Identification of molecular markers for metastasis-related genes in primary breast cancer cells

Comparing differential gene expression profiles established by cDNA microarray between normal cells (N), primary carcinoma cells (T), and metastatic carcinoma cells (M) may determine those critical genes directly associated with progression and metastasis of breast cancer. Total RNA was extracted by laser microdissection (LMD) from 20 slices of T, N and M from 6 cases. After amplification by a T7-based system, differentially expressed genes between T, N and M were identified by cDNA microarray. In addition, to clarify the mechanism for altered gene expression, we determined the methylation status by sequencing after bisulfite treatment for intriguing genes. As a result, the expression of motility related protein-1 (MRP-1/CD9), peripheral myelin protein-22 (PMP-22), and caspase 3 (CASP-3) were down-regulated in M compared to T. We focused especially on MRP-1 and found that the expression status of MRP-1 was significantly inversely associated with stage of disease in 56 cases of breast cancer (P<0.05), and the relapse free survival in 5 years was significantly higher in MRP-1 positive cases than those negative cases (P<0.05). Conversely, overexpression, by 11-fold, of signal transduction and translation factors were observed in T compared to N. The cancer specific methylation was observed only in CASP-3 in a case. In conclusion, the establishment of the present assay allows us to detect genes directly associated with each cell population within tumor tissue and gives us clues to identify metastasis-related genes comprehensively in clinical breast cancer cases.     

负性共刺激分子B7-H4在结直肠癌及其微环境中的表达及临床意义

负性共刺激分子B7-H4是新近发现的B7家族的新成员。为了检测人结直肠癌组织中肿瘤细胞及肿瘤浸润淋巴细胞(tumor-infiltrating lymphocyte,TIL)上B7-H4的表达以及分析巨噬细胞、T淋巴细胞及其亚群的浸润,进而探讨其相关临床意义,本课题采用免疫组化SP法检测98例人结直肠癌组织中肿瘤细胞及浸润淋巴细胞上B7-H4的表达以及巨噬细胞、T细胞及其亚群的浸润,统计学分析B7-H4的表达与患者临床病理参数、预后、肿瘤相关巨噬细胞(tumor-associated mac-rophage,TAM)以及CD3+T细胞、CD8+T细胞浸润程度的相关性。结果显示,结直肠癌组织中高表达B7-H4分子,而正常组织中基本不表达,并且在肿瘤细胞及浸润淋巴细胞中均检测到B7-H4的表达。统计学分析表明,浸润淋巴细胞中B7-H4的表达与患者年龄、有无淋巴结转移、是否为粘液腺癌有关(P<0.05),与患者总体生存率呈负相关(P<0.05),且B7-H4高表达组织较B7-H4低表达组织CD3+T细胞高度浸润例数显著增加(P<0.05);肠癌细胞中B7-H4的表达与患者肿瘤分期、淋巴结转移、有无远处转移以及Dukes分期有关(P<0.05)。综上所述,人结直肠癌患者的肠癌细胞及浸润淋巴细胞较正常细胞均异常表达B7-H4分子,CD3+T细胞的浸润程度与浸润淋巴细胞中B7-H4的表达呈显著相关而与肿瘤细胞上B7-H4的表达无关,而肿瘤细胞表达B7-H4与肿瘤分期、淋巴结转移、有无远处转移以及Dukes分期有关。上述结果提示肿瘤细胞表达B7-H4与浸润淋巴细胞表达B7-H4可能分别以不同机制参与了肿瘤的发生和进展,进一步分析肿瘤组织中B7-H4在不同细胞表达的生物学和临床意义具有重要的价值。     

High phospho-histone H3 expression uniquely predicts favorable survival among four markers of cellular proliferation in colorectal cancer

