Colorectal Adenocarcinoma Samples Research Articles

The Potential of PD-1 and PD-L1 as Prognostic and Predictive Biomarkers in Colorectal Adenocarcinoma Based on TILs Grading.

Colorectal cancer (CRC) is a prevalent malignancy with a high mortality rate. Tumor-infiltrating lymphocytes (TILs) play a crucial role in the immune response against tumors. Programmed death-1 (PD-1) and programmed death-ligand 1 (PD-L1) are key immune checkpoints regulating T cells in the tumor microenvironment. This study aimed to assess the relationships among PD-1 expression on TILs, PD-L1 expression in tumors, and TIL grading in colorectal adenocarcinoma. A cross-sectional design was employed to analyze 130 colorectal adenocarcinoma samples. The expression of PD-1 and PD-L1 was assessed through immunohistochemistry. A semi-quantitative scoring system was applied. Statistical analysis with the chi-square test was performed to explore correlations, with the data analyzed in SPSS version 27. PD-1 expression on TILs significantly correlated with a higher TIL grading (p < 0.001), while PD-L1 expression in tumors showed an inverse correlation with TIL grading (p < 0.001). The expression of PD-1 on TILs and PD-L1 on tumor cells correlated significantly with the grading of TILs in colorectal adenocarcinoma. This finding shows potential as a predictive biomarker for PD-1/PD-L1 blockade therapy. Further studies are needed to strengthen these results.

Generation of novel lipid metabolism-based signatures to predict prognosis and immunotherapy response for colorectal adenocarcinoma

Lipid metabolism reprogramming involves in epithelial-mesenchymal transition (EMT), cancer stemness and immune checkpoints (ICs), which influence the metastasis of cancer. This study aimed to generate lipid metabolism-based signatures to predict prognosis, immunotherapy and chemotherapy response for colorectal adenocarcinoma (COAD). Transcriptome data and clinical information of COAD patients were collected from the cancer genome atlas (TCGA) database. The expression of EMT-, stem cell-, and IC-related genes were assessed between COAD and control samples. Modules and genes correlated EMT, ICs and stemness signatures were identified through weighted gene co-expression network analysis (WGCNA). Prognostic signatures were generated and then the distribution of risk genes was evaluated using single-cell RNA sequencing (scRNA-seq) data from GSE132465 dataset. COAD patients exhibited increased EMT score and stemness along with decreased ICs. Next, 12 hub genes (PIK3CG, ALOX5AP, PIK3R5, TNFAIP8L2, DPEP2, PIK3CD, PIK3R6, GGT5, ELOVL4, PTGIS, CYP7B1 and PRKD1) were found within green and yellow modules correlated with EMT, stemness and ICs. Lipid metabolism-based prognostic signatures were generated based on PIK3CG, GGT5 and PTGIS. Patients with high-risk group had poor prognosis, elevated ESTIMATEScore and StromalScore, 100% mutation rate and higher TIDE score. Samples in low-risk group had more immunogenicity on ICIs. Notably, PIK3CG was expressed in B cells, while GGT5 and PTGIS were expressed in stromal cells. This study generates lipid metabolism-based signatures correlated with EMT, stemness and ICs for predicting prognosis of COAD, and provides potential therapeutic targets for immunotherapy in COAD.

Evaluation of PD-L1 Immunoexpression in Resected Samples of Colorectal Adenocarcinoma: A Study on 64 Cases

Evaluation of PD-L1 Immunoexpression in Resected Samples of Colorectal Adenocarcinoma: A Study on 64 Cases

Abstract 3496: EpigenPlot: A tool for the gene-level methylation analysis of colorectal tumors

