ABSTRACT
The structure-based design, synthesis, and biological evaluation of two novel series of potent and selective MEK kinase inhibitors are described herein. The elaboration of a lead pyrrole derivative to a bicyclic dihydroindolone core provided compounds with high potency and good drug-like pharmaceutical properties. Further scaffold modification afforded a series of dihydroindolizinone inhibitors, including an orally available advanced preclinical MEK inhibitor with potent in vivo antitumor efficacy.
Subject(s)
Indolizines/chemical synthesis , MAP Kinase Kinase Kinases/antagonists & inhibitors , Protein Kinase Inhibitors/chemical synthesis , Pyrroles/chemical synthesis , Administration, Oral , Animals , Antineoplastic Agents/administration & dosage , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/therapeutic use , Drug Design , HT29 Cells , Humans , Indolizines/administration & dosage , Indolizines/therapeutic use , MAP Kinase Kinase Kinases/metabolism , Molecular Structure , Protein Kinase Inhibitors/administration & dosage , Protein Kinase Inhibitors/therapeutic use , Pyrroles/administration & dosage , Pyrroles/therapeutic use , Rats , Tumor Burden/drug effects , Xenograft Model Antitumor AssaysABSTRACT
Continuing studies based on dihydroquinoline glucocorticoid receptor agonists lead to the discovery of a series of C4-oxime analogs. Representative compounds exhibited potent transrepression activity with minimal transactivation of phosphoenolpyruvate caboxykinase (PEPCK), a key protein in the gluconeogenesis pathway. These compounds represent promising leads in identifying GR agonists with high anti-inflammatory activity and attenuated potential for glucose elevation.
Subject(s)
Carboxy-Lyases/metabolism , Quinolines/pharmacology , Receptors, Glucocorticoid/agonists , Enzyme Activation , Quinolines/chemistry , Structure-Activity RelationshipABSTRACT
A series of tetrahydroquinoline derivatives were synthesized and profiled for their ability to act as glucocorticoid receptor selective modulators. Structure-activity relationships of the tetrahydroquinoline B-ring lead to the discovery of orally available GR-selective agonists with high in vivo activity.
Subject(s)
Quinolines/pharmacology , Receptors, Glucocorticoid/agonists , Administration, Oral , Animals , Drug Discovery , Enzyme-Linked Immunosorbent Assay , Humans , Quinolines/administration & dosage , Quinolines/chemistry , Rats , Rats, Sprague-Dawley , Structure-Activity RelationshipABSTRACT
We have previously disclosed a series of glucocorticoid receptor (GR) ligands derived from 6-indole-1,2,3,4-tetrahydroquinolines through structure-activity relationship (SAR) of the pendent C6-indole ring. In parallel with this effort, we now report SAR of the tetrahydroquinoline A-ring that identified the importance of a C3 hydroxyl in improving GR selectivity within a series of non-steroidal GR agonists.
Subject(s)
Quinolines/chemistry , Receptors, Glucocorticoid/agonists , Drug Evaluation, Preclinical , Protein Binding , Quinolines/chemical synthesis , Quinolines/pharmacology , Receptors, Glucocorticoid/metabolism , Structure-Activity RelationshipABSTRACT
A novel series of pyrrole inhibitors of MEK kinase has been developed using structure-based drug design. Optimization of the series led to the identification of potent inhibitors with good pharmaceutical properties.
Subject(s)
MAP Kinase Kinase Kinases/antagonists & inhibitors , Protein Kinase Inhibitors/chemistry , Pyrroles/chemistry , Animals , Crystallography, X-Ray , Drug Design , Models, Molecular , Molecular Structure , Protein Kinase Inhibitors/chemical synthesis , Protein Kinase Inhibitors/pharmacology , Pyrroles/chemical synthesis , Pyrroles/pharmacology , RatsABSTRACT
A series of nonsteroidal glucocorticoid receptor (GR) ligands based on a 6-indole-1,2,3,4-tetrahydroquinoline scaffold are reported. Structure-activity relationship (SAR) of the pendent indole group identified compound 20 exhibiting good GR binding affinity (K(i)=1.5nM) and 100- to 1000-fold selectivity over MR, PR, and AR while showing activity in an E-selectin repression assay.
