Metabolic characterization of aggressive breast cancer cells exhibiting invasive phenotype: impact of non-cytotoxic doses of 2-DG on diminishing invasiveness

Abstract

Background Metabolic reprogramming is being recognized as a fundamental hallmark of cancer, and efforts to identify drugs that can target cancer metabolism are underway. In this study, we used human breast cancer (BC) cell lines and established their invading phenotype (INV) collected from transwell inserts to compare metabolome differences and evaluate prognostic significance of the metabolome in aggressive BC invasiveness. Methods The invasiveness of seven human BC cell lines were compared using the transwell invasion assay. Among these, INV was collected from SUM149, which exhibited the highest invasiveness. Levels of metabolites in INV were compared with those of whole cultured SUM149 cells (WCC) using CE-TOFMS. The impact of glycolysis in INV was determined by glucose uptake assay using fluorescent derivative of glucose (2-NBDG), and significance of glycolysis, or tricarboxylic acid cycle (TCA) and electron transport chain (ETC) in the invasive process were further determined in aggressive BC cell lines, SUM149, MDA-MB-231, HCC1937, using invasion assays in the presence or absence of inhibitors of glycolysis, TCA cycle or ETC. Results SUM149 INV sub-population exhibited a persistent hyperinvasive phenotype. INV were hyper-glycolytic with increased glucose (2-NBDG) uptake; diminished glucose-6-phosphate (G6P) levels but elevated pyruvate and lactate, along with higher expression of phosphorylated-pyruvate dehydrogenase (pPDH) compared to WCC. Notably, inhibiting of glycolysis with lower doses of 2-DG (1 mM), non-cytotoxic to MDA-MB-231 and HCC1937, was effective in diminishing invasiveness of aggressive BC cell lines. In contrast, 3-Nitropropionic acid (3-NA), an inhibitor of succinate dehydrogenase, the enzyme that oxidizes succinate to fumarate in TCA cycle, and functions as complex II of ETC, had no significant effect on their invasiveness, although levels of TCA metabolites or detection of mitochondrial membrane potential with JC-1 staining, indicated that INV cells originally had functional TCA cycles and membrane potential. Conclusions Hyper-glycolytic phenotype of invading cells caters to rapid energy production required for invasion while TCA cycle/ETC cater to cellular energy needs for sustenance in aggressive BC. Lower, non-cytotoxic doses of 2-DG can hamper invasion and can potentially be used as an adjuvant with other anti-cancer therapies without the usual side-effects associated with cytotoxic doses.

Glucose deprivation reduces proliferation and motility, and enhances the anti-proliferative effects of paclitaxel and doxorubicin in breast cell lines in vitro

“…Several previous reports have demonstrated similar effects of 2-DG on breast cancer cells, with reduction in their migration [ 40 ] and proliferation [ 41 – 45 ]. In another recent report, enhanced glucose uptake was correlated with enhanced invasiveness of several breast cancer cell lines, whereas low doses of 2-DG (1 mM) significantly reduced their invasive capacity [ 46 ]. Our data is consistent with these reports, in the same concentration range (0.5–1 mM).…”

A Multi-Omics Study Revealing the Metabolic Effects of Estrogen in Liver Cancer Cells HepG2

“…For example, 1 mM 2-DG blocked cell viability and induced cytotoxicity in liver cancer cells HepG2 in this study. It has been reported that 1 mM 2-DG is toxic in breast cancer cell SUM149 [ 48 ], but it is not toxic in other breast cancer cells MDA-MB-231, HCC1937, HDQ-P1, and MCF-7 cells [ 48 , 49 ]. Thus, the findings from this study can be referenced in future studies.…”

Combining Carbon-Ion Irradiation and PARP Inhibitor, Olaparib Efficiently Kills BRCA1-Mutated Triple-Negative Breast Cancer Cells

“…Immunofluorescence labeling and image acquisition were performed as described previously, with some modifications. 29 Briefly, cells were fixed with 4% paraformaldehyde in phosphate-buffered saline (PBS; Nissui Pharmaceutical Co., Ltd.; Tokyo, Japan) for 15 min, and given three washes with PBS. Cells were then blocked with PBS containing 5% fetal calf serum and 0.3% Triton ×100, followed by incubation with primary antibody for 30 min at room temperature.…”