Discovery of novel N-(alkylpiperidinyl) substituted benzamides targeting the glycine transporter 2 with anti-nociceptive effects in neuropathic pain

Neuropathic pain is a global health problem, affecting approximately 7%-10% of the population worldwide. In recent years, targeting glycine transporter 2 (GlyT2) has emerged as a promising strategy for the treatment of neuropathic pain, as it enhances glycinergic neurotransmission and strengthens inhibitory control of nociceptive signal transmission. A series of N-(alkylpiperidinyl) substituted benzamides were designed, synthesized, and evaluated as potential GlyT2 inhibitors, and their binding affinities were assessed in vitro by surface plasmon resonance assay. Compound 14 was found to have potent affinity, with a K_D value of 23.7 nM. In mice, 14 exhibited robust antinociceptive effects in the formalin test (ED₅₀ = 30.5 mg/kg), abdominal contraction (ED₅₀ = 26.0 mg/kg) test, and paclitaxel-induced neuropathic pain (ED₅₀ = 12.17 mg/kg) models, with overall activity superior to the reference drug gabapentin. The antinociceptive effect of 14 was attributable to the inhibition of glycine transporters, as confirmed by combined administration of strychnine (a glycine receptor antagonist) in mice. In addition, 14 functionally inhibited GlyT2-mediated glycine uptake with an IC₅₀ value of 19.4 nM. Furthermore, 14 exhibited an improved safety profile compared to gabapentin, notably with reduced sedative effects and less impairment of motor coordination. Compound 14 also exhibited favorable pharmacokinetic properties in rats. Taken together, these findings highlight 14 as a promising lead candidate for developing safer and more effective analgesics against neuropathic pain.