Mycophenolic Acid Concentrations Research Articles

Optimal exposure of mycophenolic acid for induction therapy of childhood lupus nephritis patients: an observational cohort study.

Mycophenolic acid (MPA) is recommended for lupus nephritis (LN) treatment, but with large inter-individual variability in pharmacokinetics (PK). The aim of this study is to reveal the relationship between MPA exposure and disease response and adverse drug reactions in pediatric LN patients. This was a population-based observational cohort study. A total of 86 pediatric LN patients treated with mycophenolate mofetil (MMF) for induction therapy were enrolled. The area-under the concentration-time curve (AUC) was calculated using MPA concentrations according to a limited sampling strategy. Receiver operating characteristic analysis was performed to assess the MPA-AUC threshold values. The cumulative incidence of renal remission and inactive SLE over time was evaluated by Kaplan-Meier's analysis. MPA-AUC was identified as an independent factor associated with renal remission and lupus activity at 6 and 12 months after MMF treatment, and the improved renal remission rates was correlated with higher MPA-AUC, with thresholds of 29.81 and 30.63 μg·h·mL - 1 at 6 and 12 months, respectively. Furthermore, the thresholds for maintaining the hypoactive state of LN were 30.96 and 31.19 μg·h·mL - 1at 6 months and 12 months, respectively. Patients reaching target thresholds for MPA-AUC achieved renal response or stable disease earlier. In addition, the MPA-AUC threshold for decreasing MMF-related adverse reactions was 50.80 μg·h·mL - 1. The initial and long-term treatments of pediatric LN patients with MMF should be individualized according to the MPA-AUC, and the recommended MPA exposure is 31.19-50.80 μg·h·mL - 1.

Population Pharmacokinetics and Limited Sampling Strategy of Mycophenolate Mofetil for Indian Patients with Lupus Nephritis

Background: Mycophenolic acid is widely used to treat lupus nephritis (LN). However, it exhibits complex pharmacokinetics with large interindividual variability. This study aimed to develop a population pharmacokinetic (popPK) model and a 3-sample limited sampling strategy (LSS) to optimize therapeutic drug monitoring in Indian patients with LN. Methods: Five blood samples from each LN patient treated with mycophenolic acid were collected at steady-state predose and 1, 2, 4, and 6 hours postdose. Demographic parameters were tested as covariates to explain interindividual variability. PopPK analysis was performed using Monolix and the stochastic approximation expectation-maximization algorithm. An LSS was derived from 500 simulated pharmacokinetic (PK) profiles using maximum a posteriori Bayesian estimation to estimate individual PK parameters and area under the curve (AUC). The LSS-calculated AUC was compared with the AUC calculated using the trapezoidal rule and all the simulated samples. Results: A total of 51 patients were included in this study. Based on the 245 mycophenolic acid concentrations, a 1-compartmental model with double absorption using gamma distributions best fitted the data. None of the covariates improved the model significantly. The model was internally validated using diagnostic plots, prediction-corrected visual predictive checks, and bootstrapping. The best LSS included samples at 1, 2, and 4 hours postdose and exhibited good performances in an external dataset (root mean squared error, 21.9%; mean bias, −4.20%). Conclusions: The popPK model developed in this study adequately estimated the PK of mycophenolic acid in adult Indian patients with LN. This simple LSS can optimize TDM based on the AUC in routine practice.

The effects of food on the pharmacokinetics of mycophenolate mofetil in healthy horses

AbstractAdditional immunomodulatory treatment is needed for the management of immune‐mediated disease in horses. Mycophenolate mofetil (MMF) is an immunomodulatory agent used in human and veterinary medicine for the prevention of graft rejection and the management of autoimmune diseases. Few studies exist investigating the pharmacokinetics of MMF in horses. The aim of this study was to evaluate the pharmacokinetics of a single dose of MMF in healthy horses in the fed vs. fasted state. Six healthy Standardbred mares were administered MMF 10 mg/kg by a nasogastric (NG) tube in a fed and fasted state. A six‐day washout period was performed between the two doses. No statistically significant differences in mycophenolic acid (MPA) concentrations were seen at any time point apart from 8 h, when plasma metabolite concentrations were significantly higher in the fasted state compared to the fed state (p = .038). Evidence of enterohepatic recirculation was seen only in the fasted state; this did not yield clinical differences in horses administered a single‐dose administration but may be significant in horses receiving long‐term MMF treatment.

Topical Tacrolimus and Mycophenolic Acid Therapy Synergizes with Low Dose Systemic Immunosuppression to Sustain Vascularized Composite Allograft Survival

