Rational Design of Novel Allosteric EYA2 Inhibitors as Potential Therapeutics for Multiple Brain Cancers

The Eyes Absent (EYA) family of developmental proteins, often in partnership with the sine oculis (SIX) homeobox proteins, promote cancer metastasis and recurrence in numerous tumor types. In addition to being a transcriptional coactivator, EYA2 is a Tyr phosphatase that dephosphorylates H2AX which leads to repair instead of apoptosis upon DNA damage and ERβ which inhibits the anti‐tumor transcriptional activity of ERβ. The SIX members of the EYA‐SIX complex are difficult to target, therefore, we targeted the EYA2 to promote cell death and prevent cancer progression. We conducted structural optimization of a previously discovered allosteric inhibitor of EYA2, 9987, using the combination of in silico modeling, biochemical and cell‐based assays. A new series of compounds was developed with significantly improved cellular activity and physiochemical properties desirable for brain targets. Specifically, compound 2 e showed >30‐fold improvement in the medulloblastoma cell line D458, relative to 9987, while maintaining potent and selective inhibitory activity against EYA2 Tyr phosphatase activity and a good multiparameter optimization score for central nervous system drugs. Using in silico modeling and Structure Activity Relationship analysis, we have increased biochemical and cell‐based assay inhibition of the Eyes Absent 2 (EYA2) protein over 30‐fold relative to the previous lead inhibitor. We aim to increases survival for medulloblastoma and glioblastoma patients through EYA2 inhibition.