The role of RCAS1 as a biomarker in diagnosing CRC and monitoring tumor recurrence and metastasis

Receptor-binding cancer antigen expressed on SiSo cells (RCAS1) plays an important role in tumor progression by helping tumor cell to escape from host immunological surveillance or modifying the characteristics of connective tissue around. RCAS1 may appropriately reflect the development and prognosis of tumor. In the study, we sought to identify the clinical significance of RCAS1 in colorectal cancer (CRC) diagnosis and tumor recurrence monitoring. Immunohistochemistry (IHC) with tissue array slides was preformed to analyze RCAS1 protein expression in CRC, colorectal polyps, and normal colon tissues. RCAS1 levels in colorectal cancer were significantly higher than those in colorectal polyps and normal colon tissues (P?<?0.001). Silencing RCAS1 gene in human colonic adenocarcinoma cells decreased cell proliferation and enhanced apoptosis through the p53 signaling pathway. Further analysis by an enzyme-linked immunosorbent assay (ELISA) showed that serum RCAS1 levels in CRC are significantly higher than in healthy controls and polyps (P?<?0.05), in which the highest serum RCAS1 level is reported in the recurrence group. The serum RCAS1 levels have a significant correlation with clinical stage and pathologic grading. Furthermore, the positive rate of serum RCAS1 in CRC was 82.1 %, which was higher than carcinoembryonic antigen (CEA). Especially in CEA-negative cases, the sensitivity of RCAS1 was 88.2 %. Finally, CRC patients who were followed up showed a serum RCAS1 level which significantly decreased after surgery (P?<?0.001) and obviously increased in the recurrence group. Taken together, our data demonstrated that RCAS1 is not only a supplementary serological biomarker for CRC diagnosis but also useful for monitoring tumor recurrence. RCAS1 might be a supplementary serological marker for CRC.     

Epigenetic Silencing of HOPX Promotes Cancer Progression in Colorectal Cancer

Homeodomain-only protein X (HOPX)-β promoter methylation was recently shown to be frequent in human cancers and was suggested as tumor suppressor gene in esophageal and gastric cancer. The aim of this study was to investigate the mechanistic roles of HOPX-β promoter methylation and its clinical relevance in colorectal cancer (CRC). HOPX-β promoter methylation was assessed in human CRC cell lines and 294 CRC tissues. HOPX mRNA and protein levels were measured in relation to HOPX-β promoter methylation. The effects of forced HOPX expression on tumorigenesis were studied using in vitro and in vivo assays. The association between HOPX-β promoter methylation and clinical relevance of CRC patients was determined. HOPX-β promoter methylation is cancer-specific and frequently found in CRC cell lines and tissues, resulting in the down-regulation of HOPX mRNA and protein levels. In CRC cell lines, forced expression of HOPX suppressed proliferation, invasion, and anchorage-independent growth. DNA microarray analyses suggested critical downstream genes that are associated with cancer cell proliferation, invasion or angiogenesis. In a mouse xenograft model, HOPX inhibited tumorigenesis and angiogenesis. Finally, HOPX-β promoter methylation was associated with worse prognosis of stage III CRC patients (hazard ratio= 1.40, P = .035) and also with poor differentiation (P = .014). In conclusion, HOPX-β promoter methylation is a frequent and cancer-specific event in CRC progression. This epigenetic alteration may have clinical ramifications in the diagnosis and treatment of CRC patients.     

