Adjacent Normal Mucosa Research Articles

Multi-omic profiling reveals associations between the gut mucosal microbiome, the metabolome, and host DNA methylation associated gene expression in patients with colorectal cancer

BackgroundThe human gut microbiome plays a critical role in the carcinogenesis of colorectal cancer (CRC). However, a comprehensive analysis of the interaction between the host and microbiome is still lacking.ResultsWe found correlations between the change in abundance of microbial taxa, butyrate-related colonic metabolites, and methylation-associated host gene expression in colonic tumour mucosa tissues compared with the adjacent normal mucosa tissues. The increase of genus Fusobacterium abundance was correlated with a decrease in the level of 4-hydroxybutyric acid (4-HB) and expression of immune-related peptidase inhibitor 16 (PI16), Fc Receptor Like A (FCRLA) and Lymphocyte Specific Protein 1 (LSP1). The decrease in the abundance of another potentially 4-HB-associated genus, Prevotella 2, was also found to be correlated with the down-regulated expression of metallothionein 1 M (MT1M). Additionally, the increase of glutamic acid-related family Halomonadaceae was correlated with the decreased expression of reelin (RELN). The decreased abundance of genus Paeniclostridium and genus Enterococcus were correlated with increased lactic acid level, and were also linked to the expression change of Phospholipase C Beta 1 (PLCB1) and Immunoglobulin Superfamily Member 9 (IGSF9) respectively. Interestingly, 4-HB, glutamic acid and lactic acid are all butyrate precursors, which may modify gene expression by epigenetic regulation such as DNA methylation.ConclusionsOur study identified associations between previously reported CRC-related microbial taxa, butyrate-related metabolites and DNA methylation-associated gene expression in tumour and normal colonic mucosa tissues from CRC patients, which uncovered a possible mechanism of the role of microbiome in the carcinogenesis of CRC. In addition, these findings offer insight into potential new biomarkers, therapeutic and/or prevention strategies for CRC.

Enhanced CXCR4 Expression Associates with Increased Gene Body 5-Hydroxymethylcytosine Modification but not Decreased Promoter Methylation in Colorectal Cancer

In colorectal cancer (CRC), upregulation of the C-X-C motif chemokine receptor 4 (CXCR4) is correlated with metastasis and poor prognosis, highlighting the need to further elucidate CXCR4’s regulation in CRC. For the first time, DNA methylation and 5-hydroxymethylcytosine aberrations were investigated to better understand the epigenetic regulation of CXCR4 in CRC. CXCR4 expression levels were measured using qPCR and immunoblotting in normal colon tissues, primary colon cancer tissues and CRC cell lines. Publicly available RNA-seq and methylation data from The Cancer Genome Atlas (TCGA) were extracted from tumors from CRC patients. The DNA methylation status spanning CXCR4 gene was evaluated using combined bisulfite restriction analysis (COBRA). The methylation status in the CXCR4 gene body was analyzed using previously performed nano-hmC-seal data from colon cancers and adjacent normal colonic mucosa. CXCR4 expression levels were significantly increased in tumor stromal cells and in tumor colonocytes, compared to matched cell types from adjacent normal-appearing mucosa. CXCR4 promoter methylation was detected in a minority of colorectal tumors in the TCGA. The CpG island of the CXCR4 promoter showed increased methylation in three of four CRC cell lines. CXCR4 protein expression differences were also notable between microsatellite stable (MSS) and microsatellite instable (MSI) tumor cell lines. While differential methylation was not detected in CXCR4, enrichment of 5-hydroxymethylcytosine (5hmC) in CXCR4 gene bodies in CRC was observed compared to adjacent mucosa.

Histone deacetylase inhibitor pre-treatment enhances the efficacy of DNA-interacting chemotherapeutic drugs in gastric cancer.

