Nutrient-deprived cancer cells preferentially use sialic acid to maintain cell surface glycosylation

Abstract

Cancer is characterized by abnormal energy metabolism shaped by nutrient deprivation that malignant cells experience during various stages of tumor development. This study investigated the response of nutrient-deprived cancer cells and their non-malignant counterparts to sialic acid supplementation and found that cells utilize negligible amounts of this sugar for energy. Instead cells use sialic acid to maintain cell surface glycosylation through complementary mechanisms. First, levels of key metabolites (e.g., UDP-GlcNAc and CMP-Neu5Ac) required for glycan biosynthesis are maintained or enhanced upon Neu5Ac supplementation. In concert, sialyltransferase expression increased at both the mRNA and protein levels, which facilitated increased sialylation in biochemical assays that measure sialyltransferase activity as well as at the whole cell level. In the course of these experiments, several important differences emerged that differentiated the cancer cells from their normal counterparts including resistant to sialic acid-mediated energy depletion, consistently more robust sialic acid-mediated glycan display, and distinctive cell surface vs. internal vesicle display of newly-produced sialoglycans. Finally, the impact of sialic acid supplementation on specific markers implicated in cancer progression was demonstrated by measuring levels of expression and sialylation of EGFR1 and MUC1 as well as the corresponding function of sialic acid-supplemented cells in migration assays. These findings both provide fundamental insight into the biological basis of sialic acid supplementation of nutrient-deprived cancer cells and open the door to the development of diagnostic and prognostic tools.

Harnessing cancer cell metabolism for theranostic applications using metabolic glycoengineering of sialic acid in breast cancer as a pioneering example

“…As a brief background, OGT is an enzyme that installs a single GlcNAc on thousands of intracellular proteins to make the “ O -GlcNAc” post-translational modification originally described to have a yin-yang relation with phosphorylation [135]; it is now known that O -GlcNAc modification of proteins extends well beyond cross-talk with phosphorylation [133]). Of interest to nutrient-deprived breast cancer cells, the emergence of O -GlcNAc as a master regulator of epigenetics [136, 137] suggests that changes to flux through the HBP modulate a bevy of epigenetic-driven effects ranging from (1) direct changes to the chromatin (e.g., through changes to the methylation of promoter regions), to (2) indirect regulation via microRNAs that modulate glycosylation [138] and, (3) finally, to the expression of sialyltransferases, consistent with our finding that both UDP-levels and the these enzymes were highly affected by nutrient deprivation in breast cancer cells [76]. The O -GlcNAc protein modification ties oncogenesis to metabolism and illustrates how cancer progression does not rely solely on genetic mutations or even epigenetic mechanisms but extends to the dynamic regulation of the proteome.…”

“…Specifically, instead of changes being limited to a targeted reduction in highly-branched N-glycans due to the expected selective impact of lower UDP-GlcNAc levels on MGAT4/5 activity, dozens of additional glycan structures were affected with about half increasing in abundance; a reduction in UDP-GlcNAc could not have this effect based on any currently-known mechanism. Second, in recently-published studies we discovered links between metabolism and sialylation that also were not easily explained by direct interplay between metabolic intermediates and biosynthetic enzymes [76, 101]. In particular, cell surface sialylation was rapidly diminished upon induced nutrient deprivation of breast cancer cells evaluated in vitro , which based on the metabolic networks diagramed in Figure 4, initially appears reasonable because UDP-GlcNAc is the metabolic feedstock for sialylation.…”

“…In cancer, the neo-expression of the requisite sialyltransferase enables the production of these (or other) TACAs and is considered to be a key event in oncogenic progression. An example of this phenomenon is provided by our recent characterization of breast cancer cells where rapid increases in sialyltransferase expression was observed during recovery from nutrient deprivation [75, 76]. …”

“…We have already characterized the restoration of surface sialylation in nutrient-deprived breast cancer cells upon exogenous supplementation at a rudimentary biochemical level [75, 76, 101, 170]. These studies discounted the possibility that Neu5Ac was routed for energy production by showing that ATP levels did not increase upon supplementation but instead dramatically decreased in certain cell lines.…”

“…For example, because cell surface glycans offer a surprisingly comprehensive snapshot into the internal metabolic status of a cell and by cataloging these changes under various states of nutrient availability [101, 103, 182–186], TACA profiles that indicate the “energetics” status of cancer cell could be used to assess the stage of cancer and then to suggest a treatment strategy. In particular, we focus on nutrient-deprived breast cancer cells that efficiently take up sialic acids used to restore cell surface display of this sugar [75, 76, 101]. …”