Lymph Node Metastasis In Colorectal Cancer Research Articles

Expression profiling and intracellular localization studies of the novel Proline-, Histidine-, and Glycine-rich protein 1 suggest an essential role in gastro-intestinal epithelium and a potential clinical application in colorectal cancer diagnostics.

BackgroundThe primary function of the intestines is the absorption of water and nutrients. Although our knowledge about these processes on the cellular level is extensive, a number of important intracellular elements remain unknown. Here, we characterize the novel proline-, histidine-, glycine-rich 1 (PHGR1) mRNA and protein on the molecular level and propose a functional role of the PHGR1 protein in the intestinal and gastric epithelium.MethodsPHGR1 mRNA and protein expression in human tissues and cell lines were characterized by quantitative RT-PCR, in situ hybridization, Northern blotting, Western blotting, and immunohistochemistry. Glycosylation was assessed by a chemical deglycosylation assay, whereas intracellular localization was studied by immunofluorescent staining of cell line cells. PHGR1 mRNA levels in HT29 cells was reduced by RNA interference and the resulting global changes in gene expression assessed by microarray hybridization.ResultsPHGR1 mRNA and protein were found to be expressed specifically in epithelial cells of intestinal mucosa, with the highest expression in the most mature and differentiated cells. PHGR1 protein was found to be glycosylated and to localize to both the cytoplasm and nucleus. Transcript profiling and gene ontology analysis of HT29 cells subjected to PHGR1 knockdown suggested a functional relationship with transport and metabolic processes. Examination of PHGR1 mRNA and protein levels in lymph nodes with known colorectal cancer metastases indicated that they may serve as biomarkers for detection of such metastases.ConclusionsFunctional analyses of the novel PHGR1 mRNA and protein suggest an essential role in gastrointestinal epithelium and a clinical application in detection of colorectal cancer lymph node metastases.

BCL6B suppresses proliferation and migration of colorectal carcinoma cells through inhibition of the PI3K/AKT signaling pathway.

B-cell CLL/lymphoma 6 member B (BCL6B), a BCL6-homologous gene, has been reported to be a tumor suppressor that is silenced in a variety of human cancers, including colorectal cancer (CRC). Although it was recently demonstrated that reduced expression of BCL6B is associated with tumor stage and lymph node metastasis in CRC, little is known on whether BCL6B contributes to CRC development, or the related underlying mechanism. The aim of the present study was to detect BCL6B expression in CRC cells, and determine the molecular mechanisms underlying the role of BCL6B in CRC development by investigating cell proliferation and migration in vitro. As a result, BCL6B expression was found to be notably repressed in CRC cells compared with normal intestinal epithelial cells by reverse transcription-polymerase chain reaction and western blot analysis. CRC cell proliferation was significantly inhibited by BCL6B upregulation, as indicated by MTT and colony-forming assays. Cell apoptosis was markedly induced, as indicated by flow cytometry, and BCL6B-transfected CRC cells exhibited decreased migration ability. Additionally, BCL6B overexpression diminished the phosphorylation level of AKT in CRC cells. These effects of BCL6B were empowered by treatment with the specific phosphoinositide 3 kinase (PI3K)/AKT inhibitor LY294002. Furthermore, overexpression of BCL6B resulted in upregulation of E-cadherin and downregulation of cyclin D1 and matrix metalloproteinase-9, which were strongly enhanced by LY294002. In conclusion, the findings of the present study demonstrated that BCL6B suppressed the proliferation and migration of CRC cells indirectly, via inhibition of PI3K.

Angiotensin-Converting Enzyme Gene Deletion Polymorphism is Associated with Lymph Node Metastasis in Colorectal Cancer Patients in a Chinese Population.

BackgroundThe purpose of this study was to assess the effect of angiotensin-converting enzyme (ACE) gene insertion/deletion (I/D) polymorphism on the risk of lymph node metastasis (LNM) in colorectal cancer (CRC) patients.Material/MethodsWe enrolled 146 CRC patients and 106 healthy controls in this study. ACE gene I/D polymorphism was genotyped by polymerase chain reaction (PCR). Hardy-Weinberg equilibrium (HWE) was used to assess the goodness of fit of the genotypes. χ2 test was used to calculate the differences of genotype and allele distributions. Odds ratios (ORs) with corresponding 95% confidence intervals (95% CIs) were used to analyze the association between ACE I/D polymorphism and LNM in CRC patients.ResultsInsertion/deletion (ID) and deletion/deletion (DD) genotypes were frequently observed in CRC patients, but only DD genotype and D allele were related to the susceptibility of CRC (P=0.038, OR=2.158, 95%CI=1.039–4.480; P=0.026, OR=1.501, 95%CI=1.048–2.150). DD genotype and D allele also increased the risk of LNM in CRC patients (P=0.028, OR=2.844, 95%CI=1.107–7.038; P=0.026, OR=1.692, 95%CI=1.063–2.693).ConclusionsDD genotype and D allele of ACE gene I/D polymorphism might increase the risk of LNM in CRC patients.