Sulfonamide derivative targeting carbonic anhydrase IX as a nuclear imaging probe for colorectal cancer detection in vivo

Hypoxic microenvironment is a common situation in solid tumors. Carbonic anhydrase IX (CA9) is one of the reliable cellular biomarkers of hypoxia. The role of CA9 in colorectal cancer (CRC) remains to be clarified. CA9 inhibitor such as sulfonamides is known to block CA9 activation and reduce tumor growth consequently. Here, we aimed to investigate the CA9 expression in serum and tumor from different stages of CRC patients and utilize sulfonamide derivative with indium-111 labeling as a probe for CRC nuclear imaging detection in vivo. The serum CA9 was correlated with the tumor CA9 levels in different stages of CRC patients. Hypoxia increased cell viability and CA9 expression in colorectal cancer HCT-15 cells. Sulfonamide derivative 5-(2-aminoethyl)thiophene-2-sulfonamide (ATS) could bind with CA9 in vitro under hypoxia. Moreover, tumor tissues in HCT-15-induced xenograft mice possessed higher hypoxic fluorescence signal as compared with other organs. We also found that the radioisotope signal of indium-111 labeled ATS, which was utilized for CRC detection in HCT-15-induced xenograft mice, was markedly enhanced in tumors as compared with non-ATS control. Taken together, these findings suggest that CA9 is a potential hypoxic CRC biomarker and measurement of serum CA9 can be as a potential tool for diagnosing CA9 expressions in CRC clinical practice. The radioisotope-labeled sulfonamide derivative (ATS) may be useful to apply in CRC patients for nuclear medicine imaging.     

Ruxolitinib induces apoptosis of human colorectal cancer cells by downregulating the JAK1/2-STAT1-Mcl-1 axis

Under pathological conditions, the Janus kinase (JAK)/STAT signaling pathway can regulate the proliferation, differentiation and migration of tumor cells, including colorectal cancer (CRC). CRC is the third major types of cancer among males and the second among females worldwide. In China, CRC is the fifth common cancer among both males and females. Western blotting, flow cytometry, RNA interference, immunoprecipitation, xenografts models, and immunohistochemical staining were carried out to evaluate the possible mechanisms of acton of ruxolitinib. The present data suggested that ruxolitinib can suppress CRC cell proliferation by inducing apoptosis. Firstly, JAK1/2-STAT1 was identified as the target of ruxolitinib. Then, ruxolitinib downregulated myeloid cell leukemia-1 (Mcl-1) mRNA level and decreased its protein level, which enabled Bak to trigger CRC apoptosis. Furthermore, ruxolitinib exerted potent activity against CRC xenograft growth in vivo. High expression of phosphorylated STAT1 (S727) was also confirmed in 44 pairs of human colon carcinoma and adjacent normal tissues. Taken together, the results showed that ruxolitinib decreased JAK1/2-STAT1-Mcl-1 protein level and effectively suppressed CRC cell proliferation in vitro and in vivo. Therefore, ruxolitinib could be a promising anticancer agent for CRC treatment.     

Prognostic and immunological roles of Fc fragment of IgG binding protein in colorectal cancer

Valuable diagnostic and prognostic biomarkers are urgently needed for colorectal cancer (CRC), which is one of the leading causes of mortality worldwide. Previous studies have reported altered expression of a mucin-like protein Fc fragment of IgG binding protein (FCGBP) in various types of cancer, but its potential diagnostic, prognostic and immunological roles in CRC remain to be determined. Therefore, the aim of current study was to investigate the potential roles of FCGBP in CRC. The present study investigated FCGBP mutations and changes in its expression levels using a combination of microarray and public dataset analyses, as well as immunohistochemistry. The results demonstrated a 10.5% mutation frequency in the FCGBP coding sequence in CRC tissues, and identified decreased FCGBP mRNA or protein expression levels in colorectal adenoma and CRC (compared with those in normal colorectal tissues from healthy control subjects), including pathologically advanced CRC (stage III+IV vs. I+II). Survival analysis using the GEPIA and Kaplan-Meier Plotter databases revealed that low FCGBP expression levels were associated with short overall, disease-free, relapse-free and event-free survival times in patients with CRC. Notably, analysis using the online Tumor IMmune Estimation Resource database revealed a positive correlation between FCGBP expression levels and the extent of infiltrating immune cells, such as B cells and dendritic cells. Consistently, the expression levels of most markers (51/57) for various types of immune cells were significantly correlated with FCGBP expression levels in CRC tissues. These findings suggested that FCGBP may serve as a diagnostic and prognostic biomarker, and that FCGBP may be associated with immune infiltration in CRC.     

