Prognostic Value In Colorectal Cancer Research Articles

Identification of Hub genes with prognostic values in colorectal cancer by integrated bioinformatics analysis.

Our study aimed to investigate the Hub genes and their prognostic value in colorectal cancer (CRC) via bioinformatics analysis. The data set of colorectal cancer was downloaded from the GEO database (GSE21510, GSE110224 and GSE74602) for differential expression analysis using the GEO2R tool. Hub genes were screened by protein-protein interaction (PPI) comprehensive analysis. GEPIA was used to verify the expression of Hub genes and evaluate its prognostic value. The protein expression of Hub gene in CRC was analyzed using the Human Protein Atlas database. The cBioPortal was used to analyze the type and frequency of Hub gene mutations, and the effects of mutation on the patients' prognosis. The TIMER database was used to study the correlation between Hub genes and immune infiltration in CRC. Gene set enrichment analysis (GSEA) was used to explore the biological function and signal pathway of the Hub genes and corresponding co-expressed genes. We identified 346 differentially expressed genes (DEGs), including 117 upregulated and 229 downregulated. Four Hub genes (AURKA, CCNB1, EXO1 and CCNA2) were selected by survival analysis and differential expression validation. The protein and mRNA expression levels of AURKA, CCNB1, EXO1 and CCNA2 were higher in CRC tissues than in adjacent tissues. There were varying degrees of immune cell infiltration and gene mutation of Hub genes, especially B cells and CD8+ T cells. The results of GSEA showed that Hub genes and their co-expressed genes mainly participated in chromosome segregation, DNA replication, translational elongation and cell cycle. Overexpression of AURKA, CCNB1, CCNA2 and EXO1 had a better prognosis for CRC and this effect was correlation with gene mutation and infiltration of immune cells.

Exploring the prognostic significance of iron death‑related lncRNAs in colorectal cancer: A systematic review and meta‑analysis.

Colorectal cancer is a significant health challenge globally, with its prognosis associated with the profile of molecular biomarkers. Recently, the role of iron death-associated long non-coding (lnc)RNAs in cancer development has garnered attention; however, their expression patterns and prognostic value in colorectal cancer remain poorly elucidated. The present study aimed to assess the expression levels of iron death-related lncRNAs in colorectal cancer tissues and evaluate their relationship with patient outcomes through a comprehensive meta-analysis. Systematic searches were performed across multiple databases, including PubMed, Embase, Web of Science, Cochrane Library, CNKI, Wanfang and VIP databases, to identify studies relevant to the objective of the present study. Selected studies met predetermined inclusion criteria, from which data were extracted. R software version 4.3.1 was used for the meta-analysis, evaluating the association between iron death-related lncRNAs expression and colorectal cancer prognosis. Publication bias was assessed using funnel plot analysis, and Begg's and Egger's test. A total of 11 studies, including 3,984 patients with colorectal cancer, were included in the present meta-analysis. The results demonstrated a significant association between iron death-related lncRNAs and tumor stage classification [odds ratio (OR)=2.00; 95% confidence interval (CI), 1.77-2.24]. Notably, a significant association was also revealed between iron death-related lncRNAs and T stage classification in colorectal cancer (OR=1.82; 95% CI, 1.50-2.20). Furthermore, a statistically significant association was demonstrated between iron death-related lncRNAs and lymph node metastasis in colorectal cancer (OR=1.31; 95% CI, 1.03-1.66). The findings also highlighted a significant association between iron death-associated lncRNA and distant metastasis in colon cancer (OR=2.04; 95% CI, 1.62-2.56). Moreover, a significant association between iron death-related lncRNAs and risk score in colorectal cancer was revealed (OR=1.75; 95% CI, 1.25-2.46). In conclusion, the findings of the present meta-analysis underscore the potential of high ferroptosis-associated lncRNA expression as an indicator of adverse outcomes in colorectal cancer, suggesting their viability as biomarkers for cancer progression and prognosis. This insight opens potential new avenues for clinical application and further research into colorectal cancer management.

ZNF26-Associated Genes as Prognostic Signatures in Colorectal Cancer with Broad Therapeutic Implications.

The identification of biomarkers correlated with colorectal cancer (CRC) prognosis holds substantial importance from both clinical and scientific perspectives. Zinc finger protein 26 (ZNF26) has not been previously investigated or documented in solid tumors; thus, further research is necessary to ascertain its prognostic value in CRC. Gene expression profiles and clinicopathological data were acquired from The Cancer Genome Atlas (TCGA) database. Subsequently, expression correlation was assessed utilizing the TCGA CRC cohort. The prognostic value of ZNF26 was evaluated through Kaplan-Meier (KM) and ROC curve analyses. Following this, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were conducted to perform enrichment analysis between high- and low-ZNF26 expression groups. The association between immune cells, immune checkpoint genes, and ZNF26 expression levels was examined. Lastly, the research findings were further validated using CRC tissue samples. The results revealed that, in comparison to healthy controls, CRC significantly reduced ZNF26 expression. Elevated ZNF26 expression was associated with poorer overall survival in CRC patients. Additionally, high ZNF26 expression exhibited an inverse relationship with the immunological score and immune checkpoint gene expression in CRC patients. The findings from the TCGA data analysis were corroborated by the PCR results obtained from CRC tissue samples. ZNF26 is markedly upregulated in colorectal cancer tissues, potentially serving as a biomarker for CRC.

