Colon Cancer Tissues Research Articles

Abstract PR016: Dynamic chromatin landscapes of colorectal cancer development and cell-free DNA fragmentation

Abstract The stepwise progression of colorectal cancer (CRC) from healthy epithelium, to premalignant adenoma, to cancer is accompanied by genome-wide epigenetic reprogramming which may be reflected in circulating cell-free DNA (cfDNA). However, current analyses of these processes have been hampered by complex mixtures of cells in normal, adenoma and cancer tissue samples, as well as absence of suitable adenoma model systems. Cultured colorectal organoids, comprised of pure epithelial cells, could enable insights on the epigenetic changes leading to progression from healthy tissue to cancer. Such features could be instrumental in the early cancer detection using genome-wide analyses of cfDNA fragmentation. Using 43 patient-derived colorectal organoids derived from healthy, adenoma, and cancer tissues, as well as 10 freshly collected peripheral blood mononuclear cell (PBMC) control samples, we examined the chromatin landscape of tumor progression using transposase accessible chromatin analyses with next-generation sequencing. We defined a consensus set of nucleosome depleted regions for each disease state and identified loci with differential accessibility between PBMCs, colon healthy, colon adenoma, and colon cancer tissues. We analyzed these differential loci for fragmentation characteristics in the circulating cfDNA of 250 healthy individuals and 51 patients with metastatic CRC. Across all organoid samples, we identified >35 million nucleosome depleted regions that we coalesced into a consensus set of 62,708 regions. As 78% of these regions could be linked to genes, we assessed whether these signatures identify known pathways involved in colorectal cancer. Within these regions, we identified unique colon tumor-related signatures comprising genes of known and novel pathways, including those involved in WNT, hippo, and RAS signaling. While the majority of chromatin accessibility differences were shared between adenomas and cancers, we identified 895 chromatin changes in adenomas that were not present in either healthy or malignant tissues. In healthy individuals, we found that the cfDNA coverage was negatively correlated with accessibility of white blood cell accessibility loci (Spearman ρ= - 0.60) while in individuals with cancer, changes in chromatin accessibility were associated with independent circulating tumor DNA abundance by droplet digital PCR (R2=0.69, p<0.001). Chromatin accessibility profiles were correlated with genome-wide cell-free DNA fragmentation features and could be used to distinguish healthy individuals from those with colorectal cancer (AUC=0.97). These analyses are consistent with dynamic chromatin remodeling during colorectal tumor development and may provide an avenue for evaluating these through genome-wide analyses of cfDNA fragmentation. Citation Format: Nicholas A. Vulpescu, Zachariah H. Foda, Pieter H.A. Wisse, Christopher Cherry, Jaime E. Medina, Vilmos Adleff, Remond J.A. Fijneman, Robert B. Scharpf, Gerrit A. Meijer, Beatriz Carvalho, Victor E. Velculescu. Dynamic chromatin landscapes of colorectal cancer development and cell-free DNA fragmentation [abstract]. In: Proceedings of the AACR Special Conference: Liquid Biopsy: From Discovery to Clinical Implementation; 2024 Nov 13-16; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(21_Suppl):Abstract nr PR016.

Multi-omics analysis of the biological function of the VEGF family in colon adenocarcinoma.

The vascular endothelial growth factor (VEGF) family plays a crucial role in cancer progression, but the prognostic significance and biological functions of VEGF family members in colon adenocarcinoma (COAD) remain unclear. Using data from The Cancer Genome Atlas, Gene Expression Omnibus, Gene Set Cancer Analysis, cBioPortal, GeneMANIA, String, MethSurv and starBase database, we identified vascular endothelial growth factor B (VEGFB) as a key gene associated with COAD prognosis, with its abnormal expression linked to methylation dysregulation. In vitro experiments confirmed VEGFB expression was significantly higher in colon cancer tissues compared to normal tissues, as shown by Real-time quantitative PCR and immunohistochemistry. Cell Counting Kit-8 and colony formation assay showed that decreased VEGFB expression in SW480 cells resulted in decreased cell viability and proliferation ability. Scratch assay showed that VEGFB downregulation impaired SW480 cell migration. In addition, our research suggests that VEGFB not only promotes angiogenesis but is also involved in the tumor microenvironment and immune regulation. The SHNG17-miR-375-VEGFB regulatory axis provides a potential therapeutic target for COAD, highlighting VEGFB's role in immune activation during anti-angiogenic therapy and potential reversal of drug resistance.

