Spread Of Colorectal Cancer Research Articles

CircZFR promotes colorectal cancer progression via stabilizing BCLAF1 and regulating the miR-3127-5p/RTKN2 axis.

Aberrant expression of circular RNAs (circRNAs) is frequently linked to colorectal cancer (CRC). Here, we identified circZFR as a promising biomarker for CRC diagnosis and prognosis. CircZFR was upregulated in CRC tissues and serum exosomes and its level was linked to cancer incidence, advanced-stages, and metastasis. In both in vitro and in vivo settings, circZFR promoted the growth and spread while suppressing apoptosis of CRC. Exosomes with circZFR overexpression promoted the proliferation and migration of cocultured CRC cells. Mechanistically, epithelial splicing regulatory protein 1 (ESRP1) in CRC cells may enhance the production of circZFR. BCL2-associated transcription factor 1 (BCLAF1) bound to circZFR, which prevented its ubiquitinated degradation. Additionally, circZFR sponged miR-3127-5p to boost rhotekin 2 (RTKN2) expression. Our TCP1-CD-QDs nanocarrier was able to carry and deliver circZFR siRNA (si-circZFR) to the vasculature of CRC tissues and cells, which inhibited the growth of tumors in patient-derived xenograft (PDX) models. Taken together, our results show that circZFR is an oncogenic circRNA, which promotes the development and spread of CRC in a BCLAF1 and miR-3127-5p-dependent manner. CircZFR is a possible serum biopsy marker for the diagnosis and a desirable target for further treatment of CRC.

Transcriptomics and proteomics reveal distinct biology for lymph node metastases and tumour deposits in colorectal cancer.

Both lymph node metastases (LNMs) and tumour deposits (TDs) are included in colorectal cancer (CRC) staging, although knowledge regarding their biological background is lacking. This study aimed to compare the biology of these prognostic features, which is essential for a better understanding of their role in CRC spread. Spatially resolved transcriptomic analysis using digital spatial profiling was performed on TDs and LNMs from 10 CRC patients using 1,388 RNA targets, for the tumour cells and tumour microenvironment. Shotgun proteomics identified 5,578 proteins in 12 different patients. Differences in RNA and protein expression were analysed, and spatial deconvolution was performed. Image-based consensus molecular subtype (imCMS) analysis was performed on all TDs and LNMs included in the study. Transcriptome and proteome profiles identified distinct clusters for TDs and LNMs in both the tumour and tumour microenvironment segment, with upregulation of matrix remodelling, cell adhesion/motility, and epithelial-mesenchymal transition (EMT) in TDs (all p < 0.05). Spatial deconvolution showed a significantly increased abundance of fibroblasts, macrophages, and regulatory T-cells (p < 0.05) in TDs. Consistent with a higher fibroblast and EMT component, imCMS classified 62% of TDs as poor prognosis subtype CMS4 compared to 36% of LNMs (p < 0.05). Compared to LNMs, TDs have a more invasive state involving a distinct tumour microenvironment and upregulation of EMT, which are reflected in a more frequent histological classification of TDs as CMS4. These results emphasise the heterogeneity of locoregional spread and the fact that TDs should merit more attention both in future research and during staging. © 2023 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of The Pathological Society of Great Britain and Ireland.

Novel insights on perils and promises of miRNA in understanding colon cancer metastasis and progression.

Colorectal cancer (CRC) is the third highest frequent malignancy and ultimate critical source of cancer-associated mortality around the world. Regardless of latest advances in molecular and surgical targeted medicines that have increased remedial effects in CRC patients, the 5-year mortality rate for CRC patients remains dismally low. Evidence suggests that microRNAs (miRNAs) execute an essential part in the development and spread of CRC. The miRNAs are a type of short non-coding RNA that exhibited to control the appearance of tumor suppressor genes and oncogenes. miRNA expression profiling is already being utilized in clinical practice as analytical and prognostic biomarkers to evaluate cancer patients' tumor genesis, advancement, and counteraction to drugs. By modulating their target genes, dysregulated miRNAs are linked to malignant characteristics (e.g., improved proliferative and invasive capabilities, cell cycle aberration, evasion of apoptosis, and promotion of angiogenesis). This review presents an updated summary of circulatory miRNAs, tumor-suppressive and oncogenic miRNAs, and the potential reasons for dysregulated miRNAs in CRC. Further we will explore the critical role of miRNAs in CRC drug resistance.