Colorectal cancer (CRC) is one of the most frequent gastrointestinal cancers worldwide, with high morbidity and mortality rates. Despite numerous attempts to identify prognostic markers for the CRC patients, the significance of the association of cellular proliferation markers with survival is controversial. Here we used immunohistochemistry to detect four markers of cellular proliferation expressed in primary CRC tissue specimens (n = 269) to assess their potential to serve as prognostic factors. CRC cells variably expressed phospho-histone H3 (PHH3) (range, 0–76 per high-powered field (HPF); median, 7 per HPF), cyclin A (CCNA) (range, 11.3–73.7%; median, 32%), geminin (GMNN) (range, 7.8–82.0%; median, 37.1%), and marker of proliferation Ki-67 (MKI67) (range, 4.9–96.6%; median, 49.6%). Among them, patients with PHH3-high (≥7 per HPF) tumors uniquely experienced significantly longer 5-year survival than those with PHH3-low (≤6 per HPF) (81.8% vs. 65.5%; P = 0.0047). Multivariable Cox hazards regression analysis identified PHH3-high (hazard ratio, 0.54; 95% confidence interval, 0.31–0.92; P = 0.025) as potential favorable factors. PHH3 levels inversely associated with pT stage (P < 0.0001) and were significantly and inversely associated with tumor diameter (ρ = −0.314, P < 0.0001). These findings support the use of PHH3 immunohistochemistry for predicting the prognoses of patients with CRC.     

Genome and transcriptome profiles of CD133-positive colorectal cancer cells

Colorectal carcinomas (CRC) might be organized hierarchically and contain a subpopulation of tumorigenic, putative cancer stem cells that are CD133 positive. We studied the biological and genetic characteristics of such cells in CRC cell lines and primary tumors. Three CRC cell lines were sorted in CD133 positive and negative fractions. The respective genetic aberration profiles were studied using array comparative genomic hybridization (aCGH) and expression profiling. Tumorigenicity for each cellular population was tested by injection into nude mice. Additionally, we compared CD133+ and CD133- cells of 12 primary colorectal tumors using laser capture microdissection and aCGH. Three of five CRC cell lines displayed both CD133+ and CD133- cells, but tumorigenicity of these subfractions did not differ significantly and aCGH revealed essentially identical genomic imbalances. However, 96 genes were differentially expressed between the two populations. Array comparative genomic hybridization analysis after laser capture microdissection of CD133+ and CD133- areas in primary colorectal tumors revealed genetic differences in 7 of 12 cases. The use of cell lines for studying genomic alterations that define cancer stem cell characteristics, therefore, seems questionable. In contrast, CD133+ cells in primary cancer samples showed a unique genomic aberration profile. In conclusion, our data suggest that CD133 positivity defines a genetically distinct cellular compartment in primary CRC, which potentially includes tumor initiating cells.     

Focal enhancement of expression of c-Met/hepatocyte growth factor receptor in the myocardium in human myocardial infarction.

To determine the distribution and expression level of hepatocyte growth factor (HGF) specific receptor, c-Met, in human myocardial infarction. Autopsies of 13 patients who died without heart diseases (control) and 13 patients with a history of myocardial infarction (2 h to 10 years before death). The harvested myocardial tissues were stained with hematoxylin-eosin (H&E) and immunohistochemically stained for c-Met expression by the avidin-biotin-horseradish peroxidase complex method using an antibody to c-Met. C-Met expression was only slightly increased in control subjects and in noninfarcted myocardium of the test group. In contrast, high expression was noted in the peripheral region of the myocardial infarction and in some hypertrophic myocardial cells. C-Met was not expressed in the infarcted myocardium, but overexpression was noted in the surrounding myocardial cells of blood vessels and in the subendocardium and subepicardium in a band-like pattern. The expression level of c-Met was most enhanced at the time of appearance of coagulative necrosis and least in the myocardium of subjects with old infarcts. Our results indicate that HGF preferentially reaches the ischemic regions of the myocardium and has local and direct effects on the myocardium in patients with myocardial infarction.     