SIRT2 deacetylates and decreases the expression of FOXM1 in colon cancer.

New FOXM1-specific inhibitors with the potential to be used for therapeutic purposes are under extensive research. We hypothesized that deacetylation of FOXM1 would decrease protein expression, thus providing novel therapeutic management of colon cancers. Immunostaining was used to determine FOXM1 and SIRT2 expressions in human colon cancer tissue microarrays (n = 90) from Stage I to Stage IV. SIRT2-FOXM1 interaction was evaluated in colon cancer cells using immunoprecipitation. Deacetylation of FOXM1 via SIRT2 was determined using in vitro deacetylation assays. FOXM1 could be hyper-acetylated when p300 and pCAF histone acetyltransferases were administered alongside deacetylase inhibitors. We detected that SIRT2 and FOXM1 physically interacted, and SIRT2 deacetylated FOXM1 in vitro. SIRT2 overexpression led to a significant decrease while knockdown of SIRT2 increased the FOXM1 expression in HCT116 human colon carcinoma cells. In the analysis of 90 human colorectal cancer samples, high SIRT2 expression was observed in about 49% of colorectal cancer, intermediate in 29%, and low or no staining in 22%. Strong SIRT2 expression was found to be negatively associated with the FOXM1 staining in our clinical cohort. This study reveals a molecular interaction and association between SIRT2 and FOXM1 expression in colon cancer cell lines and human colon cancer samples, and suggests that targeting SIRT2 activity using small molecule modulators may be a promising therapeutic approach for colorectal cancer.

The eIF3a translational control axis in the Wnt/β-catenin signaling pathway and colon tumorigenesis

Translational initiation in protein synthesis is an important regulatory step in gene expression and its dysregulation may result in diseases such as cancer. Translational control by eIF4E/4E-BP has been well studied and contributes to mTOR signaling in various biological processes. Here, we report a novel translational control axis in the Wnt/β-catenin signaling pathway in colon tumorigenesis by eIF3a, a Yin-Yang factor in tumorigenesis and prognosis. We show that eIF3a expression is upregulated in human colon cancer tissues, pre-cancerous adenoma polyps, and associates with β-catenin level and APC mutation in human samples, and that eIF3a overexpression transforms intestinal epithelial cells. We also show that eIF3a expression is regulated by the Wnt/β-catenin signaling pathway with an active TCF/LEF binding site in its promoter and that eIF3a knockdown inhibits APC mutation-induced spontaneous colon tumorigenesis in APCmin/+ mice. Together, we conclude that eIF3a upregulation in colon cancer is due to APC mutation and it participates in colon tumorigenesis by adding a translational control axis in Wnt/β-catenin signaling pathway and that it can serve as a potential target for colon cancer intervention.

Increased MAGE-C Family Gene Expression Levels as a Biomarker of Colon Cancer Through the Demethylation Mechanism

Background/Objectives: Colon cancer (CC) in Saudi Arabia is associated with a high death rate and is commonly identified at a more progressive stage. Therefore, it is critical to identify and characterize potential novel cancer-specific biomarkers to enhance early CC diagnosis. The goal was to assess their potential use as cancer biomarkers for the early detection and improvement of CC treatment. Methods: MAGE-C1, MAGE-C2, and MAGE-C3 family gene expression levels were examined using RT-PCR and qRT-PCR assays in 26 adjacent normal colon (NC) and CC tissue samples from male and female Saudi patients. Using several cell lines and the qRT-PCR technique, epigenetic control was also investigated to determine whether reduced treatment with 5-aza-2′-deoxycytidine, which reduces DNA methyltransferase, can increase the expression of the MAGE-C gene. The expression levels, promoter methylation, and prognostic significance of MAGE-C1, MAGE-C2, and MAGE-C3 genes across various cancers were analyzed using The Cancer Genome Atlas (TCGA) data. Additionally, the prognostic significance of these genes was assessed through Kaplan–Meier survival analysis. Results: The RT-PCR results showed that MAGE-C1, MAGE-C2, and MAGE-C3 gene expressions were significantly higher in the CC and NC tissues. The MAGE-C1 expression level was the highest in CC tissues (p < 0.0001), followed by MAGE-C3 (p = 0.0004) and MAGE-C2 (p = 0.0020) in descending order. The 5-aza-2′-deoxycytidine treatment significantly increased the mRNA expression levels of the MAGE-C1, MAGE-C2, and MAGE-C3 genes in HCT116, Caco-2, MCF-7, and MCF-10A cells. Expression analyses of TCGA samples revealed significant upregulation of these genes in several cancer types, with notable differences between normal, tumor, and metastatic tissues. Promoter methylation indicates hypomethylation in cancerous tissues. Survival analyses show that high expression levels of MAGE-C1 correlate with better prognosis, while MAGE-C3 is associated with poorer outcomes. Conclusions: These results demonstrate that MAGE-C genes are viable prospective biomarkers of CC controlled by hypomethylating drugs, consequently offering a possible treatment target for CC in a specific population.

