Exploring glioblastoma microenvironment using organoids: opportunities, limitations, and emerging concepts

Glioblastoma (GBM) remains the most lethal primary brain tumor in adults. These aggressive tumors evolve as dynamic, spatially organized ecosystems in which tumor cells continuously interact with the surrounding brain parenchyma and systemic environment. These reciprocal interactions actively drive invasion, therapeutic resistance, and ultimately, inevitable recurrence. Modelling this level of complexity has long required significant compromise. GBM organoids have emerged as a promising intermediate platform, bridging the gap between costly, low-throughput animal models and overly simplistic two-dimensional in vitro cultures. In this review, we summarize the diverse protocols currently used for GBM organoid derivation and long-term maintenance, focusing on the recapitulation of microenvironmental traits. We further discuss how these systems enable the investigation of tumor niche architecture and dynamic crosstalk with key components of the tumor microenvironment, including neural and immune elements, vascular-associated signals, and extracellular matrix cues. Although the inherent limitations of ex vivo systems must be carefully considered, increasingly advanced and well-designed protocols will enable robust interrogation of interactions within the GBM ecosystem and provide powerful platforms for therapeutic testing.