Transcriptomic and Cellular Content Analysis of Colorectal Cancer by Combining Multiple Independent Cohorts.

INTRODUCTION: By linking cellular content and molecular subtypes of colorectal cancer (CRC), we aim to uncover novel features useful for targeted therapy. Our first goal was to evaluate gene expression alterations linked to CRC pathogenesis, and then, we aimed to evaluate the cellular composition differences between normal colon mucosa and tumor and between different colon cancer molecular subtypes. METHODS: We collected microarray and RNA sequencing data of patients with CRC from the Genome Expression Omnibus and The Cancer Genome Atlas. We combined all cases and performed quantile normalization. Genes with a fold change of >2 were further investigated. We used xCell for cellular decomposition and CMScaller for molecular subtyping. For statistical analyses, the Kruskal-Wallis H test and Mann-Whitney U tests were performed with Bonferroni correction. RESULTS: We established an integrated database of normal colon and CRC using transcriptomic data of 1,082 samples. By using this data set, we identified genes showing the highest differential expression in colon tumors. The top genes were linked to calcium signaling, matrix metalloproteinases, and transcription factors. When compared with normal samples, CD4+ memory T cells, CD8+ naive T cells, CD8+ T cells, Th1 cells, Th2 cells, and regulatory T cells were enriched in tumor tissues. The ImmuneScore was decreased in tumor samples compared with normal samples. The CMS1 and CMS4 molecular subtypes were the most immunogenic, with the highest ImmuneScore but also high infiltration by CD8+ T cells, Th1 cells, and Th2 cells in CMS1 and B-cell subtypes and CD8+ T cells in CMS4. DISCUSSION: Our analysis uncovers features enabling advanced treatment selection and the development of novel therapies in CRC.

A Detailed Overview About the Single-Cell Analyses of Solid Tumors Focusing on Colorectal Cancer.

In recent years, the evolution of the molecular biological technical background led to the widespread application of single-cell sequencing, a versatile tool particularly useful in the investigation of tumor heterogeneity. Even 10 years ago the comprehensive characterization of colorectal cancers by The Cancer Genome Atlas was based on measurements of bulk samples. Nowadays, with single-cell approaches, tumor heterogeneity, the tumor microenvironment, and the interplay between tumor cells and their surroundings can be described in unprecedented detail. In this review article we aimed to emphasize the importance of single-cell analyses by presenting tumor heterogeneity and the limitations of conventional investigational approaches, followed by an overview of the whole single-cell analytic workflow from sample isolation to amplification, sequencing and bioinformatic analysis and a review of recent literature regarding the single-cell analysis of colorectal cancers.

Folic Acid Treatment Directly Influences the Genetic and Epigenetic Regulation along with the Associated Cellular Maintenance Processes of HT-29 and SW480 Colorectal Cancer Cell Lines.

Folic acid (FA) is a synthetic form of vitamin B9, generally used as a nutritional supplement and an adjunctive medication in cancer therapy. FA is involved in genetic and epigenetic regulation; therefore, it has a dual modulatory role in established neoplasms. We aimed to investigate the effect of short-term (72 h) FA supplementation on colorectal cancer; hence, HT-29 and SW480 cells were exposed to different FA concentrations (0, 100, 10,000 ng/mL). HT-29 cell proliferation and viability levels elevated after 100 ng/mL but decreased for 10,000 ng/mL FA. Additionally, a significant (p ≤ 0.05) improvement of genomic stability was detected in HT-29 cells with micronucleus scoring and comet assay. Conversely, the FA treatment did not alter these parameters in SW480 samples. RRBS results highlighted that DNA methylation changes were bidirectional in both cells, mainly affecting carcinogenesis-related pathways. Based on the microarray analysis, promoter methylation status was in accordance with FA-induced expression alterations of 27 genes. Our study demonstrates that the FA effect was highly dependent on the cell type, which can be attributed to the distinct molecular background and the different expression of proliferation- and DNA-repair-associated genes (YWHAZ, HES1, STAT3, CCL2). Moreover, new aspects of FA-regulated DNA methylation and consecutive gene expression were revealed.