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1.
Cancer Cell ; 29(6): 889-904, 2016 06 13.
Article in English | MEDLINE | ID: mdl-27265506

ABSTRACT

Activating mutations in Gαq proteins, which form the α subunit of certain heterotrimeric G proteins, drive uveal melanoma oncogenesis by triggering multiple downstream signaling pathways, including PLC/PKC, Rho/Rac, and YAP. Here we show that the small GTPase ARF6 acts as a proximal node of oncogenic Gαq signaling to induce all of these downstream pathways as well as ß-catenin signaling. ARF6 activates these diverse pathways through a common mechanism: the trafficking of GNAQ and ß-catenin from the plasma membrane to cytoplasmic vesicles and the nucleus, respectively. Blocking ARF6 with a small-molecule inhibitor reduces uveal melanoma cell proliferation and tumorigenesis in a mouse model, confirming the functional relevance of this pathway and suggesting a therapeutic strategy for Gα-mediated diseases.


Subject(s)
ADP-Ribosylation Factors/metabolism , GTP-Binding Protein alpha Subunits, Gq-G11/genetics , Melanoma/drug therapy , Small Molecule Libraries/administration & dosage , Uveal Neoplasms/drug therapy , beta Catenin/metabolism , ADP-Ribosylation Factor 6 , ADP-Ribosylation Factors/antagonists & inhibitors , ADP-Ribosylation Factors/genetics , Animals , Cell Line, Tumor , Cell Membrane/metabolism , Cell Nucleus/metabolism , Cell Proliferation/drug effects , Cytoplasm/metabolism , GTP-Binding Protein alpha Subunits, Gq-G11/metabolism , Humans , Melanoma/genetics , Melanoma/metabolism , Mice , Neoplasm Transplantation , Protein Transport/drug effects , Signal Transduction/drug effects , Small Molecule Libraries/pharmacology , Uveal Neoplasms/genetics , Uveal Neoplasms/metabolism
2.
Bioorg Med Chem Lett ; 23(24): 6829-33, 2013 Dec 15.
Article in English | MEDLINE | ID: mdl-24183538

ABSTRACT

Mps1, also known as TTK, is a mitotic checkpoint protein kinase that has become a promising new target of cancer research. In an effort to improve the lead-likeness of our recent Mps1 purine lead compounds, a scaffold hopping exercise has been undertaken. Structure-based design, principles of conformational restriction, and subsequent scaffold hopping has led to novel pyrrolopyrimidine and quinazoline Mps1 inhibitors. These new single-digit nanomolar leads provide the basis for developing potent, novel Mps1 inhibitors with improved drug-like properties.


Subject(s)
Cell Cycle Proteins/antagonists & inhibitors , Protein Kinase Inhibitors/chemistry , Protein Kinase Inhibitors/pharmacology , Protein Serine-Threonine Kinases/antagonists & inhibitors , Protein-Tyrosine Kinases/antagonists & inhibitors , Purines/chemistry , Pyrimidines/chemistry , Pyrimidines/pharmacology , Pyrroles/chemistry , Pyrroles/pharmacology , Quinazolines/chemistry , Cell Cycle Proteins/metabolism , Cell Proliferation/drug effects , Drug Design , HCT116 Cells , Humans , Models, Molecular , Protein Binding , Protein Kinase Inhibitors/metabolism , Protein Serine-Threonine Kinases/metabolism , Protein-Tyrosine Kinases/metabolism , Purines/metabolism , Purines/pharmacology , Pyrimidines/metabolism , Pyrroles/metabolism , Quinazolines/metabolism , Quinazolines/pharmacology , Structure-Activity Relationship
3.
J Med Chem ; 55(17): 7480-501, 2012 Sep 13.
Article in English | MEDLINE | ID: mdl-22913511

