INVESTIGATION OF MECHANISMS UNDERLYING SPON1+ INFLAMMATORY MONOCYTE DRIVEN LUNG CANCER

Non-small cell lung cancers (NSCLC) largely consists of lung squamous (LUSC) and lung adenocarcinoma (LUAD) and alterations in the TRP53 and PTEN tumor suppressors are common in both subtypes. While Sox2 is now considered a LUSC proto-oncogene, the relationship of p53 and Pten with Sox2 isn’t well understood. We deleted Trp53, or Pten in a Sox2High/Nkx2-1-/-/Lkb-1-/- (SNL) genetic background. In parallel, we generated a highly metastatic LUSC cell line, LN2A, that grows in C57/B6 mice. Loss of TRP53 or PTEN resulted in significantly reduced survival, increased tumor burden and a pure LUAD histology. Using single cell RNAseq, we found that LUSC tumors recruit high levels of pro-tumorigenic immune infiltrates, such as tumor infiltrating inflammatory monocytes (TIMs) and macrophages (Ccr5+ Mafb+) and have identified a new subcluster of CD38+ TIMs. Previous work in our lab identified a subset of LUSC characterized by IM infiltration that was associated with poor survival. RNA-seq data from LUSC mouse models identified Spon1, a secreted extracellular matrix glycoprotein, upregulation in TIMs as compared with healthy IMs. TCGA data revealed that patients expressing high SPON1 levels showed enrichment in collagen extracellular matrix gene-sets, which are widely associated with poor survival. In vitro treatment of NSCLC spheroids with recombinant SPON1 led to increased spheroid formation and fibrillar collagen production. Adding wildtype monocytes elicited the same response while adding Spon1-/- monocytes did not. In vivo, we observed significant reduction of tumor formation and collagen content of tumors in a Spon1-/- mouse when compared to wildtype controls. An adoptive transfer of wildtype IMs delivered into Spon1-/- mice rescued tumor formation. Spon1 is known to bind to the receptor LRP8. LRP8 loss led to reduced collagen production in NSCLC spheroids in vitro and loss of tumor formation and collagen deposition in vivo. These tumors demonstrated evidence of Spon1 induced TGFβ1 activation. Collagen in the tumor microenvironment has been of growing interest for cancer biologists. Taken together, our work elicits important mechanisms explaining how collagen deposition is triggered by inflammatory monocytes to promote tumor metastasis and disease progression and immune TME remodeling as mechanisms by which p53 and Pten promote SOX2-mediated NSCLC.