Upregulation of lncRNA BANCR associated with the lymph node metastasis and poor prognosis in colorectal cancer

BackgroundGrowing evidence has supported that long non-coding RNAs (lncRNAs) could play vital roles in the development, progression, and prognosis of colorectal cancer (CRC). However, little is known about the clinical significance of BRAF-activated non-coding RNA (BANCR) in CRC. The aim of this study is to explore the clinical value of lncRNA BANCR in CRC patients.MethodsThe expression of lncRNA BANCR was measured in 106 CRC tissues and 65 adjacent normal tissues using the quantitative real-time PCR.ResultsThe study showed that lncRNA BANCR was highly expressed in CRC tissues compared with adjacent normal tissues (P < 0.001). In addition, high expression of lncRNA BANCR was positively correlated with the lymph node metastasis (P < 0.001). Kaplan–Meier analysis showed that patients with high lncRNA BANCR expression had a shorter overall survival (OS) compared with the low lncRNA BANCR expression group (P = 0.001). Interestingly, for the group of patients with the lymph node metastasis, we found the similar result that high lncRNA BANCR expression was related to poor OS (P = 0.004). Furthermore, the multivariate Cox regression model analysis indicated that high expression of lncRNA BANCR was an independent poor prognostic factor in CRC patients (HR 2.24, 95% CI 1.22–4.16, P = 0.009).ConclusionsUpregulation of lncRNA BANCR may be associated with the lymph node metastasis and poor survival of CRC. LncRNA BANCR could be served as a novel and useful biomarker for CRC lymph node metastasis and prognosis.

MicroRNA-130a is upregulated in colorectal cancer and promotes cell growth and motility by directly targeting forkhead box F2.

Colorectal cancer (CRC) is one of the most prevalent cancers among males and females worldwide. Despite progress in diagnostic and therapeutic strategies for CRC patients, the prognosis for patients with advanced CRC remains poor. MicroRNAs (miRNAs/miRs) are a class of highly conserved short, endogenously expressed and single-stranded non-coding RNAs. In recent years, increasing studies have demonstrated that dysregulation of miRNAs is closely associated with CRC carcinogenesis and progression. The aim of the present study was to explore the expression, roles and underlying molecular mechanism of miR-130a in CRC. The results indicated that miR-130a was significantly upregulated in CRC, and that miR-130a expression levels were correlated with TNM stage and lymph node metastasis of CRC. Inhibition of miR-130a markedly suppressed colorectal cancer cell proliferation, migration and invasion. Furthermore, forkhead box F2 (FOXF2) was identified as a direct downstream target gene of miR-130a in colorectal cancer. Downregulation of FOXF2 could partially reverse the functions induced by miR-130a under-expression in CRC cells. These findings suggested that miR-130a can regulate FOXF2 and function as an oncogene in CRC. Therefore, miR-130a may serve as a useful therapeutic agent for miRNA-based CRC targeted therapy.

MicroRNA-30a-5p (miR-30a) regulates cell motility and EMT by directly targeting oncogenic TM4SF1 in colorectal cancer.

Colorectal cancer (CRC) is one of the leading causes of cancer death worldwide, and many oncogenes and tumor suppressor genes are involved in CRC. MicroRNAs (miRNAs) are small non-coding RNAs that can negatively regulate gene expression. Previous studies have revealed that miRNAs regulate the development and progression of many cancers. In this study, we investigated the role of microRNA-30a-5p (miR-30a) in CRC and its unknown mechanisms. qRT-PCR was used to detect miR-30a and TM4SF1 mRNA expression in CRC specimens and cell lines. CRC cell migration and invasion were assessed after transfection with miR-30a or TM4SF1 using wound healing and trans-well migration and invasion assays. Transmembrane-4-L-six-family protein (TM4SF1) was validated as a target of miR-30a in CRC through luciferase reporter assay and bioinformatics algorithms. Moreover, two EMT regulators, E-cadherin and VEGF, were also identified using Western blotting and immunohistochemistry. We found that miR-30a was down-regulated in CRC tumor tissues and cell lines, and miR-30a was inversely associated with advanced stage and lymph node metastatic status compared with normal tissues. miR-30a decreased migration and invasion in CRC cell lines, and miR-30a overexpression not only down-regulated TM4SF1 mRNA and protein expression, but also inhibited the expression of VEGF and enhanced expression of E-cadherin. We also showed that TM4SF1 was up-regulated in CRC tumor specimens compared with adjacent normal tissues, and TM4SF1 expression was significantly associated with advanced stage and lymph node status compared with adjacent normal tissues. These results suggest that miR-30a is an important regulator of TM4SF1, VEGF, and E-cadherin for CRC lymph node metastasis, a potential new therapeutic target in CRC.

