Time-dependent cytotoxicity of dichloroacetate and metformin against Lewis lung carcinoma

Abstract

Summary. The use of inhibitors of energy metabolism of malignant cells is a new promising trend in the treatment of cancer patients, based on one of the unique features of the malignant cell, namely the dominance of glycolysis over oxidative phosphorylation, even in the presence of oxygen, the so-called Warburg effect. Aim: To study time-dependent cytotoxicity of sodium dichloroacetate (DCA) and metformin (MTF) against metastatic tumor cells and action of these agents on tumor cell migration. Materials and Methods: In the study low metastatic LLC/R9 variant of Lewis lung carcinoma was used. The number of living cells in the cytotoxic test was evaluated using sulforhodamine B after 1, 2 and 3 days of cell incubation in vitro. The parameters of the sensitivity of tumor cells to the action of DCA and MTF in vitro were calculated using nonlinear and linear regression of experimental data. The effect of DCA and MTF on cellular motility in vitro was evaluated using a Boyden chamber by calculation of the number of cells that migrated to the bottom side of the filter within 3 days of incubation. The statistical analysis of the data was carried out with the use of descriptive methods, Student’s t-criterion, nonlinear, and linear regression analysis. Results: IC50 of DCA was found to be equal to 50.8 ± 7.6 mM at the first day of incubation with LLC/R9 cells and decreased by 1.9 (p < 0.05) and 2.1 (p < 0.05) times at the 2nd and 3rd days, respectively. Despite the almost identical ІC50 at the 2nd and 3rd days, an increase in the incubation period of cells with DCA for up to 3 days increased the C0 parameter, which reflects the maximum concentration of the agent that does not exhibit cytotoxic effects, by 93% (p < 0.05) compared to this at the 2nd day (16.2 ± 1.4 mM vs 8.4 ± 1.0 mM, correspondently). Unlike DCA, the LLC/R9 cell population was not homogeneous by the sensitivity to the action of MTF; at least at the 3rd day, an appearance of MTF-resistant subpopulation was observed, accounting for 35% of all cells. IC50 of MTF was equal to 12.1 ± 0.6 mM, and unlike DCA, this index progressively decreased at the 2nd and 3rd days by 1.4 (p < 0.05) and 9.3 times (p < 0.05) respectively. Action of DCA at a concentration of 25 mM alone and in combination with MTF at the concentrations of 0.1 mM and 0.7 mM resulted in an increase in cell migration by 65% (p < 0.05), 63% (p < 0.05) and 78.5% (p < 0.05), respectively. There was no significant effect of MTF on the tumor cell migration. Conclusions: The sensitivity of the metastatic Lewis lung carcinoma cells to the action of the modifiers of the energy metabolism increased significantly with an increase in the incubation period, apparently, primarily due to the shortage of nutrient substrates and, in particular, glucose, indicating the relevance of their combined use as well as with other agents, which promote the deficiency of glucose in the tumor microenvironment.

Dichloroacetate (DCA) and Cancer: An Overview towards Clinical Applications

“…Similar results were obtained in melanoma cells in which the administration of retinoic acid receptor β (RAR β ) inhibitors confer sensitization to DCA [41]. A positive effect of DCA coadministration with metformin, a hypoglycaemic drug widely used to treat diabetes was demonstrated in a preclinical model of glioma [42] as well as in a low metastatic variant of Lewis lung carcinoma (LLC) [43]. Jiang and colleagues investigated the effects of phenformin, a metformin analog, and DCA in glioblastoma, demonstrating that contemporary inhibition of complex I and PDK by phenformin and DCA, respectively, decreased self-renewal and viability of glioma stem cells (GSCs), thus suggesting their possible employment to affect cancer stem cell fraction [44].…”

Cancer cachexia has many symptoms but only one cause: anoxia

“…• DCA targets mainly cells that cannot use the oxidative metabolism 248 . This concept is further confirmed by the synergy between DCA and metformin [267][268][269][270][271][272][273][274] . The fundamentals of this association stem from the fact that metformin inhibits mitochondrial Complex I and reduces oxidative metabolism while DCA inhibits glycolysis.…”

Metformin and sodium dichloroacetate effects on proliferation, apoptosis, and metabolic activity tested alone and in combination in a canine prostate and a bladder cancer cell line

“…To reduce the required DCA dose and associated side effects and to avoid resistance development, the combination therapy with a synergistic substance is a promising approach in cancer therapy [ 55 ]. In vitro studies demonstrated a synergistic effect of DCA and metformin in human breast cancer cells [ 13 ], human ovarian cancer cells [ 14 ], Lewis lung carcinoma [ 56 ], and murine glioblastoma cells [ 15 ]. Both substances are easy to synthesize and metformin is already approved as therapeutic for human medicine [ 56 , 57 ], while DCA has been used in humans for the treating lactic acidosis [ 24 , 58 , 59 ].…”