BACKGROUNDThe prognosis of gastric cancer continues to remain poor, and epigenetic drugs like histone deacetylase inhibitors (HDACi) have been envisaged as potential therapeutic agents. Nevertheless, clinical trials are facing issues with toxicity and efficacy against solid tumors, which may be partly due to the lack of patient stratification for effective treatments.AIMTo study the need of patient stratification before HDACi treatment, and the efficacy of pre-treatment of HDACi as a chemotherapeutic drug sensitizer.METHODSThe expression activity of class 1 HDACs and histone acetylation was examined in human gastric cancer cells and tissues. The potential combinatorial regime of HDACi and chemotherapy drugs was defined on the basis of observed drug binding assays, chromatin remodeling and cell death.RESULTSIn the present study, the data suggest that the differential increase in HDAC activity and the expression of class 1 HDACs are associated with hypo-acetylation of histone proteins in tumors compared to normal adjacent mucosa tissue samples of gastric cancer. The data highlights for the first time that pre-treatment of HDACi results in an increased amount of DNA-bound drugs associated with enhanced histone acetylation, chromatin relaxation and cell cycle arrest. Fraction-affected plots and combination index-based analysis show that pre-HDACi chemo drug combinatorial regimes, including valproic acid with cisplatin or oxaliplatin and trichostatin A with epirubicin, exhibit synergism with maximum cytotoxic potential due to higher cell death at low combined doses in gastric cancer cell lines.CONCLUSIONExpression or activity of class 1 HDACs among gastric cancer patients present an effective approach for patient stratification. Furthermore, HDACi therapy in pre-treatment regimes is more effective with chemotherapy drugs, and may aid in predicting individual patient prognosis.

Exosomal markers (CD63 and CD9) expression and their prognostic significance using immunohistochemistry in right-sided and left-sided colon cancer.

182 Background: Exosomes play pivotal roles in cancer progression, metastasis and chemoresistance. CD63 and CD9 are widely accepted exosomal markers. Their pattern of expression and prognostic significance in patients with RSCC and LSCC is unknown. This study explored CD63 and CD9 expression and prognostic significance in patients with RSCC and LSCC using immunohistochemistry (IHC). Methods: Between 2015 and 2018, 63 patients underwent surgical resection of colon cancer for whom we had available tissues for CD63 and CD9 IHC staining. Two pathologists independently scored the CD63 and CD9 expression in the tumor and adjacent normal mucosa (ANM). Staining intensity was graded 1-3 and staining percentage was estimated in 10% increments. Mean Quick-score (Q-score) (intensity X percentage of staining) was calculated. Results: RSCC and LSCC represented 52% and 48% of the patients respectively. The ANM and Tumor CD63 Q-scores were 225 vs 191 (p = 0.009) in RSCC and 224 vs 154 (p = 0.0001) in LSCC, respectively. The ANM and Tumor CD9 Q-scores were 134 vs 152 (p = 0.142) in RSCC and 135 vs 154 (p = 0.137) in LSCC, respectively. In patients with RSCC and LSCC, the mean Tumor CD63 Q-score was 191 vs 154 (p = 0.024), while the mean ANM CD63 Q-score was 225 vs 224 (p = 0.920). The mean Tumor CD9 Q-score was 152 and 154 (p = 0.883), and the mean ANM CD9 Q-score was 134 vs 135 (p = 0.926). In our cohort, there was no difference in progression free survival (PFS) between patients with RSCC and LSCC (p = 0.2349). In all patients, there was no difference in PFS in patients with CD63 expression < 100 and ≥100 (p = 0.8284). Among patients with RSCC, there was a significantly lower PFS in patients with CD63 expression < 100 vs. ≥100 (p = 0.0259). However, among patients with LSCC, there was no difference in PFS in patients with CD63 expression < 100 vs. ≥100 (p = 0.3494). Conclusions: To our knowledge, this is the first study to show a difference in exosomal marker (CD63) expression pattern and its prognostic significance in patients with RSCC and LSCC. There was a significant positive correlation between progression free survival in patients with RSCC and higher exosomal expression.

Expression Patterns of Xenobiotic-Metabolizing Enzymes in Tumor and Adjacent Normal Mucosa Tissues among Patients with Colorectal Cancer: The ColoCare Study.

