Discovery of 5-(5-fluoro-1H-pyrrolo[2,3-b]pyridin-3-yl)pyrazin-2(1H)-one derivatives as new potent PB2 inhibitors

PB2 is an important subunit of influenza RNA-dependent RNA polymerase (RdRP) and has been recognized as a promising target for the treatment of influenza. We herein report the discovery of a new series of PB2 inhibitors containing the skeleton 5-(5-fluoro-1H-pyrrolo[2,3-b]pyridin-3-yl)pyrazin-2(1H)-one. Compound 12b is the most potent one, which showed K_D values of 0.11 μM and 0.19 μM in surface plasmon resonance (SPR) and isothermal titration calorimetry (ITC) assays, respectively. In antiviral activity and cellular cytotoxicity assays, compound 12b showed an EC₅₀ value of 1.025 μM and a CC₅₀ value greater than 100 μM. Molecular docking was also used to predict the binding mode of 12b with PB2. Collectively, this study provides a promising lead compound for subsequent anti-influenza drug discovery targeting PB2.     

Discovery of 5-(4-methylpiperazin-1-yl)-2-nitroaniline derivatives as a new class of SIRT6 inhibitors

SIRT6 is a deacetylase of histone H3 and inhibitors of SIRT6 have been thought as potential agents for treatment of diabetes. Herein we report the discovery of a series of new SIRT6 inhibitors containing the skeleton 1-phenylpiperazine. Among them, compound 5-(4-methylpiperazin-1-yl)-2-nitroaniline (6d) is the most potent one, which showed an IC₅₀ value of 4.93 μM against SIRT6 in the Fluor de Lys (FDL) assay. It displayed K_D values of 9.76 μM and 10 μM in surface plasmon resonance (SPR) and isothermal titration calorimetry (ITC) assays, respectively. In selectivity assay, 6d showed no activity against other members of the HDAC family (SIRT1-3 and HDAC1-11) at concentrations up to 200 µM. In a mouse model of type 2 diabetes, 6d could significantly increase the level of glucose transporter GLUT-1, thereby reducing blood glucose. Overall, this study provides a promising lead compound for subsequent drug discovery targeting SIRT6.     

Synthesis and antibacterial activity of N-(5-benzylthio-1,3,4-thiadiazol-2-yl) and N-(5-benzylsulfonyl-1,3,4-thiadiazol-2-yl)piperazinyl quinolone derivatives

A series of N-(5-benzylthio-1,3,4-thiadiazol-2-yl) and N-(5-benzylsulfonyl-1,3,4-thiadiazol-2-yl) derivatives of piperazinyl quinolones was synthesized and evaluated for antibacterial activity against Gram-positive and Gram-negative microorganisms. Some of these derivatives exhibit high activity against Gram-positive bacteria; Staphylococcus aureus and Staphylococcus epidermidis, comparable or more potent than their parent N-piperazinyl quinolones norfloxacin and ciprofloxacin as reference drugs. The SAR of this series indicates that both the structure of the benzyl unit and the S or SO(2) linker dramatically impact antibacterial activity.     

Synthesis, bioactivity and SAR study of N'-(5-substituted-1,3,4-thiadiazol-2-yl)-N-cyclopropylformyl-thioureas as ketol-acid reductoisomerase inhibitors

Ketol-acid reductoisomerase (KARI; EC 1.1.1.86) catalyzes the second common step in branched-chain amino acid biosynthesis. The catalyzed process consists of two steps, the first of which is an alkyl migration from one carbon atom to its neighboring atom. The likely transition state is a cyclopropane derivative, thus a new series of cyclopropanecarbonyl thiourea derivatives were designed and synthesized involving a one-pot phase transfer catalyzed reaction. Rice KARI inhibitory activity of these compounds were evaluated and the 5-butyl substituted (3e) and 3-pyridinyl substituted (3n) compounds reached 100% at 100 microg x mL(- 1). Structure-activity relationship shows that longer chain derivatives had higher KARI inhibitory activity. Meanwhile substitution of the 4-position of the benzene ring had higher KARI inhibitory activity than that of the 2 and 3-position. Auto-Dock was used to predict the binding mode of 3n. This was done by analyzing the interaction of compound 3n with the active sites of the available spinach KARI. This was in accord with the results analyzed by the frontier molecular orbital theory.     

