<scp>KLF9</scp> Inhibits Brain Metastasis of Non‐Small Cell Lung Cancer by Regulating Ceramide Synthase 1 Synthesis

The regulatory mechanisms driving brain metastasis (BM) in non-small cell lung cancer (NSCLC) are complex, with Ceramide synthase 1 (Cers1) playing a critical role. However, the upstream factors controlling Cers1 expression remain unclear. Additionally, Kruppel-like factor 9 (KLF9) has been implicated as a potential tumor suppressor transcription factor (TF) in lung cancer. Our study aims to explore the physiological effects and molecular mechanisms of Cers1 upstream regulatory TFs in NSCLC BM, focusing on the link between KLF9 and Cers1. TFs regulating Cers1 expression were screened via GENECARD and UCSC databases. KLF9 expression and survival analysis in NSCLC were analyzed using TCGA and GEO data sets. Dual-luciferase assays were conducted to investigate the specific binding site of KLF9 on the Cers1 promoter. KLF9 genetic modulation impact on NSCLC BM was assessed through in vitro assays for proliferation, migration, and invasion, along with in vivo orthotopic xenograft models to evaluate tumor growth and metastasis. Our study revealed a significant downregulation of KLF9 in vitro, correlating with poor prognosis. KLF9 has a direct positive transcriptional regulation on Cers1, particularly on its promoter region (-711 nt/+22 nt). In vitro, KLF9 overexpression inhibited the ability of cells to penetrate a blood-brain barrier model. Moreover, in vivo experiments demonstrated a marked suppression of BM tumor formation upon KLF9 expression. This study reports KLF9's direct control over Cers1 expression by acting on its promoter. KLF9 demonstrates inhibitory effects on NSCLC BM, presenting a promising therapeutic avenue for NSCLC BM treatment.