Human cytomegalovirus (HCMV) terminase complex entails a metal-dependent endonuclease at the C-terminus of pUL89 (pUL89-C). We report herein the design, synthesis, and characterization of dihydroxypyrimidine (DHP) acid (<b>14</b>), methyl ester (<b>13</b>), and amide (<b>15</b>) subtypes as inhibitors of HCMV pUL89-C. All analogs synthesized were tested in an endonuclease assay and a thermal shift assay (TSA) and subjected to molecular docking to predict binding affinity. Although analogs inhibiting pUL89-C in the sub-μM range were identified from all three subtypes, acids (<b>14</b>) showed better overall potency, substantially larger thermal shift, and considerably better docking scores than esters (<b>13</b>) and amides (<b>15</b>). In the cell-based antiviral assay, six analogs inhibited HCMV with moderate activities (EC<sub>50</sub> = 14.4-22.8 μM). The acid subtype (<b>14</b>) showed good <i>in vitro</i> ADME properties, except for poor permeability. Overall, our data support the DHP acid subtype (<b>14</b>) as a valuable scaffold for developing antivirals targeting HCMV pUL89-C.