ABSTRACT

Modulation of Hsp90 (heat shock protein 90) function has been recognized as an attractive approach for cancer treatment, since many cancer cells depend on Hsp90 to maintain cellular homeostasis. This has spurred the search for small-molecule Hsp90 inhibitors. Here we describe our lead optimization studies centered on the purine-based Hsp90 inhibitor 28a containing a piperidine moiety at the purine N9 position. In this study, key SAR was established for the piperidine N-substituent and for the congeners of the 1,3-benzodioxole at C8. These efforts led to the identification of orally bioavailable 28g that exhibits good in vitro profiles and a characteristic molecular biomarker signature of Hsp90 inhibition both in vitro and in vivo. Favorable pharmacokinetic properties along with significant antitumor effects in multiple human cancer xenograft models led to the selection of 28g (MPC-3100) as a clinical candidate.


Subject(s)
Adenine/analogs & derivatives , Benzodioxoles/chemistry , Benzodioxoles/pharmacology , HSP90 Heat-Shock Proteins/antagonists & inhibitors , Adenine/chemistry , Adenine/pharmacokinetics , Adenine/pharmacology , Animals , Benzodioxoles/pharmacokinetics , Chromatography, High Pressure Liquid , Drug Discovery , Inhibitory Concentration 50 , Mice , Models, Molecular , Structure-Activity Relationship
4.
Bioorg Med Chem Lett ; 22(13): 4377-85, 2012 Jul 01.
Article in English | MEDLINE | ID: mdl-22632936

ABSTRACT

Efforts to optimize biological activity, novelty, selectivity and oral bioavailability of Mps1 inhibitors, from a purine based lead MPI-0479605, are described in this Letter. Mps1 biochemical activity and cytotoxicity in HCT-116 cell line were improved. On-target activity confirmation via mechanism based G2/M escape assay was demonstrated. Physico-chemical and ADME properties were optimized to improve oral bioavailability in mouse.


Subject(s)
Adenine/analogs & derivatives , Morpholines/chemistry , Protein Kinase Inhibitors/chemistry , Protein Serine-Threonine Kinases/antagonists & inhibitors , Purines/chemistry , Adenine/chemistry , Adenine/pharmacokinetics , Adenine/toxicity , Administration, Oral , Animals , Apoptosis/drug effects , Binding Sites , Crystallography, X-Ray , G2 Phase Cell Cycle Checkpoints/drug effects , HCT116 Cells , Humans , M Phase Cell Cycle Checkpoints/drug effects , Mice , Molecular Conformation , Morpholines/pharmacokinetics , Morpholines/toxicity , Protein Kinase Inhibitors/pharmacokinetics , Protein Kinase Inhibitors/toxicity , Protein Serine-Threonine Kinases/metabolism , Structure-Activity Relationship
5.
J Antibiot (Tokyo) ; 65(5): 237-43, 2012 May.
Article in English | MEDLINE | ID: mdl-22377538

ABSTRACT

Der is an essential and widely conserved GTPase that assists assembly of a large ribosomal subunit in bacteria. Der associates specifically with the 50S subunit in a GTP-dependent manner and the cells depleted of Der accumulate the structurally unstable 50S subunit, which dissociates into an aberrant subunit at a lower Mg(2+) concentration. As Der is an essential and ubiquitous protein in bacteria, it may prove to be an ideal cellular target against which new antibiotics can be developed. In the present study, we describe our attempts to identify novel antibiotics specifically targeting Der GTPase. We performed the structure-based design of Der inhibitors using the X-ray crystal structure of Thermotoga maritima Der (TmDer). Virtual screening of commercially available chemical library retrieved 257 small molecules that potentially inhibit Der GTPase activity. These 257 chemicals were tested for their in vitro effects on TmDer GTPase and in vivo antibacterial activities. We identified three structurally diverse compounds, SBI-34462, -34566 and -34612, that are both biologically active against bacterial cells and putative enzymatic inhibitors of Der GTPase homologs. We also presented the possible interactions of each compound with the Der GTP-binding site to understand the mechanism of inhibition. Therefore, our lead compounds inhibiting Der GTPase provide scaffolds for the development of novel antibiotics against antibiotic-resistant pathogenic bacteria.


