Skip to main content
Premium Trial:

Request an Annual Quote

Single-Cell Analysis of Colorectal Cancer Uncovers Two Epithelial-Cell Based Subtypes

NEW YORK — Researchers have identified two epithelial cell-based colorectal cancer subtypes using single-cell transcriptome analyses, suggesting an update to how the disease is classified.

An international consortium of scientists previously identified four consensus molecular subtypes of colorectal cancer based on bulk tumor analysis. These subtypes, CMS1 though CMS4, are distinguished by their levels of immune infiltration, activation of WNT and Myc, metabolic dysregulation, and mesenchymal fibrotic reaction. But as Iain Beehuat Tan from the National Cancer Center Singapore noted, at the bulk tumor level, the transcriptomes of the different cell types are obscured.

He and his colleagues examined the single-cell transcriptomes of more than 140 tumor samples from 63 patients to study the contribution of epithelial cells. As they reported on Thursday in Nature Genetics, they uncovered two distinct groups of malignant epithelial cells, which they dubbed iCMS2 and iCMS3, that corresponded with CMS classifications but also refined them.

"Putting things together, integrating epithelial subtypes with bulk tumor phenotypes, we propose IMF, a sequential layered classification structure based on intrinsic epithelial subtypes (l), microsatellite status (M), and the presence of fibrosis (F)," Tan, the co-senior author on the paper, said in an email.

For their analysis, the researchers examined single cells isolated from 141 tumors from 63 patients alongside 39 normal tissue samples and nine lymph node samples. The samples came from five cohorts from Singapore, Belgium, and Korea.

They focused on a set of 49,155 epithelial cells. Principal components analysis uncovered two epithelial subgroups within samples from one of the Singapore cohorts. Using the 848 genes that were differentially expressed between those subgroups, the researchers could identify them within the other four sample cohorts. A combined analysis of all five groups could also tease out the iCMS2 and iCMS3 subtypes.

The two subtypes also emerged when the researchers instead analyzed single-cell activity scores of the regulons of about 350 transcription factors, as well as when they examined copy number changes. iCMS2 tumors, they found, were marked by certain chromosomal arm gains and losses, particularly the gain of 7pq, 8q, 13q, and 20pq and the loss of 1p, 4pq, 8p, 14q, 15q, 17p, and 18pq. iCMS3 tumors, on the other hand, were more often diploid or had infrequent copy number alterations.

The researchers further identified 715 iCMS marker genes that clustered cells into normal, iCMS2, or iCMS3. When they applied these marker genes to a set of 3,614 bulk tumor transcriptomes from 15 different datasets, they could also identify the two groups.

When the researchers then examined these colorectal cancer subtypes in relation to other subtypes, they found that the iCMS3 subtype encompasses microsatellite unstable (MSI-H) cancers as well as a third of the microsatellite stable (MSS) tumors. The iCMS3-MSS tumors were more similar, transcriptionally speaking, to MSI-H tumors than to other MSS tumors.

At the same time, the vast majority of CMS1 and CMS3 tumors had iCMS3-type epithelium, and most CMS2 tumors were iCMS2. Tumors falling in the CMS4 or fibrosis subtype, meanwhile, could have either iCMS2 or iCMS3 type epithelium, suggesting that the intrinsic epithelial subtypes and fibrosis are decoupled. The researchers further noted that the fibrotic CMS4 cancers with iCMS3 epithelial cells had the worst prognosis.

Based on these findings, they suggested that colorectal cancer classification should be updated to take intrinsic epithelial subtype, microsatellite status, and presence of fibrosis into account.

"This new classification system could give us the functional units that explain the diversity of colorectal cancer: intrinsic epithelial axis with the distinct biological programs active in i2 and i3 cancers, microsatellite status and its impact on tumor immunogenicity and response to immunotherapy, and fibrosis and its potential impact on propensity to metastasize," Tan said.