Clearance Prediction Methodology Needs Fundamental Improvement: Trends Common to Rat and Human Hepatocytes/Microsomes and Implications for Experimental Methodology

Abstract

Although prediction of clearance using hepatocytes and liver microsomes has long played a decisive role in drug discovery, it is widely acknowledged that reliably accurate prediction is not yet achievable despite the predominance of hepatically cleared drugs. Physiologically mechanistic methodology tends to underpredict clearance by several fold, and empirical correction of this bias is confounded by imprecision across drugs. Understanding the causes of prediction uncertainty has been slow, possibly reflecting poor resolution of variables associated with donor source and experimental methods, particularly for the human situation. It has been reported that among published human hepatocyte predictions there was a tendency for underprediction to increase with increasing in vivo intrinsic clearance, suggesting an inherent limitation using this particular system. This implied an artifactual rate limitation in vitro, although preparative effects on cell stability and performance were not yet resolved from assay design limitations. Here, to resolve these issues further, we present an up-to-date and comprehensive examination of predictions from published rat as well as human studies (where = 128 and 101 hepatocytes and = 71 and 83 microsomes, respectively) to assess system performance more independently. We report a clear trend of increasing underprediction with increasing in vivo intrinsic clearance, which is similar both between species and between in vitro systems. Hence, prior concerns arising specifically from human in vitro systems may be unfounded and the focus of investigation in the future should be to minimize the potential in vitro assay limitations common to whole cells and subcellular fractions.

Species differences in metabolism of a new antiepileptic drug candidate, DSP‐0565 [2‐(2′‐fluoro[1,1′‐biphenyl]‐2‐yl)acetamide]

“…Although IVIVE approaches are considered useful even in the case of species‐associated differences in metabolism, the prediction of clearance is still challenging compared with other PK parameters, such as the volume of distribution. For example, the predicted human CL int, in vitro in hepatocytes or subcellular fractions tends to be underestimated, and the reason has not been well clarified (Rawden et al, ; Riley, McGinnity, & Austin, ; Wood et al, ). Indeed, DSP‐0565 CL int, in vitro in hepatocytes was 11–15 fold smaller than CL int, in vivo in rats and dogs (Table ).…”

“…The prediction of new drug candidates pharmacokinetics (PK) in humans is important in both drug discovery and drug development. Among human PK parameters, the prediction of drug clearance is still challenging (Di et al, ; Wood, Houston, & Hallifax, ). The methodologies for the prediction of metabolic clearance can roughly be divided into two approaches: allometric scaling and in vitro – in vivo extrapolation (IVIVE).…”

Mechanistic Assessment of Extrahepatic Contributions to Glucuronidation of Integrase Strand Transfer Inhibitors

“…The in vitro Cl int,u was used to estimate whole organ Cl int as follows: in vitro Cl int,u * scaling factor (MPPGL, MPPGK, or MPPI) * organ weight (liver or kidney), where MPPGL is the microsomal protein per gram of liver, MPPGK is the microsomal protein per gram of kidney, and MPPI is the microsomal protein per total intestine. The following scaling factors were used: MPPGL of 37.69 mg mics/g of liver tissue (Wood et al, 2017) (total liver weight = 1800 g) (Davies and Morris, 1993); MPPGK of 12.8 mg mics/g of renal tissue (Al-Jahdari et al, 2006) (total kidney weight = 310 g) (Davies and Morris, 1993); and MPPI of 2935.17 mg mics/total intestine (Paine et al, 1997). The microsomal scaling factors are imbedded in the SimCYP software.…”

Hepatic Organic Anion Transporting Polypeptide–Mediated Clearance in the Beagle Dog: Assessing In Vitro–In Vivo Relationships and Applying Cross-Species Empirical Scaling Factors to Improve Prediction of Human Clearance

“…In contrast, there is minimal information on or systematic evaluation of activity of hepatic transporters in beagle dogs (Wilby et al, 2011). Studies in rats have been limited to hepatocyte investigations, and predictions of in vivo clearance have been inconsistent (Huang et al, 2010;Wood et al, 2017). Recent protein quantification by mass spectrometry has revealed interspecies differences in absolute expression levels of hepatobiliary transporters in the liver and drugmetabolizing enzymes in various species (Heikkinen et al, 2015;Wang et al, 2015).…”