2’,3’-O-Substituted ATP derivatives as potent antagonists of purinergic P2X3 receptors and potential analgesic agents

Compounds able to block membrane currents induced by the stimulation of the purinergic P2X3 receptor localized on nociceptive sensory fibres may be useful as novel and promising tools to treat chronic pain conditions including neuropathic pain, migraine, and inflammatory pain. Inhibitors of these receptors have been reported in literature, among which the ATP analogue 2’,3’-O-(2,4,6-trinitrophenyl)-ATP (TNP-ATP) represent a reference tool with nanomolar potency at the P2X3. With the aid of molecular modelling studies performed at P2X4-based homology models of the human, rat, and mouse P2X3 receptor [1-3], we analysed the binding mode of the TNP-ATP at this protein and designed novel adenosine nucleotide analogues (“PF compounds”, Figure 1) bearing alkyl or arylalkyl substituents of various hindrances and replacing the trinitrophenyl moiety of TNP-ATP. These compounds were synthesised and then functionally evaluated on native P2X3 receptors from mouse trigeminal ganglion (TG) sensory neurons using patch clamp recordings under voltage clamp configuration. The results show that these molecules are not endowed with agonist activity at the P2X3 while they are able to inhibit with nanomolar potency currents evoked by α,β-methyleneATP-induced stimulation of this protein. The P2X3 inhibition is reversible and its potency appears influenced by the volume of the 2’,3’-O-substituent introduced in the ATP moiety. Finally, these molecules do not show any apparent effect on trigeminal GABAA and 5-HT3 receptors that are expressed by the same neurons [4]. The activity of these compounds was also analysed at the recently published crystal structures of the P2X3 receptor [5].