Time-dependent Inhibition (TDI)

Time-dependent inhibition often involves mechanism-based inhibition (MBI), where the inhibitor is converted into an active form during the enzyme's catalytic process, leading to irreversible inactivation of the enzyme. This process requires time, as the inhibitor must first bind to the enzyme and undergo metabolic transformation. The inhibitory effect increases over time, with significant inhibition typically observed after preincubation, and this type of inhibition may be irreversible.

TDI differs from other types of inhibition, such as direct inhibition and slow-binding inhibition, primarily in terms of the mechanism and temporal characteristics of the inhibition. Direct inhibition occurs when an inhibitor binds to the active site or another site on the enzyme, immediately reducing its activity. This type of inhibition is typically reversible, meaning the inhibitor can quickly affect the enzyme's activity without requiring time. Slow-binding inhibition refers to a process where the inhibitor binds to the enzyme slowly, potentially involving multiple steps or conformational changes. While it ultimately leads to decreased enzyme activity, the effect takes time to manifest.

Time-dependent Cytochrome P450 (CYP) Inhibition Assessment at Creative Biolabs

CYP TDI refers to a phenomenon where the effectiveness of CYP enzyme inhibitors increases during a period of in vitro incubation or in vivo dosing. This can occur due to the formation of more potent inhibitory metabolites or through mechanism-based inhibition (MBI), where the drug's metabolic products irreversibly inactivate the CYP enzymes.

Fig. 1 Time-dependent inhibitor inactivates CYP in the organs.^1,2

In TDI, the relationship between CYP (cytochrome P450) and NADPH (reduced nicotinamide adenine dinucleotide phosphate) is important. During TDI, the inhibitor first binds to the CYP enzyme and is metabolically converted into an active form in the presence of NADPH. These active forms can form covalent or non-covalent bonds with the CYP enzyme, leading to irreversible inhibition of enzyme activity. The occurrence of TDI is usually NADPH-dependent, since without NADPH, the inhibitor may not be metabolized into the active form, making it ineffective at inhibiting CYP enzyme activity.

Creative Biolabs evaluates whether the test compound is an irreversible, time-dependent inhibitor of the CYP enzyme. The assay is measured by CYPs' known substrate under conditions with and without the NADPH in human liver microsomes by HPLC-MS/MS. The CYPs we assessed contain CYP1A, CYP2C9, CYP3A, etc.

Creative Biolabs is committed to offering reliable assistance for your research. If you would like to know more about our services, please don't hesitate to contact us. At the same time, the following services in ADME we offer may also be worth your consideration.

• Drug-Drug Interactions
• Enzyme Reaction Phenotyping
• Transporter Interaction Assessment

References

1. Filppula, A. M., et al. "Improved predictions of time-dependent drug-drug interactions by determination of cytosolic drug concentrations." Scientific reports 9.1 (2019): 5850.
2. Under Open Access license CC BY 4.0, without modification.

For Research Use Only | Not For Clinical Use