Human cortico-vascular assembloids reveal a CELF2-AHNAK-dependent switch from neuronal to endothelial tropism in glioblastoma cells

Glioblastoma (GB) remains one of the most aggressive human cancers, driven by profound cellular plasticity and dynamic interactions with the neurovascular niche. Yet, existing preclinical models fail to reproduce the human brain-vascular interface, limiting studies of tumor-host crosstalk and invasion. To overcome this gap, we developed a human induced pluripotent stem cell (hiPSC)-derived cortico-endothelial (CO+EO) assembloid model by fusing cortical and endothelial organoids from the same genetic background. These assembloids spontaneously form branched vascular networks enriched in tight junction proteins (CLDN5, OCLN, ZO-1), thereby recapitulating a key blood-brain barrier (BBB)-like property and providing a physiologically relevant human platform for exploring glioblastoma-neurovascular interactions. Using this system, we uncover a previously unrecognized CELF2-dependent glioblastoma stem cell (GSC) tropism. CELF2-expressing GSCs preferentially infiltrate neural regions, where they trigger neuronal apoptosis and disrupt endothelial tight junction integrity, supporting an aggressive phenotype. Conversely, CELF2-deficient GSCs lose neurotropism, acquire mesenchymal features, and are redirected toward vascular compartments. This endothelial affinity requires the scaffold protein AHNAK, strongly expressed at the plasma membrane of CELF2-deficient cells and enriched at tumor-endothelial interfaces. AHNAK knockdown abolished endothelial infiltration, demonstrating its critical role in vascular tropism. Analysis of patient GB samples confirmed that CELF2-positive tumor cells are enriched in poorly vascularized, mitotically active areas and excluded from vessel-rich zones, closely paralleling assembloid findings. Transcriptomic profiling further revealed that CELF2 promotes a neuronal progenitor-like program while repressing mesenchymal and vascular-associated gene expression, thereby shaping tumor identity, invasive behavior, and tissue preference. Collectively, our study introduces CO+EO assembloids as an original human model of glioblastoma plasticity at the neurovascular interface. We identify CELF2 as a master regulator of GSC tropism and AHNAK as a mediator of vascular affinity, unveiling a novel molecular axis that governs glioblastoma invasion and highlighting new therapeutic opportunities.