Liquid Chromatography/tandem Mass Spectrometry-based Target Engagement of Lysine-specific Demethylase 1 Inhibitor TAK-418 Through Detecting Covalent Adducts in the Brain.

Translating pharmacokinetics (PK) and pharmacodynamics (PD) from preclinical to clinical is important in successful development of drugs for central nervous system (CNS) diseases. TAK-418 is a clinical candidate of lysine-specific demethylase 1 (LSD1) inhibitor with a unique mechanism of action; it covalently binds to flavin adenine dinucleotide (FAD) to form a compact adduct formylated FAD (F-FAD), thereby minimizing hematological side effect risks. In this study, we established a liquid chromatography-tandem mass spectrometry (LC/MS/MS)-based approach for evaluating F-FAD as a target engagement marker for TAK-418 to advance PK/PD translation and clinical dose setting. In rats, LC/MS/MS enabled detection of TAK-418-induced F-FAD formation in the brain with chromatographic separation from native FAD. F-FAD levels were positively associated with LSD1 protein abundance across brain regions and increased with increasing TAK-418 dose. LSD1 occupancy calculated from F-FAD concentrations was correlated with the inhibition level of LSD1 enzyme activity in rat hippocampus. This mechanism-based approach provides a link between covalent adduct formation, target engagement, and enzyme inhibition in the brain. In addition, similar point estimates of the half-maximal effective concentration (EC50)​ for F-FAD formation in rat and human hippocampal homogenates support the potential utility of F-FAD measurements for translational assessment of LSD1 target engagement. These findings provide useful information for evaluating the translational relationship between TAK-418 exposure and LSD1 target engagement and may support future PK/PD modeling and clinical dose selection for TAK-418. Moreover, our methodology for measuring covalent adducts would be applicable for multiple CNS drug candidates with covalent binding property.