1,2-Diamines as inhibitors of co-activator associated arginine methyltransferase 1 (CARM1)

We have identified the N¹-benzyl-N²-methylethane-1,2-diamine unit as a substitute for the (S)-alanine benzylamide moiety for the design of co-activator associated arginine methyltransferase 1 (CARM1) inhibitors. The potency of these inhibitors is in the same order of magnitude as their predecessors and their clearance, volume of distribution, and half lives were greatly improved.     

Pyrazole inhibitors of coactivator associated arginine methyltransferase 1 (CARM1)

This study reports the identification and Hits to Leads optimization of inhibitors of coactivator associated arginine methyltransferase (CARM1). Compound 7b is a potent, selective inhibitor of CARM1.     

Synthesis of new pyridothienopyrimidinone derivatives as Pim-1 inhibitors

Four series of pyridothienopyrimidin-4-one derivatives were designed and prepared to improve the pim-1 inhibitory activity of the previously reported thieno[2,3-b]pyridines. Significant improvement in the pim-1 inhibition and cytotoxic activity was achieved using structure rigidification strategy via ring closure. Six compounds (6c, 7a, 7c, 7d, 8b and 9) showed highly potent pim-1 inhibitory activity with IC₅₀ of 4.62, 1.18, 1.38, 1.97, 8.83 and 4.18?μM, respectively. Four other compounds (6b, 6d, 7b and 8a) showed moderate pim-1 inhibition. The most active compounds were tested for their cytotoxic activity on three cell lines [MCF7, HCT116 and PC3]. Compounds 7a [the 2-(2-chlorophenyl)-2,3-dihydro derivative] and 7d [the 2-(2-(trifluoromethyl)-phenyl)-2,3-dihydro derivative] displayed the most potent cytotoxic effect on the three cell lines tested consistent with their highest estimated pim-1 IC₅₀ values.     

Kinetic mechanism of protein arginine methyltransferase 1

Protein arginine methyltransferases (PRMTs) are SAM-dependent enzymes that catalyze the mono- and dimethylation of peptidyl arginine residues. Although all PRMTs produce monomethyl arginine (MMA), type 1 PRMTs go on to form asymmetrically dimethylated arginine (ADMA), while type 2 enzymes form symmetrically dimethylated arginine (SDMA). PRMT1 is the major type 1 PRMT in vivo, thus it is the primary producer of the competitive NOS inhibitor, ADMA. Hence, potent inhibitors, which are highly selective for this particular isozyme, could serve as excellent therapeutics for heart disease. However, the design of such inhibitors is impeded by a lack of information regarding this enzyme's kinetic and catalytic mechanisms. Herein we report an analysis of the kinetic mechanism of human PRMT1 using both an unmethylated and a monomethylated substrate peptide based on the N-terminus of histone H4. The results of initial velocity and product and dead-end inhibition experiments indicate that PRMT1 utilizes a rapid equilibrium random mechanism with the formation of dead-end EAP and EBQ complexes. This mechanism is gratifyingly consistent with previous results demonstrating that PRMT1 catalyzes substrate dimethylation in a partially processive manner.     

N-Benzyl-1-heteroaryl-3-(trifluoromethyl)-1H-pyrazole-5-carboxamides as inhibitors of co-activator associated arginine methyltransferase 1 (CARM1)

A series of N-benzyl-1-heteroaryl-3-(trifluoromethyl)-1H-pyrazole-5-carboxamides targeting co-activator associated arginine methyltransferase 1 (CARM1) have been designed and synthesized. The potency of these inhibitors was influenced by the nature of the heteroaryl fragment with the thiophene analogues being superior to thiazole, pyridine, isoindoline and benzofuran based inhibitors.     

Thiophene and bioisostere derivatives as new MMP12 inhibitors

A new MMP12 inhibitor series has been identified containing a thiophene moiety. Different approaches have been considered to replace this potential toxicophore. α-Fluorothiophene derivatives were the most interesting compounds. Their synthesis is presented.     

Discovery of new potent protein arginine methyltransferase 5 (PRMT5) inhibitors by assembly of key pharmacophores from known inhibitors

Protein arginine methyltransferase 5 (PRMT5) is an epigenetics related enzyme that has been validated as a promising therapeutic target for human cancer. Up to now, two small molecule PRMT5 inhibitors has been put into phase I clinical trial. In the present study, a series of candidate molecules were designed by combining key pharmacophores of formerly reported PRMT5 inhibitors. The in vitro PRMT5 inhibitory testing of compound 4b14 revealed an IC₅₀ of 2.71?μM, exhibiting high selectivity over PRMT1 and PRMT4 (>70-fold selective). As expected, 4b14 exhibited potent anti-proliferative activity against a panel of leukemia and lymphoma cells, including MV4-11, Pfeiffer, SU-DHL-4 and KARPAS-422. Besides, 4b14 showed significant cell cycle arrest and apoptosis-inducing effects, as well as reduced the cellular symmetric arginine dimethylation level of SmD3 protein. Finally, affinity profiling analysis indicated that hydrophobic interactions, π-π stacking and cation-π actions made the major contributions to the overall binding affinity. This scaffold provides a new chemical template for further development of better lead compounds targeting PRMT5.     

