Skip to content
Licensed Unlicensed Requires Authentication Published by De Gruyter December 15, 2020

Immunosuppressant quantification in intravenous microdialysate – towards novel quasi-continuous therapeutic drug monitoring in transplanted patients

  • Susanne Weber ORCID logo , Sara Tombelli , Ambra Giannetti , Cosimo Trono , Mark O’Connell , Ming Wen , Ana B. Descalzo , Heike Bittersohl , Andreas Bietenbeck ORCID logo , Pierre Marquet , Lutz Renders , Guillermo Orellana , Francesco Baldini and Peter B. Luppa EMAIL logo



Therapeutic drug monitoring (TDM) plays a crucial role in personalized medicine. It helps clinicians to tailor drug dosage for optimized therapy through understanding the underlying complex pharmacokinetics and pharmacodynamics. Conventional, non-continuous TDM fails to provide real-time information, which is particularly important for the initial phase of immunosuppressant therapy, e.g., with cyclosporine (CsA) and mycophenolic acid (MPA).


We analyzed the time course over 8 h of total and free of immunosuppressive drug (CsA and MPA) concentrations measured by liquid chromatography-tandem mass spectrometry (LC-MS/MS) in 16 kidney transplant patients. Besides repeated blood sampling, intravenous microdialysis was used for continuous sampling. Free drug concentrations were determined from ultracentrifuged EDTA-plasma (UC) and compared with the drug concentrations in the respective microdialysate (µD). µDs were additionally analyzed for free CsA using a novel immunosensor chip integrated into a fluorescence detection platform. The potential of microdialysis coupled with an optical immunosensor for the TDM of immunosuppressants was assessed.


Using LC-MS/MS, the free concentrations of CsA (fCsA) and MPA (fMPA) were detectable and the time courses of total and free CsA comparable. fCsA and fMPA and area-under-the-curves (AUCs) in µDs correlated well with those determined in UCs (r≥0.79 and r≥0.88, respectively). Moreover, fCsA in µDs measured with the immunosensor correlated clearly with those determined by LC-MS/MS (r=0.82).


The new microdialysis-supported immunosensor allows real-time analysis of immunosuppressants and tailor-made dosing according to the AUC concept. It readily lends itself to future applications as minimally invasive and continuous near-patient TDM.

Corresponding author: Peter B. Luppa, Institute of Clinical Chemistry and Pathobiochemistry, Klinikum rechts der Isar, Technische Universität München, Ismaninger Str. 22, 81675 Munich, Germany, Phone: +49 89 4140 4759, E-mail:

Funding source: European Commission


We thank Christine Grubmüller and Christine Völkl for their excellent technical assistance, Francesca Salis for her help in the synthesis of CsA–O–CO2H, and Dr. Jutta Redlin for preparation of the perfusion solutions. We thank also all enrolled patients for their dedication and provision of clinical material.

  1. Research funding: This work was funded by the European Union (EU) within the project NANODEM – Nanophotonic device for multiple therapeutic drug monitoring (GA 318372) of the 7th Framework Programme. P. Marquet received research grants from Sandoz and Chiesi.

  2. Author contributions: All authors have accepted responsibility for the entire content of this submitted manuscript and approved submission.

  3. Competing interests: M. O’Connell, Chief Technical Officer and shareholder of Cornel Medical Limited. Cornel holds the IP and assets of Probe Scientific Limited, the developer of the MicroEye microdialysis device. All other authors state no conflict of interest.

  4. Informed consent: Informed consent was obtained from all individuals included in this study.

  5. Ethical approval: Research involving human subjects complied with all relevant national regulations, institutional policies and is in accordance with the tenets of the Helsinki Declaration (as revised in 2013). The Ethics Committee of the Klinikum rechts der Isar (TUM) has approved the study (565/19 S).


1. Freudenberger, K, Hilbig, U, Gauglitz, G. Recent advances in therapeutic drug monitoring of immunosuppressive drugs. Trend Anal Chem 2016;79:257–68. in Google Scholar

2. Gross, AS. Best practice in therapeutic drug monitoring. Br J Clin Pharmacol 2001;52:5–9. in Google Scholar

3. Tett, SE, Saint-Marcoux, F, Staatz, CE, Brunet, M, Vinks, AA, Miura, M, et al.. Mycophenolate, clinical pharmacokinetics, formulations, and methods for assessing drug exposure. Transplant Rev 2011;25:47–57. in Google Scholar PubMed

4. Kahan, BD, Welsh, M, Rutzky, LP. Challenges in cyclosporine therapy: the role of therapeutic monitoring by area under the curve monitoring. Ther Drug Monit 1995;17:621–4. in Google Scholar PubMed

