phenoxodiol and sphingosine-1-phosphate

A major limitation in the treatment of cancers is the prevalence of chemoresistant tumors. Chemotherapy agents induce cell death by activating apoptosis. However, most cancer cells express high levels of antiapoptotic proteins and, hence, are chemoresistant. Phenoxodiol, a novel isoflavone derivative, has been shown to induce apoptosis both in vitro and in vivo, even in chemoresistant cancer cells. In addition, phenoxodiol has been shown to chemosensitize resistant cancer cells to commonly used chemotherapy agents, such as carboplatin and paclitaxel. This review will discuss the characterization of phenoxodiol's molecular mechanism and its current state in the clinic.

Topics: Angiogenesis Inhibitors; Antineoplastic Agents; Apoptosis; Cell Cycle; Cell Line, Tumor; Clinical Trials as Topic; Drug Resistance, Neoplasm; Drug Screening Assays, Antitumor; Enzyme Inhibitors; Female; Humans; Isoflavones; Lysophospholipids; Male; Models, Biological; Neoplasm Proteins; Neoplasms; Neovascularization, Pathologic; Sphingosine; Topoisomerase II Inhibitors

Phenoxodiol, a synthetic analog of the plant isoflavone genistein, represents a new generation of oncology drugs acting as multiple signal transduction regulators. Phenoxodiol exerts its effect mainly by the induction of apoptosis through multiple mechanisms resulting in degradation of antiapoptotic proteins, with increased levels being linked to chemoresistance in tumor cells. Preclinical studies with this agent showed promising anticancer activity leading to a potential role in the treatment of a wide range of solid and hematologic cancers. Early clinical studies, especially in chemotherapy-resistant ovarian cancer, showed minimal toxicity with minor antitumor activity. Hormone-refractory prostate cancer is another promising area in which phenoxodiol is being actively tested. Studies are ongoing to define the optimal use of this novel anticancer agent.

Topics: Animals; Antineoplastic Agents; Apoptosis; CASP8 and FADD-Like Apoptosis Regulating Protein; Caspases; Cell Line, Tumor; Clinical Trials as Topic; Cyclin-Dependent Kinase Inhibitor p21; Down-Regulation; Female; Humans; Intracellular Signaling Peptides and Proteins; Isoflavones; Lysophospholipids; Male; Neoplasms; Sphingosine; Topoisomerase II Inhibitors; X-Linked Inhibitor of Apoptosis Protein

Constituents and inhibitors of intermediary metabolism resulting in alterations in levels of cytosolic NADH, stimulation of sphingomyelinase and inhibition of sphingosine kinase were evaluated for effects on growth inhibition and induction of apoptosis by the ENOX2 inhibitors EGCG, the principal catechin of green tea, and phenoxodiol, a naturally occurring isoflavone.. Responses were evaluated from dose-response curves of the metabolites and metabolic inhibitors in which growth of HeLa cells, apoptosis based on DAPI fluorescence and cytosolic NADH levels were correlated with sphingomyelinase and spingosine kinase activities and levels of ceramide and sphingosine1-phosphate.. Growth inhibition correlated with the modulation of localized cytosolic NADH levels by metabolites and metabolic inhibitors, the response of sphingomyelinase and sphingosine kinase located near the inner surface of the plasma membrane, and apoptosis.. Based on findings with metabolites, we conclude that apoptosis in cancer cell lines caused by ENOX2 inhibitors such as EGCG and phenoxodiol is a direct response to elevated levels of cytosolic NADH that result from ENOX2 inhibition.. The findings help to explain why increased NADH levels resulting from ENOX2 inhibition result in decreased prosurvival sphingosine-1-phosphate and increased proapoptotic ceramide, both of which may be important to initiation of the ENOX2 inhibitor-induced apoptotic cascade.

Topics: Anticarcinogenic Agents; Apoptosis; Catechin; Cell Membrane; Cell Survival; Enzyme Inhibitors; HeLa Cells; Humans; Isoflavones; Lysophospholipids; NADH, NADPH Oxidoreductases; NADP; Sphingosine

ENOX2 (tNOX), a tumor-associated cell surface ubiquinol (NADH) oxidase, functions as an alternative terminal oxidase for plasma membrane electron transport. Ubiquitous in all cancer cell lines studied thus far, ENOX2 expression correlates with the abnormal growth and division associated with the malignant phenotype. ENOX2 has been proposed as the cellular target for various quinone site inhibitors that demonstrate anticancer activity such as the green tea constituent epigallocatechin-3-gallate (EGCg) and the isoflavone phenoxodiol (PXD). Here we present a possible mechanism that explains how these substances result in apoptosis in cancer cells by ENOX2-mediated alterations of cytosolic amounts of NAD(+) and NADH. When ENOX2 is inhibited, plasma membrane electron transport is diminished, and cytosolic NADH accumulates. We show in HeLa cells that NADH levels modulate the activities of two pivotal enzymes of sphingolipid metabolism: sphingosine kinase 1 (SK1) and neutral sphingomyelinase (nSMase). Their respective products sphingosine 1-phosphate (S1P) and ceramide (Cer) are key determinants of cell fate. S1P promotes cell survival and Cer promotes apoptosis. Using plasma membranes isolated from cervical adenocarcinoma (HeLa) cells as well as purified proteins of both bacterial and human origin, we demonstrate that NADH inhibits SK1 and stimulates nSMase, while NAD(+) inhibits nSMase and has no effect on SK1. Additionally, intact HeLa cells treated with ENOX2 inhibitors exhibit an increase in Cer and a decrease in S1P. Treatments that stimulate cytosolic NADH production potentiate the antiproliferative effects of ENOX2 inhibitors while those that attenuate NADH production or stimulate plasma membrane electron transport confer a survival advantage.