Abstract 4635: RNF208: A novel diagnostic and prognostic biomarker for colorectal cancer and key regulator of malignant behavior

Abstract Background: RNF208, a member of the RING finger protein family, has been implicated in ubiquitin-mediated degradation of Vimentin, regulating invasion in breast cancer, and serves as a prognostic factor in uveal melanoma. However, its diagnostic and prognostic value in colorectal cancer (CRC), as well as its role in regulating malignant behavior in CRC, remains unclear. This study aims to reveal the potential of RNF208 as a diagnostic and prognostic marker for CRC and its function in regulating malignant behavior in CRC through bioinformatics analysis and experimental validation. Methods: In this study, based on the ExoRbase2.0 human plasma exosome database (CRC=35; Healthy=118) and TCGA-COAD dataset (CRC=478; Healthy=41), we conducted bioinformatics analysis of the transcriptomic differences in plasma exosomes between CRC patients and normal individuals using ExoRbase2.0. Survival data from the TCGA-COAD dataset were integrated to select key molecules for CRC diagnosis and prognosis. Plasma samples from 39 CRC patients and 16 normal individuals were collected from hospital, and exosomes were isolated and purified using a reagent kit. qPCR was employed to validate the levels of RNF208 in plasma exosomes, confirming the results of bioinformatics analysis. Biological behavior was assessed through CCK-8 proliferation assays, flow cytometry experiments, and Transwell migration assays to investigate the relationship between RNF208 and malignant behavior in the HCT116 colorectal cancer cell line. Results: Bioinformatics analysis revealed a significant correlation between elevated RNF208 expression in plasma exosomes and the incidence of CRC, as well as poor prognosis. The qPCR results from clinical samples further confirmed this correlation, and ROC curves demonstrated high sensitivity and accuracy, suggesting RNF208's potential as a diagnostic and independent prognostic factor for CRC. CCK-8 experiments showed that RNF208 knockdown suppressed proliferation activity in HCT116 cells, flow cytometry indicated that RNF208 knockdown promoted apoptosis in HCT116 cells, and Transwell migration assays demonstrated that RNF208 knockdown inhibited the migration of HCT116 cells. Conclusion: RNF208 serves as a novel diagnostic and prognostic biomarker for colorectal cancer, promoting the proliferation and migration of colorectal cancer cells and contributing to the further deterioration of the disease. Citation Format: Fangyue Guo, Shanfeng Zhang, Pei Li. RNF208: A novel diagnostic and prognostic biomarker for colorectal cancer and key regulator of malignant behavior [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4635.

Relationship Between Tim-3 and Galectin-9 Expression Levels, Clinical Pathological Characteristics, and Prognosis in Patients After Radical Resection of Colorectal Cancer