Abstract Introduction. Aberrant DNA methylation patterns are frequent in colorectal adenocarcinoma (CRC) tissues. The investigation of this layer of epigenetic regulation holds the promise of a better understanding of tumour progression and the identification of novel biomarker candidates. Purpose. Our goal was to collect and analyse publicly available DNA methylation data of colorectal samples and create a user-friendly tool for exploring and visualizing the CRC methylome. Methods. We used Illumina Infinium Methylation Arrays, which is a popular platform for genome-wide methylation analysis of large cohorts. Gene Expression Omnibus database (GEO) and TCGA-COAD (The Cancer Genome Atlas Colon Adenocarcinoma) datasets including normal colorectal mucosa, adenoma, and adenocarcinoma samples were systematically collected and filtered. Differentially methylated regions were identified using the Kruskal-Wallis test. Genes with significant methylation changes were further analysed using ROC (Receiver Operator Characteristic) analysis. Finally, we established a web application using the shiny R package. Results. We assembled a database including methylation data of 2,586 samples. Adenoma and adenocarcinoma samples were globally hypomethylated. Hypermethylation (p &amp;lt; 0.05, Δβ &amp;gt;= 0.2) in adenocarcinoma samples was predominantly present in regions near proximal promoters. Out of the 74 previously proposed biomarkers examined, 50 had a cross-validated AUC of over 0.8. FDA-approved biomarkers BMP3, NDRG4, and SEPT9 have shown the highest performance in the TSS200 region (cvAUC = 0.8, 0.83, and 0.76, respectively). Genes with the highest performance were ITGA4 (5’UTR, first exon: cvAUC = 0.9), MDFI (5’UTR: cvAUC = 0.9), CNRIP1 (5’UTR, first exon: cvAUC = 0.89). The established web application can visualize methylation at CpG sites and gene regions, and KEGG pathway genes. The platform is available at https://epigenplot.com. Conclusion. We collected and analysed a sizeable database of colorectal tissue data. Gene regions with the best performance include previously proposed and novel diagnostic biomarker candidates. The web platform provides an tool for the efficient exploration of the assembled data. Citation Format: Dalma Müller, Balazs Gyorffy. EpigenPlot: A tool for the gene-level methylation analysis of colorectal tumors [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 3496.

Abstract 5074: Profiling the splicing landscape in solid tumors in a large, real-world dataset

Abstract Introduction Aberrant pre-mRNA splicing is common across cancers. Molecular regulation of splicing is an attractive drug target and clinical trials are currently evaluating splice-targeted therapies (STTs). Due in part to the complex and pleiotropic nature of splicing regulation, molecular biomarkers to identify patients who may benefit from STTs are lacking. While mutations in splicing factors (SFs) are promising biomarkers, these are rare, leaving many patients with unmet need. In this study, we identified and characterized splicing patterns (SPs) in a large, multi-cancer cohort. Methods De-identified patient records with splicing data from the Tempus xR whole transcriptome assay were extracted from the Tempus clinicogenomic database (N=2918). A total of 1052 alternative splicing events were detected in at least 90% of the samples with at least 10 supporting reads. Leiden clustering was applied to the percent spliced-in (PSI) values of these events to identify SPs. Fisher’s exact test was used to assess clinical and molecular enrichment in SPs, and Cox proportional-hazards models were used to assess associations with rwOS and derive hazard ratios (HRs). Results From a list of 404 SF genes 7.7% of patients had one or more pathogenic or likely pathogenic (P/LP) SF variants, and 2.2% had variants in the key biomarker SF genes SF3B1, SRSF2, or U2AF1. Clustering revealed 15 SPs containing 43 to 353 samples. One SP had a significant enrichment of SF variants (19.7% of SP, P &amp;lt; 0.0005). SPs were associated with significant differences in rwOS, and many were enriched for samples from a single cancer type, biopsy, or histology - principally by prostate cancer (97.5% of SP), melanoma (94.3% of SP), urothelial carcinoma (93.4% of SP), and renal clear cell carcinoma (93.0% of SP), as well as samples collected from liver biopsies (93.9% of SP). In contrast, colorectal and lung adenocarcinoma samples were present in a wide variety of SPs. One SP was enriched for squamous histologies (66.6% of SP, P &amp;lt; 0.0005) and associated with shorter rwOS (HR = 1.33, 95% CI [1.11, 1.59], P &amp;lt; 0.0005). This association remained after controlling for squamous histology (HR = 1.45, 95% CI [1.02, 2.05], P = 0.04). Samples in this SP were characterized by increased expression of MYC- and proliferation-related genes, and lacked enrichment for SF variants (P = 0.51, 6.6%). Conclusions Mutations in SFs were rare in our cohort, highlighting the need for additional biomarkers related to STTs. This work revealed the presence of multiple SPs - characterized by distinct clinical and molecular traits. Further work will be able to contextualize STT response data using SPs to facilitate STT biomarker discovery. Citation Format: Mario G. Rosasco, Andrew J. Sedgewick, Justin Guinney, Michelle M. Stein. Profiling the splicing landscape in solid tumors in a large, real-world dataset [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 5074.

Automated Image Analysis for Characterization of Circulating Tumor Cells and Clusters Sorted by Magnetic Levitation.

