ORCID ID

0000-0002-2991-9622

Graduation Date

Fall 12-20-2019

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Programs

Pharmaceutical Sciences

First Advisor

Dr. Aaron M Mohs

Abstract

This work presents research using the naturally available non- sulfated carbohydrate glycosaminoglycan hyaluronic acid (HA) for the synthesis of different chemical derivatives of HA for evaluation of binding kinetics with CD44 and P- selectin proteins for applications in fluorescence image-guided surgery. Chemical derivatives of HA such as deacetylated HA (deHA), sulfated HA (sHA), and deacetylated and sulfated HA (s-deHA) were synthesized by modulating sulfating and deacetylating reagents to alter binding specificities to CD44. Modified HA derivatives and CD44 biophysical interactions were assessed by fluorescence polarization. In silico techniques were also used to determine binding using molecular docking and MM-PBSA approaches. Both modifications represent the first step to effectively re-target HA-based materials. Re- targeting HA was successfully conducted by tuning the sulfate distribution on HA to P-selectin. Three derivatives of HA were synthesized by controlling the sulfation agent to give selectively sulfated HA (ssHA), moderately sulfated HA (msHA), and completely sulfated HA (csHA). The strongest binding was observed for the csHA with recombinant P-selectin when compared with fucoidan. Amphiphilic conjugates were prepared for the three derivatives of HA by conjugating a hydrophobic ligand (1- pyrenebutanamide- PBA) followed by chemical conjugation of Cyanine7.5 dye to give near-infrared fluorescent nanoparticles. Modified HA nanoparticles were assessed for uptake in P-selectin and CD44 positive cells along with knockout cell lines. Preference for P-selectin mediated uptake was observed in vitro binding assays. In vivo performance for image-guided surgery applications indicated higher uptake of sulfated HA nanoparticles compared to non-sulfated HA nanoparticles in S2-013 tumors grown subcutaneously in nude mice. HA as a non- toxic, biocompatible material is an ideal precursor for generating newer imaging probes for biomedical applications. HA as a non-fluorescent precursor was conjugated with non-fluorescent biocompatible amino acids to form a novel fluorescent organic nanoparticle (FONPs) using one-pot green chemistry conditions. The FONPs exhibited high water solubility due to the existence of hydrophilic functional groups on its surface, acceptable stability profiles, superior biocompatibility, and high quantum yield photo-luminescent properties. Also, the FONPs demonstrated drug carrier properties with enhanced drug loading and delivery of the anticancer drug doxorubicin (DOX) in cancer cells and 3D tumor spheroid model. Results demonstrate the efficacy of a novel fluorescent HA-based with potential future applications in image-guided drug delivery.

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