Long non-coding RNA FBXL19-AS1 plays oncogenic role in colorectal cancer by sponging miR-203

Long non-coding RNAs (lncRNAs) have emerged as critical regulators of the progression of human cancers, including colorectal cancer (CRC). The study of genome-wide lncRNA expression patterns in metastatic CRC could provide novel mechanism underlying CRC carcinogenesis. In here, we determined the lncRNA expression profiles correlating to CRC with or without lymph node metastasis (LNM) based on microarray analysis. We found that 2439 lncRNAs and 1654 mRNAs were differentially expressed in metastatic CRC relative to primary CRC. Among these dysregulated lncRNAs, FBXL19-AS1 was the most significantly upregulated lncRNA in metastatic tumors. Functionally, knockdown of FBXL19-AS1 played tumor-suppressive effects by inhibiting cell proliferation, migration and invasion in vitro and tumor growth and metastasis in vivo. Overexpression of FBXL19-AS1 was markedly correlated with TNM stage and LNM in CRC. Bioinformatics analysis predicted that miR-203 was potentially targeted by FBXL19-AS1, which was confirmed by dual-luciferase reporter assay. Pearson's correlation analysis showed that miR-203 expression was negatively related to FBXL19-AS1 in tumor tissues. Finally, miR-203 inhibition abrogated the effect of FBXL19-AS1 knockdown on the proliferation and invasion of LoVo cells. Our results reveal the cancer-promoting effect of FBXL19-AS1, acting as a molecular sponge in negatively modulating miR-203, which might provide a new insight for understanding of CRC development.

Genetic variation of long non-coding RNA TINCR contribute to the susceptibility and progression of colorectal cancer.

Colorectal cancer (CRC) accounts for the leading causes of cancer-related morbidity and mortality. However, a large part of heritable factors are warranted to be explored. Long non-coding RNAs (lncRNAs) serve critical roles in cancer development and progression. Herein, we explored effect of genetic variants of Tissue differentiation-inducing non-protein coding RNA (TINCR), a key lncRNA required for somatic tissue differentiation and tumor progression, on risk and progression of CRC. Three tagSNPs, including rs2288947, rs8105637, and rs12610531, were evaluated in in a two-stage, case-control study. Two SNPs, rs2288947 and rs8105637, were significantly associated with susceptibility of CRC in both stages. When pooled together, the allele G was significantly associated with 23% decreased risk of CRC (OR=0.77; 95% CI=0.67-0.88; P value = 1.2×10−4)for SNP rs2288947. While for SNP rs8105637, the allele A was significantly associated with 22% increased risk of CRC (OR=1.22; 95% CI=1.09-1.37; P value = 6.2×10−4). The two SNPs were also statistically associated with occurrence of lymph node metastasis of CRC. The carriers of allele G are less likely to get lymph node metastasis (OR=0.77; 95% CI=0.63-0.94; P value = 0.011) for rs2288947, and the carriers of allele A are more likely to get lymph node metastasis (OR=1.22; 95% CI=1.03-1.43; P value = 0.019) for rs8105637. These results suggest that lncRNA TINCR polymorphisms may be implicated in the development and progression of CRC.

