Modeling Sitagliptin Effect on Dipeptidyl Peptidase 4 (DPP4) Activity in Adults with Hematological Malignancies After Umbilical Cord Blood (UCB) Hematopoietic Cell Transplant (HCT)

Background and Objectives— Dipeptidyl peptidase-4 (DPP4) inhibition is a potential strategy to increase the engraftment rate of hematopoietic stem/progenitor cells. A recent clinical trial using sitagliptin, a DPP4 inhibitor approved for type 2 diabetes mellitus, has shown to be a promising approach in adults with hematological malignancies after umbilical cord blood (UCB) hematopoietic cell transplant (HCT). Based on data from this clinical trial, a semi-mechanistic model was developed to simultaneously describe DPP4 activity after multiple doses of sitagliptin in subjects with hematological malignancies after a single-unit UCB HCT. Methods— The clinical study included 24 patients that received myeloablative conditioning followed by 4 oral sitagliptin 600mg with single-unit UCB HCT. Using a nonlinear mixed effects approach, a semi-mechanistic pharmacokinetic/pharmacodynamic model was developed to describe DPP4 activity from this trial data using NONMEM 7.2. The model was used to drive Monte-Carlo simulations to probe various dosage schedules and the attendant DPP4 response. Results— The disposition of sitagliptin in plasma was best described by a 2-compartment model. The relationship between sitagliptin concentration and DPP4 activity was best described by an indirect response model with a negative feedback loop. Simulations showed that twice a day or three times a day dosage schedules were superior to once daily schedule for maximal DPP4 inhibition at the lowest sitagliptin exposure. Conclusion— This study provides the first pharmacokinetic/pharmacodynamic model of sitagliptin in the context of HCT, and provides a valuable tool for exploration of optimal dosing regimens, critical for improving time to engraftment in patients after UCB HCT

Development and validation of an HPLC-UV method for iodixanol quantification in human plasma

Iodixanol is a widely used iso-osmolar contrast medium agent. Similar to iohexol, it can also be a good exogenous marker for the measurement of glomerular filtration rate (GFR). This article describes the development and validation of an HPLC-UV method for quantification of iodixanol in human plasma. Internal standard, iohexol (20 μl, 1 mg/ml), and perchloric acid (30 μl, 20%, v/v) were added to plasma samples (300 μl), followed by neutralization with 10 μl potassium carbonate (5 M). Samples were centrifuged and 10 μl of the supernatant was injected onto a C18 EPS analytical column (3 μm particle size, 150 mm × 4.6 mm). The extraction method yielded \u3e95% recovery for both iodixanol and iohexol. The mobile phase consisted of 0.1% (w/v) sodium formate buffer and acetonitrile. Iohexol and iodixanol peaks were eluted at ∼5 and 9 min, respectively using a fast gradient method. The assay lower limit of detection was 2.0 μg/ml and lower limit of quantification was 10 μg/ml. The calibration curves, assessed in six replicates, were linear over an iodixanol concentration range of 10-750 μg/ml. Intra- and inter-day accuracy was \u3e95% and precision expressed as % coefficient of variation was \u3c10%. This method is simple, accurate, precise and robust and can potentially be used for iodixanol quantification in large-scale clinical studies. © 2008 Elsevier B.V. All rights reserved

Multidrug resistance-associated protein 2 (MRP2/ABCC2) haplotypes significantly affect the pharmacokinetics of tacrolimus in kidney transplant recipients

