Abstract
Betulinic acid (BetA) is a plant-derived pentacyclic triterpenoid that exerts potent anti-cancer effects in vitro and in vivo. It was shown to induce apoptosis via a direct effect on mitochondria. This is largely independent of proapoptotic BAK and BAX, but can be inhibited by cyclosporin A (CsA), an inhibitor of the permeability transition (PT) pore. Here we show that blocking apoptosis with general caspase inhibitors did not prevent cell death, indicating that alternative, caspase-independent cell death pathways were activated. BetA did not induce necroptosis, but we observed a strong induction of autophagy in several cancer cell lines. Autophagy was functional as shown by enhanced flux and degradation of long-lived proteins. BetA-induced autophagy could be blocked, just like apoptosis, with CsA, suggesting that autophagy is activated as a response to the mitochondrial damage inflicted by BetA. As both a survival and cell death role have been attributed to autophagy, autophagy-deficient tumor cells and mouse embryo fibroblasts were analyzed to determine the role of autophagy in BetA-induced cell death. This clearly established BetA-induced autophagy as a survival mechanism and indicates that BetA utilizes an as yet-undefined mechanism to kill cancer cells.
Highlights
Alternative cell death pathways include necrosis, necroptosis, lysosomal membrane permeabilization (LMP) and autophagy
We have shown, in Jurkat cells, that Betulinic acid (BetA)-induced apoptosis is blocked by co-treatment with zVAD.fmk, but that this pan-caspase inhibitor does not prevent cell death as measured by propidium iodide (PI) exclusion.[4]
As this combination did not inhibit BetA-induced cell death, our data point to a cell death mechanism that is independent of caspases and necroptosis
Summary
Alternative cell death pathways include necrosis, necroptosis, lysosomal membrane permeabilization (LMP) and autophagy. Specific autophagy can include ubiquitinated proteins, peroxisomes and mitochondria.[19,20,21] Mitophagy involves the selective degradation of mitochondria and is among others used for clearance of damaged mitochondria.[17,22] Previously, BetA and a derivative of BetA, B10, have been shown to induce autophagosome formation in multiple myeloma cells and glioblastoma cells.[23,24] In these studies it was suggested that the autophagic flux was prevented, leading to an accumulation of undigested autophagosomes Both studies observed autophagy, the role of autophagy as a cell death mechanism was not addressed for BetA.[23,24]. With the use of knockout and knockdown studies of key regulators of the autophagy pathway, we demonstrate that autophagy serves as a rescue pathway and is not responsible for the cell death induced by BetA
Sign-in/Register to view highlights & summary 
Published Version (
Free)
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call