic properties, and solubility of this class of JNK inhibitors. Optimization of these properties led to the identification of the adamantyl analogue, 4. 4 achieved proof of mechanism in both rat and mouse TNF-α challenge models.
RÉSUMÉ
The MAP kinase pathway is one of the most important pathways involved in cell proliferation and differentiation, and its components are promising targets for antitumor drugs. Design and synthesis of a novel MEK inhibitor, based on the 3D-structural information of the target enzyme, and then multidimensional optimization including metabolic stability, physicochemical properties and safety profiles were effectively performed and led to the identification of a clinical candidate for an orally available potent MEK inhibitor, CH4987655, possessing a unique 3-oxo-oxazinane ring structure at the 5-position of the benzamide core structure. CH4987655 exhibits slow dissociation from the MEK enzyme, remarkable in vivo antitumor efficacy both in mono- and combination therapy, desirable metabolic stability, and insignificant MEK inhibition in mouse brain, implying few CNS-related side effects in human. An excellent PK profile and clear target inhibition in PBMC were demonstrated in a healthy volunteer clinical study.
Sujet(s)
Antinéoplasiques/composition chimique , Benzamides/composition chimique , MAP Kinase Kinase Kinases/antagonistes et inhibiteurs , Oxazines/composition chimique , Inhibiteurs de protéines kinases/composition chimique , Administration par voie orale , Régulation allostérique , Antinéoplasiques/administration et posologie , Antinéoplasiques/pharmacologie , Benzamides/administration et posologie , Benzamides/pharmacologie , Humains , Modèles moléculaires , Oxazines/administration et posologie , Oxazines/pharmacologie , Inhibiteurs de protéines kinases/administration et posologie , Inhibiteurs de protéines kinases/pharmacologieRÉSUMÉ
IL-2-inducible T cell kinase plays an essential role in T cell receptor signaling and is considered a drug target for the treatment of Th2-mediated inflammatory diseases. By applying high-throughput protein engineering and crystallization, we have determined the X-ray crystal structures of IL-2-inducible T cell kinase in complex with its selective inhibitor BMS-509744 and the broad-spectrum kinase inhibitors sunitinib and RO5191614. Sunitinib uniquely stabilizes IL-2-inducible T cell kinase in the helix C-in conformation by inducing side chain conformational changes in the ATP-binding site. This preference of sunitinib to bind to an active kinase conformation is reflective of its broad-spectrum kinase activity. BMS-509744 uniquely stabilizes the activation loop in a substrate-blocking inactive conformation, indicating that structural changes described for Src family kinases are also involved in the regulation of IL-2-inducible T cell kinase activity. The observed BMS-509744 binding mode allows rationalization of structure-activity relationships reported for this inhibitor class and facilitates further structure-based drug design. Sequence-based analysis of this binding mode provides guidance for the rational design of inhibitor selectivity.
Sujet(s)
Conception de médicament , Inhibiteurs de protéines kinases/composition chimique , Protein-tyrosine kinases/antagonistes et inhibiteurs , Sites de fixation , Cristallographie aux rayons X , Indoles/composition chimique , Indoles/pharmacologie , Ingénierie des protéines , Inhibiteurs de protéines kinases/pharmacologie , Structure tertiaire des protéines , Protein-tyrosine kinases/métabolisme , Pyrroles/composition chimique , Pyrroles/pharmacologie , Relation structure-activité , Sunitinib , src-Family kinases/métabolismeRÉSUMÉ
High-throughput screening for inhibitors of the human metalloprotease, methionine aminopeptidase-2 (MetAP2), identified a potent class of 3-anilino-5-benzylthio-1,2,4-triazole compounds. Efficient array and interative synthesis of triazoles led to rapid SAR development around the aniline, benzylthio, and triazole moeities. Evaluation of these analogs in a human MetAP2 enzyme assay led to the identification of several inhibitors with potencies in the 50-100 picomolar range. The deleterious effects on inhibitor potency by methylation of the anilino-triazole nitrogens, as well as the X-ray crystal structure of triazole 102 bound in the active site of MetAP2, confirm the key interactions between the triazole nitrogens, the active site cobalt atoms, and the His-231 side-chain. The structure has also provided a rationale for interpreting SAR within the triazole series. Key aniline (2-isopropylphenyl) and sulfur substituents (furanylmethyl) identified in the SAR studies led to the identification of potent inhibitors (103 and 104) of endothelial cell proliferation. Triazoles 103 and 104 also exhibited dose-dependent activity in an aortic ring tissue model of angiogenesis highlighting the potential utility of MetAP2 inhibitors as anticancer agents.