Some colorectal cancer patients still face high recurrence rates and poor prognoses even after they have undergone the surgical treatment of radical resection. Identifying potential biochemical markers and therapeutic targets for the prognostic evaluation of patients undergoing radical resection of colorectal cancer is crucial for improving their clinical outcomes. Recently, it has been reported that the T cell immunoglobulin and mucin domain protein 3 (Tim-3) and its ligand galactose lectin 9 (galectin-9) play crucial roles in immune dysfunction caused by various tumors, such as colorectal cancer. However, their expressions, biological functions, and prognostic value in colorectal cancer are still unclear. This study aims to investigate the relationship between Tim-3 and galectin-9 expression levels and the clinicopathological characteristics and prognosis of patients undergoing radical resection of colorectal cancer. A total of 171 patients who underwent radical resection of colorectal cancer at Chengdu Fifth People's Hospital between February 2018 and March 2019 were selected. Immunohistochemistry was performed to assess the expression levels of Tim-3 and galectin-9 in the cancer tissue samples and the paracancerous tissue samples of the patients. The relationship between Tim-3 and galectin-9 expression levels and the baseline clinical parameters of the patients was analyzed accordingly. Kaplan-Meier analysis was performed to assess the association between Tim-3 and galectin-9 expression levels and the relapse-free survival (RFS) and the overall survival (OS) of colorectal cancer patients. Cox regression analysis was conducted to identify factors associated with adverse prognosis in the patients. The immunohistochemical results showed that the high expression levels of Tim-3 and galectin-9 were observed in 70.18% (120/171) and 32.16% (55/171), respectively, of the colorectal cancer tissues, whereas the low expression levels were 29.82% (51/171) and 67.84% (116/171), respectively. Furthermore, the expression score of Tim-3 was significantly higher in colorectal cancer tissues than that in the paracancerous tissues, while the expression score of galectin-9 was lower than that in the paracancerous tissues (P<0.05). Further analysis revealed that the expression of Tim-3 and galectin-9 was associated with the depth of tumor infiltration, vascular infiltration, and clinical staging (P<0.05). During the follow-up period of 14-63 months, 7 out of 171 patients were lost to follow-up. Among the remaining patients, 49 and 112 cases presented abnormally low expression of Tim-3 and galectin-9, respectively, whereas 115 and 52 cases presented high expression of Tim-3 and galectin-9, respectively. Kaplan-Meier survival analysis demonstrated that patients with high Tim-3 expression in colorectal cancer tissues had significantly lower RFS and OS than those with low expression did (RFS: log-rank=22.66, P<0.001; OS: log-rank=19.71, P<0.001). Conversely, patients with low galectin-9 expression had significantly lower RFS and OS than those with high expression did (RFS: log-rank=19.45, P<0.001; OS: log-rank=22.24, P<0.001). Cox multivariate analysis indicated that TNM stage Ⅲ (HR=2.26, 95% CI: 1.20-5.68), high expression of Tim-3 (HR=0.80, 95% CI: 0.33-0.91), and low expression of galectin-9 (HR=1.80, 95% CI: 1.33-4.70) were independent risk factors affecting RFS and OS in patients (P<0.05). Aberrant expression of Tim-3 and galectin-9 is observed in colorectal cancer tissues. High expression of Tim-3 and low expression of galectin-9 are closely associated with adverse clinico-pathological characteristics and prognosis. They are identified as independent influencing factors that may trigger adverse prognostic events in patients. These findings suggest that Tim-3 and galectin-9 have potential as new therapeutic targets and clinical indicators.

Using machine learning approach for screening metastatic biomarkers in colorectal cancer and predictive modeling with experimental validation

Colorectal cancer (CRC) liver metastasis accounts for the majority of fatalities associated with CRC. Early detection of metastasis is crucial for improving patient outcomes but can be delayed due to a lack of symptoms. In this research, we aimed to investigate CRC metastasis-related biomarkers by employing a machine learning (ML) approach and experimental validation. The gene expression profile of CRC patients with liver metastasis was obtained using the GSE41568 dataset, and the differentially expressed genes between primary and metastatic samples were screened. Subsequently, we carried out feature selection to identify the most relevant DEGs using LASSO and Penalized-SVM methods. DEGs commonly selected by these methods were selected for further analysis. Finally, the experimental validation was done through qRT-PCR. 11 genes were commonly selected by LASSO and P-SVM algorithms, among which seven had prognostic value in colorectal cancer. It was found that the expression of the MMP3 gene decreases in stage IV of colorectal cancer compared to other stages (P value < 0.01). Also, the expression level of the WNT11 gene was observed to increase significantly in this stage (P value < 0.001). It was also found that the expression of WNT5a, TNFSF11, and MMP3 is significantly lower, and the expression level of WNT11 is significantly higher in liver metastasis samples compared to primary tumors. In summary, this study has identified a set of potential biomarkers for CRC metastasis using ML algorithms. The findings of this research may provide new insights into identifying biomarkers for CRC metastasis and may potentially lay the groundwork for innovative therapeutic strategies for treatment of this disease.

Tissue cadherin 17 (CDH17): A favorable prognostic determinant of colorectal cancer using digital image analysis.

e14651 Background: Colorectal cancer (CRC) is the third most diagnosed malignancy and the second leading cause of cancer-death globally. The identification of robust prognostic markers for risk stratification of CRC would facilitate the accurate selection of high-risk patients for adjuvant therapies, hence improving patients’ survival and quality of living. Cadherin 17 (CDH17) is a validated biomarker for CRC characterized by its overexpression in tumor tissues (Zheng et al., 2021). CDH17 expression in tissue samples can be reliably detected by immunohistochemistry (IHC), which could potentially provide useful information for malignancy monitoring and prognosis. Nevertheless, the implementation of IHC in clinical settings is hindered by the lack of standardized guidelines. In this regard, the present study employed a standardized automated IHC platform and a digitalized sample scoring system to investigate CDH17 tissue expression and examine its clinical prognostic value in CRC. Methods: IHC was performed to assess CDH17 expression on paired tumor and adjacent non-tumor tissues from a cohort of 198 retrospective CRC cases, using the Lic3 antibody and an automated IHC system integrated with AI-digital imaging analysis. The M-score was generated for CDH17 membrane staining according to the formula (1 × % of weakly stained cells + 2 × % moderately stained cells + 3 × % of strongly stained cells)/6. Clinical correlation analysis with TNM, differentiation, and metastasis was determined. The prognostic value of tissue CDH17 was evaluated by Kaplan-Meier curves and Cox regression analyses on patients’ OS and RFS. Comparison of M-score and pathologist visual scoring was made to assess the validity of software-derived IHC scoring. Results: A significantly higher tissue CDH17 level was observed in CRC patients with advanced tumour staging (Early vs. Late stage, p = 0.0493; Mann-Whitney U test) and distant metastasis (p = 0.0188; Mann-Whitney U test). CRC patients with high M-score (M-ScoreHigh; cut-off = 30) were associated with poorer OS and shorter RFS (OS, p = 0.015; RFS, p = 0.018; Log-rank test). Subgroup analysis revealed that M-ScoreHigh in early-stage CRC predicts patients’ disease recurrence of less than 2 years after curative surgery (p = 0.025; Log rank test). Multivariate cox regression analysis showed that CDH17 was an independent prognostic factor for OS (HR = 1.918, 95% CI = 1.001-3.676, p = 0.05). A comparison of software-derived CDH17 M-score with pathologist scoring demonstrated good correlation of 0.7257 (p &lt; 0.0001, n = 36; Pearson correlation). Conclusions: The present study suggests that CDH17 is a potential biomarker of poor prognosis for CRC. Quantification of tissue CDH17 level using a standardized automated IHC platform and digital image analysis could aid in patient stratification which allows efficient treatment strategies for better survival outcomes.

