Nanoparticle Delivery of Alu RNA Adjuvants Enhances Vaccine Immunogenicity

Vaccine adjuvants stimulate innate immunity to enhance and shape adaptive immune responses. However, approved adjuvants typically elicit weak CD8⁺ T cell responses to protein- and peptide-based vaccines, motivating an investigation into the discovery and testing of new adjuvants. Unedited forms of endogenous Alu RNAs are sensed by pattern recognition receptors (PRRs) to trigger sterile inflammation, and therefore we hypothesized that synthetic Alu RNA molecules could be harnessed as vaccine adjuvants. To enhance their intracellular delivery, Alu RNA was copackaged with a model antigen into polymer nanoparticles that promoted endosomal escape of Alu RNA to the cytosol. Using this nanovaccine formulation, we found that Alu RNA activated antigen-presenting cells in vitro and in vaccine-site draining lymph nodes in vivo. Furthermore, we demonstrated that vaccine formulations containing Alu RNA as an adjuvant elicited comparable CD8⁺ T cell responses to those containing the common but highly heterogeneous RNA adjuvant PolyIC, and that this response protected mice from tumor challenge. Based on this, we further evaluated the antitumor efficacy of nanovaccine formulations containing Alu RNA adjuvants in mice with established tumors, again observing comparable responses to formulations containing PolyIC. Finally, we found that nanovaccines adjuvanted with Alu RNA could improve responses to anti-PD-1 immune checkpoint blockade in tumor-bearing mice. Overall, this study demonstrates that unedited Alu RNA coformulated with antigen in polymer nanoparticles can be harnessed as an effective vaccine adjuvant for stimulating CD8⁺ T cell responses with antitumor function and may offer a sequence-defined alternative to PolyIC.