Overexpression of nuclear NHERF1 in advanced colorectal cancer: association with hypoxic microenvironment and tumor invasive phenotype

Over 57% of colorectal cancer (CRC) patients have regional or distant spread of their disease at the time of diagnosis. Despite recent advances, there is a compelling need to better characterize prognostic markers for advanced CRC. The present study investigates protein expression of NHERF1, HIF-1α and TWIST1 and their relationship in distant normal mucosa (DNM), tumor (T) and adjacent normal mucosa (ANM), lymph node metastasis (LNM) and liver metastasis (LM), determining their role as potential markers in advanced stages of human CRC. Overexpression of nuclear NHERF1 was shown in 47% of tumors, which exhibited a significant association with poor histological grade (P=0.0346). Nuclear NHERF1 showed a higher expression in T, LNM and LM than both DNM (P<0.0001) and ANM (P<0.05). Nuclear HIF-1α was significantly higher in T, LNM and LM than DNM and ANM (P<0.05, P<0.001, P<0.0001, respectively). A positive correlation between nuclear NHERF1 and nuclear HIF-1α was found in LNM (r=0.331, P=0.020), where an extended co-localization of the two proteins was demonstrated. TWIST1 was more expressed in T than DNM and ANM (P<0.0001) and was higher in T than LNM and LM (P<0.0001). Moreover, nuclear NHERF1 was directly correlated to TWIST1 (r=0.339, P=0.015) in T samples, where a high co-expression of the two proteins was demonstrated both in no longer polarized epithelial cells and in invasive mesenchymal elements adjacent to hypoxic and perinecrotic colonic areas. Overall, nuclear NHERF1 expression was associated with poorer differentiation grade and with higher expression both of HIF-1α in lymphatic metastasis and TWIST1 in invasive front of tumor. Our results support the oncogenic role of NHERF1 and promote nuclear NHERF1 as a potential new biomarker of advanced CRC.     

Expression of c-Met and heparan-sulfate proteoglycan forms of CD44 in colorectal cancer

In colorectal cancer patients, prognosis is not determined by the primary tumor but by the formation of distant metastases. Molecules that have been implicated in the metastatic process are the proto-oncogene product c-Met and CD44 glycoproteins. Recently, we obtained evidence for functional collaboration between these two molecules: CD44 isoforms decorated with heparan sulfate chains (CD44-HS) can bind the c-Met ligand, the growth and motility factor hepatocyte growth factor/scatter factor (HGF/SF). This interaction strongly promotes signaling through the receptor tyrosine kinase c-Met. In the present study, we explored the expression of CD44-HS, c-Met, and HGF/SF in the normal human colon mucosa, and in colorectal adenomas and carcinomas, as well as their interaction in colorectal cancer cell lines. Compared to the normal colon, CD44v3 isoforms, which contain a site for HS attachment, and c-Met, were both overexpressed on the neoplastic epithelium of colorectal adenomas and on most carcinomas. Likewise, HGF/SF was expressed at increased levels in tumor tissue. On all tested colorectal cancer cell lines CD44v3 and c-Met were co-expressed. As was shown by immunoprecipitation and Western blotting, CD44 on these cells lines was decorated with HS. Interaction with HS moieties on colorectal carcinoma (HT29) cells promoted HGF/SF-induced activation of c-Met and of the Ras-MAP kinase pathway. Interestingly, survival analysis showed that CD44-HS expression predicts unfavorable prognosis in patients with invasive colorectal carcinomas. Taken together, our findings indicate that CD44-HS, c-Met, and HGF/SF are simultaneously overexpressed in colorectal cancer and that HS moieties promote c-Met signaling in colon carcinoma cells. These observations suggest that collaboration between CD44-HS and the c-Met signaling pathway may play an important role in colorectal tumorigenesis.     

Cyclooxygenase-2 expression correlates with uPAR levels and is responsible for poor prognosis of colorectal cancer

Considering recent findings that cyclooxygensase-2 (COX-2) is involved in the progression of colorectal carcinoma (CRC), the role of COX-2 in promoting invasion and angiogenesis was investigated by evaluating the relationship of COX-2 expression to various clinicopathological variables, including plasminogen activating system (PA system) and vascular endothelial growth factor (VEGF). Tumor tissues from 71 patients with CRC were assayed to determine the antigen levels of urokinase-type plasminogen activator (uPA), uPA receptor (uPAR), and plasminogen activator inhibitor-1 and -2 (PAI-1 and PAI-2), as well as immunohistochemical expression of VEGF. COX-2 was assayed immunohistochemically in 56 patients. COX-2 expression was detected in cancer cells and it was also expressed by stromal cells in some patients. Fourteen patients (25%) were COX-2 positive, whereas 42 were negative. COX-2 expression was significantly related to lymphatic invasion (P=0.0317), but was not related to microvessel density or VEGF expression. In the PA system, uPAR antigen levels were significantly higher in tumors with COX-2 expression than in tumors without (P=0.0233). Univariate analysis showed that significant prognostic variables for survival were tumor size, lymph node involvement, lymphatic invasion, vascular invasion, liver metastasis, high uPAR level, and COX-2 expression, but only liver metastasis was an independent prognostic factor (P=0.0065) in multivariate analysis. COX-2 expression was a more important prognostic indicator than any other factor except liver metastasis (P=0.0526). The significant relationship between the presence of COX-2 protein and uPAR antigen levels contributed to the enhancement of tumor invasion and the poor outcome in patients with CRC. This revised version was published online in July 2006 with corrections to the Cover Date.     

