preparation-characterization-lipophilic-doxorubicin-pro-drug

Preparation and Characterization of Lipophilic Doxorubicin Pro-drug Micelles

Micelles have been successfully used for the delivery of anticancer drugs. Amphiphilic polymers form core-shell structured micelles in an aqueous ...

Hampton University

A kinetic model was developed to describe recombinant human protein disulphide isomerase (rhPDI)-assisted folding of a substrate protein, C125A recombinant human interleukin-2 (C125A rhIL-2). A series of progress curves showing native C125A rhIL-2 formation under different reaction conditions were generated. Non-linear regression analysis of the progress curves of rhPDI-assisted C125A rhIL-2 folding was used to fit the differential equations of the described kinetic models. The goodness-of-fit of the model to the experimental datasets was used to support or exclude a particular kinetic model of rhPDI-assisted C125A rhIL-2 folding. The results suggest that the formation of native C125A rhIL-2 results from both glutathione-dependent oxidative folding and rhPDI-catalysed folding reactions. During oxidative folding of C125A rhIL-2, both rhPDI and reduced C125A rhIL-2 aggregated in folding buffer. The aggregation rates of rhPDI and C125A rhIL-2 followed second-order kinetics. Guanidinium chloride inactivated rhPDI but also decreased the aggregation of reduced C125A rhIL-2. These results demonstrate that during rhPDI-assisted C125A rhIL-2 folding, reduced C125A rhIL-2 aggregation competes with the productive folding pathway. While rhPDI enhances the oxidative folding of C125A rhIL-2, inactivation of rhPDI by the residual guanidinium chloride compromises its catalytic efficiency. The established model can be used to optimize the folding components in the folding mixture, and thus improve the folding efficiency.

Kinetic study of recombinant human protein disulphide isomerase-assisted C125A recombinant human interleukin-2 folding.

Initial velocity enzyme kinetics was used to study the inhibition mechanism of guanidine hydrochloride (Gdm.Cl) on catalytic activity of recombinant human protein disulfide isomerase (rhPDI) in protein folding. Reduced C125A recombinant human interleukin 2 (C125A rhIL-2), the substrate, was dissolved in 8 M Gdm.Cl before it was diluted into the folding buffer to initiate the folding reactions. The final Gdm.Cl concentrations in the folding buffer were fixed at 0.2 M, 0.4 M, 0.6 M and 0.8 M. The reduced and native C125A rhIL-2 were resolved by reversed phase-high performance liquid chromatography (RP-HPLC). The simultaneous nonlinear fitting of the initial velocities of the native C125A rhIL-2 formation vs the reduced C125A rhIL-2 concentrations in the presence of different Gdm.Cl concentrations shows that the inhibition mechanism of Gdm.Cl on the catalytic activities of rhPDI is a mixed-type noncompetitive nonlinear inhibition.

The inhibition mechanism of guanidine hydrochloride on the catalytic activity of recombinant human protein disulfide isomerase.

Recent studies show that quantitative and qualitative differences in amyloid beta (Abeta ) peptides may be implicated in the development of Alzheimer's disease. New evidence seems to support the existence of a dynamic equilibrium between Abeta peptide in the brain and peripheral blood circulation. The quantitation of Abeta in the blood may allow the development of the potential value of Abeta peptides as a biomarker in the development of Alzheimer's disease. In this communication, quantitation of Abeta peptides using high-performance liquid chromatography coupled with tandem mass spectrometry in a linear ion trap mode is presented. RP-HPLC was performed using a Waters Xterra MS C8 column (3.0 mm x 150 mm). Abeta(1-40) peptide was eluted using a gradient elution program. Eluate from the RP-HPLC column was split to both the UV detector and electrospray ionization MS source. The product ion scan was performed in a linear ion trap mode utilizing the transition of a multiply charged molecular ion of Abeta(1-40) to a singly charged product ion. The detection limit of 31.25 ng in column load using a 3.0-mm-diameter conventional C8 column was achieved. The Abeta(1-40) standard calibration curves show excellent linearity from 34 ng to 2500 ng Abeta(1-40) of column sample load. The product ion scan enhances sensitivity 10 times compared with the best previously achieved by a single-quadrupole instrument in the selective ion monitoring mode. Moreover, the product ion scan of Abeta(1-40) provides superior selectivity and specificity, which is very important in the quantitation of Abeta(1-40) in a complex biological matrix.

High-performance liquid chromatography coupled with tandem mass spectrometry for the detection of amyloid Beta peptide related with Alzheimer's disease.

Our objective was to design a polymeric micelle-based doxorubicin and lapatinib combination therapy for treating multidrug resistant (MDR) breast cancers. Poly(ethylene glycol)-block-poly(2-methyl-2-benzoxycarbonylpropylene carbonate) (PEG-PBC) polymers were synthesized for preparing doxorubicin and lapatinib loaded micelles using a film dispersion method. Micelles were characterized by determining critical micelle concentration (CMC), particle size distribution, and drug loading. The anticancer effects were determined in vitro with MTT assays as well as with lactate dehydrogenase (LDH) release studies. In addition, the cellular uptake of drug-loaded micelles was determined with fluorescence microscopy and flow cytometry. Finally, in vivo anticancer activity and tolerance of developed formulations were evaluated in resistant breast tumor bearing mice. PEG5K-PBC7K polymer synthesized in this study had a low CMC value (1.5 mg/L) indicating an excellent dynamic stability. PEG-PBC micelles could efficiently load both doxorubicin and lapatinib drugs with a loading density of 21% and 8.4%, respectively. The mean particle size of these micelles was 100 nm and was not affected by drug loading. The use of lapatinib as an adjuvant sensitized drug resistant MCF-7/ADR cells to doxorubicin treatment. Cellular uptake studies showed enhanced doxorubicin accumulation in MCF-7/ADR cells in the presence of lapatinib. The doxorubicin and lapatinib combination therapy showed a significant decrease in tumor growth compared to doxorubicin monotherapy. In conclusion, we have developed PEG-PBC micelle formulations for the delivery of doxorubicin and lapatinib. The combination therapy of doxorubicin plus lapatinib has a great potential for treating MDR breast cancer.

