Poster Z3 - Development of Targeted Nanoparticles to Enhance Drug Delivery for Peripheral Arterial Disease Treatment

Introduction: Peripheral arterial disease (PAD) is a condition marked by the narrowing of arteries from a buildup of plaque in the lower extremities. About 6.5 million Americans, primarily the elderly, are affected by PAD with high mortality and mobility rates. Current treatments for PAD such as bypasses, stents, gene therapy, stem cell therapy, and others consist of several limitations including high rates of restenosis, and limited drug availability over time, thus, show poor therapeutic effects. Recently, nanotechnology has gained significant attention with potential for controlled and localized therapeutic agents to improve treatment efficacy. In this study, by utilizing upregulated molecules in PAD’s tissues, we have developed biocompatible nanocarriers to improve cellular absorption, bioavailability and enable therapeutic efficacy for PAD treatments.

Materials and

Methods: Poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) were synthesized by using a standard emulsion technique and conjugated to targeting moiety through covalent coupling. The supernatant was retained for assessing conjugation efficiency via HPLC-LCMS 9030. Sizing, charge, and polydispersity (PDI) of these NPs were quantified using Dynamic Light Scattering (DLS). The in vitro binding properties of the targeting NPs were investigated via cellular uptake studies in static and flow conditions. After NPs’ treatment, activated cells were washed and imaged under fluorescent microscope. Afterward, cells were lysed to evaluate NPs’ fluorescent signals using a spectrophotometer. The measured fluorescence intensity was normalized by a protein concentration using BCA protein assays. The biodistribution of targeting NPs in vivo was studied using the PAD mouse models. The ischemic hindlimb injury was induced by ligating the left femoral artery and its branches, while the right hindlimb was left untreated as a control. The NPs were administered intravenously (IV) via the tail vein. NPs’ accumulation in vivo and ex vivo was assessed using the Kodak in vivo imaging System. The tissues were further harvested to evaluate the amount of these NPs at PAD legs and different organs.

Results, Conclusions, and Discussions: Results and Discussions: We had successfully engineered and incorporated targeting moiety to PLGA NPs, which had average diameters of around 200 nm, surface charges of around -25 mV, and were highly stable up to 3 days. Our binding studies of targeting system in vitro showed higher cellular uptake at least more than 50% in static condition in the first 10 minutes and highest under flow conditions in activated cells. Our in vivo, ex vivo, and muscle homogenization data demonstrated significantly higher NPs accumulation around 2-fold in ischemic sites in our targeting NPs compared to a control. The NPs distribution in ischemic muscles observed under fluorescent microscope also supported the promising capability of our targeting system via high localization at ischemic tissues.

Conclusion: Our studies conducted in vitro and in vivo have demonstrated that NPs specifically targeted activated cells under flow conditions and ischemic hindlimb legs in mice, thus offering the potential for site specific drug delivery in PADs. More in vitro studies will be conducted to assess NPs uptake pathways and the cytocompatibility of the NPs such as cell viability and hemocompatibility. In vivo toxicity will also be investigated by monitoring body weight, hematocrit, and toxicity biomarkers such as troponin, serum creatinine, and lactate dehydrogenase in other organs.

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