The sub-chronic impact of mPEG2k-PCLx polymeric nanocarriers on cytochrome P450 enzymes after intravenous administration in rats.

Recent studies indicated obvious impacts of nano-carriers on cytochrome P450 enzymes (CYP450s) in vitro, but the effects in vivo are still unknown. In the present research, mPEG2k-PCLx micelles with different length of hydrophobic block (2000-10,000 Da) were intravenously administrated into rats for 14 days to evaluate the sub-chronic influences in the metabolic function of hepatic CYP450s. Although CYP1A1/B2 was susceptible to mPEG2k-PCLx micelles compared with other CYP isoenzymes, induction was mainly observed and varied with micelle type, administration dose, and CYP isoform. Interestingly, mPEG2k-PCL3.5k micelles at 5 mg/kg increased the activity of CYP1A2, CYP2B1, CYP2C6, CYP2C11, and CYP3A1/2 while mPEG2k-PCL5k micelles only induced the latter three enzymes at 75 mg/kg. The mRNA expression of corresponding CYPs was mostly up-regulated by mPEG2k-PCL3.5k micelles whilst less effect in protein level except for CYP3A1/2. Moreover, mPEG2k-PCL3.5k micelles could affect the pharmacokinetic properties of phenacetin (CYP1A2), tolbutamide (CYP2C6), omeprazole (CYP2C11), and midazolam (CYP3A1/2) with a decrease of 19.6% in Cmax, 20.5% in AUC0-t, 31.6% in AUC0-t, and 40.1% in Cmax at 5 mg/kg, respectively (P < 0.05 or P < 0.01). These results unveiled nano-DDS might be involved in nanocarrier-drug interaction by intervening in the activity of CYP450s.

In vitro effect of mPEG2k-PCLx micelles on rat liver cytochrome P450 enzymes.

The present study was aimed to evaluate the effects of amphiphilic copolymer micelles on six major hepatic cytochrome P450 (CYP) isoforms. A series of mPEG2k-PCLx polymeric micelles (mPEG2k-PCL2k, mPEG2k-PCL3.5k, mPEG2k-PCL5k and mPEG2k-PCL10k) ranging from 20 to 100 nm were prepared to investigate the inhibitory or inductive activities by in vitro incubations of rat liver microsomes and primary rat hepatocytes. Inhibition of these polymeric micelles on CYP1A2, CYP2B1, CYP2C6, CYP2C11, CYP2D2 and CYP3A1/2 isoenzymes were observed above their critical micelle concentrations (>10 µg·mL-1) and in a concentration-dependent manner. The mPEG2k-PCL2k micelles showed the strongest inhibition of CYP1A2, followed by CYP2C11. The micelles with lower molecular weight PCL segment exhibited more potent inhibitory potential. Induction on CYP1A2, CYP2B1 and CYP3A1/2 activity (2.1-7.2-fold, 1.5-2.4-fold and 1.3-3.0-fold, respectively) were detected at all tested concentrations (0.1-1000 µg·mL-1 or 0.1-100 µg·mL-1). Accordingly, most of the mRNA levels were upregulated. As demonstrated in ex vivo fluorescence imaging results, the mPEG2k-PCLx micelles mainly accumulated in the liver after intravenous administration. In conclusion, mPEG2k-PCLx micelles can interfere with the normal metabolic function of CYP450s in vitro, indicating polymeric micelles as promising drug nano-carriers might cause micelle-drug interaction and the in vivo interaction deserves further investigation.

LC-MS/MS assay of ropinirole in rat biological matrices: elimination of lysoglycerophosphocholines-based matrix effect.