ALDH2 promotes cancer stemness and metastasis in colorectal cancer through activating β-catenin signaling.

Colorectal cancer (CRC) is the primary cause of death from gastrointestinal cancers. Aldehyde dehydrogenase 2 (ALDH2), a crucial mitochondrial enzyme for the oxidative pathway of alcohol metabolism, plays a dual role in cancer progression. In some cancers, it is tumor suppressive; in others, it drives cancer progression. However, whether targeting ALDH2 has any therapeutic implications or prognostic value in CRC is still unclear. Here, we investigated the role of ALDH2 in CRC progression by targeting its enzymatic activity rather than gene expression. We found that inhibiting ALDH2 by CVT-10216 and daidzein significantly decrease migration and stemness properties of both DLD-1 and HCT 116 cells, whereas activating ALDH2 by Alda-1 enhances migration rate. Concomitantly, ALDH2 inhibition by both CVT-10216 and daidzein downregulates the mRNA levels of fibronectin, snail, twist, MMP7, CD44, c-Myc, SOX2, and OCT-4, which are oncogenic in the advanced stage of CRC. Furthermore, Gene Set Enrichment Analysis (GSEA) on ALDH2 co-expressed genes from The Cancer Genome Atlas (TCGA) revealed that MYC target gene sets are upregulated. We found that ALDH2 inhibition decreased the nuclear protein levels of pGSK3β serine 9 and c-Myc. This suggests that ALDH2 probably targets β-catenin signaling in CRC cells. Together, our results demonstrate the prognostic value of ALDH2 in CRC as it regulates both CRC stemness and migration. Our findings also propose that the plant-derived isoflavone daidzein could be a potential chemotherapeutic drug targeting ALDH2 in CRC.

TIGD1 Function as a Potential Cuproptosis Regulator Following a Novel Cuproptosis-Related Gene Risk Signature in Colorectal Cancer

Cuproptosis is a new form of copper-dependent programmed cell death commonly occurring within the body. There is emerging evidence indicating that cuproptosis has a significant regulatory function in the onset and progression of cancer. However, it is still unclear how cuproptosis regulates cancer and whether other genes are involved in the regulation. Using the TCGA-COAD dataset of 512 samples, we found that seven of ten cuproptosis markers showed prognostic value in colorectal cancer (CRC) using Kaplan-Meier survival analysis. Furthermore, 31 prognostic cuproptosis-related genes were identified using weighted gene co-expression network analysis and univariate Cox analysis. Subsequently, we constructed a 7-PCRG signature using least absolute shrinkage and selection operator (LASSO)-Cox regression analysis. The risk score predicting survival in patients with CRC was evaluated. Two risk groups were classified based on their risk scores. The two groups revealed a significant difference in immune cells, such as B and T cells. Furthermore, we identified differences in many immune functions and checkpoints, including CD276 and CD28. In vitro experiments showed that a hub cuproptosis-related gene, TIGD1, could significantly regulate cuproptosis in CRC after exposure to elesclomol. This study validated that cuproptosis was closely related to the progression of CRC. Seven new cuproptosis-related genes were identified, and the function of TIGD1 in cuproptosis was preliminarily understood. Since a certain concentration of copper in CRC cells is important, cuproptosis may provide a new target for cancer therapy. This study may provide novel insights into the treatment of CRC.

