[¹⁸F]ALX5406: A Brain-Penetrating Prodrug for GlyT1-Specific PET Imaging

Selective inhibition of glycine transporter 1 (GlyT1) has emerged as a potential approach to alleviate N-methyl-d-aspartate receptor (NMDAR) hypofunction in patients with schizophrenia and cognitive decline. ALX5407 is a potent and selective inhibitor of GlyT1 derived from the metabolic intermediate sarcosine (N-methylglycine) that showed antipsychotic potential in a number of animal models. Whereas clinical application of ALX5407 is limited by adverse effects on motor performance and respiratory function, a suitably radiolabeled drug could represent a promising PET tracer for the visualization of GlyT1 in the brain. Herein, [¹⁸F]ALX5407 and the corresponding methyl ester, [¹⁸F]ALX5406, were prepared by alcohol-enhanced copper mediated radiofluorination and studied in vitro in rat brain slices and in vivo in normal rats. [¹⁸F]ALX5407 demonstrated accumulation consistent with the distribution of GlyT1 in in vitro autoradiographic studies but no brain uptake in μPET experiments in naı̈ve rats. In contrast, the methyl ester [¹⁸F]ALX5406 rapidly entered the brain and was enzymatically transformed into [¹⁸F]ALX5407, resulting in a regional accumulation pattern consistent with GlyT1 specific binding. We conclude that [¹⁸F]ALX5406 is a promising and easily accessible PET probe for preclinical in vivo imaging of GlyT1 in the brain.