Abstract PR07: Complex sub-clonal populations in colorectal cancer lymph node metastasis

Abstract A number of studies have highlighted the role of sub-clones in tumor progression across multiple cancer types including colorectal cancer (CRC). However, the genetic origin of lymph node (LN) metastasis is not well understood. Given that the prognosis for CRC patients worsens once metastasis occurs, we aimed to better understand the relationship between detected sub-clones in the primary tumor and the LN metastasis. Two or three spatially disparate regions from each of 5 colon cancers were dissected and subjected to next-generation whole exome DNA sequencing (average read depth of 50x) followed by targeted confirmatory deep sequencing (average read depth of 420x). The number of mutations per tumor ranged from 95 to 334 in the 4 non-hypermutated tumors and was 1208 in the single hypermutated tumor. DNA was then isolated from the metastatic areas of two to four matched metastatic LN's from each tumor. Deep-targeted sequencing was also performed on the LN samples targeting the same somatic variant locations identified in the primary tumors. Affymetrix OncoScan SNP arrays were performed on all primary tumor and LN samples to assess copy number (CN) variation and to permit more accurate inference of the cellular frequency of each somatic variant using the computational method PyClone. The PyClone software implements a probabilistic model to estimate the cellular frequency of each mutation in a population of cells using observed alternate allele frequencies as well as copy number and loss of heterozygosity (LOH) information, resulting in a clustering of variants based on the cellular frequency patterns of variants in each sample. Sub-clones are inferred corresponding to the distinct groupings of variants across all tumor and LN samples from a single patient. Additionally, the Clomial software was utilized to complement and validate PyClone results. Clomial implements an expectation-maximization algorithm to infer, given a count of distinct clones expected in a sample, profiles of the variant genotypes of each sample and sample clonal compositions. Generating Clomial profiles at several possible expected clone counts provides an orthogonal measure of clonal genotypes, as well as generating tumor purity estimates and facilitating phylogenetic profiling of variants and clones. CN and LOH differences were identified between different spatial locations in some individual tumors as well as between different LN's from the same tumor. Some LN's contained the same CN changes as a particular spatial region of the matched primary tumor and others contained CN variants that were distinct from the tested regions of the primary tumor. As defined by PyClone, each region in the primary tumor contained multiple (2–13, average 6.1) distinct sub-clones and each LN contained 3 - 13 sub-clones (average 5.9). We demonstrate evidence that individual LN metastasis are comprised of multiple sub-clones from the primary tumor. When there are multiple independent LN metastases in an individual patient, they do not all have the same composition of sub-clones. Some metastatic sub-clones were shared among all of the LN's from the same tumor but there were also sub-clones that were private to a portion of the tumor and to an individual LN. This may be evidence of individual sub-clones metastasizing at different times to different LN's. Further study of the origins of metastasis may help elucidate how to better target it therapeutically. This abstract is also being presented as Poster A10. Citation Format: Karin M. Hardiman, Peter J. Ulintz, Joel K. Greenson, Rong Wu, Eric R. Fearon. Complex sub-clonal populations in colorectal cancer lymph node metastasis. [abstract]. In: Proceedings of the AACR Special Conference on Colorectal Cancer: From Initiation to Outcomes; 2016 Sep 17-20; Tampa, FL. Philadelphia (PA): AACR; Cancer Res 2017;77(3 Suppl):Abstract nr PR07.

Clostridium difficile colonization in preoperative colorectal cancer patients

The entire process of Clostridium difficile colonization to infection develops in large intestine. However, the real colonization pattern of C. difficile in preoperative colorectal cancer patients has not been studied. In this study, 33 C. difficile strains (16.1%) were isolated from stool samples of 205 preoperative colorectal cancer patients. C. difficile colonization rates in lymph node metastasis patients (22.3%) were significantly higher than lymph node negative patients (10.8%) (OR=2.314, 95%CI=1.023-5.235, P =0.025). Meanwhile, patients positive for stool occult blood had lower C. difficile colonization rates than negative patients (11.5% vs. 24.0%, OR=0.300, 95%CI=0.131-0.685, P =0.019). A total of 16 sequence types were revealed by multilocus sequence typing. Minimum spanning tree and time-space cluster analysis indicated that all C. difficile isolates were epidemiologically unrelated. Antibiotic susceptibility testing showed all isolates were susceptible to vancomycin and metronidazole. The results suggested that the prevalence of C. difficile colonization is high in preoperative colorectal cancer patients, and the colonization is not acquired in the hospital. Since lymph node metastasis colorectal cancer patients inevitably require adjuvant chemotherapy and C. difficile infection may halt the ongoing treatment, the call for sustained monitoring of C. difficile in those patients is apparently urgent.

High expression of ADAMTS5 is a potent marker for lymphatic invasion and lymph node metastasis in colorectal cancer.

