Abstract: Colorectal cancer is the third most common cancer and the second cause of cancer-related deaths. Rectal cancer presents roughly one-third of all colorectal cancer cases and differs from it on both anatomical and molecular levels. While standard treatment of colon cancer patients is radical surgery, rectal cancer is usually treated with pre-operative chemoradiotherapy followed by total mesorectal excision, which requires precise estimation of TNM staging. Unfortunately, stage evaluation is based solely on imaging modalities, and they often do not correlate with postoperative pathological findings. Moreover, approximately half of rectal cancer patients do not respond to such pre-operative therapy, so they are exposed to its toxic effects without any clinical benefit. Thus, biomarkers that could precisely predict pre-operative TNM staging, and especially response to therapy, would significantly advance rectal cancer treatment-but till now, no such biomarker has been identified. In cancer research, microRNAs are emerging biomarkers due to their connection with carcinogenesis and exceptional stability. Circulating miRNAs are promising non-invasive biomarkers that could allow monitoring of a patient throughout the whole therapeutic process. This mini-review aims to summarize the current knowledge on miRNAs and circulating miRNAs involved in the prediction of response to treatment and pre-operative staging in rectal cancer patients.

• Klíčová slova
• biomarker, circulating microRNA, colorectal cancer, miRNA, microRNA,
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

Blood plasma microRNAs (miRNAs) are emerging as a clinically useful tool for non-invasive detection and prognosis estimation in various cancer types including pancreatic ductal adenocarcinoma (PDAC). The aim of the present study was to provide an independent validation of circulating miRNAs identified in previous studies as diagnostic and/or prognostic biomarkers in PDAC. Based on the literature search, 6 miRNAs were chosen as candidates for independent validation; miR-21-5p, miR-375, miR-155, miR-17-5p, miR-126-5p and miR-1290. Validation of these miRNAs was performed in a cohort of 25 patients with PDAC undergoing surgical resection and 24 healthy donors. Plasma levels of miRNAs were determined using quantitative real-time PCR. We confirmed significantly higher levels of all tested miRNA in blood plasma of PDAC patients in comparison to healthy controls with miR-21-5p showing the highest analytical performance (p<0.001; AUC>0.99). Increased levels of miR-21-5p (p=0.045) and miR-375 (p=0.013) were significantly associated with overall survival. Multivariate analysis demonstrated that miR-21-5p is a significant unfavorable prognostic factor independent on other clinical variables including adjuvant chemotherapy (hazard ratio 2.95; 95% CI 1.06-8.18; p=0.038). Our preliminary data indicate promising diagnostic and prognostic utility of plasma miR-21-5p in PDAC patients.

• Klíčová slova
• Pancreatic ductal adenocarcinoma, diagnosis, miR-21-5p, microRNAs, plasma, prognosis,
• MeSH
• adenokarcinom krev   patologie   chirurgie   MeSH
• duktální karcinom slinivky břišní krev   patologie   chirurgie   MeSH
• kohortové studie MeSH
• lidé středního věku MeSH
• lidé MeSH
• mikro RNA krev   MeSH
• míra přežití MeSH
• nádorové biomarkery metabolismus   MeSH
• nádory slinivky břišní krev   patologie   chirurgie   MeSH
• následné studie MeSH
• předoperační péče * MeSH
• prognóza MeSH
• senioři MeSH
• studie případů a kontrol MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• mikro RNA MeSH
• MIRN21 microRNA, human MeSH Prohlížeč
• nádorové biomarkery MeSH

BACKGROUND: Senescence is a fundamental biological process implicated in various pathologies, including cancer. Regarding carcinogenesis, senescence signifies, at least in its initial phases, an anti-tumor response that needs to be circumvented for cancer to progress. Micro-RNAs, a subclass of regulatory, non-coding RNAs, participate in senescence regulation. At the subcellular level micro-RNAs, similar to proteins, have been shown to traffic between organelles influencing cellular behavior. The differential function of micro-RNAs relative to their subcellular localization and their role in senescence biology raises concurrent in situ analysis of coding and non-coding gene products in senescent cells as a necessity. However, technical challenges have rendered in situ co-detection unfeasible until now. METHODS: In the present report we describe a methodology that bypasses these technical limitations achieving for the first time simultaneous detection of both a micro-RNA and a protein in the biological context of cellular senescence, utilizing the new commercially available SenTraGorTM compound. The method was applied in a prototypical human non-malignant epithelial model of oncogene-induced senescence that we generated for the purposes of the study. For the characterization of this novel system, we applied a wide range of cellular and molecular techniques, as well as high-throughput analysis of the transcriptome and micro-RNAs. RESULTS: This experimental setting has three advantages that are presented and discussed: i) it covers a "gap" in the molecular carcinogenesis field, as almost all corresponding in vitro models are fibroblast-based, even though the majority of neoplasms have epithelial origin, ii) it recapitulates the precancerous and cancerous phases of epithelial tumorigenesis within a short time frame under the light of natural selection and iii) it uses as an oncogenic signal, the replication licensing factor CDC6, implicated in both DNA replication and transcription when over-expressed, a characteristic that can be exploited to monitor RNA dynamics. CONCLUSIONS: Consequently, we demonstrate that our model is optimal for studying the molecular basis of epithelial carcinogenesis shedding light on the tumor-initiating events. The latter may reveal novel molecular targets with clinical benefit. Besides, since this method can be incorporated in a wide range of low, medium or high-throughput image-based approaches, we expect it to be broadly applicable.