Synthesis and evaluation of 4-(1,3,4-oxadiazol-2-yl)-benzenesulfonamides as potent carbonic anhydrase inhibitors

Human Carbonic anhydrase (hCA) I and II are crucial targets for anti-acute mountain sickness. Twenty-one 4-(1,3,4-oxadiazol-2-yl) benzenesulfonamides were synthesized and screened against these two isoforms. The results illustrated that 5c, 5g, 5h, 5k were more potent against both hCA I and II than clinical drug AAZ. In particular, the value of compound 5c with hCA I (18.08 nM) was over 84-fold more than of AAZ with hCA I. The data of docking simulations were also in accord with the tendency of inhibitive activities. Furthermore, compound 6h, the methanesulfonate of 5h, showed better anti-hypoxia activity than AAZ in vivo, making it interesting lead compound.     

Identification of N-(5-(phenoxymethyl)-1,3,4-thiadiazol-2-yl)acetamide derivatives as novel protein tyrosine phosphatase epsilon inhibitors exhibiting anti-osteoclastic activity

Cytosolic protein tyrosine phosphatase epsilon (cyt-PTPε) plays a central role in controlling differentiation and function of osteoclasts, whose overactivation causes osteoporosis. Based on our previous study reporting a number of cyt-PTPε inhibitory chemical compounds, we carried out a further and extended analysis of our compounds to examine their effects on cyt-PTPε-mediated dephosphorylation and on osteoclast organization and differentiation. Among five compounds showing target selectivity to cyt-PTPε over three other phosphatases in vitro, two compounds exhibited an inhibitory effect against the dephosphorylation of cellular Src protein, the cyt-PTPε substrate. Moreover, these two compounds caused destabilization of the podosome structure that is necessary for the bone-resorbing activity of osteoclasts, and also attenuated cellular differentiation of monocytes into osteoclasts, without affecting cell viability. Therefore, these findings not only verified anti-osteoclastic effects of our cyt-PTPε inhibitory compounds, but also showed that cyt-PTPε expressed in osteoclasts could be a putative therapeutic target worth considering.     

Design and synthesis of novel 3-(benzo[d]oxazol-2-yl)-5-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyridin-2-amine derivatives as selective G-protein-coupled receptor kinase-2 and -5 inhibitors

G-protein-coupled receptor kinase (GRK)-2 and -5 are emerging therapeutic targets for the treatment of cardiovascular disease. In our efforts to discover novel small molecules to inhibit GRK-2 and -5, a class of compound based on 3-(benzo[d]oxazol-2-yl)-5-(1-(piperidin-4-yl)-1H-pyrazol-4-yl)pyridin-2-amine was identified as a novel hit by high throughput screening campaign. Structural modification of parent benzoxazole scaffolds by introducing substituents on phenyl displayed potent inhibitory activities toward GRK-2 and -5.     

Discovery of novel 2-[2-(3-hydroxy-pyridin-2-yl)-thiazol-4-yl]-acetamide derivatives as HIF prolyl 4-hydroxylase inhibitors; SAR,synthesis and modeling evaluation

The design, synthesis, and capacity to inhibit HIF prolyl 4-hydroxylases (PHDs) are described for 2-[2-(3-hydroxy-pyridin-2-yl)-thiazol-4-yl]-acetamide analogs. These analogs revealed two kinds of novel scaffolds as PHD2 inhibitors. Synthetic routes were developed for the preparation of their analogs containing the new scaffolds. In addition, the structure–activity relationship (SAR) of the 2-[2-(3-hydroxy-pyridin-2-yl)-thiazol-4-yl]-acetamide derivatives and their biological activities were reported. The complex structure of compound 18 with PHD2 was also obtained for the purpose of more efficient lead optimization.     

Synthesis,molecular modeling and biological evaluation of N-benzylidene-2-((5-(pyridin-4-yl)-1,3,4-oxadiazol-2-yl)thio)acetohydrazide derivatives as potential anticancer agents

A series of new 1,3,4-oxadiazole derivatives (6a–6x) containing pyridine and acylhydrazone moieties were synthesized and developed as potential telomerase inhibitors. The bioassay tests demonstrated that compounds 6n, 6o, 6q, 6s and 6t exhibited significant broad-spectrum anticancer activity with IC₅₀ range from 0.76 to 9.59 μM against the four cancer cell lines (HEPG2, MCF7, SW1116 and BGC823). Moreover, all the title compounds were assayed for telomerase inhibition using the TRAP-PCR-ELISA assay. Compound 6s showed the highest anticancer activity with IC₅₀ of 0.76–1.54 μM against the tested cancer cell lines and exhibited the most potent telomerase inhibitory activity with IC₅₀ of 1.18 ± 0.14 μM. The docking simulation was carried out to investigate a possible binding mode of compound 6s into the active site of telomerase (pdb. 3DU6) while the QSAR model was built to check the previous work as well as to introduce new directions.     