Members of the ADAMTS family contain propeptide, metalloproteinase and disintegrin domains and serve key roles for cancer cell proliferation, progression and metastasis. Although overexpression of ADAMTS5 has been reported in glioblastoma, and head and neck cancer, little has been demonstrated in colorectal cancer types. The present study aimed to clarify the significance of ADAMTS5 for clinicopathological factors and prognosis in colorectal cancer. The mRNA expression of ADAMTS5 was measured in 143 colorectal cancer specimens. ADAMTS5 expression was increased as the pathological stage increased. The expression of ADAMTS5 in stage III-IV colorectal cancer was significantly greater compared with that of stage 0-II colorectal cancer (P=0.0003). The median expression of ADAMTS5 was used to divide the ADAMTS5 higher expressing group and the ADAMTS5 lower expressing group to assess the correlation of ADAMTS5 expression with clinicopathological factors and prognosis. The proportions of lymphatic invasion and lymph node metastasis were significantly greater in the ADAMTS5 higher expressing group (P=0.0214 and P=0.0289 respectively). Overall survival and disease-free survival were assessed by the Kaplan-Meier curve with calculation of significance by the log-rank test; however, no significant difference was observed between the ADAMTS5 higher expressing group and the ADAMTS5 lower expressing group (P=0.7490 and P=0.9455, respectively). The present study confirmed high expression of ADAMTS5 as a potent marker for lymphatic invasion and lymphnode metastasis in colorectal cancer. To clarify the function of ADAMTS5 in colorectal cancer, further molecular investigations are required.

Midkine is up-regulated in both cancerous and inflamed bowel, reflecting lymph node metastasis in colorectal cancer and clinical activity of ulcerative colitis

Midkine is a multifunctional cytokine and growth factor displaying proinflammatory and pro-tumorigenic activity. Its association with bowel diseases has not been fully elucidated. Our purpose was to delineate midkine expression pattern by RT-qPCR in inflamed/cancerous bowel (n=208) and whole blood (n=150) in colorectal cancer (CRC), Crohn’s disease (CD), and ulcerative colitis (UC) and to evaluate midkine dynamics in early postoperative period following colorectal surgery. The expression of midkine was significantly up-regulated in stage III CRC and independently associated with lymph node metastasis. The expression of midkine in whole blood was up-regulated solely in N1 CRC. Midkine expression in cancer-free tissue (CRC) was also elevated and dependent on CRC advancement. In IBD, inflammation increased the bowel expression of midkine solely in UC, in a manner proportional to the disease clinical activity. Large and small bowel differed with respect to the expression of midkine in quiescent tissue (higher in small bowel) and to its correlation pattern with chemokines (in a large bowel) and angiogenic factors and cell cycle regulators (in a small bowel). Circulating midkine and its expression in whole blood dropped directly following colorectal surgery; however, the concentration of midkine in serum was restored on postoperative day three.Midkine is involved in bowel inflammation in UC and lymph node metastasis in CRC, rendering midkine an attractive target for their treatment. Owing to midkine elevation in early postoperative period and its overexpression in tumor-adjacent tissue, targeting midkine might be considered also as a prevention of CRC recurrence following curative tumor resection.

Correlation between quantitative expression of T lymphoma invasion and metastasis inducing factor 1 mRNA and lymph node micrometastasis and prognosis in patients with colorectal cancer

