The precise causes of psoriasis, a relatively common, chronic, inflammatory and hyperproliferative skin disease, are still not known, thus making it very difficult for treatment. We previously found an in vitro anti-psoriatic activity in ethanolic extract of Annona squamosa L. leaves. Epidermal growth factor receptor (EGFR) and its ligand, transforming growth factor (TGF)-α, have been demonstrated to be strikingly upregulated in active psoriatic plaques, thus implicating their functional roles in psoriatic hyperplasia. The objective of this study was to investigate the molecular effect of ethanolic extract derived from Annona squamosa L. leaves on the EGFR expression using HaCaT keratinocyte cell line as a model. Based on RT-PCR, concentrations at 3.15 and 1.575 μg/mL (IC50= 6.3 μg/mL) significantly reduced the EGFR mRNA expression (P<0.05) as compared to control (HaCaT pretreated with IFN-γ and TNF-α, 10 ng/mL for each) although all three concentrations tested showed a similar tendency of inhibitory effect. Using confocal immunofluorescence microscopy as a qualitative measurement, we also found the inhibitory effect of Annona squamosa L. leaf extract on the expression of EGFR protein. Western blot analysis confirmed that all concentrations of the extract tested markedly inhibited the expression of EGFR (P<0.05). Taken together, this suggest that the ethanolic extract of Annona squamosa leaves could exert its biological effect by suppressing the expression of EGFR biomarker, thus highlighting the potential therapeutic development of this extract for future applications.