Novel 5-(benzyloxy)pyridin-2(1H)-one derivatives as potent c-Met inhibitors

A series of novel 5-(benzyloxy)pyridin-2(1H)-ones were designed, synthesized and biologically evaluated for c-Met inhibition. Various amides and benzoimidazoles at C-3 position were investigated. A potent compound 12b with a c-Met IC₅₀ of 12 nM was identified. This compound exhibited potent inhibition of EBC-1 cell associated with c-Met constitutive activation and showed high selectivity for c-Met than other tested 11 kinases. The binding model 12b with c-Met was disclosed by docking analysis.     

Synthesis of (1,3,4-thiadiazol-2-yl)-acrylamide derivatives as potential antitumor agents against acute leukemia cells

A lead compound with the (1,3,4-thiadiazol-2-yl)-acrylamide scaffold was discovered to have significant cytotoxicity on several tumor cell lines in an in-house cell-based screening. A total of 60 derivative compounds were then synthesized and tested in a CCK-8 cell viability assay. Some of them exhibited improved cytotoxic activities. The most potent compounds had IC₅₀ values of 1–5 μM on two acute leukemia tumor cell lines, i.e. RS4;11 and HL-60. Flow cytometry analysis of several active compounds and detection of caspase activation indicated that they induced caspase-dependent apoptosis. It was also encouraging to observe that these compounds did not have obvious cytotoxicity on normal cells, i.e. IC₅₀ > 50 μM on HEK-293T cells. Although the molecular targets of this class of compound are yet to be revealed, our current results suggest that this class of compound represents a new possibility for developing drug candidates against acute leukemia.     

New heterocyclic analogues of 4-(2-chloro-5-methoxyanilino)quinazolines as potent and selective c-Src kinase inhibitors

A series of 5,7-disubstituted quinazolines, bearing 4-heteroaryl substituents such as 2-pyridinylamine or 2-pyrazinylamine, has been synthetised and evaluated as c-Src kinase inhibitors. Highly potent inhibition, high selectivity and physical properties suitable for oral dosing were achieved within this series: 23d and 42 were identified as sub-0.1 μM inhibitors in a c-Src-driven cell proliferation assay and displayed adequate rat pharmacokinetics after oral administration.     

Novel and orally active 5-(1,3,4-oxadiazol-2-yl)pyrimidine derivatives as selective FLT3 inhibitors

5-(1,3,4-Oxadiazol-2-yl)pyrimidine derivative 1 was identified as a new class of FLT3 inhibitor from our compound library. With the aim of enhancement of antitumor activity of 2 prepared by minor modification of 1, structure optimization of side chains at the 2-, 4-, and 5-positions of the pyrimidine ring of 2 was performed to improve the metabolic stability. Introduction of polar substituents on the 1,3,4-oxadiazolyl group contributed to a significant increase in the metabolic stability. As a result, a series of compounds showed increased efficacy against MOLM-13 xenograft model in mice by oral administration.     

5-(2-amino-pyrimidin-4-yl)-1H-pyrrole and 2-(2-amino-pyrimidin-4-yl)-1,5,6,7-tetrahydro-pyrrolo[3,2-c]pyridin-4-one derivatives as new classes of selective and orally available Polo-like kinase 1 inhibitors

The discovery and characterization of two new chemical classes of potent and selective Polo-like kinase 1 (PLK1) inhibitors is reported. For the most interesting compounds, we discuss the biological activities, crystal structures and preliminary pharmacokinetic parameters. The more advanced compounds inhibit PLK1 in the enzymatic assay at the nM level and exhibit good activity in cell proliferation on A2780 cells. Furthermore, these compounds showed high levels of selectivity on a panel of unrelated kinases, as well as against PLK2 and PLK3 isoforms. Additionally, the compounds show acceptable oral bioavailability in mice making these inhibitors suitable candidates for further in vivo activity studies.     

