Volume 15, Issue 9 , Pages 1134-1139, September 2009
Pharmacokinetics of Mycophenolic Acid Administered 3 Times Daily after Hematopoietic Stem Cell Transplantation with Reduced-Intensity Regimen
Article Outline
Mycophenolate mofetil (MMF) is an immunosuppressive drug used as a prophylactic agent to prevent acute graft-versus-host disease (aGVHD) after hematopoietic stem cell transplantation (HSCT). After reduced-intensity conditioning (RIC) regimen, administration of MMF orally 3 times a day (tid) seems to be more beneficial than twice a day (bid). However, information regarding the pharmacokinetic (PK) parameters of mycophenolic acid (MPA), the active metabolite of MMF, administered in this regimen are very limited. We performed a prospective study in 15 patients for whom 3 sets of sampling were performed: at the beginning of the treatment, after 1 week, and after 1 month. Two consecutive 8-hour sets of sampling were performed at day 0 (D0) and D7. Plasma concentrations of MPA were quantified and areas under the curve for 8
hours (AUC0-8), and maximal and through concentrations were calculated. The results show that AUC0-8 increases between the beginning of treatment and the end of the first week, but remains stable thereafter. Moreover, a trend to lower AUC0-8 was observed for the patients who experienced GVHD ≥2 compared to those patients who did not. The other PK parameters are not associated with pharmacodynamic events. A limited sampling strategy with Bayesian estimators is currently under investigation to confirm these data and the role of D7 AUC0-8 as a potential target of therapeutic drug monitoring (TDM).
key Words: Mycophenolate mofetil, Pharmacokinetics, Reduced-intensity hematopoietic cell transplantation, GVHD
Introduction
Mycophenolic acid (MPA), the active metabolite of mycophenolate mofetil (MMF), is widely used as an immunosuppressive drug to prevent graft rejection in solid organ transplantation 1, 2. Because of a better tolerance, MMF has also been introduced in replacement of methotrexate (MTX) in the prophylaxis of graft-versus-host disease (GVHD) after hematopoietic stem cell transplantation (HSCT) 3, 4. The administration of MMF as prophylaxis of GVHD is of particular interest after reduced-intensity conditioning (RIC) regimens. These regimens, by decreasing the intensity of myeloablation, allow performing HSCT in patients who are not suitable candidates for conventional conditioning because of age or comorbidities, but induce higher rates of GVHD 5, 6, 7. In routine practice, because of a shorter half-life (3hours) after HSCT, the dose of MMF progressively shifted from 15mg/kg twice a day (bid) to 15mg/kg 3 times a day (tid), resulting in a trend to a decrease of the GHVD rate 5, 8, 9. Moreover, this trend is supported by lower exposition to MPA (1g bid) after HSCT than after solid organ transplantation 5, 10, 11. There are few data regarding the evolution of the pharmacokinetics (PK) of MPA administered orally tid after RIC HSCT 12, 13. In this study, we extensively describe the results of the PK parameters of MPA in such situations and their potential impact on acute GVHD (aGVHD) occurrence, in a view to standardize the best time to perform therapeutic drug monitoring (TDM).
Patients and Methods
Patients
For this prospective study, patients were required to be candidates for RIC HSCT as defined by the European Group for Blood and Marrow Transplantation (EBMT) [14], aged between 18 and 70 years, and to provide written informed consent. Exclusion criteria were progressive disease, hypersensitivity to MMF, impossibility to perform the planned follow-up, and patient's refusal. Patient details (pathologies, conditioning regimens, source of hematopoietic stem cells, and cytomegalovirus [CMV] status) and MMF doses are presented in Table 1. None of the patients received antithymocyte globulin (ATG). Oral cyclosporine (CsA; 5mg/kg bid) was administered together with oral MMF (15mg/kg tid) for GVHD prophylaxis to all patients, starting 3 days before stem cell infusion. MMF was administered according to the patient's body weight (BW); 750mg tid if BW was <70kg, 1g tid if BWwas ≥70kg and adjusted to a trough blood concentration not exceeding 3.5mg/L. CsA was targeted to maintain a 150-300 ng/mL blood level. The cell infusion was performed 24-48hours after the end of conditioning regimen. aGVHD was graded using Glucksberg's criteria [15]. Institutional ethical review committee (Comité de Protection des Personnes de Franche-Comté) approved the study.