Topics: Anticarcinogenic Agents; Apoptosis; Catechin; Cell Line; Cell Line, Tumor; Cell Membrane; Cell Proliferation; Cell Survival; Ceramides; Chromatography, Thin Layer; Cytosol; Dose-Response Relationship, Drug; HeLa Cells; Humans; Isoflavones; Lysophospholipids; Multienzyme Complexes; NAD; NADH, NADPH Oxidoreductases; Phosphotransferases (Alcohol Group Acceptor); Sphingolipids; Sphingomyelin Phosphodiesterase; Sphingosine

Phenoxodiol, an ENOX2 inhibitor, alters cytosolic NADH levels to initiate a regulatory cascade linking sphingolipid metabolism and the PI3K/Akt pathway to programmed cell death. Specifically, the pyridine nucleotide products of plasma membrane redox, NAD+ and NADH, directly modulate in a recriprocal manner two key plasma membrane enzymes. NADH stimulation of sphingomyelinase and NADH inhibition of sphingosine kinase potentially lead to G1 arrest (increase in ceramide) and apoptosis (loss of sphingosine-1-phosphate). The findings link plasma membrane electron transport and the anticancer action of several clinically-relevant anticancer agents targeted to ENOX2 such as phenoxodiol. Growth inhibition by phenoxodiol is unaffected by inhibitors of protein or mRNA synthesis. Findings with okadiaic acid, an inhibitor of serine/threonine phosphatases, suggest that hyperphosphorylation of intracellular substrates does not affect the action of phenoxodiol on ENOX2. Our findings and those of others are consistent with operation of the FAS signaling pathway of apoptosis and its suppression by sphingosine-1-phosphate. The prevailing hypothesis is that products of Akt activation, c-FLIP and XIAP, which exhibit anticaspase activities to block FAS signaling when sphingosine-1-phospate is elevated, are down regulated to permit apoptosis when sphingosine-1-phosphate is decreased by inhibition of sphingosine kinase under conditions of elevated cytosolic NADH associated with anticancer drug inhibition of ENOX2.

Topics: Blotting, Western; Cell Survival; Ceramides; Enzyme Inhibitors; HeLa Cells; Humans; Isoflavones; Lysophospholipids; Models, Biological; NADH, NADPH Oxidoreductases; Okadaic Acid; Phosphoproteins; Signal Transduction; Sphingolipids; Sphingosine

To elucidate possible biochemical links between growth arrest from antiproliferative chemotherapeutic agents and apoptosis, our work has focused on agents (EGCg, capsaicin, cis platinum, adriamycin, anti-tumor sulfonylureas, phenoxodiol) that target tNOX. tNOX is a cancer-specific cell surface NADH oxidase (ECTO-NOX protein), that functions in cancer cells as the terminal oxidase for plasma membrane electron transport. When tNOX is active, coenzyme Q(10) (ubiquinone) of the plasma membrane is oxidized and NADH is oxidized at the cytosolic surface of the plasma membrane. However, when tNOX is inhibited and plasma membrane electron transport is diminished, both reduced coenzyme Q(10) (ubiquinol) and NADH would be expected to accumulate. To relate inhibition of plasma membrane redox to increased ceramide levels and arrest of cell proliferation in G(1) and apoptosis, we show that neutral sphingomyelinase, a major contributor to plasma membrane ceramide, is inhibited by reduced glutathione and ubiquinone. Ubiquinol is without effect or stimulates. In contrast, sphingosine kinase, which generates anti-apoptotic sphingosine-1-phosphate, is stimulated by ubiquinone but inhibited by ubiquinol and NADH. Thus, the quinone and pyridine nucleotide products of plasma membrane redox, ubiquinone and ubiquinol, as well as NAD(+) and NADH, may directly modulate in a reciprocal manner two key plasma membrane enzymes, sphingomyelinase and sphingosine kinase, potentially leading to G(1) arrest (increase in ceramide) and apoptosis (loss of sphingosine-1-phosphate). As such, the findings provide potential links between coenzyme Q(10)-mediated plasma membrane electron transport and the anticancer action of several clinically-relevant anticancer agents.

Topics: Alkyl and Aryl Transferases; Apoptosis; Cell Membrane; Ceramides; Electron Transport; G1 Phase; HeLa Cells; Humans; Isoflavones; Lysophospholipids; NAD; NADH, NADPH Oxidoreductases; Phosphotransferases (Alcohol Group Acceptor); Sphingomyelin Phosphodiesterase; Sphingosine; Ubiquinone