Macrocyclization of acyclic compounds is a powerful strategy for improving inhibitor potency and selectivity. Here we have optimized 2-aminopyrimidine-based macrocycles to use these compounds as chemical tools for the ephrin kinase family. Starting with a promiscuous macrocyclic inhibitor, 6, we performed a structure-guided activity relationship and selectivity study using a panel of over 100 kinases. The crystal structure of EPHA2 in complex with the developed macrocycle 23 provided a basis for further optimization by specifically targeting the back pocket, resulting in compound 55, a potent inhibitor of EPHA2/A4 and GAK. Subsequent front-pocket derivatization resulted in an interesting in cellulo selectivity profile, favoring EPHA4 over the other ephrin receptor kinase family members. The dual EPHA2/A4 and GAK inhibitor 55 prevented dengue virus infection of Huh7 liver cells. However, further investigations are needed to determine whether this was a compound-specific effect or target-related.