Design, synthesis, and biological evaluation of naphthalene imidazo[1,2-b]pyridazine hybrid derivatives as VEGFR selective inhibitors.

The vascular endothelial growth factor receptor (VEGFR) is a receptor tyrosine kinase that is regarded as an emerging target for abnormal angiogenesis diseases. In this study, novel naphthalene imidazo[1,2-b]pyridazine hybrids as VEGFR selective inhibitors were designed and synthesized using a scaffold hopping strategy based on ponatinib, a multitarget kinase inhibitor. Among the evaluated compounds, derivative 9k (WS-011) demonstrated the most potent inhibitory potency against VEGFR-2 (IC50 = 8.4 nM) and displayed superior VEGFR selectivity over a panel of 70 kinases compared with ponatinib. Furthermore, 9k possessed good cytotoxic effects on various cancer cell lines, especially the colon cancer HT-29 cells, with an acceptable oral bioavailability. Moreover, 9k significantly inhibited the migration and invasion of human umbilical vein endothelial cells (HUVEC) cells and induced apoptosis through the upregulation of apoptotic proteins in HT-29 cells. 9k also effectively suppressed the activation of VEGFR-2 signaling pathways, which in turn inhibited the growth of HT-29 cells and the tube formation of HUVECs in vitro. All of the findings revealed that 9k could be considered a promising antiangiogenesis lead that merits further investigation.

Design, synthesis and biological evaluation of 2,4-pyrimidinediamine derivatives as ALK and HDACs dual inhibitors for the treatment of ALK addicted cancer.

Simultaneous inhibition of histone deacetylases (HDACs) and anaplastic lymphoma kinase (ALK) could enhance therapeutic activity against ALK addicted cancer cells. Herein, a new series of 2,4-pyrimidinediamine derivatives as ALK and HDACs dual inhibitors were designed, synthesised and evaluated. Compound 12a which possessed good inhibitory potency against ALKwt and HDAC1, exhibited stronger antiproliferative activity than Ceritinib on ALK positive cancer cell lines though inducing cell apoptosis and cell cycle arrest in vitro and in vivo. In addition, the mechanism is further verified by the down-regulation of p-ALK protein, and up-regulation of Acetylated histone 3 (Ac-H3) protein in cancer cells. These results suggested that 12a would be a potential candidate for the ALK addicted cancer treatment.

T-17, a novel cyclin-dependent kinases/histone deacetylases dual inhibitor, induces cancer cells death through cell cycle arrest and apoptosis.

Combination of cyclin-dependent kinases (CDKs) and histone deacetylases (HDACs) inhibitors may have statistical synergy in suppressing cancer cell proliferation. Herein, a novel CDKs/HDACs dual inhibitor T-17 was rationally designed, synthesized, and evaluated. Our results demonstrated that T-17 concurrently exhibited potent and balanced inhibitory activity against CDKs (IC50 = 18.0 nM) and HDACs (IC50 = 6.6 nM) and also displayed good cell viability inhibitory effect on four cancer cell lines. Meanwhile, T-17 blocked the MDA-MB-231 and A549 cell cycle at G1 phase and S phase, respectively. In addition, T-17 induced MDA-MB-231 cells apoptosis and inhibited the HDACs and CDKs mediated signaling pathways. Finally, we also found that T-17 had good antitumor activity in vivo. In summary, these results indicated that T-17 would be a promising lead compound which deserves further research.

The Biological Fate of a Novel Anticancer Drug Candidate TNBG-5602: Metabolic Profile, Interaction with CYP450, and Pharmacokinetics in Rats.

TNBG-5602, a novel anticancer drug candidate, may induce the expression of PPARγ, causing targeted lipotoxicity in cancer tissues. In this study, the in vivo metabolism in rats, in vitro metabolism in recombinant cytochromes, molecular docking for the CYP binding site, and pharmacokinetics in rats were explored to better understand TNBG-5602's in vivo fate and behavior. Thirteen metabolites were identified using a high-resolution mass spectrometry method, and metabolizing pathways of TNBG-5602 were proposed. Results suggest that TNBG-5602 could be metabolized by CYP450s, while CYP2D6 may play an important role in its in vivo metabolism. The main metabolizing sites of TNBG-5602 are the amino group on the side chain and rings A and E in the molecule. TNBG-5602 is a potent CYP2D6 inhibitor, with an IC50 value of 2.52 µM. An interaction responsible for its metabolism is formed by the NH on the side chain bonding with the ASP301 on the CYP2D6. The pharmacokinetics in rats after a single intravenous administration were fitted to a two-compartment model. The clearance was 0.022 L min-1, and the elimination half-life was 710.9 min. The distribution volume of the peripheral compartment was 1.88-fold that of the central compartment, while the K12 was 1.5-fold that of K21. In conclusion, these studies have not only revealed the metabolizing pathways of TNBG-5602 using in vivo and in vitro methodology, but they have also provided the pharmacokinetic characteristics of TNBG-5602 in rats. The results suggest that TNBG-5602 has good drug developability in terms of pharmacokinetic behaviors.

