Therapeutic drug monitoring (TDM) of Leflunomide as an immunosuppressive treatment in thoracic (cardiac and/or lung) transplant recipients at The Georges POMPIDOU European Hospital-Paris-France

 Corresponding author: Ismail Faiz. Copyright © 2025 Author(s) retain the copyright of this article. This article is published under the terms of the Creative Commons Attribution Liscense 4.0. Therapeutic drug monitoring (TDM) of Leflunomide as an immunosuppressive treatment in thoracic (cardiac and/or lung) transplant recipients at The Georges POMPIDOU European Hospital-Paris-France Ismail Faiz 1, 5, 6, *, Eliane M. Billaud 1, Sandrine Lefeuvre 1, Catherine Amrein 2, Romain Guillmain 3, Agnès LILLO-LE Louet 4, El-Houcine Sebbar 5, 6, Dounia EL-Moujtahide 5, 6 and Mohammed Choukri 5, 6 1 Toxicology-Pharmacology Laboratory, Georges POMPIDOU European Hospital, Paris, France. 2 Thoracic Surgery and Cardiopulmonary Transplantation Department, Georges POMPIDOU European Hospital, Paris, France. 3 Functional thoracic transplantation unit (FTTU), Georges POMPIDOU European Hospital, Paris, France. 4 Pharmacovigilance Department, Georges POMPIDOU European Hospital, Paris, France. 5 Biochemistry Laboratory, Mohammed VI University Hospital, Oujda, Morocco. 6 Faculty of Medicine and Pharmacy, Mohammed the First University, Oujda, Morocco. World Journal of Biology Pharmacy and Health Sciences, 2025, 21(03), 018-031 Publication history: Received on 11 January 2025; revised on 24 February 2025; accepted on 27 February 2025 Article DOI: https://doi.org/10.30574/wjbphs.2025.21.3.0220 Abstract Leflunomide is an immunosuppressant indicated in the treatment of rheumatoid arthritis. This drug has a particular pharmacokinetics. Therapeutic Drug Monitoring (TDM) has been recommended for this drug because of its hepatic and hematological toxicities. We present here the prospective and retrospective analysis of a preliminary experience of the use of leflunomide as an alternative immunosuppressant in heart and/or lung transplantation of patients with or without cystic fibrosis. This study was conducted in 17 heart transplant patients (n=8) and/or lung transplant patients (n=9, 7 of whom had cystic fibrosis) who received treatment with leflunomide between April 2005 and June 2008. The indication for leflunomide was generally intolerance to the other immunosuppressants. The residual concentrations measured in patients with cystic fibrosis (C0 = 12.8 ± 5.5 mg/L) were statistically lower than those measured in patients without cystic fibrosis (C0 = 44.0 ± 24.2 mg/L) (p < 0.05). However, the dose related to weight in patients with cystic fibrosis (D = 0.32 ± 0.08 mg/Kg) tends to be slightly higher than that in patients without cystic fibrosis (D = 0.26 ± 0.10 mg/Kg). In terms of evolution, two patients died, one patient was lost to follow-up and leflunomide was stopped in 2 patients. With a mean follow-up of 12 months, the outcome was acceptable in the 12 patients in whom treatment was maintained. This experience must be evaluated over the longer term so that it can be extended to a larger cohort or proposed earlier after transplantation. Keywords: Therapeutic Drug Monitoring; Leflunomide; Immunosuppressant; Cystic Fibrosis; Transplantation 1. Introduction Therapeutic Drug Monitoring (TDM) consists of monitoring the blood concentrations of some drugs to be in the therapeutic range to avoid a possible underdose that would lead to therapeutic ineffectiveness or a possible overdose that would increase toxicity. The TDM mainly concerns drugs with a narrow therapeutic index with a large interindividual variability in terms of pharmacokinetics and/or pharmacodynamics. This process allows the individualization of treatment to find the