Quantification of T Cell Binding Polyclonal Rabbit Anti-thymocyte Globulin in Human Plasma with Liquid Chromatography Tandem-Mass Spectrometry.

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From: The AAPS Journal(Vol. 22, Issue 2)
Publisher: Springer
Document Type: Report
Length: 406 words

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Keywords: KEY WORDS Anti-thymocyte globulin; Immunoaffinity interaction; Jurkat T cell line; Liquid chromatography tandem-mass spectrometry; Polyclonal antibody; Quantification Abstract The addition of rabbit anti-human thymocyte globulin (ATG) to the conditioning regimen prior to allogeneic hematopoietic cell transplantation has significantly reduced the risk of graft-versus-host disease (GvHD) and graft failure. However, ATG has a small therapeutic window. Overexposure of ATG post-HCT hampers T cell immune reconstitution and has been associated with increased relapse rates and viral reactivations, whereas underexposure has been associated with an increased incidence of GvHD, both of which lead to increased mortality. Therapeutic drug monitoring of T cell binding ATG plasma levels provides a means to optimize dosing for patients at high risk for graft failure to ensure timely T cell immune reconstitution and subsequently increase survival chances. This manuscript describes the first liquid chromatography tandem-mass spectrometry (LC-MS/MS) method to quantify the pharmacologically active fraction of polyclonal ATG in plasma. This was achieved through immunoaffinity purification of active ATG from plasma with Jurkat T cells. After the binding and washing, samples were eluted, denatured, and trypsin-digested. Signature peptides originating from the IgG constant chain were measured with LC-MS/MS. Critical method parameters were optimized, and the method was successfully validated following European Medicines Agency (EMA) guidelines. The method covered the therapeutic range of ATG and was validated at a lower limit of quantification (LLOQ) of 1 AU/mL with an overall CV and bias of 11.8% and -2.5%, respectively. In conclusion, we developed a LC-MS/MS-based method to quantify active polyclonal rabbit ATG in human plasma. We suggest that this novel assay can be used to monitor and optimize dosing of ATG in clinical practice. Author Affiliation: (1) Department of Clinical Pharmacy, Division of Laboratory Medicine and Pharmacy, University Medical Center Utrecht, Utrecht University, P.O. Box 85500, Heidelberglaan 100, Room no. D.00.318A, Internal post no. D.00.204, 3508 GA, Utrecht, The Netherlands (2) Center for Translational Immunology, University Medical Center Utrecht, Utrecht University, Utrecht, The Netherlands (3) Princess Máxima Center for Pediatric Oncology, Utrecht, The Netherlands (4) Department of Pharmacy & Pharmacology, Netherlands Cancer Institute, Amsterdam, The Netherlands (i) E.M.vanMaarseveen@umcutrecht.nl Article History: Registration Date: 01/08/2020 Received Date: 10/22/2019 Accepted Date: 01/07/2020 Online Date: 02/06/2020 Byline: Mohsin El Amrani (1), Rick Admiraal (2, 3), Lobke Willaert (1), Lysette J. C. Ebskamp-van Raaij (2), Amelia M. Lacna (2), C. Erik Hack (2), Alwin D. R. Huitema (1, 4), Stefan Nierkens (2, 3), Erik M. van Maarseveen (corresponding author) (1, i)

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Gale Document Number: GALE|A628195942