Mitochondrial-mediated apoptosis in lymphoma cells by the diterpenoid lactone andrographolide, the active component of Andrographis paniculata.
Clin Cancer Res. 2010 Aug 26. Epub 2010 Aug 26. PMID: 20798229
Medicine, Northwestern University.
PURPOSE: Andrographolide is a diterpenoid lactone isolated from Andrographis paniculata, an herbal medicine used in Asia. It has anti-inflammatory, antihypertensive, anti-viral and immuno-stimulant properties. Furthermore, it has been shown to inhibit cancer cell proliferation and induce apoptosis in leukemia and solid tumor cell lines. EXPERIMENTAL DESIGN: We studied the Burkitt p53 mutated Ramos cell line, the mantle-cell lymphoma (MCL) line Granta, the follicular lymphoma (FL) cell line HF-1 and the diffuse large B-cell lymphoma (DLBCL) cell line SUDHL4, as well as primary cells from patients with FL, DLBCL, and MCL. RESULTS: We found that andrographolide resulted in dose- and time-dependent cell death as measured by MTT. Andrographolide significantly increased reactive oxygen species (ROS) production in all cell lines. To determine mechanism of cell death, we measured apoptosis by Annexin-V/propidium iodide (PI) in the presence and absence of the antioxidant N-acetyl-L-cysteine (NAC), the glutathione-depleting agent buthionine sulfoxamine (BSO), or caspase inhibitors. We found that apoptosis was greatly enhanced by BSO, blocked by NAC, and accompanied by PARP cleavage and activation of caspases 3, 8 and 9. We measured BAX conformational change, and mitochondrial membrane potential, and using mouse embryonic fibroblast (MEF) Bax/Bak double knockouts (MEFBax-/-/Bak-/-), we found that apoptosis was mediated through mitochondrial pathways, but dependent on caspases in both cell lines and in patient samples. CONCLUSIONS: Andrographolide caused ROS-dependent apoptosis in lymphoma cell lines and in primary tumor samples, which was enhanced by depletion of GSH and inhibited by NAC or the pan-caspase inhibitor Z-VAD-FMK. Further studies of diterpenoid lactones in lymphoma are warranted.