Protopine Exerts Anti-Leukemic Effects by Promoting ROS-mediated Inhibition of Survival Signals in Acute Myeloid Leukemia Cells

The inhibition of differentiation in myeloid progenitor cells is a hallmark of acute myeloid leukemia (AML). The chemotherapy regimen for AML patients, the so-called 7+3 protocol and the survival rate, have remained unchanged for decades, leaving AML one of the most lethal diseases among adults. Differentiation therapy represents a promising approach that aims to induce the maturation of leukemic progenitor cells into non-proliferative, terminally differentiated cells, thereby directly targeting the underlying pathogenesis of the disease. The anti-leukemic effects of protopine were evaluated using flow cytometry, MTS assay, cell proliferation assay, and trypan blue exclusion test. Mitochondrial reactive oxygen species (ROS) levels were assessed using MitoSOX Red staining. Apoptosis- and cell survival-related proteins were analyzed via Western blotting. Disruption of mitochondrial membrane potential was determined using the JC-1 dye. Protopine exhibited differentiation-inducing activity in AML cells, which was closely associated with its inhibitory effects on cell proliferation and viability. Mechanistically, increased ROS levels and subsequent up-regulation of cyclin-dependent kinase inhibitors p16 and p21 played a critical role in protopine-induced differentiation. Additionally, protopine suppressed the AKT and ERK survival signaling pathways and down-regulated anti-apoptotic Bcl-2 family proteins. These changes were associated with mitochondrial membrane potential disruption and the induction of apoptosis. Protopine represents a promising candidate for differentiation-based therapy in AML.