Involvement of BH3-only proapoptotic proteins in mitochondrial-dependent Phenoxodiol-induced apoptosis of human melanoma cells

Abstract

Phenoxodiol is a chemically modified analogue of the plant hormone isoflavone with antitumour activities. In the present study, we have examined its ability to induce apoptosis in human melanoma cells and the mechanisms involved. Apoptosis was observed in Phenoxodiol-treated cells by using annexin V/propidium iodide staining and determining mitochondrial membrane potential. To determine which caspase pathways were involved in Phenoxodiol-induced apoptosis, studies were performed using specific caspase inhibitors. Western studies were performed to ascertain which proteins of the apoptosis cascade were affected to cause Phenoxodiol-induced apoptosis. We found that induction of apoptosis by Phenoxodiol was maximal at 48 h with a range of apoptosis of 12+/-4 to 48+/-5% in different melanoma lines. This apoptosis was mainly dependent on activation of caspase-3 and caspase-9. Apoptosis was associated with induction of changes in mitochondrial membrane potential and was inhibited by over-expression of Bcl-2. Variation in sensitivity to Phenoxodiol appeared related to events upstream of the mitochondria and the degree of conformational change in Bax. The p53-regulated BH3-only proteins (Bad, PUMA and Noxa) were increased in the sensitive, but not in the resistant lines, whereas Bim was increased in all the lines tested. Bim appeared, however, to be partially involved because reduction of Bim by RNA interference resulted in decreased levels of apoptosis. Together, these studies suggest that Phenoxodiol induces apoptosis of melanoma cells by induction of p53-dependent BH3 proteins (Bad, PUMA and Noxa) and the p53-independent Bim protein, resulting in activation of Bax and its downstream events.

Citrinin induces apoptosis via a mitochondria-dependent pathway and inhibition of survival signals in embryonic stem cells, and causes developmental injury in blastocysts

“…As shown in Figure 3(A), 10-30 µM CTN directly induced a ∼ 7-18-fold increase in the intracellular ROS content of ESC-B5 cells, compared with untreated control cells. Since the protein expression ratio of Bax versus Bcl-2 is relevant to the mitochondria-dependent apoptotic pathway [30,31], with high and low Bax/Bcl-2 ratios associated with lower and higher apoptotic thresholds respectively, I determined whether CTN induced apoptosis via modulation of the Bax/Bcl-2 ratio. Immunoblotting revealed that CTN treatment of ESC-B5 cells caused the Bax and Bcl-2 expression levels to increase and decrease respectively ( Figure 3B), and densitometric analysis confirmed that CTN-treated ESC-B5 cells had a higher Bax/Bcl-2 ratio, which is indicative of apoptosis ( Figure 3C).…”

The anti-cancer drug, phenoxodiol, kills primary myeloid and lymphoid leukemic blasts and rapidly proliferating T cells

“…1,2 Phenoxodiol has been referred to as a pan-cancer drug, 3 causing apoptosis in cancer cell lines and murine cancer models via both intrinsic and extrinsic pathways. [4][5][6][7][8][9][10][11][12][13][14] Plasma membrane electron transport (PMET) has been shown to be a target of phenoxodiol in several studies with cancer cell lines. 3,13,15 PMET consists of an outer membrane (surface) NADH-oxidoreductase, redox-recycling plasma membrane ubiquinone and an inner membrane NADH-oxidoreductase.…”

The BH3-only protein Noxa is stimulated during apoptosis of chronic lymphocytic leukemia cells triggered by M2YN, a new plant-derived extract

“…It is possible that a number of flavonoids (quercetin, apigenin, myricetin) known for their capacity to inhibit proteasome activity (28) would be able to induce apoptosis by enhancing Noxa expression. In addition, Noxa upregulation was reported to be associated with the proapoptotic effects of several polyphenols such as resveratrol, phenoxodiol or tetramethoxystilbene as well as the flavonoid isoliquiritigenin (37)(38)(39)(40). Therefore, such a family of natural specific Noxa upregulators might be useful for the development of novel combining therapeutic protocols in CLL.…”