ABSTRACT
A novel series of 2-amino-1,3,5-triazines bearing a tricyclic moiety as heat shock protein 90 (Hsp90) inhibitors is described. Molecular design was performed using X-ray cocrystal structures of the lead compound CH5015765 and natural Hsp90 inhibitor geldanamycin with Hsp90. We optimized affinity to Hsp90, in vitro cell growth inhibitory activity, water solubility, and liver microsomal stability of inhibitors and identified CH5138303. This compound showed high binding affinity for N-terminal Hsp90α (Kd=0.52nM) and strong in vitro cell growth inhibition against human cancer cell lines (HCT116 IC50=0.098µM, NCI-N87 IC50=0.066µM) and also displayed high oral bioavailability in mice (F=44.0%) and potent antitumor efficacy in a human NCI-N87 gastric cancer xenograft model (tumor growth inhibition=136%).
Subject(s)
Benzopyrans/pharmacology , Drug Design , HSP90 Heat-Shock Proteins/antagonists & inhibitors , Neoplasms, Experimental/drug therapy , Triazines/pharmacology , Administration, Oral , Animals , Benzopyrans/administration & dosage , Benzopyrans/chemical synthesis , Biological Availability , Cell Line, Tumor , Cell Proliferation/drug effects , Crystallography, X-Ray , Dose-Response Relationship, Drug , Drug Screening Assays, Antitumor , Female , HCT116 Cells , Humans , Mice , Mice, Inbred BALB C , Mice, Nude , Models, Molecular , Molecular Structure , Structure-Activity Relationship , Triazines/administration & dosage , Triazines/chemical synthesisABSTRACT
Macrocyclic compounds bearing a 2-amino-6-arylpyrimidine moiety were identified as potent heat shock protein 90 (Hsp90) inhibitors by modification of 2-amino-6-aryltriazine derivative (CH5015765). We employed a macrocyclic structure as a skeleton of new inhibitors to mimic the geldanamycin-Hsp90 interactions. Among the identified inhibitors, CH5164840 showed high binding affinity for N-terminal Hsp90α (K(d)=0.52nM) and strong anti-proliferative activity against human cancer cell lines (HCT116 IC(50)=0.15µM, NCI-N87 IC(50)=0.066µM). CH5164840 displayed high oral bioavailability in mice (F=70.8%) and potent antitumor efficacy in a HCT116 human colorectal cancer xenograft model (tumor growth inhibition=83%).
Subject(s)
Antineoplastic Agents/pharmacology , Drug Design , HSP90 Heat-Shock Proteins/antagonists & inhibitors , Macrocyclic Compounds/pharmacology , Pyrimidines/pharmacology , Animals , Antineoplastic Agents/administration & dosage , Antineoplastic Agents/chemical synthesis , Cell Proliferation/drug effects , Crystallography, X-Ray , Dose-Response Relationship, Drug , Humans , Macrocyclic Compounds/administration & dosage , Macrocyclic Compounds/chemical synthesis , Mice , Mice, Nude , Mice, SCID , Models, Molecular , Molecular Structure , Pyrimidines/administration & dosage , Pyrimidines/chemical synthesis , Stereoisomerism , Structure-Activity Relationship , Xenograft Model Antitumor AssaysABSTRACT
Heat shock protein 90 (Hsp90) is a molecular chaperone which regulates maturation and stabilization of its substrate proteins, known as client proteins. Many client proteins of Hsp90 are involved in tumor progression and survival and therefore Hsp90 can be a good target for developing anticancer drugs. With the aim of efficiently identifying a new class of orally available inhibitors of the ATP binding site of this protein, we conducted fragment screening and virtual screening in parallel against Hsp90. This approach quickly identified 2-aminotriazine and 2-aminopyrimidine derivatives as specific ligands to Hsp90 with high ligand efficiency. In silico evaluation of the 3D X-ray Hsp90 complex structures of the identified hits allowed us to promptly design CH5015765, which showed high affinity for Hsp90 and antitumor activity in human cancer xenograft mouse models.
Subject(s)
Antineoplastic Agents/chemical synthesis , Benzopyrans/chemistry , Benzopyrans/chemical synthesis , Computer Simulation , Drug Design , Drug Discovery/methods , HSP90 Heat-Shock Proteins/antagonists & inhibitors , Triazines/chemistry , Triazines/chemical synthesis , Adenosine Triphosphatases/metabolism , Administration, Oral , Animals , Antineoplastic Agents/chemistry , Antineoplastic Agents/metabolism , Antineoplastic Agents/pharmacology , Benzopyrans/metabolism , Benzopyrans/pharmacokinetics , Dose-Response Relationship, Drug , Escherichia coli/genetics , HSP90 Heat-Shock Proteins/chemistry , HSP90 Heat-Shock Proteins/metabolism , Humans , Hydrophobic and Hydrophilic Interactions , Mice , Neoplasms/drug therapy , Reproducibility of Results , Structure-Activity Relationship , Surface Plasmon Resonance , Triazines/metabolism , Triazines/pharmacokinetics , Xenograft Model Antitumor AssaysABSTRACT
A series of benzofuran-based farnesyltransferase inhibitors have been designed and synthesized as antitumor agents. Among them, 11f showed the most potent enzyme inhibitory activity (IC(50)=1.1nM) and antitumor activity in human cancer xenografts in mice.
