Abstract
Acute myeloid leukemia (AML) is an aggressive disease and, despite advances, its treatment remains challenging. Therefore, it remains important to identify new agents for the management of this disease. Withanolides, a group of steroidal lactones found in Solanaceae plants are of potential interest due to their reported anticancer activities in different settings. In this study we investigated the anti-proliferative effects and mode of action of Solanaceae-derived withanolides in AML cell models; these metabolites include withametelin (WTH) and Coagulansin A (CoA) isolated from Datura innoxia and Withania coagluanse, respectively. Both withanolides inhibited the proliferation of AML cells and induced cell death, with WTH being more potent than CoA in the AML models tested. Quantitative label-free proteomics and phosphoproteomics were employed to define the mechanism of action of the studied withanolides. We identified and quantified 5269 proteins and 17,482 phosphosites in cells treated with WTH, CoA or vehicle control. Withanolides modulated the expression of proteins involved in regulating key cellular processes including cell cycle, metabolism, signaling, protein degradation and gene expression. Enrichment analysis of the phosphoproteomics data against kinase substrates, kinase-kinase relationships and canonical pathways showed that the withanolides decreased the activity of kinases such as phosphoinositide 3-kinase (PI3K), protein kinase B (PKB; also known as RAC-alpha serine/threonine-protein kinase or AKT), mammalian target of rapamycin (mTOR), extracellular signal-regulated protein kinase 1 and 2 (ERK1/2) and the serine/threonine-protein kinase A-Raf (ARAF), while increasing the activation of DNA repair kinases. These results indicate that withanolide metabolites have pleiotropic effects in the modulation of oncogenic pro-survival and pro-apoptotic signaling pathways that regulate the induction of apoptosis. Withanolide mediated apoptosis was confirmed by immunoblotting showing increased expression of cleaved PARP and Caspases 3, 8 and 9 as a result of treatment. Overall, our results suggest that WTH and CoA have therapeutic potential against AML with WTH exhibiting more potent effects and should be explored further.
Highlights
Acute myeloid leukemia (AML) is the most common form of acute leukemia and a leading cause of cancer-related mortality [1]
We evaluated the pharmacological potential of withametelin (WTH) and coagulansin A (CoA) in AML cell models and investigated their mode of action using proteomic and computational biology techniques
Our results indicate that WTH and coagulansin A withanolides have potent anti-leukemic activity by inhibiting several oncogenic pathways downstream growth factor receptors, including the phosphoinositide 3-kinase (PI3K)/protein kinase B (PKB; known as AKT) and mitogen-activated protein kinase (MAPK) pro-survival pathways, leading to apoptosis
Summary
Acute myeloid leukemia (AML) is the most common form of acute leukemia and a leading cause of cancer-related mortality [1]. AML is, a highly aggressive disease, and drugs that target different pathways (e.g., FLT3 tyrosine kinas, BCL-2 and IDH1/2) are approved for the treatment of AML, with others being in different stages of development [9], overall survival rates are still low, and show complications resulting from therapy and from the emergence of resistance [10] This highlights the need for new therapeutic agents that can complement current targeted therapies to enable a sustained recovery of AML patients. Our results indicate that WTH and coagulansin A withanolides have potent anti-leukemic activity by inhibiting several oncogenic pathways downstream growth factor receptors, including the PI3K/protein kinase B (PKB; known as AKT) and MAPK pro-survival pathways, leading to apoptosis
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