AimThe goal of this study was to evaluate whether topical administration of tacrolimus (TAC) and mycophenolic acid (MPA) at the transplant site enables vascularized composite allograft (VCA) survival with significant minimization of the dose and adverse effects of systemic TAC (STAC) immunosuppression. Materials and methodsLewis (Lew) rats received orthotopic hind limb allotransplants from fully mismatched Brown Norway (BN) donors. Group 1 (Controls) received no treatment. Other groups were treated with STAC at a dose of 1 mg/kg/day for 7 days. On post-operative day (POD) 8, the STAC dose was dropped to 0.1 mg/kg/day for Group 2 and maintained at 1 mg/kg for Group 3. Group 4 received topical application of TAC and MPA on the transplanted (Tx) limb starting POD 8 without STAC. Group 5 received topical TAC and MPA on the contralateral non-Tx limb and Group 6 received topical TAC and MPA on the Tx limb starting POD 8 along with low dose STAC (0.1 mg/kg/day). Treatment was continued until the study end point was reached, defined as either grade 3 rejection or allograft survival exceeding 100 days. .We conducted sequential LC-MS/MS measurements to assess TAC and MPA concentrations in both blood/plasma and allograft tissues. Additionally, we evaluated markers indicative of organ toxicity associated with STAC immunosuppression. ResultsCompared to controls, topical therapy with TAC+MPA significantly prolonged allograft survival beyond 100 daysat very low dose STAC (0.1 mg/kg/day) (Group 6). The histopathological assessment of the grafts was consistent with the clinical outcomes. .Drug levels in blood/plasma remained low or undetectable, while allograft tissues showed higher drug concentrations compared to contralateral limb tissues (P<0.05). . Urinary creatinine clearance remained within the normal range at 2.5 mL/min. ConclusionCombination therapy with topical TAC and MPA synergizes with a very low dose, corticosteroid- free-STAC regimen and facilitates rejection-free, prolonged VCA survival without morbidity.

Therapeutic drug monitoring of mycophenolic acid and clinical outcomes of lupus nephritis: a systematic review and meta-analysis.

Mycophenolic acid (MPA) is a primary immunosuppressive agent used in the treatment of lupus nephritis (LN). While therapeutic drug monitoring (TDM) of MPA is well established in organ transplantation, its role in LN treatment remains uncertain. Our objective was to review and summarise current knowledge on TDM of MPA in the LN treatment. A systematic search was conducted in the online databases, specifically targeted patients diagnosed with LN receiving MPA treatment. The included studies had to report both MPA pharmacokinetic parameters and renal outcomes. A random-effects model meta-analysis was conducted to assess the relationship between clinical responses and MPA pharmacokinetics. A total of 1507 studies were initially screened, resulting in the inclusion of 16 studies for meta-analysis, encompassing 433 patients. The response group exhibited significantly higher MPA area under the concentration-time curve (AUC) compared with the non-response group (51.44±21.73 mg·h/L vs 30.30±16.24 mg·h/L). The weighted mean difference (WMD) of MPA-AUC between responders and non-responders was 16.83 mg·h/L (95% CI 10.59 to 23.06; p<0.001). Similarly, trough concentration (C0) of MPA showed a strong association with renal response, evidenced by C0 values of 2.50±1.73 mg/L in the response group vs 1.51±1.33 mg/L in the non-response group (WMD 1.37 mg/L; 95% CI 0.77 to 1.97; p<0.001). There was no significant relationship identified between MPA-AUC and adverse events. This meta-analysis emphasised the meaningful correlation between MPA AUC and C0 with renal response in LN treatment. Randomised controlled trials are necessary to validate this approach and determine its superiority over fixed dosing in the context of LN treatment.

Evaluation of mycophenolic acid exposure in a patient with immune-related hepatotoxicity caused by nivolumab and ipilimumab therapy for malignant melanoma: a case report.

Guidelines such as the National Comprehensive Cancer Network recommend mycophenolate mofetil (MMF) for the treatment of severe steroid-refractory immune-related hepatotoxicity. Mycophenolic acid (MPA) is an active form of MMF that suppresses T- and B-lymphocyte proliferation and immune-related adverse events caused by immune checkpoint inhibitors. MPA has a narrow therapeutic range (37-70µg·h/mL) and overexposure increases the risk of leukopenia in transplantation. However, the optimal use of MMF in oncology has not yet been established; thus, monitoring plasma MPA concentrations is necessary to avoid excessive immunosuppression in oncology practice. We evaluated plasma MPA concentration in a 75-year-old man with immune-related hepatotoxicity following nivolumab and ipilimumab combination therapy for malignant melanoma. The patient developed severe hepatotoxicity after immunotherapy, and immunosuppressant therapy with corticosteroids was initiated. The patient then developed steroid-refractory immune-related hepatotoxicity; therefore, MMF (1,000mg twice daily) was co-administered. Seven days after the administration of MMF, the plasma MPA concentration was measured using an enzyme multiplied immunoassay technique. The area under the plasma concentration-time curve for MPA from 0 to 12h was 41.0µg·h/mL, and the same dose of MMF was continued. Grade 2 lymphocytopenia, which could be attributed to MMF, was observed during the administration period. Unfortunately, the patient was infected with SARS-CoV-2 and died from respiratory failure. Our patient did not exceed the upper limit of MPA levels as an index of the onset of side effects of kidney transplantation and achieved rapid improvement in liver function. Prompt initiation of MMF after assessment of the steroid effect leads to adequate MPA exposure. Therapeutic drug monitoring should be considered when MMF is administered, to avoid overexposure.

Quantification of Mycophenolic Acid in Plasma by High Performance Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS).