Magnetic levitation-based sorting technologies have revolutionized the detection and isolation of rare cells, including circulating tumor cells (CTCs) and circulating tumor cell clusters (CTCCs). Manual counting and quantification of these cells are prone to time-consuming processes, human error, and inter-observer variability, particularly challenging when heterogeneous cell types in 3D clusters are present. To overcome these challenges, we developed "Fastcount," an in-house MATLAB-based algorithm for precise, automated quantification and phenotypic characterization of CTCs and CTCCs, in both 2D and 3D. Fastcount is 120 times faster than manual counting and produces reliable results with a ±7.3% deviation compared to a trained laboratory technician. By analyzing 400 GB of fluorescence imaging data, we showed that Fastcount outperforms manual counting and commercial software when cells are aggregated in 3D or staining artifacts are present, delivering more accurate results. We further employed Fastcount for automated analysis of 3D image stacks obtained from CTCCs isolated from colorectal adenocarcinoma and renal cell carcinoma blood samples. Interestingly, we observed a highly heterogeneous spatial cellular composition within CTCCs, even among clusters from the same patient. Overall, Fastcount can be employed for various applications with lab-chip devices, such as CTC detection, CTCC analysis in 3D and cell detection in biosensors.

Integrative analysis of KRAS wildtype metastatic pancreatic ductal adenocarcinoma reveals mutation and expression-based similarities to cholangiocarcinoma

Oncogenic KRAS mutations are absent in approximately 10% of patients with metastatic pancreatic ductal adenocarcinoma (mPDAC) and may represent a subgroup of mPDAC with therapeutic options beyond standard-of-care cytotoxic chemotherapy. While distinct gene fusions have been implicated in KRAS wildtype mPDAC, information regarding other types of mutations remain limited, and gene expression patterns associated with KRAS wildtype mPDAC have not been reported. Here, we leverage sequencing data from the PanGen trial to perform comprehensive characterization of the molecular landscape of KRAS wildtype mPDAC and reveal increased frequency of chr1q amplification encompassing transcription factors PROX1 and NR5A2. By leveraging data from colorectal adenocarcinoma and cholangiocarcinoma samples, we highlight similarities between cholangiocarcinoma and KRAS wildtype mPDAC involving both mutation and expression-based signatures and validate these findings using an independent dataset. These data further establish KRAS wildtype mPDAC as a unique molecular entity, with therapeutic opportunities extending beyond gene fusion events.

Abstract 979: Analysis of metabolic and genetic changes in human colorectal cancer and colorectal metastatic tissue by combined NanoString technology and mass spectrometry imaging (MSI)

Abstract Introduction: The development of normal colon tissue to carcinoma can progress through several pathways, which exhibit different clinical, pathological and genetic features. Increasing evidence indicates that tumor development and metastases are not solely driven by genetic alterations but also the tumor microenvironment which is associated with changes in metabolic processes. While many studies show the importance of gene expression profiling in cancer progression, little is known about how this correlates with altered lipid metabolism at metastatic sites within the peritoneal cavity. NanoString nCounter provides specific and sensitive genetic profiling data identifying up to 800 genes simultaneously from small amounts of RNA. For biomolecule analysis, MSI is a powerful tool as it enables the rapid identification of phenotype-dependent MS signatures from tissues with little sample preparation. The aim of this study was to investigate the characteristics of secondary tumors at different metastatic sites compared to the primary tumors using a combination of immunohistochemistry, NanoString analysis and Desorption Electrospray Ionization (DESI) MSI. Methods: 12 fresh-frozen human colorectal adenocarcinoma and metastases samples were cut into 12μm sections. DESI-MSI (Xevo G2-XS, Waters) was performed in both polarities with 50μm spatial resolution. SCiLS Lab (Bruker) software were used for data analysis. Histologically-stained (H&amp;E) adjacent sections were used for microscopy. For genomic analysis, RNA was extracted using AllPrep DNA/RNA micro kit from 20μm sections. Following this, RNA abundance was measured using Qubit fluorimeter. Lastly, mRNA hybridisation, detection and scanning was performed on a NanoString nCounter mRNA Gene Expression system and analysed using nSolver 4.0 software. Results: H&amp;E staining allowed the visualisation of different tissue histopathologies. These differences are echoed in selected DESI-MSI ion images. DESI spectra from distinct regions of the tissue show differences in biomolecule expression especially for phospholipids (m/z 600-900). Ions such as m/z 812 correlated with the non-cancerous region whereas others such as m/z 861 were only observed in the cancerous regions of the tissue. Gene expression analysis showed concordance between primary and metastatic tumors such as in RAS/RAF genes. Others, such as IL-6 expression was seen to upregulated in the metastatic sites compared to the primary tumor. Multivariate analysis enabled identification of biomolecules representative of different tissue phenotypes, with the data showing putative MS biomarkers associated with genetic mutations. Conclusion: Results confirmed the advantage of the combined techniques to reveal spatial correlations between changes in lipid metabolism, tumor heterogeneity and gene expression. Citation Format: Yasmin Shanneik, Emrys Jones, Michal Smiga, Bipasha Chakrabarty, Omer Aziz, Steven Pringle, Kaye Williams, Adam McMahon. Analysis of metabolic and genetic changes in human colorectal cancer and colorectal metastatic tissue by combined NanoString technology and mass spectrometry imaging (MSI) [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 979.