AIM: To adequately support PK evaluation of ropinirole in rats following intranasal administration, it was desirable to determine ropinirole concentrations in rat plasma, brain homogenate and microdialysate. Results & methodology: A robust LC-MS/MS method has been developed for the determination of ropinirole in biological samples. Liquid-liquid extraction using ethyl acetate eliminated matrix effect due to significantly reduced levels of lysoglycerophosphocholines. The assay was fully validated with dynamic ranges of 0.01-20 ng/ml for plasma and brain homogenate samples and 0.1-200 ng/ml for microdialysate samples. CONCLUSION: The proposed method was accurate and precise for the quantification of ropinirole in biological samples and was successfully applied to a microdialysis study of ropinirole in rats.

Screening of stabilizers for LC-MS/MS analysis of clevidipine and its primary metabolite in dog whole blood.

BACKGROUND: Clevidipine is an ester-containing antihypertensive agent that undergoes rapid hydrolysis in blood. A reliable stabilizer cocktail containing citric acid and ascorbic acid was established and the LC-MS/MS method was validated for simultaneous determination of clevidipine and its major metabolite in beagle dog whole blood. RESULTS: The stabilizer could nearly completely inhibit the esterase activity. Both analytes were extracted from whole blood by toluene and detected by MS/MS in positive ESI mode. The linearity range was 0.1-100.0 ng/ml for clevidipine and 1.0-1000.0 ng/ml for the primary metabolite. CONCLUSION: The stabilizer cocktail was able to effectively suppress the activity of esterase in blood. The method was successfully applied to a PK study of clevidipine in beagle dogs.

Epidermal growth factor-ferritin H-chain protein nanoparticles for tumor active targeting.

Human ferritin H-chain protein (FTH1)-based nanoparticles possess a precisely assembled nanometer-scale structure and high safety. However, their applications for imaging and drug delivery towards cancer cells remain limited due to a lack of target specificity. Epidermal growth factor receptor (EGFR) is overexpressed in many malignant tissues including breast cancer, and has been used as a therapeutic target for cancer treatment. Herein, a genetic method is shown to generate EGF-FTH1 chimeric proteins. EGF-FTH1 nanoparticles with EGF on the surface are then produced. The data demonstrate that EGF-FTH1 nanoparticles, with a small size (11.8 ± 1.8 nm), narrow size distribution, and high biosafety, can specifically bind to and then be taken up by breast cancer MCF-7 cells and MDA-MB-231 cells, but not normal breast epithelial MCF-10A cells. In contrast, binding and absorption of nontargeted ferritin-based nanoparticles to breast cancer cells are negligible. In vivo studies show that EGF-FTH1 nanoparticles are accumulated in breast tumors in a mouse xenograft model. Interestingly, the concentration of EGF-FTH1 nanoparticles in the tumor site is significantly reduced when mice are pretreated with an excess of free EGF. These results imply that EGF-EGFR interaction plays an important role in regulating the tumor retention of EGF-FTH1 nanoparticles.

[Proteins in saliva and dental plaque fluid in relation to caries susceptibility].

OBJECTIVE: To investigate the relationship of the levels of proteins in parotid saliva, whole saliva and dental plaque fluid with caries susceptibility. METHODS: Sixty-six of university students were selected as subjects, 39 in caries-free group (CF, DMFS = 0) and 27 in caries-susceptible group (CS, DMFS >/= 8 and DT >/= 3). Total protein concentration was detected with Lowry method. Protein compositions were separated with SDS-PAGE and alkaline electrophoresis. The gels were analyzed using an image evaluation system. RESULTS: Total protein level in dental plaque fluid was about 10-fold higher than that in saliva. Whole saliva was closely related to dental plaque fluid in terms of proteins only in CF group (r = 0.804), but there was little relation in CS group. The proteins that occurred in all three fluids were 14 000, 66 000 and 76 000 proteins. The 14 000, 15 000 and 38 000 proteins level in dental plaque fluid and 14 000 protein level in whole saliva were significantly lower in CS group than in CF group. CONCLUSIONS: The proteins of dental plaque fluid are influenced significantly by whole saliva in CF group. The results suggest some kinds of proteins in dental plaque fluid and in whole saliva might play important roles against caries.