Strong epidemiological and molecular data associate cervical cancer (CC) with high-risk human papillomavirus (HPV) infections. The carcinogenic mechanism depends mainly on the expression of E6 and E7 oncoproteins encoded by the viral genome. Using a cell-based high-throughput assay, an in-house library of compounds was screened identifying the 1,8-naphthyridone 1 that efficiently inhibited the transcription driven by the long control region of the HPV genome. A series of analogues were then synthesized, obtaining more potent derivatives able to downregulate E6 and E7 transcripts in HPV-16-positive CC CaSki cells. An unusual structural insight emerged for the C-3 position of the 1,8-naphthyridone core, where the ethyl carboxylate esters, but not the carboxylic acids, are responsible for the activity. In vitro uptake studies showed that the 3-ethyl carboxylates do not act as prodrugs. The 1,8-naphthyridones emerged as valid starting points for the development of innovative agents potentially useful for the treatment of HPV-induced CC.