Systematic Evaluation of Tyrosine Kinase Inhibitors as OATP1B1 Substrates Using a Competitive Counterflow Screen

Area of Science:

• Pharmacology
• Hepatology
• Drug Metabolism

Background:

• The cellular uptake mechanism for most tyrosine kinase inhibitors (TKIs) into hepatocytes is not well understood, despite their primary elimination via CYP3A4 metabolism.
• Unpredictable pharmacokinetic profiles and hepatotoxicity associated with TKIs highlight the need to investigate their hepatic disposition pathways.

Purpose of the Study:

• To develop and validate a competitive counterflow (CCF) assay for identifying TKIs as substrates of the hepatic uptake transporter OATP1B1.
• To investigate the role of OATP1B1 in the hepatic uptake and potential toxicity of TKIs.

Main Methods:

• A competitive counterflow (CCF) assay was optimized and validated using OATP1B1-overexpressing HEK293 cells and radiolabeled estradiol-17β-glucuronide.
• Sixty-two approved TKIs were screened using the CCF assay to identify OATP1B1 substrates.
• Pazopanib was selected for further validation, including in vitro transport assays, molecular docking, and in vivo studies in mice with deficient orthologous transporters.

Main Results:

• The CCF assay identified 13 out of 62 evaluated TKIs as putative substrates of OATP1B1.
• Pazopanib transport by OATP1B1 was confirmed, with molecular docking suggesting overlapping binding sites with known substrates.
• In vivo studies showed decreased liver-to-plasma ratio and reduced pazopanib-induced hepatotoxicity in mice lacking orthologous transporters.

Conclusions:

• The CCF assay is a valuable tool for assessing TKI substrate affinity for OATP1B1.
• OATP1B1-mediated hepatic uptake is a significant mechanism for certain TKIs, influencing their disposition and contributing to hepatotoxicity.
• Understanding OATP1B1's role provides insights into TKI pharmacokinetics and potential adverse effects.