PET imaging with [18F]FP-(+)-DTBZ in 6-OHDA-induced partial and full unilaterally-lesioned model rats of Parkinson's disease and the correlations to the biological data.

OBJECTIVE: The deficit of dopaminergic neurons in the nigrostriatal pathway is one of the pathological features of Parkinson's disease (PD). The decline of vesicular monoamine transporter type 2 (VMAT2) has been verified to relate with the severity of PD. The purpose of this study was to evaluate the ability of [18F]fluoropropyl-(+)-dihydrotetrabenazine ([18F]FP-(+)-DTBZ) to detect dopaminergic neuron dysfunction in a standard rat model of PD using PET imaging. Specifically, two different doses of 6-hydroxydopamine (6-OHDA) were injected unilaterally into the medial forebrain bundle (MFB) to create the models with two different severities. METHODS: Male Sprague-Dawley rats were intracranially injected with 8 µg 6-OHDA (partial lesion group), 16 µg 6-OHDA (full lesion group) and vehicle (sham group) into MFB, respectively. Thirty minutes static [18F]FP-(+)-DTBZ microPET scanning was performed to determine the dopaminergic neuron integrity on the 28th day post-injection and the behavioral tests were carried out in the next two days. Then, the rats were decapitated, and the brains were collected for biogenic amines content analysis or dissected for autoradiography and immunohistochemical (IHC) staining. The correlations of PET results to the behavioral, biological, histological, autoradiography results were analyzed, respectively. RESULTS: The standardized uptake value ratio (ST to CB) of [18F]FP-(+)-DTBZ in the ipsilateral striata decreased significantly in partial lesion group and full lesion group. Compared with the sham group, the ratio of the standardized uptake value in ipsilateral striatum to that in contralateral striatum decreased by 57.09 ± 2.30% (full lesion group) and 25.31 ± 5.70% (partial lesion group), respectively. The dopaminergic neuronal dysfunction was corroborated by in vitro autoradiography, IHC, and quantitative analysis of DA as well as its metabolites concentration tests. The motor function impairments of 6-OHDA-treated animals were manifested by a series of behavioral tests. The results of microPET imaging were linearly correlated with behavioral, biological, histological, and autoradiography results, respectively. CONCLUSION: Our data suggest that [18F]FP-(+)-DTBZ may be useful for detecting different degrees of dopaminergic neuronal lesions by PET imaging in PD models induced by 6-OHDA.

Insights into anticancer activity and mechanism of action of a ruthenium(II) complex in human esophageal squamous carcinoma EC109 cells.

A ruthenium(II) complex [Ru(p-cymene)(NHC)Cl2] (NHC=1,3-bis(4-(tert-butyl)benzylimidazol-2-ylidene), referred to as L-4, has been designed and synthesized recently in order to look for new anticancer drugs with high efficacy and low side effects. The anticancer activity and mechanism of action of L-4 in human esophageal squamous carcinoma EC109 cells were systematically investigated. The results revealed that L-4 exerted strong inhibitory effect on the proliferation of EC109 cells, and it arrested EC109 cells at G2/M phase, accompanied with the up-regulation of p53 and p21 and the down-regulation of cyclin D1. The results also showed that the reactive oxygen species (ROS)-dependent apoptosis of EC109 can be induced by L-4 via inhibiting the activity of glutathione reductase (GR), decreasing the ratio of glutathione to oxidized glutathione (GSH/GSSG), and leading to the generation of reactive oxygen species. The mitochondria-mediated apoptosis of EC109 induced by L-4 was also observed from the increase of Bax/Bcl-2 ratio, overload of Ca(2+), disruption of mitochondrial membrane potential (MMP), redistribution of cytochrome c, and activation of caspase-3/-9. However, the effects of L-4 on the cell viability, GR activity, GSH/GSSG ratio, reactive oxygen species level, mitochondria dysfunction and apoptosis induction were remarkably attenuated by adding the reactive oxygen species scavenger, NAC. Therefore, it was concluded that L-4 can inhibit the proliferation of EC109 cells via blocking cell cycle progression and inducing reactive oxygen species-dependent and mitochondria-mediated apoptosis. These findings suggested that the ruthenium(II) complex might be a potential effective chemotherapeutic agent for human esophageal squamous carcinoma (ESCC) and worthy of further investigation.

Synthesis and biological evaluation of novel platinum complexes of imidazolyl-containing bisphosphonates as potential anticancer agents.

