The sequential deadhesion/readhesion of primary human astrocytes resulted in the clonal expansion of astrocyte-derived GFAP-positive cells with malignant traits, including high proliferative and migratory rates—comparable to a human GBM cell line (U87MG)—in addition to clonogenic capability and anchorage-independent growth. Here, GFAP is linked to glioblastoma.