Anti-myeloma Effect of Xanthohumol on Multiple Myeloma Cells

Introduction: Multiple myeloma (MM) is characterized as a type of blood cell cancer, specifically affecting plasma cells within bone marrow. This disease results in approximately 13,000 deaths each year in the United States. The over-accumulation of malignant plasma cells crowds out normal, healthy blood and immune cells, leaving the host potentially immunocompromised and prone to weakness, fatigue, weight loss and broken bones. Xanthohumol (XN), a prenylated flavonoid derived from the common hop plant Humulus lupulus, has been shown with profound anti-obesity, anti-inflammatory and anti-cancer effects. However, its effect on MM was rarely evaluated. The purpose of our study was to examine not only the cytotoxic and apoptogenic effects of XN on human MM cells, but also whether the 5’ AMP-activated protein kinase (AMPK) pathway plays a role in mediating those effects in a simulated bone marrow microenvironment. Methods: Cytotoxicity of XN on RPMI-8226 and U266 MM cell lines was investigated using PrestoBlue™ Cell Viability assay. The apoptotic effects of XN were investigated on the RPMI-8226 cell after incubation for 24 hours and measured through flow cytometry and western blotting with pro-apoptotic protein markers such as caspase 3, caspase 8, caspase 9, and poly-(­­ADP-ribose) polymerase (PARP). Involvement of the AMPK pathway was presently evaluated through western blot analysis with consideration for future experimentation utilizing AMPK-specific activators and inhibitors. To simulate a bone marrow microenvironment, a TransWell™ plate was used to assess the cell viability of MM cells when co-cultured with human bone marrow stromal cells HS-5. Results: Our results show that XN decreased MM cell viability in a time (24, 48, 72 hours) and dose (1, 5, 10, 20 and 30 μM) -dependent manner in RPMI-8226 and U266 MM cell lines. The apoptogenic effect of XN was confirmed by flow cytometry analysis and our data demonstrated that, as the concentration of XN administered increased, the number of apoptotic MM cells increased in a simulated bone marrow microenvironment. Protein expression of apoptosis-related factors including caspase 3, caspase 8, caspase 9, poly-(­­ADP-ribose) polymerase (PARP) and AMPK was examined and showed that the amounts of cleaved caspase 3, PARP, and AMPK were increased corresponding to an increase in XN. Conclusions:­ Our data suggests that XN inhibits MM cell proliferation through induction of apoptosis and possibly AMPK activation