Four novel platinum complexes, [Pt(en)]2ZL (1), [Pt(en)]2IPrBP (2), [Pt(en)]2MIBP (3) and [Pt(en)]2EIBP (4) [en = ethylenediamine; ZL = 1-hydroxy-3-(1H-imidazol-1-yl)ethane-1,1-diylbisphosphonic acid, commonly known as zoledronic acid; IPrBP = 1-hydroxy-3-(1H-imidazol-1-yl)propane-1,1-diylbisphosphonic acid; MIBP = 1-hydroxy-2-(2-methyl-1H-imidazol-1-yl)ethane-1,1-diylbisphosphonic acid; EIBP = 1-hydroxy-2-(2-ethyl-1H-imidazol-1-yl)ethane-1,1-diylbisphosphonic acid], were prepared and evaluated against five human cancer cell lines, including U2OS, A549, HCT116, MDA-MB-231 and HepG2. While exhibiting lower efficacy on the inhibition of cancer cell lines than cisplatin (CDDP), four complexes showed higher cytotoxicity than the corresponding ligands and relatively stronger cytotoxic effect on the hepatoma cell lines HepG2, and the complex 1 showed higher cytotoxicity than others on the whole. These complexes have better selectivity than the corresponding ligands in inhibiting hepatocarcinoma cells rather than normal liver cells, and the selective inhibitory effect of the complex 1 at the high concentration (100 µM) is better than that at the low concentration. Morphology studies exhibited typical characteristics of cell apoptosis and the cell cycle distribution analysis indicated that the complexes can inhibit cancer cells by inducing the cell cycle arrest at the G2/M phase, exhibiting a similar mechanism of action to CDDP. The binding interaction of complex with DNA has been explored by circular dichroism (CD) and UV-Vis absorption spectra, demonstrating these new complexes have moderate binding affinity for DNA.

Synthesis, crystal structure and antitumor effect of a novel copper(II) complex bearing zoledronic acid derivative.

A great majority of Cu(II) complexes currently studied in the anticancer research field exert their antiproliferative activities through ligand exchange. In this work, we present the synthesis and structural characterization of two novel Cu(II) complexes, {[Cu3(ZL)2(H2O)6]·6H2O}n (1) (ZL = 1-hydroxy-2-(1H-imidazol-1-yl)ethane-1,1-diyldiphosphonic acid) and [Cu(IPrDP)2]·3H2O (2) (IPrDP = 1-hydroxy-3-(1H-imidazol-1-yl)propane-1,1-diyldiphosphonic acid). Due to the insolubility of polymer 1 in common solvents, only the biological activities of complex 2 were investigated. The antitumor activity of complex 2 was evaluated against a panel of human cancer cell lines, including U2OS, A549, HCT116, MDA-MB-231 and HepG2. Complex 2 exhibited comparable cytotoxic effect to cisplatin (CDDP) against the human colon carcinoma cells HCT116, and superior selectivity for inhibiting human hepatocarcinoma cells rather than normal liver cells. The cell cycle distribution analysis indicates that complex 2 inhibits human carcinoma cells by inducing the cell cycle arrest at the G2/M phase, showing a similar mechanism of action to that of CDDP. The binding interaction of complex 2 with calf thymus DNA (CT-DNA) has been explored by UV-vis absorption and circular dichroism (CD), demonstrating complex 2 has a moderate binding affinity for DNA through intercalation.

Generation of a liver targeting fusion interferon and its bioactivity analysis in vitro.

The objective of this study was to generate a liver targeting fusion interferon, galactosyl-human serum albumin-interferon alpha2b (G-HSA-IFN) and to evaluate its bioactivity in vitro on HepG2.2.15 cells which express hepatitis B surface antigen (HBsAg) and hepatitis B e antigen (HBeAg). The cell proliferation was determined by Sulpho Rhodamine B (SRB) staining method and flow cytometry (FCM) assay. Hochest33342 and Propidium Iodide (PI) double staining and Western blot analysis of Bcl-2/Bax were also performed to evaluate cell lethality and apoptosis. The concentrations of HBsAg and HBeAg secreted in culture supernatant were detected using Enzyme-Linked Immunosorbent Assay (ELISA). The results demonstrated that G-HSA-IFN could inhibit the proliferation of HepG2.2.15 cells and the cell cycle was arrested at G0/G1 phase. Western blotting results showed that the expression of Bcl-2 was inhibited in a dose-dependent manner while the expression of Bax was enhanced. The expression of HBsAg was inhibited by G-HSA-IFN in a dose-dependent manner, while no significant inhibiting effect on the expression of HBeAg was observed. Conclusively, G-HSA-IFN could not only significantly inhibit the HBsAg expression and the proliferation of HepG2.2.15 cells, but also induce the apoptosis of the target cells, rendering it a promising drug candidate for hepatitis B.

