Synthesis and Antimicrobial Activity Evaluation of Pyridine Derivatives Containing Imidazo[2,1-b][1,3,4]Thiadiazole Moiety.

Four series of novel pyridine derivatives (17 a-i, 18 a-i, 19 a-e, and 20 a-e) were synthesized and their antimicrobial activities were evaluated. Of all the target compounds, almost half target compounds showed moderate or high antibacterial activity. The 4-F substituted compound 17 d (MIC=0.5 µg/mL) showed the highest antibacterial activity, its activity was twice the positive control compound gatifloxacin (MIC=1.0 µg/mL). For fungus ATCC 9763, the activities of compounds 17 a and 17 d are equivalent to the positive control compound fluconazole (MIC=8 µg/mL). Furthermore, compounds 17 a and 17 d showed little cytotoxicity to human LO2 cells, and did not show hemolysis even at ultra-high concentration (200 µM). The results indicate that these compounds are valuable for further development as antibacterial and antifungal agents.

Synthesis and Antibacterial Activity Evaluation of Imidazole Derivatives Containing 6-Methylpyridine Moiety.

A series of 2-cyclopropyl-5-(5-(6-methylpyridin-2-yl)-2-substituted-1H-imidazol-4-yl)-6-phenylimidazo[2,1-b][1,3,4]thiadiazoles (15a-t and 16a-f) were synthesized and their antibacterial activities were evaluated. More than half of the compounds showed moderate or strong antibacterial activity. Among them, compounds 15t (MIC=1-2 µg/mL) and 16d (MIC=0.5 µg/mL) showed the strongest antibacterial activities. Notably, compound 16d did not exhibit cytotoxicity in HepG2 cells and did not show hemolysis like the positive control compound Gatifloxacin. The results suggest that compound 16d should be further investigated as a candidate antibacterial agent.

Synthesis and antimicrobial activity evaluation of pyrazole derivatives containing the imidazo[2,1-b][1,3,4]thiadiazole moiety.

Four series of novel pyrazole derivatives (compounds 17a-m, 18a-m, 19a-g, and 20a-g) were synthesized, and their antibacterial and antifungal activities were evaluated. Most of the target compounds (17a-m, 18k-m, and 19b-g) showed strong antifungal activity and high selectivity relative to both Gram-positive and Gram-negative bacteria. Among them, compounds 17l (minimum inhibitory concentration [MIC] = 0.25 µg/mL) and 17m (MIC = 0.25 µg/mL) showed the strongest antifungal activity, being 2- and 4-fold more active than the positive controls gatifloxacin and fluconazole, respectively. In particular, compound 17l showed little cytotoxicity against human LO2 cells and did not exhibit hemolysis at ultrahigh concentrations, as did the positive control compounds gatifloxacin and fluconazole. These results indicate that these compounds are valuable for further development as antifungal agents.

Synthesis and biological evaluation of novel benzo[c][1,2,5]thiadiazol-5-yl and thieno[3,2-c]- pyridin-2-yl imidazole derivatives as ALK5 inhibitors.

Transforming growth factor (TGF-ß), a key mediator of tumor growth and metastasis, has been recognized as an important cancer drug target. A series of benzo[c][1,2,5]thiadiazol-5-yl imidazoles (14a-g) and thieno[3,2-c]-pyridin-2-yl imidazoles (20a-g) were designed, synthesized, and evaluated for their activin receptor-like kinase 5 (ALK5) activities. Among these compounds, 14c showed the highest activity (IC50 = 0.008 µM) against ALK5 kinase, which was 16.1-fold and 1.8-fold higher than those of positive control compounds LY-2157299 (IC50 = 0.129 µM) and EW-7197 (IC50 = 0.014 µM), respectively. Compound 14g (350) showed the highest selectivity index of ALK5 against p38α MAP kinase, which was significantly higher than that of positive control compounds LY-2157299 (4) and EW-7197 (211). The inhibitory effects of compound 14c on TGF-ß-induced Smad signaling and cell motility were studied in SPC-A1, HepG2 and HUVEC cells using western blot analysis and wound healing assay. ADMET prediction analysis showed that compounds 14c and 14g had good pharmacokinetics and drug-likeness behaviors.

MRI-visible liposome-polyethylenimine complexes for DNA delivery: preparation and evaluation.