Olfactomedin 4 associates with expression of differentiation markers but not with properties of cancer stemness, EMT nor metastatic spread in colorectal cancer.

Tumor stem cells play a pivotal role in carcinogenesis and metastatic spread in colorectal cancer (CRC). Olfactomedin 4 (OLFM4) is co-expressed with the established stem cell marker leucine-rich repeat-containing G protein-coupled receptor 5 at the bottom of intestinal crypts and has been suggested as a surrogate for cancer stemness and a biomarker in gastrointestinal tumors associated with prognosis. Therefore, it was the aim of the present study to clarify whether OLFM4 is involved in carcinogenesis and metastatic spread in CRC. We used a combined approach of functional assays using forced OLFM4 overexpression in human CRC cell lines, xenograft mice, and an immunohistochemical approach using patient tissues to investigate the impact of OLFM4 on stemness, canonical Wnt signaling, properties of metastasis and differentiation as well as prognosis. OLFM4 expression correlated weakly with tumor grade in one patient cohort (metastasis collection: p= 0.05; pooled analysis of metastasis collection and survival collection: p= 0.19) and paralleled the expression of differentiation markers (FABP2, MUC2, and CK20) (p= 0.002) but did not correlate with stemness-associated markers. Further analyses in CRC cells lines as well as xenograft mice including forced overexpression of OLFM4 revealed that OLFM4 neither altered the expression of markers of stemness nor epithelial-mesenchymal transition, nor did OLFM4 itself drive proliferation, migration, or colony formation, which are all prerequisites of carcinogenesis and tumor progression. In line with this, we found no significant correlation between OLFM4 expression, metastasis, and patient survival. In summary, expression of OLFM4 in human CRC seems to be characteristic of differentiation marker expression in CRC but is not a driver of carcinogenesis nor metastatic spread.

Abstract 987: Proteomics identifies differences in the biological phenotype of tumor deposits and lymph node metastases in colorectal cancer

Abstract Background: Distant metastatic disease remains the main cause of colorectal cancer (CRC) mortality, but our knowledge on how cancer cells travel from the primary tumor to a secondary site is limited. Lymph node metastases (LNM) play a central role in the staging of CRC patients since they are currently viewed as the gateway to distant metastases. However, in recent years additional forms of locoregional spread with varying prognostic impact have been identified, with tumor deposits (TD) having significantly more impact than LNM. The biological explanation for the difference in prognostic impact between LNM and TD is lacking, while this is essential for a more comprehensive understanding of the role of these biomarkers CRC spread. Therefore, in this study, shotgun proteomics were used to compare the biological phenotype of TD and LNM. Methods: From 12 CRC patients, one TD and one LNM were included for paired protein analyses. Proteins were isolated from formalin-fixed paraffin-embedded tissue after which a filter aided sample preparation method was performed. Then, the samples were analyzed by liquid-chromatography tandem mass-spectrometry, and after data handling and filtering, 5578 differentially expressed proteins were used for downstream analyses. Differences in expression were visualized using heatmaps and volcano plots, and enrichment analyses as well as gene set enrichment was performed. All analyses were performed using R version 4.1.2. Results: Among the main findings was that the proteins ASPN, MRC2, SPON1, MXRA5, BGN, LUM, and PLOD2 were overexpressed in TD. These proteins play a role in stimulating pro-tumorigenic processes such as migration, invasion, and epithelial to mesenchymal transition. For the enrichment analyses all proteins with a Fold change of less than -1 and more than 1 were included. Enrichment analyses using the KEGG pathways showed more immune activity in the LNM, with upregulated B-cell and T-cell signaling (p&amp;lt;0.001), while TD showed more interaction with the extracellular matrix in the form of proteoglycans, focal adhesion, and ECM-receptor interaction (p&amp;lt;0.01). In addition, when hallmarks from the Molecular Signatures Database (MSigDB) were used for the enrichment analyses, TD showed the hallmark of epithelial mesenchymal transition (p &amp;lt;0.005). Conclusions: This study shows that TD have a more aggressive and invasive phenotype compared to LNM on protein level. TD had a higher expression of proteins that promote pro-tumorigenic processes and a more extensive interaction with the extracellular matrix. Furthermore, the hallmark of epithelial mesenchymal transition is enriched in TD compared to LNM. These findings form a possible explanation for the strong prognostic impact of TD, and give insight into the heterogeneity of different modes of locoregional spread in CRC. Citation Format: Nelleke P. Brouwer, Loth Webbink, Shannon Van Lent-Van Vliet, Pascal W. Jansen, Femke Simmer, Daniele V. Tauriello, Michiel Vermeulen, Iris D. Nagtegaal. Proteomics identifies differences in the biological phenotype of tumor deposits and lymph node metastases in colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 987.

