Ian S. Westley

Opportunities to optimize tacrolimus therapy in solid organ transplantation: report of the European consensus conference.

In 2007, a consortium of European experts on tacrolimus (TAC) met to discuss the most recent advances in the drug/dose optimization of TAC taking into account specific clinical situations and the analytical methods currently available and drew some recommendations and guidelines to help clinicians with the practical use of the drug. Pharmacokinetic, pharmacodynamic, and more recently pharmacogenetic approaches aid physicians to individualize long-term therapies as TAC demonstrates a high degree of both between- and within-individual variability, which may result in an increased risk of therapeutic failure if all patients are administered a uniform dose. TAC has undoubtedly benefited from therapeutic drug monitoring, but interpretation of the blood concentration is confounded by the relative differences between the assays. Single time points, limited sampling strategies, and area under concentration-time curve have all been considered to determine the most appropriate sampling procedure that correlates with efficacy. Therapeutic trough TAC concentration ranges have changed since the initial introduction of the drug, while still maintaining adequate immunosuppression and avoiding drug-related adverse effects. Pharmacodynamic markers have also been considered advantageous to the clinician, which may better reflect efficacy and safety, taking into account the between-individual variability rather than whole blood concentrations. The choice of method, differences between methods, and potential pitfalls of the method should all be considered when determining TAC concentrations. The recommendations of this consensus meeting regarding the analytical methods include the following: encourage the development and promote the use of analytical methods displaying a lower limit of quantification (1 ng/mL), perform careful validation when implementing a new analytical assay, participate in external proficiency testing programs, promote the use of certified material as calibrators in high-performance liquid chromatography with mass spectrometric detection methods, and take account of the assay and intermethod bias when comparing clinical trial outcomes. It is also important to consider that TAC concentrations may also be influenced by other factors such as specific pharmacokinetic characteristics associated with the population, drug interactions, pharmacogenetics, adverse events that may alter TAC concentrations, and any change in the oral formulation that may result in pharmacokinetic changes. This meeting emphasized the importance of obtaining multicenter prospective trials to assess the efficacy of alternative strategies to TAC trough concentrations whether it is other single time points or area under the concentration-time curve Bayesian estimation using limited sampling strategies and to select, standardize, and validate routine biomarkers of TAC pharmacodynamics.

Role of MRP2 in the hepatic disposition of mycophenolic acid and its glucuronide metabolites : Effect of cyclosporine

Mycophenolic acid (MPA) is part of the immunosuppressant therapy for transplant recipients. This study examines the role of the canalicular transporter, Mrp2, and the effect of cyclosporin A (CsA), on the biliary secretion of the ether (MPAGe) and acyl (MPAGa) glucuronides of MPA. Isolated livers from Wistar rats (n = 6), or Wistar TR– rats (n = 6) were perfused with MPA (5 mg/l). A third group of Wistar rats (n = 6) was perfused with MPA and CsA (250 μg/l). There was no difference in the half-life, hepatic extraction ratio (EH), clearance or partial clearance of MPA to MPAGe, but there was a difference in partial clearance to MPAGa between control and CsA groups (0.9 ± 0.4 versus 0.5 ± 0.1 ml/min). TR– rats had a lower EH (0.59 ± 0.30 versus 0.95 ± 0.30), a lower clearance (18 ± 8 versus 29 ± 7 ml/min), and a longer half-life (19.5 ± 10.3 versus 10.1 ± 2.4 min) than controls. Compared to controls, MPAGe and MPAGa biliary excretion was reduced by 99% and 71.8%, respectively, in TR– rats, and 17.5% and 53.8%, respectively, in the MPA-CsA group. The biliary excretion of MPAGe is mediated by Mrp2, whereas that of MPAGa seems to depend on both Mrp2 and another unidentified canalicular transporter. Although CsA can inhibit Mrp2, our data suggest that it may also inhibit the hepatic glucuronidation of MPA in Wistar rats.

Clinical study of Lamotrigine and valproic acid in patients with epilepsy : Using a drug interaction to advantage?

