Since it has been reported that Perilla leaves (Perilla frutescens) have antimutagenic, antioxidant, and anti-inflammatory properties, we hypothesized that Perilla leaves may have a potential anticancer activity. Therefore, we examined the possibility that cancer cell growth is reduced by treatment with a Perilla leaf ethanol extract (PLE) using human leukemia HL-60 cells and then investigated the mechanism of the growth inhibition. We found that PLE treatment suppressed cell viability in a dose-dependent manner. Flow cytometric analysis revealed that PLE treatment caused the appearance of a sub-G1 DNA peak and induced cell cycle arrest at the G1 phase. We detected DNA ladders in PLE-treated cells by agarose gel electrophoresis, and the cleavage of pro-caspase-3 and poly(ADP-ribose) polymerase with remarkable activation of caspase-8, -9, and -3. Western blot analysis revealed dose-dependent increases in Bax and cytochrome c in cytosol fractions and decreased Bid and pro-caspase-8 and -3 in PLE-treated cells. In addition, glucose-regulated protein 78, phosphorylated eukaryotic translation initiation factor 2 subunit $alpha$, phosphorylated c-jun N-terminal kinase, and p21 levels were increased by PLE treatment in a dose-dependent manner, whereas the p27 level was not changed. We concluded that PLE induced apoptosis through the combinations of mitochondrial, death receptor-mediated, and endoplasmic reticulum pathways and suppressed the cell proliferation via p21-mediated G1 phase arrest in HL-60 cells.