Controlled release of peptide from biodegradable platforms for cardiovascular applications
Ng, Xu Wen
Date of Issue2013
School of Materials Science and Engineering
Cenderitide, also known as CD-NP, is a designer peptide developed by Mayo Clinic for treating cardiovascular pathologies. CD-NP is synthesized by combining native mammalian c-type natriuretic peptide (CNP) and a C-terminus isolated from the dendroapis natriuretic peptide (DNP) of the venom of green mamba. Currently, the clinical means of administering CD-NP is via intravenous or subcutaneous injection. However, such administration is low in patient compliance and results in high wastage. This thesis presents the development of a controlled release system for the administration of CD-NP over a sustained period of time. Due to difficulty in solubilizing hydrophilic CD-NP, two co-solvent systems were explored for the encapsulation of CD-NP in polymeric matrix. In the first co-solvent system consisting of ethanol and dichloromethane (DCM), sustained release of CD-NP from PCL-based formulation was achieved for 30 days. The release rates of CD-NP were further manipulated through the addition of polyethylene glycol (PEG) and copolymers of poly (e-caprolactone) with PEG. The addition of these excipients resulted in increase in the initial CD-NP release. A detailed investigation on the surface morphology of films, degradation characteristics and kinetics of release was carried out and the coleaching of CD-NP with excipient was singled out as the key factor for the increase in the initial release. In the second co-solvent system consisting of water and DCM, two emulsification conditions were studied due to the immiscibility of the two solvents. The results demonstrate that different emulsification conditions dramatically affected the initial release of peptide, where a difference of 5-fold was observed. This observation was attributed to difference in peptide aggregation and degree of porous formation under the two emulsification conditions. Subsequently, the combined effects of emulsification condition and excipient addition were investigated. The addition of PEG and copolymer reduced the effects of emulsification on peptide release due to colocalization of excipients with peptide, which redistributed CD-NP throughout the matrix. In literature, CD-NP had been reported to possess anti-fibrotic and anti-proliferative properties. However, these studies lack systematic understanding on the extent of CDNP's actions on cardiac cells in vitro. In this work, the anti-fibrotic capability of CDNP was explored through the study of CD-NP's interactions with the human cardiac fibroblast (HCF) cells. First, we studied the effects of CD-NP on HCF and elucidated the extent of CD-NP inhibition on HCF. Single dose CD-NP exhibited inhibition of hyperplasia HCF for up to 24 hours but was ineffective on hypertrophic HCF. Multiple doses CD-NP sustained the inhibition of hyperplasia HCF for 5 days for the highest concentration. Moreover, multiple doses of CD-NP at highest concentration exhibit inhibition of hypertrophic HCF by the third dose. We also studied the anti-proliferative capability of CD-NP by incubating CD-NP with the human coronary smooth muscle cells (HCaSMCs). We found that the inhibitory effects of CD-NP on HCaSMCs were limited compared to HFC. After elucidating the effects of CD-NP on the two cardiac cell types, we moved to understand the effects of CD-NP released from the controlled release systems on HCF and HCaSMCs. The bioactivity of released peptide was verified from the elevateproduction of cyclic guanosine monophospate (cGMP) in both HCF and HCaSMCs. To understand the effects of continuous release CD-NP, selected films releasing CDNP were incubated with the HCF. The sustained delivery ofCD-NP from formulations manufactured from Water/DCM co-solvent system showed superior suppressive actions on HCF compared to daily infusion of CD-NP. Furthermore, we investigated the effects of excipient addition on encapsulated CD-NP. The addition of excipients did not result in loss of bioactivity in the released CD-NP, which signified that the addition of excipients could be used as modifier or stabilizer in peptide release profiles. The results suggest that development of CD-NP releasing films or devices could be used to inhibit fibrosis and reduce cardiac remodeling in future studies.