Preparation of 99mTc-PQQ and preliminary biological evaluation for the NMDA receptor.

Pyrroloquinoline quinone (PQQ), an essential nutrient, antioxidant, redox modulator and nerve growth factor found in a class of enzymes called quinoproteins, was labeled with 99mTc by using stannous fluoride (SnF2) method. Radiolabeling qualification, quality control and characterization of 99mTc-PQQ and its biodistribution studies in mice were performed and discussed. Effects of pH values, temperature, time and reducing agents concentration on the radiolabeling yield were investigated. The quality control procedure of 99mTc-PQQ was determined by thin layer chromatography (TLC), radio high-performance liquid chromatography (RHPLC) and paper electrophoresis methods. The average radiolabeling yield was 94 ± 1% under optimum conditions of 0.99 mg of PQQ, 30 µg of SnF2, 0.5 mg of ethylenediaminetetraacetic acid disodium salt (EDTA-2Na) and 18.5 MBq of Na99mTcO4 at pH 6 and 25 °C with a response volume of 1 ± 0.1 mL. 99mTc-PQQ was stable and anionic. Lipid-water partition coefficient of 99mTc-PQQ was -1.49 ± 0.16. The pharmacokinetics parameters of 99mTc-PQQ were t1/2α = 18.16 min, t1/2ß = 100.45 min, K12 = 0.013 min-1, K21 = 0.017 min-1, Ke = 0.016 min-1, AUC (area under the curve) = 1040.78 ID% g-1 min and CL (plasma clearance) = 0.096 mL min-1. The dual-exponential equation was Y = 10.88e-0.038t  + 5.21e-0.0069t . The biodistribution of 99mTc-PQQ was studied in ICR (Institute for Cancer Research 7701 Burhelme Are., Fox Chase, Philadelphia, PA 1911 USA) mice. In vitro autoradiographic studies clearly showed that the 99mTc-PQQ radioactivity accumulated predominantly in the hippocampus and cortex, which had a high density of N-methyl-d-aspartate Receptor (NMDAR). The enrichment can be blocked by NMDAR redox modulatory site antagonists-ebselen (EB) and 99mTc-PQQ is therefore a promising candidate for the molecular imaging of NMDAR. To date, however, there have been no studies characterizing 99mTc-PQQ.

[Influence of betulinic acid on proliferation, migration, cell cycle and apoptosis of pancreatic cancer cells].

OBJECTIVE: To investigate the effect of betulinic acid (BA) on the proliferation, migration, apoptosis and cell cycle of pancreatic cancer cells (BxPC-3) in vitro and elucidate the underlying. METHOD: The effect of BA on the proliferation of BxPC-3 was measured by using sulforhodamine B (SRB) assay. Migratory ability of BxPC3 cells were detected by wound healing assay, and the morphological change was observed with light microscope. The influence of BA on cell cycle of BxPC-3 cells was tested by flow cytometry (FCM). Apoptosis was analyzed by using Hochest33342-PI double staining. Western blot technologies were applied to detect the expression of Bcl-2 and Bax. RESULT: BA exhibited significant cell proliferation and migration inhibition, as well as its potency of inducing apoptosis in BxPC-3 cells in vitro in a dose-dependent manner. The IC50 value for 72 h was 16.54 mg x L(-1). Cell migration was significantly inhibited at 5 mg x L(-1) of BA. Cells treated with BA showed increased cell population in G0 phase, with decreased G2/M phase population. The expression of Bax and Bcl-2 was up and down-regulated respectively in BA-treated BxPC-3 cells in a dose-dependent manner. CONCLUSION: BA exerted potent effect on growth inhibition, G0 cell cycle arrest and induction of apoptosis in BxPC-3 cells in vitro, possibly associated with the down-regulation of Bcl-2 and up-regulation of Bax expression. The potent antitumor capacity of BA suggested that it could be a promising new anticancer agent in human pancreatic cancer treatment.