Design,synthesis, and biological activity of novel,potent, and highly selective fused pyrimidine-2-carboxamide-4-one-based matrix metalloproteinase (MMP)-13 zinc-binding inhibitors

Matrix metalloproteinase-13 (MMP-13), a member of the collagenase family of enzymes, has been implicated to play a key role in the pathology of osteoarthritis. Recently, we have reported the discovery of a series of quinazoline-2-carboxamide based non-zinc-binding MMP-13 selective inhibitors, as exemplified by compound 1. We then continued our research of a novel class of zinc-binding inhibitors to obtain follow-up compounds with different physicochemical, pharmacokinetic, and biological activity profiles. In order to design selective MMP-13 inhibitors, we adopted a strategy of connecting a zinc-binding group with the quinazoline-2-carboxamide system, a unique S1′ binder, by an appropriate linker. Among synthesized compounds, a triazolone inhibitor 35 exhibited excellent potency (IC₅₀ = 0.071 nM) and selectivity (greater than 170-fold) over other MMPs (MMP-1, 2, 3, 7, 8, 9, 10, 12, and 14) and tumor necrosis factor-α converting enzyme (TACE). In this article, the design, synthesis, and biological activity of novel zinc-binding MMP-13 inhibitors are described.     

Novel tricyclic pyrazolo[1,5-d][1,4]benzoxazepin-5(6H)-one: Design,synthesis, model and use as hMAO-B inhibitors

Based on our recently reported selective hMAO-A inhibitors, on which, the intramolecular cyclization led to a very interesting change of isoform selectivity. A series of selective hMAO-B inhibitors (3a–3u) with novel scaffold of tricyclic pyrazolo[1,5-d][1,4]benzoxazepin-5(6H)-one were designed and synthesized. Compound 3u (IC₅₀ = 221 nM) exhibited the best inhibitory activity and isoform selectivity against hMAO-B, superior to selegiline (IC₅₀ = 321 nM), which is a commercial selective hMAO-B inhibitor used to Parkinson’s disease. Modeling study indicated that the selectivity of our compounds to hMAO-B is determined by at least two residues, i.e., Ile 199 and Cys 172 (or corresponded Phe 208 and Asn 181 of hMAO-A). These data support further studies to assess rational design of more efficiently selective hMAO-B inhibitors.     

Discovery and structure-guided drug design of inhibitors of 11β-hydroxysteroid-dehydrogenase type I based on a spiro-carboxamide scaffold

Spiro-carboxamides were identified as inhibitors of 11β-hydroxysteroid-dehydrogenase type 1 by high-throughput screening. Structure-based drug design was used to optimise the initial hit yielding a sub-nanomolar IC₅₀ inhibitor (0.5 nM) on human 11β-HSD1 with a high binding efficiency index (BEI of 32.7) which was selective against human 11β-HSD2 (selectivity ratio > 200000).     

Design and synthesis of novel 5-(3,4,5-trimethoxybenzoyl)-4-aminopyrimidine derivatives as potent and selective phosphodiesterase 5 inhibitors: Scaffold hopping using a pseudo-ring by intramolecular hydrogen bond formation

5-(3,4,5-Trimethoxybenzoyl)-4-amimopyrimidine derivatives were found as a novel chemical class of potent and highly selective phosphodiesterase 5 inhibitors. A pseudo-ring formed by an intramolecular hydrogen bond constrained the conformation of 3-chloro-4-methoxybenzylamino and 3,4,5-trimethoxybenzoyl substituents and led to the discovery of T-6932 (19a) with a potent PDE5 inhibitory activity (IC₅₀ = 0.13 nM) and a high selectivity over PDE6 (IC₅₀ ratio: PDE6/PDE5 = 2400). Further modification at the 2-position of T-6932 resulted in the finding of 26, which exhibited potent relaxant effects on isolated rabbit corpus cavernosum (EC₃₀ = 11 nM) with a high PDE5 selectivity over PDE6 (IC₅₀ ratio: PDE6/PDE5 = 2800).     

Adamantyl carboxamides and acetamides as potent human 11β-hydroxysteroid dehydrogenase type 1 inhibitors

The modulation of 11β-HSD1 activity with selective inhibitors has beneficial effects on various metabolic disorders including insulin resistance, dyslipidemia and obesity. Here we report the discovery of a series of novel adamantyl carboxamide and acetamide derivatives as selective inhibitors of human 11β-HSD1 in HEK-293 cells transfected with the HSD11B1 gene. Optimization based on an initially identified 11β-HSD1 inhibitor (3) led to the discovery of potent inhibitors with IC₅₀ values in the 100 nM range. These compounds are also highly selective 11β-HSD1 inhibitors with no activity against 11β-HSD2 and 17β-HSD1. Compound 15 (IC₅₀ = 114 nM) with weak inhibitory activity against the key human cytochrome P450 enzymes and moderate stability in incubation with human liver microsomes is worthy of further development. Importantly, compound 41 (IC₅₀ = 280 nM) provides a new lead that incorporates an adamantyl group surrogate and should enable further series diversification.     

