Ano1 is a Prognostic Biomarker That is Correlated with Immune Infiltration in Colorectal Cancer

Abstract

Background: Anoctamin 1 (ANO1) has been observed to be overexpressed in gastrointestinal and pulmonary epithelial cells, as well as in a number of cancers. Although Ano1 is involved in the prognosis of colorectal cancer (CRC), its mechanism of action in metastatic CRC has not been fully elucidated. Methods: The expression of Ano1 was assessed in samples obtained from The Cancer Genome Atlas (TCGA) database. Then, we used Gene Ontology (GO) enrichment, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, Gene set enrichment analysis (GSEA), Gene set variation analysis (GSVA), and Weighted Correlation Network Analysis (WGCNA) to determine the functions of Ano1. Additionally, random survival forest, Cox multivariate analysis, Kaplan Meier analysis, and ROC were used to determine the predictive value of Ano1 on clinical outcomes in CRC patients. Finally, HE staining, immunohistochemical (IHC) analysis and qRT-PCR were used to explore the expression of the Ano1 gene in CRC tissue. Results:The expression level of Ano1 in CRC was significantly elevated, and the prognosis was poor. The modules with a higher proportion of upregulated genes tended to be positively correlated with Ano1-high. KNG1, GNG4, F2, POSTN, THBS2, SPP1 and FGA were identified as hub proteins of the PPI network. The heatmap showed that the expression level of the Ano1-high group was significantly negatively correlated with immune infiltrate. The overexpression of the Ano1 gene in CRC tissue samples was also confirmed by HE staining, immunohistochemical (IHC) analysis and qRT-PCR. Conclusion: High expression of Ano1 is closely related to a poor prognosis in patients with colorectal cancer. Ano1 may participate in the metastasis and progression, as well as the immune regulation of CRC. In summary, Ano1 can act as a potential prognostic biomarker and a novel target for CRC therapy.

Enhancing Proteome Coverage by Using Strong Anion-Exchange in Tandem with Basic-pH Reversed-Phase Chromatography for Sample Multiplexing-Based Proteomics

“…Interestingly, the protein level of ERMN were particularly enriched in the near-haploid leukemia cell line HAP1 (Figure B). Anoctamin1, ANO1, a calcium-activated chloride channel that is highly expressed in epithelial cells, is also overexpressed in many types of cancers that originated from epithelia, such as colon cancer, and has been implicated as a prognostic biomarker for colorectal cancer . It follows that ANO1 was expressed at higher levels in the colorectal HCT116 cell line (Figure C).…”

“…Anoctamin1, ANO1, a calcium-activated chloride channel that is highly expressed in epithelial cells, is also overexpressed in many types of cancers that originated from epithelia, such as colon cancer, and has been implicated as a prognostic biomarker for colorectal cancer. 52 It follows that ANO1 was expressed at higher levels in the colorectal HCT116 cell line (Figure 3C). HHIP is a membrane protein that regulates hedgehog signaling and is important for the development of brain and lung tissue.…”

FOXD subfamily genes serve as biomarkers and therapy targets in colorectal cancer

“…The protocol was approved by the ethics committee (approval number:2021-079). IHC experiments were performed according to the steps described previously (Chen et al 2022). The primary antibodies used were anti-FOXD1 (1:200 dilution, abs102294, Absin), anti-FOXD2 (1:200, TA322739, OriGene), anti-FOXD3 (1:200 dilution, abs133773, Absin), and anti-FOXD4 (1:200, orb156929, Biorbyt).…”

“…The total RNA from fresh tissues and cell lines was extracted using RNAiso plus ( Dalian, China). We performed the experiment according to the recommended system of instructions (Chen et al 2022). The primer sequences are listed in Supplementary Table 1.…”