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Metabolism of remimazolam in primary human hepatocytes during continuous long-term infusion in a 3-D bioreactor system

Authors Freyer N, Knöspel F, Damm G, Greuel S, Schneider C, Seehofer D, Stöhr T, Petersen KU, Zeilinger K

Received 7 September 2018

Accepted for publication 18 January 2019

Published 2 April 2019 Volume 2019:13 Pages 1033—1047


Checked for plagiarism Yes

Review by Single anonymous peer review

Peer reviewer comments 3

Editor who approved publication: Dr Sukesh Voruganti

Nora Freyer,1 Fanny Knöspel,1 Georg Damm,2 Selina Greuel,1 Christin Schneider,1 Daniel Seehofer,2 Thomas Stöhr,3 Karl-Uwe Petersen,3 Katrin Zeilinger1

1Berlin Brandenburg Center for Regenerative Therapies (BCRT), Charité – Universitätsmedizin Berlin, Berlin 13353, Germany; 2Department of Hepatobiliary Surgery and Visceral Transplantation, University of Leipzig, Leipzig 04103, Germany; 3PAION Deutschland GmbH (Non-clinical development), Aachen 52062, Germany

Background: Remimazolam is an ultra-short acting benzodiazepine under development for procedural sedation and general anesthesia. It is hydrolyzed by CES1 to an inactive metabolite (CNS7054).
Purpose: In this study, the effect of continuous remimazolam exposure on its metabolism and on CES1 expression was investigated in a dynamic 3-D bioreactor culture model inoculated with primary human hepatocytes.
Methods: Remimazolam was continuously infused into bioreactors for 5 days at a final concentration of 3,000 ng/ml (6.8 µM). In parallel, 2-D cultures were run with cells from the same donors, but with discontinuous exposure to remimazolam.
Results: Daily measurement of clinical chemistry parameters (glucose, lactate, urea, ammonia, and liver enzymes) in culture supernatants indicated no noxious effect of remimazolam on hepatocyte integrity as compared to untreated controls. Concentrations of remimazolam reached steady-state values of around 250 ng/ml within 8 hours in 3-D bioreactors whereas in 2-D cultures remimazolam concentrations declined to almost zero within the same time frame. Levels of CNS7054 showed an inverse time-course reaching average values of 1,350 ng/ml in perfused 3-D bioreactors resp. 2,800 ng/ml in static 2-D cultures. Analysis of mRNA expression levels of CES1 indicated no changes in gene expression over the culture period.
Conclusion: The results indicated a stable metabolism of remimazolam during 5 days of continuous exposure to clinically relevant concentrations of the drug. Moreover, there was no evidence for a harmful effect of remimazolam exposure on the integrity and metabolic activity of in vitro cultivated primary human hepatocytes.

Keywords: benzodiazepine, metabolite, CNS7054, carboxylesterase 1, steady state

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