Optimizing CpG spatial distribution with DNA origami for Th1-polarized therapeutic vaccination

Multivalent presentation of ligands often enhances receptor activation and downstream signaling. DNA origami offers precise nanoscale spacing of ligands, a potentially useful feature for therapeutic nanoparticles. Here we introduce a square-block DNA origami platform to explore the importance of spacing of CpG oligonucleotides, which engage Toll-like receptors and thereby act as danger signals for dendritic cells. Through in vitro cell-culture studies and in vivo tumor-treatment models, we demonstrate that square blocks induce Th1 immune polarization when CpG is spaced at 3.5 nm. We observe that this DNA origami vaccine enhances DC activation, antigen cross-presentation, CD8 T cell activation, Th1-polarized CD4 activation and NK cell activation. The vaccine also synergizes effectively with anti-PD-L1 for improved cancer immunotherapy in melanoma and lymphoma models and induced long-term T cell memories. Our results suggest that DNA origami may serve as an advanced vaccine platform for controlling adjuvant spacing and co-delivering antigens. ### Competing Interest Statement W.M.S, J.H.R. and Y.C.Z. are inventors on U.S. patent application PCT/US2020/036281 held by Dana-Farber Cancer Institute, Korea Institute of Science & Technology, and Wyss Institute (filed on 6/5/2020) partially based on this work. All other authors declare that they have no competing interests.