Abstract LB129: Analysis of KRAS wildtype pancreatic ductal adenocarcinoma reveals mutation and expression-based similarities to cholangiocarcinoma

Abstract Introduction Oncogenic KRAS mutations are absent in 10% of patients with pancreatic ductal adenocarcinoma (PDAC) and may represent a subgroup of PDAC with therapy options beyond standard-of-care chemotherapy. While distinct gene fusions have been implicated in KRAS wildtype (wt) PDAC, other traits unique to KRAS wt PDAC remain unexplored. Methods Patients with treatment-naive metastatic PDAC (n=63) received matched tumor/normal whole-genome and RNA sequencing and clinical follow-up as part of the PanGen trial (NCT02869802). Clinical data and mutation, gene fusion and copy alteration frequency was compared between KRAS wt (n=9) and mutant (n=54) groups. Metastatic colorectal adenocarcinoma and cholangiocarcinoma samples from the POG trial (NCT02155621) were incorporated for sequencing-based comparison. Results KRAS wt tumors showed lower frequency of TP53 SNV/indels (p=0.01) and distinct amplifications affecting chr1q genes NR5A2 (p=9.0e-4), MPC2 (p=0.002) and MCL1 (p=0.02) and chr8q gene SQLE (p=0.02). Oncogenic fusions involving NRG1, FGFR2, NTRK2 or BRAF were unique to KRAS wt tumors (67% of samples; p=1e-6). KRAS wt tumors showed expression patterns indicative of cholangiocytes and, when clustered alongside other tumor types, formed a distinct cluster with cholangiocarcinoma samples. Conclusion These data provide the first identification of distinct chr1q/chr8q amplification events unique to KRAS wt PDAC while highlighting novel and striking molecular similarities to cholangiocarcinoma. Taken together, results of the PanGen trial generate new hypotheses relevant to therapeutic targeting of KRAS wt PDAC and rationale towards incorporating KRAS mutation status as part of routine clinical testing in PDAC. Citation Format: James Topham, Erica Tsang, Joanna Karasinska, Andrew Metcalfe, Hassan Ali, Steve Kalloger, Veronika Csizmok, Laura Williamson, Emma Titmuss, Hui-Li Wong, Richard Moore, Andrew Mungall, Jonathan Loree, Oliver Bathe, Patricia Tang, Rachel Goodwin, Janessa Laskin, Marco Marra, Steven Jones, David Schaeffer, Daniel Renouf. Analysis of KRAS wildtype pancreatic ductal adenocarcinoma reveals mutation and expression-based similarities to cholangiocarcinoma [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 LB129.

Clinicopathological Significance of PTEN Expression and Its Prognostic Effect in Colorectal Adenocarcinoma Patients.

Background & Objective:Phosphatase and tensin homolog (PTEN) is a tumor suppressor gene located at chromosome 10. PTEN is a regulator of the PI3K/AKT signaling pathway that inhibits cell proliferation and promotes apoptosis. PTEN loss of function occurs in a spectrum of cancers, including colorectal adenocarcinoma. This study aimed to investigate the probable correlation of negative PTEN expression with clinicopathological features and colorectal adenocarcinoma (CRC) patients' survival. Methods:In this cross-sectional study using Immunohistochemistry stainingPTEN expression status on 151 CRC tissues was evaluated. Then the results of IHC staining was compared to those of clinicopathological features. The relationship between PTEN and KRAS mutation status was also investigated.Results:Of 151 CRC samples, 89 (58.9%) were negative for PTEN expression. Loss of PTEN expression was associated with KRAS mutation (P<0.0001), lymph node metastasis (P=0.002), and advanced tumor stage (P=0.016), whereas no significant association was found with other clinicopathological features. Multivariate analysis indicated that tumor site and KRAS mutation were independent prognostic CRC patients (P<0.05). The Kaplan-Meier analysis indicated a correlation between loss of PTEN expression and overall survival of patients with colorectal adenocarcinoma (P= 0.01). Conclusion:The current study suggests that decreasing PTEN expression or its negative expression may be associated with a higher stage and poor prognosis. Combined analysis of mutated KRAS and PTEN expression could be a good predictor of disease prognosis as well as its clinical outcomes.

