Neutral polymer micelle carriers with pH-responsive, endosome-releasing activity modulate antigen trafficking to enhance CD8(+) T cell responses.
Synthetic subunit vaccines need to induce CD8(+) cytotoxic T cell (CTL) responses for effective vaccination against intracellular pathogens. Most subunit vaccines primarily generate humoral immune responses, with a weaker than desired CD8(+) cytotoxic T cell response. Here, a neutral, pH-responsive polymer micelle carrier that alters intracellular antigen trafficking was shown to enhance CD8(+) T cell responses with a correlated increase in cytosolic delivery and a decrease in exocytosis. Polymer diblock carriers consisted of a N-(2-hydroxypropyl) methacrylamide corona block with pendent pyridyl disulfide groups for reversible conjugation of thiolated ovalbumin, and a tercopolymer ampholytic core-forming block composed of propylacrylic acid (PAA), dimethylaminoethyl methacrylate (DMAEMA), and butyl methacrylate (BMA). The diblock copolymers self-assembled into 25-30nm diameter micellar nanoparticles. Conjugation of ovalbumin to the micelles significantly enhanced antigen cross-presentation in vitro relative to free ovalbumin, an unconjugated physical mixture of ovalbumin and polymer, and a non-pH-responsive micelle-ovalbumin control. Mechanistic studies in a murine dendritic cell line (DC 2.4) demonstrated micelle-mediated enhancements in intracellular antigen retention and cytosolic antigen accumulation. Approximately 90% of initially internalized ovalbumin-conjugated micelles were retained in cells after 1.5h, compared to only ~40% for controls. Furthermore, cells dosed with conjugates displayed 67-fold higher cytosolic antigen levels relative to soluble ovalbumin 4h post uptake. Subcutaneous immunization of mice with ovalbumin-polymer conjugates significantly enhanced antigen-specific CD8(+) T cell responses (0.4% IFN-γ(+) of CD8(+)) compared to immunization with soluble protein, ovalbumin and polymer mixture, and the control micelle without endosome-releasing activity. Additionally, pH-responsive carrier facilitated antigen delivery to antigen presenting cells in the draining lymph nodes. As early as 90min post injection, ova-micelle conjugates were associated with 28% and 55% of dendritic cells and macrophages, respectively. After 24h, conjugates preferentially associated with dendritic cells, affording 30-, 3-, and 3-fold enhancements in uptake relative to free protein, physical mixture, and the non-pH-responsive conjugate controls, respectively. These results demonstrate the potential of pH-responsive polymeric micelles for use in vaccine applications that rely on CD8(+) T cell activation.
Department of Bioengineering, University of Washington, Box 355061, Seattle, WA 98195-1721, USA. Electronic address: salka@uw.edu.
Salka
S
Keller
Department of Bioengineering, University of Washington, Box 355061, Seattle, WA 98195-1721, USA. Electronic address: wilsonjt@uw.edu.
John T
JT
Wilson
Department of Bioengineering, University of Washington, Box 355061, Seattle, WA 98195-1721, USA. Electronic address: patilg@uw.edu.
Gabriela I
GI
Patilea
Department of Bioengineering, University of Washington, Box 355061, Seattle, WA 98195-1721, USA. Electronic address: kernh@uw.edu.
Hanna B
HB
Kern
Department of Bioengineering, University of Washington, Box 355061, Seattle, WA 98195-1721, USA. Electronic address: aconv@uw.edu.
Anthony J
AJ
Convertine
Department of Bioengineering, University of Washington, Box 355061, Seattle, WA 98195-1721, USA. Electronic address: stayton@uw.edu.
Patrick S
PS
Stayton
D000818:Animals; D000941:Antigens; D002460:Cell Line; D002626:Chemistry, Pharmaceutical; D003692:Delayed-Action Preparations; D003713:Dendritic Cells / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D004337:Drug Carriers; D011992:Endosomes / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D005260:Female; D006863:Hydrogen-Ion Concentration; D007114:Immunization; D008198:Lymph Nodes / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D008213:Lymphocyte Activation / Q000187:drug effects*; D008264:Macrophages / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D008810:Mice, Inbred C57BL; D008823:Micelles; D053758:Nanoparticles; D010047:Ovalbumin / Q000737:chemistry / Q000276:immunology / Q000494:pharmacology*; D010316:Particle Size; D011108:Polymers / Q000737:chemistry*; D021381:Protein Transport; D013602:T-Lymphocytes, Cytotoxic / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D013678:Technology, Pharmaceutical / Q000379:methods; D013997:Time Factors; D022223:Vaccines, Subunit / Q000494:pharmacology
D000941:Antigens; D003692:Delayed-Action Preparations; D004337:Drug Carriers; D008823:Micelles; D011108:Polymers; D022223:Vaccines, Subunit; D010047:Ovalbumin
CD8(+) T cell response; Nanoparticles; Polymer micelles; Subunit vaccine; pH-responsive