Solute carrier family 4 member 4 (SLC4A4) is associated with cell proliferation, migration and immune cell infiltration in colon cancer

BackgroundSolute Carrier Family 4 Member 4 (SLC4A4) is a membrane protein‐coding gene for a Na+/HCO3− cotransporter and plays a crucial role in regulating pH, bicarbonate secretion and homeostasis. However, the prognostic and immunological role of SLC4A4 in colon cancer remains unknown.MethodIn this study, expression profiles of SLC4A4 were retrieved from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO) databases, to which a variety of bioinformatic analyses were performed. Sangerbox, Xiantao, ESTIMATE and TIMER online tools were used to delve into the relationship between SLC4A4 expression and immune cell infiltration. The role of SLC4A4 in the proliferation and migration of colon cancer cells was verified by CCK8, EdU and wound healing assays. The related molecules and pathways that SLC4A4 may affect were validated by bioinformatic prediction and western blotting analysis.ResultsThe expression levels of SLC4A4 were significantly lower in colon cancer tissues than in normal tissues and its low expression was positively correlated with poor prognosis. TIMER and ESTIMATE showed that SLC4A4 broadly influenced immune cell infiltration. Experiments in vitro demonstrated that SLC4A4 inhibited partial epithelial-mesenchymal transition (EMT) phenotypes.ConclusionsTo conclude, our study revealed that SLC4A4 is lowly expressed in colon cancer tissues, and SLC4A4 may inhibit the progression of colon cancer via regulating partial EMT phenotypes and immune cell infiltration, which may provide new perspectives for the development of more precise and personalized immune anti-tumor therapies.

MiR‑106b‑5p in stage II left‑sided and right‑sided colon cancer and its association with the prognostic characteristics of patients.

MicroRNA (miR)-106b-5p is highly expressed in colon cancer; however, data on its expression levels in left-sided colon cancer (LCC) vs. right-sided colon cancer (RCC) is lacking. The present study aimed to assess the differences in miR-106b-5p expression in stage II LCC and RCC, as well as its relationship with patient prognosis. From August 2018 to February 2020, 40 specimens of primary stage II colon cancer were collected from Huizhou First Hospital (Huizhou, China), which included 20 cases of LCC and 20 cases of RCC. The miR-106b-5p expression levels in cancer tissues were compared with normal adjacent tissues, as well as between LCC and RCC tissues, and survival outcomes were assessed. miR-106b-5p expression was significantly higher in stage II LCC tissues compared with RCC tissues. However, no significant difference in 5-year survival was observed between the two groups. Notably, 5-year survival was significantly lower in the high miR-106b-5p expression group compared with the low expression group among patients with RCC. By contrast, there were no survival differences between the high and low miR-106b-5p expression groups in LCC. Multivariate analysis indicated that miR-106b-5p expression was an independent prognostic factor for patients with RCC. In conclusion, miR-106b-5p expression was significantly upregulated in colon cancer tissues, with higher expression levels demonstrated in LCC compared with RCC. High miR-106b-5p expression in RCC was identified as an independent prognostic factor, whilst its expression in LCC did not show a significant association with prognosis.

Augmented histopathology: Enhancing colon cancer detection through deep learning and ensemble techniques.

