Host–Guest Assembly of Transmucosal Molecular Spherical Nucleic Acids for Oral Leukoplakia Immunotherapy and Microbiota Homeostasis Restoration

Oral leukoplakia (OLK) is a prevalent oral mucosal disorder and is at risk of malignant transformation. While nanomedicines hold great promise for OLK therapy, their current nanoformulations for topical treatment of OLK typically suffer from poor transmucosal penetration, neglect immunomodulation for suppressing epithelial hyperproliferation, and lack insights into restoration of oral microbiota homeostasis. Herein, we leverage host-guest molecular recognition for the precise construction of an unprecedented molecular spherical nucleic acid (SNA), supramolecular SNA (sSNA). Briefly, β-cyclodextrin (β-CD) and indocyanine green are comodified on the spherical poly-(amidoamine) dendrimer. The β-CD-grafted dendrimers enable high-performance radial arrangement of ferrocene-tagged CpG immunoadjuvant and PD-L1 aptamer through host-guest recognition. The thus-formed sSNA-CpG/Apt_PD‑L1, when applied topically to OLK lesions, exhibits superior transmucosal penetration through the epithelial barrier. Upon ultrasound irradiation, sSNA-CpG/Apt_PD‑L1 generates cytotoxic reactive oxygen species (ROS), suppressing the OLK cell hyperproliferation and even triggering immunogenic cell death. Concurrently, ROS-mediated dissociation of the supramolecular assembly releases CpG oligonucleotides, which enhance antigen-presenting function in dendritic cells and potentiate the activation of CD4⁺/CD8⁺ T cells. Simultaneously, the released PD-L1 aptamers block PD-1/PD-L1 checkpoint signaling, sustaining T-lymphocyte activation, ultimately inhibiting pathological OLK progression. Importantly, this sonodynamic immunotherapy establishes adaptive immunological memory by expanding central and effector memory T cell populations, which prevents OLK recurrence and impedes malignant transformation. Furthermore, this therapeutic modality remolds the composition and function of oral microbiota, restoring the homeostasis of oral ecosystem. In short, this study proposes an unprecedented sSNA, establishes the first immunotherapeutic approach against OLK, and restores oral microbiota homeostasis during therapy.