Subject(s)
Anti-Bacterial Agents/pharmacology , Drug Design , Enzyme Inhibitors/pharmacology , GTP Phosphohydrolases/antagonists & inhibitors , Thermotoga maritima/enzymology , Anti-Bacterial Agents/chemistry , Binding Sites , Crystallography, X-Ray , Drug Delivery Systems , Drug Resistance, Bacterial , GTP Phosphohydrolases/metabolism , Structure-Activity Relationship , Thermotoga maritima/drug effects
6.
J Med Chem ; 53(24): 8734-46, 2010 Dec 23.
Article in English | MEDLINE | ID: mdl-21080724

ABSTRACT

We have shown previously that the target of the potent cytotoxic agent 4-[(7-bromo-2-methyl-4-oxo-3H-quinazolin-6-yl)methyl-prop-2-ynylamino]-N-(3-pyridylmethyl)benzamide (CB38065, 1) is nicotinamide phosphoribosyltransferase (Nampt). With its cellular target known we sought to optimize the biochemical and cellular Nampt activity of 1 as well as its cytotoxicity. It was found that a 3-pyridylmethylamide substituent in the A region was critical to cellular Nampt activity and cytotoxicity, although other aromatic substitution did yield compounds with submicromolar enzymatic inhibition. Small unsaturated groups worked best in the D-region of the molecule, with 3,3-dimethylallyl providing optimal potency. The E region required a quinazolin-4-one or 1,2,3-benzotriazin-4-one group for activity, and many substituents were tolerated at C² of the quinazolin-4-one. The best compounds showed subnanomolar inhibition of Nampt and low nanomolar cytotoxicity in cellular assays.


Subject(s)
Antineoplastic Agents/chemical synthesis , Benzamides/chemical synthesis , Nicotinamide Phosphoribosyltransferase/antagonists & inhibitors , Quinazolines/chemical synthesis , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacology , Benzamides/chemistry , Benzamides/pharmacology , Drug Screening Assays, Antitumor , HCT116 Cells , Humans , Models, Molecular , Quinazolines/chemistry , Quinazolines/pharmacology , Structure-Activity Relationship
7.
Protein Sci ; 14(7): 1870-8, 2005 Jul.
Article in English | MEDLINE | ID: mdl-15937277

ABSTRACT

We have identified a rare HIV-1 protease (PR) mutation, I47A, associated with a high level of resistance to the protease inhibitor lopinavir (LPV) and with hypersusceptibility to the protease inhibitor saquinavir (SQV). The I47A mutation was found in 99 of 112,198 clinical specimens genotyped after LPV became available in late 2000, but in none of 24,426 clinical samples genotyped from 1998 to October 2000. Phenotypic data obtained for five I47A mutants showed unexpected resistance to LPV (86- to >110-fold) and hypersusceptibility to SQV (0.1- to 0.7-fold). Molecular modeling and energy calculations for these mutants using our structural phenotyping methodology showed an increase in the binding energy of LPV by 1.9-3.1 kcal/mol with respect to the wild type complex, corresponding to a 20- to >100-fold decrease in binding affinity, consistent with the observed high levels of LPV resistance. In the WT PR-LPV complex, the Ile 47 side chain is positioned close to the phenoxyacetyl moiety of LPV and its van der Waals interactions contribute significantly to the ligand binding. These interactions are lost for the smaller Ala 47 residue. Calculated binding energy changes for SQV ranged from -0.4 to -1.2 kcal/mol. In the mutant I47A PR-SQV complexes, the PR flaps are packed more tightly around SQV than in the WT complex, resulting in the formation of additional hydrogen bonds that increase binding affinity of SQV consistent with phenotypic hypersusceptibility. The emergence of mutations at PR residue 47 strongly correlates with increasing prescriptions of LPV (Spearman correlation r(s) = 0.96, P < .0001).