Role of Claudin-4 and Matrix Metalloproteinase-2 in Tumor Invasion of Colorectal Adenocarcinoma

Colorectal adenocarcinoma is positioned as the third of most common cancer which the cases rise in Indonesia lately. More than 90% of colorectal carcinoma are adenocarcinoma type. One of prognostic factor of colorectal adenocarcinoma is invasion state of the tumor (T). Uncontrollable proliferation of tumor cell causes transformation of paracellular permeability that increase claudin-4 expression (a protein located on main integral membrane). Claudin-4 activation influence the expression and activity of MMP-2 directly or by altering transduction signal. Expression of claudin-4 and MMP-2 play a role in tumor invasion. Analyzing correlation of claudin-4 and MMP-2 toward invasion state of the tumor (T stadium) on colorectal adenocarcinoma. Analytical observation was conducted on 47 samples of colorectal adenocarcinoma with different invasion state collected by Laboratory of Pathological Anatomy, Dr. Soetomo General Academic Hospital during 2015-2018. Immunohistochemistry was conducted using both claudin-4 and MMP-2 antibodies. Expression of claudin-4 and MMP-2 were semiquantitatively measured then statistically analyzed. Significant result could be obtained in comparison between claudin-4 and tumor invasion state (p=0.773). The significant result could be obtained in comparison between MMP-2 and tumor invasion state (p=0.920). It also could be observed in comparison between claudin-4 and MMP-2 (p=0.638). In summary, claudin-4 and MMP-2 play a role on tumor invasion colorectal adenocarcinoma. It was showed by significant result between claudin-4 and MMP-2 expressions compared to invasion state of colorectal adenocarcinoma.

Analysis of SYK Gene as a Prognostic Biomarker and Suggested Potential Bioactive Phytochemicals as an Alternative Therapeutic Option for Colorectal Cancer: An In-Silico Pharmaco-Informatics Investigation.

Background: SYK gene regulates the expression of SYK kinase (Spleen tyrosine kinase), an important non-receptor protein-tyrosine kinase for immunological receptor-mediated signaling, which is also considered a tumor growth metastasis initiator. An onco-informatics analysis was adopted to evaluate the expression and prognostic value of the SYK gene in colorectal cancer (CRC), the third most fatal cancer type; of late, it may be a biomarker as another targeted site for CRC. In addition, identify the potential phytochemicals that may inhibit the overexpression of the SYK kinase protein and minimize the human CRC. Materials & Methods: The differential expression of the SYK gene was analyzed using several transcriptomic databases, including Oncomine, UALCAN, GENT2, and GEPIA2. The server cBioPortal was used to analyze the mutations and copy number alterations, whereas GENT2, Gene Expression Profiling Interactive Analysis (GEPIA), Onco-Lnc, and PrognoScan were used to examine the survival rate. The protein-protein interaction network of SYK kinase and its co-expressed genes was conducted via Gene-MANIA. Considering the SYK kinase may be the targeted site, the selected phytochemicals were assessed by molecular docking using PyRx 0.8 packages. Molecular interactions were also observed by following the Ligplot+ version 2.2. YASARA molecular dynamics simulator was applied for the post-validation of the selected phytochemicals. Results: Our result reveals an increased level of mRNA expression of the SYK gene in colorectal adenocarcinoma (COAD) samples compared to those in normal tissues. A significant methylation level and various genetic alterations recurrence of the SYK gene were analyzed where the fluctuation of the SYK alteration frequency was detected across different CRC studies. As a result, a lower level of SYK expression was related to higher chances of survival. This was evidenced by multiple bioinformatics platforms and web resources, which demonstrated that the SYK gene can be a potential biomarker for CRC. In this study, aromatic phytochemicals, such as kaempferol and glabridin that target the macromolecule (SYK kinase), showed higher stability than the controls, and we have estimated that these bioactive potential phytochemicals might be a useful option for CRC patients after the clinical trial. Conclusions: Our onco-informatics investigation suggests that the SYK gene can be a potential prognostic biomarker of CRC. On the contrary, SYK kinase would be a major target, and all selected compounds were validated against the protein using in-silico drug design approaches. Here, more in vitro and in vivo analysis is required for targeting SYK protein in CRC.

