2-Methoxyestradiol induces apoptosis in Ewing sarcoma cells through mitochondrial hydrogen peroxide production

Abstract

The Ewing sarcoma is the second most common bone tumor in children and young adults. Despite the advances in therapy, the 5-year survival rate for patients with metastatic disease is poor, indicating the need for alternative treatments. Here, we report that 2-methoxyestradiol (2-Me), a natural estrogen metabolite, induced a caspase-dependent apoptosis of Ewing sarcoma-derived cells independently of their p53 status. 2-Me-induced apoptosis occurred through the mitochondrial death pathway as evidenced by reduction of the mitochondrial transmembrane potential, cytochrome c release and caspase-9 activation. Treatment of cells with 2-Me resulted in generation of intracellular H 2 O 2 , which occurred earlier than caspase-9 activation. The H 2 O 2 -reducing agent Ebselen and the lipid peroxidation inhibitor vitamin E decreased both 2-Me-induced caspase-9 activation and cell death, thus providing evidence for a role of H 2 O 2 and lipid peroxides in the initiation of this process. Rotenone, an inhibitor of the mitochondrial respiratory chain, abolished both apoptosis and H 2 O 2 production, thereby identifying mitochondria as the source of H 2 O 2 . Moreover, we observed that treatment of cells with 2-Me or H 2 O 2 induced activation of the c-Jun N-terminal kinase (JNK). Overexpression of a dominantnegative mutant of JNK1 reduced 2-Me-induced apoptosis indicating that JNK participates in this process. Altogether, our results provide evidence that 2-Me triggers apoptosis of Ewing sarcoma cells through induction of a mitochondria redox-dependent mechanism and suggest that this compound or other agents that selectively increase the level of reactive oxygen species may prove useful to the development of novel strategies for treatment of Ewing tumors.

In vitro and in vivo antitumor effect of 2‐methoxyestradiol on human melanoma

“…28 In melanoma cells, we found that, similarly to some other tumor types, 37,41 mitochondria, caspase-9, and, to a lesser extent, caspase-8 were involved in the process of 2ME 2 -induced apoptosis. Moreover, the inhibition of caspases did not block apoptosis induction completely, indicating the possible involvement of other, caspaseindependent, pathways.…”

“…A mitochondria-dependent mechanism was demonstrated in leukemia, pancreatic carcinoma, Ewing sarcoma and multiple myeloma cells. 19,27,37,41 On the other hand, the use of the FIGURE 3 -Effect of 2ME 2 on the microtubular organization in HT168-M1 human melanoma cells. Cells were cultured on coverslips and treated with Taxol (100 nM, 10 min) or 2ME 2 (5 M, 1 hr).…”

2-Methoxyestradiol-induced apoptosis in human leukemia cells proceeds through a reactive oxygen species and Akt-dependent process

“…Currently, investigation of signaling events in 2-MEtreated cells has emphasized activation of stress-related pathways (e.g. JNK, H1Fa) implicated in oxidative injury (Djavaheri-Mergny et al, 2003;Mooberry, 2003). Presently, little information is available concerning the functional role of the Akt pathway in mediating 2-MEinduced lethality, particularly in malignant hematopoietic cells.…”

“…For example, in Ewing's sarcoma cells, 2-ME has been shown to induce apoptosis in association with hydrogen peroxide (H 2 O 2 ) generation and JNK activation (Djavaheri-Mergny et al, 2003). In prostate cancer cells, the PI3K pathways has been implicated in 2-ME-induced engagement of the extrinsic apoptotic pathway (Shimada et al, 2004), suggesting that certain cytoprotective signal transduction cascades might also influence 2-ME lethality.…”

Effect of AC-264, a Novel Indole Derivative, on Apoptosis in HL-60 Cells

“…[15][16] Initiation of apoptosis is controlled by regulation of the balance a These authors equally contributed to this work. between life and death signals received by the cells.…”