# Lung Adenocarcinoma Promotes NETosis via the NPM1–TNFAIP6–CD44–SPP1 Axis

## TL;DR

This study identifies a new pathway in lung adenocarcinoma that promotes cancer-linked NET formation, offering a potential target for treatment.

## Contribution

The novel NPM1–TNFAIP6–CD44–SPP1 axis is identified as a key driver of NETosis in lung adenocarcinoma.

## Key findings

- TNFAIP6 from lung adenocarcinoma induces NETosis by interacting with CD44 and increasing SPP1 levels.
- NPM1 enhances TNFAIP6 transcription by binding to its promoter region.
- The NPM1–TNFAIP6–CD44–SPP1 axis is critical for tumor-promoting NET formation.

## Abstract

Neutrophil extracellular traps (NETs) promote cancer progression, but the regulatory mechanism underlying their formation remains unclear. This study shows that tumor necrosis factor alpha-inducible protein 6 (TNFAIP6) derived from lung adenocarcinoma induces NET formation (NETosis) in vitro and in vivo. TNFAIP6 interacts with CD44, leading to increased extracellular secreted phosphoprotein 1 (SPP1) levels and subsequent NETosis. Additionally, nucleophosmin 1 (NPM1) upregulates the transcriptional activation of TNFAIP6 by interacting with the −2000 to −1700 bp region of its promoter. Together, these results indicate that the NPM1–TNFAIP6–CD44–SPP1 axis is a critical regulator of NETosis in lung adenocarcinoma, highlighting this pathway as a potential therapeutic target for suppressing tumor progression.

Background: While neutrophil extracellular traps (NETs) have been shown to contribute to cancer progression, including that of lung adenocarcinoma, the mechanisms underlying NET formation within the tumor immune microenvironment (TIME) remain incompletely understood. Notably, neutrophil infiltration has been strongly linked to tumor necrosis factor alpha-inducible protein 6 (TNFAIP6) expression. Methods: In vitro and in vivo experiments were performed. DNA pulldown coupled with mass spectrometry, bioinformatics analyses, immunohistochemistry, and dual-luciferase reporter assays were conducted. DNA pulldown Western blotting, chromatin immunoprecipitation–quantitative PCR, and dual-luciferase reporter assays using truncated promoter constructs were also employed. Results: TNFAIP6 expressed by lung adenocarcinoma cells was shown to induce NET formation (a form of programmed cell death called NETosis). Mechanistically, TNFAIP6 interacted with CD44 in lung adenocarcinoma cells, leading to increased extracellular availability of secreted phosphoprotein 1 (SPP1) within the TIME and the subsequent promotion of NETosis. Additionally, nucleophosmin 1 (NPM1) significantly enhances the transcriptional activation of TNFAIP6 and associates with the −2000 to −1700 bp region of its promoter. Conclusions: These findings delineate a regulatory model in which lung adenocarcinoma cells directly stimulate NETosis through the NPM1–TNFAIP6–CD44–SPP1 axis, suggesting that therapeutic targeting of this pathway may attenuate tumor progression.