Author ORCID Identifier

https://orcid.org/0000-0001-5700-6954

Year of Publication

2021

Degree Name

Doctor of Philosophy (PhD)

Document Type

Doctoral Dissertation

College

Medicine

Department/School/Program

Toxicology and Cancer Biology

First Advisor

Dr. Mark Evers

Abstract

Neurotensin (NT) is a nutrient-regulated gut hormone that plays important roles in lipid absorption, obesity, metabolic disorders, and normal and neoplastic growth in the intestine. In this study, we 1) elucidate the mechanisms regulating NT release from endocrine cells, 2) examine the role of NT on proliferation and stem cell function in the small intestine and 3) define the effects of NT on colorectal cancer stem cells. We report that NT release from endocrine cells is enhanced by the MAPK scaffold protein Kinase Suppressor of Ras 1 (KSR1) and the exocyst complex component 70 (Exo70). Moreover, free fatty acid stimulated release of NT is attenuated by inhibition of KSR1, MEK, ERK1/2, and Exo70, indicating these proteins as possible therapeutic targets for modulating the negative effects of NT on lipid absorption, obesity, and metabolic dysfunction.

Functionally, we show that NT plays an evolutionarily conserved role in maintenance of intestinal stem cells during nutritional stress. NT is required for the induction of WNT/β-catenin signaling and ISC-specific gene expression during fasting and promotes intestinal stem cell self-renewal. In the murine small intestine, loss of NT impairs crypt progenitor cell proliferation via downregulation of ERK1/2 signaling, cyclin D1 expression, and cell cycle related gene expression programs. Loss of NT impairs intestinal stem cell-specific gene expression and intestinal stem cell function after fasting, whereas NT overexpression prevents intestinal stem cell depletion in the midgut of Drosophila fed a nutrient-reduced diet.

We extend these findings to reveal a similar role for NT in regulation of stem cell function in colorectal cancer. NT enhances WNT/β-catenin signaling and cancer stem cell self-renewal in the human colon carcinoma cell line HCT116 and in intestinal tumor organoids derived from APC mutant models of colorectal cancer. Mechanistically, we show that NT induces phosphorylation of LRP6 in an ERK1/2 dependent manner in colorectal cancer cells and intestinal tumor organoids, indicating that NT promotes WNT/β-catenin signaling via activation of the ERK1/2 signaling pathway. In contrast, loss of NT impairs cancer stem cell function in tumor organoids derived from APCmin/NT-/-mice, and our preliminary findings indicate that NT-deficiency significantly increases lifespan in APC mutant mice.

Collectively, the studies within this dissertation identify NT as a positive regulator of normal and neoplastic growth within the small intestine through regulation of stem cell function and activation of the WNT/β-catenin pathway and identify potential therapeutic targets for modulating the effects of NT in the small intestine.

Digital Object Identifier (DOI)

https://doi.org/10.13023/etd.2021.219

Funding Information

1. National Institutes of Health grant R01 DK112034 2016-2021

2. National Institutes of Health grant R01 CA208343 2019-2021

3. National Institutes of Health grant R01 CA133429 2019-2021

4. National Institutes of Health grant R35 GM131807 2020-2021

5. National Institutes of Health grant T32 DK007778 2018-2020

6. National Cancer Institute grant P30 CA177558 2016-2021

Available for download on Thursday, December 16, 2021

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