Lamotrigine (LTG) is one of the newer antiepileptic drugs which has been shown to have a spectrum of drug interactions (including with other epilepsy drugs) that can have a pronounced effect on LTG kinetics. The present study examined the LTG metabolic inhibition dose–response relationship with valproic acid (VPA) in eight patients with epilepsy with a view to using this to benefit the patient. This could benefit the patient not only by attaining higher plasma LTG concentrations with “standard” dosages of LTG, but also possibly by achieving better seizure control through providing a less variable peak-to-trough fluctuation in LTG concentrations as a result of extending the half-life of LTG. The dosages of VPA trialed were 0, 200, 500, and 1,000 mg/d which resulted in a mean increase in LTG area under the curve of 83.7 ± 14.7% at 200 mg VPA/d, to and 160 ± 37.9% at 1,000 mg VPA/d. The presence of concomitant enzyme inducers in some patients did not influence the percentage increase from baseline in half-life observed, although clearly those on inducers started from a lower absolute half-life as a result of the induction. The effect was shown to be quite variable, particularly at the highest dosage of VPA tested (1,000 mg/d), suggesting that this effect could be best applied with the support of the therapeutic drug monitoring laboratory determining plasma LTG concentrations to allow individualization of the LTG dosage.

Cloned enzyme donor immunoassay tacrolimus assay compared with high-performance liquid chromatography-tandem mass spectrometry and microparticle enzyme immunoassay in liver and renal transplant recipients.

The immunosuppressant drug tacrolimus has a narrow therapeutic index and is subject to a large variation in individual bioavailability and clearance. With its narrow therapeutic index, therapeutic drug monitoring is standard clinical practice in the management of transplant recipients. In this study, we report the evaluation of the cloned enzyme donor immunoassay (CEDIA) for the determination of whole-blood tacrolimus concentrations compared with high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) and microparticle enzyme immunoassay (MEIA) using samples obtained from liver (n = 100) and renal (n = 88) transplant recipients. Linear regression analysis showed a relationship of CEDIA = 1.24 HPLC-MS/MS −0.18 (r2 = 0.81). The mean bias (±SEM) for all patients when compared with HPLC-MS/MS was 22.2% (±2.1%). The precision of the CEDIA method for all samples showed a root mean square error of 3.1 μg/L. Liver transplant recipient samples showed a mean (±SEM) bias compared with HPLC-MS/MS of 12.5% (±1.6%). The precision of the CEDIA method for these samples showed a root mean square error of 1.5 μg/L. The data suggest that in the renal transplant group, the CEDIA and MEIA methods have a bias of 33.3% and 20.1%, respectively, compared with HPLC-MS/MS. The CEDIA tacrolimus immunoassay has been shown to be a rapid method for the determination of whole-blood tacrolimus concentrations and may be considered when HPLC-MS/MS is not available. When used in the clinical setting with other parameters, it would be a useful adjunct in the management of liver transplant recipients, but a significant bias in renal transplant patients needs to be further investigated.

CEDIA mycophenolic acid assay compared with HPLC-UV in specimens from transplant recipients.

Routine monitoring of mycophenolic acid (MPA) has been accepted as an essential tool in the management of this therapy in transplant recipients. The availability of simple, sensitive assays that measure MPA in plasma permits individualization of dosing regimens according to pharmacokinetic principles. We report the results of an evaluation of the CEDIA® Mycophenolic Acid Immunoassay (Microgenics Corporation, Fremont, California) for the measurement of plasma MPA concentrations in a range of transplant indications and compare its performance and specificity to an established HPLC/UV method. Precision and accuracy were determined both within and between runs using the quality control materials provided with the CEDIA® MPA assay, which produced within run (n = 21) coefficients of variation (CV%) and biases of less than 5%. The between run analyses, performed over consecutive days following daily calibration of the assay, showed CVs and biases of less than 7%. Routine patient samples (n = 298) from 142 patients of varying transplant type were analyzed using the CEDIA® MPA kit and HPLC/UV methods. Regression analysis of the patient samples gave an equation of CEDIA® = 1.18 HPLC/UV + 0.45 (r2 = 0.83). According to the manufacturers product information, there is 192% cross reactivity with the active mycophenolate acyl glucuronide. The data presented suggest that the CEDIA® MPA immunoassay, run on the Hitachi 911 analyzer, over-estimates plasma MPA concentrations with a magnitude that is influenced by transplant type. Hence, users must interpret the immunoassay results with caution and not assume that the metabolite fraction is constant in recipients of the same organ type or in different organ transplant populations.