LINC00178 promotes cell growth,migration, and invasion in colorectal cancer in vitro and in vivo

BackgroundThis study aimed to determine the role of LINC00178 in colorectal cancer (CRC) cell invasion and migration by examining its expression in CRC cells and tissues.MethodsCancer tissues and corresponding adjacent tissue specimens were collected from 45 patients who experienced radical CRC resection in the hospital from March to September 2021. The expression of LINC00178 was measured in both CRC cells and tissues and normal human colorectal mucosal cells using quantitative fluorescence polymerase chain reaction (QF-PCR). Cell Counting Kit-8 (CCK-8), clonogenic, and transwell assays were used to assess the impact of LINC00178 overexpression or knockdown on the CRC cells invasion and proliferation. In addition, the expression levels of vimentin, E-cadherin, and N-cadherin in CRC cells were determined after either LINC00178 knockdown or overexpression was performed using western blotting.ResultsThe experiments revealed that LINC00178 was over expressed in CRC cells and tissues. Over-expression of LINC00178 could significantly promote the propagation, clone formation, invasion, and transportation of CRC, whereas knockdown of LINC00178 had the opposite function. When LINC00178 was expressed at high levels, it suppressed the vimentin and N-cadherin expression and prevented the upregulation of E-cadherin. In vivo (nude mouse) studies showed that the over expression of LINC00178 could significantly promote the propagation in CRC cells.ConclusionsLINC00178 is overexpressed in CRC cells and tissues. In vivo and in vitro experiments showed that LINC00178 can significantly promote the propagation of CRC cells, so it may develop a potential biological site for targeted therapy of CRC patients.     

Applications of novel monoclonal antibodies specific for synuclein-gamma in evaluating its levels in sera and cancer tissues from colorectal cancer patients

Overexpressions of synuclein-gamma (SNCG) in different cancers display stage-specific patterns. At present, appropriate anti-SNCG monoclonal antibodies (mAbs) with high specificity and affinity are unavailable for different immunoassays in clinical applications. In this study, we generated 10 mAbs against endogenous SNCG and evaluated SNCG levels in several colorectal cancer cell lines, serum samples and tumor tissues from colorectal cancer (CRC) patients. Elevated SNCG levels in cancer cell lines evaluated by a novel sandwich ELISA were consistent with data obtained from Western blot. Secreted SNCG protein levels in sera from CRC patients could be detected by the sandwich ELISA and were further confirmed by Western blot analysis following SNCG enrichment. Immunohistochemical results showed that SNCG was highly expressed in tumor cells of CRC patients, but was undetectable in the adjacent normal epithelium. Taken together, these novel anti-SNCG mAbs specifically recognized endogenous SNCG and were suitable for measuring SNCG levels in cell lysates, human serum samples, and tumor tissues. Elevated serum SNCG and overexpressed SNCG in tumor tissue from CRC patients suggest SNCG is a potential biomarker for CRC.     

MicroRNA-145 inhibits tumour growth and metastasis in colorectal cancer by targeting fascin-1

Background:

Recent studies have reported miR-145 dysregulated in colorectal cancer (CRC). In this study, miR-145 profiles were compared between CRC and corresponding non-tumour tissues.

Methods:

The expression levels of miR-145 were analysed in CRC cell lines and tumour tissues by real-time PCR. A luciferase reporter assay confirmed direct targets. The functional effects of miR-145 were examined in transfected CRC cells in vitro and in vivo using established assays.

Results:

Downregulation of miR-145 was detected in most primary CRC tumours, and was significantly correlated with a more aggressive phenotype of CRC in patients. In CRC cell lines, ectopic overexpression of miR-145 inhibited cell proliferation, motility and invasion in vitro. Stable overexpression of miR-145 suppressed tumour growth and pulmonary metastasis in vivo. Further studies indicated that miR-145 may directly interact with the 3′-untranslated region (3′-UTR) of Fascin-1 messenger RNA (mRNA), downregulating its mRNA and protein expression levels. In clinical specimens, Fascin-1 expression was negatively correlated with miR-145 expression.

