Revelation of Proteomic Indicators for Colorectal Cancer in Initial Stages of Development

Abstract

Background: Colorectal cancer (CRC) at a current clinical level is still hardly diagnosed, especially with regard to nascent tumors, which are typically asymptotic. Searching for reliable biomarkers of early diagnosis is an extremely essential task. Identification of specific post-translational modifications (PTM) may also significantly improve net benefits and tailor the process of CRC recognition. We examined depleted plasma samples obtained from 41 healthy volunteers and 28 patients with CRC at different stages to conduct comparative proteome-scaled analysis. The main goal of the study was to establish a constellation of protein markers in combination with their PTMs and semi-quantitative ratios that may support and realize the distinction of CRC until the disease has a poor clinical manifestation. Results: Proteomic analysis revealed 119 and 166 proteins for patients in stages I–II and III–IV, correspondingly. Plenty of proteins (44 proteins) reflected conditions of the immune response, lipid metabolism, and response to stress, but only a small portion of them were significant (p < 0.01) for distinguishing stages I–II of CRC. Among them, some cytokines (Clusterin (CLU), C4b-binding protein (C4BP), and CD59 glycoprotein (CD59), etc.) were the most prominent and the lectin pathway was specifically enhanced in patients with CRC. Significant alterations in Inter-alpha-trypsin inhibitor heavy chains (ITIH1, ITIH2, ITIH3, and ITIH4) levels were also observed due to their implication in tumor growth and the malignancy process. Other markers (Alpha-1-acid glycoprotein 2 (ORM2), Alpha-1B-glycoprotein (A1BG), Haptoglobin (HP), and Leucine-rich alpha-2-glycoprotein (LRG1), etc.) were found to create an ambiguous core involved in cancer development but also to exactly promote tumor progression in the early stages. Additionally, we identified post-translational modifications, which according to the literature are associated with the development of colorectal cancer, including kininogen 1 protein (T327-p), alpha-2-HS-glycoprotein (S138-p) and newly identified PTMs, i.e., vitamin D-binding protein (K75-ac and K370-ac) and plasma protease C1 inhibitor (Y294-p), which may also contribute and negatively impact on CRC progression. Conclusions: The contribution of cytokines and proteins of the extracellular matrix is the most significant factor in CRC development in the early stages. This can be concluded since tumor growth is tightly associated with chronic aseptic inflammation and concatenated malignancy related to loss of extracellular matrix stability. Due attention should be paid to Apolipoprotein E (APOE), Apolipoprotein C1 (APOC1), and Apolipoprotein B-100 (APOB) because of their impact on the malfunction of DNA repair and their capability to regulate mTOR and PI3K pathways. The contribution of the observed PTMs is still equivocal, but a significant decrease in the likelihood between modified and native proteins was not detected confidently.

Identification of Significant Genes with poor prognosis in Liver Metastasis of Colorectal Cancer via Bioinformatical Analysis

“…APOE, which has been proved to be up regulated in patients with colorectal cancer (Borgquist, Butt et al 2016). It has been reported that overexpression of APOE was related to the growth of tumor in the advanced stage and increaseds the risk of tumor metastasis and invasion (Kopylov, Stepanov et al 2020), which may be related to the key role of APOE in cell proliferation and DNA synthesis (Niemi and Yla-Herttuala 2002). Moreover,APOE is one of the regulators of PI3K/Akt/mTOR pathway, which plays an important role in cell migration and proliferation, therefore, APOE can promote tumor progression by enhancing cell polarity (Vergadi, Ieronymaki et al 2017).…”

“…The complex of ITIH2 and hyaluronan plays a signi cant role in the in ammatory response and the high expression of ITIH2 is associated with poor outcome of NSCLC (Wu, Wang et al 2013). In addition, a strong link between the expression levels of ITIH2 and estrogen levels has been demonstrated, since ITIH2 contains an estrogen-binding domain which might be critical for metastasis and tumor growth because estrogen has a profound effect on extracellular matrix integrity (Kopylov, Stepanov et al 2020). At present, there is not much research on the role of ITIH2 in tumor growth and metastasis, which is also a potential direction for the study of tumor metastasis mechanism in the future CDH2 is a member of the cadherin family, which regulates many cellular processes including apoptosis, angiogenesis, and chemoresistance (Miao, Wang et al 2018).…”

Super Secondary Structures of Proteins with Post-Translational Modifications in Colon Cancer

“…We identified a portion of 57 believable PTM peptides corresponding to 29 proteins. Further data curation left only 25 proteins containing 42 PTMs in total, and of them 20 proteins with 29 PTMs were finally recovered as the most related to the group of CRC patients [17]. Among them, a conformational analysis was performed on six proteins containing 12 peptides with modified amino acid residues, for which intact 3D structures were annotated in the Protein Database (see Supplementary materials, Table S3).…”

“…Samples were collected into pre-chilled EDTA-2K tubes and centrifuged at 4 • C for 10 min at 1500 g after gentle agitation. Plasma fraction (1 mL) was transferred into the new clean cryovials (nominal volume is 2 mL) and frozen at −80 • C until the use [17].…”

“…Digestion was performed with trypsin (200 ng/µL supplemented in 30 mM acetic acid) in two stages: at the first stage trypsin was added at a ratio of 1:50 (w/w) and the reaction incubated for 3 h at 37 • C. At the second stage the enzyme was added at a ratio of 1:100 (w/w) and the reaction incubated at 37 • C for 12 h (approximately) [17]. Details of the sample preparation protocol are presented in the PRIDE Project PXD015163.…”