triacsin-c and Colonic-Neoplasms

It is well documented that arachidonic acid (AA) and its metabolites are intimately linked to cancer biology. However, the downstream mechanism(s) that link AA levels to cancer cell proliferation remain to be elucidated. Initial experiments in the current study showed that exogenous AA and inhibitors of AA metabolism that lead to the accumulation of unesterified AA are cytotoxic to the colon cancer cell line, HCT-116. Additionally, exogenous AA and triacsin C, an inhibitor of AA acylation, induced apoptosis and related caspase-3 activity in a transcriptionally dependent manner. Gene array analysis revealed that both exogenous AA and triacsin C alter the expression of similar genes in HCT-116 cells. For example, both downregulate several genes with well-documented roles in cell survival and apoptotic resistance. Conversely, both upregulate genes encoding activator protein-1 (AP-1) transcription factors, which have known roles in inducing apoptosis, and genes that counteract ras (Erk/MAPK) growth signaling pathways. Real-time polymerase chain reaction and immunoblotting demonstrated that mRNA and protein levels of one of the major AP-1 transcription factors, c-Jun, is markedly elevated by exogenous AA and triacsin C. Additionally, the cyclooxygenase inhibitor, sulindac sulfide, increases c-Jun mRNA levels. Together, these studies reveal that the generation of intracellular AA and its subsequent impact on gene expression probably represents a critical step that regulates colon cancer cell proliferation.

Topics: Apoptosis; Arachidonic Acid; Cell Survival; Colonic Neoplasms; Gene Expression Regulation, Neoplastic; HCT116 Cells; Humans; Proto-Oncogene Proteins c-jun; Transcription Factor AP-1; Transcription, Genetic; Triazenes

Although cancer cells appear to maintain the machinery for intrinsic apoptosis, defects in the pathway develop during malignant transformation, preventing apoptosis from occurring. How to specifically induce apoptosis in cancer cells remains unclear.. We determined the apoptosome activity and p53 status of normal human cells and of lung, colon, stomach, brain, and breast cancer cells by measuring cytochrome c-dependent caspase activation and by DNA sequencing, respectively, and we used COMPARE analysis to identify apoptosome-specific agonists. We compared cell death, cytochrome c release, and caspase activation in NCI-H23 (lung cancer), HCT-15 (colon cancer), and SF268 (brain cancer) cells treated with Triacsin c, an inhibitor of acyl-CoA synthetase (ACS), or with vehicle. The cells were mock, transiently, or stably transfected with genes for Triacsin c-resistant ACSL5, dominant negative caspase-9, or apoptotic protease activating factor-1 knockdown. We measured ACS activity and levels of cardiolipin, a mitochondrial phospholipid, in mock and ACSL5-transduced SF268 cells. Nude mice carrying NCI-H23 xenograft tumors (n = 10) were treated with Triacsin c or vehicle, and xenograft tumor growth was assessed. Groups were compared using two-sided Student t tests.. Of 21 p53-defective tumor cell lines analyzed, 17 had higher apoptosome activity than did normal cells. Triacsin c selectively induced apoptosome-mediated death in tumor cells (caspase activity of Triacsin c-treated versus untreated SF268 cells; means = 1020% and 100%, respectively; difference = 920%, 95% CI = 900% to 940%; P<.001). Expression of ACSL5 suppressed Triacsin c-induced cytochrome c release and subsequent cell death (cell survival of Triacsin c-treated mock- versus ACSL5-transduced SF268 cells; means = 40% and 83%, respectively; difference = 43%, 95% CI = 39% to 47%; P<.001). ACS was also essential to the maintenance of cardiolipin levels. Finally, Triacsin c suppressed growth of xenograft tumors (relative tumor volume on day 21 of Triacsin c-treated versus untreated mice; means = 4.6 and 9.6, respectively; difference = 5.0, 95% CI = 2.1 to 7.9; P = .006).. Many p53-defective tumors retain activity of the apoptosome, which is therefore a potential target for cancer chemotherapy. Inhibition of ACS may be a novel strategy to induce the death of p53-defective tumor cells.

Topics: Animals; Antineoplastic Agents; Apoptosis; Apoptosis Inducing Factor; Apoptotic Protease-Activating Factor 1; Blotting, Western; Brain Neoplasms; Breast Neoplasms; Cardiolipins; Caspases; Coenzyme A Ligases; Colonic Neoplasms; Cytochromes c; Enzyme Activation; Enzyme Inhibitors; Female; Flavoproteins; Gene Transfer Techniques; Humans; Lung Neoplasms; Membrane Proteins; Mice; Mice, Nude; Mitochondria; Neoplasms, Experimental; Proteins; RNA, Small Interfering; Sequence Analysis, DNA; Stomach Neoplasms; Transfection; Transplantation, Heterologous; Triazenes; Tumor Suppressor Protein p53