Objective To investigate the relationship between the changes of expression of T lymphoma invasion and metastasis inducing factor 1(Tiam1) mRNA and lymph node micrometastasis and prognosis in patients with node-negative colorectal cancer(CRC). Methods From June 2008 to August 2010, a total of 63 patients with lymph node-negative CRC, 20 patients with lymph node-positive CRC and 25 patients with colorectal benign lesions were enrolled. The relative quantity expression (RQ) of Tiam1 mRNA in CRC cancer tissues and colorectal benign lesions were detected by real-time quantitative polymerase chain reaction (PCR). The lymph node micrometastasis was determined by cytokeratin 20 detection in lymph nodes tissues. The five-year postoperative prognosis was evaluated by followed-up in the patients with lymph node-negative CRC. The correlation between the expression of Tiam1 mRNA and lymph node micrometastasis and prognosis was analyzed. The t-test, univariate analysis, multivariate analysis and Log-rank test were performed for statistical analysis. Results Lymphnode micrometastasis occurred in 33 from 63 patients with lymph node-negative CRC. The RQ of Tiam1 mRNA in the patients with lymph node-positive CRC was 9.84±2.36, which was higher than that of patients with lymph node-negative CRC (5.15±3.58), and the difference was statistically significant (t=5.479, P<0.01). The RQ of Tiam1 mRNA of these two groups were both higher than that of colorectal benign lesion (0.30±0.21), and the difference was statistically significant(t=20.169, 6.745; both P<0.01). The RQ of Tiam1 mRNA of positive lymph node micrometastasis group was 6.30±1.95, which was higher than that of negative lymph node micrometastasis group (3.88±1.63), and the difference was statistically significant (t=5.330, P<0.01). In the patients with lymph node-negative CRC, the lower degree of tumor difference, the deeper invasion, the shorter survival time after surgery and postoperative recurrence and metastasis were related with the higher RQ of Tiam1 mRNA in tumor tissues (t=2.536, 3.112, 3.213, 2.676; all P<0.05). Univariate analysis revealed that the expression of Tiam1 mRNA in cancer tissues was correlated with lymph node micrometastases(χ2 =11.878, P=0.001). Multivariate analysis showed that it was an independent risk factors of lymph node micrometastases in CRC(relative risk: 9.782). The five-year postoperative cumulative survival rate of high Tiam1 mRNA expression group was 75.8%, which was lower than that of low expression group (97.1%) , and the difference was statistically significant (χ2 =4.575, P<0.05). Conclusions Tiam1 may participate in the regulation of lymph node metastasis in CRC. Elevated expression of Tiam1 promotes lymph node metastasis and is closely relatived with poor prognosis of node-negative CRC, and which can be considered as an indicator of prognosis. Key words: Tiam1 mRNA; Colorectal cancer; Lymph node micrometastases; Prognosis

Immunohistochemical expression of Notch1, Jagged1, NF-κB and MMP-9 in colorectal cancer patients and the relationship to clinicopathological parameters.

Activation of Notch and NF-κB signaling has been frequently observed in colorectal cancer (CRC) patients and contributes to the chemo-resistance and treatment failure. However, the relationship between these signaling pathways and CRC has not been clearly described. To investigate the expression of Notch1, Jagged1, NF-κB and MMP-9 in CRC patients and analyze their correlation with clinicopathological factors. Expression of Notch1, Jagged1, NF-κB and MMP-9 was visualized by immune-histology in the tumor tissue, adjacent and distant normal tissues from 47 CRC patients without receiving chemotherapy or radiotherapy. Notch1 (80.8%), Jagged1 (80.8%), NF-κB (70.2%) and MMP-9 (76.6%) were overexpressed in cancer tissues compared normal tissues (P< 0.05). The intensity of Notch1, Jagged1 and NF-κB expression was associated with histological grading, depth of invasion, TNM staging and lymph node metastasis of CRC. MMP-9 was intimately correlated with depth of invasion, TNM staging and lymph node metastasis. NF-κB, MMP-9 and Notch1 expression was positively correlated (P< 0.05), and the same positive correlation was found among NF-κB, MMP-9 and Jagged1 expression (P< 0.05). Notch1, Jagged1, NF-κB and MMP-9 probably play a pivotal role during the CRC development, serving as biomarkers for early detection of the recurrence and metastasis and prognosis of CRC.

A High RORγT/CD3 Ratio is a Strong Prognostic Factor for Postoperative Survival in Advanced Colorectal Cancer: Analysis of Helper T Cell Lymphocytes (Th1, Th2, Th17 and Regulatory T Cells).

Tumor-infiltrating lymphocytes (TILs), part of the host immune response, have been widely reported as influential factors in the tumor microenvironment for the clinical outcome of colorectal cancer (CRC). However, the network of helper T cells is very complex, and which T-cell subtypes affect the progression of CRC and postoperative prognosis remains unclear. This study investigated the expression of several subtypes of TILs including T helper type 1 (Th1), Th2, Th17, and regulatory T (Treg) cells to determine their correlation with clinicopathologic features and postoperative prognosis. The study investigated the expression of TILs using immunohistochemistry of tissue microarray samples for 199 CRC patients. The number of each T-cell subtype infiltrating tumors was counted using ImageJ software. The relationship between TIL marker expression, clinicopathologic features, and prognosis was analyzed. A high RORγT/CD3 ratio (Th17 ratio) was significantly correlated with lymph node metastasis (p = 0.002), and a high of Foxp3/CD3 ratio (Treg ratio) was correlated with tumor location in the colon (p = 0.04), as shown by the Chi square test. In multivariate analysis, a high RORγT/CD3 ratio was the only independent prognostic factor for overall survival (p = 0.04; hazard ratio [HR], 1.84; 95% confidence interval [CI] 1.02-3.45). This study confirmed a high RORγT/CD3 ratio as a strong prognostic marker for postoperative survival. The immunohistochemistry results suggest that Th17 may affect lymph node metastasis in CRC. If new immunotherapies reducing Th17 expression are established, they may improve the efficiency of cancer treatment and prolong the survival of patients with CRC.