Detection of B-type natriuretic peptide by establishing a low-cost and replicable fluorescence resonance energy transfer platform.

Aiming at the establishment of a sensitive and specific diagnostic method for early heart failure (HF), we developed a cost-effective fluorescence resonance energy transfer (FRET) platform for the detection of B-type natriuretic peptide (BNP), a characteristic biomarker of HF. Graphene oxide (GO) was selected as the FRET receptor in view of its advantages including commercial availability, low-cost and chemical stability, and dye-modified aptamer was used as the energy donor of FRET as well as in charge of the specific recognition of BNP. Based on the ON (strong emission) and OFF (quenching) states of FRET in the presence and absence of BNP, respectively, specific detection of BNP was achieved in the range 0.074-0.56 pg/mL with a limit of detection as low as 45 fg/mL (3σ). This FRET platform was applied to detect BNP in 45 blood samples to demonstrate its practicability in clinical diagnosis. Compared to the commonly used Siemens method (chemiluminescence immunoassay, CLIA) in hospital, our approach is more accurate and specific for HF diagnosis with areas under the receiver operating characteristic curves of 0.869 (95% CI 0.733-1.00, P < 0.05) vs 0.850 (95% CI 0.703-0.997, P < 0.05) and specificity of 68.8% vs 65.6%. This platform is promising in early diagnosis of HF through ultrasensitive and specific detection of BNP. Graphical abstract To solve the clinical diagnostic problem for early heart failure (HF) which lacks sensitivity and specificity, we established a cost-effective and rapid fluorescence analysis method based on fluorescence resonance energy transfer (FRET) platform for the detection of B-type natriuretic peptide (BNP), a characteristic biomarker of HF.

Discovery of novel 9H-purin derivatives as dual inhibitors of HDAC1 and CDK2.

HDAC and CDK inhibitors have been demonstrated to be synergistically in suppressing cancer cell proliferation and inducing apoptosis. In this work, we incorporated the pharmacophore groups of HDACs and CDKs inhibitors into one molecule to design and synthesize a series of purin derivatives as HDAC/CDK dual inhibitors. The lead compound 6d, showing good HDAC1 and CDK2 inhibitory activity with IC50 values of 5.8 and 56 nM, respectively, exhibited attractive potency against several cancer cell lines in vitro. This work may lead to the discovery of a novel scaffold andpotentialdual HDAC/CDK inhibitors.

Neural mechanisms underlying the deficit of learning and memory by exposure to Di(2-ethylhexyl) phthalate in rats.

Di(2-ethylhexyl) phthalate (DEHP), as one of the most broadly representative phthalic acid esters, is used as a plasticizer in Polyvinyl chloride production. The exact neurotoxicologically effects of DEHP to human have not been adequately researched. In order to investigate the effects and mechanisms of DEHP exposure on neural circuit, the spatial learning and memory of Sprague Dawley (SD) rats was measured, and the cellular mechanisms underlying synaptic plasticity, cellular excitability and ion channels were detected. Our data showed that the spatial learning and memory was changed by DEHP (100 and 300 mg) treatment. Meanwhile, the frequency of mini Excitatory Postsynaptic Current (mEPSC) from CA3 pyramidal cells were significantly decreased by DEHP exposure (0.1 and 0.3 M); the firing threshold, membrane potential threshold, number, amplitude and latency of Action Potentials (Aps) of CA1 pyramidal cells were altered with the application of DEHP (0.1 and 0.3 M); furthermore, DEHP, both in 0.1 and 0.3 M could inhibit the voltage-gated potassium channel of CA1 pyramidal cells. Our results indicated that DEHP could impair the spatial learning and memory, and this impairment might due to the DEHP-induced suppression of the neuronal excitability and synaptic plasticity by inhibiting the voltage-gated potassium channel, supporting the hypothesis that DEHP could cause the disruption of neural function.

Cellular Mechanism Underlying rTMS Treatment for the Neural Plasticity of Nervous System in Drosophila Brain.

