Glioblastoma-Neuroblastoma co-cultured multicellular spheroid model for anti-cancer drug screening

Preclinical drug screening often relies on 2D mono-cultures and animal models, which fail to replicate the complexity of the human brain tumor microenvironment. This study presents a novel 3D co-culture spheroid model combining glioblastoma (U87-MG) and neuroblastoma (SH-SY5Y) cells to better mimic the cellular and molecular dynamics of brain tumors. Using the liquid overlay technique, reproducible and scalable spheroids were developed and characterized for growth, morphology, and drug sensitivity to temozolomide (TMZ), an FDA-approved brain cancer drug. Our findings demonstrate that glioblastoma spheroids thrive under hypoxic conditions and show significant resistance to TMZ, maintaining [~]88% viability even at 1000 {micro}M. In contrast, neuroblastoma spheroids exhibit marked sensitivity, with viability reducing to [~]67% at the same concentration. Co-cultured spheroids displayed intermediate drug sensitivity ([~]68% viability), highlighting the influence of neuroblastoma cells on glioblastomas drug response. Notably, neuroblastoma cells in the co-culture model exhibited neurite-like morphologies, suggesting differentiation and enhanced tumor-tumor interactions. This model addresses key limitations of existing preclinical systems by bridging the gap between simplistic 2D cultures and time-intensive organoids. It provides a biologically relevant and time-efficient platform for investigating tumor-stroma interactions, drug responses, and the role of neuron-glia dynamics in glioblastoma progression. Therefore, this co-culture spheroid system represents a critical step forward in developing more effective therapeutic strategies for brain tumors.