Die funktionelle Rolle nicht-kodierender RNAs in gastrointestinalen Stromatumoren

Gastrointestinal stromal tumours (GISTs) are the most common mesenchymal tumours of the gastrointestinal tract. Despite the fact that the vast majority of GISTs harbour common activating mutations of KIT and PDGFRA, heterogeneous clinical behaviour with tumours of distinct aggressiveness is a feature of the disease. These observations suggest the possible role of other molecular factors beyond KIT and PDGFRA mutations that may contribute to the tumour phenotype. Different types of non-coding transcripts are being increasingly recognised as crucial components of regulatory networks that can be deregulated in cancer. While the functionality and molecular mechanism of miRNAs are relatively well described, the role of long non-coding transcripts remains only incompletely understood given more complicated and diverse regulatory functions. Even less is known if epigenetic-related lncRNAs are capable of affecting DNA methylation patterns in oncogenesis. Ascertainment of the role of non-coding transcripts in GIST is important for deeper understanding of the pathological mechanism of the disease identification of therapeutic targets and molecular markers. In this context, the aim of the study was to analyse the contribution of different classes of ncRNAs in development and progression of GIST. In the first part of the work, we took advantage of the laboratory’s preliminary data on miRNA and mRNA expression in distinct clinic-pathological subgroups of GIST. We predicted in silico and experimentally validated miR-665/CD34 and miR-409-3p/PRKCA pairs that are functionally relevant in GIST. MiR-409-3p regulates cell viability and apoptosis in imatinib-resistant GIST cells via targeting Protein Kinase C alpha. A similar phenotypic effect was also achieved by inhibition of PRKCA with a low molecular weight inhibitor dequalinium chloride suggesting PRKCA as a potential druggable target in imatinib resistant GIST. The expression of the second target, the histopathological biomarker of GIST CD34, demonstrated the anatomic localisation-specific pattern that is a risk factor for tumour recurrence. In GIST cell line models, CD34 was targeted by miR-665. Additionally, the gene’s expression was regulated by promoter DNA methylation with tumour localization-dependent inverse methylation levels. In the second part of the project, we set off to identify different classes of non-coding transcripts that may be of potential relevance for development of an aggressive phenotype of GIST by performing first genome-wide array-based expression analysis of ncRNAs in GIST patient samples of different risk of recurrence. Numerous transcripts were associated with low and high risk GIST. A prominent long non-coding RNA candidate HOTAIR was selected as a representative epigenetic-related transcript for testing a hypothesis about its capability of specifically patterning DNA methylation in GIST. Global alterations of DNA methylation patterns were detected in GIST cell line model by means of HumanMethylation450 BeadChip array upon stable knock-down of HOTAIR. These results were additionally validated for functionally relevant genes DPP4 and ALDH1A3 by using bisulfite pyrosequencing. While the molecular mechanism remains to be determined, it is plausible to assume a recruitment of DNA methyltransferases by the Polycomb repressive complex 2, which target specificity is determined by HOTAIR. This transcript was functionally important in GIST cells by having an effect on cell migration and affecting the expression of multiple genes that may be relevant for invasive properties. Other deregulated in patient samples non-coding transcripts belonged to classes of lincRNAs, snoRNAs and snRNAs. The expression of most prominent candidates in patient samples was validated by qRT-PCR. Pending confirmation on a larger sample sets, these transcripts may be of interest as potential biomarkers of different clinic-pathological subgroups of GIST. Taken together, our results indicate significant deregulation of different classes of non-coding transcripts in GIST. We have shown the functional importance of two miRNA species, miR-665 and miR-409-3p, and the role of the lncRNA HOTAIR that affects DNA methylation patterns of multiple genes. While the functionality of other lncRNAs and small transcripts such as sno- and snRNA remains to be elucidated, their global deregulation suggests the functional contribution to the tumour phenotype

Methylation and Noncoding RNAs in Gastric Cancer: Everything Is Connected

Despite recent progress, gastric cancer remains one of the most common cancers and has a high mortality rate worldwide. Aberrant DNA methylation pattern and deregulation of noncoding RNA expression appear in the early stages of gastric cancer. Numerous investigations have confirmed their significant role in gastric cancer tumorigenesis and their high potential as diagnostic and prognostic biomarkers. Currently, it is clear that these epigenetic regulators do not work alone but interact with each other, generating a complex network. The aim of our review was to summarize the current knowledge of this interaction in gastric cancer and estimate its clinical potential for the diagnosis, prognosis, and treatment of the disease

Mutual Regulation of ncRNAs and Chromatin Remodeling Complexes in Normal and Pathological Conditions