Discovery of 2-((4-resorcinolyl)-5-aryl-1,2,3-triazol-1-yl)acetates as potent Hsp90 inhibitors with selectivity over TRAP1

As the most abundant heat shock protein (HSP), Hsp90 is actively involved in tumor cell growth and various responses to anti-carcinogenic stress. Hsp90 has thus emerged as a potential drug target. A structure-based drug design approach was applied to develop novel resorcinolyltriazole derivatives as Hsp90 inhibitors. Structure-activity relationships (SARs) and molecular docking were investigated to provide a rationale for binding affinity and paralog selectivity. Click chemistry between iodoethynylresorcinol and an azido derivative was used to synthesize a new family of 2-((4-resorcinolyl)-5-aryl-1,2,3-triazol-1-yl) acetates that exhibited Hsp90 binding affinities of 40–100 nM (IC₅₀). Among the synthesized molecules, the triazole alkyl acetates displayed the highest Hsp90 binding affinities. Their potency against Hsp90 was over 100-fold stronger than against TRAP1 and 1–3-fold stronger than against Grp94. In particular, compounds 18, 19, and 30 had Hsp90 inhibitory activities of ~45 nM (IC₅₀) and they displayed over 350-fold selectivity for Hsp90 over TRAP1.     

Discovery and preclinical studies of 5-isopropyl-6-(5-methyl-1,3,4-oxadiazol-2-yl)-N-(2-methyl-1H-pyrrolo[2,3-b]pyridin-5-yl)pyrrolo[2,1-f][1,2,4]triazin-4-amine (BMS-645737), an in vivo active potent VEGFR-2 inhibitor

We report herein a series of substituted N-(1H-pyrrolo[2,3-b]pyridin-5-yl)pyrrolo[2,1-f][1,2,4]triazin-4-amines as inhibitors of vascular endothelial growth factor receptor-2 tyrosine kinase. Through structure–activity relationship studies, biochemical potency, pharmacokinetics, and kinase selectivity were optimized to afford BMS-645737 (13), a compound with good preclinical in vivo activity against human tumor xenograft models.     

Synthesis and anti-angiogenic activity of 6-(1,2,4-thiadiazol-5-yl)-3-amino pyridazine derivatives

General screening for inhibitors of microvessel growth in vitro in the rat aortic ring assay led to the discovery of a novel series of thiadiazole pyridazine compounds with potential anti-angiogenic activity. Chemical optimization produced orally active compounds with potent in vitro and in vivo anti-angiogenesis and anti-tumor activities.     

N-(pyrimidin-4-yl) and N-(pyridin-2-yl) phenylalanine derivatives as VLA-4 integrin antagonists

The SAR studies to optimise both potency and rate of clearance in the rat for a series of pyrimidine and pyridine based VLA-4 antagonists are described.     

Synthesis of new N-(2-(trifluoromethyl)pyridin-4-yl)anthranilic acid derivatives and their evaluation as anticancer agents

The N-(2-(trifluoromethyl)pyridin-4-yl)anthranilic acid 6 and a series of its ester and amide derivatives were synthesized and evaluated for their in vitro cytotoxic activity against human cancer cells. Ester derivatives 13 and 18 exhibited potent growth inhibitory activity with GI(50) values at nanomolar concentrations. Among amide derivatives, N-anthraniloylglycinate 19 shown moderate inhibitory activity in the full panel cancer cell line screening.     

Quinoxalinone (Part II). Discovery of (Z)-3-(2-(pyridin-4-yl)vinyl)quinoxalinone derivates as potent VEGFR-2 kinase inhibitors

Inhibition of VEGFR-2 kinase has been highlighted as one of the well-defined strategies to suppress tumor growth via blockade of angiogenesis. Guided by the principles of bioisosteric replacement and pharmacophoric fragment migration, a series of novel quinoxalinone derivates were designed, synthesized and evaluated for their VEGFR-2 inhibitory potencies. Among them, compounds 7c, 8b, 8c, 8e and 10b displayed antiangiogenic abilities via the in vitro tube formation assay (cellular level) and ex vivo rat aortic ring assay (tissue level) at a low concentration (0.1 μM). By means of in vivo zebrafish embryo model, two (Z)-3-(2-(pyridin-4-yl)vinyl)quinoxalinone derivates 8c and 8e showed significant antiangiogenesis effects, suggesting they have potentials to be developed into antiangiogenesis agents via further structural optimization. Moreover, these two compounds also demonstrated potent inhibition toward VEGFR-2 and B-raf kinases in a low concentration (1 μM). A possible interpretation of our evaluation result has been presented by a molecular docking study by docking representative compound 8c with VEGFR-2.