Novel insights into the molecular mechanisms underlying risk of colorectal cancer from smoking and red/processed meat carcinogens by modeling exposure in normal colon organoids

Abstract

Tobacco smoke and red/processed meats are well-known risk factors for colorectal cancer (CRC). Most research has focused on studies of normal colon biopsies in epidemiologic studies or treatment of CRC cell lines in vitro . These studies are often constrained by challenges with accuracy of self-report data or, in the case of CRC cell lines, small sample sizes and lack of relationship to normal tissue at risk. In an attempt to address some of these limitations, we performed a 24-hour treatment of a representative carcinogens cocktail in 37 independent organoid lines derived from normal colon biopsies. Machine learning algorithms were applied to bulk RNA-sequencing and revealed cellular composition changes in colon organoids. We identified 738 differentially expressed genes in response to carcinogens exposure. Network analysis identified significantly different modules of co-expression, that included genes related to MSI-H tumor biology, and genes previously implicated in CRC through genome-wide association studies. Our study helps to better define the molecular effects of representative carcinogens from smoking and red/processed meat in normal colon epithelial cells and in the etiology of the MSI-H subtype of CRC, and suggests an overlap between molecular mechanisms involved in inherited and environmental CRC risk.

DNA methylation analysis of normal colon organoids from familial adenomatous polyposis patients reveals novel insight into colon cancer development

“…Colon organoids derived from normal-appearing colons of FAP patients and healthy subjects were grown in complex media, as described previously [13][14][15][16]. All colon organoids used within this study were derived from distal (left) colon biopsies.…”

“…FAP and healthy colon organoids were grown at the same time, under the same conditions to minimize the effects of batch on our dataset. Our procedure for establishing colon organoids has been described elsewhere [13][14][15][16]. All colon organoids considered for this study were between passages eight-ten.…”

“…For viability analysis, Matrigel was scratched off and resuspended in fresh culture medium, described elsewhere [13][14][15][16]. Organoids were broken down into single cells by washing each well with 500 μL DPBS (Gibco, ref: 10,010-031) and 10 μM Y27632 (R&D Systems ref: 1254) and transfer the additional 500 μL/well to the same tubes.…”

“…In recent years, our group has successfully employed the normal colon organoid model [12] to study the role of CRC risk factors in colon epithelial cell biology [13][14][15][16]. The colon organoid system serves as a model of the epithelial cells of the colonic crypt, particularly the colon crypt stem cell niche.…”

Insights into Early Onset Colorectal Cancer through Analysis of Normal Colon Organoids of Familial Adenomatous Polyposis Patients

“…Colon organoids represent a model of the colon stem cell niche, where CRC is believed to originate [ 17 ]. Previously, our group has used this model to investigate how differences in CRC environmental risk factors may alter gene expression of genes implicated in CRC [ 18 , 19 , 20 , 21 , 22 ]. In this study, we have applied the colon organoid model to identify gene expression differences between FAP subjects and healthy controls.…”