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Dichloroacetate and Salinomycin Exert a Synergistic Cytotoxic Effect in Colorectal Cancer Cell Lines.

ABSTRACT: In the present study, we examined a hypothesis that dichloroacetate, a metabolic inhibitor, might efficiently potentiate the cytotoxic effect of salinomycin, an antibiotic ionophore, on two human colorectal cancer derived cell lines DLD-1 and HCT116. First, we performed a series of dose response experiments in the 2D cell culture by applying mono- and combination therapy and by using the Chou-Talalay method found that salinomycin in combination with dichloroacetate acted synergistically in both cell lines. Secondly, in order to recapitulate the in vivo tumor architecture, we tested various doses of these compounds, alone and in combination, in the 3D multicellular spheroid culture. The effect of combination of dichloracetate and salinomycin on multicellular spheroid size was stronger than the sum of both monotherapies, particularly in HCT116 cells. Further, we demonstrate that the synergistic effect of compounds may be related to the inhibitory effect of dichloroacetate on multidrug resistance proteins, and in contrast, it is not related to dichloroacetate-induced reduction of intracellular pH. Our findings indicate that the combination therapy of salinomycin and dichloroacetate could be an effective option for colorectal cancer treatment and provide the first mechanistic explanation of the synergistic action of these compounds.

SUBMITTER: Skeberdyte A

PROVIDER: S-EPMC6288092 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundTumors are heterogeneous in nature, composed of different cell populations with various mutations and/or phenotypes. Using a single drug to encounter cancer progression is generally ineffective. To improve the treatment outcome, multiple drugs of distinctive mechanisms but complementary anticancer activities (combination therapy) are often used to enhance antitumor efficacy and minimize the risk of acquiring drug resistance. We report here the synergistic effects of salinomycin (a polyether antibiotic) and dasatinib (a Src kinase inhibitor).MethodsFunctionally, both drugs induce cell cycle arrest, intracellular reactive oxygen species (iROS) production, and apoptosis. We rationalized that an overlapping of the drug activities should offer an enhanced anticancer effect, either through vertical inhibition of the Src-STAT3 axis or horizontal suppression of multiple pathways. We determined the toxicity induced by the drug combination and studied the kinetics of iROS production by fluorescence imaging and flow cytometry. Using genomic and proteomic techniques, including RNA-sequencing (RNA-seq), reverse transcription-quantitative polymerase chain reaction (RT-qPCR), and Western Blot, we subsequently identified the responsible pathways that contributed to the synergistic effects of the drug combination.ResultsCompared to either drug alone, the drug combination showed enhanced potency against MDA-MB-468, MDA-MB-231, and MCF-7 human breast cancer (BC) cell lines and tumor spheroids. The drug combination induces both iROS generation and apoptosis in a time-dependent manner, following a 2-step kinetic profile. RNA-seq data revealed that the drug combination exhibited synergism through horizontal suppression of multiple pathways, possibly through a promotion of cell cycle arrest at the G1/S phase via the estrogen-mediated S-phase entry pathway, and partially via the BRCA1 and DNA damage response pathway.ConclusionTranscriptomic analyses revealed for the first time, that the estrogen-mediated S-phase entry pathway partially contributed to the synergistic effect of the drug combination. More importantly, our studies led to the discoveries of new potential therapeutic targets, such as E2F2, as well as a novel drug-induced targeting of estrogen receptor β (ESR2) approach for triple-negative breast cancer treatment, currently lacking of targeted therapies.

Project description:Introduction: Colorectal cancer remains the third most common cancer diagnosis and fourth leading cause of cancer-related mortality worldwide. Purified cannabinoids have been reported to prevent proliferation, metastasis, and induce apoptosis in a variety of cancer cell types. However, the active compounds from Cannabis sativa flowers and their interactions remain elusive. Research Aim: This study was aimed to specify the cytotoxic effect of C. sativa-derived extracts on colon cancer cells and adenomatous polyps by identification of active compound(s) and characterization of their interaction. Materials and Methods: Ethanol extracts of C. sativa were analyzed by high-performance liquid chromatography and gas chromatograph/mass spectrometry and their cytotoxic activity was determined using alamarBlue-based assay (Resazurin) and tetrazolium dye-based assay (XTT) on cancer and normal colon cell lines and on dysplastic adenomatous polyp cells. Annexin V Assay and fluorescence-activated cell sorting (FACS) were used to determine apoptosis and cell cycle, and RNA sequencing was used to determine gene expression. Results: The unheated cannabis extracts (C2F), fraction 7 (F7), and fraction 3 (F3) had cytotoxic activity on colon cancer cells, but reduced activity on normal colon cell lines. Moreover, synergistic interaction was found between F7 and F3 and the latter contains mainly cannabigerolic acid. The F7 and F7+F3 cytotoxic activity involved cell apoptosis and cell cycle arrest in S or G0/G1 phases, respectively. RNA profiling identified 2283 differentially expressed genes in F7+F3 treatment, among them genes related to the Wnt signaling pathway and apoptosis-related genes. Moreover, F7, F3, and F7+F3 treatments induced cell death of polyp cells. Conclusions:C. sativa compounds interact synergistically for cytotoxic activity against colon cancer cells and induce cell cycle arrest, apoptotic cell death, and distinct gene expression. F3, F7, and F7+F3 are also active on adenomatous polyps, suggesting possible future therapeutic value.

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Dichloroacetate and Salinomycin Exert a Synergistic Cytotoxic Effect in Colorectal Cancer Cell Lines.

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