Abstract
The antitumor enzyme asparaginase, which targets essential amino acid L-asparagine and catalyzes it to L-aspartic acid and ammonia, has been used for years in the treatment of acute lymphoblastic leukemia (ALL), subtypes of myeloid leukemia and T-cell lymphomas, whereas the anti-chronic myeloid leukemia (CML) effect of asparaginase and its underlying mechanism has not been completely elucidated. We have shown here that asparaginase induced significant growth inhibition and apoptosis in K562 and KU812 cells. Apart from induction of apoptosis, we reported for the first time that asparaginase induced autophagic response in K562 and KU812 cells as evidenced by the formation of autophagosome, microtubule-associated protein light chain 3 (LC3)-positive autophagy-like vacuoles, and the upregulation of LC3-II. Further study suggested that the Akt/mTOR (mammalian target of rapamycin) and Erk (extracellular signal-regulated kinase) signaling pathway were involved in asparaginase-induced autophagy in K562 cells. Moreover, blocking autophagy using pharmacological inhibitors LY294002, chloroquine (CQ) and quinacrine (QN) enhanced asparaginase-induced cell death and apoptosis, indicating the cytoprotective role of autophagy in asparaginase-treated K562 and KU812 cells. Together, these findings provide a rationale that combination of asparaginase anticancer activity and autophagic inhibition might be a promising new therapeutic strategy for CML.
Highlights
chronic myeloid leukemia (CML) is a myeloproliferative neoplasm with an incidence of 1–2 cases per 100,000 adults, and accounts for ~15% of newly diagnosed cases of leukemia in adults
Western blot analysis illustrated that the level of cleaved-caspase 3 and cleaved-PARP increased in a dose- and time-dependent manner, indicating the apoptosis was induced by asparaginase in K562 and KU812 cells (Figure 1C and Supplementary Figure 1D)
The results demonstrated that asparaginase in combination with LY294002, CQ or QN induced a higher percentage of apoptotic cells (Figure 4D, 4E and Supplementary Figures 3D, 3E, 4D, 4E) and more cleavage of caspase 3 and PARP (Figure 4F and Supplementary Figures 3F, 4F) when compared with asparaginase-treated alone, whereas cells treatment with LY294002, CQ and QN alone showed limited apoptosis-inducing effects on K562 and KU812 cells
Summary
CML is a myeloproliferative neoplasm with an incidence of 1–2 cases per 100,000 adults, and accounts for ~15% of newly diagnosed cases of leukemia in adults. Targeting amino acid metabolism has been safely and effectively employed for tumor therapy [6]. Leukemia cells, require huge amounts of asparagine to keep up with their rapid malignant growth. L-asparagine is an essential amino acid for the growth of tumor cells, whereas the growth of normal cells is not dependent on its requirement as it can be synthesized in amounts sufficient for their metabolic needs with their own enzyme L-asparagine synthetase (ASNS) [8, 9]. The presence of therapeutic asparaginase deprives tumor cells of an important growth factor by hydrolyzing L-asparagine www.impactjournals.com/oncotarget into L-aspartic acid and ammonia, afterwards tumor cells fail to survive because of their reduced ASNS levels [10]. Primarily used as a chemotherapeutic agent against ALL [11, 12], asparaginase is used in other types of leukemia such as non-Hodgkin’s lymphoma [13], subtypes of myelocytic leukemia [14] and chronic lymphocytic leukemia, sarcomas such as lymphosarcoma, reticulosarcoma and melanosarcoma [15], ovarian cancer [16] and brain cancer [6] with a potential role for its glutaminase activity [10]
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