Repetitive transcranial magnetic stimulation (rTMS) is used as a research tool and clinical treatment for the non-clinical and clinical populations, to modulate brain plasticity. In the case of neurologic and psychiatric disease, there is significant evidence to suggest that rTMS plays an important role in the functional recovery after neurological dysfunction. However, the causal role for rTMS in the recovery of nervous dysfunction remains unclear. The purpose of the present study is to detect the regulation of rTMS on the excitatory neuronal transmission and specify the mode of action of rTMS on the neural plasticity using Drosophila whole brain. Therefore, we identified the effects of rTMS on the neural plasticity of central neural system (CNS) by detecting the electrophysiology properties of projection neurons (PNs) from adult Drosophila brain after rTMS. Using patch clamp recordings, we recorded the mini excitatory postsynaptic current (mEPSC) of PNs after rTMS at varying frequencies (1 Hz and 100 Hz) and intensities (1%, 10%, 50%, and 100%). Then, the chronic electrophysiology recordings, including mEPSC, spontaneous action potential (sAP), and calcium channel currents from PNs after rTMS at low frequency (1 Hz), with low intensity (1%) were detected and the properties of the recordings were analyzed. Finally, the frequency and decay time of mEPSC, the resting potential and frequency of sAP, and the current density and rise time of calcium channel currents were significantly changed by rTMS. Our work reveals that rTMS can be used as a tool to regulate the presynaptic function of neural circuit, by modulating the calcium channel in a frequency-, intensity- and time-dependent manner.

A Glimepiride-Metformin Multidrug Crystal: Synthesis, Crystal Structure Analysis, and Physicochemical Properties.

A multidrug crystal based on drug combinations was synthesized by the solvent evaporation method. This multicomponent crystal consisted of antidiabetic drugs Glimepiride (Gli) and Metformin (Met), which was performed by single crystal X-ray structure analysis. The results showed an enhancement of the pharmaceutical properties such as lower hygroscopicity and greater accelerated stability than the parent drug Met, and a higher solubility and dissolution rate than Gli.

One-Step Regioselective Synthesis of Benzofurans from Phenols and α-Haloketones.

Reported here is the direct synthesis of naphthofurans and benzofurans from readily available phenols and α-haloketones promoted by titanium tetrachloride which combines Friedel-Crafts-like alkylation and intramolecular cyclodehydration into one step. This simple protocol allows for the formation of a variety of high value naphthofurans and benzofurans within which a series of cyclic and acyclic groups are readily incorporated. This process demonstrates the advantages of high levels of regioselectivity, broad substrate scope, and moderate to excellent yields.

Eco-Friendly Syntheses of 2-Substituted Benzoxazoles and 2-Substituted Benzothiazoles from 2-Aminophenols, 2-Aminothiophenols and DMF Derivatives in the Presence of Imidazolium Chloride.

A simple, economical and metal-free approach to the synthesis of 2-substituted benzoxazoles and 2-substituted benzothiazoles from 2-aminophenols, 2-aminothiophenols and DMF derivatives, only using imidazolium chloride (50% mmol) as promoter without any other additive, was reported. Various 2-substituted benzoxazoles and 2-substituted benzothiazoles were thus prepared in moderate to excellent yields.

Imidazolium Chloride: An Efficient Catalyst for Transamidation of Primary Amines.

A highly efficient and convenient protocol of imidazolium chloride (30 mol %) catalyzed amidation of amines with moderate to excellent yields was reported. The protocol shows broad substrate scope for aromatic, aliphatic, and heterocyclic primary amines.

Synthesis and characterization of four process impurities in pazopanib.

Pazopanib (trade name Votrient®) is a potent and selective multi-targeted tyrosine kinase inhibitor that blocks tumor growth and inhibits angiogenesis. Based on a recently reported procedure, we herein report the first synthesis of four potential process impurities generated in the production of pazopanib. The structure of these impurities were synthesized and characterized by 1H NMR, 13C NMR and HRMS data. The possible formation mechanisms of these impurities were also elucidated. These findings should be useful for the quality control of pazopanib in manufacture.

Design, synthesis and biological evaluation of novel tetrahydroisoquinoline quaternary derivatives as peripheral κ-opioid receptor agonists.