Seradyn Quantitative Microsphere System Lamotrigine Immunoassay on a Hitachi 911 Analyzer Compared With HPLC-UV

Lamotrigine (LTG) is used currently as monotherapy or, more frequently, as add-on therapy with other antiepileptic drugs. It demonstrates efficacy against partial seizures, primary and secondary tonic clonic seizures, absence seizures, and drop attacks. LTG pharmacokinetics is complicated by coadministration with other antiepileptic drugs such as valproic acid, phenytoin, or carbamazepine. The wide interpatient variability in LTG dosage required to attain therapeutic plasma LTG concentrations for seizure control suggests that LTG is a good candidate for therapeutic drug monitoring (TDM). In this study, we compared the quantitative microsphere system (QMS) LTG immunoassay with the LTG high-performance liquid chromatography-ultra violet (HPLC-UV) assay routinely employed for TDM in our laboratory. Samples tested by these methods were patient samples presented for TDM and from a quality assurance program. Quality control material demonstrated within- and between-run (n = 6) coefficient of variation and biases of less than 10%. Patient samples demonstrated a Deming regression of QMS = 1.09 HPLC-UV − 0.17 and quality assurance program samples had a Deming regression of QMS = 1.03 HPLC-UV − 0.11. Patient samples demonstrated a mean bias of 6.1% and quality assurance program samples had a mean bias of 0.2%. The QMS LTG assay had a clinically small but significant overestimation of plasma LTG concentrations. It may be useful as a convenient alternative method that would provide TDM guidance if a chromatographic assay was not available.

CEDIA sirolimus assay compared with HPLC-MS/MS and HPLC-UV in transplant recipient specimens.

The role of the therapeutic drug monitoring laboratory in support of immunosuppressant drug therapy is well established, and the introduction of sirolimus (SRL) is a new direction in this field. The lack of an immunoassay for several years has restricted the availability of SRL assay services. The recent availability of a CEDIA® SRL assay has the potential to improve this situation. The present communication has compared the CEDIA® SRL method with 2 established chromatographic methods, HPLC-UV and HPLC-MS/MS. The CEDIA® method, run on a Hitachi 917 analyzer, showed acceptable validation criteria with within-assay precision of 9.1% and 3.3%, and bias of 17.1% and 5.8%, at SRL concentrations of 5.0 μg/L and 20 μg/L, respectively. The corresponding between-run precision values were 11.5% and 3.3% and bias of 7.1% and 2.9% at 5.0 μg/L and 20 μg/L, respectively. The lower limit of quantification was found to be 3.0 μg/L. A series of 96 EDTA whole-blood samples predominantly from renal transplant recipients were assayed by the 3 methods for comparison. It was found that the CEDIA® method showed a Deming regression line of CEDIA = 1.20 × HPLC-MS/MS − 0.07 (r = 0.934, SEE =1.47), with a mean bias of 20.4%. Serial blood samples from 8 patients included in this evaluation showed that the CEDIA® method reflected the clinical fluctuations in the chromatographic methods, albeit with the variable bias noted. The CEDIA® method on the H917 analyzer is therefore a useful adjunct to SRL dosage individualization in renal transplant recipients.

Interaction of Terbinafine (Anti-fungal agent) with Perhexiline: A Case Report

Perhexiline is a unique anti-anginal agent that is frequently used in the treatment of chronic refractory angina. Its utility has been limited because of its complex pharmacokinetics that were only appreciated following the development of a therapeutic perhexiline assay. Perhexiline is cleared primarily via formation of mono-hydroxy metabolites (OH-perhexiline) by cytochrome P450 2D6 (CYP2D6). Drugs that are inhibitors of CYP2D6 may therefore inhibit perhexiline metabolism, increase plasma perhexiline concentration and may consequently increase the risk of toxicity. We report a case of a rise in perhexiline plasma concentration to a toxic level following the introduction of terbinafine hydrochloride; a moderate CYP2D6 inhibiting drug.