Subject(s)
Antineoplastic Agents/chemical synthesis , Benzofurans/chemical synthesis , Chemistry, Pharmaceutical/methods , Farnesyltranstransferase/antagonists & inhibitors , Quinolones/chemical synthesis , Animals , Antineoplastic Agents/pharmacology , Benzofurans/pharmacology , Crystallography, X-Ray , Drug Design , Drug Screening Assays, Antitumor , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/pharmacology , Farnesyltranstransferase/chemistry , Humans , Inhibitory Concentration 50 , Mice , Neoplasm Transplantation , Quinolones/pharmacology , Structure-Activity RelationshipABSTRACT
The C-4 side chain modification of lead compound 1 has resulted in the identification of a potent and selective Candida albicans N-myristoyltransferase (CaNmt) inhibitor RO-09-4609, which exhibits antifungal activity against C. albicans in vitro. Further modification of its C-2 substituent has led to the discovery of RO-09-4879, which exhibits antifungal activity in vivo. The drug design is based on X-ray crystal analysis of a CaNmt complex with benzofuran derivative 4a. The optimization incorporates various biological investigations including a quasi in vivo assay and pharmacokinetic study. The computer aided drug design, synthesis, structure-activity relationships, and biological properties of RO-09-4879 are described in detail.
Subject(s)
Acyltransferases/antagonists & inhibitors , Antifungal Agents/chemical synthesis , Benzofurans/chemical synthesis , Animals , Antifungal Agents/pharmacokinetics , Antifungal Agents/pharmacology , Benzofurans/pharmacokinetics , Benzofurans/pharmacology , Candida albicans/drug effects , Candida albicans/enzymology , Candidiasis/drug therapy , Drug Design , Drug Resistance, Fungal/genetics , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/pharmacokinetics , Enzyme Inhibitors/pharmacology , Fungal Proteins/antagonists & inhibitors , Inhibitory Concentration 50 , Male , Protein Binding , Rats , Rats, Inbred F344 , Structure-Activity RelationshipABSTRACT
The (6R)-2,2,6-trimethyl-1,4-cyclohexanedione (levodione) reductase (LVR) of the soil isolate bacterium Corynebacterium aquaticum M-13 is a NAD(H)-linked enzyme that catalyzes reversible oxidoreduction between (4R)-hydroxy-(6R)-2,2,6-trimethylcyclohexanone (actinol) and levodione. Here the crystal structure of a ternary complex of LVR with NADH and its inhibitor 2-methyl-2,4-pentanediol has been determined by molecular replacement and refined at 1.6-A resolution with a crystallographic R factor of 0.199. The overall structure is similar to those of other short-chain alcohol dehydrogenase/reductase enzymes. The positions of NADH and 2-methyl-2,4-pentanediol indicate the binding site of the substrate and identify residues that are likely to be important in the catalytic reaction. Modeling of the substrate binding in the active site suggests that the specificity of LVR is determined by electrostatic interactions between the negatively charged surface of Glu-103 of LVR and the positively charged surface on the re side of levodione. Mutant LVR enzymes in which Glu-103 is substituted with alanine (E103A), glutamine (E103Q), asparagines (E103N), or aspartic acid (E103D) show a 2-6-fold increase in Km values as compared with wild-type LVR and a much lower enantiomeric excess of the reaction products (60%) than the wild-type enzyme (95%). Together, these data indicate that Glu-103 has an important role in determining the stereospecificity of LVR.
Subject(s)
Corynebacterium/enzymology , Oxidoreductases/chemistry , Oxidoreductases/metabolism , Binding Sites , Catalysis , Circular Dichroism , Crystallization , Crystallography , Enzyme Inhibitors/metabolism , Escherichia coli/genetics , Gene Expression , Glutamic Acid , Glycols/metabolism , Hydrogen Bonding , Models, Molecular , Molecular Structure , Mutation , NAD/metabolism , Oxidoreductases/genetics , Protein Structure, Secondary , Recombinant Proteins/chemistry , Static Electricity , Stereoisomerism , Structure-Activity Relationship , Substrate SpecificityABSTRACT
A new series of acid-stable antifungal agents having strong inhibitory activity against Candida albicans N-myristoyltransferase (CaNmt) has been developed starting from acid-unstable benzofuranylmethyl aryl ether 2. The inhibitor design is based on X-ray crystallographic analysis of a CaNmt complex with aryl ether 3. Among the new inhibitors, pyridine derivative 8b and benzimidazole derivative 8k showed clear antifungal activity in a murine systemic candidiasis model.
