Determination of low intrinsic clearance in vitro: the benefit of a novel internal standard in human hepatocyte incubations

Abstract

1. A novel method utilizing an internal standard in hepatocytes incubations has been developed and demonstrated to decrease the variability in the determination of intrinsic clearance (CL_int) in this system. The reduced variability was shown to allow differentiation of lower elimination rate constants from noise.

2. The suggested method was able to compensate for a small but systematic error (0.5 µL/min/10⁶ cells) caused by an evaporation of approximately 15% of the volume during the incubation time.

3. The approach was validated using six commercial drugs (ketoprofen, tolbutamide, phenacetin, etodolac and quinidine) which were metabolized by different pathways.

4. The suggested internal standard, MSC1815677, was extensively characterized and the acquired data suggest that it fulfills the requirements of an internal standard present during the incubation. The proposed internal standard was stable during the incubation and showed a low potential to inhibit drug metabolizing enzymes and transporters. With MSC1815677 we propose a novel simple, robust and cost-effective method to address the challenges in the estimation of low clearance in hepatocyte incubations.

Keywords:

• Word
• hepatocyte
• low clearance
• internal standard

Acknowledgments

We thank Lassina Badolo and Robert L. Walsky for their insightful scientific review and comments of the work described herein.

Disclosure statement

All authors were employed by Merck at the time this study was performed.

Additional information

Funding

This work was supported by Merck.