Comparison of cyclooxygenase-2 and CD44 mRNA expression in colorectal cancer and its relevance for prognosis

This study evaluated CD44 and COX-2 expression in colorectal cancer (CRC) and analyzed its relationship with the clinicopathological characteristics. The prognostic impact on patient survival was compared between the two proteins. CD44 and COX-2 mRNA levels in 42 primary CRCs were analyzed using quantitative real-time PCR, with normalization relative to GAPDH. The cycle threshold (Ct) values were measured, and results are expressed as the Ct ratios of CD44 or COX-2 to GAPDH. The COX-2 Ct ratio was much lower in cases of lymphovascular invasion by the tumor than for no invasion (P = 0.004). During follow-up for a median of 40 months, there was no significant difference in the median CD44 Ct ratio between survivors and non-survivors (P = 0.362), whereas the COX-2 Ct ratio was significantly associated with survival at the time of data analysis (P = 0.042). The survival of colorectal cancer patients with a high COX-2 Ct ratio was significantly longer than that of patients with a low COX-2 Ct ratio (P = 0.048). This study suggests that COX-2 expression has a more significant impact than CD44 expression on the survival of CRC patients. Further studies are needed to resolve these issues with a large sample size.     

Peroxiredoxin 1 promoted tumor metastasis and angiogenesis in colorectal cancer

Peroxiredoxin1 (Prdx1) is a member of the PrdxS family, and it regulates cellular signaling and differentiation. The role of Prdx1in colorectal cancer (CRC) remains unclear. In this study, we investigated the relevance of Prdx1 in the metastasis and angiogenesis of CRC. The expression of Prdx1 in 60 cases human CRC tissues was detected through immunohistochemistry. The tumors that highly expressed Prdx1 (42/60) exhibited higher tumor grade and lymph node metastasis than those with low expression of Prdx1 (18/60) (p?<?0.05). Kaplan–Meier survival analysis showed that the survival time of thePrdx1-positive group was shorter than that of thePrdx1-negative group (p?=?0.046).Moreover, a statistically significant correlation was observed between the Prdx1 expression and microvessel density (p?=?0.004). Transwell migration assay revealed that Prdx1 was down-regulated in the CRC cell line HCT116, thereby suppressing the invasion and migration capacities of tumor cells, whereas Prdx1was up-regulated in HT29 cells, thereby increasing the invasion and migration capacities of tumor cells. The tube formation capacity of human umbilical vein endothelial cells cultured in 3D medium was increased after conditioned medium from overexpressed Prdx1cancer cells was added relative to that when down-regulated Prdx1 cell medium was added (p?<?0.05). In addition, up-regulated Prdx1 increased the protein expression of MMP2, MMP9, and VEGFA. These data suggested that Prdx1 expression predicted poor prognosis by regulating the tumor metastasis and angiogenesis of CRC. Therefore, Prdx1 may serve as a potential therapeutic target.     

