Quantitation of putative colorectal cancer biomarker candidates in serum extracellular vesicles by targeted proteomics

Abstract Background: Colorectal cancer (CRC) is a common malignant tumor with unsatisfactory overall prognosis. CircRNAs could be promising prognostic biomarkers in cancers, and play important role in the process of tumorigenesis and progression. Here, we explored the role of hsa_circ_0004831 in blood extracellular vesicles and its prognostic value in CRC. Methods: The circRNA and mRNA expression level matrix in extracellular vesicles of CRC and normal samples were obtained from the exoRBase database. The corresponding miRNA expression level matrix in extracellular vesicles was downloaded from the BBCancer database. Differentially expressed circRNAs, miRNAs and mRNAs were identified using the limma package of R software at the cut-off criteria of fold change (FC) > 2 and adj. p < 0.05. RT-qPCR assay was conducted to measure hsa_circ_0004831 expression level in CRC blood samples. A circRNA-miRNA-mRNA regulatory network of hsa_circ_0004831 was constructed based on competitive endogenous RNA mechanism and differentially expressed genes. The mRNAs co-expressed with hsa_circ_0004831 were screened at the cut-off criteria of pearson |r| > 0.3 and p < 0.05. Gene set enrichment analysis (GSEA) based on co-expressed mRNAs was used to explore the potential molecular function of hsa_circ_0004831. Results: Differentially expressed circRNAs, miRNAs and mRNAs were identified and hsa_circ_0004831 had a FC value of 3.92 in CRC blood extracellular vesicles. The RT-qPCR assay showed that the hsa_circ_0004831 was up-regulated in CRC blood samples. The overall survival analysis found that high expression of hsa_circ_0004831 was linked with poorer prognosis. Finally, a circRNA-miRNA-mRNA regulatory network of hsa_circ_0004831 was constructed based on down-regulated miR-4326 and 12 up-regulated mRNAs. GSEA indicated that mRNAs co-expressed with hsa_circ_0004831 were involved in EMT, WNT and p53 signaling pathways.Conclusions: The study confirmed the up-regulation of hsa_circ_0004831 in CRC, and it may act as a vital prognostic biomarker. The circRNA-miRNA-mRNA regulatory network of hsa_circ_0004831 could be used to uncover the tumorigenesis and progression of CRC.

Download Full-text

EGFL6 Promotes Colorectal Cancer Cell Growth and Mobility and the Anti-cancer Property of Anti-EGFL6 Antibody

10.21203/rs.3.rs-92169/v1 ◽

2020 ◽

Author(s):

Ting-Yi Sung ◽

Han-Li Huang ◽

Chun-Chun Cheng ◽

Fu-Ling Chang ◽

Po-Li Wei ◽

...

Keyword(s):

Colorectal Cancer ◽

Cancer Cell ◽

Animal Experiments ◽

Patient Characteristics ◽

Cancer Biomarker ◽

Blue Staining ◽

Migration And Invasion ◽

Metal Stents ◽

Αvβ3 Integrin ◽

Phage Display Technology

Abstract BackgroundA reliable cancer biomarker will be critical for advanced colorectal cancer (CRC) therapeutic approaches since current treatments are limited to certain patient characteristics, such as age, sex, comorbidities and patients received self-expandable metal stents implantations. The association of epidermal growth factor-like domain 6 (EGFL6) with cancer development has been reported. Here, we focused on the role of EGFL6 in CRC progression and its clinical relevance. MethodsAn anti-EGFL6 antibody was generated by phage display technology to investigate the potential therapeutic efficacy in CRC. Methylene blue staining was applied to investigate the EGFL6 expression in CRC patients and animal tissue. Protein and DNA level of EGFL6 expression as well as related pathway investigation were determined by western blot and quantitative real time PCR. Silence EGFL6 by siRNA transfection, transwell migration and invasion assay were performed to verify EGFL6 function. Student t test, Kruskal-Wallis test and multiple comparisons were used to statistical analysis results. ResultsSignificant EGFL6 expression was found in colon tissues from patients and spontaneous tumorigenesis mouse but not in normal tissue. Furthermore, we found EGFL6 could enhance cancer cell migration, invasion and proliferation in CRC via up-regulating ERK/ AKT pathway, as well as reducing ADAMTS1 and Snail expression. We also found EGFL6 regulates cell abilities through EGFR/αvβ3 integrin receptors. By conducting animal experiments, our anti-EGFL6 antibody, EGFL6-E5-IgG, showed tumor inhibition and anti-metastasis ability. Furthermore, no impact on angiogenesis and wound healing by using EGFL6-E5-IgG were observed. ConclusionsWe demonstrated that EGFL6 plays a role in CRC tumorigenesis and tumor progression, indicating that EGFL6 is a potential cancer biomarker and therapeutic target worth further investigation.

Download Full-text