Optimization of Stability, Encapsulation, Release, and Cross-Priming of Tumor Antigen-Containing PLGA Nanoparticles
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In order to investigate Poly (lactic-co-glycolic acid) (PLGA) nanoparticles (NP) as potential vehicles for efficient tumor antigen (TA) delivery to dendritic cells (DC), this study aimed to optimize encapsulation/release kinetics before determining immunogenicity of antigen-containing NP.
Various techniques were used to liberate TA from cell lines. Single (gp100) and multiple (B16-tumor lysate containing gp100) antigens were encapsulated within differing molecular weight PLGA co-polymers. Differences in morphology, encapsulation/release and biologic potency were studied. Findings were adopted to encapsulate fresh tumor lysate from patients with advanced tumors and compare stimulation of tumor infiltrating lymphocytes (TIL) against that achieved by soluble lysate.
Four cycles of freeze-thaw + 15 s sonication resulted in antigen-rich lysates without the need for toxic detergents or protease inhibitors. The 80KDa polymer resulted in maximal release of payload and favorable production of immunostimulatory IL-2 and IFN-γ. NP-mediated antigen delivery led to increased IFN-γ and decreased immunoinhibitory IL-10 synthesis when compared to soluble lysate.
Four cycles of freeze-thaw followed by 15 s sonication is the ideal technique to obtain complex TA for encapsulation. The 80KDa polymer has the most promising combination of release kinetics and biologic potency. Encapsulated antigens are immunogenic and evoke favorable TIL-mediated anti-tumor responses.
Key Wordsantigen delivery dendritic cell immunotherapy molecular weight nanoparticles
antigen presenting cell
American Type Culture Collection
bone marrow dendritic cells
bovine serum albumin
cytometric bead array
cluster of differentiation
cytotoxic T cells
- FT + S
freeze-thaw + sonication
granulocyte macrophage-colony stimulating factor
head and neck squamous cell carcinoma
phosphate buffered saline
poly (lactic-co-glycolic acid)
sodium dodecyl sulfate polyacrylamide gel electrophoresis
scanning electron microscopy
Statistical Package for the Social Sciences
tumor associated antigen
tumor infiltrating lymphocytes
tumor necrosis factor-α
Acknowledgments and Disclosures
The authors thank Prof Mark Saltzman and Dr Camille Solbrig in Biomedical Engineering for assistance and supply of reagents associated with NP encapsulation, Dr Michael Girardi and Kacie Carlson in Dermatology for their help in animal handling, and members of the Girardi and Cresswell laboratory for helpful discussions. We also thank Prof Sasaki and Shelley Jolie in Otolaryngology Head and Neck Surgery, Dr Diane Kowalski and Lori Patruno in Surgical Pathology, and members of the team of anaesthetists involved in care of patients recruited to this study. This work was partially funded by intra mural grant available to the Department of Dermatology, Yale University School of Medicine and partially by a NCI/NTRAC grant managed by Cancer Research UK. There is no perceived, potential or real conflict of interest.
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