Innovation in mRNA Vaccines and RNAi via Protein Nanocages

Abstract

Self-assembling protein nanocages (SAPNs) are distinct natural structures formed by the self-assembly of identical subunits, providing a highly efficient platform and a novel strategy for vaccine development and RNAi therapy. Their internal cavity allows for precise cargo encapsulation, while the externally modifiable surface supports multivalent antigen presentation, thereby enhancing stability, targeted delivery, and immune activation. In addition to serving as stable subunit vaccines with multivalent antigen display, SAPNs can be incorporated into mRNA vaccines (SAPN-RNA vaccines) by pre-fusing with the antigen. This strategy stabilizes secreted antigenic proteins with prolonged presentation to the immune system, and improves vaccine efficacy while reducing off-target effects and minimizing required doses. Additionally, SAPNs can overcome cellular uptake barriers, enhance DNA vaccine efficacy, and enable the co-delivery of antigens and adjuvants. Functionalization with adjuvants or targeting ligands further improves their immunostimulatory properties and specificity. The SAPN-RNAi strategy optimizes siRNA delivery by promoting lysosomal escape, enhancing targeted uptake, and protecting siRNA from degradation through SAPN encapsulation. This review examines the structural and functional properties of protein nanocages and their applications in vaccine design and RNAi delivery, emphasizing their synergistic effects, and exploring current progress, challenges, and future directions. In conclusion, SAPNs represent a versatile multifunctional platform with broad applicability across subunit, mRNA and DNA vaccines, adjuvant co-delivery, and RNAi therapeutics, with significant potential against viral infections.

Keywords: ferritin; mRNA vaccines; protein nanocages; siRNA delivery; vaccine platform; virus.

Figure 1

Self-assembling protein nanocages (SAPNs) for multivalent antigen display and cargo encapsulation. (A) Schematic representation of ferritin nanocages and a monomeric subunit. Ferritin assembles into a 24-subunit spherical cage with inner and outer diameters of approximately 8 nm and 12 nm, respectively. Multivalent antigen display can be achieved via genetic fusion of antigen-encoding sequences to the C-terminal region of ferritin subunits. (B) Schematic illustration of a core encapsulin operon and targeting peptide (TP)-mediated cargo encapsulation. Encapsulin monomers self-assemble into icosahedral protein cages ranging from 20 to 42 nm in diameter, composed of 60, 180, or 240 identical subunits. A domain-level view of the encapsulin monomer is shown. Antigen display on the encapsulin surface can also be achieved through C-terminal genetic fusion. Created with BioRender.com.

Figure 2

Self-assembling protein nanocages (SAPNs) as versatile platforms for subunit, RNA, and DNA vaccines. (A) SAPN-Subunit Vaccines: antigens are covalently linked to protein nanocages using the SpyTag/SpyCatcher system, enabling a highly ordered, multivalent display with enhanced stability. This promotes robust immune responses by directly activating naïve B cells through efficient BCR cross-linking and facilitating antigen uptake by antigen-presenting cells (APCs) such as such as dendritic cells (DCs), leading to strong cellular immunity and durable, high-affinity antibody production. (B) SAPN-RNA Vaccines: RNA platforms, including conventional mRNA, self-amplifying RNA (saRNA), trans-amplifying RNA (taRNA), and circular RNA (circRNA), encode SAPN-antigen fusion proteins. Delivered via lipid nanoparticles (LNPs), these RNAs are translated in situ, generating stabilized antigens that enhance immune presentation, cellular uptake, and overall vaccine efficacy while reducing off-target effects and required doses. (C) SAPN-DNA Vaccines: plasmid DNA encoding SAPN–antigen fusions is delivered intramuscularly (IM), leading to in situ antigen production within host cells. This forms multivalent antigen displays with enhanced stability, cellular uptake, and immunogenicity. The SAPN platform is based on lumazine synthase, which self-assembles into 60-subunit icosahedral nanocages (~16 nm in diameter; PDB ID: 8F25). Created with BioRender.com.

Figure 3

Schematic of SAPN-RNAi delivery for antiviral therapy. (A) Self-assembling protein nanocages (SAPNs) for RNAi encapsulation: siRNA is encapsulated within ferritin via reversible disassembly at pH 2 and reassembly at pH 7. (B) Antiviral mechanism of SAPN-RNAi: SAPN protects siRNA from degradation and facilitates cellular uptake via endocytosis. After endosomal escape, Dicer processes long dsRNA into siRNAs that guide Ago2-RISC to degrade viral mRNA and inhibit protein synthesis. Targeting-ligands on the SAPN surface ensure cell-specific delivery. Created with BioRender.com.