Doxorubicin and lapatinib combination nanomedicine for treating resistant breast cancer.

The objective of this study was to determine the compatibility of amiodarone hydrochloride with vasopressin during simulated Y-site administration. Amiodarone hydrochloride is compatible with vasopressin under simulated-use conditions when administered according to the Advanced Cardiac Life Support Guidelines. A simulated-use approach is a more appropriate way to evaluate compatibility of drug solutions and to assure safe and adequate drug delivery to the patient, particularly when the drug being evaluated is well-documented to have physical or chemical incompatibility concerns with various drugs. When amiodarone hydrochloride (50 mg/mL) was diluted to 6 mg/mL with 5% dextrose injection, then mixed with an equal volume of vasopressin (0.2 units/mL in normal saline) in a glass test tube, no visual incompatibility was observed for up to 24 hours. The purpose of our study was to assess compatibility of the two drugs under simulated-use conditions and to measure the recovery of amiodarone by high-performance liquid chromatography after sequential administration of vasopressin and amiodarone hydrochloride according to the Advanced Cardiac Life Support Guidelines.

Compatibility of amiodarone hydrochloride with vasopressin during simulated Y-site administration.

Users of the illicit drug, 3,4-methylenedioxymethamphetamine (MDMA), show signs of neurotoxicity. However, the precise mechanism of neurotoxicity caused by use of MDMA has not yet been elucidated. Synthetic glutathione (GSH) conjugates of MDMA are transported into the brain by the GSH transporter and subsequently produce neurotoxicity. The objective of this research is to show direct evidence of the formation of GSH adducts of MDMA in human hepatocytes. High-performance liquid chromatography coupled with tandem mass spectrometry was utilized to examine in vitro incubations of MDMA with cryopreserved human hepatocytes. The use of hydrophilic liquid chromatography in combination with linear ion trap mass spectrometry permitted the identification of two possible GSH metabolites. Enhanced product ion scans of m/z = 499 and 487 amu of extracts from hepatocytes treated with 1.0 mM MDMA show a distinct fragmentation pattern (m/z 194.2, 163, 135, 105), suggesting the formation of MDMA-GSH conjugate, MDMA-SG and 3,4-dihydroxymethamphetamine-SG. The formation of an MDMA-GSH conjugate was further supported by the apparent lack of the same fragmentation pattern from hepatocyte samples without MDMA treatment. The results generated from this study yield valuable qualitative and quantitative information about the neurotoxic thioether metabolites formed from MDMA in humans.

In vitro metabolism of 3,4-methylenedioxymethamphetamine in human hepatocytes.

Genetic information, such as single nucleotide polymorphism (SNP) data, has been widely recognized as useful in prediction of disease risk. However, how to model the genetic data that is often categorical in disease class prediction is complex and challenging. In this paper, we propose a novel class of nonlinear threshold index logistic models to deal with the complex, nonlinear effects of categorical/discrete SNP covariates for Schizophrenia class prediction. A maximum likelihood methodology is suggested to estimate the unknown parameters in the models. Simulation studies demonstrate that the proposed methodology works viably well for moderate-size samples. The suggested approach is therefore applied to the analysis of the Schizophrenia classification by using a real set of SNP data from Western Australian Family Study of Schizophrenia (WAFSS). Our empirical findings provide evidence that the proposed nonlinear models well outperform the widely used linear and tree based logistic regression models in class prediction of schizophrenia risk with SNP data in terms of both Types I/II error rates and ROC curves.

Article	Year
Kinetic study of recombinant human protein disulphide isomerase-assisted C125A recombinant human interleukin-2 folding. Biotechnology and applied biochemistry\| PubMed ID: 12630900	2003
The inhibition mechanism of guanidine hydrochloride on the catalytic activity of recombinant human protein disulfide isomerase. Journal of enzyme inhibition and medicinal chemistry\| PubMed ID: 12751820	2003
High-performance liquid chromatography coupled with tandem mass spectrometry for the detection of amyloid Beta peptide related with Alzheimer's disease. Journal of biomolecular techniques : JBT\| PubMed ID: 16522857	2005
Doxorubicin and lapatinib combination nanomedicine for treating resistant breast cancer. Molecular pharmaceutics\| PubMed ID: 24405470	2014
Compatibility of amiodarone hydrochloride with vasopressin during simulated Y-site administration. International journal of pharmaceutical compounding\| PubMed ID: 24459790	2013
In vitro metabolism of 3,4-methylenedioxymethamphetamine in human hepatocytes. Journal of analytical toxicology\| PubMed ID: 24682111	2014
Analysis of schizophrenia data using a nonlinear threshold index logistic model. PloS one\| PubMed ID: 25330160	2014

Chengan Du

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