Colon cancer poses a significant threat to human life with a high global mortality rate. Early and accurate detection is crucial for improving treatment quality and the survival rate. This paper presents a comprehensive approach to enhance colon cancer detection and classification. The histopathological images are gathered from the CRC-VAL-HE-7K dataset. The images undergo preprocessing to improve quality, followed by augmentation to increase dataset size and enhance model generalization. A deep learning based transformer model is designed for efficient feature extraction and enhancing classification by incorporating a convolutional neural network (CNN). A cross-transformation model captures long-range dependencies between regions, and an attention mechanism assigns weights to highlight crucial features. To boost classification accuracy, a Siamese network distinguishes colon cancer tissue classes based on probabilities. Optimization algorithms fine-tune model parameters, categorizing colon cancer tissues into different classes. The multi-class classification performance is evaluated in the experimental evaluation, which demonstrates that the proposed model provided highest accuracy rate of 98.84%. In this research article, the proposed method achieved better performance in all analyses by comparing with other existing methods. RESEARCH HIGHLIGHTS: Deep learning-based techniques are proposed. DL methods are used to enhance colon cancer detection and classification. CRC-VAL-HE-7K dataset is utilized to enhance image quality. Hybrid particle swarm optimization (PSO) and dwarf mongoose optimization (DMO) are used. The deep learning models are tuned by implementing the PSO-DMO algorithm.

LncRNA HAND2-AS1 Inhibited Colon Cancer Progression By Regulating miR-3118/ZG16 Axis.

LncRNA HAND2-AS1 is a novel cancer regulator, but the role and mechanisms of HAND2-AS1 involved with colon cancer (CC) progression remains unknown. The purpose of this research was to figure out how HAND2-AS1 regulates the progression of CC. Using qRT-PCR, we studied expression levels of miR-3118, HAND2-AS1, and ZG16 in CC tissues and cells. Protein levels of apoptosis-related proteins (Bax and Bcl-2) and ZG16 were quantified by western blotting. In vitro function analysis referred to western blotting, wound healing assay and CCK-8. The binding association among miR-3118, HAND2-AS1, and ZG16 was investigated using luciferase reporter and RIP assays. The functional role of HAND2-AS1 was analyzed using xenograft tumor models in vivo. In tissues and cells of CC, HAND2-AS1 was downregulated. We observed that HAND2-AS1 overexpression declined CC cell proliferation and migration while facilitating apoptosis. We further verified that when HAND2-AS1 is overexpressed it reduced CC tumor development in vivo. In CC cells and tissues, miR-3118 competed with HAND2-AS1 and was elevated. Further it was noted that the HAND2-AS1 when overexpressed, lessened the survival of CC cells, however overexpression of miR-3118 restored these changes. ZG16 was shown to be a target of miR-3118, it was found that ZG16 was downregulated in CC tissue and cells. We observed, high expression of ZG16 partially restored the enhanced malignant phenotype caused by miR-3118 overexpression. HAND2-AS1 inhibited CC progression by upregulating ZG16 expression through sponging miR-3118. Hence, HAND2-AS1/miR-3118/ZG16 axis could be a possible new target for CC treatment.

Covalent assembly-based two-photon fluorescent probes for in situ visualizing nitroreductase activities: From cancer cells to human cancer tissues

Nitroreductase (NTR) is widely regarded as a biomarker whose enzymatic activity correlates with the degree of hypoxia in solid malignant tumors. Herein, we utilized 2-dimethylamino-7-hydroxynaphthalene as fluorophore linked diverse nitroaromatic groups to obtain four NTR-activatable two-photon fluorescent probes based on covalent assembly strategy. With the help of computer docking simulation and in vitro assay, the sulfonate-based probe XN3 was proved to be able to identify NTR activity with best performances in rapid response, outstanding specificity, and sensitivity in comparison with the other three probes. Furthermore, XN3 could detect the degree of hypoxia by monitoring NTR activity in kinds of cancer cells with remarkable signal-to-noise ratios. In cancer tissue sections of the breast and liver in mice, XN3 had the ability to differentiate between healthy and tumorous tissues, and possessed excellent fluorescence stability, high tissue penetration and low tissue autofluorescence. Finally, XN3 was successfully utilized for in situ visualizing NTR activities in human transverse colon and rectal cancer tissues, respectively. The findings suggested that XN3 could directly identify the boundary between cancer and normal tissues by monitoring NTR activities, which provides a new method for imaging diagnosis and intraoperative navigation of tumor tissue.

