S100a9 plays a cell specific role in the bone marrow - lung cancer crosstalk

Introduction

A high influx of immunosuppressive myeloid cells in lung tumors hinders effective immunotherapy. The bone marrow niche (BMN), a major source of myeloid cells, is understudied due to its inaccessibility, a low hematopoietic stem cell (HSPCs) fraction, and the dynamic nature of hematopoiesis. Recent single-cell RNA sequencing of bone marrow from Lewis lung carcinoma (LLC)-bearing mice allowed us to identify S100a9 as a potential driver of emergency myelopoiesis. However, its spatiotemporal cell-specific expression and protein-level function within the tumor and the BMN remain unknown.

Methods

To validate alterations in the expression of S100a9 and study its role in emergency hematopoiesis and lung cancer progression, in vitro and in vivo assays were conducted using LLC and KP (KrasG12D; p53ko/ko) cell lines. In vitro, bone marrow was cultured in tumor-conditioned medium or co-cultured with tumor cells, with or without the S100a9 inhibitor Tasquinimod. In vivo, bone marrow and lung tumors were collected at three disease stages determined by μCT imaging. Subsequently, tumors and femoral bones underwent flow cytometry and immunohistochemistry to assess phenotype, spatial organization, and S100a9 expression. We plan to treat LLC-bearing mice with Tasquinimod while HSPC changes and tumor progression are monitored via calvarial intravital imaging and μCT imaging resp.

Results/Discussion

In vitro validation was performed using flow cytometry of bone marrow cultured with control or tumor-conditioned medium (CM) from two lung cancer cell lines: LLC and KP (KrasG12D; p53ko/ko). Bone marrow cultured with CM revealed significant changes in the progenitor compartment, with increased neutrophils and Ly6c+ monocytes and reduced MHCII-low macrophages. These findings aligned with ex vivo observations, which additionally showed a reduction in B cells but no changes in the monocytic compartment. scRNA-seq data indicated increased progenitors, basophils/eosinophils, and erythroid compartments, along with transcriptomic upregulation of S100a9 in these populations. While flow cytometry, from in vivo and ex vivo experiments, confirmed an increase of S100a9 in differentiated immune cells, it failed to detect S100a9 upregulation at the protein level within the HSPC compartment. These findings warrant further investigation of spatiotemporal S100a9-driven HSPC alterations in the BMN.

Conclusion

S100a9 plays a critical role in lung cancer progression by driving shifts in hematopoiesis that favor myeloid cell production, yet its underlying mechanisms remain poorly understood. Traditional methods like flow cytometry and ex vivo imaging offer limited insight into the complex dynamics of the BMN. Advanced imaging techniques, such as intravital imaging, offer a promising approach to uncover S100a9-driven HSPC alterations within the BMN.

Novelty

This work will offer valuable insights into the crosstalk between the BMN and lung cancer, highlighting the role of S100a9 in this process.

Impact

Targeting S100a9 signaling could represent a promising therapy for lung cancer by impacting the BMN, as well as the local tumor microenvironment.