Automated assessment of necrosis tumor ratio in colorectal cancer using an artificial intelligence‐based digital pathology analysis

AbstractBackgroundWith the advance in digital pathology and artificial intelligence (AI)‐powered approaches, necrosis is proposed as a marker of poor prognosis in colorectal cancer (CRC). However, most previous studies quantified necrosis merely as a tissue type and patch‐level segmentation. Thus, it was worth exploring and validating the prognostic and predictive value of necrosis proportion with a pixel‐level segmentation in large multicenter cohorts.MethodsA semantic segmentation model was trained with 12 tissue types labeled by pathologists. Segmentation was performed using the U‐net model with a subsequently derived necrosis tumor ratio (NTR). We proposed the NTR score (NTR‐low or NTR‐high) to evaluate the prognostic and predictive value of necrosis for disease‐free survival (DFS) and overall survival (OS) in the development (N = 443) and validation cohorts (N = 333) using 75% as a threshold.ResultsThe 2‐category NTR was an independent prognostic factor and NTR‐low was associated with significant prolonged DFS (unadjusted HR for high vs. low 1.72 [95% CI 1.19–2.49] and 1.98 [1.22–3.23] in the development and validation cohorts). Similar trends were observed for OS. The prognostic value of NTR was maintained in the multivariate analysis for both cohorts. Furthermore, a stratified analysis showed that NTR‐high was a high risk with adjuvant chemotherapy for OS in stage II CRC (p = 0.047).ConclusionAI‐based pixel‐level quantified NTR has a stable prognostic value in CRC associated with unfavorable survival. Additionally, adjuvant chemotherapy provided survival benefits for patients with a high NTR score in stage II CRC.

Statistical Modeling of Relations Between PET/CT Parameters and CEA in Recurrent and Metastatic Colorectal Cancer.

Colorectal cancer (CRC) is a diverse disease with various clinical, pathological and molecular features that affect tumor biological behavior, treatment response and prognosis. The aim of this study was to evaluate the correlation between metabolic 18F-FDG PET/CT parameters (SUVmax, MTV and TLG) and CEA in recurrent and metastatic CRC and to evaluate prognostic value of metabolic 18F-FDG PET/CT parameters in recurrent and metastatic CRC. A descriptive study of 100 patients with previously detected and surgically treated CRC referred to PET/CT with a suspicion of recurrent or metastatic CRC. CEA was measured within three months from the imaging. A low-dose PET/CT was performed per institutional protocol. For each hypermetabolic lesion, metabolic PET/CT parameters (SUVmax, MTV, TLG) were calculated semiautomatically. Pathohistology or clinical data from the follow-up were used as the gold standard. Sensitivity, specificity, PPV and NPV for 18F-FDG PET/CT and CEA in detection of recurrent or metastatic CRC were calculated. Correlation between CEA and SUVmax, MTV and TLG was calculated, separately. To assess the prognostic values of metabolic parameters in CRC, survival analysis with 18-month progression-free survival (PFS) as an endpoint was performed. Microsoft Excel sheets, ROC and Kaplan-Meier curves were used to present the data. Logrank and Tarone-Ware test and Cox model of proportional hazards were used to compare the groups. Study included 100 patients, 45 males and 55 females, age range 36-81 years, mean age 61,4 years. Cancer site was colon in 56% and rectum in 44%. Sensitivity, specificity, PPV and NPV of 18F-FDG PET/CT in detection of recurrent or metastatic CRC was 95%, 73%, 70% and 95%, respectively. Sensitivity, specificity, PPV and NPV of CEA in detection of recurrent or metastatic CRC was 58%, 96%, 91% and 78%, respectively. SUVmax, MTV and TLG positively correlated with CEA, but only CEA-TLG correlation was considered significant (r=0,67). The regression model analysis revealed: SUVmax (HR=0,63, 95%CI=0,28-1,41, p=0,214), MTV (0,59, 95%CI=0,28-1,22, p=0,111) and TLG (HR=0,45 95%CI=0,21-0,99, p=0,028), and the prognostic role in CRC was proven for TLG only. Metabolic 18F-FDG PET/CT parameters may have the prognostic value in CRC, but further multicentric prospective studies are required for validation.

T cell-inflamed gene expression profile is associated with favorable disease-specific survival in non-hypermutated microsatellite-stable colorectal cancer patients.

The anti-tumor immune response plays a key role in colorectal cancer (CRC) progression and survival. The T cell-inflamed gene expression profile (GEP) is a biomarker predicting response to checkpoint inhibitor immunotherapy across immunogenic cancer types, but the prognostic value in CRC is unknown. We evaluated associations with disease-specific survival, somatic mutations, and examined its differentially expressed genes and pathways among 84 sporadic CRC patients from the Seattle Colon Cancer Family Registry. Gene expression profiling was performed using Nanostring's nCounter PanCancer IO 360 panel. Somatic mutations were identified by a targeted DNA sequencing panel. The T cell-inflamed GEP was positively associated with tumor mutation burden and microsatellite instability high (MSI-H). Higher T cell-inflamed GEP had favorable CRC-specific survival (hazard ratio [HR] per standard deviation unit=0.50, p=0.004) regardless of hypermutation or MSI status. Analysis of recurrently mutated genes having at least 10 mutation carriers, suggested that the T cell-inflamed GEP is positively associated with RYR1, and negatively associated with APC. However, these associations were attenuated after adjusting for hypermutation or MSI status. We also found that expression of genes RPL23, EPCAM, AREG and ITGA6, and the Wnt signaling pathway was negatively associated with the T cell-inflamed GEP, which might indicate immune-inhibitory mechanisms. Our results show that the T cell-inflamed GEP is a prognostic biomarker in non-hypermutated microsatellite-stable CRC. This also suggests that patient stratification for immunotherapy within this CRC subgroup should be explored further. Moreover, reported immune-inhibitory gene expression signals may suggest targets for therapeutic combination with immunotherapy.

