Unique gene patterns lead to distinct functional phenotypes and chemosensitivity profiles among subclones obtained from a single glioblastoma cell line

• Single-cell-derived subclones from the U87-MG glioblastoma cell line exhibit distinct functional phenotypes, with CF5 showing enhanced proliferation and chemoresistance while G11 displays greater motility and invasion. • Transcriptomic analysis reveals significant gene expression differences among subclones, with downregulated genes enriched in extracellular matrix-related pathways, highlighting the role of integrins in tumor heterogeneity. • ITGA11 and ITGA6 are identified as key regulators of functional phenotypes and chemosensitivity in specific subclones, and their combined knockdown in parental U87 cells inhibits tumor growth and enhances chemotherapy sensitivity. • The study underscores the persistence of tumor heterogeneity even under uniform growth conditions, emphasizing the need for personalized therapeutic strategies targeting specific genetic backgrounds.