Xenobiotic-metabolizing enzymes (XME) play a critical role in the activation and detoxification of several carcinogens. However, the role of XMEs in colorectal carcinogenesis is unclear. We investigated the expression of XMEs in human colorectal tissues among patients with stage I-IV colorectal cancer (n = 71) from the ColoCare Study. Transcriptomic profiling using paired colorectal tumor and adjacent normal mucosa tissues of XMEs (GSTM1, GSTA1, UGT1A8, UGT1A10, CYP3A4, CYP2C9, GSTP1, and CYP2W1) by RNA microarray was compared using Wilcoxon rank-sum tests. We assessed associations between clinicopathologic, dietary, and lifestyle factors and XME expression with linear regression models. GSTM1, GSTA1, UGT1A8, UGT1A10, and CYP3A4 were all statistically significantly downregulated in colorectal tumor relative to normal mucosa tissues (all P ≤ 0.03). Women had significantly higher expression of GSTM1 in normal tissues compared with men (β = 0.37, P = 0.02). By tumor site, CYP2C9 expression was lower in normal mucosa among patients with rectal cancer versus colon cancer cases (β = -0.21, P = 0.0005). Smokers demonstrated higher CYP2C9 expression levels in normal mucosa (β = 0.17, P = 0.02) when compared with nonsmokers. Individuals who used NSAIDs had higher GSTP1 tumor expression compared with non-NSAID users (β = 0.17, P = 0.03). Higher consumption of cooked vegetables (>1×/week) was associated with higher CYP3A4 expression in colorectal tumor tissues (β = 0.14, P = 0.007). XMEs have lower expression in colorectal tumor relative to normal mucosa tissues and may modify colorectal carcinogenesis via associations with clinicopathologic, lifestyle, and dietary factors. Better understanding into the role of drug-metabolizing enzymes in colorectal cancer may reveal biological differences that contribute to cancer development, as well as treatment response, leading to clinical implications in colorectal cancer prevention and management.

Claudin-3 expression in gastric cancer

To evaluate the expression of claudin-3 in gasric cancer and in adjacent normal mucosa and its association with clinical and pathological parameters. Tissue specimens from a total of 69 patients with gastric cancer were obtained. Immunohistochemical reactions were performed using mouse polyclonal antibodies to claudin-3. The expression of claudin-3 in gastric cancer was significantly higher than in adjacent normal mucosa (p<0,05). The absence of claudin-3 was significantly associated with poor differentiation (p<0,05). An abnormal nuclear expression of claudin-3 was observed in 69.6% cases. A significant association was found between nuclear expression and the absence of membranous claudin-3 expression (p<0,05).

High potential of SOX21 gene promoter methylation as an epigenetic biomarker for early detection of colorectal cancer.

Despite the advances in screening during the past decades, colorectal cancer (CRC) still is a leading cause of cancer deaths worldwide. Therefore, the development of new diagnostic methods is necessary. The aim of this study was to compare methylation changes of SRY-Box 21 (SOX21) gene promoter in tumor tissues and their normal adjacent mucosa in patients with CRC and to examine the relationship between the methylation levels and demographic/clinicopathological factors. A total of 41 CRC patients participated in the present study. After the extraction of DNA and bisulfite treatment of the samples, the methylation levels were determined by using the MethyLight method. Two-sided Mann-Whitney U test was used to compare the median level of methylation in tumor tissues and their adjacent normal mucosa. The methylation rates in tumor tissue samples were significantly higher compared to their adjacent normal mucosa (P < 0.0001). No association between demographic/clinicopathological factors and methylation status observed in tumor tissues. A receiver operating characteristics curve was constructed and tissue samples exhibited a sensitivity of 80.5% and specificity of 97.6% for SOX21 promoter methylation. The results of this study indicated the high potential of SOX21 gene promoter methylation as a candidate noninvasive diagnostic biomarker in stool and plasma of colorectal cancer patients. However, further studies with larger sample sizes are required to evaluate the specific role of SOX21 methylation as a biomarker for early detection of CRC.