Carbonic Anhydrase IX and Survivin in Colorectal Adenocarcinoma Cells: Slovakian Population Study.

Simple SummaryThis retrospective study (Slovakian population study) brings information about immunohistochemical detection of CAIX and survivin in 74 samples of human colorectal adenocarcinoma and comparison their expression with expression in healthy colon tissue. Our results show that all of samples with healthy colon tissue were CAIX and survivin-negative and there is no statistically significant dependence of these proteins and the chosen clinicopathological parameters. These findings demonstrate that detection of these proteins could be useful for tumor diagnostic and prognostic and CAIX and survivin could represent independent negative prognostic markers of colorectal cancer.The aim of this study was to detect carbonic anhydrase IX (CAIX) and survivin in the colorectal adenocarcinoma cells of the Slovakian population. We used an indirect three-step immunohistochemical method with DAB staining for the localization of the proteins and investigation their expression. We compared their expression with expression in healthy colorectal tissue. In 74 tissues of colorectal adenocarcinomas, 42% showed CAIX positivity and 20% showed survivin positivity. Brown membrane immunostaining was visible in CAIX-positive tumors. Survivin-positive tumors had strong brown cytoplasmic immunostaining. Co-expression of both proteins was present in five specimens (7%). The samples of normal colorectal tissue (without carcinoma) were CAIX-negative and survivin-negative. We also applied the Chi-squared test for evaluation statistically significant association between the expression of proteins and selected clinical and histopathological parameters. We did not find any statistically significant correlations between CAIX or survivin expression and sex of patients, the grade of the tumor, nodal status and presence of metastasis (p > 0.05). The fact that all samples of normal colorectal tissue were CAIX- and survivin-negative could lead to the possibility of using these two proteins as potential tumor diagnostic markers. On the basic of the available publications and data, we suggest that CAIX and survivin could be negative independent prognostic markers of colorectal cancer, which could affect response to therapy.

Pharmacologically targeting tribbles gene expression in colorectal cancer

Abstract Background TRIB1, TRIB2 and TRIB3 belong to the mammalian Tribbles family of pseudokinases proteins. Several studies reported Tribbles oncogenic role in different types of cancer, including colorectal cancer (CRC). Though current CRC treatment can be curative, patients are in risk of disease recurrence, meaning novel pharmacological targets and strategies are required. Our goal was to analyze Tribbles gene expression in CRC in response to different drugs. Methods Tribbles transcript levels were obtained from GEO profiles database (NCBI). Gene data sets (GDS) were selected based on experimental drug treatment description. Statistical analysis was performed at GraphPadPrism. Results Compared to non-treated control, TRIB2 expression was ∼2-fold increased in colorectal adenocarcinoma samples from patients treated with cyclooxygenase-2 inhibitor celecoxib (GDS3384), though not statistically significant (P &amp;lt; 0.1). TRIB1 was unaltered and data for TRIB3 was not available. By contrast, all Tribbles showed differential expression after treatment of SW620 colon cancer cells with supercritical rosemary extract in progressive increasing doses (0, 30, 60, 100 μg/mL) (P &amp;lt; 0.01;GDS5416). While both TRIB1 and TRIB3 were moderately increased in a dose-dependent manner (∼18% and 13%, respectively), TRIB2 was maximally down-regulated by ∼15% after 60 μg/mL. Conclusions Although celecoxib exhibits antiproliferative effects in different cancer cell types, TRIB2 gene expression showed a trend to be induced after treatment, in contrast to several genes involved in fatty acid oxidation that were down-regulated, which could result from a compensatory mechanism based on a metabolic shift. Since TRIB1/TRIB3 and TRIB2 were oppositely modulated in response to rosemary extract, additional studies are needed to validate its specific pharmacological potential interest for CRC treatment.

ETV4 plays a role on the primary events during the adenoma-adenocarcinoma progression in colorectal cancer