NR3C2 affects the proliferation and invasiveness of colon cancer cells through the Wnt/β-Catenin signaling pathway

PurposeThe aim of this study was to explore the potential correlation between the nuclear receptor subfamily 3 group C member 2 (NR3C2) and outcomes of colon cancer, along with the mechanisms underlying this association.MethodmRNA (messenger RNA) data and clinical records pertaining to colon cancer were retrieved from The Cancer Genome Atlas (TCGA) database. The analysis of NR3C2 expression discrepancies between normal colon and tumor tissues was conducted using R software. In addition, we also studied the relationship between NR3C2 expression and prognosis, pathological parameters. The relative role of NR3C2 were further predicted through bioinformatics methods and receiver operating characteristic (ROC) curve was used to evaluate the diagnostic value of NR3C2 in colon cancer. Single-cell data from colon cancer samples in the GEO (Gene Expression Omnibus) database further investigated the mechanism of the lower survival associated with NR3C2 dysregulation. NR3C2 expression in three fresh colon cancer samples and their respective paracancer samples was determined. Furthermore, colon cancer cell models overexpressing NR3C2 and with knockdown NR3C2 were constructed by lentiviral vector transfection. Cell Counting Kit-8 assay, transplantation of tumors in nude mice and transwell assays were used to examine the proliferation, migration and invasion of colon cancer cells. The effect on the Wnt/β-catenin pathway, activities of cellular autophagy and cell apoptosis were examined by assessing the expression levels of several key proteins, including Bcl-2, Bax, and LC3.ResultsWe found that NR3C2 was found a significantly lower level in colon cancer tissues than in adjacent tissues, which was associated with distant and lymphatic metastases, clinical stage, and poor clinical outcome, and it was an independent prognostic factor and potential marker of colon cancer. Single-cell transcriptome data identified the subset of circulating T and B cells with high expression of NR3C2, which is involved in TNF signaling pathway. Functional experiments show that downregulation of NR3C2 resultsed in the activation of the Wnt/β-catenin signaling pathway, and promotesd the proliferation and invasion of colon cancer cells while suppressing cell autophagy and apoptosis.ConclusionNR3C2 may regulate Wnt/β-catenin to affect the proliferation, invasion apoptosis and autophagy of colon cancer, and this axis is a potential target for the treatment of colon cancer.

The Role of miR-132-3p in Inhibiting the Proliferation, Invasion and Metastasis of Colon Cancer by Targeted Regulation of PDGF

This study investigated the impact of miR-132-3p on colon cancer cell behavior by targeting PDGF, offering insights into potential approaches for understanding colon cancer development and gene-targeted therapy. The study involved qRT-PCR analysis to assess miR-132-3p expression in colon cancer and adjacent tissues. Overexpression of miR-132-3p in HCT-116 cells was examined through CCK-8 assays for proliferation, flow cytometry for apoptosis, and Transwell/scratch assays for invasion/metastasis. Pearson correlation analysis evaluated miR-132-3p-PDGF relationship, validated via luciferase assays. qRT-PCR/Western blot assessed PDGF mRNA/protein levels. In vivo tumorigenesis assays in nude mice confirmed miR-132-3p’s inhibitory effect on HCT-116 cells. Results showed reduced miR-132-3p expression in colon cancer tissues (vs. adjacent tissues), correlating with decreased cell proliferation and invasion upon miR-132-3p overexpression. Luciferase activity confirmed PDGF as a miR-132-3p target. Additionally, miR-132-3p inhibited tumor growth, while miR-132-3p+PDGF reversed these effects. In conclusion, miR-132-3p regulates PDGF, suppressing colon cancer cell proliferation, invasion, and metastasis, thereby promoting apoptosis. This highlights the potential of miR-132-3p as a therapeutic target for colon cancer treatment.

Cystathionine-β-synthase expression correlates with tumour progression and adverse prognosis in patients with colon cancer.

To investigate the levels of cystathionine-β-synthase (CBS) in colon cancer tissues compared with adjacent control tissues; and to examine the relationship between CBS level and clinical characteristics and prognosis. This retrospective study enrolled patients with primary colon cancer. Paraffin-embedded specimens were used to create pathological tissue microarrays. Immunohistochemistry was performed on the microarray to detect the levels of CBS in colon cancer tissues and normal adjacent tissues. Analyses were undertaken to examine the relationship between the level of CBS and clinical characteristics and prognosis. A total of 216 patients (107 males and 109 females) were included in the study. The level of CBS in cancer tissues was found to be significantly increased compared with normal adjacent control tissues. There were significant differences in tumour location, tumour-node-metastasis stage and survival rate between the CBS-negative and CBS-positive groups. Positive CBS immunostaining was associated with decreased survival in colon cancer patients. The results of multivariate Cox regression analysis revealed that tumour location and positive CBS immunostaining were independent prognostic factors for survival. Positive CBS immunostaining was closely associated with colon cancer and high levels of CBS might accelerate tumour development and affect patient prognosis in colon cancer.