Subject(s)
Acyltransferases/antagonists & inhibitors , Antifungal Agents/chemical synthesis , Benzofurans/chemical synthesis , Fungal Proteins/antagonists & inhibitors , Animals , Antifungal Agents/pharmacokinetics , Antifungal Agents/pharmacology , Benzofurans/pharmacokinetics , Benzofurans/pharmacology , Candida albicans/drug effects , Candida albicans/enzymology , Disease Models, Animal , Drug Design , Drug Stability , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/pharmacokinetics , Enzyme Inhibitors/pharmacology , Inhibitory Concentration 50 , Mice , Models, Molecular , Structure-Activity RelationshipABSTRACT
A highly potent water soluble triazole antifungal prodrug, RO0098557 (1), has been identified from its parent, the novel antifungal agent RO0094815 (2). The prodrug includes a triazolium salt linked to an aminocarboxyl moiety, which undergoes enzymatic activation followed by spontaneous chemical degradation to release 2. Prodrug 1 showed high chemical stability and water solubility and exhibited strong antifungal activity against systemic candidiasis and aspergillosis as well as pulmonary aspergillosis in rats.
Subject(s)
Antifungal Agents/chemical synthesis , Antifungal Agents/pharmacology , Prodrugs/chemical synthesis , Prodrugs/pharmacology , Triazoles/chemical synthesis , Triazoles/pharmacology , Animals , Antifungal Agents/pharmacokinetics , Aspergillosis/drug therapy , Aspergillosis/microbiology , Biotransformation , Candidiasis/drug therapy , Candidiasis/microbiology , Chemical Phenomena , Chemistry, Physical , Drug Design , Half-Life , Haplorhini , Humans , Hydrogen-Ion Concentration , In Vitro Techniques , Models, Molecular , Molecular Conformation , Prodrugs/pharmacokinetics , Rats , Solubility , Solvents , Triazoles/pharmacokinetics , WaterABSTRACT
Myristoyl-CoA:protein N-myristoyltransferase (Nmt) is a monomeric enzyme that catalyzes the transfer of the fatty acid myristate from myristoyl-CoA to the N-terminal glycine residue of a variety of eukaryotic and viral proteins. Genetic and biochemical studies have established that Nmt is an attractive target for antifungal drugs. We present here crystal structures of C. albicans Nmt complexed with two classes of inhibitor competitive for peptide substrates. One is a peptidic inhibitor designed from the peptide substrate; the other is a nonpeptidic inhibitor having a benzofuran core. Both inhibitors are bound into the same binding groove, generated by some structural rearrangements of the enzyme, with the peptidic inhibitor showing a substrate-like binding mode and the nonpeptidic inhibitor binding differently. Further, site-directed mutagenesis for C. albicans Nmt has been utilized in order to define explicitly which amino acids are critical for inhibitor binding. The results suggest that the enzyme has some degree of flexibility for substrate binding and provide valuable information for inhibitor design.
Subject(s)
Acyltransferases/antagonists & inhibitors , Acyltransferases/chemistry , Candida albicans/enzymology , Enzyme Inhibitors/chemistry , Acyltransferases/metabolism , Amino Acids/chemistry , Amino Acids/metabolism , Binding Sites , Crystallography, X-Ray , Enzyme Inhibitors/metabolism , Imidazoles/metabolism , Models, Molecular , Mutagenesis, Site-Directed , Oligopeptides/metabolism , Protein Conformation , Substrate SpecificityABSTRACT
Pharmacokinetic (PK) parameters of N-myristoyltransferase (Nmt) inhibitors were measured, and a multivariate quantitative structure-pharmacokinetic relationship (QSPKR) model for predicting rat elimination half-life (t(1/2)) values was constructed. One hundred seven benzofuran derivatives have been selected as the data set for QSPKR analysis. The correlation between the t(1/2) values and 30 physicochemical descriptors was examined by a stepwise multiple linear regression method. The statistical analysis gives a significant QSPKR model (r = 0.843) with the following three variables: partial negative surface area (PNSA), atomic-based octanol/water partition coefficient (AlogP), and the number of rotational bonds (Rotlbonds). The QSPKR model obtained is predictive and simple, and would give a direction for designing new Nmt inhibitors having good PK profiles.
Subject(s)
Acyltransferases/antagonists & inhibitors , Antifungal Agents/chemistry , Antifungal Agents/pharmacokinetics , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/pharmacokinetics , Animals , Benzofurans/chemistry , Benzofurans/pharmacokinetics , Computer Simulation , Drug Design , Male , Models, Chemical , Quantitative Structure-Activity Relationship , Rats , Rats, Inbred F344ABSTRACT
Modification of the C-2 position of a benzofuran derivative 6 (RO-09-4609), an N-myristoyltransferase (Nmt) inhibitor, has led us to discover antifungal agents that are active in a murine systemic candidiasis model. The drug design is based on the analysis of a crystal structure of a Candida Nmt complex with 2. The optimization has been guided by various biological evaluations including a quasi in vivo assay and pharmacokinetic analysis.