ERK5在结直肠癌组织中的表达及其与结直肠癌转移的关系

目的: 检测和分析细胞外信号调节激酶5(ERK5)蛋白在原发性结直肠癌组织和癌旁正常黏膜组织中的表达情况及其与结直肠癌临床病理参数的关系,探讨ERK5的临床意义。方法: 使用组织芯片和免疫组化的方法检测338例原发性结直肠癌癌组织和80例癌旁正常组织中ERK5的表达,分析ERK5表达与结直肠癌的临床病理参数及预后的相关性。结果:80例癌旁正常黏膜中,ERK5低表达59例(73.8%)、高表达为21例(26.2%);338例结直肠癌组织中ERK5低表达204例(60.4%)、高表达134例(39.6%);二者中的ERK5蛋白表达有显著差异(P<0.05)。在338例结直肠癌组织中,27例发生远处转移患者的癌组织中ERK5高表达率为63.0%,而311例未发生远处转移患者的癌组织中ERK5高表达率为37.6%,二者之间有显著差异(P<0.05)。但癌组织中ERK5的表达水平与患者性别、年龄、体重指数、肿瘤位置、T分期、N分期、TNM分期和分化程度无明显相关性(P>0.05)。Kaplan-Meier生存分析显示在结直肠癌组织中ERK5高表达的病人总体5年生存率与低表达组无明显差异(P>0.05)。结论:ERK5蛋白在癌组织中的表达较癌旁正常组织高;发生远处转移的结直肠癌组织中ERK5蛋白的表达水平明显高于未发生远处转移的癌组织中ERK5水平。ERK5可能是一个促进肿瘤远处转移的因子。     

Expression of c-Met in laryngeal carcinoma

 Expression of c-Met, a gene for the hepatocyte growth factor/scatter factor (HGF/SF) receptor, is known to be associated with tumour development in several human carcinomas. The expression of c-Met was examined using immunohistochemistry in 82 cases of primary laryngeal carcinoma to evaluate the tissue distribution of c-Met and the clinicopathological significance of c-Met expression. In normal larynx, c-Met expression was observed only in some minor salivary glands. Positive reaction for c-Met in neoplastic epithelium was noted in 45 out of 82 (54.9%) cases. In 44 cases, structures adjacent to the carcinoma (noncancerous squamous epithelium, some stromal fibroblastic cells, and endothelial cells) showed positive reaction for c-Met. c-Met expression in cancerous epithelium was significantly correlated with lymph node status (P<0.04) and proliferating activity expressed by the Ki-67 labelling index (P<0.02). There was no correlation between c-Met expression and age, sex, histological type, T category, distant metastasis or clinical stage. The data suggest that overexpression of c-Met in laryngeal carcinomas represents a growth advantage for cancer cells, which may be conferred by the mitogenic effect of HGF/SF. Simultaneous c-Met expression in noncancerous components of the larynx may represent a paracrine modification of c-Met. Received: 7 October 1997 / Accepted: 4 November 1997     

The expression and clinical significance of ribophorin II (RPN2) in human breast cancer

Ribophorin II (RPN2), part of the N‐oligosaccharyltransferase complex, is highly expressed in breast cancer stem cells and is associated with tumor metastasis through interaction with mutant p53. The clinicopathological implication of RPN2 expression is undetermined. We examined immunohistochemically the expression levels of RPN2 and p53 in primary breast cancer tissues surgically resected from 218 patients. The correlations of RPN2 expression with the intrinsic subtype defined by hormone receptors (HRs) and HER2, clinicopathological parameters, p53 expression, and patients’ clinical outcomes were examined. RPN2 was positive in 139 (64%), and the incidence of RPN2 expression was higher in the triple‐negative breast cancer (TNBC) (HR‐/HER2‐) (65%) and HER2‐enriched (HR‐/HER2+) subtype (95%) than in the luminal A‐like (HR+/HER2‐) subtype (58%) (P = 0.0009). RPN2 expression was also correlated with p53 nuclear accumulation (P = 0.04). The RPN2‐positive/p53‐positive patient group showed significantly poorer prognosis than the RPN2‐negative group for disease‐free survival (P = 0.05) and for overall survival (P = 0.02). By multivariate analyses, the combination of RPN2 and p53 was not an independent prognostic factor. RPN2 expression was correlated with clinically aggressive features of breast cancer. These data support the further clinical application of anti‐RPN2 therapy and the development of personalized medicine.     

