Synthesis and structure-activity relationship for a novel class of potent and selective carbamoyl-triazole based inhibitors of hormone sensitive lipase

The central role of the intracellular enzyme hormone-sensitive lipase (HSL) in regulating fatty acid metabolism makes it an interesting pharmacological target for the treatment of insulin resistant and dyslipidemic disorders where a decrease in delivery of fatty acids to the circulation is desirable, e.g., in individuals with type 2 diabetes, metabolic syndrome, or impaired glucose tolerance. On the basis of a lead structure from high throughput screening, we have identified a very potent type of carbamoyl-triazole inhibitors of HSL. As part of the lead optimization program, four new classes of carbamoyl-triazoles were synthesized and tested with respect to potency, efficacy and selectivity. Methyl-phenyl-carbamoyl-triazoles were identified as potent and efficacious HSL inhibitors. These compounds do not inhibit other hydrolases such as hepatic lipase, lipoprotein lipase, pancreatic lipase, and butyrylcholine esterase. However, the inhibitors 4b and 4g with IC(50) values for HSL of 0.17 and 0.25 microM, respectively, were the only inhibitors selective against acetylcholine esterase. A reversible pseudosubstrate inhibition mechanism is proposed for this class of inhibitors.     

The first potent inhibitors for human glutaminyl cyclase: synthesis and structure-activity relationship

The first effective inhibitors for human glutaminyl cyclase (QC) are described. The structures are developed by applying a ligand-based optimization approach starting from imidazole. Screening of derivatives of that heterocycle led to compounds of the imidazol-1-yl-alkyl thiourea type as a lead scaffold. A library of thiourea derivatives was synthesized, resulting in an inhibitory improvement by 2 orders of magnitude, leading to 1-(3-(1H-imidazol-1-yl)propyl)-3-(3,4-dimethoxyphenyl)thiourea as a potent inhibitor. Systematic exploitation of the scaffold revealed a strong impact on the inhibitory efficacy and resulted in the development of imidazole-propyl-thioamides as another new class of potent inhibitors. A flexible alignment of the most potent compounds of the thioamide and thiourea class and a QC substrate revealed a good match of characteristic features of the molecules, which suggests a similar binding mode of both inhibitors and the substrate to the active site of QC.     

Design,synthesis, and structure-activity relationship of 6-alkynylpyrimidines as potent adenosine kinase inhibitors

Adenosine (ADO) is an extracellular signaling molecule within the central and peripheral nervous system. Its concentration is increased at sites of tissue injury and inflammation. One of the mechanisms by which antinociceptive and antiinflammatory effects of ADO can be enhanced consists of inhibition of adenosine kinase (AK), the primary metabolic enzyme for ADO. Novel nonnucleoside AK inhibitors based on 4-amino-6-alkynylpyrimidines were prepared, and the importance of the length of the linker at the 5-position for high affinity AK inhibition was demonstrated. Compounds with 2- and 3-atom linkers were the most potent AK inhibitors. Optimization of their physicochemical properties led to 31a and 37a that effectively reduced pain and inflammation in animal models.     

Design and structure-activity relationship of a new class of potent VEGF receptor tyrosine kinase inhibitors

A series of substituted 4-anilinoquinazolines and related compounds were synthesized as potential inhibitors of vascular endothelial growth factor (VEGF) receptor (Flt and KDR) tyrosine kinase activity. Enzyme screening indicated that a narrow structure-activity relationship (SAR) existed for the bicyclic ring system, with quinazolines, quinolines, and cinnolines having activity and with quinazolines and quinolines generally being preferred. Substitution of the aniline was investigated and clearly indicated that small lipophilic substituents such as halogens or methyl were preferred at the C-4' position. Small substituents such as hydrogen and fluorine are preferred at the C-2' position. Introduction of a hydroxyl group at the meta position of the aniline produced the most potent inhibitors of Flt and KDR tyrosine kinases activity with IC(50) values in the nanomolar range (e.g. 10, 12, 13, 16, and 18). Investigation of the quinazoline C-6 and C-7 positions indicates that a large range of substituents are tolerated at C-7, whereas variation at the C-6 is more restricted. At C-7, neutral, basic, and heteroaromatic side chains led to very potent compounds, as illustrated by the methoxyethoxy derivative 13 (IC(50) < 2 nM). Our inhibitors proved to be very selective inhibitors of Flt and KDR tyrosine kinase activity when compared to that associated with the FGF receptor (50- to 3800-fold). Observed enzyme profiles translated well with respect to potency and selectivity for inhibition of growth factor stimulated proliferation of human umbilical vein endothelial cells (HUVECs). Oral administration of selected compounds to mice produced total plasma levels 6 h after dosing of between 3 and 49 microM. In vivo efficacy was demonstrated in a rat uterine oedema assay where significant activity was achieved at 60 mg/kg with the meta hydroxy anilinoquinazoline 10. Inhibition of growth of human tumors in athymic mice has also been demonstrated: compound 34 inhibited the growth of established Calu-6 lung carcinoma xenograft by 75% (P < 0.001, one tailed t-test) following daily oral administration of 100 mg/kg for 21 days.     

