MicroRNAs biomarkers for early screening of colorectal cancer

Colorectal cancer (CRC) is the most incident neoplasia in Portugal. When diagnosed early, the 5-year cancer survival rate increases to 90% [2]. However, the current noninvasive screening method for CRC, Fecal Immunochemical Test (FIT), has low sensitivity and specificity for detecting precancerous lesions. Therefore, it is necessary to develop a new screening method for CRC. MicroRNAs (miRs) play a role in genetic events associated with carcinogenesis, and their disrupted expression in tumors can be readily detected in biological fluids [5-8]. This characteristic offers a promising tool for CRC screening. Review the existing literature to assess the advancements made in recent years in the potential use of miRs as a biomarker to improve the CRC screening. A comprehensive literature review was conducted, analyzing a total of 54 studies that investigated miRs expression in stool and blood samples and evaluated is potential as biomarkers for CRC identification. In our search, we identified a total of 104 miRs with potential relevance to CRC screening in both stool and blood samples. Among these miRs, miR-21-5p and miR-92a-3p, along their cluster including miR-29a-3p, miR-20a-5p, and miR-18-5p, emerged as the most frequently mentioned and promising candidates. Furthermore, is reported a differential expression of miR-135b-5p, miR-223-3p, and miR-451 only in stool specimens, while miR-139-3p and miR-4516 exhibit this altered expression in blood samples. Other notable miRs, including miR-146a-5p, miR-199a-5p, miR-421, miR-27a-3p, and miR-221-3p, have shown promising results in detecting advanced adenomas, exhibiting a better performance compared to FIT. However, these findings require further validation in a larger patient cohort and across different biological samples to confirm their significance for CRC and precancerous lesions detection. Therefore, miRs are regarded as a promising approach for enhancing the detection of CRC, particularly in the identification of precancerous lesions. Nevertheless, further studies are required to assess the accuracy of these molecules as biomarkers.info:eu-repo/semantics/publishedVersio

Glycan affinity magnetic nanoplatforms for urinary glycobiomarkers discovery in bladder cancer

Bladder Cancer (BC) presents one of the highest recurrence rates amongst solid tumours and constitutes the second deadliest disease of the genitourinary track. Non-invasive identification of patients facing disease recurrence and/or progression remains one of the most critical and challenging aspects in disease management. To contribute to this goal, we demonstrate the potential of glycan-affinity glycoproteomics nanoplatforms for urinary biomarkers discovery in bladder cancer. Briefly, magnetic nanoprobes (MNP) coated with three broad-spectrum lectins, namely Concanavalin A (ConA; MNP@ConA), Wheat Germ Agglutinin (WGA; MNP@WGA), and Sambucus nigra (SNA; MNP@SNA), were used to selectively capture glycoproteins from the urine of low-grade and high-grade non-muscle invasive as well as muscle-invasive BC patients. Proteins were identified by nano-LC MALDI-TOF/TOF and data was curated using bioinformatics tools (UniProt, NetOGlyc, NetNGlyc, ClueGO app for Cytoscape and Oncomine) to highlight clinically relevant species. Accordingly, 63 glycoproteins were exclusively identified in cancer samples compared with healthy controls matching in age and gender. Specific glycoprotein sets exclusively found in low-grade non-muscle invasive bladder tumours may aid early diagnosis, while those only found in high-grade non-invasive and muscle-invasive tumours hold potential for accessing progression. Amongst these proteins is bladder cancer stem-cell marker CD44, which has been associated with poor prognosis. Orthogonal validation studies by slot-blotting demonstrated an elevation in urine CD44 levels of high-grade patients, which became more pronounced upon muscle-invasion, in mimicry of the primary tumour. These observations demonstrate the potential of MNP@lectins for identification of clinically relevant glycoproteomics signatures in bladder cancer. Future clinical validation in a larger and well characterized patient subset is required envisaging clinical translation of the results.publishe

Abnormal Protein Glycosylation and Activated PI3K/Akt/mTOR Pathway: Role in Bladder Cancer Prognosis and Targeted Therapeutics

Muscle invasive bladder cancer (MIBC, stage >= T2) is generally associated with poor prognosis, constituting the second most common cause of death among genitourinary tumours. Due to high molecular heterogeneity significant variations in the natural history and disease outcome have been observed. This has also delayed the introduction of personalized therapeutics, making advanced stage bladder cancer almost an orphan disease in terms of treatment. Altered protein glycosylation translated by the expression of the sialyl-Tn antigen (STn) and its precursor Tn as well as the activation of the PI3K/Akt/mTOR pathway are cancer-associated events that may hold potential for patient stratification and guided therapy. Therefore, a retrospective design, 96 bladder tumours of different stages (Ta, T1-T4) was screened for STn and phosphorylated forms of Akt (pAkt), mTOR (pmTOR), S6 (pS6) and PTEN, related with the activation of the PI3K/Akt/mTOR pathway. In our series the expression of Tn was residual and was not linked to stage or outcome, while STn was statically higher in MIBC when compared to non-muscle invasive tumours (p = 0.001) and associated decreased cancer-specific survival (log rank p = 0.024). Conversely, PI3K/Akt/mTOR pathway intermediates showed an equal distribution between non-muscle invasive bladder cancer (NMIBC) and MIBC and did not associate with cancer-specif survival (CSS) in any of these groups. However, the overexpression of pAKT, pmTOR and/or pS6 allowed discriminating STn-positive advanced stage bladder tumours facing worst CSS (p = 0.027). Furthermore, multivariate Cox regression analysis revealed that overexpression of PI3K/Akt/mTOR pathway proteins in STn+ MIBC was independently associated with approximately 6-fold risk of death by cancer (p = 0.039). Mice bearing advanced stage chemically-induced bladder tumours mimicking the histological and molecular nature of human tumours were then administrated with mTOR-pathway inhibitor sirolimus (rapamycin). This decreased the number of invasive lesions and, concomitantly, the expression of STn and also pS6, the downstream effector of the PI3K/Akt/mTOR pathway. In conclusion, STn was found to be marker of poor prognosis in bladder cancer and, in combination with PI3K/Akt/mTOR pathway evaluation, holds potential to improve the stratification of stage disease. Animal experiments suggest that mTOR pathway inhibition could be a potential therapeutic approach for this specific subtype of MIBC