To noninvasively monitor the effect of gene therapy and achieve an optimal therapeutic effect, liposomes encapsulated polyethylenimine (PEI)-coated superparamagnetic iron oxide nanoparticles (SPION) with dual functions of MRI diagnosis and gene therapy were prepared. SPION was synthesized via co-precipitation, and then modified with PEI via thiourea reaction. The liposomes encapsulating PEI-SPION (LP-PEI-SPION) were prepared by ethanol injection. Fourier transform infrared spectra confirmed that PEI was successfully modified onto SPION, and thermogravimetric analysis indicated that the PEI content was about 17.1%. The LP-PEI-SPION/DNA had a small particle size of 253.07 ± 0.90 nm. LP-PEI-SPION/DNA had low cytotoxicity with more than 80% of the cell survival rates and high transfection efficiency compared with Lipofectamine® 2000/DNA. Additionally, it also showed good MRI effect on three cell lines. The liposomes encapsulating PEI-SPION (lipopolyplexes) have been successfully prepared as MRI contrast agents and gene delivery vectors, which may have great theoretical research significance and clinical potentials. Abbreviations: PEI, polyethylenimine; SPION, superparamagnetic iron oxide nanoparticles; LP-PEI-SPION, liposomes encapsulating PEI-SPION; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide; ICP-MS, inductively coupled plasma mass spectrometry; XRD, X-ray diffraction; TEM, transmission electron microscope; TGA, thermogravimetric analysis; DOTAP, 1,2-dioleoyl-3-trimethylammonium-propane; DOPE, 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine; Chol, cholesterol.

Lipid-Albumin Nanoparticles (LAN) for Therapeutic Delivery of Antisense Oligonucleotide against HIF-1α.

Lipid-albumin nanoparticles (LAN) were synthesized for delivery of RX-0047, an antisense oligonucleotide (ASO) against the hypoxia inducible factor-1 alpha (HIF-1α) to solid tumor. These lipid nanoparticles (LNs) incorporated a human serum albumin-pentaethylenehexamine (HSA-PEHA) conjugate, which is cationic and can form electrostatic complexes with negatively charged oligonucleotides. The delivery efficiency of LAN-RX-0047 was investigated in KB cells and a KB murine xenograft model. When KB cells were treated with LAN-RX-0047, significant HIF-1α downregulation and enhanced cellular uptake were observed compared to LN-RX-0047. LN-RX-0047 and LAN-RX-0047 showed similar cytotoxicity against KB cells with IC50 values of 19.3 ± 3.8 and 20.1 ± 4.2 µM, respectively. LAN-RX-0047 was shown to be taken up by the cells via the macropinocytosis and caveolae-mediated endocytosis pathways while LN-RX-0047 was taken up by cells via caveolae-mediated endocytosis. In the KB xenograft tumor model, LAN-RX-0047 exhibited tumor suppressive activity and significantly reduced intratumoral HIF-1α expression compared to LN-RX-0047. Furthermore, LAN-RX-0047 greatly increased survival time of mice bearing KB-1 xenograft tumors at doses of either 3 mg/kg or 16 mg/kg. These results indicated that LAN-RX-0047 is a highly effective vehicle for therapeutic delivery of antisense agents to tumor.

CD33-Targeted Lipid Nanoparticles (aCD33LNs) for Therapeutic Delivery of GTI-2040 to Acute Myelogenous Leukemia.

CD33-targeted lipid nanoparticles (aCD33LNs) were synthesized for delivery of GTI-2040, an antisense oligonucleotide (ASO) against the R2 subunit of ribonucleotide reductase, to acute myelogenous leukemia (AML). These LNs incorporated a deoxycholate-polyethylenimine (DOC-PEI) conjugate, which has shown significant activity to facilitate oligonucleotide delivery. Anti-CD33 scFv (aCD33) was added as a targeting ligand. The delivery efficiency of this system was investigated both in vitro and in vivo. When cells were treated with aCD33LN/GTI-2040, significant uptake was observed in CD33 positive Kasumi-1 cells. aCD33LNs loaded with GTI-2040 induced significant down-regulation of R2 mRNA and protein levels in AML cells. Moreover, aCD33LN/GTI-2040 showed a 15-fold reduction in the IC50 of antileukemic drug Ara-C in Kasumi-1 cells. In Kasumi-1 xenograft model, aCD33LN/GTI-2040 showed significant R2 downregulation compared to LN/GTI-2040. Furthermore, aCD33LN/GTI-2040 coadministered with Ara-C was shown to be highly effective in tumor growth inhibition and to greatly increase survival time of mice bearing Kasumi-1 xenograft tumors. The conjugate DOC-PEI has shown an ability to include calcein release from lipid nanoparticles, suggesting a potential mechanism contributing to efficient endosome release by DOC-PEI2K. These results indicate that aCD33LNs are a highly effective vehicle for the therapeutic delivery of antisense agents to AML.