Synthesis and structure-activity relationship for a novel class of potent and selective carbamate-based inhibitors of hormone selective lipase with acute in vivo antilipolytic effects

Hormone-sensitive lipase (HSL) is an intracellular enzyme that has a central role in the regulation of fatty acid metabolism. The enzyme, therefore, is a potentially interesting pharmacological target for the treatment of insulin resistance and dyslipidemic disorders. Based on a high throughput screening, a carbamate based HSL inhibitor was identified and optimized into the selective HSL inhibitors 4-hydroxymethyl-piperidine-1-carboxylic acid 4-(5-trifluoromethylpyridin-2-yloxy)-phenyl ester (13f) and 4-hydroxy-piperidine-1-carboxylic acid 4-(5-trifluoromethylpyridin-2-yloxy)-phenyl ester (13g), with IC50 values of 110 and 500 nM, respectively. Both inhibitors were active in acute antilipolytic experiments in vivo and none of the inhibitors inhibited the cytochrome P450 (CYP) isoforms 2D6, 3A4, and 1A2.     

Design, synthesis, and structure-activity relationship studies of highly potent novel benzoxazinyl-oxazolidinone antibacterial agents

A series of novel benzoxazinyl-oxazolidinones bearing nonaromatic heterocycle or aryl groups were designed and synthesized. Their in vitro and in vivo antibacterial activities were investigated. Most of the (3S, 3aS) biaryl benzoxazinyl-oxazolidinones exhibited potent activity against Gram-positive pathogens. SAR trends were observed; a pyridyl C ring was preferable to other 5- or 6-member aryl C rings, while fluorine substitution on the B ring generated derivatives with reduced activity. Various substituent group positions on the pyridyl ring were also evaluated. The resulting compounds displayed excellent activity against linezolid-resistant strains. Compound 45 exhibited excellent in vitro activity, with a MIC value of 0.25-0.5 μg/mL against MRSA and an activity against linezolid-resistant strains of 8-16-fold higher potency than linezolid. In a MRSA systemic infection model, compound 45 displayed an ED(50) < 5.0 mg/kg, a potency that is nearly 3-fold better than that of linezolid. This compound also showed excellent pharmacokinetic profiles, with a half-life of more than 5 h as well as an oral bioavailability of 81% in rats.     

Design, synthesis, and 3D QSAR of novel potent and selective aromatase inhibitors

The design, synthesis, and biological evaluation of a series of new aromatase inhibitors bearing an imidazole or triazole ring linked to a fluorene (A), indenodiazine (B), or coumarin scaffold (C) are reported. Properly substituted coumarin derivatives displayed the highest aromatase inhibitory potency and selectivity over 17-alpha-hydroxylase/17-20 lyase. The modeling of the aromatase inhibition data by Comparative Molecular Field Analysis (CoMFA/GOLPE 3D QSAR approach) led to the development of a PLS model with good fitting and predictive powers (n = 22, ONC = 3, r(2) = 0.949, s = 0.216, and q(2) = 0.715). The relationship between aromatase inhibition and the steric and electrostatic fields generated by the examined azole inhibitors enables a clear understanding of the nature and spatial location of the main interactions modulating the aromatase inhibitory potency.     

Synthesis and structure-activity relationship of novel, highly potent metharyl and methcycloalkyl cyclooxygenase-2 (COX-2) selective inhibitors

