Mn porphyrin in combination with ascorbate acts as a pro-oxidant and mediates caspase-independent cancer cell death

Abstract

Resistance to therapy-mediated apoptosis in inflammatory breast cancer (IBC), an aggressive and distinct subtype of breast cancer, was recently attributed to increased superoxide dismutase (SOD) expression, glutathione (GSH), and decreased accumulation of reactive species. In the present study, we demonstrate the unique ability of two Mn(III) N-substituted pyridylporphyrin (MnP)-based SOD mimics (MnTE-2-PyP5+ and MnTnBuOE-2-PyP5+) to catalyze oxidation of ascorbate, leading to the production of excessive levels of peroxide and in turn cell death. The accumulation of peroxide, as a consequence of MnP + ascorbate treatment, was fully reversed by the administration of exogenous catalase showing that ROS was essential for cell death. Cell death as a consequence of the action of MnP/ascorbate corresponded with decreases in GSH levels, pro-survival signaling (pNF-κB, pERK1/2), and in expression of X-linked inhibitor of apoptosis protein (XIAP), the most potent caspase inhibitor. Although markers of classical apoptosis were observed, including PARP cleavage and Annexin V staining, administration of a pan-caspase inhibitor, QVD-OPh, did not reverse the observed cytotoxicity. MnP + ascorbate treated cells showed nuclear translocation of apoptosis inducing factor (AIF), suggesting the possibility of a mechanism of caspase-independent cell death. Pharmacological ascorbate has already shown promise in recently completed Phase I Clinical Trials, where its oxidation and subsequent peroxide formation was catalyzed by endogenous metalloproteins. The catalysis of ascorbate oxidation by an optimized metal-based catalyst (such as Mn porphyrin) carries a large therapeutic potential as a sole anticancer agent or in combination with other modalities such as radio- and chemo- therapy.

SOD Therapeutics: Latest Insights into Their Structure-Activity Relationships and Impact on the Cellular Redox-Based Signaling Pathways

“…A much higher yield of peroxide may be achieved if catalysts are optimized for ascorbate oxidation. Such are isomeric Mn(III) N-substituted pyridylporphyrins; their H 2 O 2 -producing potency has already been shown by us and others (82,220,273,299) (Fig. 11).…”

“…In subsequent reoxidation of MnP with O 2 or O 2 $ -, and oxidation of ascorbate radical to dehydroascorbate, the peroxide is produced. MnP can employ peroxide and/or GSH to inactivate nuclear factor jB (NF-jB) (82,125,127,238,240), suppress mitochondrial respiration (124) and glycolysis (70,124), or induce adaptive responses (52, 75). Such pro-oxidative actions agree well with what Forman et al recently put forward as the mechanism of action of natural antioxidants (89).…”

“…Ascorbate was added soon after MnP. The effect of MnP/ascorbate on several other cancer cell lines has also been demonstrated (82,299).…”

Simple Biological Systems for Assessing the Activity of Superoxide Dismutase Mimics

“…The E. coli-based discovery of MnP-ascorbate cytotoxicity (92) brought up the idea that the combination can be used for anticancer treatment (48,95,117,130). The therapeutic potential of pharmacological doses of ascorbate alone has already been demonstrated in several preclinical and clinical trials (46,61,83,127) and is based on the increased production of reactive species due to ascorbate oxidation, catalyzed by endogenous metalloproteins (36).…”

“…Consequently, a maximal production of H 2 O 2 and other reactive species can be achieved, causing a more efficient tumor destruction. Promising results of such combinational catalytic therapy have been reported (48,95,130). In vitro experiments have shown that some cancer cell lines are more sensitive to either cationic MnPs or Fe porphyrins (FePs) alone and to the MnP-ascorbate combination than are normal cells (121).…”