Myeloid-derived suppressor-like cells - a potential biomarker for prognosis of colorectal cancer?

Background/Aim. Myeloid-derived suppressor cells (MDSC) represent a heterogeneous group of immature cells of myeloid origin, which have the ability to inhibit both the innate and adaptive immune response. Due to its immunosuppressive effect, MDSC can promote the growth and progression of cancer. Colorectal cancer (CRC) is one of the most common cancers in the general population for whose advanced stages of the disease there is still no good therapy. In addition to contributing to the development and spread of CRC, MDSC could potentially be markers of its prognosis. The aim of the study was to examine the potential prognostic role of peripheral blood MDSC counts in CRC patients. Methods. In a prospective study we analyzed the possibility of using CD16low granulocytes and mononuclear (M-MDSC) like cells as well as neutrophyl to lymphocyte ratio (NLR), lymphocite to monocyte ratio (LMR), CD16hi/CD16low granulocytes ratio and M/MMDSC like cells ratios before starting of the treatment as a biomarker for overall survival (OS) in patients with CRC. Hazards ratios (HRs) with the corresponding 95% confidence intervals (95%CIs) were calculated to evaluate the prognostic role of MDSCs in cancer. Results. The analysis was performed in 47 patients in stage III and IV CRC according to the TNM/AJCC disease classification. Dependable data were obtained in 32 patients. Patients blood samples are taken before the possible start of treatment (surgery, chemotherapy). Increased percentage and absolute values of CD16low granulocytes as well as absolute values of M-MDSC-like cells have been shown to be associated with shorter overall survival (OS) (p &lt; 0.0066, p &lt; 0.0013 and p &lt; 0.0119, respectively). The relationship of CD16hi/CD16low granulocytes ratio and M/M-MDSC like cells ratio with OS indicated the existence of positive correlations between these parameters, which higher value of this relationship indicated longer survival of patients (p &lt; 0.0054 and p &lt; 0.0426, respectively). Relationship between OS and NLR indicators found statistically significant inverse correlation with NLR (p = 0.0349). No statistical significance was found in the relationship between OS and LMR. Conclusion. This study shows that percent and absolute numbers of CD16low granulocytes, as well as absolute number of M-MDSC like cells, CD16hi/CD16low granulocytes ratio, M/M-MDSC like cells and NLR ratio ratio may be reliable indicators of survival in patients with CRC.