appropriate dose for each patient that allows a target concentration to be reached within the therapeutic range while avoiding toxic thresholds (1)(2). At the Georges POMPIDOU European Hospital, the Cardiovascular Surgery department adopted a distinct immunosuppressive strategy depending on the type World Journal of Biology Pharmacy and Health Sciences, 2025, 21(03), 018-031 19 of transplant (cardiac or pulmonary). Indeed, heart transplant patients were usually put on corticosteroids and ciclosporin ± mycophenolate mofetil, and in case of intolerance, patients were switched to an inhibitor of proliferation signal (everolimus or sirolimus). While lung transplant patients were put on corticosteroids and tacrolimus ± mycophenolate mofetil and as previously, directed towards everolimus or sirolimus in case of intolerance or failure. But the department's strategy had changed following the observation of serious adverse effects, especially with the inhibitors of proliferation signal, which prompted doctors to prescribe leflunomide as an immunosuppressive alternative to these patients. Leflunomide (N-(4-trifluoromethylphenyl)-5-methylisoxazol-4-carboxamide) is an immunosuppressive drug from the malononitrilamides class, initially developed for organ transplantation (kidneys), but this development had been stopped, in particular due to a risk of liver toxicity. However, the development of this drug continued in the indication of autoimmune diseases such as rheumatoid arthritis, leading to marketing in this indication (MA obtained on 09/02/1999) (3). But its use is complicated by specific pharmacokinetics because it is a prodrug that will release an active metabolite, teriflunomide "A77 1726" which has a very long half-life (2 weeks). It is necessary to wait several weeks (12 weeks) to reach the plasma balance state, which will complicate the TDM. In addition, this product has a strong binding to plasma proteins, particularly to albumin. Elimination is mainly by metabolization in the liver with the formation of a glucuron-conjugated derivative which will be excreted in the urine. There is a part of biliary excretion of teriflunomide in the unchanged form which will be found in the feces, accompanied by a significant enterohepatic cycle. The clearance of this product is mainly hepatic, also requiring 12 weeks after stopping administration for the body to eliminate this drug, which penalizes the flexibility of use (4). The therapeutic dose indicated in rheumatoid arthritis is 10 mg to 20 mg per day after a loading dose of 100 mg per day for 3 consecutive days, to reach a steady-state concentration of around 35 mg/L (3)(5). The monitoring methods include a TDM due to his hepatotoxicity and also his hematotoxicity and nephrotoxicity, which implies the monitoring of hepatic and renal functions as well as the blood count. This TDM is also necessary because of the teratogenic risk which, combined with the long half-life, requires information on the negative circulating concentrations in both men and women before starting procreation. In case of lung transplantation in patients with cystic fibrosis, particularly pediatric patients, the risk of prolonged underdosing due to increased clearance and potential decreased absorption must be assessed, which will justify special monitoring of blood concentrations in these patients to avoid possible graft rejection or poor control of a possible BK virus infection. The objective of our study is to retrospectively describe the cohort of cardiac and/or pulmonary transplant patients at The Georges POMPIDOU European Hospital who received leflunomide as an alternative immunosuppressant in order to determine the optimal dosage regimen and monitoring methods during treatment. 