A label-free and signal-amplifiable fluorescent biosensor based on aptamer-conjugated gold nanoparticles and hybridization chain reaction for determination of carcinoembryonic antigen.

The sensitive detection of cancer biomarkers is crucial for early accurate diagnostics and therapy of cancer patients. Carcinoembryonic antigen (CEA) is a tumor-associated antigen derived from colon cancer and embryonic tissues. In this study, we have developed a label-free fluorescence biosensing platform for the quantification of CEA with the "turn-on" signal output. This platform employs a label-free strategy that incorporates an aptamer-modified gold nanoparticle (Apt@AuNP) probe for the recognition of CEA, in combination with hybridization chain reaction (HCR) amplification. In the presence of target CEA, Apt@AuNPs selectively capture CEA, resulting in a reduction of subsequent complementary chains (CP) binding on Apt@AuNPs. The remaining CP, acting as the initiator sequence for HCR, triggers the HCR, leading to the formation of abundant G-quadruplex structures. By employing Thioflavin T (ThT) for the formation of G-quadruplex/ThT complexes, the biosensor exhibits a significant enhancement of the fluorescence signal. Under optimized conditions, the biosensor platform demonstrates a limit of detection of 0.03 nM and a linear range from 0.1 to 2.5nM. Additionally, the specificity investigation reveals the high selectivity of this fluorescent biosensor. Finally, the performance of this method has been validated by successfully detecting CEA in real-life samples, highlighting its potential for clinical applications.

OTU deubiquitinase, ubiquitin aldehyde binding 2 (OTUB2) modulates the stemness feature, chemoresistance, and epithelial-mesenchymal transition of colon cancer via regulating GINS complex subunit 1 (GINS1) expression

BackgroundColon cancer is one of the most prevalent tumors in the digestive tract, and its stemness feature significantly contribute to chemoresistance, promote the epithelial-mesenchymal transition (EMT) process, and ultimately lead to tumor metastasis. Therefore, it is imperative for researchers to elucidate the molecular mechanisms underlying the enhancement of stemness feature, chemoresistance, and EMT in colon cancer.MethodsSphere-formation and western blotting assays were conducted to assess the stemness feature. Edu, flow cytometry, and cell viability assays were employed to evaluate the chemoresistance. Immunofluorescence and western blotting assays were utilized to detect EMT. Immunoprecipitation, ubiquitination, agarose gel electrophoresis, chromatin immunoprecipitation followed by quantitative PCR (chip-qPCR), and dual luciferase reporter gene assays were employed for mechanistic investigations.ResultsWe demonstrated a markedly higher expression level of OTUB2 in colon cancer tissues compared to adjacent tissues. Furthermore, elevated OTUB2 expression was closely associated with poor prognosis and distant tumor metastasis. Functional experiments revealed that knockdown of OTUB2 attenuated stemness feature of colon cancer, enhanced its sensitivity to oxaliplatin, inhibited its EMT process, ultimately reduced the ability of tumor metastasis. Conversely, overexpression of OTUB2 exerted opposite effects. Mechanistically, we identified OTUB2 as a deubiquitinase for SP1 protein which bound specifically to SP1 protein, thereby inhibiting K48 ubiquitination of SP1 protein. The SP1 protein functioned as a transcription factor for the GINS1, exerting its regulatory effect by binding to the 1822–1830 region of the GINS1 promoter and enhancing its transcriptional activity. Ultimately, alterations in GINS1 expression directly regulated stemness feature, chemosensitivity, and EMT progression in colon cancer.ConclusionCollectively, the OTUB2/SP1/GINS1 axis played a pivotal role in driving stemness feature, chemoresistance, and EMT in colon cancer. These results shed new light on understanding chemoresistance and metastasis mechanisms involved in colon cancer.

Hsa-miR-181a-2-3p inhibits the oncogenicity of colon cancer by directly targeting STING.