5. Marquet, P, Åsberg, A. Individualizing transplant therapy. In: Jelliffe, RW, Nelly, M, editors. Individualized drug therapy for patients. Boston: Academic Press; 2017.10.1016/B978-0-12-803348-7.00016-2Search in Google Scholar

6. Ting, LSL, Villeneuve, E, Ensom, MHH. Beyond cyclosporine: a systematic review of limited sampling strategies for other immunosuppressants. Ther Drug Monit 2006;28:419–30. in Google Scholar PubMed

7. Nowak, I, Shaw, LM. Mycophenolic acid binding to human serum albumin: characterization and relation to pharmacodynamics. Clin Chem 1995;41:1011–7. in Google Scholar

8. Mandla, R, Line, PD, Midtvedt, K, Bergan, S. Automated determination of free mycophenolic acid and its glucuronide in plasma from renal allograft recipients. Ther Drug Monit 2003;25:407–14. in Google Scholar PubMed

9. Lemaire, M, Tillement, JP. Role of lipoproteins and erythrocytes in the in vitro binding and distribution of cyclosporin A in the blood. J Pharm Pharmacol 1982;34:715–8. in Google Scholar PubMed

10. Legg, B, Rowland, M. Cyclosporin: measurement of fraction unbound in plasma. J Pharm Pharmacol 1987;39:599–603. in Google Scholar PubMed

11. Ghareeb, M, Akhlaghi, F. Alternative matrices for therapeutic drug monitoring of immunosuppressive agents using LC-MS/MS. Bioanalysis 2015;7:1037–58. in Google Scholar

12. Akhlaghi, F, Trull, AK. Distribution of cyclosporin in organ transplant recipients. Clin Pharmacokinet 2002;41:615–37. in Google Scholar

13. Chan, S, Gerson, B. Free drug monitoring. Clin Lab Med 1987;7:279–87. in Google Scholar

14. Dasgupta, A. Usefulness of monitoring free (unbound) concentrations of therapeutic drugs in patient management. Clin Chim Acta 2007;377:1–13. in Google Scholar PubMed

15. Dasgupta, A. Monitoring free mycophenolic acid concentration: is there any clinical advantage? In: Oellerich, M, Dasgupta, A, editors. Personalized immunosuppression in transplantation. San Diego: Elsevier Inc.; 2016.10.1016/B978-0-12-800885-0.00004-7Search in Google Scholar

16. Gruzdys, V, Merrigan, SD, Johnson-Davis, KL. Feasibility of immunosuppressant drug monitoring by a microsampling device. J Appl Lab Med 2019;4:241–6. in Google Scholar PubMed

17. Avataneo, V, D’Avolio, A, Cusato, J, Cantù, M, De Nicolò, A. LC-MS application for therapeutic drug monitoring in alternative matrices. J Pharm Biomed Anal 2019;166:40–51. in Google Scholar PubMed

18. Ates, HC, Roberts, JA, Lipman, J, Cass, AEG, Urban, GA, Dincer, C. On-site therapeutic drug monitoring. Trends Biotechnol 2020. [Epub ahead of print].Search in Google Scholar PubMed

19. NANODEM European Project. NANOphotonic DEvice for Multiple therapeutic drug monitoring [Online]. Available from: [Accessed 1 Oct 2020].Search in Google Scholar

20. Berrettoni, C, Berneschi, S, Bernini, R, Giannetti, A, Grimaldi, IA, Persichetti, G, et al.. Optical monitoring of therapeutic drugs with a novel fluorescence-based POCT device. Procedia Eng 2014;87:392–5. in Google Scholar

21. Berrettoni, C, Trono, C, Tombelli, S, Giannetti, A, Berneschi, S, Baldini, F, et al.. A point-of-care device for immunosuppressants monitoring in transplanted patients. In: Compagnone, D, Baldini, F, Di Natale, C, Betta, G, Siciliano, P, editors. Sensors lecture notes in electrical engineering. Cham: Springer International Publishing; 2015.10.1007/978-3-319-09617-9_6Search in Google Scholar

22. Nandi, P, Lunte, SM. Recent trends in microdialysis sampling integrated with conventional and microanalytical systems for monitoring biological events: a review. Anal Chim Acta 2009;651:1–14. in Google Scholar PubMed PubMed Central

23. Baldini, F. Microdialysis-based sensing in clinical applications. Anal Bioanal Chem 2010;397:909–16. in Google Scholar PubMed