Expression of CD147 and matrix metalloproteinase-11 in colorectal cancer and their relationship to clinicopathological features.

BackgroundThis study aimed to investigate the expression of CD147 and MMP-11 in human colorectal cancer (CRC) and to evaluate their clinical significance.MethodsReal-time polymerase chain reaction was used to evaluate CD147 and MMP-11 mRNA level in 56 pairs of fresh CRC samples matched with adjacent normal mucosa. The protein expression of CD147 and MMP-11 in CRC specimens and corresponding normal colorectal mucosa were evaluated by immunohistochemistry on CRC tissue microarrays. Expression and co-localization of these two proteins in human colorectal cancer tissue were also evaluated by laser scanning confocal microscopy. Furthermore, their correlations with clinicopathological factors and overall survival after surgery were evaluated.ResultsBoth CD147 and MMP-11 were demonstrated to be over-expressed at mRNA level (P < 0.001, both) and protein level (P < 0.001, both) in CRC tissue than paired normal mucosa. Spearman rank test showed a positive correlation between these two proteins (P = 0.025). Immunofluorescence double staining confirmed the co-localization of CD147 and MMP-11 in paraffin-embedded tissues of CRC patients. Expression of CD147 and MMP-11 were both correlated with CRC lymph node metastasis (P = 0.021 and P = 0.031, respectively), distant metastasis (P < 0.001 and P = 0.013, respectively) and TNM stage (P = 0.006 and P = 0.049, respectively). Univariate survival analysis showed that both CD147 and MMP11 expression was significantly associated with shorter survival time (P = 0.001 and P = 0.009, respectively). Additionally, in multivariate analysis, both CD147 and MMP-11 were proved to be independent prognostic factors (P = 0.009, 0.028, respectively).ConclusionsThese results indicated that both CD147 and MMP-11 may be involved in the progression of colorectal cancer, and they are potential prognostic factors and might become new therapeutic targets for CRC patients.

Pri-miR-124 rs531564 polymorphism and colorectal cancer risk.

MiR-124 functions as a tumor suppressor and plays an important role in tumorigenesis. A common polymorphism (rs531564, C>G) in the pri-miR-124 has been recently studied in connection with cancer risk. The aim of the present study was to investigate the association between pri-miR-124 rs531564 polymorphism and the risk and clinicopathological characteristics of colorectal cancer (CRC). Two case-control studies involving 900 CRC patients and 1110 cancer-free controls showed that pri-miR-124 rs531564 polymorphism was significantly associated with the decreased risk of CRC in Xuzhou population [GG vs. CC: OR = 0.25, 95%CI = 0.09–0.67, P = 0.003; (CG+GG) vs. CC: OR = 0.73, 95%CI = 0.56–0.94, P = 0.01; GG vs. (CC+CG): OR = 0.27, 95%CI = 0.10–0.70, P = 0.004; G vs. C: OR = 0.70, 95%CI = 0.56–0.89, P = 0.003], Bengbu population [GG vs. CC: OR = 0.20, 95%CI = 0.04–0.90, P = 0.02; GG vs. (CC+CG): OR = 0.21, 95%CI = 0.05–0.95, P = 0.03; G vs. C: OR = 0.72, 95%CI = 0.54–0.98, P = 0.03] and pooled population [GG vs. CC: OR = 0.26, 95%CI = 0.11–0.59, P<0.001; (CG+GG) vs. CC: OR = 0.76, 95%CI = 0.62–0.93, P = 0.008; GG vs. (CC+CG): OR = 0.27, 95%CI = 0.12–0.62, P < 0.001; G vs. C: OR = 0.71, 95%CI = 0.59–0.85, P<0.001]. Additionally, pri-miR-124 rs531564 polymorphism was significantly associated with the decreased risk of poor differentiation and lymph node metastasis of CRC. Our results suggest that pri-miR-124 rs531564 polymorphism may be a genetic modifier for developing CRC. However, further studies are needed to validate our findings.

Dysregulation of over-expressed IL-32 in colorectal cancer induces metastasis.

