Potent, Selective, and Brain Penetrant Ether-Linked 2-Aminopyridine Inhibitors of Human Neuronal Nitric Oxide Synthase with Excellent Oral Bioavailability

Neuronal nitric oxide synthase (nNOS) is a therapeutic target for the treatment of various neurological disorders and for melanoma. As part of our ongoing efforts to develop potent and selective nNOS inhibitors, we modified our previously reported compound <b>3</b> to <b>4</b> by introducing an ether linker, leading to a new series of ether-linked 2-aminopyridine-based compounds that exhibit high potency, isoform selectivity, and membrane permeability. Among them, lead compound <b>4</b> inhibits human nNOS with a <i>K</i>_i of 25 nM and exhibits 2300-fold selectivity over human endothelial NOS (eNOS) while also displaying high effective permeability in the parallel artificial membrane permeability assay for the blood-brain barrier (PAMPA-BBB) assay (<i>P</i>_e = 16.67 × 10^-6 cm/s), indicating favorable blood-brain barrier penetration. Pharmacokinetic evaluation confirmed the brain penetrance of <b>4</b> and demonstrated a high oral bioavailability (77%). Moreover, the X-ray crystal structures of representative compounds bound to three NOS isoforms (hnNOS, rnNOS, and heNOS) revealed key binding interactions that contribute to both potency and selectivity.