Chromatin remodeling is the one of the main epigenetic mechanisms of gene expression regulation both in normal cells and in pathological conditions. In recent years, a growing number of investigations have confirmed that epigenetic regulators are tightly connected and form a comprehensive network of regulatory pathways and feedback loops. Genes encoding protein subunits of chromatin remodeling complexes are often mutated and change their expression in diseases, as well as non-coding RNAs (ncRNAs). Moreover, different mechanisms of their mutual regulation have already been described. Further understanding of these processes may help apply their clinical potential for establishment of the diagnosis, prognosis, and treatment of the diseases. The therapeutic targeting of the chromatin structure has many limitations because of the complexity of its regulation, with the involvement of a large number of genes, proteins, non-coding transcripts, and other intermediary molecules. However, several successful strategies have been proposed to target subunits of chromatin remodeling complexes and genes encoding them, as well as the ncRNAs that regulate the operation of these complexes and direct them to the target gene regions. In our review, we focus on chromatin remodeling complexes and ncRNAs, their mutual regulation, role in cellular processes and potential clinical application

Histone Modifications and Non-Coding RNAs: Mutual Epigenetic Regulation and Role in Pathogenesis

In the last few years, more and more scientists have suggested and confirmed that epigenetic regulators are tightly connected and form a comprehensive network of regulatory pathways and feedback loops. This is particularly interesting for a better understanding of processes that occur in the development and progression of various diseases. Appearing on the preclinical stages of diseases, epigenetic aberrations may be prominent biomarkers. Being dynamic and reversible, epigenetic modifications could become targets for a novel option for therapy. Therefore, in this review, we are focusing on histone modifications and ncRNAs, their mutual regulation, role in cellular processes and potential clinical application

Roles of E-cadherin and Noncoding RNAs in the Epithelial–mesenchymal Transition and Progression in Gastric Cancer

The epithelial–mesenchymal transition (EMT) is thought to be at the root of invasive and metastatic cancer cell spreading. E-cadherin is an important player in this process, which forms the structures that establish and maintain cell–cell interactions. A partial or complete loss of E-cadherin expression in the EMT is presumably mediated by mechanisms that block the expression of E-cadherin regulators and involve the E-cadherin-associated transcription factors. The protein is involved in several oncogenic signaling pathways, such as the Wnt/β-catenin, Rho GTPase, and EGF/EGFR, whereby it plays a role in many tumors, including gastric cancer. Such noncoding transcripts as microRNAs and long noncoding RNAs—critical components of epigenetic control of gene expression in carcinogenesis—contribute to regulation of the E-cadherin function by acting directly or through numerous factors controlling transcription of its gene, and thus affecting not only cancer cell proliferation and metastasis, but also the EMT. This review focuses on the role of E-cadherin and the non-coding RNAs-mediated mechanisms of its expressional control in the EMT during stomach carcinogenesis

Genetic and Clinical Factors Associated with Olokizumab Treatment in Russian Patients with Rheumatoid Arthritis

Rheumatoid arthritis (RA) is a chronic systemic inflammatory disease and its treatment is an urgent problem of rheumatology. Olokizumab (OKZ) is a new humanized monoclonal antibody targeting IL-6 and is one of the few promising drugs for RA therapy. One-hundred-and-twenty-five DNA samples from Russian patients with RA, treated with olokizumab, were genotyped with an NGS panel containing 60 single nucleotide polymorphisms (SNPs) and the whole coding sequences of IL6, IL6R, TNFRSF1A, CTLA4, IL10, IL23R, and PADI4; and by RT-PCR for HLA-DRB1 and HLA-B. Associations of polymorphic variants with olokizumab efficacy according to the scores ACR20, ACR50, and DAS28-CRP were determined. We analyzed the obtained data by using logistic regression, ROC curves, and multivariate ANOVA. A high predictive value of the response to olokizumab therapy at 24 weeks was found for the combination of HLA-DRB1*04 and HLA-B*27 alleles with SNPs located in non-HLA genes (IL1B, IL17A, PADI4, DHODH, GLCCI1, IL23R, and TNFAIP3), and clinical characteristics (age, RA duration, and intensity) according to ACR20. Thus, the comprehensive assessment of polymorphic variants of HLA and non-HLA genes considering population characteristics in combination with clinical parameters allows for the elaboration of an RA prognostic panel

Genetic Polymorphisms Associated with Rheumatoid Arthritis Development and Antirheumatic Therapy Response

Rheumatoid arthritis (RA) is the most common inflammatory arthropathy worldwide. Possible manifestations of RA can be represented by a wide variability of symptoms, clinical forms, and course options. This multifactorial disease is triggered by a genetic predisposition and environmental factors. Both clinical and genealogical studies have demonstrated disease case accumulation in families. Revealing the impact of candidate gene missense variants on the disease course elucidates understanding of RA molecular pathogenesis. A multivariate genomewide association study (GWAS) based analysis identified the genes and signalling pathways involved in the pathogenesis of the disease. However, these identified RA candidate gene variants only explain 30% of familial disease cases. The genetic causes for a significant proportion of familial RA have not been determined until now. Therefore, it is important to identify RA risk groups in different populations, as well as the possible prognostic value of some genetic variants for disease development, progression, and treatment. Our review has two purposes. First, to summarise the data on RA candidate genes and the increased disease risk associated with these alleles in various populations. Second, to describe how the genetic variants can be used in the selection of drugs for the treatment of RA