Conclusions:

MiR-145 has a critical role in the inhibition of invasive and metastatic capacities of CRC, probably through directly targeting Fascin-1. This miRNA may be involved in the development and progression of CRC.     

miR-191 promotes tumorigenesis of human colorectal cancer through targeting C/EBPβ

MicroRNA-191 (miR-191), a small non-coding RNA, is involved in disease development and cancer diagnosis and prognosis. However, how miR-191 functions in colorectal cancer remains largely unclear. In this study, we show that miR-191 is highly expressed in colon tumor tissues, and that inhibition of miR-191 leads to decreased cell growth, proliferation and tumorigenicity in a xenograft model. Overexpression of miR-191 in colorectal cancer cell lines alters cell cycle progression and cell resistance to 5-Fu induced cell apoptosis. Mechanistic studies demonstrated that miR-191 directly binds to the 3′UTR of the C/EBPβ mRNA and mediates a decrease in the mRNA and protein expression of C/EBPβ. We further showed that C/EBPβ induces growth arrest in a colorectal cancer cell line and that its expression is negatively correlated with the miR-191 level in patient samples. Our findings suggest that miR-191 may be a potential gene therapy target for the treatment of colorectal cancer.     

MicroRNA-143 targets DNA methyltransferases 3A in colorectal cancer

Background:

MicroRNAs (miRNAs) are 19-25-nucleotides regulatory non-protein-coding RNA molecules that regulate the expressions of a wide variety of genes, including some involved in cancer development. In this study, we investigated the possible role of miR-143 in colorectal cancer (CRC).

Methods:

Expression levels of human mature miRNAs were examined using real-time PCR-based expression arrays on paired colorectal carcinomas and adjacent non-cancerous colonic tissues. The downregulation of miR-143 was further evaluated in colon cancer cell lines and in paired CRC and adjacent non-cancerous colonic tissues by qRT–PCR. Potential targets of miR-143 were defined. The functional effect of miR-143 and its targets was investigated in human colon cancer cell lines to confirm miRNA–target association.

Results:

Both real-time PCR-based expression arrays and qRT–PCR showed that miR-143 was frequently downregulated in 87.5% (35 of 40) of colorectal carcinoma tissues compared with their adjacent non-cancerous colonic tissues. Using in silico predictions, DNA methyltranferase 3A (DNMT3A) was defined as a potential target of miR-143. Restoration of the miR-143 expression in colon cell lines decreased tumour cell growth and soft-agar colony formation, and downregulated the DNMT3A expression in both mRNA and protein levels. DNMT3A was shown to be a direct target of miR-143 by luciferase reporter assay. Furthermore, the miR-143 expression was observed to be inversely correlated with DNMT3A mRNA and protein expression in CRC tissues.

Conclusion:

Our findings suggest that miR-143 regulates DNMT3A in CRC. These findings elucidated a tumour-suppressive role of miR-143 in the epigenetic aberration of CRC, providing a potential development of miRNA-based targeted approaches for CRC therapy.     

Depletion of insulin-like growth factor 1 receptor increases radiosensitivity in colorectal cancer

