ABSTRACT
The design, synthesis, and evaluation of dipeptide analogues as ligands for the pp60c-src SH2 domain are described. The critical binding interactions between Ac-Tyr-Glu-N(n-C5H11)2 (2) and the protein are established and form the basis for our structure-based drug design efforts. The effects of changes in both the C-terminal (11-27) and N-terminal (51-69) portions of the dipeptide are explored. Analogues with reduced overall charge (92-95) are also investigated. We demonstrate the feasibility of pairing structurally diverse subunits in a modest dipeptide framework with the goal of increasing the druglike attributes without sacrificing binding affinity.
Subject(s)
Dipeptides/pharmacology , Enzyme Inhibitors/pharmacology , Proto-Oncogene Proteins pp60(c-src)/antagonists & inhibitors , src Homology Domains , Crystallography, X-Ray , Dipeptides/chemical synthesis , Dipeptides/chemistry , Dipeptides/metabolism , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/chemistry , Enzyme Inhibitors/metabolism , Enzyme-Linked Immunosorbent Assay , Humans , Ligands , Models, Molecular , Molecular Conformation , Proto-Oncogene Proteins pp60(c-src)/metabolism , Structure-Activity RelationshipABSTRACT
A variety of C17 amide-substituted 6-azaandrost-4-en-3-ones were prepared and tested versus human type 1 and 2 steroid 5 alpha-reductase (5AR) and human adrenal 3 beta-hydroxy-delta 5-steroid dehydrogenase/3-keto-delta 5-steroid isomerase (3BHSD) in order to optimize potency versus both isozymes of 5AR and selectivity versus 3BHSD. Two series of potent and selective C17 amides were discovered, 2,5-disubstituted anilides and (arylcycloalkyl)amides. Compounds from each series with picomolar IC50's versus human type 2 5AR and low nanomolar to picomolar IC50's versus human type 1 5AR possessing 100-500-fold selectivity versus 3BHSD were identified. A conformational model to predict 3BHSD potency was developed which could rationalize 3BHSD potency within three different series of compounds. Evaluation of some optimal compounds from this series in a chronic castrated rat model of 5AR inhibitor induced prostate involution, and pharmacokinetic measurements identified compounds (9, 12, 16, and 29) with good in vivo efficacy and half-life in the dog. An intact rat model of in vivo selectivity for 5AR versus 3BHSD inhibition was also developed. Dual inhibitors of both human 5AR's may show advantages over type 2 selective 5AR inhibitors, such as finasteride (1), in the treatment of disease states which depend upon dihydrotestosterone.
Subject(s)
3-Hydroxysteroid Dehydrogenases/antagonists & inhibitors , 5-alpha Reductase Inhibitors , Adrenal Glands/enzymology , Azasteroids/pharmacology , Steroid Isomerases/antagonists & inhibitors , Animals , Azasteroids/chemistry , Azasteroids/pharmacokinetics , Dogs , Humans , Male , Models, Molecular , Rats , Rats, Sprague-Dawley , Structure-Activity RelationshipABSTRACT
6-Azaandrost-4-en-3-ones were synthesized and tested versus human type 1 and 2 steroid 5 alpha-reductase (5AR) and human adrenal 3 beta-hydroxy-delta 5-steroid dehydrogenase/3-keto-delta 5-steroid isomerase (3BHSD) to explore the structure-activity relationship of this novel series in order to optimize potency versus both isozymes of 5AR and selectivity versus 3BHSD. Compounds with picomolar IC50's versus human type 2 5AR and low nanomolar Ki's versus human type 1 5AR with 100-fold selectivity versus 3BHSD were identified (70). Preliminary in vivo evaluation of some optimal compounds from this series in a chronic castrated rat model of 5AR inhibitor-induced prostate involution and dog pharmacokinetic measurements identified a series of 17 beta-[N-(diphenylmethyl)carbamoyl]-6-azaandrost-4-en-3-ones (compounds 54, 66, and 67) with good in vivo efficacy and half-life in the dog. Inhibitors with, at the minimum, low nanomolar potency toward both human 5AR's and selectivity versus 3BHSD may show advantages over previously known 5AR inhibitors in the treatment of disease states which depend upon dihydrotestosterone, such as benign prostatic hyperplasia.
Subject(s)
3-Hydroxysteroid Dehydrogenases/antagonists & inhibitors , 5-alpha Reductase Inhibitors , Adrenal Glands/enzymology , Azasteroids/pharmacology , Isoenzymes/antagonists & inhibitors , Steroid Isomerases/antagonists & inhibitors , Animals , Azasteroids/chemistry , Dogs , Humans , Rats , Rats, Sprague-Dawley , Structure-Activity RelationshipSubject(s)
5-alpha Reductase Inhibitors , Azasteroids/chemical synthesis , Isoenzymes/antagonists & inhibitors , Animals , Azasteroids/pharmacology , Baculoviridae/genetics , Cell Line , Dihydrotestosterone/metabolism , Humans , Male , Moths , Orchiectomy , Rats , Rats, Sprague-Dawley , Recombinant Proteins/antagonists & inhibitors , Testosterone/metabolism , TransfectionABSTRACT
4,4'-Methylenedianiline and its derivatives were assayed for mutagenicity in the Salmonella/microsomal mutagenicity assay develop by Ames. A specificity to revert strain TA98 suggests a mechanism of frameshift mutagenesis. Liver microsomal preparations (S-9) from rats induced with phenobarbital were most effective for metabolic activation. Alkyl substitution of 4,4'-methylenedianiline did not alter its mutagenic activity; however, substitution of both positions ortho to the amino group eliminated mutagenic activity. Substitution with alkoxy-carbonyl groups eliminated mutagenic activity, whereas halogen substitution (chlorine, fluorine) enhanced the mutagenic activity. The results presented here show the use of structure-activity studies as predictive tools for the assessment of genotoxic properties of industrial chemicals.
