The effect of insulin and insulin-like growth factors on colonic crypt stem cell proliferation/survival and colorectal cancer

 

Trainee:

Sarah Bortvedt

Bortvedt

Research Mentor:

Dr. P. Kay Lund, PhD

Lund

Clinical Co-mentor:

Dr. Robert Sandler, MD

sandler

Additional Co-Mentor:

Dr. Temitope Keku, PhD

keku
Home Department Cell & Molecular Physiology
Project Description Colorectal cancer remains a prominent cause of cancer-related death, despite the prevalence of colonoscopy screening. This necessitates the discovery of biomarkers that can be used to predict early stages of colorectal cancer (CRC) or risk of precancerous adenoma. Previously, studies conducted by my mentors have shown colorectal cancer risk to be associated with an increase in insulin/insulin-like growth factor (IGF) levels or IGF axis activity. According to the working model for adenoma formation, reduced apoptosis and inappropriate survival and expansion of damaged crypt stem cells promotes adenoma. We predict that insulin/IGF axis activity promotes survival and expansion of genetically damaged stem cells, ultimately leading to adenoma formation and CRC. Further investigation of this hypothesis requires stem cell biomarkers. Sox9 and Lgr5 are two recently described biomarkers known to mark different stem cell populations within the intestine and to be downstream targets of the Wnt pathway. The Wnt pathway activates β-catenin, a pathway frequently disregulated in CRC or adenoma and linked to stem cell marker expression. Finally, we would like to determine whether the insulin/IGF axis promotes stem cell survival or adenoma via the insulin receptor (IR) or the IGF-I receptor (IGF-IR). We will use colon cancer cell lines, mouse models, and samples from routine colonscopy patients to examine the role of insulin and/or IGFs acting via the IGF-IR or IR to enhance survival and promote expansion of Sox9 and Lgr5 positive stem cells, in normal crypts and in precancerous adenomas.

We will investigate whether reduced apoptosis is correlated with Sox9/Lgr5 positive stem cell expansion using RNA and tissue samples collected from patients in the Diet and Health Study (DHS) IV directed by my clinical mentor Dr. Sandler. Quantitative PCR (qPCR) and immunohistochemistry will be used to examine Sox9/Lgr5 expression. The role of insulin/IGF on Lgr5-positive stem cells will be explored using cells sorted from Lgr5-LacZ mice. These cells will be FACS isolated and expression of IGF axis components, such as IR, IGF-IR and downstream mediators, IRS-I and IRS-2, will be quantified using qPCR. Insulin and IGFs will be administered to the cells to determine the relative roles of IGF-IR and IR in stem cell survival and expansion. The effects of IGF-IR or IR modulation on cancer-associated stem cells will be examined in vivo in a mouse model of spontaneous adenoma generated by crossing Lgr5-LacZ mice with APC Min/+ mice and in CRC cell lines, whose degree of stem cell biomarker expression I am currently measuring by qPCR. These investigations will allow us to elucidate the direct role of insulin/IGFs acting via IGF-IR or IR on intestinal and cancer-associated stem cells.