Construction of GPC3-modified Lipopolymer SiRNA Delivery System.

BACKGROUND: Gene therapy has been widely concerned because of its unique therapeutic mechanism. However, due to the lack of safe and effective carries, it has not been widely used in clinical practice. Glypican 3 (GPC3) is a highly specific proteoglycan for hepatocellular carcinoma and is a potential diagnostic and therapeutic target for hepatocellular carcinoma. Herein, to monitor the effect of gene therapy and enhance the transfection efficiency of gene carriers, GPC3-modified lipid polyethyleneimine-modified superparamagnetic nanoparticle (GLPS), a type of visualized carrier for siRNA (small-interfering RNA) targeting the liver, was prepared. METHODS: We performed in vitro gene silencing, cytotoxicity, and agarose gel electrophoresis to identify the optimal GLPS formulation. In vitro MRI and Prussian blue staining verified the liver-targeting function of GLPS. We also analyzed the biocompatibility of GLPS by co-culturing with rabbit red blood cells. Morphological changes were evaluated using HE staining. RESULTS: The GLPS optimal formulation consisted of LPS and siRNA at a mass ratio of 25:1 and LPS and DSPE-PEG-GPC3 at a molar ratio of 2:3. GLPS exhibited evident liver-targeting function. In vitro, we did not observe morphological changes in red blood cells or hemolysis after co-culture. In vivo, routine blood analysis revealed no abnormalities after GLPS injection. Moreover, the tissue morphology of the kidney, spleen, and liver was normal without injury or inflammation. CONCLUSION: GLPS could potentially serve as an effective carrier for liver-targeted MRI monitoring and siRNA delivery.

Synthesis and anti-liver fibrosis activity of imidazole and thiazole compounds containing amino acids.

Four series of imidazoles (15a-g, 20c, and 20d) and thiazoles (18a-g, 22a, and 22b) possessing various amino acids were synthesized and evaluated for activin receptor-like kinase 5 (ALK5) inhibitory activities in an enzymatic assay. Among them, compounds 15g and 18c showed the highest inhibitory activity against ALK5, with IC50 values of 0.017 and 0.025 µM, respectively. Compounds 15g and 18c efficiently inhibited extracellular matrix (ECM) deposition in TGF-ß-induced hepatic stellate cells (HSCs), and eventually suppressed HSC activation. Moreover, compound 15g showed a good pharmacokinetic (PK) profile with a favorable half-life (t1/2 = 9.14 h). The results indicated that these compounds exhibited activity targeting ALK5 and may have potential in the treatment of liver fibrosis; thus they are worthy of further study.

Preparation and evaluation of LA-PEG-SPION, a targeted MRI contrast agent for liver cancer.

This study aims to synthesize a magnetic resonance imaging (MRI) contrast agent that can specifically target the asialoglycoprotein receptor of liver cancer cells and evaluate its ability as a targeted MRI contrast agent. Lactobionic acid (LA) and polyethylene glycol (PEG) were used to modify superparamagnetic iron oxide nanoparticles (SPION) to obtain LA-PEG-SPION. LA-PEG-SPION was uniformly spherical under the electron microscope, with regular morphology and good dispersion. The particle size of LA-PEG-SPION was about 30 ± 4.5 nm, and its surface potential was about 31 ± 1.5 mV. LA-PEG-SPION had no toxicity or low toxicity to HepG2 cells and HeLa cells, even at 400 µg/mL. The uptake of LA-PEG-SPION by HepG2 cells was higher than that of SPION, with increased blue-stained particles. The fluorescent labeling rate of HepG2 cells reached 68.8%, which was higher than that of the control group. In vitro, MRI showed that the T2-weighted signal intensity of HepG2 cells was lower than that of the control group. Conclusively, LA-PEG-SPION nanoparticles are synthesized in a simple and efficient way. They are successfully applied to the T2-weighted contrast-enhanced MRI in liver cancer in vitro, and they have the potential to be used for in vivo research and clinical studies.