结直肠癌术后循环肿瘤细胞检测的临床意义

目的 探讨结直肠癌(CRC)患者术后外周血循环肿瘤细胞(CTCs)阳性率及其计数与临床病理特征、肿瘤复发转移及预后之间的关系及其临床意义。方法 回顾性分析2016年1月—2017年7月上海交通大学医学院附属第九人民医院首诊并经病理确诊的99例CRC患者的临床资料,其中男66例、女33例,年龄37~79岁。检测患者术后4~8周且辅助治疗前CTCs计数和血清癌胚抗原(CEA)水平,随访患者生存情况,比较患者不同临床病理学特征患者间CTCs表达及计数值的统计学差异,包括性别、年龄、原发灶部位、肿瘤分化程度、原发灶浸润深度(T分期)、区域淋巴结侵袭程度(N分期)、远处转移(M分期)、TNM总分期;观察患者生存情况;采用COX比例风险回归模型进行无进展生存期、总生存期的单因素和多因素分析。结果 99例CRC患者CTCs阳性率为60.6%(60/99),CTCs计数值为0~24(4.909±5.518)。患者的CTCs阳性率和CTCs计数值在T1+T2+T3期、N0期、M0期、Ⅰ~Ⅱ期、CEA＜5 ng/mL和中高分化组患者低于T4期、N1+N2期、M1期、Ⅲ~Ⅳ期、CEA≥5 ng/mL、低分化组患者,差异均有统计学意义(P值均＜0.05)。患者随访3~20个月,死亡22例(22/99,22.2%),其中CTCs阳性21例;疾病进展32例(32/99,32.3%),其中CTCs阳性29例。单因素分析显示:CTCs表达、N分期、M分期、肿瘤分期、和高CEA水平是无进展生存期的影响因素(P值均＜0.05);CTCs表达、N分期、M分期、肿瘤原发灶位于直肠、高CEA水平是总生存期的影响因素(P值均＜0.05)。CTCs和远处转移是CRC患者无进展生存期和总生存期的独立预后因素(HR= 5.418、2.254,95%可信区间:1.595~8.403、1.227~7.986,P值均＜0.05)。结论 在术后CRC患者中,CTCs表达与预后不良有关,对肿瘤复发转移评估具有一定的价值和临床意义。     

Correlation between Mcl-1 and pAKT protein expression in colorectal cancer

Mcl-1 inhibits apoptosis in well-differentiated cells by sequestering BAD, BID, and BAX and other apoptotic molecules. pAKT blocks apoptotsis by facilitating the interaction of BAD with BCL-XL. Expression of pAKT and Mcl-1 have been described in colon cancer, however, the relationship between pAKT and Mcl-1 has not. Mcl-1 and pAKT immunohistochemistry was performed using colorectal cancer tissue microarray (TMA). The Holm step-down method was used to adjust for multiple testing. Mcl-1 and pAKT scores, stage, and grade were compared using Spearman's correlation coefficient. Metastasis and no metastasis groups were compared using the Wilcoxon rank sum test. Mcl-1 and pAKT scores were compared for normal colorectal mucosa (NR), adenoma (AD), and colorectal cancer (CRC) cohorts. The mean (SD) pAKT expression in NR (14) was 2.0 (1.4), in AD (8) was 3.0 (1.7), and in CRC (101) was 5.6 (2.4). These differences were statistically significant. For Mcl-1 the mean (SD) expression was 4.1 (1.7) in NR, 3.2 (1.2) in AD, and 3.3 (2.6) in CRC. Mcl-1 and pAKT scores were directly correlated during various stages of colon car-cinogenesis (p = 0.04). Mcl-1 showed direct correlation with tumor grade (p = 0.001) and tumor stage (p = 0.02) and with presence of metastasis (p = 0.008). We report the correlation of Mcl-1 protein expression with higher grade and stage in colorectal cancer. Mcl-1 correlated also with pAKT expression. We also report the up regulation of pAKT during the transition from NR to CRC.     