Tumour break load is a biologically relevant feature of genomic instability with prognostic value in colorectal cancer

BackgroundClinically implemented prognostic biomarkers are lacking for the 80% of colorectal cancers (CRCs) that exhibit chromosomal instability (CIN). CIN is characterised by chromosome segregation errors and double-strand break repair defects that lead to somatic copy number aberrations (SCNAs) and chromosomal rearrangement-associated structural variants (SVs), respectively. We hypothesise that the number of SVs is a distinct feature of genomic instability and defined a new measure to quantify SVs: the tumour break load (TBL). The present study aimed to characterise the biological impact and clinical relevance of TBL in CRC. MethodsDisease-free survival and SCNA data were obtained from The Cancer Genome Atlas and two independent CRC studies. TBL was defined as the sum of SCNA-associated SVs. RNA gene expression data of microsatellite stable (MSS) CRC samples were used to train an RNA-based TBL classifier. Dichotomised DNA-based TBL data were used for survival analysis. ResultsTBL shows large variation in CRC with poor correlation to tumour mutational burden and fraction of genome altered. TBL impact on tumour biology was illustrated by the high accuracy of classifying cancers in TBL-high and TBL-low (area under the receiver operating characteristic curve [AUC]: 0.88; p < 0.01). High TBL was associated with disease recurrence in 85 stages II–III MSS CRCs from The Cancer Genome Atlas (hazard ratio [HR]: 6.1; p = 0.007) and in two independent validation series of 57 untreated stages II–III (HR: 4.1; p = 0.012) and 74 untreated stage II MSS CRCs (HR: 2.4; p = 0.01). ConclusionTBL is a prognostic biomarker in patients with non-metastatic MSS CRC with great potential to be implemented in routine molecular diagnostics.

Pan-Cancer Analysis Reveals SH3TC2 as an Oncogene for Colorectal Cancer and Promotes Tumorigenesis via the MAPK Pathway.

Simple SummarySH3 domain and tetrapeptide repeat 2 (SH3TC2) is a protein-encoding gene and has previously been described as a critical signaling hub for neurological disorders, but no systematic analysis of SH3TC2 is available in cancer research. We analyzed SH3TC2 in various kinds of cancer to find its tumorigenic role in one or more specific cancers and further explored the mechanism of SH3TC2 in colorectal cancer (CRC). Our research revealed that higher expression of SH3TC2 indicated poor disease-free survival and promoted CRC progression and invasion via the MAPK signaling pathway.SH3 domain and tetrapeptide repeat 2 (SH3TC2) is a protein-encoding gene and has previously been described as a critical signaling hub for neurological disorders. Although increasing evidence supports a vital role of SH3TC2 in the tumorigenesis of various kinds of cancer, no systematic analysis of SH3TC2 is available. The function and mechanism of SH3TC2 in other cancers remain unknown. Thus, this study aimed to analyze SH3TC2 in various kinds of cancer to find its tumorigenic role in one or more specific cancers. In the current study, we analyzed the expression level and prognostic value of SH3TC2 in different tumors in the TCGA-GTEx pan-cancer dataset. Subsequently, the prognostic role and mechanism of SH3TC2 in colorectal cancer (CRC) were further explored via clinical samples and in vitro and in vivo experiments. We observed differential expression of SH3TC2 in colon adenocarcinoma (COAD), acute myeloid leukemia (LAML), READ (rectum adenocarcinoma), SKCM (skin cutaneous melanoma), and TGCT (testicular germ cell tumors). Subsequently, SH3TC2 showed a significant effect on the clinical stage and prognostic value in CRC, LAML, and SKCM. Moreover, we found in the TCGA database and seven GEO datasets that SH3TC2 was significantly highly expressed in tumor tissue. Through enrichment analysis of SH3TC2 and its co-expressed genes, we found that SH3TC2 may play a role in the MAPK signaling pathway. Correlation analysis indicated that SH3TC2 was significantly associated with multiple key factors in the MAPK signaling pathway. Additionally, higher expression of SH3TC2 was found in tumor tissue in our cohort including 40 CRC patients. Overexpression of SH3TC2 may imply poor prognosis. Knockdown of SH3TC2 significantly inhibited tumor invasion, migration, and proliferation. More importantly, knockdown of SH3TC2 inhibited tumor growth in a CRC mouse model. The study preliminarily conducted a pan-cancer study of SH3TC2 and further explored the mechanism of SH3TC2 in CRC. Our research revealed that higher expression of SH3TC2 may promote CRC progression and invasion via the MAPK signaling pathway.

