Three-dimensional (3D) in vivo cell culture modeling is quickly emerging as a platform to replace two-dimensional (2D) monolayer cell culture in vitro tests. Three-dimensional tumor models mimic physiological conditions and provide valuable insight of the tumor cell response to drug discovery application. In this study, we used poly(ethylene glycol) (PEG) hydrogel microwells to generate 3D brain cancer spheroids and studied their treatment with anticancer drugs in single or combination treatment. Glioblastoma (GBM) spheroids were grown through 14 days before infecting with two drugs: Pitavastatin and Irinotecan at various concentrations. A significant cell lysis was observed and cell viability decreased to lower than 7% when drugs were combined at the concentration Pitavastatin 10 μM and Irinotecan 50 μM to infect after 7 days. These findings demonstrate a promising platform—PEG hydrogel microwells—that should be an efficient way to test the drug sensitivity in vitro as well as application in different studies.