Discovery of 2-arylquinazoline derivatives as a new class of ASK1 inhibitors

The development of a new series of apoptosis signal-regulating kinase 1 (ASK1) inhibitors is described. Starting from purine, pyrimidine and quinazoline scaffolds identified by high throughput screening, we used tools of structure-based drug design to develop a series of potent kinase inhibitors, including 2-arylquinazoline derivatives 12 and 23, with submicromolar inhibitory activities against ASK1. Kinetic analysis demonstrated that the 2-arylquinazoline scaffold ASK1 inhibitors described herein are ATP competitive.     

Design and synthesis of new tetrahydroquinolines derivatives as CETP inhibitors

This letter describes the discovery and SAR optimization of tetrazoyl tetrahydroquinoline derivatives as potent CETP inhibitors. Compound 6m exhibited robust HDL-c increase in hCETP/hApoA1 double transgenic model and favorable pharmacokinetic properties.     

Evaluation of new chalcone derivatives as polyphenol oxidase inhibitors

A newly series of 4-(phenylurenyl)chalcone (4a–j) and 4′-(phenylurenyl/thiourenyl)chalcone (9a–l) derivatives were synthesized and their inhibitory effects on the diphenolase activity of banana tyrosinase were evaluated. Tyrosinase has been purified from banana on an affinity gel comprised of Sepharose 4B-l-tyrosine-p-aminobenzoic acid. The result showed that 4a–j inhibited the PPO enzyme activity. Conversely, 9a–h and 9i–l showed activator effect on tyrosinase enzyme activity.     

Phenanthrene derivatives from Appendicula reflexa as new CDK1/cyclin B inhibitors

The chemical investigation of the epiphytic orchid Appendicula reflexa led to the isolation of one new phenanthrene, 3,4,6,7-tetramethoxyphenanthrene-2,8-diol (1) and one new bisphenanthrene ether, named blestrin E (6), together with the known monomeric phenanthrenes, nudol (2), 3,4,6-trimethoxyphenanthrene-2,7-diol (3), coelonin (4) and 6-methoxycoelonin (5). Their structural elucidation was established on the basis of spectroscopic data analysis. The monomeric phenanthrenes 1 and 3 showed moderate cytotoxic activities against KB, MCF7 and K562 cells, and potent inhibiting activity on CDK1/cyclin B (IC₅₀s = 0.07 and 0.2 μM, respectively).     

The structure and oligomerization of the yeast arginine methyltransferase, Hmt1

Protein methylation at arginines is ubiquitous in eukaryotes and affects signal transduction, gene expression and protein sorting. Hmt1/Rmt1, the major arginine methyltransferase in yeast, catalyzes methylation of arginine residues in several mRNA-binding proteins and facilitates their export from the nucleus. We now report the crystal structure of Hmt1 at 2.9 A resolution. Hmt1 forms a hexamer with approximate 32 symmetry. The surface of the oligomer is dominated by large acidic cavities at the dimer interfaces. Mutation of dimer contact sites eliminates activity of Hmt1 both in vivo and in vitro. Mutating residues in the acidic cavity significantly reduces binding and methylation of the substrate Npl3.     

Oxindole derivatives as inhibitors of TAK1 kinase

Several series of oxindole analogues were synthesized and screened for inhibitory activity against transforming growth factor-β-activating kinase 1 (TAK1). Modifications around several regions of the lead molecules were made, with a distal hydroxyl group in the D region being critical for activity. The most potent compound 10 shows an IC₅₀ of 8.9 nM against TAK1 in a biochemical enzyme assay, with compounds 3 and 6 showing low micromolar cellular inhibition.     

Identification of pyrimidine derivatives as hSMG-1 inhibitors

hSMG-1 kinase plays a dual role in a highly conserved RNA surveillance pathway termed nonsense-mediated RNA decay (NMD) and in cellular genotoxic stress response. Since deregulation of cellular responses to stress contributes to tumor growth and resistance to chemotherapy, hSMG-1 is a potential target for cancer treatment. From our screening efforts, we have identified pyrimidine derivatives as hSMG-1 kinase inhibitors. We report structure-based optimization of this pan-kinase scaffold to improve its biochemical profile and overall kinome selectivity, including mTOR and CDK, to generate the first reported selective hSMG-1 tool compound.     

Synthesis of new thieno[2,3-b]pyridine derivatives as pim-1 inhibitors

Three series of 5-bromo-thieno[2,3-b]pyridines bearing amide or benzoyl groups at position 2 were prepared as pim-1 inhibitors. All the prepared compounds were tested for their pim-1 enzyme inhibitory activity. Two compounds (3c and 5b) showed moderate pim-1 inhibitory activity with IC₅₀ of 35.7 and 12.71?μM, respectively. Three other compounds (3d, 3g and 6d) showed poor pim-1 inhibition. The most active compounds were tested for their cytotoxic activity on five cell lines [MCF7, HEPG2, HCT116, A549 and PC3]. Compound 3g was the most potent cytotoxic agent on almost all the cell lines tested.     