Nuclear factor κB -inducing kinase is a diagnostic marker of gastric cancer.

Nuclear factor-κB-inducing kinase (NIK) is a new regulator of nuclear factor-κB signaling, which plays an important role in tumorigenesis. This study aimed to examine the expression of NIK in gastric cancer and investigate its clinical significance.Tumor issues were collected from 80 gastric cancer patients who received surgery and the diagnosis was confirmed by postoperative pathological analysis. The expression of NIK in gastric cancer tissues and adjacent normal mucosa was detected by immunohistochemical analysis. The associations between NIK expression and clinicopathological features of the patients were further analyzed.NIK expression was significantly higher in gastric cancer tissues than in adjacent normal tissues (P < .05). Furthermore, NIK expression showed significant association with UICC stage, T status, and differentiation, but not with age and gender of gastric cancer patients.NIK is overexpressed in gastric cancer and is a potential diagnostic marker of gastric cancer.

A comparative study of the clinical andhistopathological features of normal labial mucosa and redundant tissue in double lip

Introduction: double lip is one of lip abnormalities that may be congenital or acquired. It consists of a fold of excess tissue on the mucosal side of the lip. It causes functional and aesthetic problems to affected patientsAim of the work : The present study was performed to compare clinically and histopathologically between redundant tissue in double lip and normal labial mucosa.Subjects and methods: Seventeen patients with congenital non syndromic double lip were selected. Redundant tissues were removed surgically together with part of adjacent normal mucosa as part of the surgical manipulation. Hematoxylin and eosin staining was done to compare both tissues microscopically. Expression of ki-67 and p53 was evaluated in both tissue by immunohistochemistryResults: Clinically, redundant tissue appeared as thick extra fold with normal mucosal color and soft consistency analogous to normal labial mucosa. This redundant tissue was removed surgically without any complications. Histopathologically, normal labial mucosa revealed non keratinized stratified squamous epithelium and delicate underlying connective tissue. Excess tissue exhibited parakeratosis, acanthosis, elongation of rete pegs. The underlying connective tissue was fibrotic with inflammatory cell infiltration. In normal labial mucosa, ki-67 was detected only in parabasal layer of epithelium whereas it was localized in parabasal and superficial layers of epithelium in redundant tissue. No expression of p53 was detected in both normal and excess tissue. For both ki-67 and p53, the difference in expression between normal and redundant tissue was significant.Conclusion : Redundant tissue is a benign hyperplastic tissue that require surgical removal. So surgical excision here is the treatment of choice when excess tissue causes functional and aesthetic problems. Moreover, the high proliferation of redundant tissue may necessitate the surgical excision of this tissue.

Impact of the ESM-1 Gene Expression on Outcomes in Stage II/III Gastric Cancer Patients Who Received Adjuvant S-1 Chemotherapy.

Endothelial cell-specific molecule-1 (ESM-1) is a soluble proteoglycan which has important role in various biological events. We investigated the impact of the ESM-1 expression in cancer tissues on outcomes in stage II/III gastric cancer patients who received adjuvant S-1 chemotherapy. The ESM-1 mRNA expression in cancerous tissues and adjacent normal mucosa from 253 patients was measured. The associations between the ESM-1 gene expression and the survival and clinicopathological features were investigated. A significant association was observed between high ESM-1 expression and undifferentiated adenocarcinoma. The overall survival curve was significantly lower in patients with high ESM-1 expression than in those with low expression (p=0.005). High ESM-1 expression was a significant independent prognosticator (HR=2.291, p=0.007). ESM-1 gene expression in cancerous tissues is an important prognosticator in stage II/III gastric cancer patients who received adjuvant S-1 chemotherapy.

