The heterogeneous nature of tumour microenvironment (TME) underlying diverse treatment responses remains unclear in nasopharyngeal carcinoma (NPC). Here, we profile 176,447 cells from 10 NPC tumour-blood pairs, using single-cell transcriptome coupled with T cell receptor sequencing. Our analyses reveal 53 cell subtypes, including tumour-infiltrating CD8⁺T, regulatory T (Treg), and dendritic cells (DCs), as well as malignant cells with different Epstein-Barr virus infection status. Trajectory analyses reveal exhausted CD8⁺T and immune-suppressive TNFRSF4⁺Treg cells in tumours might derive from peripheral CX3CR1⁺CD8⁺T and naïve Treg cells, respectively. Moreover, we identify immune-regulatory and tolerogenic LAMP3⁺DCs. Noteworthily, we observe intensive inter-cell interactions among LAMP3⁺DCs, Treg, exhausted CD8⁺T, and malignant cells, suggesting potential cross-talks to foster an immune-suppressive niche for the TME. Collectively, our study uncovers the heterogeneity and interacting molecules of the TME in NPC at single-cell resolution, which provide insights into the mechanisms underlying NPC progression and the development of precise therapies for NPC.

Significance: Our study has identified a minimalist nanovaccine with C1 lipid nanoparticle (LNP) that effectively promotes mRNA delivery and antigen presentation with a self-adjuvant feature through activating TLR4 signaling. The C1 LNP mRNA nanovaccine exhibited significant in vivo efficacy in both tumor prevention and therapeutic vaccine settings. Therefore, our work presents a mRNA nanovaccine platform for developing cancer immunotherapy for a wide range of tumor types.

Acute myeloid leukemia (AML) is one of the most common types of adult acute leukemia. Standard chemotherapies can induce complete remission in selected patients; however, a majority of patients eventually relapse and succumb to the disease. Thus, the development of novel therapeutics for AML is urgently needed. Human C-type lectin-like molecule-1 (CLL-1) is a type II transmembrane glycoprotein, and its expression is restricted to myeloid cells and the majority of AML blasts. Moreover, CLL-1 is expressed in leukemia stem cells (LSCs), but absent in hematopoietic stem cells (HSCs), which may provide a potential therapeutic target for AML treatment. We tested the expression of CLL-1 antigen on peripheral blood cells and bone marrow cells in healthy donor and AML patients. Then, we developed a chimeric antigen receptor (CAR) containing a CLL1-specific single-chain variable fragment, in combination with CD28, 4-1BB costimulatory domains, and CD3-ζ signaling domain. We further investigate the function of CLL-1 CAR-T cells. The CLL-1 CAR-T cells specifically lysed CLL-1+ cell lines as well as primary AML patient samples in vitro. Strong anti-leukemic activity was observed in vivo by using a xenograft model of disseminated AML. Importantly, CLL-1+ myeloid progenitor cells and mature myeloid cells were specifically eliminated by CLL-1 CAR-T cells, while normal HSCs were not targeted due to the lack of CLL-1 expression. CLL-1 CAR-T represents a promising immunotherapy for the treatment of AML.

Tertiary lymphoid structures are immune cell aggregates linked with cancer outcomes, but their interactions with tumour cell aggregates are unclear. Using nasopharyngeal carcinoma as a model, here we analyse single-cell transcriptomes of 343,829 cells from 77 biopsy and blood samples and spatially-resolved transcriptomes of 31,316 spots from 15 tumours to decipher their components and interactions with tumour cell aggregates. We identify essential cell populations in tertiary lymphoid structure, including CXCL13⁺ cancer-associated fibroblasts, stem-like CXCL13⁺CD8⁺ T cells, and B and T follicular helper cells. Our study shows that germinal centre reaction matures plasma cells. These plasma cells intersperse with tumour cell aggregates, promoting apoptosis of EBV-related malignant cells and enhancing immunotherapy response. CXCL13⁺ cancer-associated fibroblasts promote B cell adhesion and antibody production, activating CXCL13⁺CD8⁺ T cells that become exhausted in tumour cell aggregates. Tertiary lymphoid structure-related cell signatures correlate with prognosis and PD-1 blockade response, offering insights for therapeutic strategies in cancers.