Expression of Robo protein in bladder cancer tissues and its effect on the growth of cancer cells by blocking Robo protein

This study aimed to detect the expression of Slit signaling protein ligand Robo protein in human bladder cancer and para-carcinoma tissue, and observe the tumor cell survival and growth by inoculating the bladder cancer cells with the blocked signaling protein into the subcutaneous tissue of nude mice. The expression of Robo protein was detected in T24 cells in human bladder uroepithelium carcinoma and cultivated human bladder uroepithelium carcinoma confirmed by pathological diagnosis. The cultivated T24 cells were coated by the protein antibody and human bladder uroepithelium carcinoma T24 tumor-bearing mice model was established. The tumor cell survival and growth were observed in the antibody coating group and non-coating group. The tumor body size was measured. The immunohistochemical detection showed that Robo protein isoforms Robo1 and Robo 4 were expressed in T24 cells of cancer tissues, paracarcinoma tissues and cultured human uroepithelium carcinoma. The expression of Robo1 was significantly higher than that of Robo4 (P<0.05). The cancer cells could be detected in nodular tumor of mice in each group. The volume of the tumor-bearing mice in the nodular tumor of the non-coating group was larger than that of anti-Robol antibody coating group and the difference was statistically significant (P<0.01). There was no significant difference in tumor volume between anti-Robo4 antibody coating group and non-coating group (P>0.05); The difference was statistically significant compared with the anti-Robo1 antibody coating group (P<0.01). In conclusion, Robo protein isoforms Robo1 and Robo4 were expressed in human bladder cancer T24 cells. To block Robo4 signal protein had little effect on the survival and growth of the transplantation tumor and to block Robo1 signal protein would seriously affect the survival and growth of the transplantation tumor, suggesting that Robo1 might play an important role in the growth and metastasis of bladder cancer, and might become a new target for the treatment of human bladder cancer and drug research.     

Low expression of MicroRNA‐126 is associated with poor prognosis in colorectal cancer

MicroRNA‐126 (miR‐126) has been reported to be a tumor suppressor that targets CXCR4 in colorectal cancer (CRC) cells. This study investigated whether miR‐126 has any prognostic impact in patients with CRC. MiR‐126 and CXCR4 mRNA expression in 92 pairs of CRC and adjacent nontumorous tissues was examined using quantitative real‐time PCR, and CXCR4 protein expression was assessed by immunohistochemistry (IHC) and Western blotting. The correlation between miR‐126 and CXCR4 protein expression and clinicopathological features and overall survival rate was determined. MiR‐126 was downregulated in CRC tissues that expressed high levels of CXCR4 mRNA. IHC and Western blotting detected high expression of CXCR4 protein in CRC tissues. An inverse correlation was observed between miR‐126 and CXCR4 protein expression in CRC tissues. Moreover, low miR‐126 and high CXCR4 protein expression was associated with distant metastasis, clinical TNM stage, and poor survival. Multivariate analysis indicated that miR‐126 was an independent prognostic factor for overall survival, suggesting its clinical significance as a prognostic predictor in CRC patients. © 2014 Wiley Periodicals, Inc.     

Lemur tyrosine kinase-3 is a significant prognostic marker for patients with colorectal cancer

Lemur tyrosine kinase-3 (LMTK3) belongs to the family of serine-threonine-tyrosine kinases and the aberrant expression of LMTK3 was observed in several human malignancies. However, the association of LMTK3 with clinical outcomes in colorectal cancer patients is unclear. Thus, this present study was to evaluate the association of LMTK3 expression level with clinicopathologic factors and prognosis of patients with colorectal cancer (CRC). The expression level of LMTK3 in 69 archival paraffin-embedded colorectal tumor tissue specimens was examined by immunohistochemistry (IHC). As a result, we found that the LMTK3 expression level was significantly elevated in CRC tissues as compared with Crohn’s disease or colorectal polyp tissues (P<0.0001, P<0.0001, respectively). Positive LMTK3 signals in the colorectal cancer cells were observed in about 89.9% (62 of 69) CRC tissue specimens. Additionally, LMTK3 expression was significantly correlated with lymph node metastasis and tumor-node-metastasis (TNM) classification (P=0.003, and P=0.008, respectively), but not with sex, age, tumor location, histological differentiation, tumor size, or depth of tumor invasion (all P>0.05). Kaplan-Meier survival curves showed that the overall survival rate was significantly higher in the patients with low expression of LMTK3 when compared with those patients with high LMTK3 (P=0.010). Moreover, multivariate analysis revealed that LMTK3 expression was an independent prognostic factor for CRC patients (P=0.047). These results suggest that LMTK3 protein could serve as a prognostic marker for CRC patients.