Lumican: A new inhibitor of matrix metalloproteinase-14 activity

We previously showed that lumican regulates MMP-14 expression. The aim of this study was to compare the effect of lumican and decorin on MMP-14 activity. In contrast to decorin, the glycosylated form of lumican was able to significantly decrease MMP-14 activity in B16F1 melanoma cells. Our results suggest that a direct interaction occurs between lumican and MMP-14. Lumican behaves as a competitive inhibitor which leads to a complete blocking of the activity of MMP-14. It binds to the catalytic domain of MMP-14 with moderate affinity (K_D ∼ 275 nM). Lumican may protect collagen against MMP-14 proteolysis, thus influencing cell–matrix interaction in tumor progression.     

Synthesis and structure–activity relationships of 4-fluorophenyl-imidazole p38α MAPK,CK1δ and JAK2 kinase inhibitors

The synthesis and structure–activity relationships of novel 4-(4′-fluorophenyl)imidazoles as selective p38α MAPK, CK1δ and JAK2 inhibitors with improved water solubility are described. Microwave-assisted multicomponent reactions afforded 4-fluorophenyl-2,5-disubstituted imidazoles. Carboxylate and phosphonate groups were introduced via ‘click’ reactions. The kinase selectivity was influenced by the heteroaryl group at imidazole C-5 and the position of a carboxylic acid or tetrazole at imidazole C-2. For example, pyrimidines 15 and 34 inhibited p38α MAPK with IC₅₀ = 250 nM and 96 nM, respectively. Pyridine 3 gave CK1δ inhibition with IC₅₀ = 89 nM and pyridin-2-one 31 gave JAK2 inhibition with IC₅₀ = 62 nM.     

The discovery of potent and selective 4-aminothienopyridines as B-Raf kinase inhibitors

A series of novel, potent 4-aminothienopyridine B-Raf kinase inhibitors was designed and synthesized using knowledge-based design. Compounds 5f, and 6k exhibited not only excellent potency in both enzyme assay (IC₅₀ = 5.1, 16.6 nM) and cellular assay (IC₅₀ = 0.2, 0.2 μM), but also had an outstanding selectivity profile against other kinases.     

Potent and selective inhibitors of the TASK-1 potassium channel through chemical optimization of a bis-amide scaffold

TASK-1 is a two-pore domain potassium channel that is important to modulating cell excitability, most notably in the context of neuronal pathways. In order to leverage TASK-1 for therapeutic benefit, its physiological role needs better characterization; however, designing selective inhibitors that avoid the closely related TASK-3 channel has been challenging. In this study, a series of bis-amide derived compounds were found to demonstrate improved TASK-1 selectivity over TASK-3 compared to reported inhibitors. Optimization of a marginally selective hit led to analog 35 which displays a TASK-1 IC₅₀ = 16 nM with 62-fold selectivity over TASK-3 in an orthogonal electrophysiology assay.     

Discovery of new [1,4]dioxino[2,3-f]quinazoline-based inhibitors of EGFR including the T790M/L858R mutant

A novel series of 2,3-dihydro-[1,4]dioxino[2,3-f]quinazoline derivatives were designed, synthesized and evaluated as reversible and noncovalent epidermal growth factor receptor (EGFR) inhibitors. Most of the compounds exhibited good potency against EGFR^wt and some showed moderate to excellent potency against EGFR^T790M/L858R mutant. The half-maximal inhibitory concentration (IC₅₀) values of twenty-one compounds against EGFR^wt were less than 50 nM, and those of six compounds were less than 10 nM. The IC₅₀ values of eleven compounds against EGFR^T790M/L858R were less than 100 nM. Among these, compound b1 displayed the most potent inhibitory activity against EGFR^wt (IC₅₀ = 2.0 nM) and EGFR^T790M/L858R (IC₅₀ = 6.9 nM). Compounds with excellent inhibitory activities against EGFR^wt and EGFR^T790M/L858R kinase inhibitory activities showed good antiproliferative activities against H358 and A549 cells. Docking study was performed to position compound b1 into the EGFR active pocket to determine the probable binding conformation.     

