7P - Tumor cell-released autophagosomes (TRAPs) promote breast cancer lung metastasis by modulating neutrophil extracellular traps formation

Background

Neutrophil extracellular traps (NETs) are involved in the progression and metastasis of a variety of malignancies. Our previous studies have confirmed that tumor cell-released autophagosomes (TRAPs) induced immunosuppression TME formation. However, it remains to be investigated whether TRAPs-treated neutrophils contribute to the metastatic colonization of the lungs by tumor cells.

Methods

NETs were observed by scanning electron microscopy (SEM) and Confocal Microscope. Western blot and ELISA were used to quantify MPO-DNA, NE, and cit-H3 which are important components of NETs. In vivo, TRAPs were injected into the tail vein of mice and Beclin1 knockdown 4T1 tumor cells engineering to reduce TRAPs release were injected into mice subcutaneously. The characteristic molecules of NETs in plasma were detected. The study used antibody blocking assays and HMGB1 knockdown cell lines to identify key DAMPs on the surface of TRAPs. Inhibitor-treated neutrophils and TLR4 knockout mice were used to identify functional receptors on neutrophils. Flow cytometry was used to evaluate the effect of TRAPs on T cell function and lung infiltrating T cell function, as well as to monitor late lung metastases in neutrophils treated with TRAPs suppressor.

Results

Numerous reticular structures significantly increased in the cell culture supernatant after TRAPs treatment. In vivo, NETs were significantly increased in plasma after tail vein injection of TRAPs as well as in 4T1 tumor-bearing mice. Conversely, NETs were significantly decreased in the plasma of Beclin1 knockdown 4T1 tumor-bearing mice. TRAPs derived from breast tumor cell lines induced neutrophil formation of NETs via the HMGB1-TLR4-MyD88-ERK/p38 pathway. This process inhibited the proliferation and secretion of IFN-γ in CD4⁺ and CD8⁺ T cells, ultimately leading to increased lung metastasis.

Conclusions

Tumor cell-released autophagosomes (TRAPs) promote breast cancer lung metastasis by modulating neutrophil extracellular traps formation. Overall, these findings define a novel mechanism mediated by TRAPs in neutrophils, which may suppress anti-tumor T cell immunity and highlight TRAPs as an important target for future tumor immunotherapy.

Clinical trial identification

Editorial acknowledgement

Legal entity responsible for the study

C. Wu, X. Wang.

Funding

Supported by National Natural Science Foundation of China.

Disclosure

The author has declared no conflicts of interest.