(704a) Targeting Dendritic Cells with Functionalized Polyanhydride Nanoparticles

The design of single dose vaccines is particularly important in providing immunological defense against priority diseases (e.g., influenza, measles, hepatitis, etc.) and diseases with bioterrorism potential (i.e., plague, anthrax, avian influenza, etc.). The use of vaccine adjuvants, which trigger early innate immune responses to aid in the generation of robust and long-lasting adaptive immune responses, is crucial to vaccine efficacy. Polyanhydride nanoparticles are promising platforms for use as single dose vaccine delivery systems. These vehicles have been shown to: a) stabilize protein antigens; b) provide sustained release of antigens; and c) enhance the adjuvant effect by modulating the immune response. Dendritic cells (DCs) are antigen presenting cells that play a major role in connecting the innate and adaptive immune systems. Receptors involved in the recognition and uptake of pathogens by DCs are crucial for establishing a robust immune response. C-type lectin receptors (CLRs) (e.g., mannose receptor and DC-SIGN) recognize carbohydrate structures on pathogens. Targeting CLRs on DCs leads to the internalization of the pathogen, its degradation, and subsequent antigen presentation by specific CLR pathways or by crosstalk with other receptors such as Toll-Like Receptors (TLRs).

This study outlines a novel approach to target CLRs such as the mannose receptor and DC-SIGN on DCs by functionalizing the surface of polyanhydride nanoparticles with specific carbohydrates (i.e., lactose and di-mannose) in order to mimic pathogen surfaces. An anti-solvent nanoencapsulation method was used to fabricate nanoparticles based on a 50:50 ratio of 1,8-bis(p-carboxyphenoxy)-3,6-dioaoctane (CPTEG) and 1,6-bis(p-carboxyphenoxy)hexane (CPH). The surface of polyanhydride nanoparticles was modified by attaching either lactose or di-mannose residues by an amine-carboxylic acid coupling reaction. Particle morphology was characterized by scanning electron microscopy and particle size and charge were measured using quasi-elastic light scattering. The presence of carbohydrates on the surface was quantified by X-ray photoelectron spectroscopy (XPS) and a phenol-sulfuric acid quantification assay. Modified and unmodified nanoparticles were added to in vitro cultures of bone marrow-derived dendritic cells from C57BL/6 mice. Flow cytometry was used to assess for the expression of MHC II, CD86, CD40, mannose receptor (CD206), and DC-SIGN (CD209) on DCs co-incubated with the various nanoparticles. Production of the cytokines IL-1β, IL-10, TNF-α, IL-6, and IL-12p40 was assayed using the Luminex® Multiplex assay.

The targeted activation of murine DCs by novel ?pathogen like? polyanhydride nanoparticles was demonstrated by the increased expression of CD206 and CD209. Such a targeted of antigen-loaded nanoparticles is an important determinant of antigen processing and presentation by DCs, which ultimately affects the nature of the immune response. This is also highlighted by the accompanying increased expression of MHC II and CD86 on the DCs. These studies provide key insights into the rational design of targeted nanovaccine platforms that will enhance the induction of antigen-specific immune responses.