2. Materials and Methods 2.1. Inclusion criteria Heart and/or lung transplant patients treated with leflunomide as part of immunosuppressive therapy and/or patients with BK virus infection between April 2005 and June 2008. 2.2. Studied population This study was conducted on 17 heart transplant patients (n=8) and/or lung transplant patients (n=9, 7 of whom had cystic fibrosis) from The Georges POMPIDOU European Hospital who received treatment with leflunomide. This treatment with leflunomide was always associated with other immunosuppressants (Ciclosporin for heart transplant patients and Tacrolimus for lung transplant patients, ± mycophenolate mofetil ± corticosteroids ± everolimus ± sirolimus. The demographic characteristics of this population are detailed (Table 1). Table 1 Demographic characteristics of the studied population are listed in chronological order to the initiation of leflunomide in our center Patient Sex Age (years) Weight (kg) Cystic fibrosis Type of transplantation Transplantation date Hemodialysis 1 M 54 92 no Heart-Lung 01/31/2004 2 M 61 78 no Heart 10/07/2003 3 M 30 54 yes Lung-Liver 05/02/2005 4 M 33 54 yes Lung 07/05/2005 yes 5 M 68 93 no Heart 01/04/1990 World Journal of Biology Pharmacy and Health Sciences, 2025, 21(03), 018-031 20 6 M 19 60 yes Lung 06/24/2004 7 F 55 70 no Heart-Lung 03/17/1999 8 M 53 112 no Heart 11/10/1994 9 M 58 69 no Heart 09/09/1995 10 M 37 47 yes Lung 08/22/1995 11 M 55 67 no Heart 01/01/2006 12 F 48 48 no Lung 10/18/2003 13 M 31 61 yes Lung 08/24/2001 14 M 14 25 yes Lung 05/03/2002 yes 15 F 60 52 no Lung 12/30/2006 16 M 60 75 no Heart 10/30/1988 17 M 24 53 yes Lung-Liver 11/21/2006 2.3. Data collection The study was based on the collection of patient data from their medical records, this collection highlighted the following information: individual clinical indication for leflunomide, date of initiation and duration of treatment, initial dose as well as any subsequent dosage adjustments, residual plasma concentrations of teriflunomide "active metabolite of leflunomide", assessment of hepatotoxicity "dosing of AST, ALT, and total bilirubin", assessment of hematotoxicity "leukocytes, platelets, and hemoglobin level", assessment of nephrotoxicity "creatininemia, estimated glomerular filtration rate", collection of other unusual or serious adverse effects affecting patients during treatment, analysis of the outcome of patients under leflunomide, collection of doses and corresponding concentrations of other immunosuppressants before the administration of leflunomide and at 3 months "considered the time for plasma balance" after the introduction of leflunomide). 2.4. Plasma dosing of teriflunomide The dosage of teriflunomide, the active metabolite of leflunomide, was carried out by High-Performance Liquid Chromatography (HPLC) on the plasma of patients after deproteinization of the sample by acetonitrile (6). Detection is done in UV at the wavelength λ = 295 nm. 2.5. Statistical analysis of data Comparison of mean doses between patients with and without cystic fibrosis was performed by a student’s t-test on an unpaired series (p<0.05). A comparison of mean concentrations between patients with and without cystic fibrosis was performed by a student’s t-test on an unpaired series with Welch's correction (p<0.05). 