A novel series of benzo-1,3-dioxolane metharyl derivatives was synthesized and evaluated for cyclooxygenase-2 (COX-2) and cyclooxygenase-1 (COX-1) inhibition in human whole blood (HWB). In the present study, structure-activity relationships (SAR) in the metharyl analogues were investigated. The spacer group and substitutions in the spacer group were found to be quite important for potent COX-2 inhibition. Compounds in which a methylene group (8a-c), carbonyl group (12a-c), or methylidene group (7a-c) connected cycloalkyl groups to the central benzo-1,3-dioxolane template were found to be potent and selective COX-2 inhibitors. Aryl-substituted compounds linked to the central ring by either a methylene or a carbonyl spacer resulted in potent, highly selective COX-2 inhibitors. In this series of substituted-(2H-benzo[3,4-d]1,3-dioxolan-5-yl))-1-(methylsulfonyl)benzene compounds, SAR studies demonstrated that substitution at the 3-position of the aryl group optimized COX-2 selectivity and potency, whereas substitution at the 4-position attenuated COX-2 inhibition. Mono- or difluoro substitution at meta position(s), as in 22c and 22h, was advantageous for both in vitro COX-2 potency and selectivity (e.g., COX-2 IC(50) for 22c = 1 microM and COX-1 IC(50) for 22c = 20 microM in HWB assay). Several novel compounds in the (2H-benzo[3,4-d]1,3-dioxolan-5-yl))-1-(methylsulfonyl)benzene series, as shown in structures 7c, 8a, 12a, 21c, 22c, 22e, and 22h, selectively inhibited COX-2 activity by 40-50% at a test concentration of 1 microM in an in vitro HWB assay.     

Design, synthesis, and structure-activity relationship studies of 3,4,6-triphenylpyran-2-ones as selective cyclooxygenase-2 inhibitors

A group of regioisomeric 3,4,6-triphenylpyran-2-ones with a MeSO(2) pharmacophore at the para-position of either a C-3 phenyl or a C-4 phenyl substituent on the central six-membered pyran-2-one ring were prepared and evaluated in vitro for their abilities to inhibit the isozymes COX-1 and COX-2. Structure-activity relationship (SAR) data, acquired by substituent modification at the para-position of the C-6 phenyl ring attached to the central pyranone, showed that 6-(4-methoxyphenyl)-3-(4-methanesulfonylphenyl)-4-phenylpyran-2-one (12e) was the most potent and selective COX-2 inhibitor (COX-2 IC(50) = 0.02 microM; COX-1 IC(50) > 100 microM) with a high COX-2 selectivity index (SI > 5000) relative to the reference drugs celecoxib (COX-2 IC(50) = 0.07 microM; SI = 474) and rofecoxib (COX-2 IC(50) = 0.50 microM; SI > 200). 6-(4-Methoxyphenyl)-3-(4-methanesulfonylphenyl)-4-phenylpyran-2-one (12e) was a more potent oral antiinflammatory agent (ID(50) = 5.6 mg/kg) than celecoxib (ID(50) = 10.8 mg/kg) in a carrageenan-induced rat paw edema assay. In a 4% NaCl-induced abdominal constriction assay, a 5 mg/kg oral dose of 12e exhibited good analgesic activity at different time intervals producing 37.5 and 69% inhibition of writhing at 30 and 60 min, respectively. In contrast, the corresponding 6-(4-methoxyphenyl)-4-(4-methanesulfonylphenyl)-3-phenylpyran-2-one regiosiomer (12o) was a less potent and selective COX-2 inhibitor (COX-2 IC(50) = 0.45 microM; SI = 70). A molecular modeling study for 12e indicated that the p-OMe substituent on the C-6 phenyl ring interacts with the COX-2 binding site amino acids Ile(345), Val(349), Leu(359), Leu(531), and Met(535) and that the OMe substituent may be responsible for proper orientation of the C-3 p-SO(2)Me-phenyl ring within the COX-2 secondary pocket (Gln(192), Arg(513), and Phe(518)). These results show that the COX-2 selectivity and potency of 3,4,6-triphenylpyranone regioisomers can be modulated by appropriate placement of the p-SO(2)Me pharmacophore on either the C-3 or C-4 phenyl moiety. In addition, electronic properties at the para-position of a C-6 phenyl substituent on the central pyranone ring govern COX-2 inhibitory potency and selectivity by controlling the orientation of the p-SO(2)Me pharmacophore within the COX-2 secondary pocket.     

Design of potent and selective hybrid inhibitors of the mitotic kinase Nek2: structure-activity relationship, structural biology, and cellular activity

We report herein a series of Nek2 inhibitors based on an aminopyridine scaffold. These compounds have been designed by combining key elements of two previously discovered chemical series. Structure based design led to aminopyridine (R)-21, a potent and selective inhibitor able to modulate Nek2 activity in cells.     