Elevated expression of AGGF1 predicts poor prognosis and promotes the metastasis of colorectal cancer

BackgroundAngiogenic factor with G-patch and FHA domains 1 (AGGF1) can promote angiogenesis and increasing evidence has highlighted the important roles of AGGF1 in tumorigenesis. However, the differential expression as well as the biological functions of AGGF1 in colorectal cancer (CRC) remain to be established. The purpose of the present study is therefore to identify the effect of AGGF1 on prognosis and metastasis in CRC patients.MethodsThe expression level of AGGF1 in CRC was examined by qPCR, western blot and immunohistochemistry in a tissue microarray containing 236 CRC specimens and paired normal mucosae. And the effect of AGGF1 on CRC cell malignance was investigated in our established stable AGGF1 upregulated and knockdown CRC cell lines.ResultsThe expression level of AGGF1 in CRC tissue was not significantly different to that in adjacent normal mucosa at the mRNA level. However, at the protein level, AGGF1 expression in CRC tissues was significantly higher than in paired normal mucosa, which showed a clear association with TNM stage, AJCC stage, vascular invasion, and differentiation. Further, we revealed an apparent correlation between AGGF1 expression and poorer disease-free survival and overall survival of CRC patients. In addition, we discovered that AGGF1 significantly promoted CRC cell wound healing, migration, and invasion in vitro and distant metastasis in vivo.ConclusionsOur study demonstrates the aberrant overexpression of AGGF1 in CRC and provides a basis on which to explore the application of AGGF1 as a potential therapeutic target for CRC patients, especially for CRC patients with distant metastasis.

Clinical Significance of KIAA1199 as a Novel Target for Gastric Cancer Drug Therapy.

The KIAA1199 gene has been associated with cancer-cell proliferation, but its functions remain poorly studied. Here, we examined the clinical significance of the KIAA1199 mRNA levels in locally advanced gastric cancer (GC). Materials and Methods/Results: Using samples from 254 patients with stage II/III GC, we found significantly higher KIAA1199 levels in cancerous tissues compared to adjacent normal mucosa (ANM). There was no significant relationship between KIAA1199 expression and clinical features. Although overall survival rates (OSR) of patients, who underwent surgery did not correlate with KIAA1199 expression, patients who underwent adjuvant chemotherapy with S-1 and had high KIAA1199 levels displayed significantly lower OSR. KIAA1199 knock down (KIAA1199-KD) suppressed proliferation, invasiveness, and sensitivity of GC cells to 5-fluorouracil (5-FU). KIAA1199 expression appears to be a promising prognostic marker in patients with locally advanced GC, who underwent postoperative adjuvant chemotherapy with S-1. KIAA1199 may represent a novel target for GC pharmacotherapy.

Identification and Verification of the Main Differentially Expressed Proteins in Gastric Cancer via iTRAQ Combined with Liquid Chromatography-Mass Spectrometry.

Background To find the potential intersections between the differentially expressed proteins and abnormally expressed genes in gastric cancer (GC) patients. Methods Gastric cancer tissue and adjacent normal mucosa tissue were used for iTRAQ analysis. Gene ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, and protein-protein interaction (PPI) analysis were used to evaluate gene function. Western blotting and immunohistochemistry (IHC) were applied to verify the protein expression. Results A total of 2770 proteins were identified, of which 147 proteins were upregulated and 159 proteins were downregulated. GO analysis revealed that the differentially expressed genes were mainly enriched for the terms “cellular process,” “binding,” and “cell.” The results of the KEGG analysis showed that the most abundantly enriched proteins were involved in the “focal adhesion” pathway. The results of the PPI analysis showed that VCAM1 was located at the center of the PPI network. Western blotting and IHC analysis demonstrated that VCAM1, FLNA, VASP, CAV1, PICK1, and COL4A2 were differentially expressed in GC and adjacent normal tissues, which was consistent with the results of the iTRAQ analysis. Conclusion In conclusion, 6 highly differentially expressed proteins were identified as novel differentially expressed proteins in human GC. This exploratory research may provide useful information for the treatment of gastric cancer in the clinic.