Background and Objective: Tacrolimus is an immunosuppressive drug used for the prevention of the allograft rejection in kidney transplant recipients. It exhibits a narrow therapeutic index and large pharmacokinetic variability. Tacrolimus is mainly metabolized by cytochrome P450 (CYP) 3A4 and 3A5 and effluxed via ATP-binding cassette (ABC) transporters such as P-glycoprotein (P-gp), encoded by ABCB1 gene. The influence of CYP3A53 on the pharmacokinetics of tacrolimus has been well characterized. On the other hand, the contribution of polymorphisms in other genes is controversial. In addition, the involvement of other efflux transporters than P-gp in tacrolimus disposition is uncertain. The present study was designed to investigate the effects of genetic polymorphisms of CYP3As and efflux transporters on the pharmacokinetics of tacrolimus. Subjects and Methods: A total of 500 blood concentrations of tacrolimus from 102 adult stable kidney transplant recipients were included in the analyses. Genetic polymorphisms in CYP3A4 and CYP3A5 genes were determined. In addition, the genes of efflux transporters including P-gp (ABCB1), multidrug resistance-associated protein (MRP2/ABCC2) and breast cancer resistance protein (BCRP/ABCG2) were genotyped. For ABCC2 gene, haplotypes were determined as follows: H1 (wild type), H2 (1249G\u3eA), H9 (3972C\u3eT) and H12 (-24C\u3eT and 3972C\u3eT). Population pharmacokinetic analysis was performed using nonlinear mixed effects modeling. Results: Analyses revealed that the CYP3A5 expressers (CYP3A51 carriers) and MRP2 high-activity group (ABCC2 H2/H2 and H1/H2) showed a decreased dose-normalized trough concentration of tacrolimus by 2.3-fold (p \u3c 0.001) and 1.5-fold (p = 0.007), respectively. The pharmacokinetics of tacrolimus were best described using a two-compartment model with first order absorption and an absorption lag time. In the population pharmacokinetic analysis, CYP3A5 expressers and MRP2 high-activity groups were identified as the significant covariates for tacrolimus apparent clearance expressed as 20.7 × (age/50)-0.78 × 2.03 (CYP3A5 expressers) × 1.40 (MRP2 high-activity group). No other CYP3A4, ABCB1 or ABCG2 polymorphisms were associated with the apparent clearance of tacrolimus. Conclusions: This is the first report showing that MRP2/ABCC2 has a crucial impact on the pharmacokinetics of tacrolimus in a haplotype-specific manner. Determination of the ABCC2 as well as CYP3A5 genotype may be useful for more accurate tacrolimus dosage adjustment. © 2013 Springer International Publishing Switzerland

Development and validation of a rapid and sensitive assay for simultaneous quantification of midazolam, 1′-hydroxymidazolam, and 4-hydroxymidazolam by liquid chromatography coupled to tandem mass-spectrometry

Midazolam is an ultra short acting benzodiazepine derivative and a specific probe for phenotyping cytochrome P450 (P450) 3A4/5 activity. A rapid, sensitive, and selective LC-MS/MS method was developed for simultaneous quantitation of midazolam and its metabolites (1′-hydroxymidazolam and 4-hydroxymidazolam). Deuterated (D5) analog of midazolam was utilized as an internal standard. Sample preparation either from human plasma (100 μL) or liver microsomal incubations involved a simple protein precipitation using acetonitrile (900 μL) with an average recovery of \u3e90% for all compounds. The chromatographic separation was achieved using Zorbax-SB Phenyl, Rapid Resolution HT (2.1 mm × 100 mm, 3.5 μm) and a gradient elution with 10 mM ammonium acetate in 10% methanol (A) and acetonitrile (B). The flow rate was 0.25 mL/min and total run time was 5.5 min. Calibration curves were linear over the concentration range of 0.100-250 ng/mL. The lower limit of quantitation (LLOQ) was 0.1 ng/mL for all three analytes. The accuracy and precision, estimated at LLOQ and three concentration levels of quality control samples in six replicates, were within 85-115%. In conclusion, a robust, simple and highly sensitive analytical method was developed and validated for the analysis of midazolam and its metabolites. This method is suitable for characterizing the P450 3A4/5 activity in vitro or in human pharmacokinetic studies allowing administration of smaller doses of midazolam. © 2010 Elsevier B.V. All rights reserved

Concentration of tacrolimus and major metabolites in kidney transplant recipients as a function of diabetes mellitus and cytochrome P450 3A gene polymorphism

