Abstract
In vitro-in vivo extrapolation of drug metabolism data obtained in enriched preparations of subcellular fractions rely on robust estimates of physiologically relevant scaling factors for the prediction of clearance in vivo. The purpose of the current study was to measure the microsomal and cytosolic protein per gram of kidney (MPPGK and CPPGK) in dog and human kidney cortex using appropriate protein recovery marker and evaluate functional activity of human cortex microsomes. Cytochrome P450 (CYP) content and glucose-6-phosphatase (G6Pase) activity were used as microsomal protein markers, whereas glutathione-S-transferase activity was a cytosolic marker. Functional activity of human microsomal samples was assessed by measuring mycophenolic acid glucuronidation. MPPGK was 33.9 and 44.0 mg/g in dog kidney cortex, and 41.1 and 63.6 mg/g in dog liver (n = 17), using P450 content and G6Pase activity, respectively. No trends were noted between kidney, liver, and intestinal scalars from the same animals. Species differences were evident, as human MPPGK and CPPGK were 26.2 and 53.3 mg/g in kidney cortex (n = 38), respectively. MPPGK was 2-fold greater than the commonly used in vitro-in vivo extrapolation scalar; this difference was attributed mainly to tissue source (mixed kidney regions versus cortex). Robust human MPPGK and CPPGK scalars were measured for the first time. The work emphasized the importance of regional differences (cortex versus whole kidney–specific MPPGK, tissue weight, and blood flow) and a need to account for these to improve assessment of renal metabolic clearance and its extrapolation to in vivo.
Highlights
Microsomal and cytosolic protein contents in tissues of human and preclinical species are used as scaling factors for in vivo extrapolation (IVIVE) of microsomal metabolism data to predict drug in vivo clearance
Data on the cytosolic protein in human kidney and the microsomal and cytosolic protein in preclinical species are lacking
The microsomal and cytosolic protein content was measured in 38 human kidney cortex samples
Summary
Part of this work was previously presented as a poster presentation at the 2015 American Association of Pharmaceutical Scientists annual meeting (Oct. 25–29, 2015, Orlando, FL). S This article has supplemental material available at dmd.aspetjournals.org This approach relies on robust estimates of physiologically relevant scaling factors, including the protein content of the subcellular fraction in the tissue of interest. Liver scaling factors have been well characterized [e.g., microsomal (MPPGL) and cytosolic (CPPGL) protein per gram of liver] for human and several preclinical species (Houston, 1994; Barter et al, 2007; Smith et al, 2008; Cubitt et al, 2011), fewer data have been reported for extrahepatic tissues, such as the kidney (Gill et al, 2012; Scotcher et al, 2016a,b). No data exist for CPPGK in humans, and an estimate of cytosolic protein content of liver is currently used as a surrogate for IVIVE (Säll et al, 2012; Nishimuta et al, 2014)
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