We have previously reported that insulin-like growth factor-I (IGF-I) supports growth and survival of mouse and human medulloblastoma cell lines, and that IGF-I receptor (IGF-IR) is constitutively phosphorylated in human medulloblastoma clinical samples. Here, we demonstrate that a specific inhibitor of insulin-like growth factor-I receptor (IGF-IR), NVP-AEW541, attenuated growth and survival of mouse (BsB8) and human (D384, Daoy) medulloblastoma cell lines. Cell cycle analysis demonstrated that G1 arrest and apoptosis contributed to the action of NVP-AEW54. Interestingly, very aggressive BsB8 cells, which derive from cerebellar tumors of transgenic mice expressing viral oncoprotein (large T-antigen from human polyomavirus JC) became much more sensitive to NVP-AEW541 when exposed to anchorage-independent culture conditions. This high sensitivity to NVP-AEW54 in suspension was accompanied by the loss of GSK-3β constitutive phosphorylation and was independent from T-antigen-mediated cellular events (Supplementary Materials). BsB8 cells were partially rescued from NVP-AEW541 by GSK3β inhibitor, lithium chloride and were sensitized by GSK3β activator, sodium nitroprusside (SNP). Importantly, human medulloblastoma cells, D384, which demonstrated partial resistance to NVP-AEW541 in suspension cultures, become much more sensitive following SNP-mediated GSK3β dephosphorylation (activation). Our results indicate that hypersensitivity of medulloblastoma cells in anchorage-independence is linked to GSK-3β activity and suggest that pharmacological intervention against IGF-IR with simultaneous activation of GSK3β could be highly effective against medulloblastomas, which have intrinsic ability of disseminating the CNS via cerebrospinal fluid.