Discovery and structure–activity relationships of pentanedioic acid diamides as potent inhibitors of 11β-hydroxysteroid dehydrogenase type I

Benzylamides of pentanedioic acid were identified as inhibitors of 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) by high-throughput screening. Optimisation to 2-adamantyl amides yielded inhibitors with single digit nanomolar IC₅₀s on the 11β-HSD1 human isoform. The hydroxy adamantyl amide lead compound was selective against 11β-hydroxysteroid dehydrogenase type 2 (selectivity ratio >1000) and displayed good inhibition of 11β-HSD1 (IC₅₀ < 0.1 μM) in a cellular model (3T3L1 adipocytes).     

Design,synthesis and primary activity evaluation of l-arginine derivatives as amino-peptidase N/CD13 inhibitors

A series of l-arginine derivatives were designed, synthesized and assayed for their activities against amino-peptidase N (APN)/CD13 and metalloproteinase-2 (MMP-2). The results showed that most compounds exhibited high inhibitory activities against APN and low activities against MMP-2. Within this series, two compounds 5q and 5s (IC₅₀ = 5.3 and 5.1 μM) showed similar inhibitory activities compared with bestatin (IC₅₀ = 3.8 μM), which could be used as novel lead compounds for the future APN inhibitors development as anticancer agents.     

Dioxo-triazines as a novel series of cathepsin K inhibitors

A novel dioxo-triazine series of cathepsin K inhibitors was identified from HTS. A rapid exploratory programme led to the discovery of potent and selective cathepsin K inhibitors, typified by compound 24 which displayed IC₅₀ values of 17 nM against catK and >10,000 nM in catL, catB and catS assays.     

Design,synthesis and biological evaluation of novel 4-phenoxy-6,7-disubstituted quinolines possessing (thio)semicarbazones as c-Met kinase inhibitors

In continuing our efforts to identify small molecules able to inhibit c-Met kinase, three series of novel 6,7-disubstituted-4-phenoxyquinoline derivatives (23a–w, 26a–d and 30a–d) bearing (thio)semicarbazone scaffold were designed, synthesized and evaluated for their cytotoxicity. The biological data revealed that most compounds exhibited moderate-to-excellent activity against HT-29, MKN-45, A549 cancer cell lines and relative poor potency toward MDA-MB-231 cell as well as hardly any cytotoxicity in normal PBL cell. Eleven compounds were further examined for their inhibitory activity against c-Met kinase and three compounds (23h, 23n and 26a) demonstrated good inhibitory activity. This work resulted in the discovery of a potent c-Met inhibitor 23n, bearing 2-hydroxy-3-allylphenyl group at R² moiety, as a valuable lead molecule, which possessed remarkable cytotoxicity and high selectivity against A549 and HT-29 cell lines with IC₅₀ values of 11 nM and 27 nM. Besides, it displayed excellent c-Met kinase inhibition on a single-digital nanomolar level (IC₅₀ = 1.54 nM). Meanwhile, the results from preliminarily in vivo study reflected that compound 23n showed promising overall PK profiles, consistent with the efficacy in both MKN-45 and HT-29 tumor xenograft mice model. These results clearly indicated that compound 23n is a potent and highly selective c-Met inhibitor and its favorable in vitro and in vivo profiles warrant further investigation.     

Synthesis,biological evaluation,and docking analysis of a novel family of 1-methyl-1H-pyrrole-2,5-diones as highly potent and selective cyclooxygenase-2 (COX-2) inhibitors

As a continuous research for discovery of new COX-2 inhibitors, we present the simple chemical synthesis, in vitro biological screening, and molecular docking study of 1H-pyrrole-2,5-dione derivatives. New synthetic compounds were evaluated for the inhibitory activities on LPS-induced PGE₂ production in RAW 264.7 macrophage cells as well as the COX-1 and COX-2 inhibitory potency. Among them, compound 9d (MPO-0029) was identified as more potent and selective COX-2 inhibitor [PGE₂ IC₅₀ = 8.7 nM, COX-2 IC₅₀ = 6.0 nM; COX-2 selectivity index (SI) = >168] than celecoxib. Molecular docking experiments were further performed against COX-2 and COX-1 isozymes to determine their probable binding models. Results of molecular docking studies revealed that compound 9d (MPO-0029) has stronger binding interaction with COX-2 than with COX-1 isozyme, and provided successfully complementary theoretical support for the obtained experimental biological data.     

