Use of Cancer Stem Cells to Investigate the Pathogenesis of Colitis-associated Cancer

FUTURE DIRECTIONS AND METHODS FOR IBD RESEARCH Use of Cancer Stem Cells to Investigate the Pathogenesis of Colitis-associated Cancer Julie M. Davies, PhD, Rebeca Santaolalla, PhD, and Maria T. Abreu, MD mutational history and tumor microenvironment observed in CAC patients is distinct from that observed in sporadic colon cancer and suggests a different etiology. Recently, much attention has been focused on understanding the cellular origin of cancer and the cancer stem cells, which is key to growth and progression. Cancer stem cells are often chemo-resistant making them attractive targets for improving patient outcomes. New techniques have rapidly been evolving allowing for a better understanding of the normal intestinal stem cell function and behavior in the niche. Use of these new technologies will be crucial to understanding cancer stem cells in both sporadic and CAC. In this review, we will explore emerging methods related to the study of normal and cancer stem cells in the intestine, and examine potential avenues of investigation and application to understanding the pathogenesis of CAC. (Inflamm Bowel Dis 2016;22:976–983) Key Words: colitis-associated cancer, stem cells, inflammation, enteroid, colonoid, APC, Lgr5, b-catenin C olon cancer is the third most prevalent cancer in both men and women worldwide. The incidence of disease in men in the United States is 1 in 56,400 and 1 in 41,900 for women.1 Genetic conditions, such as familial adenomatous polyposis and hereditary nonpolyposis colon cancer, account for ;5 to 10% of colon cancer cases, but age1 and environmental factors including diet, alcohol intake,2 obesity, and migration from a low-incidence country to a high-incidence country3,4 all confer increased risk. A Received for publication November 5, 2015; Accepted January 14, 2016. From the Division of Gastroenterology, Department of Medicine, University of Miami Miller School of Medicine, Miami, Florida. Supported by grants to MTA (National Institute of Health, National Institute of Diabetes and Digestive and Kidney Diseases 2R01DK099076-06A1; Crohn’s and Colitis Foundation of America Senior Research Award #3786; and Broad Medical Research Foundation at CCFA, award IBD-0989R). Additional funding was provided by The Micky & Madeleine Arison Family Foundation Crohn’s & Colitis Discovery Laboratory, and philanthropic support by Mrs Joanne “Joni” and Mr Dohn Trempala. M. T. Abreu has served as a consultant to AbbVie Laboratories, Prometheus Laboratories, Hospira Inc., Pfizer, Sanofi Aventis, Janssen, Ferring Pharmaceuticals, Takeda, GSK Holding Americas Inc., Focus Medical Communications, Mucosal Health Board, Shire Pharmaceuticals, Prometheus Laboratories, Eli Lilly, and UCB. MTA is also a member of the board of GI Health Foundation and the scientific advisory board of Asana Medical Inc. and Celgene Corp. J. M. Davies and R. Santaolalla declare no competing interests. The remaining authors have no conflict of interest to disclose. Reprints: Maria T. Abreu, MD, Division of Gastroenterology, Department of Medicine, University of Miami Miller School of Medicine, Miami, FL 33136 (e-mail: ). Copyright © 2016 Crohn’s & Colitis Foundation of America, Inc. This is an open access article distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivatives License 4.0 (CC BY-NC-ND), which permits downloading and sharing the work provided it is properly cited. The work cannot be changed in any way or used commercially. DOI 10.1097/MIB.0000000000000756 Published online 9 March 2016. 976 | www.ibdjournal.org significant risk factor in the development of colon cancer is the presence of intestinal inflammatory conditions, including ulcerative colitis (UC) and Crohn’s disease. Analysis of the risk of developing cancer in patients with UC has been evaluated by several studies and by meta-analyses5–7 over the years. Estimates of incidence have been associated with duration of disease and extent of inflammation and were found to be upward of 30% in patients with disease for over 35 years.6 However, recent studies using population-based patient cohorts have not revealed an increased incidence of cancer in patients with UC.8–10 Nevertheless, age of onset between 20 and 39 years continued to demonstrate an increased relative risk of developing cancer after 20 years.8 Moreover, mounting evidence demonstrates that antiinflammatory medications are protective against the development of cancer in UC.11,12 This evidence provides support for an important role for chronic immune activation in the development of colon tumors. Mutations in sporadic colon cancer occur in a generally predictable manner as outlined by Vogelstein.13,14 Initiating mutations in the adenomatous polyposis coli (APC) gene are followed by mutations in Kirsten rat sarcoma (K-RAS) and p53 which lead to the development of adenoma and then cancer. However, in colitis-associated cancer (CAC), the mutational progression through tumorigenesis is less well defined. Mutations in APC are found less frequently in both low-grade and high-grade dysplasia in patients with CAC than in patients with sporadic colon cancer.15,16 In CAC, the driving force behind tumor development is considered to be the inflammation itself.17,18 APC functions in a protein complex that controls the ubiquitination and cellular localization of the transcription cofactor b-catenin. When b-catenin is bound by the inactivating complex, it is ubiquitinated and degraded. When the complex is disrupted, either by a mutated APC protein, or through phosphorylation of another member of Inflamm Bowel Dis  Volume 22, Number 4, April 2016 Abstract: Colitis-associated cancer (CAC) can develop in patients with inflammatory bowel disease with long-term uncontrolled inflammation. The Inflamm Bowel Dis  Volume 22, Number 4, April 2016 intestinal stem cells, and how these functions may be coopted during transformation to drive cancer growth. We will also examine some of the emerging techniques that will allow a more in-depth study of the interaction of stem cells with other aspects of the environment and explore potential new uses for these technologies in understanding the pathogenesis of CAC. COLON CANCER STEM CELLS AND NORMAL STEM CELLS There are 2 modes of developing a cancer stem cell: (1) direct mutation of the native stem cell itself. In this model, the stem cell acquires mutations which results in altered self-renewal kinetics and uncontrolled growth and altered differentiation of progeny to form the bulk cancer cells, or (2) acquisition of stem properties in differentiated cells through mutations in transit amplifying precursor populations allowing them to dedifferentiate into stem cells. Identification of the initiating cancer stem cells can be performed using several methods. In vivo, assays include using human primary tumors fractionated into subsets based on surface protein expression and injected into an immunocompromised host mouse (...truncated)