The Eyes Absent family of protein phosphatases (EYA1-4) are aberrantly expressed and tumour-promoting across many devastating cancers of neurological origin affecting both children and adults. It has recently been demonstrated that EYA1 and EYA4 promote tumour cell survival by increasing the active pool of Polo-like kinase 1 (PLK1) molecules. This discovery provides a rationale for the therapeutic combination of EYA inhibitors with direct, ATP-competitive, PLK1 inhibitors. Here, we demonstrate potent and synergistic effects of EYA and PLK1 inhibition in cancer cell lines that overexpress EYA1 and/or EYA4, including in neuroblastoma and glioblastoma models. We identify decreases in PLK1 activity and RAD51 foci formation, and increases in mitotic arrest and cell death, as mechanistic contributors to combination sensitivity. Combined EYA and PLK1 inhibition is also effective in glioblastoma stem cell models that overexpress EYA1/EYA4 and specifically targets the cancer stem cell state. Finally, through multi-omic correlational analysis, we identify high levels of the NuRD complex and SOX9 as contributors to combination treatment sensitivity. Overall, this work identifies a novel synthetic lethal combination therapy with potential utility across a wide range of neurological cancers.