Association between mutational subgroups, Warburg-subtypes, and survival in patients with colorectal cancer.

Previous research suggests that Warburg-subtypes are related to potentially important survival differences in colorectal cancer (CRC) patients. In the present study, we investigated whether mutational subgroups based on somatic mutations in RAS, BRAF, PIK3CA, and MET, which are known to promote the Warburg-effect, as well as mismatch repair (MMR) status, hold prognostic value in CRC. In addition, we investigated whether Warburg-subtypes provide additional prognostic information, independent of known prognostic factors like TNM stage. CRC patients (n= 2344) from the prospective Netherlands Cohort Study (NLCS) were classified into eight mutually exclusive mutational subgroups, based on observed mutations in RAS, BRAF, PIK3CA, and MET, and MMR status: All-wild-type + MMRproficient , KRASmut + MMRproficient , KRASmut +PIK3CAmut + MMRproficient , PIK3CAmut + MMRproficient , BRAFmut + MMRproficient , BRAFmut + MMRdeficient , other + MMRproficient , and other + MMRdeficient . Kaplan-Meier curves and Cox regression models were used to investigate associations between mutational subgroups and survival, as well as associations between our previously established Warburg-subtypes and survival within these mutational subgroups. Compared to patients with all-wild-type + MMRproficient CRC, patients with KRASmut + MMRproficient , KRASmut +PIK3CAmut + MMRproficient , BRAFmut + MMRproficient , or other + MMRproficient CRC had a statistically significant worse survival (HRCRC-specific ranged from 1.29 to 1.88). In contrast, patients with other + MMRdeficient CRC had the most favorable survival (HRCRC-specific 0.48). No statistically significant survival differences were observed for the Warburg-subtypes within mutational subgroups. Our results highlight the prognostic potential of mutational subgroups in CRC. Warburg-subtypes did not provide additional prognostic information within these mutational subgroups. Future larger-scale prospective studies are necessary to validate our findings and to examine the potential clinical utility of CRC subtyping based on mutational subgroups.

P-211 Claudin-16, its clinical and prognostic value in colorectal cancer

Claudin-16, also known as CLDN-16 or paracellin-1, plays a major role in tight junction-specific occlusion of the intercellular space, through calcium-independent cell-adhesion activity, and Involved in paracellular magnesium reabsorption. The CLDN16 gene is a protein coding gene, mutations of which are associated with diseases such as Hypomagnesemia 3, renal and primary hypomagnesemia. Among its related pathways are cell junction organization and Blood-Brain Barrier and immune cell transmigration. Tight junctions represent one mode of cell-to-cell adhesion in epithelial or endothelial cell sheets, forming continuous seals around cells and serving as a physical barrier to prevent solutes and water from passing freely through the paracellular space. Evidence has now identified tight junction as key barriers to metastatic disease and cancer progression. We have previously shown that CL16 is associated with survival in breast cancer. The present study explored the expression and relevance of Claudin-16 in human colorectal cancer. Claudin-16 expression levels in human colorectal tumour and normal tissues were quantitatively analysed using gene transcript analyses. Expression was determined against the clinical and pathological parameters and clinical outcome of the patients including recurrence, metastatic disease and deaths related to colorectal cancer. The Mann Whitney U statistical test was utilised for comparisons and logistic regression, with Kaplan-Meier used for survival analyses. Expression of Claudin-16 was significantly reduced in tumour specimens in this colorectal cohort (p=0.0016, tumour n=83; normal n=41). Paired tumour/normal tissues also indicated a significantly reduces expression in tumour samples (p= 0.0287). There was a progressive reduction in Claudin-16 expression with increasing TNM status (p < 0.08). Intriguingly, expression of Claudin-16 was significantly lower in tumours originating in right colon and the rectum (p < 0.034) and expression was reduced in tumours that were both invasive/non-invasive (p < 0.026). When evaluating disease outcome, patients that remained alive and well had higher levels of Claudin-16 when compared to those with local recurrence, metastatic disease and had dies from cancer (p < 0.0238). Kaplan-Meier curves indicated that Clausin-16 was associated with recurrence of disease and death from colorectal cancer (p < 0.042). Claudin-16 has a highly differential expression in colorectal cancer and exhibits properties that in future may enable its use as a possible prognostic indicator.

Tumor-associated CD3- and CD8-positive immune cells in colorectal cancer: The additional prognostic value of CD8+-to-CD3+ ratio remains debatable.