LIN28 promotes tumorigenesis in colorectal cancer but is not associated with metastatic spread

BackgroundTumor stem cells play a role in metastatic spread in colorectal cancer (CRC). The oncogene LIN28A/B, a prognostic marker in CRC, is involved in tumorigenesis and maintains stem cell function. Therefore, it was the aim of the present study to clarify whether LIN28A/B is involved in metastatic spread in CRC. MethodsExpression of LIN28A/B was analyzed in patients with colon adenocarcinoma in a matched case-control study comparing patients with corresponding liver metastases (n = 42) and patients without hepatic spread within five years (n = 42) by applying immunohistochemistry. Further, LIN28A/B expression was correlated with stem cell associated markers (SOX2, CD133). ResultsLIN28A and B expression significantly correlated with SOX2 expression (p = .02, and p = .04 respectively) but not with CD133 expression. This correlation between LIN28 A/B and SOX2 was not reflected in differences in hepatic spread. In this respect, there was no significant association between LIN28A/B expression and liver metastases. ConclusionLIN28A/B might be involved in tumor initiation and progression in CRC but is not associated with hepatic spread.

Comprehensive characterization of neurotransmitters and neuronal signaling (NT) pathway alterations in colorectal cancer (CRC).

3537 Background: Aberrant NT signaling has been shown to activate uncontrolled proliferation and dissemination in several gastrointestinal cancer types. Neurotransmitters have been shown to affect endothelial cells and immune cells in the tumor microenvironment to promote tumor progression. We previously showed that single nucleotide polymorphisms in the dopamine and GABA pathways are associated with outcome in patients with metastatic CRC receiving first-line treatment. Here we further evaluated the distribution and molecular context of NT pathway alterations in CRC. Methods: A total of 7,595 CRC tumors tested at Caris Life Sciences (Phoenix, AZ) with NextGen Sequencing on DNA (Next Seq, 592 genes or NovaSeq, WES) and RNA (NovaSeq, WTS) were analyzed. ssGSEA (single-sample gene set enrichment analysis) was used to calculate pathway enrichment scores (ES) of 7 NT gene sets (GABA, nicotinic, muscarinic, dopamine (DA), reelin, glial cell line-derived neurotrophic factor and neurotrophins). X2/Fisher-Exact was used for comparison and significance was determined as p-value adjusted for multiple comparison of ( q) &lt; 0.05. Results: ES based on sample sites showed a substantial heterogeneity in NT enrichment. Notably, when compared to primary tumors, all 7 gene sets were significantly enriched in brain metastases (mets; ES ratio 1.14-1.55), while abdomen, liver, and peritoneal mets displayed significant decreases in most NT gene sets. DA was enriched in ovarian and lung mets (ES ratio: 1.18 and 1.09, respectively), the latter also showing increased neurotrophins ES (1.06) (all q &lt; 0.05). When investigating primary tumors grouped according to overall ES by unsupervised clustering, right-sided and CMS4 CRCs were more prevalent in the high ES cluster compared to the low ES cluster (32 vs 29%, P = 0.02 and 46 vs 30%, P &lt; 0.001, respectively). In addition, tumors in the high ES cluster showed lower prevalence of TMB-H (≥ 10mt/MB) (7 vs 10%), MSI-H (6 vs 10%) and PD-L1 (2 vs 6%), while higher CNA rates were noted in 9 genes (all q &lt; 0.05). High ES tumors showed significant positive associations with microenvironment infiltration of B cells, T cells (NK, CD4+ and CD8+ T cells, but not Treg), M2 Macrophages, Myeloid Dendritic Cell, Neutrophils, and an inverse association with M1 Macrophages, regardless of MSI status ( q &lt; 0.05). Conclusions: This is the first and most extensive molecular profiling study to investigate NT signaling pathway alterations in CRC. Our data show a distinct distribution of pathway enrichment according to metastatic site, distinct molecular features in high vs low ES clusters in primary tumors (including CMS subtypes, TMB, MSI and PD-L1 rates), and differential immune cell infiltration. These findings support the role of NT signaling in the metastatic spread of CRC and modulation of tumor immune microenvironment.

EphA3 Downregulation by Hypermethylation Associated with Lymph Node Metastasis and TNM Stage in Colorectal Cancer.

