Hyaluronan Production Regulates Metabolic and Cancer Stem-like Properties of Breast Cancer Cells via Hexosamine Biosynthetic Pathway-coupled HIF-1 Signaling.

Cancer stem cells (CSCs) represent a small subpopulation of self-renewing oncogenic cells. As in many other stem cells, metabolic reprogramming has been implicated to be a key characteristic of CSCs. However, little is known about how the metabolic features of cancer cells are controlled to orchestrate their CSC-like properties. We recently demonstrated that hyaluronan (HA) overproduction allowed plastic cancer cells to revert to stem cell states. Here, we adopted stable isotope-assisted tracing and mass spectrometry profiling to elucidate the metabolic features of HA-overproducing breast cancer cells. These integrated approaches disclosed an acceleration of metabolic flux in the hexosamine biosynthetic pathway (HBP). A metabolic shift toward glycolysis was also evident by quantitative targeted metabolomics, which was validated by the expression profiles of key glycolytic enzymes. Forced expression of glutamine:fructose-6-phosphate amidotransferase 1 (GFAT1), an HBP rate-limiting enzyme, resembled the results of HA overproduction with regard to HIF-1α accumulation and glycolytic program, whereas GFAT1 inhibition significantly decreased HIF-1α protein level in HA-overproducing cancer cells. Moreover, inhibition of the HBP-HIF-1 axis abrogated HA-driven glycolytic enhancement and reduced the CSC-like subpopulation. Taken together, our results provide compelling evidence that HA production regulates the metabolic and CSC-like properties of breast cancer cells via HBP-coupled HIF-1 signaling.

Excessive hyaluronan production promotes acquisition of cancer stem cell signatures through the coordinated regulation of Twist and the transforming growth factor ß (TGF-ß)-Snail signaling axis.

The cancer stem cell (CSC) model suggests that a small subpopulation of cancer cells possesses the ability to self-renew and give rise to malignant progeny that drive cancer progression. Recent reports have also proposed the existence of certain extra- or intracellular signals that allow cancer progenitors to dynamically revert to a stem cell state. However, the mechanisms underlying cancer cell plasticity and CSC expansion are not entirely clear. Our previous studies using a hyaluronan synthase 2 (Has2) transgenic mouse model demonstrated that hyaluronan overproduction caused rapid development of aggressive breast carcinoma at a high incidence. Thus, we hypothesize that hyaluronan overproduction may accelerate cancer progression by expanding CSC subpopulations during cancer development. Primary cancer cells were established from mammary tumors developed in the transgenic mice and subjected to the Hoechst 33342 dye exclusion assay to sort side population (SP) from non-side population (non-SP) cells. Flow cytometric analysis demonstrated the enrichment of CD44(high)/CD24(low) CSC-like cells in the SP fraction of hyaluronan-overproducing cancer cells. This subpopulation exhibited several characteristics that were similar to CSCs, including cancer-initiating and mammosphere-forming abilities. Excess hyaluronan production drove the epithelial-to-mesenchymal transition process defined as the loss of epithelial phenotypes, up-regulation of transforming growth factor ß (TGF-ß), and induction of the epithelial-to-mesenchymal transition-related transcriptional factors Snail and Twist. Inhibition of TGF-ß-Snail signaling or silencing of Twist expression abrogated the entrance into a stem cell state. Taken together, our findings suggest that hyaluronan overproduction allows plastic cancer cell populations to revert to stem cell states via Twist and the TGF-ß-Snail signaling axis.