Mycophenolate mofetil (MMF) and sodium mycophenolate are commonly prescribed immunosuppressive drugs for patients who have undergone solid organ transplant. Therapeutic drug monitoring (TDM) of these drugs is performed by assessing mycophenolic acid (MPA) in plasma. Due to the large inter-individual variability and narrow therapeutic range, the precise determination of systemic MPA concentration carries great clinical significance. We present a rapid, sensitive, specific, and robust liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the quantitation of MPA in plasma. A Waters Xevo TQ-S Micro mass spectrometer coupled to a Water's Acquity liquid chromatography system was used in positive electrospray ionization (ESI) mode. MPA quantitation was achieved using multiple reaction monitoring (MRM). Mycophenolic acid carboxybutoxy ether (MPAC) was employed as an internal standard. The method is linear from 0.25 to 40.00mg/L, has intra-assay (N=24) imprecision of 2.7% at 1.57mg/L and 3.9% at 4.61mg/L and inter-assay (N=20days) imprecision of 4.0% at 1.62mg/L and 5.6% at 4.68mg/L.

The Effects of Mycophenolate Mofetil and Cyclosporine with Concomitant Posaconazole Tablets in the Early Stage of Allogeneic Hematopoietic Stem Cell Transplantation

Background： Allogeneic hematopoietic stem cell transplantation (Allo-HSCT) mainly applied the “Beijing regimen”(cyclosporine + methotrexate + mycophenolate mofetil（MMF）). This approach significantly reduces the risk of graft-versus-host disease (GVHD), increases the risk of infection. Posaconazole can significantly reduce the incidence of invasive fungal disease (IFD) during the immunosuppressive phase. Cyclosporine and posaconazole have significant drug interactions, with MMF also have drug interactions. However, there is limited research about drug interactions of simultaneous three-drug combination. Objective： Explore the effects of MMF and cyclosporine with concomitant posaconazole in allo-HSCT patients within 30 days and the breakthrough incidence of IFD within 100 days post-transplantation. Method: A single-arm, open-label, prospective trial was conducted to evaluate posaconazole tablet (300 mg q12h oral d1,300 mg/d) as secondary antifungal prophylaxis in allo-HSCT patients (18-65 years old) with previous proven or probable IFD (ChiCTR2200059472). Posaconazole and cyclosporine were started when conditioning began, while MMF was used one day before infusion of stem cell for one month with fixed dose of 2 g/d. The plasma concentration of posaconazole and mycophenolic acid (MPA) which is the active ingredient of MMF, were monitored weekly, cyclosporine was monitored twice one week until 30 days post-transplantation. Results: Thirty patients from July 2022 to March 2023 were enrolled in the trial, included 16 males and 14 females, acute myelogenous leukemia 22, acute lymphoblastic leukemia 2, myelodysplastic syndrome 2, aplastic anemia 4. All patients underwent haploidentical HSCT. All patients had history of IFD(5 proven IFD, 13 probable IFD and 12 possible IFD). At week1, 2, 3 and 4 after conditioning, The mean plasma concentrations of posaconazole were 0.47±0.29ug/ml, 0.66±0.32ug/ml,0.72±0.40ug/ml and 0.91±0.32ug/ml（Figure 1）. The mean plasma concentrations of cyclosporine were 138.20±35.33ng/ml, 190.61±48.07ng/ml,245.46±37.31ng/ml and 253.56±45.19ng/ml（Figure 2）.The mean plasma concentrations of MPA at week 2, 3 and 4 were 0.35±0.15mg/L, 0.89±0.36mg/L and 1.18±0.58mg/L (Figure 3). After week 1, concentration of cyclosporine of 13 cases was below 150ng/ml and the dosage was increased by 25% of the original dose, at week 4, concentration of cyclosporine of 9 cases was above 250ng/ml and the dosage was decreased by 25%. After posaconazole reached steady state concentrations at week 2, while cyclosporine reached target concentrations after the dose increase, MPA did not reach target concentrations at week 2. The concentrations of MPA significant increase without changing the MMF dose and reached target concentrations at week 3 simultaneous three-drug combination. Cyclosporine concentrations increased significantly at week 4 with stable concentrations of MPA and posaconazole, and 30% of patients required dose reduction of cyclosporine. Only one patient was diagnosed as aspergillus fumigatus pneumonia through NGS of pulmonary alveolar lavage fluid. The breakthrough incidence of IFD was 3.33%. Conclusion: Different times of administration lead to different times of interaction among the three drugs. The combination of cyclosporine and MMF leads to decrease in MPA concentrations, but the concentration of MPA can rapidly increase when combined with posaconazole. When target concentration of MPA is not reached in the first week, it is recommended to suspend dose modification and TDM in the second week. However, the accumulation of cyclosporine was significant in the fourth week, and it was recommended that cyclosporine should be monitored and adjusted within three weeks after the three drugs were combined. The cumulative breakthrough incidence of 3.33% is lower than reported in literaturesuggesting that posaconazole appears to be effective for secondary prophylaxis of IFD after allo-HSCT.

Limited sampling strategy and population pharmacokinetic model of mycophenolic acid in pediatric patients with systemic lupus erythematosus: application of a double gamma absorption model with SAEM algorithm.