Sujet(s)
Aminopeptidases/antagonistes et inhibiteurs , Inhibiteurs de l'angiogenèse/synthèse chimique , Furanes/synthèse chimique , Metalloendopeptidases/antagonistes et inhibiteurs , Thiazoles/synthèse chimique , Thiophènes/synthèse chimique , Triazoles/synthèse chimique , Inhibiteurs de l'angiogenèse/composition chimique , Inhibiteurs de l'angiogenèse/pharmacologie , Animaux , Aorte thoracique/effets des médicaments et des substances chimiques , Vaisseaux capillaires/effets des médicaments et des substances chimiques , Prolifération cellulaire/effets des médicaments et des substances chimiques , Cristallographie aux rayons X , Cellules endothéliales/cytologie , Cellules endothéliales/effets des médicaments et des substances chimiques , Endothélium vasculaire/cytologie , Furanes/composition chimique , Furanes/pharmacologie , Techniques in vitro , Mâle , Modèles moléculaires , Rats , Rat Sprague-Dawley , Relation structure-activité , Thiazoles/composition chimique , Thiazoles/pharmacologie , Thiophènes/composition chimique , Thiophènes/pharmacologie , Triazoles/composition chimique , Triazoles/pharmacologieRÉSUMÉ
The syntheses, in vitro characterizations, and rat and monkey in vivo pharmacokinetic profiles of a series of 5-, 6-, and 7-methyl-substituted azepanone-based cathepsin K inhibitors are described. Depending on the particular regiochemical substitution and stereochemical configuration, methyl-substituted azepanones were identified that had widely varied cathepsin K inhibitory potency as well as pharmacokinetic properties compared to the 4S-parent azepanone analogue, 1 (human cathepsin K, K(i,app) = 0.16 nM, rat oral bioavailability = 42%, rat in vivo clearance = 49.2 mL/min/kg). Of particular note, the 4S-7-cis-methylazepanone analogue, 10, had a K(i,app) = 0.041 nM vs human cathepsin K and 89% oral bioavailability and an in vivo clearance rate of 19.5 mL/min/kg in the rat. Hypotheses that rationalize some of the observed characteristics of these closely related analogues have been made using X-ray crystallography and conformational analysis. These examples demonstrate the potential for modulation of pharmacological properties of cathepsin inhibitors by substituting the azepanone core. The high potency for inhibition of cathepsin K coupled with the favorable rat and monkey pharmacokinetic characteristics of compound 10, also known as SB-462795 or relacatib, has made it the subject of considerable in vivo evaluation for safety and efficacy as an inhibitor of excessive bone resorption in rat, monkey, and human studies, which will be reported elsewhere.
Sujet(s)
Azépines/synthèse chimique , Agents de maintien de la densité osseuse/synthèse chimique , Cathepsines/antagonistes et inhibiteurs , Sulfones/synthèse chimique , Animaux , Azépines/composition chimique , Azépines/pharmacologie , Biodisponibilité , Protéines du sang/métabolisme , Agents de maintien de la densité osseuse/composition chimique , Agents de maintien de la densité osseuse/pharmacologie , Cathepsine K , Cathepsines/composition chimique , Lignée cellulaire , Perméabilité des membranes cellulaires , Cristallographie aux rayons X , Haplorhini , Humains , Conformation moléculaire , Liaison aux protéines , Rats , Stéréoisomérie , Relation structure-activité , Sulfones/composition chimique , Sulfones/pharmacologieRÉSUMÉ
Inhibitors of human methionine aminopeptidase type 2 (hMetAP2) are of interest as potential treatments for cancer. A new class of small molecule reversible inhibitors of hMetAP2 was discovered and optimized, the 4-aryl-1,2,3-triazoles. Compound 24, a potent inhibitor of cobalt-activated hMetAP2, also inhibits human and mouse endothelial cell growth. Using a mouse matrigel model, this reversible hMetAP2 inhibitor was also shown to inhibit angiogenesis in vivo.