BackgroundInterleukin (IL)-32 is a described intracellular pluripotent pro-inflammatory mediator, characterized by the signaling of NF-κB and STAT3.MethodsOur study investigated whether IL-32 expression has clinical significance in the metastases of colorectal cancer (CRC). A total of 70 CRC patients were enrolled, 47 cases of which were single CRC organic metastasis lesions while the rest of which were primary CRC lesions (T4NxM0). IL-32 expression was detected by immunohistochemistry, and the correlation between IL-32 expression and CRC metastases was analyzed.ResultsThe positive rates of IL-32 in the CRC organic metastasis group were more severe than those in the primary CRC group (P < 0.05). The positive rate of IL-32 in primary CRC with lymph node metastasis was more severe than that of IL-32 in primary CRC without lymph node metastasis (P < 0.05).ConclusionsThe level of IL-32 expression could influence the N grade of CRC. Thus, IL-32 expression may stimulate the organic metastasis and the lymph node metastasis of CRC.

Systematic review and meta-analysis of histopathological predictive factors for lymph node metastasis in T1 colorectal cancer.

In this study we examined whether histopathological findings, specifically lymphatic vessel invasion identified by an anti-human podoplanin antibody, and several other factors are associated with lymph node metastasis in T1 colorectal cancer. We searched PubMed and Cochrane Library, and also handsearched relevant journals, for reports written in English and published between 1998 and 2012, utilizing combination headings, such as 'colorectal cancer,' 'lymph node metastasis,' and 'risk factors.' For the report to be included in our study, the following criteria had to be met: (1) data on the frequency of lymph node metastasis in T1 colorectal cancer in relation to histopathological factors were reported; (2) patients had undergone bowel resection and had histologically diagnosed T1 colorectal cancer; (3) lymphatic vessel invasion was identified by immunohistochemistry with an anti-human podoplanin antibody rather than by hematoxylin and eosin staining; (4) univariate and multivariate analyses were conducted. Studies investigating molecular markers were excluded. The independent predictive factors were confirmed in at least one study included in the meta-analysis in the present systematic review. Microsoft Excel 2013 for Windows was used for the statistical analysis. Initially, 369 publications were identified in the database searches and handsearches, of which five ultimately met all of the inclusion criteria and selected for this systematic review. The meta-analysis revealed that only two factors were significantly associated with T1 colorectal cancer lymph node metastasis: (1) lymphatic vessel invasion identified by an anti-human podoplanin antibody [Mantel-Haenszel odds ratio (OR) 5.19; (95% confidence interval (CI) 3.31-8.15; P = 0.01]; (2) tumor budding (OR 7.45; 95 % CI 4.27-13.02; P = 0.0077). Our meta-analysis revealed that lymphatic vessel invasion identified by an anti-human podoplanin antibody and tumor budding were significantly associated with T1 colorectal cancer lymph node metastasis.

Downregulation of miR-193a-5p correlates with lymph node metastasis and poor prognosis in colorectal cancer.

To investigate the correlation of miR-193a-5p with lymph node metastasis and postoperative survival of colorectal cancer (CRC) patients. A total of 304 formalin-fixed, paraffin-embedded specimens (69 paired cancer and normal tissues, 55 primary tumors of stage III CRC and matched lymph nodes, and 56 primary tumors of stage II CRC) were included in this study. The relative expression levels of miR-193a-5p in the normal mucosa, primary cancer, and metastatic lymph node lesions were measured by quantitative real-time reverse transcriptase polymerase chain reaction. We evaluated the association of its expression with colorectal cancer lymph node metastasis, clinicopathological factors, and patient survival. The relative expression level of miR-193a-5p was significantly lower in CRC tissues than in the normal mucosa (P = 0.0060). The expression levels of miR-193a-5p were lower in primary CRC tissues with lymph node metastases than in those without metastases (P = 0.0006), and decreased expression of miR-193a-5p correlated with advanced lymph node metastatic stage (P = 0.0007). Kaplan-Meier analysis showed that patients with low miR-193a-5p expression had decreased disease-free survival (DFS) (P = 0.0026) and poor overall survival (OS) (P = 0.0003). Interestingly, for the group of patients with lymph node metastases, miR-193a-5p expression was also related to survival. Patients with low miR-193a-5p expression had decreased DFS (P = 0.0262) and poor OS (P = 0.0230). Moreover, multivariate analysis indicated that downregulation of miR-193a-5p was an independent predictor of poor OS. Downregulation of miR-193a-5p correlates with lymph node metastasis and poor survival of CRC. miR-193a-5p may be a useful biomarker for CRC diagnosis, metastasis and prognosis prediction.