Mycophenolic acid (MPA), the active metabolite of mycophenolate mofetil (MMF), is widely used in the treatment of systemic lupus erythematosus (SLE). It has been shown that its therapeutic drug monitoring based on the area under the curve (AUC) improves treatment efficacy. MPA exhibits a complex bimodal absorption, and a double gamma distribution model has been already proposed in the past to accurately describe this phenomenon. These previous population pharmacokinetics models (POPPK) have been developed using iterative two stage Bayesian (IT2B) or non-parametric adaptive grid (NPAG) methods. However, non-linear mixed effect (NLME) approaches based on stochastic approximation expectation-maximization (SAEM) algorithms have never been published so far for this particular model. The objectives of this study were (i) to implement the double absorption gamma model in Monolix, (ii) to compare different absorption models to describe the pharmacokinetics of MMF, and (iii) to develop a limited sampling strategy (LSS) to estimate AUC in pediatric SLE patients. A data splitting of full pharmacokinetic profiles sampled in 67 children extracted either from the expert system ISBA (n = 34) or the hospital Saint Louis (n = 33) was performed into train (75%) and test (25%) sets. A POPPK was developed for MPA in the train set using a NLME and the SAEM algorithm and different absorption models were implemented and compared (first order, transit, or simple and double gamma). The best limited sampling strategy was then determined in the test set using a maximum-a-posteriori Bayesian method to estimate individual PK parameters and AUC based on three blood samples compared to the reference AUC calculated using the trapezoidal rule applied on all samples and performances were assessed in the test set. Mean patient age and dose was 13years old (5-18) and 18.1mg/kg (7.9-47.6), respectively. MPA concentrations (764) from 107 occasions were included in the analysis. A double gamma absorption with a first-order elimination from the central compartment best fitted the data. The optimal LSS with samples at 30min, 2h, and 3h post-dose exhibited good performances in the test set (mean bias - 0.32% and RMSE 21.0%). The POPPK developed in this study adequately estimated the MPA AUC in pediatric patients with SLE based on three samples. The double absorption gamma model developed with the SAEM algorithm showed very accurate fit and reduced computation time.

FRI530 Immune-Modulation By Rapamycin, Or Mycophenolic Acid, Of Th1 And Th2 Chemokines Secretion In Retro-orbital Cells Of Patients With Graves’ Ophthalmopathy

Abstract Disclosure: G. Elia: None. F. Ragusa: None. S. Paparo: None. V. Mazzi: None. A. Patrizio: None. P. Fallahi: None. A. Antonelli: None. S. Ferrari: None. Objectives: Cytokines and chemokines contribute to the inflammation of Graves’ ophthalmopathy (GO). Indeed cytokines stimulate the T helper (Th)1 and Th2 chemokines release from orbital cells (fibroblasts, preadipocytes and myocytes).We aim to investigate the effects of rapamycin or mycophenolic acid on the secretion of these chemokines in GO orbital cells. Methods: We obtained primary cultures of fibroblasts, preadipocytes and myoblasts from the orbits of GO patients, and we tested increasing concentrations of rapamycin and/or mycophenolic acid evaluating the secretion of the prototype Th1 [chemokine (C-X-C motif) ligand 10 (CXCL10)] and Th2 [chemokine (C-C motif) ligand 2 (CCL2)] chemokines. Results: We observed that CXCL10 secretion in the retro-orbital cells of GO was undetectable in the supernatants, whereas interferon (IFN)-gamma induce its release in a dose-dependently manner. No effect was observed under the tumor necrosis factor (TNF)-alpha stimuli, while the combination of both cytokines had a significant synergistic effect on CXCL10 secretion. As regards the effect on the CCL2 secretion, it was released in low amounts in basal condition; TNF-alpha dose-dependently induced its release, whereas IFN-gamma alone had no effect. The combination of TNF-alpha plus IFN-gamma had a significant synergistic effect on CCL2 secretion. By adding mycophenolic acid, or rapamycin, (in a pharmacological range) at the time of IFN-gamma and TNF-alpha stimulation, we observed a dose-dependently inhibitory effect on the chemokines release in the retro-orbital cells of GO. Furthermore, the combination of mycophenolic acid with rapamycin, have a synergistic inhibitory effect on the release of chemokines. Conclusion: Our data showed an immune-modulating effect of mycophenolic acid or rapamycin on the Th1 and Th2 chemokines secretion, in GO orbital cells. These findings suggest the therapeutic role of these drugs that could obtained, at least in part, through this mechanism. Presentation: Friday, June 16, 2023

A Phase 0 Study Assessing the Intracranial Activity of a Metabolic Radiosensitizer in Patients with Glioblastoma

Efforts to overcome treatment resistance in glioblastoma (GBM) have been unsuccessful due to tumor heterogeneity and poor intracranial drug penetration. Targeting altered metabolism is a promising approach to improve GBM therapy despite this heterogeneity. Mycophenolate mofetil (MMF) is an inhibitor of purine synthesis that sensitizes GBM to radiation and temozolomide (TMZ) in vitro and in vivo, but its ability to cross the blood brain barrier and inhibit GBM metabolism in patients is unknown. NCT04477200 is a phase 0/1 dose escalation study of MMF combined with radiation and temozolomide in GBM. Here we report the phase 0 results of this study assessing the intracranial activity of MMF. Purine (GTP and IMP) and mycophenolic acid (MPA, the active metabolite of MMF) concentrations were determined using mass spectrometry in flash-frozen tumor (enhancing and non-enhancing) and normal cortex obtained from 8 patients with recurrent GBM who received MMF (500, 1000, 1500 and 2000 mg BID, N = 2 patients each dose level) for 1 week prior to re-resection and 5 control patients who did not receive MMF prior to re-resection. Plasma MPA concentration was similarly quantified to calculate the enhancing tumor, non-enhancing tumor and normal cortex to plasma MPA ratios. Patients who received MMF had a mean MPA concentration of 2.2 ± 0.7 µM in the enhancing tumor samples, 1.2 ± 0.5 µM in the non-enhancing tumor samples and 1.3 ± 0.5 µM in normal cortex. MPA concentration was negligible in control patients. This corresponded to tissue/plasma MPA ratios of 0.31, 0.17 and 0.10 for enhancing tumor, non-enhancing tumor and normal cortex, respectively. The GTP/IMP ratio was decreased by 75% in enhancing tumor in MMF-treated patients compared to untreated controls (p = 0.009), indicating effective target engagement and inhibition of purine synthesis. The GTP/IMP ratio was also decreased in cortex and non-enhancing tumor, though a paucity of control samples prevented statistical analysis. Twice daily MMF treatment yields intracranial drug concentrations above 1 µM and lowers the GTP/IMP ratio in GBMs, consistent with target engagement. As we have previously observed radiosensitization in vitro with MPA concentrations of 1 µM, these data suggest that MMF may achieve adequate CNS penetration for therapeutic benefit. The Phase 1 component of this study to determine the dose limiting toxicity and maximally tolerated dose of MMF when combined with reirradiation in recurrent GBM and radiation and TMZ in newly diagnosed GBM is ongoing.

