LNX1 Modulates Notch1 Signaling to Promote Expansion of the Glioma Stem Cell Population During Temozolomide Therapy in Glioblastoma

Abstract

Glioblastoma (GBM) is the most common primary brain malignancy in adults, with a 100% recurrence rate and a 21-month median survival. Our lab and others have shown that GBM contains a subpopulation of glioma stem cells (GSCs) that expand with chemotherapy, and that may contribute to therapeutic resistance and recurrence in GBM. To investigate the mechanism behind this expansion, we applied gene set expression analysis (GSEA) to patient-derived xenograft (PDX) cells in response to temozolomide (TMZ), the most commonly used chemotherapy against GBM. Results showed significant enrichment of cancer stem cell and cell cycle pathways (FDR<0.25). The ligand of numb protein 1 (LNX1), a known regulator of Notch signaling by targeting negative regulator Numb, is strongly upregulated after TMZ therapy (p<0.0001) and is negatively correlated with survival of GBM patients. LNX1 is also upregulated after TMZ therapy in multiple PDX lines with concomitant downregulations in Numb and upregulations in intracellular Notch1 (NICD). Overexpression of LNX1 results in Notch1 signaling activation and increased CSC populations. In contrast, knocking down LNX1 reverses these changes, causing a significant downregulation of NICD, eliminating induction of functional stemness after TMZ therapy, and resulting in more prolonged median survival in a mouse model. Our data indicate that removing LNX1 activity results in a less aggressive and more chemo-sensitive tumor. Based on this, we propose that during anti-GBM chemotherapy, LNX1-regulated Notch1 signaling promotes stemness and contributes to therapeutic resistance.