DN-01 * BYPASSING GLIOBLASTOMA RESISTANCE TO EGFR INHIBITORS BY SYNTHETIC LETHALITY

Abstract

Clinical efficacy of EGFR inhibitors (EGFRi) in glioblastoma patients has been limited by a multitude of resistance mechanisms. We hypothesized that these mechanisms emerge upstream of the interface between EGFR signaling and fundamental cellular processes, such as DNA damage response (DDR), without altering this interface. Thus, synthetic lethal interactions targeting this interface should persist despite acquired resistance to EGFRi. To test this hypothesis, we performed an RNAi screen to identify DDR genes required for viability in glioblastoma cells dependent on EGFR signaling (through expression of the oncogenic EGFRvIII) and identified polo-like kinase 1 (PLK1). In synchronous and asynchronous populations, EGFRvIII expressing glioblastomas harbored increased levels of the p-Thr210 PLK1 (an activated form of PLK1). Moreover, EGFRvIII expression was associated with elevated PLK1 expression in the TCGA glioblastoma dataset (p = 0.044). PLK1 Inhibition (by BI2536) induced an increase in the proportion of cells that co-stained for p-Histone H3 and γH2AX foci, suggesting accumulation of mitotic DNA damage. This effect was exacerbated by EGFRvIII expression, implicating induction of mitotic DNA damage as a major contributor to the observed synthetic lethality. Using time-lapsed imaging, we showed that EGFRvIII expression induced the formation of aberrant mitosis as well as prolonged mitotic progression. Further supporting an essential role for PLK1 in suppressing DNA damage accumulation, BI2536 treatment significantly enhanced the in vitro and in vivo tumoricidal effects of the DNA damaging chemotherapy, temozolomide. Moreover, PLK1 inhibition suppressed the accumulation of pS14 Rad51 (an active form of Rad51 that is required for homologous recombination (HR)) as well as overall HR efficiency in the DR-GFP assay. The tumoricidal and TMZ effects of PLK1 inhibition were observed across a panel of eight independent EGFRvIII glioblastoma clones that had acquired distinct resistance mechanisms to EGFRi. These results suggest therapeutic opportunities in targeting the interface between oncogenic signaling and DDR.