A series of 2, 3-dihydroquinazolinone derivatives were synthesized, characterized and their anticancer activity was determined. Among the compounds synthesized and screened, one compound (17) showed potent anticancer activity against human head and neck squamous cell carcinoma cell line, SCC131 and was non-toxic to normal cells. The compound inhibited the growth of SCC131 cells, with an IC₅₀ of 1.75 μM, triggered apoptotic mode of cell death and caused tumor regression of SCC131 tumor xenografts in athymic mice. To decipher the target for the lead compound, a high throughput qPCR array was performed. Results showed that the compound 17, inhibited the expression of a vital transcription factor HNF4A, involved in regulation of metabolic pathways. Thus, the present work has identified a lead compound 17, with potent anticancer activity, minimal normal cell toxicity and a plausible target and hence definitely holds future prospects as an anticancer agent.