BackgroundColorectal cancer (CRC) is one of the most common cancers worldwide; it is the fourth leading cause of death in the world and the third in Brazil. Mutations in the APC, DCC, KRAS and TP53 genes have been associated with the progression of sporadic CRC, occurring at defined pathological stages of the tumor progression and consequently modulating several genes in the corresponding signaling pathways. Therefore, the identification of gene signatures that occur at each stage during the CRC progression is critical and can present an impact on the diagnosis and prognosis of the patient. In this study, our main goal was to determine these signatures, by evaluating the gene expression of paired colorectal adenoma and adenocarcinoma samples to identify novel genetic markers in association to the adenoma-adenocarcinoma stage transition.MethodsTen paired adenoma and adenocarcinoma colorectal samples were subjected to microarray gene expression analysis. In addition, mutations in APC, KRAS and TP53 genes were investigated by DNA sequencing in paired samples of adenoma, adenocarcinoma, normal tissue, and peripheral blood from ten patients.ResultsGene expression analysis revealed a signature of 689 differentially expressed genes (DEG) (fold-change> 2, p< 0.05), between the adenoma and adenocarcinoma paired samples analyzed. Gene pathway analysis using the 689 DEG identified important cancer pathways such as remodeling of the extracellular matrix and epithelial-mesenchymal transition. Among these DEG, the ETV4 stood out as one of the most expressed in the adenocarcinoma samples, further confirmed in the adenocarcinoma set of samples from the TCGA database. Subsequent in vitro siRNA assays against ETV4 resulted in the decrease of cell proliferation, colony formation and cell migration in the HT29 and SW480 colorectal cell lines. DNA sequencing analysis revealed KRAS and TP53 gene pathogenic mutations, exclusively in the adenocarcinomas samples.ConclusionOur study identified a set of genes with high potential to be used as biomarkers in CRC, with a special emphasis on the ETV4 gene, which demonstrated involvement in proliferation and migration.

Clinical Significance of a Novel Tumor Progression-Associated Immune Signature in Colorectal Adenocarcinoma.

BackgroundSome colorectal adenocarcinoma (CRC) patients are susceptible to recurrence, and they rapidly progress to advanced cancer stages and have a poor prognosis. There is an urgent need for efficient screening criteria to identify patients who tend to relapse in order to treat them earlier and more systematically.MethodsWe identified two groups of patients with significantly different outcomes by unsupervised cluster analysis of GSE39582 based on 101 significantly differentially expressed immune genes. To develop an accurate and specific signature based on immune-related genes to predict the recurrence of CRC, a multivariate Cox risk regression model was constructed with a training cohort composed of 519 CRC samples. The model was then validated using 129, 292, and 446 samples in the real-time quantitative reverse transcription PCR (qRT-PCR), test, and validation cohorts, respectively.ResultsThis classification system can also be used to predict the prognosis in clinical subgroups and patients with different mutation states. Four independent datasets, including qRT-PCR and The Cancer Genome Atlas (TCGA), demonstrated that they can also be used to accurately predict the overall survival of CRC patients. Further analysis suggested that high-risk patients were characterized by worse effects of chemotherapy and immunotherapy, as well as lower immune scores. Ultimately, the signature was identified as an independent prognostic factor.ConclusionThe signature can accurately predict recurrence and overall survival in patients with CRC and may serve as a powerful prognostic tool to further optimize cancer immunotherapy.

Immunohistochemical expression of Snail1 protein in colorectal adenocarcinoma samples and its prognostic activity in Caucasian patients.

IntroductionThe current study investigated the expression of Snail1 protein in colon adenocarcinoma samples to assess its prognostic significance by correlating its expression with the clinicopathological variables and survival of Caucasian patients.AimTo investigate the clinicopathological and prognostic roles of Snail1 expression, the immunohistochemical analysis was performed in colon tumour tissues and adjacent non- pathological mucosa.Material and methodsIt should be noted that only trace expression of this protein was revealed in adjacent non-tumour colorectal mucosa, whereas the high expression was demonstrated in well differentiated, moderately differentiated, and poorly differentiated tumours.ResultsThe Kaplan-Meier survival analysis showed that the overall survival rate in the group of patients with a low expression level of Snail1 was significantly longer than that for patients with a moderate or strong level of Snail1 immunoreactivity (p < 0.001). The 5-year overall survival for patients with a low, moderate, or strong level of Snail1 immunoexpression was 100%, 25.7%, and 2%, respectively. The Snail1-moderate patients had an average survival time of 44.886 months (95% CI: 40.855–48.916), whereas the Snail1-strong expression groups had an average survival time of 21.706 (95% CI: 17.863–25.549). The statistical analysis revealed that the level of the Snail1 immunohistochemical reactivity was correlated with the grade of the histological differentiation. A multivariate analysis demonstrated that the grade of tumour differentiation (HR = 2.150; 95% CI: 1.380–3.349, p = 0.001) and Snail1 expression (HR = 3.901; 95% CI: 2.436–6.247, p < 0.001) were the independent risk factors for worse survival.ConclusionsIn colon adenocarcinoma patients, the expression of Snail1 may act as the independent risk factors for worse survival.