Table 1. Main Characteristics of the Patients (pts) Included in the Study
| Pts | Gender | Age | Pathology | Conditioning Regimen | Origin of Stem Cells | Donor R/U | MMF Dose at D1 / D7 / M1 (g) | CMV Status (Donor/Recipient), Reactivation | GVHD Grade | Number of Days to ANC >0.5G/L |
|---|---|---|---|---|---|---|---|---|---|---|
| 1 | F | 51 | CML | Flu, Mel | BM | R | 1 / 1 / 1 | − / +, R+ | 2 | 14 |
| 2 | F | 57 | RAEB | Flu, Mel | PBMC | R | 1 / 1 / — | + / +, R+ | 0 | 14 |
| 3 | F | 57 | AML | Flu, TBI | PBMC | R | 0.75 / 0.75 / 0.75 | + / +, R− | 1 | 21 |
| 4 | M | 20 | Hodgkin's disease | Flu, TBI | PBMC | R | 1 / 0.75 / 0.75 | + / +, R− | 0 | 10 |
| 5 | M | 50 | Primary idiopathic myelofibrosis | Flu, Cy, TBI | UCB | U | 1 / 1 / 1.5 | NA | 3 | 21 |
| 6 | M | 42 | AML | Flu, Mel | PBMC | U | 1 / 1 / 1 | − /−, R− | 3 | 15 |
| 7 | F | 34 | AML | Flu, Cy, TBI | UCB | U | 1 / 1 / 1 | NA | 2 | 16 |
| 8 | F | 60 | AML | Flu, Mel | PBMC | R | 1 / — / 0.75 | + / +, R+ | 0 | 13 |
| 9 | M | 50 | CLL | Flu, Cy, TBI | PBMC | U | 1 / 1 / 1 | + / -, R− | 0 | 10 |
| 10 | M | 58 | AML | Flu, Mel | BM | R | 1 / 1 / 1 | + / +, R− | 0 | 21 |
| 11 | M | 57 | AML | Flu, Mel | PBMC | U | 1 / 0.5 / 0.5 | + /−, R− | 1 | 20 |
| 12 | M | 45 | AML | Flu, Cy, TBI | UCB | U | 1 / 1 / 1 | − /−, R− | 2 | 24 |
| 13 | M | 51 | AML | Flu, Mel | BM | U | 1 / 1 / 1 | − /−, R− | 0 | 14 |
| 14 | F | 47 | Myeloma | Flu, Mel | PBMC | R | 1 / 1 / 1 | − / +, R− | 2 | 13 |
| 15 | M | 59 | AML | Flu, Bu | PBMC | U | 1 / 1 / — | − /−, R− | 0 | 12 |
Methods
Blood samples for PK were taken using EDTA-containing tubes. Three sets of samples were planned: at the beginning of the MMF treatment (day 0 [D1]), 1 week later (D7), and 1 month after the beginning of the treatment (M1). For the 2 first sets (D0 and D7), 2 8-hour sets of samples covering 2 cycles of absorption were taken. To deal with potential PK interference because of enterohepatic recirculation, 2 complete set of samples were drawn at predose, 0.25, 0.5, 0.75, 1, 1.5, 2, 4, and 6hours after the first administration of the day (PK1), and at the same time points plus an additional sample 8hours after the second dose of the day (PK2). At M1, only PK1, extended to 8hours, was performed. After drawing, the samples were immediately centrifuged and frozen until assay.
MPA determination in plasma was performed using high-performance liquid chromatography (HPLC) with ultraviolet (UV) detection according to Na-Bangchang et al. [16]. Briefly, 250 μL of patient plasma or standard was added to 10 μL of internal standard (thiopental 400mg/L), 500 μL of HCl (0.02N), and 4mL of methylene chloride. After extraction, the tubes were centrifuged and the organic layer was evaporated with nitrogen. The samples were reconstituted with HPLC mobile phase (58/42 v/v mixture of NaH2PO4 buffer: 50mM pH 2.7 and CH3CN). The samples were then assayed using a Pursuit® 5C18 150×4.6-mm column (Varian SA, Les Ulis, France) and a photodiode array at a wavelength of 254nm. This method allows a lower limit of quantification of 0.1mg/L and a coefficient of variation of 6.0% for interday variability with the 1mg/L concentration. Linearity of the calibration curve is observed between 0.1 and 20mg/L. For each set of samples, normalized (1000mg) areas under the concentration-time curves (AUC) were calculated by trapezoidal method. Trough and maximal concentrations were graphically determined.
When indicated, nonparametric Wilcoxon, Mann-Whitney, or exact Fisher tests were performed to compare data. Unless indicated, the results are expressed as median with range.
Chimerism was assessed after 1 month and regularly thereafter on blood nucleated cells, mononucleated cells, granulocytes, and CD3+ lymphocytes as previously described [17]. Complete engraftment was defined by the presence of >95% CD3+ donor lymphocytes in the blood.