Discovery of novel selective inhibitors of human intestinal carboxylesterase for the amelioration of irinotecan-induced diarrhea: synthesis, quantitative structure-activity relationship analysis, and biological activity

The dose-limiting toxicity of the highly effective anticancer agent 7-ethyl-10-[4-(1-piperidino)-1-piperidino]carbonyloxy-camptothecin (irinotecan; CPT-11) is delayed diarrhea. This is thought to be caused by either bacteria-mediated hydrolysis of the glucuronide conjugate of the active metabolite 7-ethyl-10-hydroxycamptothecin (SN-38) or direct conversion of CPT-11 to SN-38 by carboxylesterases (CE) in the small intestine. After drug administration, a very high level of CPT-11 is present in the bile; this is deposited into the duodenum, the region of the gut with the highest levels of CE activity. Hence, it is likely that direct conversion of the drug to SN-38 is partially responsible for the diarrhea associated with this agent. In an attempt to ameliorate this toxicity, we have applied Target-Related Affinity Profiling to identify novel CE inhibitors that are selective inhibitors of the human intestinal enzyme (hiCE). Seven inhibitors, all sulfonamide derivatives, demonstrated greater than 200-fold selectivity for hiCE compared with the human liver CE hCE1, and none was an inhibitor of human acetylcholinesterase or butyrylcholinesterase. Quantitative structure-activity relationship (QSAR) analysis demonstrated excellent correlations with the predicted versus experimental Ki values (r2 = 0.944) for hiCE. Additionally, design and synthesis of a tetrafluorine-substituted sulfonamide analog, which QSAR indicated would demonstrate improved inhibition of hiCE, validated the computer predictive analyses. These and other phenyl-substituted sulfonamides compounds are regarded as lead compounds for the development of effective, selective CE inhibitors for clinical applications.     

Design, synthesis, and structure-activity relationships of phthalimide-phenylpiperazines: a novel series of potent and selective alpha(1)(a)-adrenergic receptor antagonists

Beginning from the screening hit and literature alpha(1)-adrenergic compounds, a hybridized basic skeleton A was proposed as the pharmacophore for potent and selective alpha(1a)-AR antagonists. Introduction of a hydroxy group to increase the flexibility afforded B which served as the screening model and resulted in the identification of the second-generation lead 1. Using the Topliss approach, a number of potent and selective alpha(1a)-AR antagonists were discovered. In all cases, binding affinity and selectivity at the alpha(1a)-AR of S-hydroxy enantiomers were higher than those of the R-hydroxy enantiomers. As compared to the des-hydroxy analogues, the S-hydroxy enantiomers had slightly lower binding affinity at alpha(1a)-AR but gained more than 2-fold selectivity for alpha(1a)-AR over alpha(1b)-AR, and 2- to 6-fold selectivity for alpha(1a)-AR over alpha(1d)-AR. They also had less cross activities against a panel of 25-35 peripheral and CNS receptors. The S-hydroxy enantiomers 23 and 24 (K(i) = 0.29 nM, 0.33 nM; alpha(1b)/alpha(1a) >5690, >6060; alpha(1d)/alpha(1a) = 186, 158, respectively) were slightly less potent but much more selective at alpha(1a)-AR than tamsulosin (K(i) = 0.13 nM, alpha(1b)/alpha(1a) = 14.8, alpha(1d)/alpha(1a) = 1.4). In the functional assay, the S-hydroxy enantiomers 20, 23, and 24 were less potent than tamsulosin in inhibiting contractions of rat prostate tissue but more selective in the inhibition of tissue contractions of rat prostate versus rat aorta. Compound 24 was selected as the development candidate for the treatment of BPH.     

Structure-based design, synthesis, and structure-activity relationship studies of novel non-nucleoside adenosine deaminase inhibitors

We disclose herein optimization efforts around the novel, highly potent non-nucleoside adenosine deaminase (ADA) inhibitor, 1-[(R)-1-hydroxy-4-(6-(3-(1-methylbenzimidazol-2-yl)propionylamino)indol-1-yl)-2-butyl]imidazole-4-carboxamide 1 (K(i)= 7.7 nM), which we recently reported. Structure-based drug design (SBDD) utilizing the crystal structure of the 1/ADA complex was performed in order to obtain structure-activity relationships (SAR) for this type of compound rationally and effectively. To utilize the newly formed hydrophobic space (F2), replacement of the benzimidazole ring of 1 with a n-propyl chain (4b) or a simple phenyl ring (4c) was tolerated in terms of binding activity, and the length of the methylene-spacer was shown to be optimal at two or three. Replacement of an amide with an ether as a linker was also well tolerated in terms of binding activity and moreover improved the oral absorption (6a and 6b). Finally, transformation of indol-1-yl to indol-3-yl resulted in discovery of a novel highly potent and orally bioavailable ADA inhibitor, 1-[(R)-4-(5-(3-(4-chlorophenyl)propoxy)-1-methylindol-3-yl)-1-hydroxy-2-butyl]imidazole-4-carboxamide 8c.     

