title: Optimization, Production, and Characterization of a CpG-Oligonucleotide-Ficoll Conjugate Nanoparticle Adjuvant for Enhanced Immunogenicity of Anthrax Protective Antigen.
abstract: We have synthesized and characterized a novel phosphorothioate CpG oligodeoxynucleotide (CpG ODN)-Ficoll conjugated nanoparticulate adjuvant, termed DV230-Ficoll. This adjuvant was constructed from an amine-functionalized-Ficoll, a heterobifunctional linker (succinimidyl-[(N-maleimidopropionamido)-hexaethylene glycol] ester) and the CpG-ODN DV230. Herein, we describe the evaluation of the purity and reactivity of linkers of different lengths for CpG-ODN-Ficoll conjugation, optimization of linker coupling, and conjugation of thiol-functionalized CpG to maleimide-functionalized Ficoll and process scale-up. Physicochemical characterization of independently produced lots of DV230-Ficoll reveal a bioconjugate with a particle size of approximately 50 nm and covalent attachment of more than 100 molecules of CpG per Ficoll. Solutions of purified DV230-Ficoll were stable for at least 12 months at frozen and refrigerated temperatures and stability was further enhanced in lyophilized form. Compared to nonconjugated monomeric DV230, the DV230-Ficoll conjugate demonstrated improved in vitro potency for induction of IFN-α from human peripheral blood mononuclear cells and induced higher titer neutralizing antibody responses against coadministered anthrax recombinant protective antigen in mice. The processes described here establish a reproducible and robust process for the synthesis of a novel, size-controlled, and stable CpG-ODN nanoparticle adjuvant suitable for manufacture and use in vaccines.
authors.affiliation: Dynavax Technologies Corporation , 2929 Seventh Street, Suite 100, Berkeley, California 94710, United States.
authors.forename: Bob
authors.initials: B
authors.lastname: Milley
authors.affiliation: Dynavax Technologies Corporation , 2929 Seventh Street, Suite 100, Berkeley, California 94710, United States.
authors.forename: Radwan
authors.initials: R
authors.lastname: Kiwan
authors.affiliation: Dynavax Technologies Corporation , 2929 Seventh Street, Suite 100, Berkeley, California 94710, United States.
authors.forename: Gary S
authors.initials: GS
authors.lastname: Ott
authors.affiliation: Dynavax Technologies Corporation , 2929 Seventh Street, Suite 100, Berkeley, California 94710, United States.
authors.forename: Carlo
authors.initials: C
authors.lastname: Calacsan
authors.affiliation: Dynavax Technologies Corporation , 2929 Seventh Street, Suite 100, Berkeley, California 94710, United States.
authors.forename: Melissa
authors.initials: M
authors.lastname: Kachura
authors.affiliation: Dynavax Technologies Corporation , 2929 Seventh Street, Suite 100, Berkeley, California 94710, United States.
authors.forename: John D
authors.initials: JD
authors.lastname: Campbell
authors.affiliation: MedImmune LLC , One MedImmune Way, Gaithersburg, Maryland 20878, United States.
authors.forename: Holger
authors.initials: H
authors.lastname: Kanzler
authors.affiliation: Dynavax Technologies Corporation , 2929 Seventh Street, Suite 100, Berkeley, California 94710, United States.
authors.forename: Robert L
authors.initials: RL
authors.lastname: Coffman
mesh_terms: D000276:Adjuvants, Immunologic / Q000737:chemistry*; D000818:Animals; D000942:Antigens, Bacterial / Q000276:immunology*; D001427:Bacterial Toxins / Q000276:immunology*; D015195:Drug Design; D004355:Drug Stability; D005362:Ficoll / Q000737:chemistry*; D006801:Humans; D008301:Maleimides / Q000737:chemistry; D008745:Methylation; D051379:Mice; D053758:Nanoparticles / Q000737:chemistry*; D009838:Oligodeoxyribonucleotides / Q000737:chemistry* / Q000276:immunology*; D011092:Polyethylene Glycols / Q000737:chemistry
chemical_list: D000276:Adjuvants, Immunologic; D000942:Antigens, Bacterial; D001427:Bacterial Toxins; C408982:CPG-oligonucleotide; D008301:Maleimides; D009838:Oligodeoxyribonucleotides; C030325:anthrax toxin; C043592:maleimide; D005362:Ficoll; D011092:Polyethylene Glycols