Salicylaldehyde derivatives as new protein kinase CK2 inhibitors

Protein kinase CK2 is a Ser/Thr kinase, with a constitutive activity, that is considered as a promising target for cancer therapy. The currently available CK2 inhibitors lack the potency and the pharmacological properties necessary to be suitable and successful in clinical settings. We report the development of new potent CK2 inhibitors from salicylaldehyde derivatives identified by automated screening of a proprietary small-molecule library. Docking simulations and analysis of the structure-activity relationship for the hits allowed to determine their binding modes on CK2, and to carry out the optimization of their structures. This strategy led to the discovery of potent CK2 inhibitors with novel structures, one of which was able to inhibit CK2 activity in living cells and promote tumor cell death. The essential features required for potent CK2 inhibitory activity of this class of compounds are discussed.     

Syntheses and characterization of non-bisphosphonate quinoline derivatives as new FPPS inhibitors

Background

Farnesyl pyrophosphate synthase (FPPS) is a key regulatory enzyme in the biosynthesis of cholesterol and in the post-translational modification of signaling proteins. It has been reported that non-bisphosphonate FPPS inhibitors targeting its allosteric binding pocket are potentially important for the development of promising anti-cancer drugs.

Methods

The following methods were used: organic syntheses of non-bisphosphonate quinoline derivatives, enzyme inhibition studies, fluorescence titration assays, synergistic effect studies of quinoline derivatives with zoledronate, ITC studies for the binding of FPPS with quinoline derivatives, NMR-based HAP binding assays, molecular modeling studies, fluorescence imaging assay and MTT assays.

Results

We report our syntheses of a series of quinoline derivatives as new FPPS inhibitors possibly targeting the allosteric site of the enzyme. Compound 6b showed potent inhibition to FPPS without significant hydroxyapatite binding affinity. The compound showed synergistic inhibitory effect with active-site inhibitor zoledronate. ITC experiment confirmed the good binding effect of compound 6b to FPPS, and further indicated the binding ratio of 1:1. Molecular modeling studies showed that 6b could possibly bind to the allosteric binding pocket of the enzyme. The fluorescence microscopy indicated that these compounds could get into cancer cells.

Conclusions

Our results showed that quinoline derivative 6b could become a new lead compound for further optimization for cancer treatment.

General significance

The traditional FPPS active-site inhibitors bisphosphonates show poor membrane permeability to tumor cells, due to their strong polarity. The development of new non-bisphosphonate FPPS inhibitors with good cell membrane permeability is potentially important.     

Mechanistic studies on transcriptional coactivator protein arginine methyltransferase 1

Protein arginine methyltransferases (PRMTs) catalyze the transfer of methyl groups from S-adenosylmethionine (SAM) to the guanidinium group of arginine residues in a number of important cell signaling proteins. PRMT1 is the founding member of this family, and its activity appears to be dysregulated in heart disease and cancer. To begin to characterize the catalytic mechanism of this isozyme, we assessed the effects of mutating a number of highly conserved active site residues (i.e., Y39, R54, E100, E144, E153, M155, and H293), which are believed to play key roles in SAM recognition, substrate binding, and catalysis. The results of these studies, as well as pH-rate studies, and the determination of solvent isotope effects (SIEs) indicate that M155 plays a critical role in both SAM binding and the processivity of the reaction but is not responsible for the regiospecific formation of asymmetrically dimethylated arginine (ADMA). Additionally, mutagenesis studies on H293, combined with pH studies and the lack of a normal SIE, do not support a role for this residue as a general base. Furthermore, the lack of a normal SIE with either the wild type or catalytically impaired mutants suggests that general acid/base catalysis is not important for promoting methyl transfer. This result, combined with the fact that the E144A/E153A double mutant retains considerably more activity then the single mutants alone, suggests that the PRMT1-catalyzed reaction is primarily driven by bringing the substrate guanidinium into the proximity of the S-methyl group of SAM and that the prior deprotonation of the substrate guanidinium is not required for methyl transfer.     

'Bridged' stilbene derivatives as selective cyclooxygenase-1 inhibitors

Resveratrol ((E)-3,4',5-trihydroxy-stilbene), a phytoalexin found in various plants, shows non-selective cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) inhibition. In order to find more selective COX inhibitors a series of bridged stilbene derivatives was synthesized and evaluated for their ability to inhibit both COX-1 and COX-2 in vitro. The compounds showed a high rate of COX-1 inhibition with the most potent compounds exhibiting submicromolar IC(50) values and high selectivity indices. A prediction model for COX-inhibiting activity was also developed using the classical LIE approach resulting in consistent docking data for our molecule sample. Phenyl substituted 1,2-dihydronaphthalene derivatives and 1H-indene derivatives therefore represent a novel class of highly selective COX-1 inhibitors and land promising candidates for in vivo studies.