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CNS 7056X, an ultra-short acting benzodiazepine: in vitro metabolism

Gary S Tilbrook, and Gavin J Kilpatrick

CeNeS Limited, Cambridge CB4 9ZR, Cambs, United Kingdom

Introduction: Soft drugs are compounds that are designed to be broken down rapidly in the body to pharmacologically inactive metabolites. The soft-drug approach has previously been employed in the field of anaesthesia, to generate short-acting agents such as the opiate, remifentanil. CNS 7056X is a novel benzodiazepine with an ester side-chain that is designed to be metabolised rapidly in the body by esterases. We now report on the metabolism of this compound by homogenates of tissues from various species. For comparative purposes, we also present data on the metabolism of remifentanil by liver tissue from various species.

Methods: Organs used in this study were snap-frozen after minimal post-mortem delay. Plasma was prepared and used fresh on the day of the experiment. Species used were: human (UK Human Tissue Bank, Leicester, UK), rat (Sprague Dawley), mouse (CD-1), mini-pig (Göttingen) and dog (beagle). Liver samples were pools of tissue from 3-11 separate subjects. Lung, kidney, brain and plasma were from single sources. Solutions of CNS 7056X (1uM for liver and plasma, 10uM for other tissues) or remifentanil (1uM) were added to tissue homogenates (organs: 30% w/v in phosphate buffer; plasma: 89% v/v) and incubated at 37^oC for 60min. Samples were withdrawn at various time points, these were analysed as appropriate for CNS 7056X and its carboxylic acid metabolite, CNS 7054X or remifentanil using HPLC with tandem mass spectrometric detection.

Results: In all tissues where metabolism of CNS 7056X was observed, there was a concomitant stoichiometric increase in CNS 7054X. CNS 7056X was very rapidly metabolised by human, rat, mouse and mini-pig liver (Table 1). A slower rate of metabolism was observed in dog liver. Rapid metabolism was also observed in rat and mini-pig kidney, rat lung and rat brain. CNS 7056X was stable in human, pig and dog plasma for at least 60min. CNS 7056X was metabolised in rat plasma with a half life of 15min.

Metabolism of remifentanil by the livers of several species showed a similar profile to CNS 7056X, with rapid metabolism in human, rat and mouse liver and slower metabolism by dog liver (Table 1). The metabolism of remifentanil by human liver, in particular (half life 4.2min), was very similar to that observed with CNS 7056X (half life 2.9min).

Conclusions: The carboxylic acid, CNS 7054X is the major metabolite of CNS 7056X. CNS 7056X is rapidly metabolised by human, rat, mouse and mini-pig liver tissue. A similar profile was observed for metabolism of remifentanil by liver tissue. Rapid metabolism of CNS 7056X was observed in organs other than the liver. CNS 7056X was not metabolised by plasma from human, mini-pig or dog. The lack of metabolism by human plasma indicates that CNS 7056X is a poor substrate for butyrylcholinesterase or paraoxonase (1). The tissue profile observed is in keeping with the hypothesis that CNS 7056X is a substrate for carboxylesterases (1,2).

References:

1. Li B et al., Biochem Pharmacol 70, 1673-1684 (2005).

2. Xie et al., Drug Met Dispos 30, 541-547 (2002).