Dynamic Monitoring of Intracellular Tacrolimus and Mycophenolic Acid Therapy in Renal Transplant Recipients Using Magnetic Bead Extraction Combined with LC-MS/MS.

Tacrolimus (TAC) and mycophenolic acid (MPA) are commonly used immunosuppressive therapies after renal transplant. Our objective was to quantify TAC and MPA concentrations in peripheral blood mononuclear cells (PBMCs) using liquid chromatography tandem mass spectrometry (LC-MS/MS) and to evaluate and validate the performance of the methodology. A prospective follow-up cohort study was conducted to determine whether intracellular concentrations were associated with adverse outcomes in renal transplants. PBMCs were prepared using the Ficoll separation technique and purified with erythrocyte lysis. The cells were counted using Sysmex XN-3100 and then packaged and frozen according to a 50 µL volume containing 1.0 × 106 cells. TAC and MPA were extracted using MagnaBeads and quantified using an LC-MS/MS platform. The chromatography was run on a reversed-phase Waters Acquity UPLC BEH C18 column (1.7 µm, 50 mm × 2.1 mm) for gradient elution separation with a total run time of 4.5 min and a flow rate of 0.3 mL/min. Mobile phases A and B were water and methanol, respectively, each containing 2 mM ammonium acetate and 0.1% formic acid. Renal transplant recipients receiving TAC and MPA in combination were selected for clinical validation and divided into two groups: a stable group and an adverse outcome group. The concentrations were dynamically monitored at 5, 7, 14, and 21 days (D5, D7, D14, and D21) and 1, 2, 3, and 6 months (M1, M2, M3, and M6) after operation. Method performance validation was performed according to Food and Drug Administration guidelines, showing high specificity and sensitivity. The TAC and MPA calibration curves were linear (r2 = 0.9988 and r2 = 0.9990, respectively). Both intra-day and inter-day imprecision and inaccuracy were less than 15%. Matrix effects and recoveries were satisfactory. The TAC and MPA concentrations in 304 "real" PBMC samples from 47 renal transplant recipients were within the calibration curve range (0.12 to 16.40 ng/mL and 0.20 to 4.72 ng/mL, respectively). There was a weak correlation between PBMC-C0TAC and WB-C0TAC (p < 0.05), but no correlation was found for MPA. The level of immunosuppressive intra-patient variation (IPV) was higher in PBMC at 77.47% (55.06, 97.76%) than in WB at 34.61% (21.90, 49.85%). During the dynamic change in C0TAC, PBMC-C0TAC was in a fluctuating state, and no stable period was found. PBMC-C0TAC did not show a significant difference between the stable and adverse outcome group, but the level of the adverse outcome group was generally higher than that of the stable group. Compared with conventional therapeutic drug monitoring, the proposed rapid and sensitive method can provide more clinically reliable information on drug concentration at an active site, which has the potential to be applied to the clinical monitoring of intracellular immunosuppressive concentration in organ transplantation. However, the application of PBMC-C0TAC in adverse outcomes of renal transplant should be studied further.

Significant Effects of Renal Function on Mycophenolic Acid Total Clearance in Pediatric Kidney Transplant Recipients with Population Pharmacokinetic Modeling.

