Neurotensin Analogues Containing Cyclic Surrogates of Tyrosine at Position 11 Improve NTS2 Selectivity Leading to Analgesia without Hypotension and Hypothermia

Neurotensin (NT) exerts its analgesic effects through activation of the G protein-coupled receptors NTS1 and NTS2. This opioid-independent antinociception represents a potential alternative for pain management. While activation of NTS1 also induces a drop in blood pressure and body temperature, NTS2 appears to be an analgesic target free of these adverse effects. Here, we report modifications of NT at Tyr¹¹ to increase selectivity toward NTS2, complemented by modifications at the N-terminus to impair proteolytic degradation of the biologically active NT(8-13) sequence. Replacement of Tyr¹¹ by either 6-OH-Tic or 7-OH-Tic resulted in a significant loss of binding affinity to NTS1 and subsequent NTS2 selectivity. Incorporation of the unnatural amino acid β³hLys at position 8 increased the half-life to over 24 h in plasma. Simultaneous integration of both β³hLys⁸ and 6-OH-Tic¹¹ into NT(8-13) produced a potent and NTS2-selective analogue with strong analgesic action after intrathecal delivery in the rat formalin-induced pain model with an ED₅₀ of 1.4 nmol. Additionally, intravenous administration of this NT analogue did not produce persistent hypotension or hypothermia. These results demonstrate that NT analogues harboring unnatural Amino acids at positions 8 and 11 can enhance crucial pharmacokinetic and pharmacodynamic features for NT(8-13) analogues, i.e., proteolytic stability, NTS2 selectivity, and improved analgesic/adverse effect ratio.