Results
Median age at the first PK profile was 50.7 (20-60) years. All aGVHD occurred before the M1 PK; no or grade I aGVHD in 8 patients and grade ≥II aGVHD observed in 7 patients (grade 2, n=4 and grade III, n=2). Hematopoietic recovery was observed in all patients with a median duration of 15 (10-24) days to achieve a sustained absolute neutrophil count (ANC) ≥0.5G/L (Table 1). Complete donor engraftment was documented in 11 patients at the end of the first month and for all patients before the third month after HSCT. Grade ≥3 nonhematologic toxicities were not observed during the study duration.
Three sets of sampling could not be analyzed: 1 at D7 because the patient was switched to intravenous MMF, and 2 at M1 because 1 patient was switched to tacrolimus and the other 1 presented with difficulties to venous access. Seventy-one 8-hour cycles of MPA PK were thus available for the analyses.
The mean concentration-time profiles for each set of samples show a high interpatient variability (Figure 1). The intraindividual variability was 20% and the interindividual variability 44%. No enterohepatic recirculation, which could overlap the PK2 absorption was observed, and the AUC0-8 were not statistically different between the PK1 and PK2 performed the same day. Thus, to compare the AUC0-8 observed at each administration, the means of the 2 consecutive AUC0-8 of the same day were compared. The high interpatient variability is also observed for the dose-normalized AUCs obtained after each 8-hour set (Figure 2). The median values of AUC0-8 were of 15.77 (6.99-31.62) mg/h/L and 16.69 (8.16-37.33) mg/h/L for the 2 respective PK of D0, of 21.83 (8.96-49.99) and 20.70 (16.66-40.20) for the 2 respective PK of D7, and of 24.08 (9.53-52.08) mg/h/L for the PK performed at M1. The mean AUC0-8 obtained at D7 (23.93mg/h/L) and AUC0-8 at D30 (26.20mg/h/L) showed a statistically significant increase (P < .05) compared to the mean AUC0-8 obtained at D0 (16.32mg/h/L). No difference was observed regarding the AUC between D7 and D30. Mean concentrations were statistically different between PK1 and PK2 for D7 (0.80 versus 0.99mg/L) [P < .05] only, but not for PK2 between D0, D7, and M1 [0.59 versus 0.99 versus 0.87mg/L]. Maximal concentrations were similar whatever the tested sampling period.
Figure 1
Mean concentration-time profiles of MPA for samples obtained at D0 (A), D7 (B), and M1 (C) after the beginning of MMF treatment. The results are the mean
±SD of 13 to15 patients.
Figure 2
Normalized AUC0-8 obtained at D0 during the first (D0 PK1) and the second administration (D0 PK2), at D7 during the first (D7 PK1) and the second administration (D7 PK2), and at M1. The box represents the 25th and 75th percentiles with the median as a solid line, and the whiskers the 5th/95th percentiles.
All aGHVD were observed before the M1 PK, and we studied D0 and D7 PK parameters according to the risk of aGVHD. A trend to a significant lower actual AUC0-8 for those patients who experienced aGVHD ≥ii compared to those patients who did not could be observed at D0 (P=.099) and D7 (P=.053) (Figure 3). At D7, patients with AUC0-8≥22.5mg/h/L (5/14 patients) displayed no aGVHD ≥II compared with patients with AUC0-8<22.5mg/h/L (9/14 patients; 7 of 9 had GVHD ≥2); (P < .05, exact Fisher test). No similar difference is observed with the other PK parameters and no PK parameters could be linked with engraftment kinetics.
Figure 3
Actual values of AUC0-8 obtained in patients with aGHVD ≥2 (black dots) and patients with aGVHD <2 (open circles).
Discussion
This study presents the PK parameters of MPA administered tid as GHVD prophylaxis for RIC HSCT. During the first week, the AUC0-8 increases, but remains stable thereafter. This is consistent with a 3-hour half-life for MPA as described by Maris et al. [8] during HSCT and the fact that the steady state is reached at D7. Moreover, the increase of MPA exposure after the beginning of the treatment is a known phenomenon, already described during HSCT even with i.v. administration of MMF [18]. Giaccone et al. [5] also find that exposure of MPA is stable between D7 and D21 with a tid oral administration. These results tend to show a stability of MPA exposure as soon as 1 week after oral administration and strengthen the interest of administering MMF 1 week before cell infusion that will then be performed at steady state.
