SCALING FACTORS TO RELATE DRUG METABOLIC-CLEARANCE IN HEPATIC MICROSOMES, ISOLATED HEPATOCYTES, AND THE INTACT LIVER - STUDIES WITH INDUCEDLIVERS INVOLVING DIAZEPAM

Microsomal protein recovery and hepatocellularity have been determined and investigated as scaling factors for interrelating clearance by he patic microsomes, freshly isolated hepatocytes and whole liver from un treated (UT) rats and rats treated with either the cytochrome P450 ind ucer phenobarbital (PB) or dexamethasone (DEX). Hepatocellularity in U T rats (1.1 x 10(8) hepatocytes/g liver) was not significantly differe nt after either PB or DEX induction (1.1 and 1.3 x 10(8) hepatocytes/g liver, respectively), However the microsomal protein recovery index, which provides a scaling factor that is inversely related to the effic iency of the microsomal preparation procedure, was 47 mg/g liver in bo th PB and DEX microsomes and differs from UT rats (60 mg/g liver), The se contrasting findings are consistent with the interlaboratory trends in the literature, indicating that, although hepatocellularity estima tes are in good accord, microsomal recovery can vary 2-fold; this has implications for scaling, The oxidation of diazepam to its three prima ry metabolites was measured in PB and DEX microsomes and hepatocytes a nd the scaling factors were applied to these data and previously repor ted UT data, Marked changes in kinetics occur on induction resulting i n a shift in the major pathway. In particular, 3-hydroxylation is indu ced over 20-fold by DEX. Diazepam CLint was determined in vivo after a dministration of a bolus dose into the hepatic portal vein of UT, PB, and DEX rats; values of 127, 191, and 323 ml/min/SRW (where SRW is a s tandard rat weight of 250 g), respectively, were obtained. Using these scaling factors, the hepatocyte predictions of CLint were excellent ( 99, 144, and 297 ml/min/SRW for UT, PB, and DEX, respectively), wherea s only the DEX prediction (248 ml/min/SRW) was accurate for the micros omal system, with a substantial underprediction for UT and PB (46 and 68 ml/min/SRW, respectively), Evidence is presented for product inhibi tion, resulting from accumulation of primary metabolites within the mi crosomal preparation, as the mechanism responsible for this underpredi ction, These results illustrate that the scaling factor approach is ap plicable to induced livers in which both cytochrome P450 complement an d zonal distribution are altered, These data, together with our previo us studies, demonstrate that CLint in cells (2.4-297 ml/min/SRW), micr osomes (2.7-248 ml/min/SRW), and in vivo (1.5-323 ml/min/SRW) are rela ted in a linear fashion and hence inherently both in vitro systems are of equal value in predicting in vivo CLint.