A TLR7-nanoparticle adjuvant promotes broad immune responses against heterologous strains of Influenza and SARS-CoV-2

Effective vaccines against viruses such as Influenza and SARS-CoV-2 must elicit a diverse repertoire of antibodies against multiple variant virus strains. However, antibody responses to current vaccines often lack cross-reactivity due to immunodominance. Here, we describe the synthesis of a toll-like receptor 7 agonist (TLR7)-nanoparticle adjuvant, TLR7-NP, constructed from TLR7 agonist-initiated ring-opening polymerization of lactide and self-assembly with poly(ethylene glycol)-b-poly(lactic-co-glycolic acid). TLR7-NP can enhance lymph node targeting, leading to persistent activation of immune cells. When mixed with Alum-adsorbed antigens, this TLR7-NP adjuvant elicited cross-reactive antibodies for both dominant and subdominant epitopes, as well as antigen-specific CD8+ T cell responses, in mice. TLR7-NP adjuvanted influenza subunit vaccine successfully protected mice from heterologous viral challenge. TLR7-NP also enhanced the antibody response to a SARS-CoV-2 subunit vaccine against multiple variants and revealed the mobilization of an antiviral response. We further demonstrate enhanced antigen-specific responses in human tonsil organoids with this novel adjuvant. The human tonsil cells were cultured to form organoids to test the vaccine and the adjuvants. We added a SARS-CoV-2 spike protein RBD nanoparticle (RBD-NP) with or without TLR7-NPs directly to the cultures. We harvested cells on day04 and day09. These cells were barcoded and FACS-sorted the live B cells for single-cell targeted RNA-seq with antibody-seq (CITE-seq). The library was prepared using the BD Rhapsody Targeted mRNA and AbSeq Reagent Kit (Cat. # 633774)and sequenced in one batch. Briefly, cells were captured with beads in microwell plates, followed by cell lysis, bead retrieval, cDNA synthesis, template switching and Klenow extension, and library preparation following the BD Rhapsody protocol. Libraries were prepared for sample tags, targeted mRNA using the customized Immune Response panel, and AbSeq. NovaSeq (Illumina, San Diego, CA) at Novogene USA Inc. (US Davis, CA) completed the sequencing.