Cyp3a4-Mediated in vitro Metabolism and in vivo Disposition of Lorlatinib in Rats

Purpose Lorlatinib, a third-generation inhibitor of anaplastic lymphoma kinase (ALK) and c-ros oncogene 1 (ROS1), undergoes extensive metabolism, with its disposition influenced by drug–drug interactions (DDIs). This study aimed to characterize its in vitro metabolism, in vivo pharmacokinetics (PK), tissue distribution, and the effects of cytochrome P450 3A4 (CYP3A4) modulation in rats.Methods Lorlatinib metabolism was investigated in human liver microsomes (HLM) and rat liver micro- somes (RLM) using ultra-high-performance liquid chromatography–tandem mass spectrometry (UHPLC–MS/MS) with selective CYP inhibitors. Pharmacokinetic (PK) studies were performed in Sprague–Dawley (SD) rats following oral and intravenous dosing, including coadministration with CYP3A4 inhibitors (voriconazole, itraconazole) and inducers (rifampicin, carbamazepine). Tissue distribution was assessed across major organs.Results In vitro assays confirmed CYP3A4 as the primary enzyme mediating lorlatinib metabolism, with distinct differences in kinetic parameters between HLM and RLM. In vivo, lorlatinib displayed nonlinear PK, low oral bioavailability (8.6%), and extensive first-pass metabolism. Voriconazole increased exposure [area under the curve from 0–24 h (AUC(0−24h))] by 120%, whereas rifampicin reduced it by 77%, demonstrating strong CYP3A4-dependent interactions. Lorlatinib distributed rapidly to highly perfused organs and achieved efficient brain penetration (brain-to-plasma ratio = 0.82).Conclusion Lorlatinib undergoes extensive CYP3A4-mediated metabolism, exhibits nonlinear PK, and shows poor oral bioavailability in rats. Significant interspecies differences between HLM and RLM emphasize the need for caution when translating preclinical findings. Coadministration with CYP3A4 modulators markedly altered systemic exposure, while effective brain penetration supports its therapeutic role in central nervous system (CNS) metastases.