There are few effective treatments for glioblastoma. MAPK-activated protein kinase 2 (MK2) functions as an effector kinase of cell cycle checkpoints following DNA damage. MK2 inhibitors enhance efficacy of conventional chemotherapeutic agents, but their effectiveness as single agents is not known. Furthermore, MK2 has not been investigated in brain cancers because of the lack of MK2 inhibitors with blood-brain barrier permeability. We have developed novel allosteric MK2 inhibitors with drug-like properties of CNS active drugs that inhibit the targeted kinase in a non-ATP competitive manner. We examined their effectiveness in a panel of glioblastoma cell lines (including primary GBM and stem cells) and normal cell types (primary human microglia, astrocytes, neurons and macrophages). We demonstrate that these inhibitors display single-agent activity with IC50 in the one-micromolar range. The inhibitors produce cytotoxic effects in malignant cells, whereas normal cells are only sensitive to these compounds at the higher concentrations. We attribute the single agent efficacy to the high intrinsic DNA damage found in the GBM cells. This cytotoxic effect is the consequence of inhibition of G2/M-checkpoint kinase MK2, which abrogates activation of DNA damage response pathways and pushes cells prematurely into the mitosis with unrepaired DNA, resulting in cell death through an apoptotic mechanism. This sensitivity suggests that targeting MK2 offers synthetic lethality approach to GBM therapy. Investigation of combination temozolomide+MK2 inhibitor therapy will also be discussed. Collectively, our data provide first evidence for the role of MK2 in synthetic lethality and GBM pathophysiology.