EphA3 is a member of Eph receptors, which is involved in tumorigenesis. The expression and clinical significance of EphA3 in colorectal cancer (CRC) have not been fully investigated. Four colon cancer cell lines and a set of CRC tissues were examined for EphA3 expression. The methylation status of a CpG island within the EphA3 promoter, the presence of four somatic EPHA3 mutations, and EPHA3 gene copy number variations were also analyzed in colon cancer cell lines. EphA3 expression was lost in all colon cancer cell lines examined. EphA3 expression was lower in tumor tissues when compared with normal intestinal tissues (P < 0.001). A comparison of EphA3 immunohistochemical scores for tumor and matched normal intestinal tissues revealed that the protein was downregulated in 82/164 (50.0%), unchanged in 52/164 (31.7%), and upregulated in 30/164 (18.3%) cases of CRC. EphA3 expression was negatively associated with lymph node metastasis (P =0.014, rs=- 0.192) and TNM stage (P =0.001, rs=- 0.260). Downregulation of expression was more common in older patients (P =0.013, rs=0.193). Methylated promoter DNA was detected in all four colon cancer cell lines. Somatic mutations or EphA3 gene deletion was not detected. EphA3 was downregulated in the majority of CRC. Hypermethylation of a CpG island within the EPHA3 promoter provides a possible mechanism. Loss of EphA3 expression was associated with lymph node metastasis and TNM stage and may therefore prove useful as a predictor for tumor spread in CRC.

Snail-1, ALDH1, and claudin-1 tissue proteinexpression in patients with colorectal cancer:clinicopathological and prognostic implications

BackgroundThe occurrence of distant metastases in colorectal cancer (CRC) is the most important parameter in assessment of the prognosis of patients with CRC. Epithelial–mesenchymal transition (EMT) is the process that is incriminated in the occurrence of metastatic spread in CRC. Snail-1 is a zinc-

CD166 as a Stem Cell Marker? A Potential Target for Therapy Colorectal Cancer?

CD166 (Activated leukocyte cell adhesion molecule (ALCAM)) is a member of the immunoglobulin superfamily, which expressed by various cells in several tissues including colorectal cancer (CRC). Many studies have reported the prognostic predictive value of CD166 as a cancer stem cell marker in CRC. CD166 gained increasing attention regarding tumor progression and metastatic spread in CRC. CD166 potentially represents either diagnostic or therapeutic capacities for CRC. This Mini review aimed to clarify the role and significance of CD166 expression in CRC.

Prognostic biomarkers for metastatic colorectal cancer

In colorectal cancer (CRC) distant metastases essentially determine the overall survival and prognosis. Biomarkers that correlate with the presence of distant metastases are therefore of great clinical importance for risk stratification and clinical treatment decisions. As part of the habilitation project various prognostic biomarkers were analyzed and correlated with the occurrence of distant metastases and different patterns of distant spread in CRC. It could be demonstrated that CRC with microsatellite instability (MSI), as detected by a loss of hMLH1 protein expression, have a very low risk of distant metastases. In contrast, microsatellite stable (MSS) CRC (with positive hMLH1 expression) with concurrent activation of the Wnt-beta catenin signaling pathway and strong expression of the cancer stem cell marker CD133, exhibit a very high risk for liver metastases. From these observations a two-step immunohistochemical algorithm based on three immunohistochemical stains could be derived, allowing CRC to be classified according to the risk of distant metastases. Further studies demonstrated that a deregulation of the Wnt-beta catenin signaling pathway and the expression of cancer stem cell markers, such as CD133 and CD44 correlated with hematogenous metastasis to the liver but not with peritoneal carcinomatosis or hematogenous metastasis to the central nervous system (CNS). Finally, in CRC patients with CNS metastases, increased rates of mutations in the mitogen-activated protein kinases (MAPK) pathway (KRAS and BRAF mutations) in combination with a low beta catenin expression could be detected. It can be concluded from these results that for CRC with different patterns of distant spread alternative molecular mechanisms must play a role.

