Induction of STING/IL-29 signaling in human lung adenocarcinoma/tumor-associated neutrophil cocultures

Abstract LKB1-mutant lung adenocarcinoma (LUAD), is an intractable form of lung cancer. LKB1-mutant LUAD is poor in T-cells and does not respond to checkpoint inhibitor therapy, and instead is enriched in tumor-infiltrating neutrophils (TINs). STING is a key innate sensor mediating anti-tumor responses that is inhibited in LKB1-mutant LUAD. However, the status of STING signaling and if it can be induced in TINs to antagonize tumors is unknown. Here, we developed a novel human LKB1-mutant LUAD/TIN model to tackle this question. Based on prior work from our lab (Dobosh, STAR Protoc. 2021), we grew the H1944 LKB1-mutant LUAD line at air-liquid interface on Alvetex scaffolds (Reprocell), followed by the transmigration of human blood neutrophils. The effect of the STING agonist MSA-2 was assessed in the model by a 20-plex immune mediator assay(Mesoscale). We successfully manufactured a LKB1-mutant LUAD/TIN model from H1944 cells and human blood neutrophils. MSA-2 treatment resulted in significant increases in the STING-regulated mediators IL-6, IP-10 and MIP-1beta. Most strikingly, MSA-2 treatment induced IL-29 (type III interferon) levels by 27-fold, while neither type I nor type II interferons were induced. The model showcased here mimics LKB1-mutant LUAD/TIN interplay and shows responsiveness to STING agonist treatment through induction of type III (but not type I and II) interferon and other immune mediators. Since IL-29 receptor expression is limited to epithelial and myeloid cells (including neutrophils) and IL-29 is known to mediate immunomodulatory and anti-tumor responses, future work will address if and how the large IL-29 increase observed upon MSA-2 treatment affects LUAD proliferation and TIN activation in this context.