Mycophenolic acid (MPA) is an immunosuppressant commonly prescribed in pediatric kidney transplantation to prevent graft rejection. Large variabilities in MPA plasma exposures have been observed in this population, which could result in severe adverse effects. The majority of the MPA pharmacokinetic data have been reported in adult populations, whereas information in pediatric patients is still very limited. The objective of this study was to establish a novel, nonlinear mixed-effects model for MPA and investigate the clinical variables affecting MPA population pharmacokinetics in pediatric kidney transplant recipients. Data were collected retrospectively from pediatric kidney transplant patients (≤ 18 years when MPA concentrations were initially collected; on oral administration of mycophenolate mofetil) in Calgary, Alberta, Canada. Nonlinear mixed-effect modeling was conducted using stochastic approximation expectation-maximization in Monolix 2021R2 (Lixoft SAS, France) to determine population pharmacokinetic estimates, interindividual variabilities, and interoccasional variabilities. Covariate models were constructed using the Model Proposal function in Monolix in conjunction with a systematic stepwise inclusion/elimination protocol. The best model was selected based on objective function values, relative standard errors, goodness-of-fit plots, prediction-corrected visual predictive checks, and numerical predictive checks. A total of 50 pediatric kidney transplant patients (25 female) with 219 MPA plasma concentration-time profiles were included. The average age (± standard deviation) and posttransplant time for the sample population were 12.8 ± 4.8 years and 762 ± 1160 days, respectively. The majority of study subjects (i.e., > 85% based on all occasions) were co-administered tacrolimus. A two-compartment, first-order absorption with lag time and linear elimination structural model with lognormal distributed proportional residual errors best described the MPA concentration-time data. The absorption rate constant (2.52 h-1 or 0.042 min-1), lag time (0.166 h or 9.96 min), volumes of distributions of the central (22.8 L) and peripheral (216 L) compartments, and intercompartment clearance (17.6 Lh-1 or 0.293 Lmin-1) were consistent with literature values; whereas total MPA clearance (0.72 Lh-1 or 0.012 Lmin-1) was relatively reduced, likely due to the general lack of cyclosporine interactions and the stabilized graft functions from significantly longer posttransplant time in our sample population. Of the clinical variables tested, only estimated glomerular filtration rate (eGFR) was identified a significant covariate affecting total MPA clearance with a positive, exponential relationship. The final population pharmacokinetic model was successfully evaluated/validated using a variety of complementary methods. We have successfully constructed and validated a novel population pharmacokinetic model of MPA in pediatric kidney transplant patients. A positive, nonlinear relationship between eGFR and total MPA clearance identified in our model is likely attributed to multiple concurrent mechanisms, which warrant further systematic investigations.

Therapeutic drug monitoring of mycophenolic acid (MPA) using volumetric absorptive microsampling (VAMS) in pediatric renal transplant recipients: ultra-high-performance liquid chromatography-tandem mass spectrometry analytical method development, cross-validation, and clinical application

BackgroundMycophenolic acid (MPA) is widely used in posttransplant pharmacotherapy for pediatric patients after renal transplantation. Volumetric absorptive microsampling (VAMS) is a recent approach for sample collection, particularly during therapeutic drug monitoring (TDM). The recommended matrix for MPA determination is plasma (PL), and conversion between capillary-blood VAMS samples and PL concentrations is required for the appropriate interpretation of the results.MethodsThis study aimed to validate and develop a UHPLC-MS/MS method for MPA quantification in whole blood (WB), PL, and VAMS samples, with cross and clinical validation based on regression calculations. Methods were validated in the 0.10–15 µg/mL range for trough MPA concentration measurement according to the European Medicines Agency (EMA) guidelines. Fifty pediatric patients treated with MPA after renal transplantation were included in this study. PL and WB samples were obtained via venipuncture, whereas VAMS samples were collected after the fingerstick. The conversion from VAMSMPA to PLMPA concentration was performed using formulas based on hematocrit values and a regression model.ResultsLC–MS/MS methods were successfully developed and validated according to EMA guidelines. The cross-correlation between the methods was evaluated using Passing-Bablok regression, Bland–Altman bias plots, and predictive performance calculations. Clinical validation of the developed method was successfully performed, and the formula based on regression was successfully validated for VAMSMPA to PLMPA concentration and confirmed on an independent group of samples.ConclusionsThis study is the first development of a triple matrix-based LC–MS/MS method for MPA determination in the pediatric population after renal transplantation. For the first time, the developed methods were cross-validated with routinely used HPLC–DAD protocol.Graphical

Abstract 1093: Measuring and inhibiting purine metabolism in patients with glioblastoma

Abstract Introduction: We have previously found that purine metabolism is an important metabolic mediator of treatment resistance in glioblastoma (GBM). Mycophenolate mofetil (MMF), an inhibitor of purine synthesis, synergizes with radiation and temozolomide in mouse models of GBM. No measurements of purine synthesis have been performed in human cancer patients, nor has the purine inhibitor MMF been used clinically for patients with GBM. Methods: We infused clinical-grade 13C6 glucose into patients undergoing craniotomy for presumed GBM and used mass spectrometry to measure carbon incorporation into purines in cortex and disparate tumor regions. We are conducting a phase 0/1 trial (NCT04477200) to determine the safety and potential efficacy of the purine inhibitor MMF in patients with GBM. In the phase 0 arm, patients with recurrent GBM needing tumor re-resection receive MMF for one week prior to surgery and tumor and brain tissue is analyzed by mass spectrometry to determine drug concentrations and target engagement. In the phase 1 portions, patients (age &amp;gt;18, KPS ≥ 60) with recurrent GBM are treated with MMF and a second course of radiation while patients with newly diagnosed GBM are treated with radiation (RT), temozolomide (TMZ) and MMF followed by cyclic MMF and TMZ. A TITE-CRM algorithm is employed to dose-escalate MMF (range 500-2000 mg BID) and determine its maximum tolerated dose in combination with RT and/or TMZ. Results: Stable isotope tracing has been performed on 8 patients with suspected GBM. Purine synthesis was increased in tumor tissue compared to normal cortex in all patients, with most patients having 10-30-fold increased purine labeling in tumor. We have enrolled 8 patients on our phase 0 peri-surgical study and all had active concentrations of mycophenolic acid (the active metabolite of MMF) in enhancing and non-enhancing GBM tissue. We have enrolled 28 evaluable phase 1 patients (13 recurrent, 15 new). No dose-limiting toxicities have been observed with MMF (1000-2000 mg BID) in combination with re-irradiation or cyclic TMZ. In concurrent chemoradiation with MMF, we have observed two dose limiting grade 3 toxicities at 2000 mg, both improved with treatment break. Other main toxicities are mild nausea and fatigue. Interim median overall survival in recurrent phase 1 patients is 15.6 months, which compares favorably with typical survival time of 8-10 months in this population. In the newly diagnosed cohort, median survival is not yet reached, but 14 of 15 patients are alive and without progressive tumor. Conclusions: Glucose-derived purine synthesis is elevated in human GBM compared to cortex, which could provide a favorable therapeutic index for purine synthesis inhibition. The purine synthesis inhibitor MMF achieves active concentrations in human GBM, is reasonably well tolerated in combination with standard GBM treatments, and is associated with promising preliminary clinical outcomes. Citation Format: Yoshie Umemura, Andrew J. Scott, Wajd Al-Holou, Anjali Mittal, Weihua Zhou, Kari Wilder-Romans, Jie Xu, Nathan Qi, Maureen Kachman, Michelle Kim, Larry Junck, Denise Leung, Nathan Clarke, Jason Heth, Bo Wen, Amit Pai, Vijay Tarnal, Deepak Nagrath, Theodore Lawrence, Costas A. Lyssiotis, Daniel R. Wahl. Measuring and inhibiting purine metabolism in patients with glioblastoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1093.