Shotgun lipidomics-based characterization of the landscape of lipid metabolism in colorectal cancer

Solid tumors are characterized by global metabolic alterations which contribute to their growth and progression. Altered gene expression profiles and plasma lipid composition suggested a role for metabolic reprogramming in colorectal cancer (CRC) development. However, a conclusive picture of CRC-associated lipidome alterations in the tumor tissue has not emerged. Here, we determined molar abundances of 342 species from 20 lipid classes in matched biopsies of CRC and adjacent normal mucosa. We demonstrate that in contrast to previous reports, CRC shows a largely preserved lipidome composition that resembles that of normal colonic mucosa. Important exceptions include increased levels of lyso-phosphatidylinositols in CRC and reduced abundance of ether phospholipids in advanced stages of CRC. As such, our observations challenge the concept of widespread alterations in lipid metabolism in CRC and rather suggest changes in the cellular lipid profile that are limited to selected lipids involved in signaling and the scavenging of reactive oxygen species.

YAP Levels Combined with Plasma CEA Levels Are Prognostic Biomarkers for Early-Clinical-Stage Patients of Colorectal Cancer.

Background Colorectal cancer (CRC) is one of the most common cancers worldwide. Surgical operation is routinely applied to patients with CRC. An important part of postoperative care for patients is to assess the prognosis of patients, especially those with early-stage cancers. However, effective biomarkers for CRC prognosis remain inadequate. The purpose of this study was to assess the prognostic potential of Yes-associated protein (YAP) and carcinoembryonic antigen (CEA) in early-stage CRC. Methods A total of 116 matched pairs of CRC tissues and adjacent normal mucosae as well as 73 cases of metastatic lymph nodes were analyzed. Results The results show that CRC tissues exhibited higher YAP expression compared with the adjacent normal mucosae. Immunohistochemical analysis shows that YAP expression in the CRC or lymphatic metastatic tissues was clearly higher than that in normal mucosae (P < 0.01), whereas that in CRC tissues with lymphatic metastasis was higher than that in tissues without lymphatic metastasis (P < 0.05). YAP expression is associated with serosal invasion, lymphatic metastasis, lymph node ratio, remote metastasis, Dukes stage, and CEA levels (P < 0.05). YAP and CEA are independent predictors of the survival of CRC patients (P < 0.05 and P < 0.01). YAP predicted CRC prognosis primarily for patients with late-clinical-stage CRC (P=0.002), but not for patients with early-clinical-stage CRC (P=0.083). However, patients with high YAP and high CEA levels exhibited lower overall survival rates than those with low YAP expression in early-clinical-stage CRC (P < 0.001). Conclusion High YAP levels in the cancer tissues combined with high plasma CEA levels are potential biomarkers for predicting CRC prognosis in the early clinical stage.

High gamma-glutamyl hydrolase and low folylpolyglutamate synthetase expression as prognostic biomarkers in patients with locally advanced gastric cancer who were administrated postoperative adjuvant chemotherapy with S-1

PurposeThe enzymes gamma-glutamyl hydrolase (GGH) and folylpolyglutamate synthetase (FPGS) regulate intracellular folate concentrations needed for cell proliferation, DNA synthesis, and repair. High GGH expression affects 5-FU thymidylate synthase (TS) inhibition and is a risk factor for various malignancies. Here, the clinical significance of GGH and FPGS expression was investigated in Stage II/III gastric cancer patients undergoing postoperative adjuvant chemotherapy with S-1.MethodsSurgical specimens of cancer tissue and adjacent normal mucosa, obtained from 253 patients with previously untreated gastric cancer, were examined. GGH and FPGS mRNA expression was measured by qPCR to evaluate their clinicopathological significance in gastric cancer patients after curative resection.ResultsWhile FPGS expression showed no significant differences between the cancerous and normal samples, GGH expression was higher in cancer tissue than in adjacent normal mucosa. High GGH expression was correlated with age, histological type, and vascular invasion. Overall survival (OS) of patients with high GGH mRNA expression was significantly poorer than of patients with low GGH expression. Multivariate analysis showed that high GGH expression was an independent prognostic factor of OS (HR: 2.58, 95% CI 1.29–5.16). Patients who received S-1 adjuvant treatment showed a significantly poor OS between high GGH/low FPGS and low GGH/high FPGS. Patients without adjuvant treatment showed no significant difference.ConclusionGGH expression was significantly higher in gastric cancer tissue than in adjacent normal mucosa. High GGH and low FPGS expression is a useful independent predictor of poor outcomes in stage II/III gastric cancer patients undergoing postoperative adjuvant chemotherapy with S-1.