Design, synthesis, and discovery of novel trans-stilbene analogues as potent and selective human cytochrome P450 1B1 inhibitors.

A series of trans-stilbene derivatives containing a 3,5-dimethoxyphenyl moiety were prepared through a new efficient solution phase synthetic pathway, and their inhibitory activities were evaluated on human cytochrome P450s (CYP) 1A1, 1A2, and 1B1 to find a potent and selective CYP1B1 inhibitor. We found that a substituent at the 2-position of the stilbene skeleton plays a very important role in discriminating between CYP1As and CYP1B1. Among the compounds tested, the most selective and potent CYP1B1 inhibitor was 2,3',4,5'-tetramethoxystilbene. Compound 7j, 2-[2-(3,5-dimethoxy-phenyl)vinyl]thiophene, showed greater inhibitory activities but had a lower selectivity toward all of the CYP1s tested.     

Discovery and structure-activity relationship of potent and selective covalent inhibitors of transglutaminase 2 for Huntington's disease

Tissue transglutaminase 2 (TG2) is a multifunctional protein primarily known for its calcium-dependent enzymatic protein cross-linking activity via isopeptide bond formation between glutamine and lysine residues. TG2 overexpression and activity have been found to be associated with Huntington's disease (HD); specifically, TG2 is up-regulated in the brains of HD patients and in animal models of the disease. Interestingly, genetic deletion of TG2 in two different HD mouse models, R6/1 and R6/2, results in improved phenotypes including a reduction in neuronal death and prolonged survival. Starting with phenylacrylamide screening hit 7d, we describe the SAR of this series leading to potent and selective TG2 inhibitors. The suitability of the compounds as in vitro tools to elucidate the biology of TG2 was demonstrated through mode of inhibition studies, characterization of druglike properties, and inhibition profiles in a cell lysate assay.     

Design and structure-activity relationships of potent and selective inhibitors of blood coagulation factor Xa.

The discovery of a series of non-peptide factor Xa (FXa) inhibitors incorporating 3-(S)-amino-2-pyrrolidinone as a central template is described. After identifying compound 4, improvements in in vitro potency involved modifications of the liphophilic group and optimizing the angle of presentation of the amidine group to the S1 pocket of FXa. These studies ultimately led to compound RPR120844, a potent inhibitor of FXa (K(i) = 7 nM) which shows selectivity for FXa over trypsin, thrombin, and several fibrinolytic serine proteinases. RPR120844 is an effective anticoagulant in both the rat model of FeCl(2)-induced carotid artery thrombosis and the rabbit model of jugular vein thrombus formation.     

Design, synthesis, and SAR of potent and selective dipeptide-derived inhibitors for dipeptidyl peptidases

In this paper we report the systematic search for new, potent, and selective DPP II inhibitors. A study of the structure-activity relationship was conducted starting from aminoacyl pyrrolidides as lead compounds. Rational exploration of the P(1) and P(2) building blocks led to the discovery of some very potent DPP II inhibitors which can be characterized by their high selectivity for DPP II with regard to DPP IV. Dab-Pip and Dab-Pip-2-CN were selected as the most promising inhibitors (IC(50) nM range) and will enable us to study the physiological role of DPP II and to differentiate between DPP II and DPP IV in biological systems.     

Design and synthesis of potent, selective, and orally active fluorine-containing renin inhibitors.

A series of primate renin inhibitors containing difluorocarbinol and difluoroketone groups at the P1-P1' position have been synthesized and studied both in vitro and in vivo. In vitro, the compounds were evaluated as inhibitors of monkey renin and the closely related aspartic proteinase, cathepsin D (bovine), as a measure of enzyme selectivity. Interestingly, the difluoroketone derivatives showed greatly reduced selectivity compared with the corresponding alcohols. However, selectivity could be enhanced by judicious choice of other substituents. Sites influencing selectivity, included not only P2, which is well-known to strongly affect selectivity, but also the P4, P1-P1', and P2' sites. These results make possible the design of inhibitors with a greater selectivity for either renin versus cathepsin D. In vivo several of the compounds in the difluoroketone series have shown good oral activity in the salt depleted normotensive cynomolgus monkey model.