Differences in Sustained Cellular Effects of MET inhibitors Are Driven by Prolonged Target Engagement and Lysosomal Retention

Mol Pharmacol. 2023 Feb;103(2):77-88. doi: 10.1124/molpharm.122.000590. Epub 2022 Nov 18.

Abstract

Intracellular distribution of drug compounds is dependent on physicochemical characteristics and may have a significant bearing on the extent of target occupancy and, ultimately, drug efficacy. We assessed differences in the physicochemical profiles of MET inhibitors capmatinib, crizotinib, savolitinib, and tepotinib and their effects on cell viability and MET phosphorylation under steady-state and washout conditions (to mimic an open organic system) in a human lung cancer cell line. To examine the differences of the underlying molecular mechanisms at the receptor level, we investigated the residence time at the kinase domain and the cellular target engagement. We found that the ranking of the drugs for cell viability was different under steady-state and washout conditions and that under washout conditions, tepotinib displayed the most potent inhibition of phosphorylated MET. Postwashout effects were correlated with the partitioning of the drug into acidic subcellular compartments such as lysosomes, and the tested MET inhibitors were grouped according to their ability to access lysosomes (crizotinib and tepotinib) or not (capmatinib and savolitinib). Reversible lysosomal retention may represent a valuable intracellular storage mechanism for MET inhibitors, enabling prolonged receptor occupancy in dynamic, open-physiologic systems and may act as a local drug reservoir. The use of washout conditions to simulate open systems and investigate intracellular drug distribution is a useful characterization step that deserves further investigation. SIGNIFICANCE STATEMENT: Generally, determination of potency and receptor occupancy is performed under steady-state conditions. In vivo conditions are more complex due to concentration differences between compartments and equilibrium processes. Experiments under steady state cannot explore effects such as sustained target inhibition. This study has shown that differences between MET inhibitors are observable by applying washout conditions to in vitro assays. This important finding applies to most compound classes and may inspire readers to rethink their assay designs in the future.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Carcinoma, Non-Small-Cell Lung* / metabolism
  • Crizotinib / pharmacology
  • Humans
  • Lung Neoplasms* / metabolism
  • Lysosomes / metabolism
  • Protein Kinase Inhibitors / pharmacology
  • Proto-Oncogene Proteins c-met / metabolism

Substances

  • capmatinib
  • Crizotinib
  • Proto-Oncogene Proteins c-met
  • Protein Kinase Inhibitors