A large number of infiltrating CD3- and CD8-positive inflammatory cells indicates an improved survival in colorectal cancer (CRC), similar to many other cancers. We investigated the prognostic value of different combinations of CD3- and CD8-positive immune cells in CRC patients. The densities of CD3- and CD8-positive cells in intratumoral and stromal tissues were evaluated from 539 patients, for which we calculated a CD3 tumor-stroma index, a CD8 tumor-stroma index, and a CD3-CD8 tumor-stroma index. High CD3 and CD8 tumor-stroma indices associated with stage I to II disease (p < 0.001 for both). The CD3 tumor-stroma index associated with a colonic tumor location (p = 0.006), while the CD8 tumor-stroma index associated with right-sided tumors (p < 0.001) and histological grade 3 tumors (p = 0.032). High intratumoral and stromal densities for CD3- and CD8-positive immune cells, the CD3 tumor-stroma index, the CD8 tumor-stroma index, and the CD3-CD8 tumor-stroma index all indicated a better DSS. The CD3 tumor-stroma index carries a strong prognostic value in CRC, and none of the CD3 and CD8 combinations we analyzed proved superior.

Parvimonas micra is associated with tumour immune profiles in molecular subtypes of colorectal cancer

The importance of the tumour microbiome in different aspects of colorectal cancer (CRC) has been increasingly recognised, but many questions remain. The aim of this study was to explore the effect of specific CRC associated microbes on the tumour immune response, which has a considerable prognostic value in CRC. We applied specific qPCR to detect Parvimonas micra and Fusobacterium nucleatum in tumour tissues from an immunologically well-characterised cohort of 69 CRC patients. This cohort included detailed analyses of immune profiles based on flow cytometry and transcriptomics in tumour tissue and blood, along with comprehensive analyses of molecular subtypes. P. micra and F. nucleatum were detected in 24% and 64% of tumour tissues, respectively. We found a significant association of P. micra with high-grade tumours and tumours of CMS1 subtype. F. nucleatum was significantly associated with right-sided tumours, microsatellite instability, and CMS1 tumours. The immunological analyses revealed significant associations of P. micra with activated CD69+ T lymphocytes and increased antigen-presenting HLA-DR+ B lymphocytes. P. micra was also positively associated with M1 and M2 macrophage traits. The impact of P. micra tumour colonisation on the immune response was further assessed using transcriptomics in validation of our findings. No associations were found between F. nucleatum and immune profiles in this study. Our findings support novel associations between P. micra and the immune response in CRC. A better understanding of these interactions might help to identify important predictive and prognostic tools as well as new targets for therapy.

The Prognostic Value of GNG7 in Colorectal Cancer and Its Relationship With Immune Infiltration.

Background: G Protein Subunit Gamma 7 (GNG7) is an important gene that regulates cell proliferation and induces apoptosis. However, the correlation between GNG7 expression and immune infiltration as well as patient prognosis of colorectal cancer (CRC) remains unclear. Methods: The GNG7 expression differences between tumor tissues and normal tissues were explored via the Oncomine database, Tumor Immune Estimation Resource (TIMER) site and UALCAN database. Then, the influence of GNG7 on clinical prognosis were evaluated, using the PrognoScan database. In addition, the relationship between GNG7 and tumor-related immune infiltration as well as gene marker sets of immune infiltration was investigated via TIMER, TISIDB and GEPIA. Results: We found that GNG7 expression was down-regulated in multiple malignant tumors including colorectal cancer (CRC) and the GNG7 expression was associated with tumor stage, histology subtype, lymph node metastasis and poor prognosis in colorectal cancer (CRC). In addition, the expression of GNG7 was significantly associated with infiltration level of multiple immune cells, immunomodulatory factors as well as part of the immune cell markers. Conclusion: GNG7 displays validated prognostic value in CRC and was associated with its immune cell infiltration and immunoregulation. These results suggest that GNG7 is a potential prognostic marker and is associated with tumor immune infiltration, thus providing a new perspective for the immunotherapy of CRC.

Identifying Potential Biomarkers of Prognostic Value in Colorectal Cancer via Tumor Microenvironment Data Mining.

Colorectal cancer (CRC) is a common cancer that has increased rapidly worldwide in the past decades with a relatively high mortality rate. An increasing body of evidence has highlighted the importance of infiltrating immune and stromal cells in CRC. In this study, based on gene expression data of CRC patients in TCGA database we evaluated immune and stromal scores in tumor microenvironment using ESTIMATE method. Results showed there was potential correlation between these scores and the prognosis, and that patients with higher immune score and lower stromal score had longer survival time. We found that immune score was correlated with clinical characteristics including tumor location, tumor stage, and survival time. Specifically, the right-sided colon cancer had markedly elevated immune score, compared to left-sided colon cancer and rectal cancer. These results might be useful for understanding tumor microenvironment in colorectal cancer. Through the differential analysis we got a list of genes significantly associated with immune and stromal scores. Gene Set Enrichment and protein-protein interaction network analysis were used to further illustrate these differentially expressed genes. Finally, 15 hub genes were identified, and three (CXCL9, CXCL10 and SELL) of them were validated with favorable outcomes in CRC patients. Our result suggested that these tumor microenvironment related genes might be potential biomarkers for the prognosis of CRC.