Abstract P102: Generation Of A Novel Peptoid-peptide Analog Of Alpha-calcitonin Gene Related Peptide (α-CGRP) For The Treatment And Prevention Of Cardiovascular Diseases

Abstract

Alpha-calcitonin gene related peptide (α-CGRP) is a 37-amino acid cardioprotective neuropeptide. Studies carried out in our laboratory and others establish α-CGRP as a potential therapeutic agent against a variety of cardiovascular diseases. However, the short half-life of α-CGRP limits its use in any long-term treatment regime. The goal of the present study is to develop an α-CGRP agonist analog with extended bioavailability using peptoid chemistry. A peptoid is a N -substituted glycine peptidomimetic molecule and is identical to α-amino acid except that the side chain in a peptoid is attached on the nitrogen rather than the α-carbon atom. Inclusion of a peptoid makes the native peptide protease-resistant and thus biostable in vivo . Using a solid-phase submonomer method, we synthesized a novel human α-CGRP analog containing two monomers of N -methoxyethylglycine (NMEG) peptoid at the N-terminus. Electrospray mass spectrometry (MALDI-TOF) analysis showed that the molecular mass of synthesized peptoid-peptide hybrid, NMEG-α-CGRP, was 4044 that is ~6.7% more than native peptide. An in vitro trypan blue cell exclusion assay demonstrated that incubation of NMEG-α-CGRP (5 μM) for 7 days did not affect the viability of rat H9C2 and mouse HL-1 cardiac cells. To evaluate the biological activity of NMEG-α-CGRP, a subcutaneous injection of human α-CGRP (10 μg/mice) and NMEG-α-CGRP (1, 3, 10, and 30 μg/mice) were given in 9-week-old C57BL6 mice (n=2 mice/dose), and blood pressure (BP) was measured using a tail-cuff method. A dose response curve showed that NMEG-α-CGRP decreased BP in mice in a time-dependent manner. Beginning with and injection of 3 μg of NMEG-α-CGRP, a dip in BP (85 ± 1 mmHg; in ±SD) was observed at 10 min after injection, and BP returned to baseline (125 mmHg) by 6 h, 18 h, and 24 h when injected with 3, 10, and 30 μg doses, respectively. Moreover, 10 μg of human-α-CGRP and NMEG-α-CGRP lowered BP from baseline for 2 h and 18 h, respectively, suggesting that NMEG addition increased stability, and thus bioavailability, of α-CGRP in vivo . In summary, our results show that a NMEG based α-CGRP modification is an effective approach to increase stability and, thus, bioavailability of α-CGRP in vivo making α-CGRP a viable therapeutic drug to treat cardiovascular diseases.