Treatment of LS174T colorectal cancer stem-like cells with n-3 PUFAs induces growth suppression through inhibition of survivin expression and induction of caspase-3 activation.

Abstract

Colorectal cancer stem cells (CCSCs) are thought to contribute to tumor initiation, progression, metastasis, chemo-resistance and therapy failure. Therefore, assessment of the effectiveness of agents with anti-proliferative activities against CCSCs is warranted. Several studies have shown that different tumorigenic steps, ranging from initiation to metastasis, can be affected by n-3 polyunsaturated fatty acids (PUFAs). Here, we evaluated the effects of the PUFA components docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), alone or in combination, on LS174T cells that serve as a model for colorectal cancer initiating cells with stem cell-like properties. LS174T cells were treated with 50, 100 and 150 μM DHA and EPA, or equal mixtures of DHA/EPA (i.e., 25/25, 50/50 and 75/75 μM), after which cell number, viability, growth inhibition, survivin expression, caspase-3 activation and apoptotic rate were evaluated. We found that treatment of LS174T cells with increasing PUFA concentrations significantly increased growth inhibition in a dose- and time-dependent manner. After a 72 h treatment with 150 μM DHA and EPA, or their combination (75/75 μM), growth rates were inhibited by 80.3 ± 5.5%, 79.3 ± 5% and 71.1 ± 1%, respectively, compared to untreated cells. We also found that treatment for 48 h with 100 μM DHA and EPA, or their combination (50/50 μM), resulted in 2.9-, 3- and 2.6-fold increases in caspase-3 activation, as well as 54, 62.4 and 100% decreases in survivin mRNA expression levels, respectively, compared to untreated cells. Low survivin mRNA levels combined with high caspase-3 activity levels were found to correlate with a higher growth inhibition in PUFA-treated cells. DHA appears to be a more potent growth inhibitor than EPA and the DHA/EPA combination. An increase in the number of apoptotic cells (early + late), ranging from 12.9 to 44.7%, was observed with increasing DHA doses. From our data we conclude that PUFAs induce growth inhibition via targeting survivin expression in LS174T cells, which serve as a model for CCSCs.