Abstract 423: Intramyocardial Delivery Of Protease-resistant Stromal Cell Derived Factor-1 By Self-assembling Peptide Nanofibers
A useful approach to cardiac regeneration could be intramyocardial delivery of the stem cell chemoattractant Stromal cell derived factor-1 (SDF-1). However, SDF-1 rapidly diffuses away from the site of injection. Therefore, we designed fusion proteins of SDF-1 and the sequence of self-assembling peptides, allowing prolonged delivery in the myocardium. Self-assembling peptides form a stable hydrogel of nanofibers when injected. In addition to rapid diffusion, SDF-1 delivery is also challenged by inactivation by matrix metalloproteinase-2 (MMP-2) expressed after myocardial infarction (MI).
Methods. We designed a mutant SDF-1 resistant to MMP-2. SDF-1 proteins were expressed in E coli and purified by a 4 step procedure consisting of affinity and cation exchange chromatography, oxidative refolding, and reversed phase-chromatography. The mutant SDF-1, called SDF-1(S4V), was resistant to MMP-2 cleavage but bioactive. SDF-1-RAD fusion proteins (RAD representing the sequence of self-assembling peptides) were expressed and incorporation into self-assembling peptides was measured by radioactive labeling of SDF-1. SDF-1, SDF-1(S4V), SDF-1-RAD and SDF-1(S4V)-RAD (30nM) were injected with self-assembling peptides in infarcted myocardium in a blinded and randomized study of 176 rats after ligation of the LAD. Left ventricular function at 28 d was measured by catheterization.
Results. SDF-1-RAD fusion proteins stably incorporated into self-assembling peptides. Intramyocardial injection of the MMP-2 resistant SDF-1(S4V)-RAD fusion protein with nanofibers increased ejection fraction from 34.0 ± 2.5% (n = 21) in the MI only group to 49.5 ± 2.9% (n = 18, p = 0.004). SDF-1, SDF-1(S4V), and SDF-1-RAD did not improve cardiac function. Capillary density increased from 169 ± 42 /mm2 in MI only group to 283 ± 27 /mm2 in SDF-1(S4V)-RAD +self-assembling peptide group (p =0.035). Finally, c-Kit + /Flk-1 +cells increased from 46 ± 7 cells/section in MI only to 132 ± 35 cells/section in MI + NF/SDF-1(S4V)-RAD (p =0.002).
Conclusions. By designing SDF-1 to be resistant to MMP-2 cleavage and enabling prolonged delivery by self-assembling peptides, cardiac function after MI was improved and capillary density and endothelial progenitor cell infiltration was increased.