MicroRNA biomarkers as promising tools for early colorectal cancer screening—a comprehensive review

Colorectal cancer (CRC) ranks as the third most prevalent cancer worldwide. Early detection of this neoplasia has proven to improve prognosis, resulting in a 90% increase in survival. However, available CRC screening methods have limitations, requiring the development of new tools. MicroRNA biomarkers have emerged as a powerful screening tool, as they are highly expressed in CRC patients and easily detectable in several biological samples. While microRNAs are extensively studied in blood samples, recent interest has now arisen in other samples, such as stool samples, where they can be combined with existing screening methods. Among the microRNAs described in the literature, microRNA-21-5p and microRNA-92a-3p and their cluster have demonstrated high potential for early CRC screening. Furthermore, the combination of multiple microRNAs has shown improved performance in CRC detection compared to individual microRNAs. This review aims to assess the available data in the literature on microRNAs as promising biomarkers for early CRC screening, explore their advantages and disadvantages, and discuss the optimal study characteristics for analyzing these biomarkers.info:eu-repo/semantics/publishedVersio

CD44 glycoprotein in cancer: a molecular conundrum hampering clinical applications

Abstract CD44 is a heavily glycosylated membrane receptor playing a key role in cell adhesion, signal transduction and cytoskeleton remodelling. It is also one of the most studied glycoproteins in cancer, frequently explored for stem cell identification, and associated with chemoresistance and metastasis. However, CD44 is a general designation for a large family of splicing variants exhibiting different degrees of glycosylation and, potentially, functionally distinct roles. Moreover, structural diversity associated with ambiguous nomenclature has delayed clinical developments. Herein, we attempt to comprehensively address these aspects and systematize CD44 nomenclature, setting milestones for biomarker discovery. In addition, we support that CD44 may be an important source of cancer neoantigens, most likely resulting from altered splicing and/or glycosylation. The discovery of potentially targetable CD44 (glyco)isoforms will require the combination of glycomics with proteogenomics approaches, exploring customized protein sequence databases generated using genomics and transcriptomics. Nevertheless, the necessary high-throughput analytical and bioinformatics tools are now available to address CD44 role in health and disease

Reference Genes for Addressing Gene Expression of Bladder Cancer Cell Models under Hypoxia: A Step Towards Transcriptomic Studies

<div><p>Highly aggressive, rapidly growing tumors contain significant areas of hypoxia or anoxia as a consequence of inadequate and/or irregular blood supply. During oxygen deprivation, tumor cells withstand a panoply of adaptive responses, including a shift towards anaerobic metabolism and the reprogramming of the transcriptome. One of the major mediators of the transcriptional hypoxic response is the hypoxia-inducible factor 1 (HIF-1), whose stabilization under hypoxia acts as an oncogenic stimulus contributing to chemotherapy resistance, invasion and metastasis. Gene expression analysis by qRT-PCR is a powerful tool for cancer cells phenotypic characterization. Nevertheless, as cells undergo a severe transcriptome remodeling.in response to oxygen deficit, the precise identification of reference genes poses a significant challenge for hypoxic studies. Herein, we aim to establish the best reference genes for studying the effects of hypoxia on bladder cancer cells. Accordingly, three bladder cancer cell lines (T24, 5637, and HT1376) representative of two distinct carcinogenesis pathways to invasive cancer (FGFR3/CCND1 and E2F3/RB1) were used. Additionally, we have explored the most suitable control gene when addressing the influence of Deferoxamine Mesilate salt (DFX), an iron chelator often used to avoid the proteasomal degradation of HIF-1α, acting as an hypoxia-mimetic agent. Using bioinformatics tools (GeNorm and NormFinder), we have elected <i>B2M</i> and <i>HPRT</i> as the most stable genes for all cell lines and experimental conditions out of a panel of seven putative candidates (<i>HPRT</i>, <i>ACTB</i>, <i>18S</i>, <i>GAPDH</i>, <i>TBP</i>, <i>B2M</i>, and <i>SDHA</i>). These observations set the molecular basis for future studies addressing the effect of hypoxia and particularly HIF-1α in bladder cancer cells.</p></div

geNorm and NormFinder analysis of the stability values of reference genes and <i>CA9</i>.

<p>Samples were exposed to (A) normoxia and hypoxia, (B) normoxia and DFX, and (C) all experimental conditions (normoxia, hypoxia and DFX). <i>B2M</i> and <i>HPRT</i> are constitutively top ranked while <i>ACTB</i> and <i>GAPDH</i> are low ranked regardless of the experimental condition.</p

Determination of the optimal number of control genes for accurate normalization using geNorm software.

<p>Each bar V_<i>n/n+1</i> represents the variation between the means of the <i>n</i> most stable genes versus the group of <i>n+1</i> most stable genes. For example, column 1 represents the variation between the mean of the two most stable genes, <i>B2M</i> and <i>HPRT</i>, versus the three most stable genes <i>B2M</i>, <i>HPRT</i> and <i>SDHA</i>. Because the 0.15 threshold is not exceeded at any point, the use of two reference genes would be sufficient under these conditions.</p