Sodium Butyrate Inhibits the Malignant Proliferation of Colon Cancer Cells via the miR-183/DNAJB4 Axis.

Colorectal carcinoma (CRC) is one of the most common malignant tumors in the digestive tract. It was found that butyric acid could inhibit the expression of miR-183 to slow down malignant progression of CRC in the early stage. However, its regulatory mechanism remains unclear. This study screened the IC50 value of butyrate on inhibition of CRC cells malignant progression. Its inhibitory effects were detected by MTT assay, colony formation experiment, Transwell migration experiment, and apoptosis evaluation by flow cytometry. Next, the expressions of miR-183 and DNAJB4 were, respectively, determined in butyrate treated and miR-183 analog or si-DNAJB4-transfected CRC cells to further detect the role of upregulated miR-183 or silencing DNAJB4 in CRC cells malignant progression. Subsequently, the targeted regulatory relationship between miR-183 and si-DNAJB4 was confirmed by bioinformatic prediction tools and double luciferase report genes analysis method. The regulatory mechanism of butyrate on miR-183/DNAJB4 axis signal pathway was evaluated in molecular level, and verified in nude mouse xerograft tumor model and immunohistochemical analysis tests of Ki67 positive rates. The results displayed that butyrate with increased concentration can hinder the proliferation and improve apoptosis of CRC cells by decreasing the expression of miR-183. Thus, butyrate reduces miR-183 expression and increases DNAJB4 expression via the miR-183/DNAJB4 axis, ultimately inhibiting the malignant progression and increasing apoptosis of CRC. While over expression of miR-183 downregulate the expression of DNAJB4, which can reverse the inhibitory effect of butyrate.

Single-cell sequencing reveals the immune microenvironment landscape related to anti-PD-1 resistance in metastatic colorectal cancer with high microsatellite instability.

BACKGROUND: The objective response rate of microsatellite instability-high (MSI-H) metastatic colorectal cancer (mCRC) patients with first-line anti-programmed cell death protein-1 (PD-1) monotherapy is only 40-45%. Single-cell RNA sequencing (scRNA-seq) enables unbiased analysis of the full variety of cells comprising the tumor microenvironment. Thus, we used scRNA-seq to assess differences among microenvironment components between therapy-resistant and therapy-sensitive groups in MSI-H/mismatch repair-deficient (dMMR) mCRC. Resistance-related cell types and genes identified by this analysis were subsequently verified in clinical samples and mouse models to further reveal the molecular mechanism of anti-PD-1 resistance in MSI-H or dMMR mCRC. METHODS: The response of primary and metastatic lesions to first-line anti-PD-1 monotherapy was evaluated by radiology. Cells from primary lesions of patients with MSI-H/dMMR mCRC were analyzed using scRNA-seq. To identify the marker genes in each cluster, distinct cell clusters were identified and subjected to subcluster analysis. Then, a protein‒protein interaction network was constructed to identify key genes. Immunohistochemistry and immunofluorescence were applied to verify key genes and cell marker molecules in clinical samples. Immunohistochemistry, quantitative real-time PCR, and western blotting were performed to examine the expression of IL-1ß and MMP9. Moreover, quantitative analysis and sorting of myeloid-derived suppressor cells (MDSCs) and CD8+ T cells were performed using flow cytometry. RESULTS: Tumor responses in 23 patients with MSI-H/dMMR mCRC were evaluated by radiology. The objective response rate was 43.48%, and the disease control rate was 69.57%. ScRNA-seq analysis showed that, compared with the treatment-resistant group, the treatment-sensitive group accumulated more CD8+ T cells. Experiments with both clinical samples and mice indicated that infiltration of IL-1ß-driven MDSCs and inactivation of CD8+ T cells contribute to anti-PD-1 resistance in MSI-H/dMMR CRC. CONCLUSIONS: CD8+ T cells and IL-1ß were identified as the cell type and gene, respectively, with the highest correlation with anti-PD-1 resistance. Infiltration of IL-1ß-driven MDSCs was a significant factor in anti-PD-1 resistance in CRC. IL-1ß antagonists are expected to be developed as a new treatment for anti-PD-1 inhibitor resistance.