Subject(s)
Anti-Inflammatory Agents, Non-Steroidal/pharmacology , Indoles/pharmacology , Quinolines/pharmacology , Receptors, Glucocorticoid/drug effects , Anti-Inflammatory Agents, Non-Steroidal/chemical synthesis , Anti-Inflammatory Agents, Non-Steroidal/chemistry , Binding Sites , Drug Evaluation, Preclinical , Indoles/chemical synthesis , Indoles/chemistry , Ligands , Molecular Structure , Quinolines/chemical synthesis , Quinolines/chemistry , Stereoisomerism , Structure-Activity RelationshipABSTRACT
Structure-activity relationship studies centered around 3'-substituted (Z)-5-(2'-(thienylmethylidene))1,2-dihydro-9-hydroxy-10-methoxy-2,2,4-trimethyl-5H-chromeno[3,4-f]quinolines are described. A series of highly potent and efficacious selective glucocorticoid receptor modulators were identified with in vitro activity comparable to dexamethasone. In vivo evaluation of these compounds utilizing a 28 day mouse tumor xenograft model demonstrated efficacy equal to dexamethasone in the reduction of tumor volume.
Subject(s)
Antineoplastic Agents/chemical synthesis , Benzopyrans/chemical synthesis , Multiple Myeloma/drug therapy , Quinolines/chemical synthesis , Receptors, Glucocorticoid/drug effects , Animals , Antineoplastic Agents/chemistry , Antineoplastic Agents/pharmacology , Benzopyrans/chemistry , Benzopyrans/pharmacology , Binding, Competitive , Dexamethasone/pharmacology , Humans , Mice , Mineralocorticoid Receptor Antagonists , Models, Molecular , Multiple Myeloma/pathology , Quinolines/chemistry , Quinolines/pharmacology , Receptors, Glucocorticoid/agonists , Receptors, Glucocorticoid/antagonists & inhibitors , Receptors, Mineralocorticoid/agonists , Stereoisomerism , Structure-Activity Relationship , Xenograft Model Antitumor AssaysABSTRACT
A series of androgen receptor modulators based on 8H-[1,4]oxazino[2,3-f]quinolin-8-ones was synthesized and evaluated in an androgen receptor transcriptional activation assay. The most potent analogues from the series exhibited single-digit nanomolar potency in vitro. Compound 18h demonstrated full efficacy in the maintenance of muscle weight, at 10 mg/kg, with reduced activity in prostate weight in an in vivo model of androgen action.
Subject(s)
Oxazines/chemical synthesis , Oxazines/pharmacology , Quinolones/chemical synthesis , Quinolones/pharmacology , Receptors, Androgen/drug effects , Animals , Cell Line , Dose-Response Relationship, Drug , Humans , Indicators and Reagents , Male , Models, Molecular , Orchiectomy , Rats , Receptors, Androgen/chemistry , Receptors, Progesterone/chemistry , Receptors, Progesterone/drug effects , Receptors, Somatotropin/chemistry , Receptors, Somatotropin/drug effects , Structure-Activity Relationship , Testosterone/bloodABSTRACT
Members of the large aggregating chondroitin sulfate proteoglycans are characterized by an N-terminal fragment known as G1 domain, which is composed of an immunoglobulin (IgG)-like motif and two tandem repeats (TR). Previous studies have indicated that the expressed product of aggrecan G1 domain was not secreted. Here we demonstrated that the inability of G1 secretion was associated with the tandem repeats but not the IgG-like motif, and specifically with TR1 of aggrecan. We also demonstrated that the G2 domain, a domain unique to aggrecan, had a similar effect on product secretion. The sequence of TR1 of G1 is highly conserved across species, which suggested similar functions played by these motifs. In a yeast two-hybrid assay, TR1 interacted with the calcium homeostasis endoplasmic reticulum protein. Deletion/mutation experiments indicated that the N-terminal fragment of TR1, in particular, the amino acids H(2)R(4) of this motif were key to its effect on product secretion. However, the N-terminal 55 amino acids were required to exert this function. Taken together, our study suggests a possible molecular mechanism for the function of the tandem repeats in product processing.