A novel series of tetrahydroisoquinoline quaternary derivatives 4 were synthesized as peripheral κ-opioid receptor agonists. All the target compounds were evaluated in κ-opioid receptor binding assays, and compounds 4l, 4m, and 4n exhibited high affinity for κ-opioid receptor. Furthermore, compound 4l (κKi=0.94nM) produced potent antinociceptive activity in the mouse acetic acid-induced writhing assay, with lower sedative side effects than the parent compound MB-1c.

Discovery, stereospecific characterization and peripheral modification of 1-(pyrrolidin-1-ylmethyl)-2-[(6-chloro-3-oxo-indan)-formyl]-1,2,3,4-tetrahydroisoquinolines as novel selective κ opioid receptor agonists.

A novel series of 1-(pyrrolidin-1-ylmethyl)-2-[(3-oxo-indan)-formyl]-1,2,3,4-tetrahydroisoquinoline derivatives maj-3a-maj-3u were synthesized and evaluated in vitro for their binding affinity at κ-opioid receptors. Maj-3c displayed the highest affinity for κ-opioid receptors (Ki = 0.033 nM) among all the compounds evaluated. Furthermore, all four stereoisomers of compound 3c were prepared, and (1S,18S)-3c was identified as the most potent (Ki = 0.0059 nM) κ-opioid receptor agonist among the four stereoisomers. Maj-3c produced significant antinociception (ED50 = 0.000406 mg kg(-1)) compared to U-50,488H and original BRL 52580 in the acetic acid writhing assay, but its strong sedative effect (ED50 = 0.000568 mg kg(-1)) observed in the mouse rotation test reduced its druggability. To minimize the central nervous system side effects, a series of hydroxyl-containing analogs of maj-3c were synthesized, and maj-11a was found to be a potent κ-opioid receptor agonist (Ki = 35.13 nM). More importantly, the dose for the sedative effect (ED50 = 9.29 mg kg(-1)) of maj-11a was significantly higher than its analgesic dose (ED50 = 0.392 mg kg(-1)), which made it a promising peripheral analgesic candidate compound with weak sedative side effects.

N76-1, a novel CDK7 inhibitor, exhibits potent anti-cancer effects in triple negative breast cancer.

Emerging evidence suggests that genetically highly specific triple-negative breast cancer (TNBC) possesses a relatively uniform transcriptional program that is abnormally dependent on cyclin-dependent kinase 7 (CDK7). In this study, we obtained an inhibitor of CDK7, N76-1, by attaching the side chain of the covalent CDK7 inhibitor THZ1 to the core of the anaplastic lymphoma kinase inhibitor ceritinib. This study aimed to elucidate the role and underlying mechanism of N76-1 in TNBC and evaluate its potential value as an anti-TNBC drug. The results of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and colony formation assays showed that N76-1 inhibited the viability of TNBC cells. Kinase activity and cellular thermal shift assays showed that N76-1 directly targeted CDK7. Flow cytometry results revealed that N76-1 induced apoptosis and cell cycle arrest in the G2/M phase. N76-1 also effectively inhibited the migration of TNBC cells by high-content detection. The RNA-seq analysis showed that the transcription of genes, especially those related to transcriptional regulation and cell cycle, was suppressed after N76-1 treatment. Moreover, N76-1 markedly inhibited the growth of TNBC xenografts and phosphorylation of RNAPII in tumor tissues. In summary, N76-1 exerts potent anticancer effects in TNBC by inhibiting CDK7 and provides a new strategy and research basis for the development of new drugs for TNBC.

Effects of Mono-2-ethylhexyl Phthalate on the Neural Transmission of PNs in Drosophila Antennal Lobe.

Long-term exposure to different types of chemicals is hazardous to human health. Di(2-ethylhexyl) phthalate (DEHP) could exert pleiotropic deleterious effects on nervous systems. Mono(2-ethylhexyl) phthalate (MEHP), as one of the most toxic metabolites of DEHP, may have similar effects on nervous systems. However, no effects of MEHP on neural circuits have been reported. To uncover the regulation of MEHP on neural transmission, the functional changes of neural excitability and synaptic plasticity of projection neurons (PNs) have been assessed. In the current study, we recorded the action potentials (APs), stimulate action potentials (sti-APs), mini excitement postsynaptic current (mEPSC), calcium currents, and sodium currents from PNs of isolated whole brain of Drosophila model utilizing patch clamp recordings. We found that MEHP-300 (at the concentration of 300 µM), but not MHEP-100 (at the concentration of 100 µM), significantly decreased the frequency and amplitude of APs. Besides, the amplitude and anti-amplitude of sti-APs were reduced with the application of MEHP-300. Meanwhile, MEHP-300 reduced the frequency of mEPSC, but not the amplitude. Furthermore, MEHP-300 reduced the peak current densities of sodium and calcium channels. Therefore, our results indicated that MEHP could alter the neural excitability and synaptic plasticity of PNs by inhibiting the ion channels activities, revealing the potential modulation of MEHP on neural transmission of PNs.