BackgroundAlthough radiation therapy for advanced colorectal cancer (CRC) is very effective in some patients, treatment resistance limits its efficacy. Insulin-like growth factor 1 receptor (IGF1R) can affect tumor responsiveness and sensitivity to radiation in several cancer types. Herein, we studied the underlying function of IGF1R in the resistance of advanced CRC to radiation therapy and the possible use of drugs targeting IGF1R to overcome this resistance in patients with CRC.MethodsDifferences in the expression levels of the IGF1R were assessed in CRC samples from patients who were radiosensitive or radioresistant. Two radio-resistant colorectal cancer cell lines, SW480 and HT29, were selected for in vitro studies, and the involvement of the IGF1R in their radiation resistance was elucidated by suppressing its expression through a targeted siRNA and through the use of a specific IGF1R inhibitor, BMS-754807. We assessed radiosensitivity in these human CRC cells lines by examining their proliferation and colony formation, as well as cell cycle analysis. Activation of the Akt pathway was assessed using western blotting.ResultsCompared with tissues from radiosensitive patients, higher IGF1R expression levels were found in patients with radiation-resistant colorectal cancer, while BMS-754807 had improved radiosensitivity and reversed radiation tolerance in both colorectal cancer cell lines. Pre-treatment with BMS-754807 prior to irradiation inhibited Akt phosphorylation, induced cell cycle arrest, and increased DNA damage. Therefore, the IGF1R contributes to radiation resistance of CRC cells in vitro.ConclusionsThis study supports the notion that the radiosensitivity of radiation-resistant colorectal cancer cells can be enhanced by directly targeting IGF1R expression or activity. Ultimately, the combination of radiotherapy with IGF1R targeted inhibitors could potentially increase its effectiveness in the treatment of advanced colorectal cancer.     

转凝蛋白在结直肠癌组织中的表达及其对SW480细胞恶性生物学行为的影响

目的:探讨转凝蛋白(transgelin,TAGLN)在结直肠癌(colorectal cancer,CRC)组织中的表达及其对SW480细胞增殖、迁移及侵袭的影响.方法:选取郑州大学附属肿瘤医院2015年5月至2016年8月收治的97例CRC患者的癌及配对的癌旁组织标本,以及人CRC细胞系SW620、SW480、HC...     

Cdc42EP3对结直肠癌细胞迁移和侵袭的影响

目的：探讨Cdc42EP3在结直肠癌（colorectal cancer,CRC）转移中的作用及相关作用机制。方法：回顾性收集2010年12月至2011年12月于南京医科大学附属淮安第一医院诊治的97例CRC患者的术后病理蜡块,同时收集相关临床病理资料。采用免疫组织化学法检测Cdc42EP3在癌组织与相应正常组织中的表达,随后通过统计学方法分析差异表达,并评估表达水平与临床病理学参数及生存预后之间的相关性。干扰CRC细胞Cdc42EP3的表达后,应用Transwell技术检测Cdc42EP3对CRC细胞的迁移及侵袭能力的影响,并使用Western blot技术检测EMT相关蛋白的表达。通过基因芯片技术及生物信息学分析预测Cdc42EP3的下游靶点STAT1,并通过回复实验验证。结果：Cdc42EP3在癌组织中的表达水平显著高于相应正常组织（P<0.001）,并与肿瘤的淋巴结转移（P=0.011）、TNM分期（P=0.008）及患者生存预后（P<0.001）之间呈显著相关性。干扰Cdc42EP3表达后,CRC细胞的迁移及侵袭能力均明显减弱（P<0.01）。预测出Cdc4...     

Circular RNA circ0104103 inhibits colorectal cancer progression through interactions with HuR and miR-373-5p

Emerging evidence has suggested that circular RNAs (circRNAs) have vital functions during the initiation and progression of various diseases. However, circRNA potential mechanisms in colorectal cancer (CRC) are largely unknown. Here, we sought to investigate the role and underlying regulatory mechanism of circ0104103 in CRC. circ0104103 was validated by quantitative RT-PCR (qRT-PCR) and Sanger sequencing. Gain- and loss-of-function assays in cell lines and mouse xenograft models were utilized to investigate the effects of circ0104103 in CRC. RNA pull-down assays, RNA immunoprecipitation assays, bioinformatics analyses, RNA FISH, and luciferase reporter assays were used to elucidate the potential mechanism of circ0104103 in CRC. We identified circ0104103, which is strongly downregulated in CRC tissues and cell lines. Functional studies revealed that circ0104103 inhibited CRC cell growth, migration, and invasion both in vitro and in vivo. Mechanistically, circ0104103 binds to HuR, a functional RNA-binding protein commonly expressed in CRC. HuR binds to the 3′UTR of LACTB mRNA to facilitate stabilization and increase its expression. Moreover, circ0104103 was verified as a competing endogenous RNA (ceRNA) via negative regulation of miR-373-5p to increase LACTB expression, resulting in inhibiting the occurrence and progression of CRC. Taken together, our study revealed that circ0104103 acts as a tumor suppressor and may be a novel biomarker and therapeutic target in CRC.     

