DDIT4 Downregulation by siRNA Approach Increases the Activity of Proteins Regulating Fatty Acid Metabolism upon Aspirin Treatment in Human Breast Cancer Cells

Abstract

Repositioning of aspirin for a more effective breast cancer (BC) treatment requires identification of predictive biomarkers. However, the molecular mechanism underlying the anticancer activity of aspirin remains fully undefined. Cancer cells enhance de novo fatty acid (FA) synthesis and FA oxidation to maintain a malignant phenotype, and the mechanistic target of rapamycin (mTORC1) is required for lipogenesis. We, therefore, aimed to test if the expression of mTORC1 suppressor DNA damage-inducible transcript (DDIT4) affects the activity of main enzymes in FA metabolism after aspirin treatment. MCF-7 and MDA-MB-468 human BC cell lines were transfected with siRNA to downregulate DDIT4. The expression of carnitine palmitoyltransferase 1 A (CPT1A) and serine 79-phosphorylated acetyl-CoA carboxylase 1 (ACC1) were analyzed by Western Blotting. Aspirin enhanced ACC1 phosphorylation by two-fold in MCF-7 cells and had no effect in MDA-MB-468 cells. Aspirin did not change the expression of CPT1A in either cell line. We have recently reported DDIT4 itself to be upregulated by aspirin. DDIT4 knockdown resulted in 1.5-fold decreased ACC1 phosphorylation (dephosphorylation activates the enzyme), 2-fold increased CPT1A expression in MCF-7 cells, and 2.8-fold reduced phosphorylation of ACC1 following aspirin exposure in MDA-MB-468 cells. Thus, DDIT4 downregulation raised the activity of main lipid metabolism enzymes upon aspirin exposure which is an undesired effect as FA synthesis and oxidation are linked to malignant phenotype. This finding may be clinically relevant as DDIT4 expression has been shown to vary in breast tumors. Our findings justify further, more extensive investigation of the role of DDIT4 in aspirin’s effect on fatty acid metabolism in BC cells.

Advanced Molecular Solutions for Cancer Therapy—The Good, the Bad, and the Ugly of the Biomarker Paradigm

“…The authors have contributed their best efforts, either by performing accurate experiments, reanalyzing publicly accessible genomic datasets (including The Cancer Genome Atlas (TCGA) [ 2 ]), or writing comprehensive reviews of the literature. Among the interesting proposed solutions, of note are: the targeted delivery of chimeric antigen receptor into T cells via CRISPR [ 3 ], the metabolic silencing via methionine-based amino acid restriction [ 4 ], the inhibition of ERK5 [ 5 ], the activation of ADRA2A [ 6 ], and the aspirin treatment of breast cancer [ 7 ].…”