Erlotinib inhibits colon cancer metastasis through inactivation of TrkB-dependent ERK signaling pathway.

The distal metastasis is the main cause of death in patients with colon cancer. Tyrosine receptor kinase B (TrkB) and ERK signals may be the potential targets for the treatment of colon cancer metastasis. This study aims to investigate whether erlotinib inhibits distant metastasis of colon cancer by regulating TrkB and ERK signaling pathway. Human colon adenocarcinoma cell lines (SW480 and Caco-2) pretreated with exogenous C-X-C motif chemokine ligand 8 (CXCL8) were used to assess the suppressive effect of erlotinib on tumor metastasis, including anoikis, epithelial-mesenchymal transformation (EMT), migration, and invasion. Through TrkB overexpression, Akt suppression, and ERK suppression, the roles of TrkB, Akt, and ERK in erlotinib-induced metastasis inhibition of colon cancer cells were explored. The results showed that erlotinib alleviated CXCL8-induced metastasis of the colon cancer cells. Overexpression of TrkB in colon cancer cells eliminated the effect of erlotinib on anoikis, inhibition of EMT, migration, and invasion, and downregulation of p-ERK and p-Akt. Furthermore, the inhibition of ERK activation instead of Akt activation was found to participate in erlotinib-mediated metastasis resistance, including anoikis, inhibition of EMT, migration, and invasion. In conclusion, erlotinib inhibits colon cancer cell anoikis resistance, EMT, migration, and invasion by inactivating TrkB-dependent ERK signaling pathway.

Identification of lncRNA-mRNA network linking ferroptosis and immune infiltration to colon adenocarcinoma suppression.

Background: Colon adenocarcinoma (COAD) is one of the most common malignant tumors. The interplay involving ferroptosis between tumor and immune cells plays a crucial in cancer progression. However, the biological basis of this interplay in COAD development remains elusive. Methods: Transcriptome data of COAD samples were obtained from The Cancer Genome Atlas and National Center for Biotechnology Information databases. Using single-sample gene set enrichment analysis, we calculated the ferroptosis score (FS) and immune cell infiltration levels for each sample, leveraging the expression levels of genes related to ferroptosis and various immune cell types. Samples with FSs greater than the 75th percentile were classified into the high-FS subgroup, while those below the 25th percentile were categorized as the low-FS subgroup. Moreover, tumor tissue samples and adjacent normal tissue samples were collected from twenty colon patients. Using real-time quantitative polymerase chain reaction, we validated the expression of certain genes in these samples. Results: The COAD samples with high FSs experienced favorable survival probability and heightened sensitivity to anticancer drugs, with FSs negatively associated with the pathological stages. Moreover, the up-regulated genes in high-FS subgroup exhibited enrichment in immune-related pathways, suggesting a correlation between immunity and ferroptosis. Importantly, we discovered a key lncRNA-mRNA co-expression network linking tumor cell ferroptosis and immune infiltration (e.g., neutrophil) in the progression and classification of COAD. Further analysis identified several ferroptosis-related lncRNAs (e.g., RP11-399O19.9) within this network, indicating their potential roles in COAD progression and deserving in-depth study. Conclusions: Our findings provide novel insights into the underlying biological basis, particularly involving lncRNAs, at gene expression level associated with ferroptosis in COAD and cancer therapy. Nevertheless, further analysis and validation are required to expand the findings.

Abnormal Expression of N6-Methyladenosine RNA Methylation Regulator IGF2BP3 in Colon Cancer Predicts a Poor Prognosis.

The value of insulin-like growth factor 2 mRNA-binding protein 3 (IGF2BP3), an N6-methyladenosine (m6A) RNA methylation regulatory factor, in the prognosis of colon cancer was still unclear. High levels of IGF2BP3 were expressed in colon adenocarcinoma (COAD) samples and in human colon cancer tissues, which was associated with poorer overall survival (OS). We validated IGF2BP3 as an independent prognostic risk biomarker in COAD patients. Moreover, functional enrichment analysis suggested that differentially expressed genes (DEGs) of groups with high versus low IGF2BP3 expression were related to immune- and cancer-related pathways. Furthermore, the tumor microenvironments of high- versus low-IGF2BP3 expression groups showed significant differences and IGF2BP3 predicted the efficiency of immunotherapy. Finally, protein-protein interaction network analysis suggested that there was a direct or indirect interaction among IGF2BP3, WNT7B, VANGL2, NKD1, AXIN2, RNF43, and CDKN2A. In brief, IGF2BP3 was confirmed as an independent prognostic signature in COAD patients and might be a therapeutic target in this study. Moreover, IGF2BP3 could be used in personalized immunotherapy for COAD.