Colon cancer is a common malignant tumor of the gastrointestinal system, which is characterized by high morbidity and mortality. The purpose of this study was to analyze the expression and biological role of miR-181a-2-3p in colon cancer and to investigate the molecular mechanism of its regulatory effect on colon cancer through stimulator of interferon genes (STING). Real-time reverse transcription polymerase chain reaction (qRT-PCR) assay was used to detect the expression of miR-181a-2-3p in colon cancer cell lines and normal intestinal epithelial cells. After overexpression of miR-181a-2-3p in colon cancer cell lines SW480 and HT29, cells were examined by CCK8, Transwell, and flow cytometry assays for alterations in proliferation, migration, apoptosis, and cell cycle. Target genes of miR-181a-2-3p were predicted by bioinformatics and validated by dual luciferase assays. Rescue experiments were performed to explore the role of STING in the effect of miR-181a-2-3p. The effect of miR-181a-2-3p on colon cancer proliferation in vivo was validated by nude mouse tumorigenicity assay. miR-181a-2-3p was lowly expressed in both colon cancer tissues and cell lines. Overexpression of miR-181a-2-3p led to reduced proliferation and migration, increased apoptosis, and altered cell cycle in colon cancer cell lines SW480 and HT29. STING was a target gene of miR-181a-2-3p. Increased STING expression partially counteracted the effect of overexpression of miR-181a-2-3p on colon cancer cell lines. miR-181a-2-3p also suppressed colon cancer proliferation in vivo. miR-181a-2-3p inhibits the proliferation and oncogenicity of colon cancer, and its molecular mechanism could be inhibited by STING.

RBM25 depletion suppresses the growth of colon cancer cells through regulating alternative splicing of MNK2.

Increasing evidence suggests that deregulated RNA splicing factors play critical roles in tumorigenesis; however, their specific involvement in colon cancer remains largely unknown. Here we report that the splicing factor RBM25 is overexpressed in colon cancer, and this increased expression correlates with a poor prognosis of patients with colon cancer. Functionally, RBM25 ablation suppresses the growth of colon cancer cells both in vitro and in vivo. Mechanistically, our transcriptome-wide analysis of splicing events revealed that RBM25 regulates a large number of cancer-related alternative splicing events across the human genome in colon cancer. Particularly, RBM25 regulates the splicing of MNK2 by interacting with the poly G rich region in exon 14a, thereby inhibiting the selection of the proximal 3' splice site (ss), resulting in the production of the oncogenic short isoform, MNK2b. Knockdown of RBM25 leads to an increase in the MNK2a isoform and a decrease in the MNK2b isoform. Importantly, re-expression of MNK2b or blocking the 3' ss of the alternative exon 14a with ASO partially reverses the RBM25 knockdown mediated tumor suppression. Moreover, MNK2b levels were significantly increased in colon cancer tissues, which is positively correlated with the expression level of RBM25. Collectively, our findings uncover the critical role of RBM25 as a key splicing factor in colon cancer, suggesting its potential as a prognostic marker and therapeutic target.

Unique miRNA Expression Profile in MSI- and EMAST-Unstable Sporadic Colon Cancer.

MicroRNAs (miRNAs) are critical post-transcriptional gene regulators and their involvement in sporadic colon cancer (CRC) tumorigenesis has been confirmed. In this study we investigated differences in miRNA expression in microsatellite stable (MSS/EMAST-S), microsatellite unstable marked by high elevated microsatellite alterations at selected tetranucleotide repeats (MSS/EMAST-H), and high microsatellite unstable (MSI-H/EMAST-H) tumor subgroups as well as in tumors with different clinicopathologic characteristics. An RT-qPCR analysis of miRNA expression was carried out on 45 colon cancer and adjacent normal tissue samples (15 of each group). Overall, we found three differentially expressed miRNAs between the subgroups. miR-92a-3p and miR-224-5p were significantly downregulated in MSI-H/EMAST-H tumors in comparison to other subgroups. miR-518c-3p was significantly upregulated in MSS/EMAST-H tumors in comparison to stable and highly unstable tumors. Furthermore, we showed that miR-143-3p and miR-145-5p were downregulated in tumors in comparison to normal tissues in all subgroups. In addition, we showed overexpression of miR-125b-5p in well-differentiated tumors and miR-451a in less advanced tumors. This is the first report on differences in miRNA expression profiles between MSS/EMAST-S, MSS/EMAST-H, and MSI-H/EMAST-H colorectal cancers. Our findings indicate that the miRNA expression signatures differ in CRC subgroups based on their instability status.