Design,synthesis, and docking studies of afatinib analogs bearing cinnamamide moiety as potent EGFR inhibitors

Two series of afatinib derivatives bearing cinnamamide moiety (10a–n and 11a–h) were designed, synthesized and evaluated for the IC₅₀ values against four cancer cell lines (A549, PC-3, MCF-7 and Hela). Two selected compounds (10e, 10k) were further evaluated for the inhibitory activity against EGFR and VEGFR2/KDR kinases. Seven of the compounds showed excellent cytotoxicity activity and selectivity with the IC₅₀ values in single-digit μM to nanomole range. Three of them are equal to more active than positive control afatinib against one or more cell lines. The most promising compound 10k showed the best activity against A549, PC-3, MCF-7 and Hela cancer cell lines and EGFR kinase, with the IC₅₀ values of 0.07 ± 0.02 μM, 7.67 ± 0.97 μM, 4.65 ± 0.90 μM and 4.83 ± 1.28 μM, which were equal to more active than afatinib (0.05 ± 0.01 μM, 4.1 ± 2.47 μM, 5.83 ± 1.89 μM and 6.81 ± 1.77 μM), respectively. Activity of compounds 10e (IC₅₀ 9.1 nM) and 10k (IC₅₀ 3.6 nM) against EGFR kinase were equal to the reference compound afatinib (IC₅₀ 1.6 nM). Structure–activity relationships (SARs) and docking studies indicated that replacement of the aqueous solubility 4-(dimethylamino)but-2-enamide group by cinnamamide moiety didn’t decrease the antitumor activity. The results suggested that methoxy substitution had a significant impact on the activity and methoxy substituted on C-4 or C-2,3,4 position was benefit for the activity.     

Synthesis and SAR of alkanediamide-linked bisbenzamidines with anti-trypanosomal and anti-pneumocystis activity

A series of alkanediamide-linked bisbenzamidines was synthesized and tested in vitro against a drug-sensitive strain of Trypanosoma brucei brucei, a drug-resistant strain of Trypanosoma brucei rhodesiense and Pneumocystis carinii. Bisbenzamidines linked with longer alkanediamide chains were potent inhibitors of both strains of T. brucei. However, bisbenzamidines linked with shorter alkanediamide chains were the most potent compounds against P. carinii. N,N′-Bis[4-(aminoiminomethyl)phenyl] hexanediamide, 4 displayed potent inhibition (IC₅₀ = 2–3 nM) against T. brucei and P. carinii, and was non-cytotoxic in the A549 human lung carcinoma cell line. The inhibitory bioactivity was significantly reduced when the amidine groups in 4 were moved from the para to the meta positions or replaced with amides.     

Synthesis and biological evaluation of N-mercaptoacylcysteine derivatives as leukotriene A4 hydrolase inhibitors

We studied synthetic modifications of N-mercaptoacylamino acid derivatives to develop a new class of leukotriene A₄ (LTA₄) hydrolase inhibitors. S-(4-Dimethylamino)benzyl-l-cysteine derivative 2a (SA6541) showed inhibitory activity against LTA₄ hydrolase (IC₅₀, 270 nM) and selectivity over other metallopeptidases except angiotensin-converting enzyme (ACE, IC₅₀, 520 nM). Modification at the para-substituent of the phenyl ring of compound 2a improved LTA₄ hydrolase inhibitory activity as well as selectivity over ACE. Finally, we obtained S-(4-cyclohexyl)benzy-l-cysteine derivatives 11l and 16i as potent and selective LTA₄ hydrolase inhibitors.     

Novel HDAC6 isoform selective chiral small molecule histone deacetylase inhibitors

In an effort to identify HDAC isoform selective inhibitors, we designed and synthesized novel, chiral 3,4-dihydroquinoxalin-2(1H)-one and piperazine-2,5-dione aryl hydroxamates showing selectivity (up to 40-fold) for human HDAC6 over other class I/IIa HDACs. The observed selectivity and potency (IC₅₀ values 10–200 nM against HDAC6) is markedly dependent on the absolute configuration of the chiral moiety, and suggests new possibilities for use of chiral compounds in selective HDAC isoform inhibition.     

Development and characterization of endocannabinoid hydrolases FAAH and MAGL inhibitors bearing a benzotriazol-1-yl carboxamide scaffold

A series of (1H-benzo[d][1,2,3]triazol-1-yl)(4-benzylpiperazin-1-yl)methanones and of (1H-benzo[d][1,2,3]triazol-1-yl)(4-phenylpiperazin-1-yl)methanones has been prepared and tested on human fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL). In the benzylpiperazinyl series, compound 29 (ML30) exhibited an IC₅₀ value of 0.54 nM on MAGL, combined with a 1000-fold selectivity versus FAAH, while compounds 11 and 16 acted as potent dual FAAH-MAGL inhibitors (IC₅₀ <10 nM). In the phenylpiperazinyl series, compounds 37, 38, 42, and 43 displayed IC₅₀ values against MAGL in the nanomolar range, whilst being between one and two orders of magnitude less potent on the FAAH, while compounds 31 and 32 were potent FAAH inhibitors (IC₅₀ <20 nM) and over 12-fold selective versus MAGL. The key structural determinants driving the structure–activity relationships were explored by the minimization of the inhibitors inside the active site of both enzymes.