SYNERGISTIC APOPTOTIC EFFECT OF THE PLANT POLYPHENOLS CURCUMIN AND CARNOSIC ACID ON ACUTE MYELOID LEUKEMIA CELLS INVOLVES INTRACELLULAR CALCIUM AND ER STRESS

Nasma Aqaqe ¹ Stella Pesakhov ¹ Michael Kafka ⁴ George P. Studzinski ³ Daniel Fishman ² Michael Danilenko ¹

¹Clinical Biochemistry and Pharmacology, Ben-Gurion University of the Negev, Beer-Sheva
²Physiology and Cell Biology, Ben-Gurion University of the Negev, Beer-Sheva
³Pathology and Laboratory Medicine, UMD-New Jersey Medical School, Newark, NJ
⁴Laboratory of Hematology, Soroka University Medical Center, Beer-Sheva

Acute myeloid leukemia (AML) is the most common acute leukemia in adults with high mortality rate. Small molecule plant-derived agents have been widely employed for the treatment of several types of malignancies. However, it is still unclear whether active phytochemicals can be used for the management of AML. Recently we have demonstrated a synergistic cell growth arrest and robust apoptotic response in KG-1a and HL60 human AML cell lines induced by the combination of the plant polyphenols curcumin (CUR) and carnosic acid (CA) at low, non-cytotoxic concentrations of each compound [Pesakhov et al. Nutr. Cancer 2010, 62:811-824]. In this study, using specific inhibitors of caspase-8 and -9 as well as stable transfection of the dominant-negative mutants of these caspases and FADD (the death receptor adaptor protein) we revealed the involvement of both extrinsic and intrinsic apoptotic pathways in CUR/CA-induced cytotoxicity. We also found that the activation of both pathways was preceded by upregulation of endoplasmic reticulum (ER) stress markers (GRP78 and CHOP) and depletion of ER calcium stores, leading to sustained rise of cytosolic Ca²⁺ ([Ca²⁺]_cyt). CUR/CA-induced apoptosis was only partially attenuated by the ER stress blocker salubrinal but was fully prevented by lowering [Ca²⁺]_cytwith the membrane-permeable Ca²⁺ chelator BAPTA-AM or by the ER inositol trisphosphate receptor antagonist, 2-aminoethoxydiphenyl borate (2-APB), which blocks the main route of Ca²⁺ release from the ER. Importantly, CUR/CA failed to induce apoptosis in normal human peripheral blood mononuclear cells or murine bone marrow cells. These findings indicate that the selective and synergistic induction of apoptotic pathways by CUR/CA is mediated by disruption of cellular Ca²⁺ homeostasis and, partially, by the ER stress response. We suggest that this combination may have potential for the treatment and/or prevention of AML and, possibly, other malignant diseases. (Supported by the American Institute for Cancer Research grant #10A049 to G.P.S. and M.D.).