• Klíčová slova
• CDC6, Cancer, DNA damage response, In situ hybridization, Micro-RNAs, Oncogene-induced senescence, R loops, Replication stress, SenTraGorTM, rDNA,
• MeSH
• epitelové buňky metabolismus   MeSH
• genom MeSH
• jaderné proteiny metabolismus   MeSH
• karcinogeneze MeSH
• kultivované buňky MeSH
• lidé MeSH
• mikro RNA metabolismus   MeSH
• nádory glandulární a epitelové genetika   patologie   ultrastruktura   MeSH
• onkogeny * MeSH
• proteiny buněčného cyklu metabolismus   MeSH
• proteiny metabolismus   MeSH
• stanovení celkové genové exprese MeSH
• stárnutí buněk genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• CDC6 protein, human MeSH Prohlížeč
• jaderné proteiny MeSH
• mikro RNA MeSH
• proteiny buněčného cyklu MeSH
• proteiny MeSH

MicroRNAs are a class of small non-coding RNAs that serve as important regulators of gene expression at the posttranscriptional level. They are stable in body fluids and pose great potential to serve as biomarkers. Here, we present a highly specific, sensitive and cost-effective system to quantify miRNA expression based on two-step RT-qPCR with SYBR-green detection chemistry called Two-tailed RT-qPCR. It takes advantage of novel, target-specific primers for reverse transcription composed of two hemiprobes complementary to two different parts of the targeted miRNA, connected by a hairpin structure. The introduction of a second probe ensures high sensitivity and enables discrimination of highly homologous miRNAs irrespectively of the position of the mismatched nucleotide. Two-tailed RT-qPCR has a dynamic range of seven logs and a sensitivity sufficient to detect down to ten target miRNA molecules. It is capable to capture the full isomiR repertoire, leading to accurate representation of the complete miRNA content in a sample. The reverse transcription step can be multiplexed and the miRNA profiles measured with Two-tailed RT-qPCR show excellent correlation with the industry standard TaqMan miRNA assays (r2 = 0.985). Moreover, Two-tailed RT-qPCR allows for rapid testing with a total analysis time of less than 2.5 hours.

• MeSH
• kvantitativní polymerázová řetězová reakce metody   MeSH
• mikro RNA analýza   genetika   MeSH
• myši MeSH
• prekurzory RNA analýza   genetika   MeSH
• reprodukovatelnost výsledků MeSH
• senzitivita a specificita MeSH
• stanovení celkové genové exprese metody   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• validační studie MeSH
• Názvy látek
• mikro RNA MeSH
• prekurzory RNA MeSH

MicroRNAs (miRNAs) are small non-coding RNAs 18-25 nucleotides in length that downregulate gene expression during various crucial cell processes such as apoptosis, differentiation and development. Changes in the expression profiles of miRNAs have been observed in a variety of human tumors, including colorectal cancer (CRC). Functional studies indicate that miRNAs act as tumor suppressors and oncogenes. These findings significantly extend Vogelstein's model of CRC pathogenesis and have shown great potential for miRNAs as a novel class of therapeutic targets. Several investigations have also described the ability of miRNA expression profiles to predict prognosis and response to selected treatments in CRC patients, and support diagnosis of CRC among cancer of unknown primary site. miRNAs' occurrence has been repeatedly observed also in serum and plasma, and miRNAs as novel minimally invasive biomarkers have indicated reasonable sensitivity for CRC detection and compare favorably with the fecal occult blood test. In this review, we summarize the knowledge regarding miRNAs' functioning in CRC while emphasizing their significance in pathogenetic signaling pathways and their potential to serve as disease biomarkers and novel therapeutic targets.

• MeSH
• kolorektální nádory krev   diagnóza   etiologie   genetika   MeSH
• lidé MeSH
• mikro RNA krev   genetika   metabolismus   MeSH
• molekulární biologie * MeSH
• polymorfismus genetický MeSH
• stanovení celkové genové exprese MeSH
• translační biomedicínský výzkum * MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• mikro RNA MeSH