Evaluation of Plasma Concentrations of Mycophenolic Acid in Renal Transplant Patients Using LM1010 High-performance Liquid Chromatography

Plasma concentrations of mycophenolic acid (MPA), an immunosuppressive agent, have been measured in clinical settings using immunoassay methods or HPLC. However, immunoassay methods show cross-reactivity with metabolites of MPA glucuronide. Recently, the LM1010 high-performance liquid chromatography instrument was approved as a new general medical device. In this study, we compared the results of MPA plasma concentrations analyzed using the LM1010 method and the previously described HPLC method. Plasma samples obtained from 100 renal transplant patients (32 women and 68 men) were evaluated using both HPLC instruments. Deming regression analyses showed a very high correlation between the two instruments, with a slope of 0.9892 and an intercept of 0.0235 µg/mL (r2=0.982). Bland-Altman analysis showed an average of -0.0012 µg/mL between the LM1010 method and the previously described HPLC method. For the LM1010 method, the total run time for MPA analysis was 7 min, and the analytical time was short; however, the extraction recovery when using a spin column was extremely low for frozen plasma samples stored at -20°C for 1 month, and the volume required for the assay (150 µL) could not be collected. Thus, for the LM1010 method, analysis using fresh plasma samples was optimal. Overall, our findings showed that the LM1010 method was a rapid, accurate HPLC assay for MPA analysis and could be used in clinical practice for routine monitoring of MPA in fresh plasma samples.

The fluorescence and colorimetric dual-readout probe for clinical rapid detection of mycophenolic acid by the poly(ethylenimine)/silica-coated CdTe quantum dots

It is particularly meaningful to therapeutic drug monitoring (TDM) of mycophenolic acid (MPA) for transplant patients to maximize the drug efficacy and minimize the adverse effect. In this study, a novel fluorescence and colorimetric dual-readout probe was put forward to fast and reliable detect MPA. The blue fluorescence of MPA was largely enhanced in the presence of poly (ethylenimine) (PEI), while the red fluorescence of CdTe@SiO2 (silica-coated CdTe quantum dots) provided a reliable reference signal. Hence, combining PEI70,000 and CdTe@SiO2, a fluorescence and colorimetric dual-readout probe could be constructed. For fluorescence measurement of MPA, the linearity was obtained in the MPA concentration range of 0.5–50 μg/mL, with a limit of detection (LOD) of 33 ng/mL. For the visual detection, the fluorescent colorimetric card was established in the MPA concentration from 0.5 to 50 μg/mL corresponding to the fluorescence color from red to violet and then to blue, which could be used for semi-quantification. Furthermore, in the light of the ColorCollect APP by the smartphone, the ratio of blue and red brightness values was linear with the MPA concentration from 1 to 50 μg/mL; thus, quantification of MPA could be realized by APP with the LOD of 83 ng/mL. The developed method was successfully applied to the analysis of MPA in the plasma samples of three patients after oral administration of mycophenolate mofetil, which was the prodrug of MPA. The result was comparable to those obtained by the clinically widely-used enzyme multiplied immunoassay technique. The developed probe was fast, cost-effective and operational convenience, and possessed high potential for TDM of MPA.

Fingerprick Microsampling Methods Can Replace Venepuncture for Simultaneous Therapeutic Drug Monitoring of Tacrolimus, Mycophenolic Acid, and Prednisolone Concentrations in Adult Kidney Transplant Patients.

