Metabolism-based DDIs

Since enzyme inhibition involves reversible mechanisms, CLi , (ij may vary with regard to the type and concentration of inhibitor. The concentrations of an inhibitor (or drug) that are relevant to clinical application can be approached for the prediction in the in vivo situation. In practice, a ratio in AUC, hepatic clearance (CLhept), plasma concentration at steady state (Css), or intrinsic clearance (CLjnt) caused by metabolism-based DDIs is commonly used to assess the degree of metabolism inhibition in vivo (Eq. 16.7). If a drug is eliminated due to both metabolism and renal excretion, the fraction of the drug metabolized by the inhibited enzyme (fj ) should be introduced to the prediction. With inclusion of fj, the ratio change in AUC in the presence and absence of an inhibitor can be expressed for competitive and noncompetitive (Eq. 16.8). 

The CYP inhibition assay utilizes the 96-well plate format with a robotic system, where both incubation and analysis are performed in the same plates. The setup of the sample plates is shown in Figure 4.1. For each compound, both direct inhibition and metabolism/mechanism-based inhibition, which is caused by a metabolite of the NCE that is either a more potent direct reversible inhibitor (metabolism-based) or a time-dependent irreversible inhibitor (mechanism-based), are evaluated. Both direct and mechanism-based inhibitors could result in inhibitory DDIs [51,52],... 

P450 mechanism-based inactivations formed via either pathway illustrated in Fig. 5.7 result in clinically relevant DDIs. MI metabolic intermediate, DDI drug-drug interactions... 

In vitro techniques for studying DDI potential are based on the metabolism of known marker substrates. Two assay types are typically used to study DDIs the turnover of drug-like probes monitored by LC-MS/MS methods or the use of spectrophotometer (plate reader) based methods. As each technique has unique advantages and shortcomings, assay use has not been standardized across the industry. Although techniques based on the turnover of radiolabeled substrates have also been developed [94—97], these methods are used infrequently and will not be discussed further. 

A number of reports also describe the prediction of mechanism-based inhibition (MBI) [17,18]. In this type of model, MBI is determined in part by spectral shift and inactivation kinetics. Jones et al. applied computational pharmacophores, recursive partitioning and logistic regression in attempts to predict metabolic intermediate complex (MIC) formation from structural inputs [17]. The development of models that accurately predict MIC formation will provide another tool to help reduce the overall risk of DDI [19]. 

Time-dependent inhibition defined mainly by mechanism-based inhibition (MBI), which includes CYP suicide inactivation (irreversible inhibition, the more widely studied process) and metabolite-intermediate (MI) complex formation (quasi-irreversible inhibition), is responsible for most clinically significant DDIs (Silverman, 1995 Waley, 1980 Zhou et al., 2005). Suicide inactivation involves the formation of a reactive intermediate that irreversibly inactivates the CYP in the process of catalytic turnover. Quasi-irreversible inhibition occurs when the CYP produces a metabolite (e.g., nitroso intermediate) with the capacity to bind tightly to the CYP heme. TDI (time-dependent inhibition) can be characterized (1) to be dose dependent, (2) to be preincubation time dependent, (3) to have bioactivation of the inhibitor that is required for inactivation of the target enzyme, (4) to have de novo protein synthesis that is required to recover metabolic capacity, and (5) to have potentially slow onset of the effects but be more profound than reversible inhibition. If present, then TDI is the major component of overall enzyme inhibition and frequently leads to clinically relevant DDIs. Table 4.5 contains a list of inhibitors of TDI observed in vitro and in vivo. 

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