24. Gowers, SAN, Hamaoui, K, Vallant, N, Hanna, GB, Darzi, A, Casanova, D, et al.. An improved rapid sampling microdialysis system for human and porcine organ monitoring in a hospital setting. Anal Methods 2018;10:5273–81. in Google Scholar PubMed PubMed Central

25. Hammarlund-Udenaes, M. Microdialysis as an important technique in systems pharmacology—a historical and methodological review. AAPS J 2017;19:1294–303. in Google Scholar PubMed

26. Deters, M, Kirchner, G, Resch, K, Kaever, V. Simultaneous quantification of sirolimus, everolimus, tacrolimus and cyclosporine by liquid chromatography-mass spectrometry (LC-MS). Clin Chem Lab Med 2002;40:285–92. in Google Scholar PubMed

27. Ceglarek, U, Lembcke, J, Fiedler, GM, Werner, M, Witzigmann, H, Hauss, JP, et al.. Rapid simultaneous quantification of immunosuppressants in transplant patients by turbulent flow chromatography combined with tandem mass spectrometry. Clin Chim Acta 2004;346:181–90. in Google Scholar PubMed

28. Bittersohl, H, Herbinger, J, Wen, M, Renders, L, Steimer, W, Luppa, PB. Simultaneous determination of protein-unbound cyclosporine A and mycophenolic acid in kidney transplant patients using liquid chromatography-tandem mass spectrometry. Ther Drug Monit 2017;39:211–9. in Google Scholar PubMed

29. Giannetti, A, Adinolfi, B, Berneschi, S, Berrettoni, C, Chiavaioli, F, Tombelli, S, et al.. Optical sensing in POCT: the contribution of the Institute of Applied Physics of the Italian CNR. J Lab Med 2017;41:251–6. in Google Scholar

30. Adinolfi, B, Baldini, F, Berrettoni, C, Berneschi, S, Giannetti, A, Tombelli, S, et al.. Total internal reflection fluorescence-based optical biochip for the detection of immunosuppressants in transplanted patients. In: 2015 1st workshop on nanotechnology in instrumentation and measurement (NANOFIM). Lecce; 2015:39–42 pp.10.1109/NANOFIM.2015.8425324Search in Google Scholar

31. Leger, F, Debord, J, Le Meur, Y, Rousseau, A, Büchler, M, Lachâtre, G, et al.. Maximum a posteriori Bayesian estimation of oral cyclosporin pharmacokinetics in patients with stable renal transplants. Clin Pharmacokinet 2002;41:71–80. in Google Scholar PubMed

32. Wiesen, MHJ, Farowski, F, Feldkötter, M, Hoppe, B, Müller, C. Liquid chromatography-tandem mass spectrometry method for the quantification of mycophenolic acid and its phenolic glucuronide in saliva and plasma using a standardized saliva collection device. J Chromatogr A 2012;1241:52–9. in Google Scholar PubMed

33. Yang, H, Elmquist, WF. The binding of cyclosporin a to human plasma: an in vitro microdialysis study. Pharm Res 1996;13:622–7. in Google Scholar

34. Ward, KW, Medina, SJ, Portelli, ST, Mahar Doan, KM, Spengler, MD, Ben, MM, et al.. Enhancement of in vitro and in vivo microdialysis recovery of SB-265123 using Intralipid® and Encapsin® as perfusates. Biopharm Drug Dispos 2003;24:17–25. in Google Scholar PubMed

35. Badri, P, Dutta, S, Coakley, E, Cohen, D, Ding, B, Podsadecki, T, et al.. Pharmacokinetics and dose recommendations for cyclosporine and tacrolimus when coadministered with ABT-450, ombitasvir, and dasabuvir. Am J Transplant 2015;15:1313–22. in Google Scholar PubMed PubMed Central

36. Marquet, P. Clinical application of population pharmacokinetic methods developed for immunosuppressive drugs. Ther Drug Monit 2005;27:727–32. in Google Scholar PubMed

37. Marquet, P. Counterpoint: is pharmacokinetic or pharmacodynamic monitoring of calcineurin inhibition therapy necessary? Clin Chem 2010;56:736–9. in Google Scholar PubMed

38. Taddeo, A, Prim, D, Bojescu, E-D, Segura, J-M, Pfeifer, ME. Point-of-care therapeutic drug monitoring for precision dosing of immunosuppressive drugs. J Appl Lab Med 2020;5:738–61. in Google Scholar PubMed

Supplementary Material

The online version of this article offers supplementary material (

Received: 2020-10-16
Accepted: 2020-12-06
Published Online: 2020-12-15
Published in Print: 2021-04-27

© 2020 Walter de Gruyter GmbH, Berlin/Boston

Downloaded on 11.12.2023 from
Scroll to top button