Comprehensive analysis of bulk and single-cell RNA sequencing data reveals Schlafen-5 (SLFN5) as a novel prognosis and immunity

Abstract

Recent advancements have elucidated the multifaceted roles of the Schlafen (SLFN) family, including SLFN5, SLFN11, SLFN12, SLFN13, and SLFN14, which are implicated in immunological responses. However, little is known about the roles of this gene family in relation to malignancy development. The current study aimed to explore the diagnostic and prognostic potential of Schlafen family genes in colorectal adenocarcinoma (COAD) through bioinformatics analysis. Leveraging advanced bioinformatics tools of bulk RNA-sequencing and single-cell sequencing, we conducted in-depth analyses of gene expressions, functional enrichment, and survival patterns of patients with colorectal cancer compared to normal tissue. Among Schlafen family genes, the transcription levels of SLFN5 in COAD tissues were significantly elevated and correlated with poor survival outcomes. Furthermore, SLFN5 regulated the immune response via Janus kinase (JAK)/signal transduction and activator of transcription (STAT)/interferon (IFN)-alpha/beta signaling. These chemokines in inflammation are associated with diabetes and metabolism, suggesting their involvement in altered cellular energetics for COAD progress. In addition, an immune cell deconvolution analysis indicated a correlation between SLFN5 expression and immune-related cell populations, such as regulatory T cells (Tregs). These findings highlighted the potential clinical significance of SLFN5 in COAD and provided insights into its involvement in the tumor microenvironment and immune regulation. Meanwhile, the drug discovery data of SFLN5 with potential targeted small molecules suggested its therapeutic potential for COAD. Collectively, the current research demonstrated that SFLN5 play crucial roles in tumor development and serve as a prospective biomarker for COAD.

Keywords: Schlafen (SLFN) family genes; cell metabolism; colorectal adenocarcinoma (COAD); immune infiltration; single cell technology; tumor microenvironment.

Figure 1

Overview of the workflow of the study.

Figure 2

Pan-cancer analysis of Schlafen (SLFN) family genes from TCGA patients. (A) Transcript levels of SLFN family genes in normal and cancer tissues from TCGA patients via the UALCAN platform. (B) Box plots indicate SLFN family mRNA expression in TCGA dataset, and only SLFN5 had high expression levels in COAD patients (p < 0.05 was considered significant).

Figure 3

Schlafen 5 (SLFN5) plays an oncogenic role in COAD development. (A-B) SLFN5 promoter methylation levels in COAD were determined by a patient's tumor stage (p < 0.05). (C) Kaplan-Meier plot of SLFN5 expression in COAD patients for an overall survival analysis from TCGA patients via the UALCAN platform. (D) Analysis of SLFN5 in COAD specimens using the Human Protein Atlas (HPA). Protein levels of SLFN5 were examined in clinical COAD specimens from the HPA. IHC images of SLFN5 illustrate staining intensities. The HPA provided IHC images and patient information, including both normal and tumor samples. Bar charts present quantified IHC staining in COAD specimens, allowing for a comparative analysis.

Figure 4

IHC expression of Schlafen 5 (SLFN5) in patients at different stages of colorectal adenocarcinoma (COAD) at Kaohsiung Armed Forces General Hospital. (A) Immunohistochemical staining showing the expression of SLFN5 in normal colon tissues, and COAD tissues at different stages (I-IV). (B-C) Quantitative analysis of SLFN5 positive cell rates in normal and various stages of COAD. The bar graph represents the percentage of positive cells in each group: Normal (n=4), Stage I (n=13), Stage II (n=15), Stage III (n=14), and Stage IV (n=6). Statistical significance was noted with **p<0.001 and ***p<0.001.

Figure 5

Analysis of Schlafen 5 (SLFN5) in tumor microenvironment characteristics in colorectal adenocarcinoma (COAD). (A) Heatmap shows SLFN5 expression (on top) and well-known cell-type markers in different single cell-type clusters of this tissue. The panel on the left shows which cell type each marker is associated with. Color coding is based on cell type groups, each consisting of cell types with common functional features. (B) Correlations of SLFN5 in a pan-cancer analysis. (C) RNA expressions in the single-cell type clusters identified in this tissue were visualized by a UMAP plot (left) and a bar chart (right). (D) Immune cell expression of SLFN5 as obtained from the Human Protein Atlas.

Figure 6

DNA methylation of Schlafen 5 (SLFN5), and transcript expression levels of SLFN5 in different colorectal cancer cell lines (A) Heatmap of DNA methylation expression levels of SLFN5 in TCGA colorectal adenocarcinoma (COAD) patients. (B) RNA-sequencing analysis of SLFN5 mRNA in different colorectal cancer cell lines from the Cancer Cell Line Encyclopedia (CCLE). For each cancer cell line, high SFLN5 expression is indicated in red (left), while low expression is indicated in blue (right).

Figure 7

Hallmark signaling pathway analysis of Schlafen 5 (SLFN5) in colorectal adenocarcinoma (COAD). TCGA colorectal cancer patients were stratified into two groups based on the median SLFN5 expression value. A gene set enrichment analysis (GSEA) was then performed on these two groups using the Hallmark database. Results of the analysis revealed relevant enriched pathways in COAD groups with elevated SLFN5 expression.

Figure 8

Functional and pathway enrichment analyses were performed specifically on genes co-expressed with Schlafen 5 (SLFN5) in colorectal adenocarcinoma (COAD) TCGA patients. (A) The analysis involved gene ontology (GO) terms, including biological processes, cellular components, and molecular functions. Circle sizes in the visual representation of the results indicate the number of genes associated with each function, while colors of the bubbles correspond to p values, providing information about the statistical significance of the enrichment. (B) Pathway analysis of SLFN5 co-expressed genes in the KEGG database.

Figure 9

MetaCore pathway analysis of Schlafen 5 (SLFN5) co-expressed genes in COAD patients from TCGA. (A) The MetaCore pathway enrichment analysis was conducted for genes co-expressed with SLFN5 in COAD patients, revealing potential pathways involving these genes ranked by their log p values. (B) The "Immune response_IFN-alpha/beta signaling via JAK/STAT" is highlighted, with symbols representing proteins and arrows indicating protein interactions (green for activation and red for inhibition). Thermometer-like histograms visually represent microarray gene expressions, with blue indicating downregulation and red indicating upregulation.

Figure 10

Drug sensitivity of Schlafen 5 (SLFN5) oncogene from GSCA. (A) Correlation between Genomics of Drug Sensitivity in Cancer (GDSC) data for FDA-approved drugs according to the SLFN5 expression levels. (B) Correlation of Cancer Therapeutics Response Portal (CTRP) drug data and SLFN5.