Polyanhydride microparticles enhance dendritic cell antigen presentation and activation

Abstract

The present studies were designed to evaluate the adjuvant activity of polyanhydride microparticles prepared in the absence of additional stabilizers, excipients, or immune modulators. Microparticles composed of varying ratios of either 1,6-bis(p-carboxyphenoxy)hexane (CPH) and sebacic acid (SA) or 1,8-bis(p-carboxyphenoxy)-3,6-dioxaoctane (CPTEG) and CPH were added to in vitro cultures of bone marrow-derived dendritic cells (DCs). Microparticles were efficiently and rapidly phagocytosed by DCs in the absence of opsonization and without centrifugation or agitation. Within 2 h, internalized particles were rapidly localized to an acidic, phagolysosomal compartment. By 48 h, only a minor reduction in microparticle size was observed in the phagolysosomal compartment, indicating minimal particle erosion consistent with being localized within an intracellular microenvironment favoring particle stability. Polyanhydride microparticles increased DC surface expression of MHC II, the co-stimulatory molecules CD86 and CD40, and the C-type lectin CIRE (murine DC-SIGN; CD209). In addition, microparticle stimulation of DCs also enhanced secretion of the cytokines IL-12p40 and IL-6, a phenomenon found to be dependent on polymer chemistry. DCs cultured with polyanhydride microparticles and ovalbumin induced polymer chemistry-dependent antigen-specific proliferation of both CD4+ OT-II and CD8+ OT-I T cells. These data indicate that polyanhydride particles can be tailored to take advantage of the potential plasticity of the immune response, resulting in the ability to induce immune protection against many types of pathogens.

Mannose-Functionalized “Pathogen-like” Polyanhydride Nanoparticles Target C-Type Lectin Receptors on Dendritic Cells

“…MHC II, CD40, and CD86 play an important role in the induction of adaptive immunity through activation of CD4 + T cells, and it would be beneficial for an effective adjuvant to enhance the expression of MHC II and costimulatory molecules. 1,9,34 CIRE and the mannose receptor play an important role in antigen uptake, as well as in DC migration and initial interaction with T cells. 9À11, 13 As mentioned before, the activation of CLRinduced signaling processes can result in the induction of diverse immune responses.…”

“…Flow cytometric analysis of surface molecule expression was performed as previously described. 34 Additional antibodies used in this study include FITC antimouse CD206 (MMR, clone MR5D3) and the isotype-specific control antibody FITC rat IgG2a k (clone RTK2748) purchased from BioLegend (San Diego, CA), and biotin anti-mouse CD209 (CIRE, clone 5H10) purchased from eBioscience (San Diego, CA). As noted, the two antibodies used for FACS analysis are different from the antibodies used to block CLRs, thus preventing false negatives due to unavailable binding sites following treatment with the blocking antibody.…”

“…After stimulation for 48 h with nonfunctionalized or functionalized nanoparticles, cell-free supernatants were assayed for IL-1β, IL-10, TNF-α, IL-6, and IL-12p40 using a multiplex cytokine assay in conjunction with a Luminex 100 System (Flowmetrics, Austin, TX) as described elsewhere. 34,35 Fluorescence Microscopy Analysis of Nanoparticle Uptake by DCs. To visualize nanoparticle interactions with DCs, a previously described experimental protocol for coincubation, immunofluorescence preparation and microscopic observation was followed.…”

Novel, High Throughput Method to Study in Vitro Protein Release from Polymer Nanospheres

“…[1][2][3][4][5] Often, the dual tasks of providing an environment capable of maintaining the functionality of the protein drug and releasing it in a sustained manner are challenging. In addition, the assays available for protein release kinetics are limited in that they lack sensitivity and are resource consuming.…”

“…1,3,14,[17][18][19][20][21][22][23][24][25][26][27][28] The polymers of interest are based on 1,6-bis(p-carboxyphenoxy)hexane (CPH) and sebacic acid (SA). In addition to their biocompatibility, the degradation of these materials is base-catalyzed 6,29 and can be tuned allowing for drug delivery ranging from weeks(SA-richchemistries)tomonths(CPH-richchemistries).…”