The Role of Extrahepatic Metabolism in the Pharmacokinetics of the Targeted Covalent Inhibitors Afatinib, Ibrutinib, and Neratinib

Abstract

Despite the fact that much progress has been made recently in the development of targeted covalent inhibitors (TCIs), their pharmacokinetics (PK) have not been well characterized in the light of extrahepatic clearance (CL extH ) by glutathione (GSH)/glutathione S-transferase (GST)-dependent conjugation attributable to the unique electrophilic structure (e.g., acrylamide moiety) of TCI compounds. In the present study, CL extH values were examined in rat, dog, and monkey to predict the contribution of CL extH to the PK of the TCIs afatinib, ibrutinib, and neratinib in humans. Afatinib and neratinib both underwent extensive conjugation with GSH in buffer and cytosol fractions of liver and kidney, whereas ibrutinib showed much lower reactivity/susceptibility to GSH/GST-dependent conjugation. The CL extH in each species was calculated from the difference between observed total body clearance and predicted hepatic clearance (CL H ) in cryopreserved hepatocytes suspended in 100% serum of the corresponding species. The power-based simple allometry relating the CL extH for the unbound compound to animal body weight was applicable across species for afatinib and neratinib (R 2 ‡ 0.9) but not for ibrutinib (R 2 = 0.04). The predicted AUC after oral administration of afatinib and neratinib agreed reasonably closely with reported values in phase I dose-escalation studies. Comparisons of CL extH and CL H predicted that CL extH largely determined the PK of afatinib (>90% as a proportion of total body clearance) and neratinib (∼34%) in humans. The present method can serve as one of the tools for the optimization of PK in humans at the discovery stage for the development of TCI candidates.

Metabolic Disposition of Osimertinib in Rats, Dogs, and Humans: Insights into a Drug Designed to Bind Covalently to a Cysteine Residue of Epidermal Growth Factor Receptor

“…Shibata and Chiba have investigated the correlation between clearance predicted from hepatocyte incubations and total body clearance for three covalent binding TKIs (afatinib, ibrutinib, and neratinib). The authors noted a disconnect between in vivo and in vitro clearance attributed to extrahepatic (covalent) conjugation to glutathione not captured in the hepatocyte system (Shibata and Chiba, 2015). For osimertinib, the in vitro studies indicated that direct conjugation with glutathione would be a minor metabolic pathway in humans; however, it is clear from the human ADME study that a high proportion of [ 14 C]-osimertinib-related material is excreted bound to proteinaceous material contributing a substantial fraction of overall clearance.…”

Raman Microspectroscopic Evidence for the Metabolism of a Tyrosine Kinase Inhibitor, Neratinib, in Cancer Cells

“…Neratinib also retains electrophilic reactivity to the Cys residue of glutathione (GSH) by forming a GSH adduct 28. We investigated the chemical reactivity of neratinib with GSH (Figure S31).…”

A tag-free collisionally induced fragmentation approach to detect drug-adducted proteins by mass spectrometry

“…Even though all of these acrylamides were electrophilic and prone to Michael addition with small molecule thiols such as glutathione and cysteine (data not shown), HSA posed a different challenge as it has several lipophilic binding pockets. Many kinase inhibitors, including the ones investigated, have been shown to have high plasma protein binding affinities and we postulated that the level of conjugation of HSA by afatinib and canertinib was due to non‐covalent HSA binding, thereby lowering the free concentration of the small molecule drug available to conjugate to the surface exposed thiol . Due to the low conjugation profiles of PD‐168393, afatinib and canertinib, MS e CID fragmentation did not produce detectable unique fragment ions and further investigation with these compounds was halted.…”