3. Results 3.1. Clinical indication of leflunomide treatment The indication for leflunomide treatment collected in each patient's file is indicated (Table 2). Table 2 Clinical indication of leflunomide treatment Patient The time between transplantation and initiation of Leflunomide (years) Indication of Leflunomide 1 1.2 Leukopenia under Mycophenolate mofetil 2 1.7 Renal failure + mouth ulcers under Sirolimus + acute cellular rejections + repeated infections World Journal of Biology Pharmacy and Health Sciences, 2025, 21(03), 018-031 21 3 0.3 Mycophenolate mofetil toxicity (leukopenia, thrombocytopenia) 4 0.3 Neutropenia + thrombotic microangiopathy 5 16.2 Renal failure + impossibility of Everolimus (Italian patient and impossibility of dosage in Italy) 6 1.7 Leukopenia under Mycophenolate mofetil 7 7.6 Severe neutropenia under Mycophenolate mofetil 8 12.6 Mouth ulcers under Everolimus 9 12.7 Renal failure + drug-induced pneumonitis with Everolimus 10 12.8 Renal failure +++ (contraindication to treatment With Tacrolimus and corticosteroids) 11 12.8 Renal failure +++ and as soon as the doses of Ciclosporin are reduced -> rejection 12 12.9 Significant diarrhea with the combination of Everolimus + Mycophenolate mofetil 13 13 Episodes of severe neutropenia (bone marrow suppression) under Mycophenolate mofetil 14 13.1 BK-Virus Nephropathy 15 13.2 Renal failure (discontinuation of Everolimus) + reactivation of CMV 16 13.3 Renal failure + mouth ulcers under Everolimus 17 13.3 Leukopenia under Everolimus 3.2. Analysis of exposure to leflunomide Dates and durations of leflunomide treatment are listed in (Table 3). Treatment durations ranged from 1 to 33 months. Two patients died, one patient was lost to follow-up after a major clinical event, and a fourth patient had treatment stopped following a bi-nephrectomy for control of both severe high blood pressure and BK virus infection. Table 3 Duration and outcome of leflunomide treatment Patient Date of initiation under leflunomide Initial dose of leflunomide (mg/24h) Duration of Treatment with leflunomide (months) Becoming patients 1 04/12/2005 10 12 Dead 2 06/19/2005 10 1 Stop after one month 3 09/03/2005 10 33 current medication 4 10/27/2005 20 32 current medication 5 12/14/2005 20 30 current medication 6 03/02/2006 20 27 current medication 7 09/21/2006 20 17 lost sight of 8 04/18/2007 20 24 Stop after 24 months 9 06/20/2007 20 12 Stop after 12 months 10 09/17/2007 10 9 Stop after 9 months World Journal of Biology Pharmacy and Health Sciences, 2025, 21(03), 018-031 22 11 01/24/2008 20 6 Stop after 6 months 12 02/19/2008 20 6 Stop after 6 months 13 03/19/2008 20 6 Stop after 6 months 14 04/30/2008 10 3 Planned stop 15 05/15/2008 20 1 Dead 16 05/28/2008 20 3 Stop after 3 months 17 05/29/2008 20 3 Stop after 3 months The initiation doses and their dosage adjustments are collected (Table 4). Table 4 Adaptation of doses (mg/24h) of leflunomide based on the TDM Patient Month 1 Month 1,5 Month 2 Month 3 Month 6 Year 1 Year 2 1 10 10 20 20 20 10 dead 2 10 Stop after one month 3 10 10 10 20 20 20 20 4 20 then 10 10 10 10 20 20 20 5 20 20 20 20 20 20 40 then 20 6 20 20 20 20 20 20 20 7 20 20 20 20 20 20 lost sight of 8 20 20 20 20 20 20 20 9 20 20 20 20 20 20 Stop after 12 months 10 10 then 20 20 20 20 40 Stop after 9 months 11 20 20 20 20 20 Stop after 6 months 12 20 20 20 20 20 Stop after 6 months 13 20 20 20 20 20 Stop after 6 months 14 10 then 20 30 40 50 Planned stop 15 20 Dead 16 20 20 20 20 Stop after 3 months 17 20 20 20 20 Stop after 3 months 12 of the 17 patients included were put on leflunomide at the initial dose of 20 mg/24 h. The other 5 patients, whose initial dose was 10 mg/24 h, underwent an adaptation and an increase in the dose up to the dose of 20 mg/24 h in the majority of cases. The mean doses of 16 ± 5 mg/24 h and 18 ± 4 mg/24 h respectively in patients with and without cystic fibrosis are comparable. The dose relative to weight in patients with cystic fibrosis (D = 0.32 ± 0.08 mg/kg/24h) is not statistically different but tends to be slightly higher than that calculated in patients without cystic fibrosis (D = 0.26 ± 0.10 mg/kg/24h). The study of the evolution of leflunomide