Cell-autonomous redox-sensitive role of selenoprotein glutathione peroxidase-2 (Gpx2) in APC-driven colorectal carcinogenesis and stem cell function

• Applicant: Niyati Vachharajani, Ph.D.
• Subject Area: Gastroenterology ; Hematology, Oncology ; Immunology
• Term: from 2018 to 2020
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 400020534

Colorectal cancer (CRC) is one of the leading causes of cancer-related deaths in western countries. Studies have shown that gut inflammation increases the risk of CRC, especially when patients are deficient in Selenium - a key component of selenoproteins involved in antioxidant defenses. Glutathione peroxidase-2 (GPx2), a gastrointestinal selenoprotein, is one of several Wnt target genes that are induced upon the loss of genes like the adenomatous polyposis coli (APC) - a frequently mutated gene in CRC. Additionally, GPx2 is thought to mediate self-renewal of the intestinal epithelium, and its upregulation in response to microbial colonization of the gut suggests a role in epithelium protection. Recent studies have shown that GPx2 is highly expressed in the tumor environment and is thought to either play an anti-inflammatory or anti-apoptotic role, depending on the cancer stage. However, the role of GPx2 in intestinal crypt growth and APC-driven tumorigenesis remains elusive. The central hypothesis of this proposal is that, GPx2 may contribute to the progression of epithelial oncogenic transformation in APC-driven colon tumorigenesis, and will be tested by the following aims: 1) The function of GPx2 will be characterized in a mouse model wherein the inducible loss of APC function within traceable Lrig1+ stem cells, predominantly leads to colonic adenomas. The role of GPx2 during tumorigenesis will be evaluated by subjecting mice of three genotypes (Gpx2+/+; Lrig1cre/+; ApcΔ14/+, Gpx2+/-; Lrig1cre/+; ApcΔ14/+, Gpx2-/-; Lrigcre/+; ApcΔ14/+) to tumorigenesis. Furthermore, the mice will be subjected to different dietary Selenium concentrations before tumor induction. Tumor development will be tracked by serial colonoscopy, in addition to immunophenotyping of immune cells in the lamina propria using flow cytometry. Redox-sensitive signaling pathways known to influence cell survival will be examined in tumor samples via RNA-seq and immunoblotting. In addition to mouse tumoroids (Gpx2-/-; Lrigcre/+; ApcΔ14/+), GPx2 expression will be modified in human tumoroids, to determine if GPx2 plays a role in progression of epithelial oncogenic transformation. Additionally, 16S rRNA sequencing will be used to characterize the gut microbiomes of the aforementioned genotypes to determine if the absence of GPx2 correlates with cancer progression and a redox-dependent shift in the microbiome. 2) The function of GPx2 in epithelial crypts will be defined by performing detailed structure-function studies of Gpx2 mutants constructs in Gpx2-/- enteroids.

• DFG Programme: Research Fellowships
• International Connection: USA
• Host: Professor Dr. Christopher Williams