Similarities and differences in the practice and use of time-dependent inhibition studies in drug discovery and development

Common Practices (≥75% concordance in responses)

Divergent Practices (<75% concordance in responses)
Strategic aspects
    All assess TDI during drug discovery/development continuum Timing of definitive assays for clinical DDI predictions ranges from lead optimization through phase 1
    TDI data are used for predicting DDI
No common cut-off values for TDI data for further progression of NMEs
Use of various study designs for TDI assessment in drug discovery (e.g., IC50 shift vs. % activity loss at single NME concentration, etc.)
No common consideration of structural alerts
Technical aspects
    Pooled human liver microsomes (100%) Dilution used during IC50 shift determinations range from no dilution to greater than 10-fold
    Major P450 enzymes tested (Fig. 4)
    LC/MS/MS for measurement of probe substrates (100%) Number of NME concentrations used to determine inactivation parameters (6 or greater)
    Solvent control at each time point (- test article + NADPH) are used (100%)
Number of time points used (4 to >6)
    Determine the log-linear phase of enzyme inactivation (100%) Data analysis
    Conduct control incubations without NADPH     Log-linear regression (kobs) followed by nonlinear fitting to determine KI and kinact parameters
    Replicate determinations of KI and kinact are conducted
    Positive controls are included     Reciprocal plot (e.g., Kitz-Wilson) to estimate KI amd kinact
    Test article depletion not measured     Global nonlinear regression
Use of TDI data
    Current models cannot accurately predict DDI due to TDI Various models (static vs. dynamic, inclusion of gut first-pass vs. no gut first pass, etc.) are used for predicting DDI risk based on KI and kinact values
    Existing models can categorize compounds as weak, moderate, or potent clinical DDI risks
    DDI predictions to decide whether to conduct a DDI study and inform its design Various values used as surrogates for [I]in vivo (e.g. Cmax, free vs. total, etc.)

Microsomal and plasma protein binding corrections used by some
  • LC/MS/MS, liquid chromatography/tandem mass spectrometry.