Identification and validation of metastasis-related gene ZG16 in the prognosis and progression in colorectal cancer.

Metastasis remains the leading cause of mortality among colorectal cancer (CRC) patients. Identification of new metastasis-related genes are critical to improve colorectal cancer prognosis. Data on mRNA expression in metastatic and primary CRC was obtained from the Gene Expression Omnibus (GEO) database, including GSE81986, GSE41568, GSE71222, GSE21510, and GSE14333. Additionally, data concerning mRNA expression in colon cancer (COAD) and adjacent normal tissues were acquired from The Cancer Genome Atlas (TCGA) database. Hub genes were identified by weighted gene co-expression network analysis (WGCNA) and differential gene expression analysis. Moreover, we assessed the impact of hub gene expression on both overall survival (OS) and disease-free survival (DFS) in patients and identified ZG16 as a potential target. We generated CRC cell lines transfected with lentivirus OE-ZG16 to investigate proliferation, invasion, and migration in vitro. To further elucidate the involvement of ZG16, we utilized gene set enrichment analysis (GSEA) to identify enriched pathways, which were subsequently validated via Western blot analysis. Five datasets containing primary and metastatic CRC samples from GEO database and CRC samples from TCGA database were included in this study and 29 hub genes were identified by WGCNA and differentially expressed gene (DEG) analysis. Low expression of the hub genes (CLCA1 and ZG16) was associated with poor DFS and OS. We confirmed the low expression of ZG16 in CRC using external database and IHC analysis at both transcriptional and protein levels. In addition, the expression of ZG16 was notably elevated in NCM460 cells in comparison to CRC cell lines. The overexpression of ZG16 in CRC cells has been shown to inhibit the proliferation, invasion, and migration of CRC cells. Furthermore, the overexpression of ZG16 has been found to suppress the activation of the epithelial-mesenchymal transition (EMT) and Wnt/β-catenin signaling pathways in CRC. ZG16 may serve as a promising therapeutic target for metastatic CRC treatment.

Differential expression of long non-coding RNAs in colon cancer: Insights from transcriptomic analysis

BackgroundColon Cancer (CC) incidence has sharply grown in recent years. Long non-coding RNAs (lncRNA) are produced by a group of non-protein-coding genes, and have important functions in controlling gene expression and impacting the biological features of various malignancies including CC. MethodsOur research focused on examining the function of lncRNAs in the development of colon cancer. To this end, we selected and analyzed a dataset (GSE104836) from the GEO database, which contained information about the expression of mRNAs and lncRNAs in both colon cancer tissues and normal adjacent paired tumor tissues. The DESeq2 R package in Bioconductor was used to identify differentially expressed lncRNAs (DElncRNAs) and mRNAs (DEmRNAs) that showed differences in expression levels. Next, by literature review of previous studies, we chose two lncRNAs (FENDRR and LINC00092) for additional studies. To validate our findings, a series of tests were performed on a total of 31 tumor tissues and normal paired adjacent tumor tissues. The lncRNA expression levels were assessed in tumor tissues as well as in surrounding normal tumor tissues. ResultsThe data confirmed that just two particular lncRNAs, FENDRR and LINC00092, had considerably decreased expression levels throughout all stages of cancer. In addition, the survival assay was conducted using the GEPIA2 software, revealing that a reduced expression of FENDRR is correlated with a reduced overall survival. Furthermore, our investigation using receiver operating characteristic (ROC) methodology revealed that these two lncRNAs had significant discriminatory ability between colon cancer and normal tissues. To determine the cause of the decrease in the activity of these two long non-coding RNAs (lncRNAs), we used methylation-specific PCR (MSP) to examine the methylation pattern of their promoter regions. Our investigation revealed hypermethylation in the promoter regions of FENDRR and LINC00092 within tumor tissues compared to normal adjacent tumor tissues. ConclusionTaken together, our findings revealed the lncRNAs signatures as potential therapeutic targets and molecular diagnostic biomarkers in colon cancer. Furthermore, the evidence provided substantiates the important role of promoter methylation in regulating the expression levels for both of these lncRNAs.