Abstract
Lipidised analgesic peptide prodrugs self-assemble into peptide nanofibers; with the nanofiber morphology protecting the peptide from plasma degradation and improving therapeutic efficacy. Extending this learning, we hypothesised that a self-assembling lipidized peptide arginine vasopressin (AVP) receptor agonist, that had not been designed as a prodrug, could prove pharmacologically active and control urine production. The only approved AVP receptor agonist, desmopressin is indicated for the treatment of central diabetes insipidus (DI), bedwetting, haemophilia A and von Willebrand disease. Desmopressin is well tolerated by most patients, however adverse effects, such as hyponatraemia and water intoxication necessitate a strict fluid intake, thus motivating the search for alternative DI treatments. Selective V2 receptor agonism is required for anti-DI activity and we hypothesised that our new lipidized peptide (METx) would lead to selective AVP receptor agonism. METx was synthesised and characterised and then tested for activity against the V2, V1a and OT uterine receptors and not tested against the V1b receptor as METx was not expected to cross the blood brain barrier. METx was also tested in vivo in a healthy rat model. METx forms nanofibers and is a partial V2 receptor agonist (determined by measuring MDCK cell line cAMP accumulation), producing 57% of AVP’s maximal activity (EC50 = 2.7 nM) and is not a V1a agonist up to a concentration of 1 μM (determined by measuring A7r5 cell line D-myo-inositol-1-phosphate accumulation). METx is a weak OT receptor antagonist, reducing the frequency of OT induced contractions (EC50 = 350 nM) and increasing the OT EC50 from 0.081 nM to 21 nM at a concentration of 600 nM. METx (41 nM) had no effect on spontaneous uterine contractions and METx (100 nM) had no effect on OT induced uterine contractions. Simulated binding studies show that binding avidity to the receptors follows the trend: V2 > OT > V1a. On intravenous injection, a nanoparticle formulation of METx reduced urine production in a healthy rat model in a dose responsive manner, with 40 mg kg−1 METx resulting in no urine production over 4 hours. The lipidized self-assembling peptide – METx - is a selective competitive V2 receptor agonist and an anti-diuretic.
Highlights
Lipidized peptides self-assemble into nanofibers and such nanofibers have been used for tissue engineering and drug delivery[1,2,3]
arginine vasopressin (AVP) is active at three known receptors: V1a receptors in the smooth muscle and liver resulting in pressor activity, V1b receptors largely found in the pituitary gland resulting in adrenocorticotrophic hormone releasing activity and V2 receptors found in the kidney collecting ducts that result in anti-diuretic activity[8]
We studied the self-assembly of METx in aqueous media in order to explore if nanofibers would be formed from the lipidized peptide, as lipidized peptides are known to self-assemble into nanofibers[2,3,4]
Summary
Lipidized peptides self-assemble into nanofibers and such nanofibers have been used for tissue engineering and drug delivery[1,2,3]. In the case of drug delivery, the lipidized peptides are prodrugs as the active peptide epitopes are bound to a fatty acid chain via a cleavable ester linker[2,3]. The vasopressor effect of arginine vasopressin (AVP) was first reported in 1895 after discovering that the extracts of cow pituitary had a vasopressor effect[5]. This was followed by the work of Du Vigneaud in the 1950s, who discovered the structure of AVP and oxytocin (OT) and synthesised these compounds[6,7]. Structure activity studies have revealed the 20 member cyclic ring to be essential for both pressor and oxytocic activity[15] and that the carboxy terminus was essential for activity at the AVP receptors[16]
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