Mitochondrial mass, a new metabolic biomarker for stem-like cancer cells: Understanding WNT/FGF-driven anabolic signaling

Abstract

Here, we developed an isogenic cell model of “stemness” to facilitate protein biomarker discovery in breast cancer. For this purpose, we used knowledge gained previously from the study of the mouse mammary tumor virus (MMTV). MMTV initiates mammary tumorigenesis in mice by promoter insertion adjacent to two main integration sites, namely Int-1 (Wnt1) and Int-2 (Fgf3), which ultimately activates Wnt/β-catenin signaling, driving the propagation of mammary cancer stem cells (CSCs). Thus, to develop a humanized model of MMTV signaling, we over-expressed WNT1 and FGF3 in MCF7 cells, an ER(+) human breast cancer cell line. We then validated that MCF7 cells over-expressing both WNT1 and FGF3 show a 3.5-fold increase in mammosphere formation, and that conditioned media from these cells is also sufficient to promote stem cell activity in untransfected parental MCF7 and T47D cells, as WNT1 and FGF3 are secreted factors. Proteomic analysis of this model system revealed the induction of i) EMT markers, ii) mitochondrial proteins, iii) glycolytic enzymes and iv) protein synthesis machinery, consistent with an anabolic CSC phenotype. MitoTracker staining validated the expected WNT1/FGF3-induced increase in mitochondrial mass and activity, which presumably reflects increased mitochondrial biogenesis. Importantly, many of the proteins that were up-regulated by WNT/FGF-signaling in MCF7 cells, were also transcriptionally over-expressed in human breast cancer cells in vivo, based on the bioinformatic analysis of public gene expression datasets of laser-captured patient samples. As such, this isogenic cell model should accelerate the discovery of new biomarkers to predict clinical outcome in breast cancer, facilitating the development of personalized medicine.Finally, we used mitochondrial mass as a surrogate marker for increased mitochondrial biogenesis in untransfected MCF7 cells. As predicted, metabolic fractionation of parental MCF7 cells, via MitoTracker staining, indicated that high mitochondrial mass is a new metabolic biomarker for the enrichment of anabolic CSCs, as functionally assessed by mammosphere-forming activity. This observation has broad implications for understanding the role of mitochondrial biogenesis in the propagation of stem-like cancer cells. Technically, this general metabolic approach could be applied to any cancer type, to identify and target the mitochondrial-rich CSC population.The implications of our work for understanding the role of mitochondrial metabolism in viral oncogenesis driven by random promoter insertions are also discussed, in the context of MMTV and ALV infections.

Loss of fructose-1,6-bisphosphatase induces glycolysis and promotes apoptosis resistance of cancer stem-like cells: an important role in hexavalent chromium-induced carcinogenesis

“…In the present study we have shown that glucose uptake, lactate production, and ATP content were all remarkably reduced in BEAS-2B-Cr-CSC compared to normal cells and Cr(VI)-transformed cells, indicating metabolic inactiveness of those CSCs. The mitochondria of CSCs have an increased mass and membrane potential, which reflects mitochondrial function, higher mitochondrial ROS and enhanced oxygen consumption rates compared with the differentiated cancer cells (Lagadinou et al , 2013; Pasto et al , 2014; De Luca et al , 2015; Lamb et al , 2015; Sancho et al , 2015). Another study has reported the lower quantity of mitochondrial DNA, higher mitochondrial potential, lower oxygen/glucose consumption, and lower intracellular levels of ROS and ATP content in CSCs isolated from adenocarcinomic human alveolar basal epithelial A549 cells (Ye et al , 2011).…”

Fibroblast growth factor–mediated crosstalk in cancer etiology and treatment

“…This triggers paracrine FGF9/FGFR3 and Wnt signaling pathways, which promote the growth of the stem cell population (Fillmore et al, ). Overexpression of FGF3 and Wnt1 in MCF7 breast cancer cells increased mammosphere formation, an indicator of “stemness.” These cells exhibited up‐regulated genes relating to epithelial‐mesenchymal transition and cell migration, consistent with a cancer stem cell phenotype (Lamb et al, ). Furthermore, breast cancer cells can revert to a fetal mammary stem cell state, especially within mouse models exhibiting features of basal‐like and HER2 + tumors.…”

The Next Generation of Anticancer Metallopharmaceuticals: Cancer Stem Cell‐Active Inorganics

“…These include, but are not limited to, mitochondrial DNA content, metabolic phenotype, intracellular ATP and mitochondrial membrane potential. Furthermore, independent studies have shown that CSCs have a higher mitochondrial mass than bulk cancer cells; this highlights the significance of mitochondrial function to CSC regulation . Therefore, mitochondrial targeting could offer a viable approach for the selective elimination of CSCs over bulk cancer cells.…”