Cancer Stem Cell Markers Are Associated With Distant Hematogenous Liver Metastases But Not With Peritoneal Carcinomatosis in Colorectal Cancer

Although peritoneal carcinomatosis (PC) displays advanced stage in colorectal cancer (CRC), most patients present without distant metastases. To analyze the expression of cancer stem cell markers immunohistochemistry for CD133, CD44 and β-catenin was applied to CRC with exclusive PC, exclusive hepatic metastasis and CRC with combined spread. Expression of cancer stem cell markers correlated with hematogeneous metastases to the liver and was absent in patients with exclusive PC. Thus, expression of cancer stem cell markers correlates with different patterns of metastatic spread in CRC. These data indicate that CRC with exclusive PC lack stem cell features needed for distant dissemination.

The diagnostic accuracy of pericolonic fat extension and attenuation for colorectal tumors

ObjectiveTo evaluate the utility of quantitative analysis of the extension and attenuation of pericolonic fat in the local staging of colorectal cancer (CRC) using multi detector computed tomography (MDCT). Materials and methodsThis was a retrospective study of 110 patients who were operated due to pathologically proven CRC from January 2007 to January 2010, and who underwent preoperative MDCT of the abdomen and pelvis with administration of intravenous contrast material and image acquisition during the portal venous phase. The mean age was 69 years (range of 38–90 years). Pathological reports were reviewed for TNM staging. All MDCT studies were reviewed by two certified radiologists for maximal and minimal tumor diameter, extent of the infiltrated pericolonic fat (measured in mm), attenuation of the infiltrated pericolonic fat (measured in Hounsfield units), and attenuation of normally appearing fat next to the tumor. The sensitivity and specificity of these parameters in detecting patients with ≥ T3 CRC were calculated. ResultsThe overall sensitivity, specificity, and accuracy of pericolonic fat infiltration in detecting patients with ≥T3 stage were 95% (95% CI, 89.0–98.7%), 20% (5.8–43.7%), and 81.9% (74.7–89%) respectively. The mean extent and attenuation of the infiltrated pericolonic fat, in addition to the maximal tumor diameter, were higher in the ≥T3 group (p<0.05). By defining threshold values to these parameters, the positive predictive value for detecting ≥T3 stage tumors approaches 100%. ConclusionQuantitative analysis of pericolonic fat increased the accuracy of MDCT in the detection of local tumor spread in CRC.

Involvement of Chemokine Receptor CCR6 in Colorectal Cancer Metastasis

Various chemokine receptors, namely CXCR4, CCR6 and CCR7, have recently been shown to be involved in the regulation of metastasis in malignant tumors. However, little is known about the role of these receptors in promoting tumor metastasis of colorectal cancer (CRC) to the primary site of CRC metastasis in the liver. To investigate this issue, we analyzed the expression of the chemokine receptors CXCR4, CCR6 and CCR7 in colorectal tumors and colorectal liver metastases. In the present study, 30 human cancer samples from colorectal tissue, 30 human samples from colorectal liver metastases and the adjacent nontumorous liver tissues were screened using quantitative real-time PCR, Western blot analysis, histochemistry, microdissection and the enzyme-linked immunosorbent assay (ELISA). While an overexpression of all the chemokine receptors was found in CRC, in colorectal liver metastases only the chemokine receptors CXCR4 and CCR6 were significantly upregulated. Consequently, we investigated the expression of the corresponding ligands CXCL12/SDF1α, CCL20/MIP3α, CCL19/MIP3β and CCL21/6Ckine in various organs, such as the stomach, esophagus, pancreas, colon and rectum, in comparison with their expression in the liver as the primary site of metastatic spread in CRC. We found that only CCL20 exhibits peak levels of expression in the liver, thus indicating that an increased production of CCL20 may contribute to the selective recruitment of CCR6-expressing cancer cells in CRC. Furthermore, we could demonstrate that CRC patients who developed liver metastases express significantly more CCL20 and CCL21 in the liver in comparison with an unaffected control group. Therefore, our findings strongly suggest an association between CCL20/CCR6 expression in human CRC and the promotion of colorectal liver metastasis.