The results are close to those already published regarding the AUC0-8, when MMF is administered in similar conditions: mean values between 16.32 and 26.20 for our study versus mean values between 15.9 and 16.9 for Okamura et al. [19], between 20.99 and 21.14 for Nash et al. [3], between 24.7 to 25.0 for Giaccone et al. [5], or between 16.48 and 18.80 for Jacobson et al. [12]. Previous data about MPA pharmacokinetics show that MPA undergoes enterohepatic recirculation that is described to happen around 6hours after the absorption of MMF. However, in a tid regimen, interindividual PK variability of MPA may lead to a delayed enterohepatic recirculation, then an overlap between this phenomenon and the absorption of MPA because of its following administration. In the present study, we perform PK sampling after 2 administrations of MMF because of a potential overlap of MPA enterohepatic recirculation during the second absorption phase. We do not observe such phenomenon probably because MMF was combined with CsA, which is known to inhibit enterohepatic recirculation in kidney transplantation [20].
GVHD has a major impact on morbidity and mortality associated to HSCT 6, 21. Grade ≥II aGVHD is associated with an increased risk of nonrelapse mortality (NRM) and a decreased progression-free survival (PFS) [21]. We investigate a potential relationship between MPA exposures, expressed as actual AUC0-8, and grade ≥II aGVHD to assess the best parameters to perform TDM. Despite a low number of patients, we find that patients with aGVHD ≥II have lower exposure than those with aGVHD <II. Only few authors have found that exposures to MPA can be associated with the rate of GVHD: Jacobson et al. [22] observed that a low AUC of free MPA is associated with a higher risk of aGVHD ≥II; Osunkwo et al. [23] showed that patients with trough concentrations between 1.0 and 3.5μg/mL before M1 had a significantly reduced risk of grade ≥II aGVHD, and Okamura et al. [19] showed that patients with higher MPA exposures (MMF tid versus bid) had a trend to lower incidence of aGVHD ≥2 [19]. These results have been obtained under variable conditions in terms of administration of ATG, use of RIC or standard conditioning regimens, and variable CsA or tacrolimus protocols making comparisons difficult. Jacobson et al. [12] also stressed the differences in the conditioning regimens (including the administration of ATGs), their impact on the PK parameters of MPA, and their relations with GVHD occurrence. In the present study, the homogeneity of the studied population, the absence of ATG administration, the closely monitored dose of CsA, and the systematic use of RIC might lead to a situation with a sufficient risk of aGVHD to observe differences in the MPA exposures even if some heterogeneity (nature of conditioning regimens and source of stem cells) persist in our data. The best association between MPA exposure and aGHVD was observed on D7, this sampling time thus appeared to be optimal for PK assessment as a potential predictive factor of aGVHD. We explore 3 sets of sampling, all after oral administration and spread over 1 month, with 2 repetitive PK performed on the same day. These 2 repetitive PK allow the extensive study of the putative enterohepatic recirculation, which was only partially explored by Jacobson et al. [12]. Thus, if the global PK parameters are similar to previously published data, we provide here additional information about the best sampling time to perform TDM and an extensive study of the intraindividual variability, which is critical to perform TDM as suggested by Perez-Simon et al. [13]. Further studies are, however, needed to confirm our preliminary results and that AUC0-8 obtained at D7 is a good candidate to perform TDM for the prevention of GHVD. We choose to analyze total MPA. Indeed, if interesting results were observed by Jacobson et al. [12] regarding the relationship between free MPA and GVHD occurrence, the routine practice is to perform TDM with total MPA; we thus assess parameters that can be easily be used by most centers.
To conclude, this study suggests that performing TDM with AUC on D7 might be relevant for the clinical follow-up of these patients. Two recent studies showed the interest of TDM, especially in decreasing the interpatient variability 19, 24. Moreover, Okamura et al. [19] confirmed that MMF tid seems more effective than bid regarding GVHD prophylaxis. The need of TDM was also strengthened by the results of the study of Jacobson et al. [12], in which the authors observed that, at a fixed dose, the patients uncommonly reached the wished AUC target for total MPA (13%-27% for concentration at steady state). However, this strategy needed 6 samples to perform the AUC calculation. We are currently developing a limited sampling strategy using a Bayesian estimator as those previously published 25, 26, 27, 28. Such strategy performed at D7 may be used to perform larger prospective studies to confirm present data.
Acknowledgments
Financial disclosure: This study was granted in part by the public agency “Agence Française de Sécurité Sanitaire des Produits de Santé.” The authors declared no financial relationship.
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Financial disclosure: See Acknowledgments on page 1138.
PII: S1083-8791(09)00213-4
doi:10.1016/j.bbmt.2009.04.011
© 2009 American Society for Blood and Marrow Transplantation. Published by Elsevier Inc. All rights reserved.
Volume 15, Issue 9 , Pages 1134-1139, September 2009
