Single‐cell RNA sequencing reveals localized tumour ablation and intratumoural immunostimulant delivery potentiate T cell mediated tumour killing

Abstract

Background Metastatic breast cancer poses great challenge in cancer treatment. N‐dihydrogalactochitosan (GC) is a novel immunoadjuvant that stimulates systemic immune responses when administered intratumourally following local tumour ablation. A combination of photothermal therapy (PTT) and GC, referred to as localized ablative immunotherapy (LAIT), extended animal survival and generates an activated B cell phenotype in MMTV‐PyMT mouse mammary tumour microenvironment (TME). However, how T cell populations respond to LAIT remains to be elucidated. Methods Using depletion antibodies, we studied the contributions of CD8 + and CD4 + T cells to the therapeutic effect of LAIT. Using single‐cell RNA‐sequencing (scRNAseq), we analysed tumour‐infiltrating T cell heterogeneity and dissected their transcriptomes upon treatments of PTT, GC, and LAIT (PTT+GC). Results Loss of CD8 + T cells after LAIT abrogated the therapeutic benefits of LAIT. Ten days after treatment, proportions of CD8 + and CD4 + T cells in untreated TME were 19.2% and 23.0%, respectively. Upon LAIT, both proportions were increased to 25.5% and 36.2%, respectively. In particular, LAIT increased the proportions of naïve and memory cells from a resting state to an activated state. LAIT consistently induced the expression of co‐stimulatory molecules, type I IFN responsive genes, and a series of antitumor cytokines, Ifng , Tnf , Il1 , and Il17 in CD8 + and CD4 + T cells. LAIT also induced immune checkpoints Pdcd1, Ctla4 , and Lag3 expression, consistent with T cell activation. Relevant to clinical translation, LAIT also upregulated genes in CD8 + and CD4 + T cells that positively correlated with extended survival of breast cancer patients. Conclusions Overall, our results reveal that LAIT prompts immunological remodelling of T cells by inducing broad proinflammatory responses and inhibiting suppressive signalling to drive antitumour immunity.

A novel biopolymer for mucosal adjuvant against respiratory pathogens

“…However, we also found that GC did not cause direct, non-specific activation of T cells, but instead works through the activation the APCs (data not shown). To determine the critical roles of dendritic cells (DCs) in antigen presentation, T cell activation and differentiation, and effector and memory T cell formation, we discovered the pathways of T cell activation through DC activation, consistent with previous published results 42, 43 . We have previously shown that GC directly induces maturation of dendritic cells via increased co-stimulatory molecules CD80, CD40 and MHC-II 44 .…”

Single-cell transcriptomics reveals that tumor-infiltrating natural killer cells are activated by localized ablative immunotherapy and share anti-tumor signatures induced by immune checkpoint inhibitors

“…We previously reported the promising therapeutic effects of localized ablative immunotherapy (LAIT) on MMTV-PyMT breast tumors and uncovered the anti-tumor roles of tumor-infiltrating B cells 24 and T cells 25 in response to LAIT. Although there are many other reports revealing the transcriptional atlas and anti-tumor roles of both innate and adaptive immune cells by scRNAseq, the contributions of NK cells were less known.…”

“…We have previously developed a localized ablative immunotherapy (LAIT) that combines local photothermal therapy (PTT) and intratumoral injection of a novel immunoadjuvant, Ndilydrogalactochitosan (GC). LAIT extended animal survival in the mouse MMTV-PyMT mammary tumor model 22 as well as B16-F10 melanoma model. 23 Using single-cell RNA sequencing (scRNAseq), we showed that LAIT activates sustainable adaptive antitumor responses by increasing the proportions of naïve CD8 + T-cells and memory CD4 + T-cells from a resting state to an activated state.…”

“…Earlier clustering of scRNAseq from our MMTV-PyMT model revealed subpopulations of NK cells in tumor infiltrated lymphoid cells. 22 Given the complementary role of NK cells to T-cells and B-cells and their potential as therapeutic immune cells, we were interested in understanding NK cells activation and signaling regulation upon GC, PTT, and LAIT treatment.…”

A novel biopolymer synergizes type I IFN and IL-1β production through STING

“…Using RNA sequencing analyses, we examined the immune cells within the TME following intratumoral application of GC and found that GC drove a strong type I interferon (IFN) response in both innate and adaptive tumor-infiltrating leukocytes (TILs). The major immune stimulation in ablation-GC treatment was potentiated by GC (18,19). However, only the combination of ablation and GC was able to achieve sustained tumor regression in both treated tumors and untreated distant metastases (13)(14)(15)(16)(17).…”