concentrations over time (Table 5, Figure 1) confirms that the treatment is balanced after 3 months of exposure. World Journal of Biology Pharmacy and Health Sciences, 2025, 21(03), 018-031 23 Table 5 Evolution of leflunomide concentrations (mg/L) over time (muco + = Cystic fibrosis) Patient Month 1 Month 1,5 Month 2 Month 3 Month 6 Year 1 Year 2 1 2 3 4 (muco +) 8.2 5 197 6 (muco +) 25.5 7 8 68 9 20.5 22.5 10 (muco +) 11.4 11 38.8 67.3 79.5 12 13 (muco +) 6.7 14 5 7.8 11.3 14 19.6 15 16 19 37 17 (muco +) 16 13.8 15 Figure 1 Evolution of leflunomide concentrations (mg/L) over time in patients with (Muco+) or not (Muco-) cystic fibrosis On the other hand, the residual concentrations measured in patients with cystic fibrosis (C0 = 12.8 ± 5.5 mg/L) are statistically lower than those measured in patients without cystic fibrosis (C0 = 44.0 ± 24.2 mg/L) (p<0.05). The statistical analysis was performed by excluding patient 5 due to a significant overdose at 197 mg/L. Patient 14, although not suffering from cystic fibrosis, had a low exposure at 19.6 mg/L for which we have no explanation 3.3. Analysis of tolerance to leflunomide Tolerance to leflunomide was assessed by analyzing patients' biological constants after they were started on leflunomide (Table 6). World Journal of Biology Pharmacy and Health Sciences, 2025, 21(03), 018-031 24 Table 6 Biological parameters corresponding to each patient Creatininemia (µmol/L) GFR (ml/min) Total bilirubin (µmol/L) AST (UI/L) ALT (UI/L) Leukocytes (G/L) Platelets (G/L) Hemoglobin (g/L) Normal values M = 60 à 115 F = 45 à 105 >60 5 à 17 4 à 40 4 à 40 4 à 10 150 à 450 M= 130 à 170 F= 120 à 160 Patient Month 1 M0 139 7 15 24 7.1 132 123 M3 123 8 12 15 2.7 142 94 M6 125 10 14 17 10.1 153 136 M12 141 48 6 11 14 7.2 189 107 2 M0 254 12 21 21 2.6 148 106 M3 170 8 10 6 3.3 206 106 M6 141 9 24 26 5.2 162 102 M12 270 22 8 17 16 5.8 136 128 3 M0 114 9 30 98 10.4 20 129 M3 134 8 21 50 6.8 17 113 M6 113 70 11 14 50 3.1 19 118 M12 143 53 12 21 27 1.3 21 107 4 M0 340 10 108 311 4 153 113 M3 382 17 8 15 25 6 196 97 M6 650 9 10 30 48 6 153 129 M12 424 15 9 42 58 4.8 194 107 5 M0 118 23 22 21 7.4 186 131 M6 118 57 25 33 35 5.8 232 122 M12 117 57 19 22 11 7.1 227 121 6 M0 115 76 19 11 13 3.9 203 119 World Journal of Biology Pharmacy and Health Sciences, 2025, 21(03), 018-031 25 M3 153 54 18 14 16 3.8 181 119 M6 152 55 5.4 185 106 M12 189 42 22 19 20 5.9 151 119 7 M0 188 26 3.9 341 104 M3 132 39 11 25 25 3.9 307 106 M6 126 41 13 19 24 4.2 313 102 8 M0 168 40 13 24 18 7.6 217 138 M12 171 39 9 28 27 4.2 181 123 9 M0 189 34 16 14 15 9.7 210 153 M12 257 24 12 29 35 4 140 127 10 M0 194 36 8 33 36 5.3 161 122 11 M0 181 36 63 136 6.8 198 114 M3 183 36 58 115 4 177 107 M6 130 53 4.3 160 94 12 M0 268 18 8 30 14 7.9 260 89 M3 190 26 M6 151 34 5 34 28 2.5 369 89 13 M0 116 68 9 32 76 3.8 274 145 M6 90 91 10 31 60 7 192 154 14 M0 508 15 9 29 40 9.8 287 109 15 M0 123 41 28 24 5.7 231 88 16 M0 204 31 25 25 4.1 131 109 17 M0 89 97 13 26 2 282 91 M3 88 98 4 18 31 3.8 156 104 World Journal of Biology Pharmacy and Health Sciences, 2025, 21(03), 018-031 26 This table shows that most patients had impaired renal function at the start of treatment with 2 cases of severe renal failure. The liver function appears to be preserved except for 4 cases with a transient increase in liver enzymes. At the hematological level, we observed 7 cases of frank leukopenia with 2 severe cases that normalized following treatment with leflunomide, and a few cases of thrombocytopenia including one severe case that persisted with leflunomide. 3.4. Other immunosuppressive treatments Daily doses of immunosuppressive treatments before leflunomide administration and 3 months after its introduction are indicated (Table 7).