THE EXPRESSION OF HER-2 AND GALECTIN-3 IN COLORECTAL ADENOCARCINOMA AND THEIR RELATIONSHIP WITH CLINICOPATHOLOGICAL PARAMETERS

Colorectal carcinoma (CRC) is the third most common cancer among worldwide. Data from WHO GLOBOCAN in 2018 show there were 1.8 million cases of CRC in the world, with mortality rates 861,000. Histopathology of tumor show 90% of CRC is adenocarcinoma and predominantly in men. Some proteins that are thought to play a role in tumor development are HER2 and Galectin-3. Correlation between expression of Human Epidermal Growth Factor Receptor 2 (HER2) and Galectin-3 with clinicopathology aspect are expected to help determine treatment options. The aim of this study was to investigate the correlation between expression levels of HER-2 and Galectin-3 in vary clinicopathological aspect of colorectal adenocarcinoma. Cross sectional study was conducted in 40 samples of colorectal adenocarcinoma in dr. Kariadi Central General Hospital from 1st January 2018 until 31st December 2018. Samples were analyzed regarding the expression of HER2 and Galectin-3 by immunohistochemical staining of paraffin-embedded material. Coexpression of HER-2, Galectin-3 and the clinicopathological parameters (gender, age, location, depth of invasion, lymph nodes metastase, distant metastase and stage) were analyzed using Chi square test and Spearmans correlation test. Positive HER-2 expression was found on 5% of samples, while strong expresion of Galectin-3 was found on 12.5% of samples. There was no significant correlation between HER-2 and Galectin-3 expression and clinicopathology aspect.

Major driver mutations are shared between sinonasal intestinal-type adenocarcinoma and the morphologically identical colorectal adenocarcinoma.

The purpose of our study was to compare genomic changes in sinonasal intestinal-type adenocarcinoma (sITAC) and colorectal adenocarcinoma (CRC), as they are histomorphologically indistinguishable. This can cause diagnostic difficulties as sinonasal tumours initially diagnosed as sITAC may represent metastasis from CRC, a frequent cancer. Previous studies have not uncovered the underlying mechanism behind the histomorphological resemblance. Tissue samples from all consecutive patients with sITAC at our facility (20 patients) were compared to samples from 20 patients with CRC as well as samples from 2 patients with both CRC and sinonasal tumours. DNA sequencing was performed using Illumina TruSight Oncology 500 panel consisting of 523 cancer-associated genes. Frequent mutations were inspected manually using the Integrative Genomics Viewer. Several well-known cancer-associated genes were mutated in the CRC group, but also in the sinonasal ITAC group. These genes included APC mutated in 65% of the CRC group and 37% of the sinonasal ITAC group, and TP53 mutated in 65% of CRC samples and 58% of ITAC samples. These shared mutations may explain the histomorphological similarities. Successful DNA sequencing was performed on the colorectal sample from one of the two patients with both CRC and sinonasal tumour. Comparing mutations in these samples from one patient we have shown that the sinonasal tumour in all probability was a CRC metastasis. We have identified several genetic similarities between sITAC and CRC. This discovery brings us closer to understanding mechanisms behind the development of sITAC-and hopefully in the future targeted therapy.

Programmed death-ligand 1 expression and tumor-infiltrating lymphocytes in colorectal adenocarcinoma.

Programmed death-ligand 1 (PD-L1) expression and tumor-infiltrating lymphocytes (TILs) are considered have a prognostic value in several malignancies. This study investigated the correlation between PD-L1 expression of tumor cells with the degree of stromal TILs in colorectal adenocarcinoma. A cross sectional study design performed by taking 52 colorectal adenocarcinoma samples. The specimens were stained by immunohistochemical procedure using PD-L1 rabbit monoclonal antibody and the degrees of TILs were assessed base on hematoxylin and eosin (H&E) staining. From a total of 52 samples, the positive PD-L1 expression of tumor cells were 44 (84.6%) samples with 22 (50.0%), 18 (40.9%) and 4 (9.1%) samples had low-, moderate-, and high-degree TILs, respectively. While the negative PD-L1 expression were eight (15.4%) samples with 1 (12.5%), three (37.5%) and four (50.0%) samples had low-, moderate-, and high-degree TILs, respectively. A value of P=0.017 (P<0.05) was obtained by the Chi-square test. This study concluded that there was a significant correlation between PD-L1 expression of tumor cells and the degree of TILs in colorectal adenocarcinoma. This result indicated that the degree of TILs had the potential to be used as a predictive factor for PD-L1 expression of tumor cells in colorectal adenocarcinoma.