Identification of genes and miRNAs associated with angiogenesis, metastasis, and apoptosis in colorectal cancer

Colorectal cancer (CRC) is the main cause of cancer-related mortalities, worldwide and is the most prevalent cancer of the gastrointestinal tract. Different techniques exist to diagnose CRC including colonoscopy, sigmoidoscopy, and plasma miRNA test. This study evaluates the important miRNAs and main genes playing role in metastasis, angiogenesis and apoptosis that can be considered as prognostic and progression markers of CRC. We have measured the expression levels of the miRNAs, and multiple genes related to angiogenesis, metastasis and apoptosis in 45 paired samples of CRC tissue specimens and adjacent normal mucosa tissues obtained from 45 patients with CRC utilizing Real-time PCR. We have also analyzed the relationship between the expression of these genes and clinic pathological features. Results showed that the expression levels of angiogenesis genes (PGE-2, VEGF, Vimentin, CD105, CXCL-1, and IGFBP-7), metastatic genes (IGF-1R, Snail-1, and MMP-7) and apoptotic genes (Bcl-2, Bcl-xl, and NF-κβ1) were significantly higher in cancer tissues. But the expression levels of E-cadherin, AdipoR1, Claudin-7, Bax, Puma, and Noxa, were notably lower in cancer tissues. Additionally, it was revealed that miRNAs (miR-21, miR-19a, miR-18a, and miR-92a) expression levels were in higher levels in cancer tissues when compared to the normal tissues. In contrast, the expression levels of miR-31, and miR-139-5p were significantly lower in cancer tissues. It is hoped that, by measuring the metastasis, angiogenesis and apoptosis resistance associated miRNAs expression, better treatment results would be achieved in CRC treatment.

Evaluation of MGP gene expression in colorectal cancer

PurposeMatrix Gla protein (MGP) is a vitamin K-dependent, γ-carboxylated protein that was initially found to be a physiological inhibitor of ectopic calcifications affecting mainly cartilage and the vascular system. Mutations in the MGP gene were found to be responsible for a human pathology, the Keutel syndrome, characterized by abnormal calcifications in cartilage, lungs, brain and vascular system. MGP was recently implicated in tumorigenic processes such as angiogenesis and shown to be abnormally regulated in several tumors, including cervical, ovarian, urogenital and breast. This fact has triggered our interest in analyzing the expression of MGP and of its regulator, the transcription factor runt related transcription factor 2 (RUNX2), in colorectal cancer (CRC). MethodsMGP and RUNX2 expression were analyzed in cancer and non-tumor biopsies samples from 33 CRC patients and 9 healthy controls by RT-qPCR. Consequently, statistical analyses were performed to evaluate the clinical-pathological significance of MGP and RUNX2 in CRC. MGP protein was also detected by immunohistochemical analysis. ResultsShowed an overall overexpression of MGP in the tumor mucosa of patients at mRNA level when compared to adjacent normal mucosa and healthy control tissues. In addition, analysis of the expression of RUNX2 mRNA demonstrated an overexpression in CRC tissue samples and a positive correlation with MGP expression (Pearson correlation coefficient 0.636; p ≤ 0.01) in tumor mucosa. However correlations between MGP gene expression and clinical-pathological characteristics, such as gender, age and pathology classification did not provide relevant information that may shed light towards the differences of MGP expression observed between normal and malignant tissue. ConclusionsWe were able to associate the high levels of MGP mRNA expression with a worse prognosis and survival rate lower than five years.These results contributed to improve our understanding of the molecular mechanism underlying MGP deregulation in cancer.

Screening for differentially‑expressed microRNA biomarkers inSaudi colorectal cancer patients by small RNA deep sequencing.

