Elucidation of the sulfur oxidation pathway and sulfur metabolism alterations in colorectal cancer

Globally, colorectal cancer (CRC) is the third leading cause of cancer-related death and the incidence
is still increasing. Our gastrointestinal tract is home to hundreds of bacterial species, constituting the
gut microbiome. Intestinal cells are exposed to hydrogen sulfide (H 2 S) produced by commensal gut
bacteria contained in the microbiome. H 2 S is also produced in mammalian cells through side
reactions via the following transsulfuration (TS) pathway enzymes: cystathionine-β-synthase (CBS),
cystathionine-γ-lyase (CSE) and combined action of cysteine aminotransferase (CAT) and 3-
mercaptopyruvate sulfurtransferase (MPST). H 2 S and persulfides have been shown to affect signaling
pathways and metabolic functions involved in carcinogenesis. The sulfur oxidation pathway (SuOX)
plays a key role in the catabolism of H 2 S in mammalian cells and notably in the intestinal epithelium,
but the sequence of reactions involved in this pathway remains poorly understood.
The first objective of our project is to clarify the role of enzymes previously associated with H 2 S
oxidation (namely sulfide quinone oxidoreductase (SQRDL), persulfide dioxygenase (ETHE1),
thiosulfate sulfurtransferase (TST), and thiosulfate:glutathione sulfurtransferase (TSTD1)) utilizing
HCT116 cell lines in which these enzymes have been knocked out using the Crispr/Cas9 technology.
The second objective is to investigate alterations in sulfur metabolism and disturbance of H 2 S
homeostasis in CRC tumorigenesis and progression using the APC min/+ mouse model.
To address both of these objectives, we have successfully developed a sensitive and
quantitative LC-MS/MS-based method for analyzing thiol- and persulfide-containing metabolites in
cell and tissue extracts. We found that the SQRDL and ETHE1 enzymes are essential for H 2 S
detoxication and thiosulfate production while TSTD1 is not required. Glutathione persulfide
produced by SQRDL is used downstream by ETHE1 enzyme as substrate. In analyses of epithelial
tissue samples derived from APC min/+ mice, we found alterations in the SuOX and TS pathways in
tumorigenesis, both at mRNA and metabolite levels. We found an accumulation of cystathionine in
tumor compared to healthy tissue samples, in agreement with decreased expression of the CSE
enzyme in the tumor samples. We also found decreased ETHE1 expression and thiosulfate
production in tumor. To elucidate potential effects of H 2 S on cancer cell growth, proliferation and
invasion, we continue our experiments with HCT116 cell lines at different supplementations. Gaining
a deeper insight into H 2 S homeostasis in carcinogenesis could contribute to the efforts to develop
specific diet recommendations for CRC prevention or management and reveal potential targets for
chemotherapy in sulfur metabolism.
Keywords: Colorectal cancer, sulfur metabolism, sulfur oxidation pathway

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