Glucuronidation of mycophenolic acid by Wistar and Mrp2-deficient TR- rat liver microsomes.

In humans, mycophenolic acid (MPA) is metabolized primarily by glucuronidation in the liver to mycophenolate ether glucuronide (MPAGe) and mycophenolate acyl glucuronide (MPAGa). We have previously reported that in perfused livers of TR- rats (lacking the Mrp2 transporter), the clearance and hepatic extraction ratio of MPA were significantly lower compared with control Wistar rats, suggesting a difference in the capacity of the TR- rats to metabolize MPA in situ. There is very little information regarding the phase II metabolic capabilities of TR- rats; therefore, the aim of this study was to investigate the in vitro glucuronidation of MPA in Wistar and TR- rat liver microsomal protein. A second aim was to determine whether MPAGa, cyclosporine (CsA), and/or its metabolites AM1, AM1c, and AM9 inhibit the metabolism of MPA to MPAGe in rat liver microsomes. MPAGe formation rates by Wistar and TR- microsomes were 0.48 and 0.65 nmol/min/mg, respectively (p = 0.33). Km values for control and TR- microsomes were 0.47 and 0.50 mM, respectively (p = 0.81). The mean (S.E.M.) ratios of MPAGe formation by Wistar rat liver microsomes incubated with 50 μM MPA plus inhibitor versus 50 μM MPA alone were MPAGa 1.2 (0.1), CsA 0.7 (0.1) (p < 0.05), AM1 2.2 (0.3) (p < 0.05), AM1c 1.2 (0.2), and AM9 1.0 (0.2). Our results suggest that lower in situ glucuronidation of MPA in TR- rats may be because of inhibition of glucuronidation by endogenous and exogenous compounds that accumulate in the transporter-deficient rat. Whereas CsA inhibits glucuronidation of MPA, its metabolite AM1 enhances MPAGe formation by rat liver microsomes.

A randomized double-blind clinical trial of a continuous 96-hour levobupivacaine infiltration after open or laparoscopic colorectal surgery for postoperative pain management--including clinically important changes in protein binding.

Background: Continuous local anesthetic infiltration has been used for pain management after open colorectal surgery. However, its application to patients undergoing laparoscopic colorectal surgery has not been examined. The aim of this prospective, randomized, double-blind, placebo-controlled clinical trial was to study the use of a commercial infiltration device in patients undergoing open or laparoscopic colorectal surgery, along with plasma concentrations of levobupivacaine, its acute-phase binding protein (alpha-1 acid glycoprotein, AAG), and the stress marker, cortisol. Methods: Eligible patients were randomized (2:1) to receive a continuous infiltration of either levobupivacaine or placebo using a commercial device (ON-Q PainBuster) inserted in the preperitoneal layer at the end of surgery. Blood was sampled for determination of levobupivacaine and AAG and cortisol concentrations. Other outcomes measured were pain scores, morbidity and mortality, time to bowel movement, mobilization, and length of hospitalization. Results: In patients having open surgery, the levobupivacaine treatment showed a trend toward reduced total opioid consumption. No patients reported adverse effects attributable to levobupivacaine, despite 11 patients having concentrations at some time(s) during the 96-hour infiltration of up to 5.5 mg/L exceeding a putative toxicity threshold of 2.7 mg/L. AAG concentrations measured postsurgery increased by a mean of 55% (P < 0.001) at 48 hours. Cortisol concentrations also increased significantly by a mean of 191% at 1 hour. Conclusions: Continuous local anesthetic infiltration may be more beneficial in open surgery. The threshold for adverse effects from highly bound local anesthetic drugs established in healthy volunteers is of limited usefulness in clinical scenarios in which AAG concentration increases in response to surgical stress. Hence, there is scope to adopt higher doses to enhance therapeutic benefit.