Petra Faltejsková

Novel classes of non-coding RNAs and cancer

For the many years, the central dogma of molecular biology has been that RNA functions mainly as an informational intermediate between a DNA sequence and its encoded protein. But one of the great surprises of modern biology was the discovery that protein-coding genes represent less than 2% of the total genome sequence, and subsequently the fact that at least 90% of the human genome is actively transcribed. Thus, the human transcriptome was found to be more complex than a collection of protein-coding genes and their splice variants. Although initially argued to be spurious transcriptional noise or accumulated evolutionary debris arising from the early assembly of genes and/or the insertion of mobile genetic elements, recent evidence suggests that the non-coding RNAs (ncRNAs) may play major biological roles in cellular development, physiology and pathologies. NcRNAs could be grouped into two major classes based on the transcript size; small ncRNAs and long ncRNAs. Each of these classes can be further divided, whereas novel subclasses are still being discovered and characterized. Although, in the last years, small ncRNAs called microRNAs were studied most frequently with more than ten thousand hits at PubMed database, recently, evidence has begun to accumulate describing the molecular mechanisms by which a wide range of novel RNA species function, providing insight into their functional roles in cellular biology and in human disease. In this review, we summarize newly discovered classes of ncRNAs, and highlight their functioning in cancer biology and potential usage as biomarkers or therapeutic targets.

Identification and functional screening of microRNAs highly deregulated in colorectal cancer

MicroRNAs (miRNAs) constitute a robust regulatory network with post‐transcriptional regulatory efficiency for almost one half of human coding genes, including oncogenes and tumour suppressors. We determined the expression profile of 667 miRNAs in colorectal cancer (CRC) tissues and paired non‐tumoural tissues and identified 42 differentially expressed miRNAs. We chose miR‐215, miR‐375, miR‐378, miR‐422a and miR‐135b for further validation on an independent cohort of 125 clinically characterized CRC patients and for in vitro analyses. MiR‐215, miR‐375, miR‐378 and miR‐422a were significantly decreased, whereas miR‐135b was increased in CRC tumour tissues. Levels of miR‐215 and miR‐422a correlated with clinical stage. MiR‐135b was associated with higher pre‐operative serum levels of CEA and CA19‐9. In vitro analyses showed that ectopic expression of miR‐215 decreases viability and migration, increases apoptosis and promotes cell cycle arrest in DLD‐1 and HCT‐116 colon cancer cell lines. Similarly, overexpression of miR‐375 and inhibition of miR‐135b led to decreased viability. Finally, restoration of miR‐378, miR‐422a and miR‐375 inhibited G1/S transition. These findings indicate that miR‐378, miR‐375, miR‐422a and miR‐215 play an important role in CRC as tumour suppressors, whereas miR‐135b functions as an oncogene; both groups of miRNA contribute to CRC pathogenesis.

MicroRNAs targeting EGFR signalling pathway in colorectal cancer.

MicroRNAs (miRNAs) are short, 18–25-nucleotide long, non-coding single-stranded RNAs, which are capable to regulate gene expression on post-transcriptional level through binding to their target protein-encoding mRNAs. miRNAs regulate individual components of multiple oncogenic pathways. One of them is epidermal growth factor receptor (EGFR) signalling pathway that regulates cell proliferation, differentiation, migration, angiogenesis and apoptosis. All these processes are deregulated in colorectal cancer (CRC). Moreover, EGFR has been validated as the therapeutic target in CRC, and monoclonal antibodies cetuximab and panitumumab are used in the therapy of patients with metastatic CRC. Because of the extensive involvement of miRNAs in the regulation of EGFR signalling, it seems they could also serve as promising predictive biomarkers to anti-EGFR therapy. In this review, we summarize current knowledge about miRNAs targeting EGFR signalling pathway, their functioning in CRC pathogenesis and potential usage as biomarkers.

Circulating miR-17-3p, miR-29a, miR-92a and miR-135b in serum: Evidence against their usage as biomarkers in colorectal cancer

BACKGROUND Colorectal cancer (CRC) is one of the leading causes of cancer-related death in the world. Therefore, there is a high demand for cost-effective and non-invasive biomarkers that would enable an early detection of asymptomatic and curable disease with high sensitivity and specificity. OBJECTIVE The main objective of this study was to investigate the potential of circulating miRNAs as biomarkers of CRC. METHODS Total RNA enriched for small RNAs was isolated from 100~sera of patients with CRC and 30 sera of healthy donors. The expression levels of miR-17-3p, miR-29a, miR-92a and miR-135b were determined using quantitative real-time PCR. The average expression levels of particular miRNAs were normalized to miR-16 levels and statistically evaluated. RESULTS Using Mann-Whitney U test, no significant differences were observed in miR-17-3p (P=0.18), miR-29a (P=0.14) and miR-92a (P=0.60) levels between sera of CRC patients and controls. The levels of miR-135b in serum were too low to be quantified accurately. Subsequently, we tried to correlate expression levels of analyzed miRNAs to clinical-pathological features of CRC patients. Only levels of mir-29a were correlated with the clinical stage (P=0.04). Expression levels of the other miRNAs were correlated neither with the clinical stage, nor with the grade. CONCLUSIONS Interestingly, our results are contradictory to previous studies performed on the CRC patients from Chinese population, providing an evidence against usage of serum miR-17-3p, miR-29a, miR-92a and miR-135b as new biomarkers for early detection of CRC.

Abstract 4172: MiR-215 expression in tumor tissue and in vitro effects of its replacement in colorectal cancer.

Background: MicroRNAs (miRNAs) constitute a robust regulatory network with post-transcriptional regulatory efficiency for almost one half of human coding genes, including oncogenes and tumor suppressors. Methods: We determined the expression profile of 667 miRNAs in colorectal cancer (CRC) tissues and paired non-tumoral tissues and identified 42 differentially expressed miRNAs. One of the most significantly altered miRNAs was miR-215, significantly down-regulated in tumor tissue. We chose miR-215 for further validation on an independent cohort of 125 clinically characterized CRC patients and for in vitro functional studies on DLD1, HCT116 and HT29 cell lines. Results: MiR-215 was proved to be significantly decreased in CRC tumor tissues (p Conclusions: These findings indicate that miR 215 play an important role in CRC as tumor suppressor and contributes to CRC pathogenesis. Supported by Internal Grant Agency of Czech Ministry of Health, grant numbers: NT13549-4/2012 a NT13860 4/2012. Citation Format: Petra Faltejskova, Marek Svoboda, Jitka Mlcochova, Klara Srutova, Pavel Fabian, Rostislav Vyzula, Ondrej Slaby. MiR-215 expression in tumor tissue and in vitro effects of its replacement in colorectal cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4172. doi:10.1158/1538-7445.AM2013-4172