Z. Liu¹, L. Zhang¹, Y. Li¹, S. Joel¹, S. Deo¹, S. Daunert¹, O. C. Velazquez^{1 1}University Of Miami,Miami, FL, USA

Introduction: Intercellular adhesion molecule-1 (ICAM-1) is induced on inflamed endothelial cells (iEC) under several pathological conditions, such as atherosclerosis and inflammation. Integrin LFA-1 recognizes ICAM-1 as its ligand, and the I-domain of LFA-1 (idLFA-1) is responsible for bind to ICAM-1. idLFA-1/ICAM-1 pair is thus an attractive target for delivery of therapeutic cells to tissues where iEC are presented. We are aimed to develop a safe and targeted cell delivery method for therapeutic administration of stem/progenitor cells, which carry repair function, to atherosclerotic lesion to stave off the development of atherosclerosis. We have recently developed a novel cell delivery platform by coating the surface of the cells to be delivered with nanocarriers composed of nanoparticle–adhesion molecule complex. These nanocarriers guide the coated cells to their destination via recognition and association with their counterpart adhesion molecules, which are highly or selectively expressed on the diseased tissue, and execute their therapeutic roles. Here, we tested effectiveness of idLFA-1 complexed nanocarriers in mediating interaction of nanocarrier-coated endothelial progenitor cells (EPC) to iEC in vitro.

Methods: Recombinant human idLFA-1 protein was denatured, refolded, and purified by gel filtration. Binding activity of idLFA-1 to ICAM-1 was validated by testing association of FITC-conjugated idLFA-1 with human aorta endothelial cells (HAEC) pre-transduced with Notch1^IC/lentivirus to induce high ICAM-1 (ICAM-1^hi) expression. Nanocarriers were created by conjugation of idLFA-1 with dendrimers. Human EPC were labeled by transduction with DsRed/Lentivirus. DsRed⁺EPC were then coated with idLFA-1-dendrimers and BSA-dendrimers (control), respectively. The effectiveness of idLFA-1 complexed nanocarriers in mediating interaction of DsRed⁺EPC to ICAM-1^hiHAEC (Notch1^IC/lentivirus transduced) vs ICAM-1^loHAEC (LacZ/lentivirus transduced) was tested using in vitro cell-cell binding assay. Interaction of DsRed⁺EPC-HAEC was quantified by measuring fluorescence intensity.

Results: Purified human idLFA-1 protein was characterized based on its size. HAEC transduced with Notch1^IC/lentivirus are ‘inflamed’ since expression of ICAM-1 along with a panel of inflammatory citokines, including IL-1α, IL-6, IL-8 and RANTES, are elevated significantly. FITC-conjugated idLFA-1 preferentially bound to ‘inflamed’ HAEC compared to control HAEC (p<0.01). idLFA-1-complexed nanacarriers mediated increased interaction of DsRed⁺EPC to ICAM-1^hi HAEC compared to ICAM-1^lo HAEC (>1 fold, p<0.01).

Conclusion: We demonstrated effectiveness of idLFA-1 complexed nanocarriers in mediating interaction of nanocarrier-coated EPC to iEC in vitro. This work paves way to test feasibility of idLFA-1 complexed nanocarriers for targeted delivery of therapeutic cells to atherosclerotic lesion or other inflammation tissues in in vivo model.