Cancer cell-expressed BTNL2 facilitates tumour immune escape via engagement with IL-17A-producing γδ T cells

Therapeutic blockade of the immune checkpoint proteins programmed cell death protein 1 (PD-1) and cytotoxic T lymphocyte antigen 4 (CTLA4) has transformed cancer treatment. However, the overall response rate to these treatments is low, suggesting that immune checkpoint activation is not the only mechanism leading to dysfunctional anti-tumour immunity. Here we show that butyrophilin-like protein 2 (BTNL2) is a potent suppressor of the anti-tumour immune response. Antibody-mediated blockade of BTNL2 attenuates tumour progression in multiple in vivo murine tumour models, resulting in prolonged survival of tumour-bearing mice. Mechanistically, BTNL2 interacts with local γδ T cell populations to promote IL-17A production in the tumour microenvironment. Inhibition of BTNL2 reduces the number of tumour-infiltrating IL-17A-producing γδ T cells and myeloid-derived suppressor cells, while facilitating cytotoxic CD8+ T cell accumulation. Furthermore, we find high BTNL2 expression in several human tumour samples from highly prevalent cancer types, which negatively correlates with overall patient survival. Thus, our results suggest that BTNL2 is a negative regulator of anti-tumour immunity and a potential target for cancer immunotherapy.


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All data presented has been replicated. Most graphs display the collective data from several independent experiments and the number of replicate experiments included are given in the figure legends.
Genetic mouse experiment: Different genotypes from the same litter were compared. Wild-type mice were randomly divided into control treatment or treatment groups. Other experiments are not relevant to the application of randomization.
During assessment of the tumor size and mice survival, the assessor did not know the genotypes or treatments the mice received. For the flow cytometry experiments of Figure 2a , 2b, 2c, 4a, 4b, 4c, 4d, 4e, 4f, 4g, 4j, 4k and 5e, the person who analyzed the data by flow cytometry was blinded to the genotypes or treatments.
All the commercial antibodies were validated by the suppliers, and the validation information can be found on the company's websites. Anti-human BTNL2 polyclonal antibody (Proteintech , cat no. 25110-1-AP). Validation: https://www.ptgcn.com/products/BTNL2-Antibody-25110-1-AP.htm Cell lines were authenticated by short tandem repeat profiling.
All cell lines are negative for mycoplasma.
No commonly misidentified cell lines were used in this study.
BTNL2-KO mice were made by Cyagen Biosciences Inc by Crispr-cas9. BTNL2-KO mice were C57BL/6 background. Six to eight weeks old female C57BL/6 and BALB/c mice were obtained from Beijing Vital River Laboratory Animal Technology Co., Ltd. OT-1 transgenic mice were kindly provided by Prof. Zhengfan Jiang at Peking University and six to eight weeks old male OT-1 transgenic mice were used in our experiments. These mice were housed in SPF condition, the ambient temperature is between 20-25°C, the humidity is between 40-70%, and the environmental light/dark cycle is 12h light, 12h dark. Laboratory animals used are described in the Method section.
The study did not involve wild animals.
The study did not involve samples collected from the field.