Colorectal cancer (CRC) is mostly diagnosed at late stages leading to high mortality rates due to the scarcity of efficient screening approaches exhibiting high diagnostic utility. The current study employed a small-RNA deep-sequencing approach for screening microRNA (miRNA) differentially expressed genes (DEGs), and evaluating their potential as early diagnostic circulating biomarkers for CRC in clinical plasma and tissue samples from a Saudi patient population. The cohort followed a paired-study design composed of 20 CRC patients, providing plasma (P) and tissue (T) samples of CRC, and adjacent normal mucosa (CT). Also, control plasma (CP) samples were obtained from neoplasm-free healthy individuals to compare its miRNA levels with those in P samples. Illumina high-throughput (HiSeq 2000) sequencing was performed for the identification of known and novel miRNA genes that were differentially expressed in the plasma and tissues of CRC patients compared with CT and CP controls. While we identified only one known (hsa-miR-182-5p, significantly upregulated) and no novel DEGs at the most stringent significance level (P<0.001) in the P-CP comparison, we found 3 and none at P<0.01, 7 and 9 at P<0.05 level, respectively. In the T-CT comparison, the results revealed 24 known and 196 novel miRNA DEGs (P<0.001), 31 and 204 (P<0.01), 41 and 213 (P<0.05), respectively. Sequencing data were then analyzed by bioinformatics for potential diagnostic miRNAs. Network functional analysis for Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway implicated two pathways rooted to signal transduction [Wnt and mitogen-activated protein kinase (MAPK)] that were enriched in CRC patients. Our results suggest that characterizing plasma and tissue profiles of CRC by deep sequencing may be a good strategy for identifying known and novel miRNAs and that the validated miRNAs described here may serve as potential CRC-associated biomarkers. Further research is necessary for determining their screen index values and diagnostic utility for the diagnosis of CRC.

175 Clinical Usefulness of SMOC1 as a Diagnostic Marker for Colorectal Serrated Lesions and Colorectal Precancerous Lesions

INTRODUCTION: Colorectal serrated lesions (SLs) include hyperplastic polyp (HP), traditional serrated adenoma (TSA) and sessile serrated adenoma/polyp (SSA/P). SSA/Ps are well known precursors of colorectal cancer (CRC) characterized by BRAF mutation and microsatellite instability (MSI). However, clinical and molecular characteristics of TSAs are not fully understood. We have reported that epigenetic silencing of SMOC1 is associated with development of traditional serrated adenomas (TSAs) and that SMOC1 may play a role in neoplastic pathways in TSAs and conventional adenomas. 1 We aimed to clarify the clinical usefulness of SMOC1 in colorectal tumors. METHODS: Expression of SMOC1 was analyzed in a series of 127 colorectal tumors and adjacent normal colonic mucosa by immunohistochemistry (IHC). The staining intensity of SMOC1 was assessed as strong (3), moderate (2), weak (1) or negative (0). The proportions of positively stained tumor cells were assessed as 0 to 10 for each staining intensity. Because neoplasm heterogeneity caused varying degrees of immunoreactivity in respective specimens, we used the sum of each intensity × proportion as an IHC score (e.g., intensity × proportion = (3) × 5 + (2) × 3 + (1) × 1 + (0) × 1 = IHC score 22, scale of 0 to 30). RESULTS: Our IHC analysis revealed that SMOC1 is abundantly expressed in normal colon, HPs and SSA/Ps, while it was significantly downregulated in TSAs, high-grade adenomas and CRCs. Among serrated lesions, SMOC1 is expressed in HPs, SSA/Ps and SSA/Ps with CRC, while it was significantly downregulated in TSAs and TSAs with CRC. Notably, in TSAs which consisted of flat and protruding subcomponents, SMOC1 expression levels were lower in the protruding portions than those in the flat portions (P &lt; 0.001). CONCLUSION: These results suggest that downregulated expression of SMOC1 could be a hallmark of TSAs and conventional adenomas at high risk of developing CRC.