Proteomic Characterization of Colorectal Cancer Cells versus Normal-Derived Colon Mucosa Cells: Approaching Identification of Novel Diagnostic Protein Biomarkers in Colorectal Cancer

Abstract

In the western world, colorectal cancer (CRC) is the third most common cause of cancer-related deaths. Survival is closely related to the stage of cancer at diagnosis striking the clinical need for biomarkers capable of early detection. To search for possible biological parameters for early diagnosis of CRC we evaluated protein expression for three CREC (acronym: Cab45, reticulocalbin, ERC-55, calumenin) proteins: reticulocalbin, calumenin, and ERC-55 in a cellular model consisting of a normal derived colon mucosa cell line, NCM460, and a primary adenocarcinoma cell line of the colon, SW480. Furthermore, this cellular model was analyzed by a top-down proteomic approach, 2-dimensional polyacrylamide gel electrophoresis (2D-PAGE) and liquid chromatography–tandem mass spectrometry (LC–MS/MS) for novel putative diagnostic markers by identification of differentially expressed proteins between the two cell lines. A different colorectal carcinoma cell line, HCT 116, was used in a bottom-up proteomic approach with label-free quantification (LFQ) LC–MS/MS. The two cellular models gave sets of putative diagnostic CRC biomarkers. Various of these novel putative markers were verified with increased expression in CRC patient neoplastic tissue compared to the expression in a non-involved part of the colon, including reticulocalbin, calumenin, S100A6 and protein SET. Characterization of these novel identified biological features for CRC patients may have diagnostic potential and therapeutic relevance in this malignancy characterized by a still unmet clinical need.

Shotgun-based proteomics of extracellular vesicles in Alzheimer’s disease reveals biomarkers involved in immunological and coagulation pathways

“…Liquid chromatography–tandem mass spectrometry was performed as previously described 26 using the universal method on an Orbitrap Fusion Tribrid mass spectrometry platform from Thermo Scientific (Waltham, MA, USA). Internal mass calibration was performed by activating the EASY-IC using fluoranthene.…”

“…Replicates were injected with an intermission of several hours to days. The acquisition was performed in the first 60 min of the gradient as previously described 26 with full Orbitrap scans (375 – 1500 m/z ), a resolution of 120,000, and an automatic gain control (AGC) target of 4 × 10 5 with a maximum injection time of 50 ms. Each cycle time lasted 3 s. Precursor ions with the highest intensity were selected, with an intensity threshold set at 5 × 10 3 , and charge states 2 – 7 included. The linear ion trap was used for MS 2 scans at a rapid scan rate with a collision-induced dissociation energy at 35 % and an AGC target of 2 × 10 3 with a maximum injection time of 300 ms.…”

Artificial Intelligence in Colorectal Cancer Diagnosis Using Clinical Data: Non-Invasive Approach

“…Numerous papers in an emerging research field inspect the efficiency of blood biomarkers in the early detection of cancer [ 10 ]. A work [ 11 ] on this matter points out that there is only one biomarker that is an internationally approved clinical practice for colorectal cancer detection: carcinoembryonic antigen (CAE). In addition, by comparing healthy tissues with carcinogen ones and performing protein expression analysis on them, the study offers a novel method to search for other possible biomarkers in order to detect colorectal cancer.…”

Proteomics of Colorectal Cancer: Tumors, Organoids, and Cell Cultures—A Minireview

“… Hu et al (2018) used HCT-116 and RKO, selectively cultured to be highly invasive, to identify the Cdc42-Cdc42BPA signaling pathway as a marker for colon cancer invasion. Ludvigsen et al (2020) compared a normal derived colon mucosa cell line (NCM460) and a colon cancer cell line (HCT116) to identify protein biomarkers with increased expression in colorectal cancer, including retinol-binding protein 1. 2D cell culture has also been used to compare non-metastatic (SW480) and metastatic colon cancer cell lines (SW620) ( Zhao et al, 2007 ; Schunter et al, 2017 ; Torres et al, 2018 ).…”