2-Cyano-3,12-dioxooleana-1,9-dien-28-imidazolide (CDDO-Im) Directly Targets Mitochondrial Glutathione to Induce Apoptosis in Pancreatic Cancer

Abstract

Surgical resection is the only curative strategy for pancreatic cancer (PC). Unfortunately, >80% of pancreatic cancer patients bear inoperable, locally advanced, chemoresistant tumors demonstrating the urgent need for development of novel therapeutic approaches to treat this disease. Here we report that the synthetic triterpenoid 2-cyano-3,12 dioxooleana-1,9 dien-28-imidazolide (CDDO-Im) antagonizes PC cell growth by inducing apoptosis at submicromolar concentrations. Notably, we demonstrate for the first time that the cytotoxicity of CDDO-Im is accompanied by the rapid and selective depletion of mitochondrial glutathione that results in accumulation of reactive oxygen species, oxidation of the cellular glutathione pool, loss of mitochondrial membrane potential, and phosphatidylserine externalization. The parent compound CDDO as well as the methyl ester of CDDO also depleted mitochondrial glutathione, demonstrating that this effect is mediated by the triterpenoid nucleus of these agents. Cotreatment with sulfhydryl nucleophiles completely prevented apoptosis and loss of viability induced by CDDO-Im, whereas alkylation of intracellular thiols by diethylmaleate or cotreatment with dithiothreitol decreased the accumulation of a biotinylated derivative of CDDO, TP-301, in PC cells, suggesting that intracellular reduced thiols are functional targets of the electrophilic triterpenoid nucleus of CDDO and its derivatives. In conclusion, our report is the first to identify mitochondrial glutathione as a target of CDDO and its derivatives and demonstrates that depletion of this antioxidant in the mitochondria is an effective strategy to induce cell death in PC cells. These results suggest that CDDO and its derivatives may offer a clinical benefit for the treatment of PC. Pancreatic cancer (PC)2 is a lethal disease characterized by a high metastatic potential and rapid progression with a median survival rate of only 24 weeks in untreated cases (1, 2). Tumor resection is still the most effective treatment for PC. However, because of local invasion and/or metastasis, only ϳ15-20% of PC patients qualify for surgical intervention. For locally advanced, unresectable, and metastatic disease, treatment is palliative at best and usually consists of 5-fluorouracil or gemcitabine alone or in combination with radiotherapy. Unfortunately, despite the moderate success of gemcitabine (2Ј,2Ј-difluorodeoxycitidine), median survival rates remain under 6 months for patients with metastatic disease (2-4). Given the poor performance of existing therapies, the development of novel therapies for the treatment of advanced metastatic pancreatic cancer is of utmost importance.Glutathione (GSH) is a critical intracellular antioxidant that scavenges reactive oxygen species (ROS) directly and indirectly by serving as cofactor for glutathione peroxidases, glutaredoxins, and lipid peroxidases (5, 6). Oxidized GSH (GSSG) must be converted back to GSH by the action of glutathione reductase, an enzyme that utilizes the pyridine nucleotide NADPH as a cofac...

1,1-Bis(3′-indolyl)-1-(p-substituted phenyl)methanes inhibit ovarian cancer cell growth through peroxisome proliferator–activated receptor–dependent and independent pathways

“…Mix et al (63) recently reported that CDDO and prostaglandin J2 inhibit matrix metalloproteinases in chondrosarcoma cells and these responses were not inhibited by the PPARg antagonist GW9662. Studies in this laboratory showed that CDDO and related methyl ester and imidazole derivatives inhibited colon cancer cell growth and induced apoptosis in colon cancer cells through both PPARg-dependent and PPARg-independent pathways (58). At lower concentrations, these compounds induced the tumor suppressor gene caveolin-1, and this response was inhibited by PPARg antagonists.…”

“…However, the antitumorigenic activity of PPARg agonists such as CDDO has been linked to both receptor-dependent and receptor-independent activation of different pathways associated with growth inhibition and cell death (53, 57 -63). For example, CDDO and related compounds activate PPARg-dependent transactivation and several responses are blocked in cells after cotreatment with PPARg antagonists (57,58). Mix et al (63) recently reported that CDDO and prostaglandin J2 inhibit matrix metalloproteinases in chondrosarcoma cells and these responses were not inhibited by the PPARg antagonist GW9662.…”

Pharmacodynamic characterization of chemopreventive triterpenoids as exceptionally potent inducers of Nrf2-regulated genes

“…CDDO-Im inhibits proliferation of human cancer cell lines, induces differentiation in leukemia cell lines, and decreases tumor burden in murine models of melanoma and leukemia (8). In addition, it induces apoptosis in pancreatic cancer cells (9). CDDO-Im is more potent than CDDO and CDDO-Me at inhibiting the proinflammatory production of nitric oxide induced by IFN-g in mouse macrophages (10).…”

The synthetic oleanane triterpenoid, CDDO-methyl ester, is a potent antiangiogenic agent

“…11 -14); (d) they inhibit activity of the transcription factor nuclear factor-nB by directly inhibiting its activating kinase, InB kinase (15,16); (e) they inhibit phosphorylation of signal transducers and activators of transcription (STAT) transcription factors (17,18), which is required for transcriptional activity of the STATs; and (f) they inhibit the ability of tumor necrosis factor-a (TNF-a) to induce expression of vascular endothelial growth factor (VEGF; ref. 19).…”