Tetrazanbigen Derivatives as Peroxisome Proliferator-Activated Receptor Gamma (PPARγ) Partial Agonists: Design, Synthesis, Structure-Activity Relationship, and Anticancer Activities.

Tetrazanbigen (TNBG) is a novel sterol isoquinoline derivative with poor water solubility and moderate inhibitory effects on human cancer cell lines via lipoapoptosis induction. Herein, we developed a series of novel TNBG analogues with improved water solubility and antiproliferative activities. The CCK-8 assay enabled us to identify a novel compound, 14g, which strongly inhibited HepG2 and A549 cell growth with IC50 values of 0.54 and 0.47 µM, respectively. The anticancer effects might be explained by the partial activation and upregulation of PPARγ expression, as indicated by the transactivation assay and western blotting evaluation. Furthermore, the in vitro antiproliferative activity was verified in an in vivo xenograft model in which 14g strongly reduced tumor growth at a dose of 10 mg/kg. In line with these positive observations, 14g exhibited an excellent water solubility of 31.4 mg/mL, which was more than 1000-fold higher than that of TNBG (4 µg/mL). Together, these results suggest that 14g is a promising anticancer therapeutic that deserves further investigation.

PTEN inhibition leads to the development of resistance to novel isoquinoline derivative TNBG-5602 in human liver cancer cells.

TNBG-5602, a new synthesized derivative of tetrazanbigen, is a potential chemotherapeutic agent against cancer. However, its underlying mechanism is complex and still unknown. In this investigation, the anticancer effects of TNBG-5602 were determined in vitro and in vivo. Small RNA retroviral library plasmids that overexpress 19-bp fragments were used to generate TNBG-5602-resistant cells. After validation, the overexpressed 19-bp fragments were sequenced using next-generation sequencing (NGS) in the drug-resistant cells. Furthermore, the relationship of TNBG-5602, phosphatase and tensin homolog deleted on Chromosome 10 (PTEN), and the phosphatidylinositol 3 kinase (PI3K)/protein kinase B (Akt) pathway was explored. The results showed that TNBG-5602 can effectively inhibit cancer cell proliferation and induce apoptosis in vitro and in vivo. Drug-resistant cells were screened using the small RNA library. Compared with naïve cells, drug-resistant cells were more resistant to TNBG-5602 in vitro and in vivo. NGS results revealed that the second highest overexpressed 19-bp fragment perfectly matched the PTEN gene, so the expression of PTEN in various cells and tissues was verified. Further research showed that exogenous overexpression of PTEN strengthened the anticancer effects of TNBG-5602 on p-Akt expression, whereas silencing of PTEN weakened these effects in naïve cells. Taken together, by using this library, we confirmed that PTEN is the target gene to the anticancer effects of TNBG-5602 via the PI3K/Akt pathway.

Discovery of 2,4-pyrimidinediamine derivatives as potent dual inhibitors of ALK and HDAC.

Combination of anaplastic lymphoma kinase (ALK) inhibitor with histone deacetylases (HDAC) inhibitor could exert synergistically anti-proliferative effects on ALK positive non-small cell lung cancer (NSCLC) naïve or resistant cells. In this work, we designed and synthesized a series of 2,4-pyrimidinediamine derivatives as dual ALK and HDAC inhibitors based on pharmacophore merged strategy. Among which, compound 10f displayed the most potent and balanced inhibitory activity against ALK (IC50 = 2.1 nM) and HDAC1 (IC50 = 7.9 nM), respectively. In particular, 10f was also potent against the frequently observed Crizotinib-resistant ALKL1196M (IC50 = 1.7 nM) as well as the Ceritinib-resistant ALKG1202R (IC50 = 0.4 nM) mutants. In antiproliferative activity assay, 10f exhibited impressive activity on ALK-addicted cancer cell lines at low micromole concentrations, which was comparable to that of Crizotinib and Ceritinib. Further flow cytometric analysis indicated that 10f could effectively induce cell death via cell apoptosis and cell cycle arrest. Taken together, these results suggested 10f would be a promising lead compound for the ALK-positive NSCLC treatment, especially the Ceritinib- or Crizotinib-resistant NSCLC.