Pharmacological induction of mesenchymal-epithelial transition via inhibition of H2S biosynthesis and consequent suppression of ACLY activity in colon cancer cells

Abstract

Hydrogen sulfide (H₂S) is an important endogenous gaseous transmitter mediator, which regulates a variety of cellular functions in autocrine and paracrine manner. The enzymes responsible for the biological generation of H₂S include cystathionine-β-synthase (CBS), cystathionine-γ-lyase (CSE) and 3-mercaptopyruvate sulfurtransferase (3-MST). Increased expression of these enzymes and overproduction of H₂S has been implicated in essential processes of various cancer cells, including the stimulation of metabolism, maintenance of cell proliferation and cytoprotection. Cancer cell identity is characterized by so-called "transition states". The progression from normal (epithelial) to transformed (mesenchymal) state is termed epithelial-to-mesenchymal transition (EMT) whereby epithelial cells lose their cell-to-cell adhesion capacity and gain mesenchymal characteristics. The transition process can also proceed in the opposite direction, and this process is termed mesenchymal-to-epithelial transition (MET). The current project was designed to determine whether inhibition of endogenous H₂S production in colon cancer cells affects the EMT/MET balance in vitro. Inhibition of H₂S biosynthesis in HCT116 human colon cancer cells was achieved either with aminooxyacetic acid (AOAA) or 2-[(4-hydroxy-6-methylpyrimidin-2-yl)sulfanyl]-1-(naphthalen-1-yl)ethan-1-one (HMPSNE). These inhibitors induced an upregulation of E-cadherin and Zonula occludens-1 (ZO-1) expression and downregulation of fibronectin expression, demonstrating that H₂S biosynthesis inhibitors can produce a pharmacological induction of MET in colon cancer cells. These actions were functionally reflected in an inhibition of cell migration, as demonstrated in an in vitro "scratch wound" assay. The mechanisms involved in the action of endogenously produced H₂S in cancer cells in promoting (or maintaining) EMT (or tonically inhibiting MET) relate, at least in part, in the induction of ATP citrate lyase (ACLY) protein expression, which occurs via upregulation of ACLY mRNA (via activation of the ACLY promoter). ACLY in turn, regulates the Wnt-β-catenin pathway, an essential regulator of the EMT/MET balance. Taken together, pharmacological inhibition of endogenous H₂S biosynthesis in cancer cells induces MET. We hypothesize that this may contribute to anti-cancer / anti-metastatic effects of H₂S biosynthesis inhibitors.

Keywords: 3-mercaptopyruvate sulfurtransferase; Colon cancer; Cystathionine-β-synthase; Epithelial cells; Hydrogen sulfide; Mesenchymal cells; Nitric oxide.