Abstract
Therapeutic hypothermia represents a brain-protective strategy for multiple emergencysituations, such as stroke or traumatic injury. Neurotensin (NT), which exerts itseffects through activation of two G protein-coupled receptors, namely NTS1 andNTS2, induces a strong and long-lasting decrease in core body temperature afterits central administration. Growing evidence demonstrates that NTS1 is the receptorsubtype mediating the hypothermic action of NT. As such, potent NTS1 agonistsdesigned on the basis of the minimal C-terminal NT(8-13) bioactive fragment have beenshown to produce mild hypothermia and exert neuroprotective effects under variousclinically relevant conditions. The high susceptibility of NT(8-13) to protease degradation(half-life<2min) represents, however, a serious limitation for its usein pharmacologicaltherapy. In light of this, we report here a structure-activity relationship study in whichpairs of NT(8-13) analogs have been developed, based on the incorporation of areduced Lys8-Lys9bond. To further stabilize the peptide bonds, a panel of backbonemodifications was also inserted along the peptide sequence,including Sip10, D-Trp11,Dmt11, Tle12, and TMSAla13. Our results revealed that the combination of appropriatechemical modifications leads to compounds exhibiting improved resistance to proteolyticcleavages (>24h;16). Among them, the NT(8-13) analogs harboring the reducedamine bond combined with the unnatural amino acids TMSAla13(4) and Sip10(6) orthe di-substitution Lys11- TMSAla13(12), D-Trp11-TMSAla13(14), and Dmt11-Tle12(16) produced sustained hypothermic effects (−3◦C for at least 1h). Importantly, weobserved that hypothermia was mainly driven by the increased stability of the NT(8-13)derivatives, instead of the high binding-affinity at NTS1. Altogether, these results revealthe importance of the reduced amine bond in optimizing the metabolic properties of theNT(8-13) peptide and support the development of stable NTS1agonists as first drugcandidate in neuroprotective hypothermia