Kidney transplant patients undergo repeated and frequent venepunctures during allograft management. Microsampling methods that use a fingerprick draw of capillary blood, such as dried blood spots (DBS) and volumetric absorptive microsamplers (VAMS), have the potential to reduce the burden and volume of blood loss with venepuncture. This study aimed to examine microsampling approaches for the simultaneous measurement of tacrolimus, mycophenolic acid, mycophenolic acid glucuronide (MPAG), and prednisolone drug concentrations compared with standard venepuncture in adult kidney transplant patients. DBS and VAMS were simultaneously collected with venepuncture samples from 40 adult kidney transplant patients immediately before and 2 hours after immunosuppressant dosing. Method comparison was performed using Passing-Bablok regression, and bias was assessed using Bland-Altman analysis. Drug concentrations measured through microsampling and venepuncture were also compared by estimating the median prediction error (MPE) and median absolute percentage prediction error (MAPE). Passing-Bablok regression showed a systematic difference between tacrolimus DBS and venepuncture [slope of 1.06 (1.01-1.13)] and between tacrolimus VAMS and venepuncture [slope of 1.08 (1.03-1.13)]. Tacrolimus values were adjusted for this difference, and the corrected values showed no systematic differences. Moreover, no systematic differences were observed when comparing DBS or VAMS with venepuncture for mycophenolic acid and prednisolone. Tacrolimus (corrected), mycophenolic acid, and prednisolone microsampling values met the MPE and MAPE predefined acceptability limits of <15% when compared with the corresponding venepuncture values. DBS and VAMS, collected in a controlled environment, simultaneously measured multiple immunosuppressants. This study demonstrates that accurate results of multiple immunosuppressant concentrations can be generated through the microsampling approach, with a preference for VAMS over DBS.

Influence of SLCO1B1 Polymorphisms on the Pharmacokinetics of Mycophenolic Acid in Renal Transplant Recipients.

This study was designed to analyze the correlation between single nucleotide polymorphisms (SNP) related to drug metabolism and pharmacokinetics of mycophenolic acid (MPA) during long-term follow-up. A retrospective cohort study involving 71 renal transplant recipients was designed. Blood samples were collected to extract total DNAs, followed by target sequencing based on next-generation sequencing technology. The MPA area under the curve (AUC) was calculated according to the formula established in our center. The general linear model and linear regression model were used to analyze the association between SNPs and MPA AUC. A total of 689 SNPs were detected in our study, and 90 tagger SNPs were selected after quality control and linkage disequilibrium analysis. The general linear model analysis showed that 9 SNPs significantly influenced MPA AUC. A forward linear regression was conducted, and the model with the highest identical degree (r2=0.55) included 4 SNPs (SLCO1B1: rs4149036 [P < 0.0001], ABCC2: rs3824610 [P = 0.005], POR: rs4732514 [P = 0.006], ABCC2: rs4148395 [P = 0.007]) and 6 clinical factors (age [P < 0.0001], gender [P < 0.0001], the incident of acute rejection (AR) [P = 0.001], albumin [P < 0.0001], duration after renal transplantation [P = 0.01], lymphocyte numbers [P = 0.026]). The most relevant SNP to MPA AUC in this model was rs4149036. The subgroup analysis showed that rs4149036 had a significant influence on MPA AUC in the older group (P = 0.02), high-albumin group (P = 0.01), male group (P = 0.046), and both within-36-month group (P = 0.029) and after-36-month group (P = 0.041). The systematic review included 4 studies, and 2 of them showed that the mutation in SLCO1B1 resulted in lower MPA AUC, which was contrary to our study. A total of 4 SNPs (rs4149036, rs3824610, rs4148395, and rs4732514) were identified to be significantly correlated with MPA AUC. Rs4149036, located in SLCO1B1, was suggested to be the most relevant SNP to MPA AUC, which had a stronger influence on recipients who were elder, male, or with high serum albumin. Furthermore, 6 clinical factors, including age, gender, occurrence of acute rejection, serum albumin, time from kidney transplantation, and blood lymphocyte numbers, were found to affect the concentration of MPA.

Clinical Evidence on the Purported Pharmacokinetic Interactions between Corticosteroids and Mycophenolic Acid.

Corticosteroids (steroids) are commonly used concurrently with mycophenolic acid (MPA) as the first-line immunosuppression therapy for the prevention of rejection in solid organ transplantations. Steroids are also commonly administered with MPA in various autoimmune disorders such as systemic lupus erythematosus and idiopathic nephrotic syndrome. Despite various review articles having suggested the presence of pharmacokinetic interactions between MPA and steroids, definitive data have not yet been demonstrated. The aim of this Current Opinion is to critically evaluate the available clinical data and propose the optimal study design for characterising the MPA-steroid pharmacokinetic interactions. The PubMed and Embase databases were searched for relevant clinical articles in English as of September 29, 2022, where a total of 8 papers have been identified as supporting and 22 as non-supporting the purported drug interaction. To objectively evaluate the data, novel assessment criteria to effectively diagnose the interaction based on known MPA pharmacology were formulated, including the availability of independent control groups, prednisolone concentrations, MPA metabolite data, unbound MPA concentrations, and the characterisations of entero-hepatic recirculation and MPA renal clearance. Overall, the majority of the identified corticosteroid data were pertaining to prednisone or prednisolone. Our assessment indicated that no conclusive mechanistic data supporting the interaction are available in the current clinical literature, and further studies are required to quantify the effects/mechanisms of steroid-tapering or withdrawal on MPA pharmacokinetics. This current opinion provides justification for further translational investigations, as this particular drug interaction has the potential to exert significant adverse outcomes in patients prescribed MPA.