基于GEPIA数据库分析SLC52A3在结直肠癌组织中的表达及临床病理意义

目的:通过研究SLC52A3在结直肠癌(CRC)组织中的表达及其与临床病理学参数的相关性,阐明其在CRC预后中的重要作用。方法:运用GEPIA大数据分析SLC52A3在正常结直肠组织和结直肠癌组织中的表达差异和其对胃癌预后的影响。收集2018年6月-2020年6月经手术切除的CRC患者石蜡标本84例。应用免疫组化方法检测84例CRC患者手术切除石蜡标本中SLC52A3表达情况和细胞定位。分析SLC52A3的表达与CRC临床病理指标的相关性。结果:数据库分析结果表明,与正常的结直肠组织相比,SLC52A3在结直肠癌组织中的表达显著提高,免疫组化结果显示,SLC52A3蛋白阳性信号定位于细胞膜。SLC52A3蛋白在84例CRC组织中低表达26例,高表达58例;在对应的84例CRC癌旁组织中9例呈高表达,75例为低表达,SLC52A3蛋白在CRC组织中的表达显著高于癌旁组织(P<0.01)。SLC52A3表达水平与CRC的肿瘤部位、肿瘤病理分化程度、浸润深度、TNM分期都明显相关,差异具有统计学意义(P<0.01)。在生存预后方面：SLC52A3高表达组的结直肠癌患者的生存率较高...     

miR-106b-5p靶向调控细胞周期蛋白D1抑制结直肠癌细胞增殖、迁移与侵袭

目的:探讨miR-106b-5p对结直肠癌(colorectal cancer,CRC)细胞增殖、迁移与侵袭的影响以及其作用机制.方法:实时荧光定量PCR检测miR-106b-5p在CRC组织与相应的癌旁组织、永生化的肠上皮细胞以及肠癌细胞中的表达量;CCK8实验检测DLD1细胞增殖能力;细胞划痕实验检测DLD1细胞的...     

Anti-Mesothelin Recombinant Immunotoxin Therapy for Colorectal Cancer

BackgroundMesothelin (MSLN) is a cell surface glycoprotein expressed at a high level on many malignancies, including pancreatic adenocarcinoma, serous ovarian cancer, and epithelioid mesothelioma. MSLN-targeted recombinant immunotoxins (RITs) consist of an anti-MSLN Fv fused to the catalytic domain of Pseudomonas exotoxin A. Recent data has also shown that MSLN is expressed at clinically relevant levels on the surface of colorectal cancer (CRC). In this study, CRC cell lines were tested for MSLN expression and susceptibility to MSLN-targeted RITs.Materials and MethodsCRC cell lines were tested for membranous MSLN expression via flow cytometry. Cell lines expressing MSLN were tested by WST-8 cell viability assay for sensitivity to various RITs and chemotherapeutic agents. CRC cell line SW-48 was tested in a mouse model for response to RIT as a single agent or in combination with actinomycin D and oxaliplatin.ResultsCRC cell lines were susceptible to anti-MSLN RITs at half maximal inhibitory concentration levels comparable with those previously described in pancreatic cancer cell lines. In a nude mouse model, MSLN-targeted RIT treatment of SW48 CRC tumors resulted in a significant decrease in tumor volume. Although combination therapy with standard of care chemotherapeutic oxaliplatin did not improve tumor regressions, combination therapy with actinomycin D resulted in > 90% tumor volume reduction with 50% complete regressions.ConclusionsThese data support the development of anti-MSLN RITs as well as other MSLN-targeted therapies for CRC.