Subject(s)
Drug Resistance, Viral/genetics , HIV Protease Inhibitors/pharmacology , HIV Protease/chemistry , HIV-1/enzymology , Mutation/genetics , Pyrimidinones/pharmacology , Genotype , HIV Protease/genetics , HIV Protease/metabolism , HIV Protease Inhibitors/chemistry , HIV-1/drug effects , HIV-1/genetics , Humans , Kinetics , Lopinavir , Models, Molecular , Molecular Structure , Phenotype , Protein Binding , Protein Conformation , Pyrimidinones/chemistry , Saquinavir/chemistry , Saquinavir/pharmacology , Saquinavir/therapeutic use
8.
Protein Sci ; 12(8): 1706-18, 2003 Aug.
Article in English | MEDLINE | ID: mdl-12876320

ABSTRACT

Mutations in HIV-1 drug targets lead to resistance and consequent therapeutic failure of antiretroviral drugs. Phenotypic resistance assays are time-consuming and costly, and genotypic rules-based interpretations may fail to predict the effects of multiple mutations. We have developed a computational procedure that rapidly evaluates changes in the binding energy of inhibitors to mutant HIV-1 PR variants. Models of WT complexes were produced from crystal structures. Mutant complexes were built by amino acid substitutions in the WT complexes with subsequent energy minimization of the ligand and PR binding site residues. Accuracy of the models was confirmed by comparison with available crystal structures and by prediction of known resistance-related mutations. PR variants from clinical isolates were modeled in complex with six FDA-approved PIs, and changes in the binding energy (DeltaE(bind)) of mutant versus WT complexes were correlated with the ratios of phenotypic 50% inhibitory concentration (IC(50)) values. The calculated DeltaE(bind) of five PIs showed significant correlations (R(2) = 0.7-0.8) with IC(50) ratios from the Virco Antivirogram assay, and the DeltaE(bind) of six PIs showed good correlation (R(2) = 0.76-0.85) with IC(50) ratios from the Virologic PhenoSense assay. DeltaE(bind) cutoffs corresponding to a four-fold increase in IC(50) were used to define the structure-based phenotype as susceptible, resistant, or equivocal. Blind predictions for 78 PR variants gave overall agreement of 92% (kappa = 0.756) and 86% (kappa = 0.666) with PhenoSense and Antivirogram phenotypes, respectively. The structural phenotyping predicted drug resistance of clinical HIV-1 PR variants with an accuracy approaching that of frequently used cell-based phenotypic assays.


Subject(s)
Anti-HIV Agents/pharmacology , Drug Resistance, Viral/genetics , HIV Protease Inhibitors/pharmacology , HIV Protease/chemistry , HIV Protease/metabolism , HIV-1/drug effects , HIV-1/enzymology , Algorithms , Anti-HIV Agents/chemistry , Anti-HIV Agents/metabolism , Drug Design , HIV Protease/genetics , HIV Protease Inhibitors/chemistry , HIV Protease Inhibitors/metabolism , HIV-1/genetics , Models, Molecular , Phenotype , Protein Conformation , Structure-Activity Relationship
9.
Drug Discov Today ; 7(24): 1214-20, 2002 Dec 15.
Article in English | MEDLINE | ID: mdl-12547004

ABSTRACT

Large-scale comparative analysis of drug-target polymorphism structures enables the rational design of next generation 'super drugs'--drugs that are less prone to development of drug resistance or that work for the largest possible fraction of the patient population. Furthermore, knowledge of the drug-target-shape repertoire that exists within the patient population enables predictions of likely clinical trial outcomes and response rates for drug efficacy. This gives information on the optimal drug candidates before the initiation of clinical trials. The economic impact of incorporating pharmacogenomics insights early on in the drug discovery process will be substantial and will afford significant competitive advantages to companies that successfully incorporate this technology.


Subject(s)
Anti-HIV Agents/chemical synthesis , Drug Design , Drug Resistance, Multiple, Viral , Pharmacogenetics/methods , Animals , Drug Resistance, Multiple, Viral/physiology , Humans , Models, Chemical , Models, Molecular
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