Circ_0026344 restrains metastasis of human colorectal cancer cells via miR-183.

Background: CircRNA circ_0026344 was previously revealed as a tumour-suppressive gene in colorectal cancer (CRC) progression. The purpose of this research was to investigate the role of circ_0026344 in CRC cells metastasis induced by chemokines. Methods: Two human CRC cell lines SW480 and Caco-2 were treated by CCL20 and CXCL8. Cell proliferation, migration/invasion, expression of epithelial-mesenchymal transition (EMT) inducers and the expression of circ_0026344 were measured using sulforhodamine B assay, Transwell chamber, western blot and qRT-PCR, respectively. The effects of circ_0026344 on CRC cells migration/invasion and the expression of EMT inducers were evaluated. Moreover, the downstream miRNA and signalling pathways of circ_0026344 were studied. Results: CCL20 and CXCL8 synergized to facilitate the proliferation, migration and invasion of CRC cells. At the meantime, E-cadherin was downregulated, whereas N-cadherin, Vimentin and Snail were up-regulated by CCL20 and CXCL8 co-stimulation, which was accompanied by the mobilization of PI3K/AKT/ERK signalling. More interestingly, the expression of circ_0026344 was down-regulated by CCL20 and CXCL8 co-stimulation. Silence of circ_0026344 increased the migratory and invasive capacities of CRC cells and increased EMT process as well. Overexpression of circ_0026344 led to a contrary impact. miR-183 was negatively regulated by circ_0026344, and the inhibitory effects of circ_0026344 overexpression on Wnt/ß-catenin pathway were reversed when miR-183 was overexpressed. Conclusion: Overexpression of circ_0026344 restrained CRC metastasis and EMT induced by CCL20 and CXCL8 synergistical treatment. miR-183 was a downstream effector of circ_0026344, and the anti-tumour function of circ_0026344 might be involved in the repressed Wnt/ß-catenin signalling. Highlights CCL20 and CXCL8 synergize to decrease the expression of circ_0026344; Silence of circ_0026344 promotes CRC cells migration, invasion and EMT process; miR-183 is a downstream effector of circ_0026344.

LINC00152 is a potential biomarker involved in the modulation of biological characteristics of residual colorectal cancer cells following chemoradiotherapy.

Concurrent radiotherapy and chemotherapy is a widely used, comprehensive treatment for rectal cancer. By studying the impact of concurrent chemoradiotherapy on the invasion and migration of colorectal cancer (CRC) cells and researching the associated molecular mechanisms, the present study aimed to provide a novel method to improve the therapeutic effect of this treatment against CRC. Human HCT116 and HT29 CRC cells were simultaneously treated with 4 Gy of 6 MV X-rays and 10 µmol/l 5-fluorouracil to establish a residual cell model. Transwell migration and invasion experiments were used to analyse the invasion and migration of the cells. The expression of long non-coding (lnc)RNAs was detected using a gene chip, and reverse transcription-quantitative polymerase chain reaction analysis was used to determine lncRNA expression levels. Specific small interfering RNAs were transfected into HCT116 residual cells to silence the expression of the identified key genes. The migration and invasion of residual CRC cells were demonstrated to be significantly increased compared with the original cells. Pvt1 oncogene, long-chain non-protein-coding RNA 152 (LINC00152), and MIR22 host gene were selected as potential targets. However, the migration and invasion of residual HCT116 cancer cells were only significantly decreased following silencing of LINC00152 expression. LINC00152 may therefore be a potential biomarker involved in modulation of the biological characteristics of residual CRC cells following chemoradiotherapy.