Better drugs are needed against human cytomegalovirus (HCMV), a pathogen responsible for severe diseases in immunocompromised hosts and newborn children. We investigated whether selective inhibitors of HCMV replication could be discovered by screening for compounds that disrupt the interaction between the accessory subunit of the viral DNA polymerase, UL44, and the C-terminal 22 residues of the catalytic subunit. From approximately 50,000 small molecules, we identified 5 structurally diverse compounds that not only specifically interfere with this interaction, but also with the physical and functional interaction of UL44 with full-length catalytic subunit. These five compounds also inhibited HCMV replication with sub- to low micromolar potency, and at concentrations up to 500-fold lower than those at which they exhibited cytotoxicity. These compounds represent a promising starting point for the development of anti-HCMV drugs.