Subject(s)
Extracellular Matrix Proteins , Proteoglycans/metabolism , Aggrecans , Amino Acid Motifs , Animals , Base Sequence , COS Cells , Cattle , Chickens , Chondroitin Sulfate Proteoglycans/chemistry , Chondroitin Sulfate Proteoglycans/genetics , Chondroitin Sulfate Proteoglycans/metabolism , Endoplasmic Reticulum/chemistry , Humans , Lectins, C-Type , Membrane Proteins/metabolism , Mice , Mutagenesis/genetics , Protein Structure, Tertiary , Proteoglycans/genetics , Rats , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Repetitive Sequences, Amino Acid/physiology , Restriction Mapping , Sequence Homology, Amino Acid , Transfection , Two-Hybrid System Techniques , Yeasts/geneticsABSTRACT
OBJECTIVE: Accurate assessment of chondrocyte metabolism is a potentially valuable indicator of cartilage health in osteoarthritis (OA). This study was conducted to explore the anabolic metabolism of chondrocytes early in the natural history of an experimental canine model of OA. METHODS: Competitive reverse transcription-polymerase chain reaction was used to calculate the copy number of type II collagen and the messenger RNA (mRNA) levels of aggrecan core protein in articular cartilage samples obtained from different regions of the femorotibial joint 12 and 39 weeks after cruciate transection. RESULTS: Gene expression of both type II collagen and aggrecan in cartilage samples obtained from experimental joints at both intervals after surgery was elevated significantly compared with that in samples from contralateral control joints. The number of mRNA copies per microgram of DNA of aggrecan exceeded that of type II collagen in control cartilage, but the copy number of type II collagen mRNA exceeded that of aggrecan in OA cartilage. Thus, the ratio of type II collagen-to-aggrecan mRNA copy number (normalized to DNA) was shown to be characteristically altered in cartilage with experimental OA. CONCLUSION: Accurate assessment of multiple gene products in small samples of cartilage taken from focal areas of a joint can be used diagnostically for monitoring chondrocyte metabolism and possibly for staging at least the early phases of this joint disease.
Subject(s)
Cartilage/pathology , Collagen Type II/genetics , Extracellular Matrix Proteins , Osteoarthritis/pathology , Osteoarthritis/physiopathology , Proteoglycans/genetics , Aggrecans , Animals , Chondrocytes/pathology , Chondrocytes/physiology , Dogs , Female , Gene Expression/physiology , Joints/pathology , Lectins, C-Type , Male , RNA, Messenger/analysis , Reverse Transcriptase Polymerase Chain ReactionABSTRACT
Aggrecan, a major structural proteoglycan in cartilage, contains three globular domains, G1, G2, and G3, as well as sequences for glycosaminoglycan modification. A large number of proteases are implicated in aggrecan cleavage in normal metabolism, aging, and arthritis. These proteases are known to cleave at the IGD, KS, and CS domains. Here we report for the first time evidence of cleavage at a novel site, the carboxyl tail of aggrecan. Results from deletion mutants of the tail indicated that the likely cleavage sites were two consensus sequences, RRLXK and RSPR, present in the aggrecan analogs of many species. This was confirmed by site-directed mutagenesis. A construct containing two G3 domains (G3G3) was also found to cleave between the G3 duplicates. When G3 tail was linked to a glycosaminoglycan-modifying sequence, it was protected from cleavage. Furin inhibitor also reduced the levels of tail cleavage. The carboxyl tails of chicken and human versican were not cleaved, despite the presence of the consensus sequence. Our studies indicate that the basic amino acids present in the tail play an important role in cleavage, and this mechanism is specific to aggrecan.
Subject(s)
Chondroitin Sulfate Proteoglycans/chemistry , Extracellular Matrix Proteins , Proteoglycans/chemistry , Proteoglycans/genetics , Aggrecans , Amino Acid Sequence , Amino Acids/chemistry , Animals , Binding Sites , COS Cells , Cattle , Chickens , Chondrocytes/metabolism , Cloning, Molecular , Densitometry , Furin , Gene Deletion , Histidine/chemistry , Humans , Lectins, C-Type , Molecular Sequence Data , Mutagenesis, Site-Directed , Peptides/chemistry , Protein Structure, Tertiary , Recombinant Proteins/metabolism , Sequence Homology, Amino Acid , Subtilisins/antagonists & inhibitors , Time Factors , Transfection , VersicansABSTRACT
Aggrecan is the major proteoglycan in the extracellular matrix of cartilage. A notable exception is nanomelic cartilage, which lacks aggrecan in its matrix. The example of nanomelia and other evidence leads us to believe that the G3 domain plays an important role in aggrecan processing, and it has indeed been confirmed that G3 allows glycosaminoglycan (GAG) chain attachment and product secretion. However, it is not clear how G3, which contains at least a carbohydrate recognition domain (CRD) and a complement binding protein (CBP) motif, plays these two functional roles. The present study was designed to dissect the mechanisms of this phenomenon and specially 1) to determine the effects of various cysteine residues in GAG modification and product secretion as well as 2) to investigate which of the two processing events is the critical step in the product processing. Our studies demonstrated that removal of the two amino-terminal cysteines in the CRD motif and the single cysteine in the amino terminus of CBP inhibited secretion of CRD and CBP. Use of the double mutant CRD construct also allowed us to observe a deviation from the usual strict coupling of GAG modification and product secretion steps. The presence of a small chondroitin sulfate fragment overcame the secretion-inhibitory effects once the small chondroitin sulfate fragment was modified by GAG.