1. Disposition of tacrolimus and its major metabolites, 13-O-desmethyl tacrolimus and 15-O-desmethyl tacrolimus, was evaluated in stable kidney transplant recipients in relation to diabetes mellitus and genetic polymorphism of cytochrome P450 (CYP) 3A. 2. Steady-state concentration-time profiles were obtained for 12-hour or 2-hour post-dose, in 20 (11 with diabetes) and 32 (24 with diabetes) patients, respectively. In addition, single nucleotide polymorphisms of the following genes: CYP3A4 (CYP3A4: CYP3A4*1B,-392A \u3e G), 3A5 (CYP3A5: CYP3A5*3, 6986A \u3e G) and P-glycoprotein (ABCB1: 3435C \u3e T) were characterized. 3. Dose-normalized concentrations of tacrolimus or metabolites were higher in diabetic patients. CYP3A4*1B carriers and CYP3A5 expressers, independently or when assessed as a combined CYP3A4-3A5 genotype, had significantly lower dose-normalized pre-dose (C0/dose) and 2-hour post-dose (C2/dose) concentrations of tacrolimus and metabolites. Non-diabetic patients with at least one CYP3A4*1B and CYP3A5*1 allele had lower C0/dose as compared to the rest of the population. 4. Genetic polymorphism of CYP3A5 or CYP3A4 influence tacrolimus or metabolites dose-normalized concentrations but not metabolite to parent concentration ratios. The effect of diabetes on tacrolimus metabolism is subject to debate and requires a larger sample size of genetically stratified subjects. © 2013 Informa UK Ltd. All rights reserved

Multidrug Resistance-Associated Protein 2 (MRP2/ABCC2) Haplotypes Significantly Affect the Pharmacokinetics of Tacrolimus in Kidney Transplant Recipients

BACKGROUND AND OBJECTIVE: Tacrolimus is an immunosuppressive drug used for the prevention of the allograft rejection in the kidney allograft recipients. It exhibits a narrow therapeutic index and a large pharmacokinetic variability. Tacrolimus is mainly metabolized by cytochrome P450 (CYP) 3A4 and 3A5, and effluxed via ATP-binding cassette (ABC) transporters such as P-glycoprotein (P-gp), encoded by ABCB1 gene. The influence of CYP3A5*3 on the pharmacokinetics of tacrolimus has been well characterized. On the other hand, the contribution of polymorphisms in other genes is controversial. In addition, the involvement of other efflux transporter than P-gp in tacrolimus disposition is uncertain. The present study was designed to investigate the effects of genetic polymorphisms of CYP3As and efflux transporters on the pharmacokinetics of tacrolimus. SUBJECTS AND METHODS: A total of 500 blood concentrations of tacrolimus from 102 adult stable kidney transplant recipients were included in the analyses. Genetic polymorphisms in CYP3A4 and CYP3A5 genes as well as the genes of efflux transporters including P-gp (ABCB1), multidrug resistance-associated protein (MRP2/ABCC2) and breast cancer resistance protein (BCRP/ABCG2) were genotyped. For ABCC2 gene, haplotypes were determined as follows: H1 (wild type), H2 (1249G>A), H9 (3972C>T) and H12 (−24C>T and 3972C>T). Population pharmacokinetic analysis was performed using nonlinear mixed effects modeling. RESULTS: Analyses revealed that CYP3A5 expressers (CYP3A5*1 carriers) and MRP2 high activity group (ABCC2 H2/H2 and H1/H2) decreased the dose-normalized trough concentration of tacrolimus by 2.3-fold (p<0.001) and 1.5-fold (p=0.007), respectively. The pharmacokinetics of tacrolimus was best described using a two-compartment model with first order absorption and an absorption lag time. In the population pharmacokinetic analysis, CYP3A5 expressers and MRP2 high activity groups were identified as the significant covariates for tacrolimus apparent clearance expressed as 20.7 × (Age/50)(−0.78) × 2.03 (CYP3A5 expressers) × 1.40 (MRP2 high activity group). No other CYP3A4, ABCB1 and ABCG2 polymorphisms were associated with the apparent clearance of tacrolimus. CONCLUSIONS: This is the first report that MRP2/ABCC2 has crucial impacts on the pharmacokinetics of tacrolimus in a haplotype specific manner. Determination of ABCC2 as well as CYP3A5 genotype may be useful for more accurate tacrolimus dosage adjustment