Department of Biology
Library of the School of Science

2022-05-03

2022

Diamantopoulos Ioannis-Marios

1. Ανδρέας Σκορίλας, (Επιβλέπων), Καθηγητής, Τμήμα Βιολογίας, Ε.Κ.Π.Α
2. Δημήτριος Γουργιώτης, Ομότιμος Καθηγητής, Ιατρική Σχολή, Ε.Κ.Π.Α
3. Διαμάντης Σίδερης, Αναπληρωτής Καθηγητής, Τμήμα Βιολογίας, Ε.Κ.Π.Α
4. Μαργαρίτης Αυγέρης, Αναπληρωτής Καθηγητής, Ιατρική Σχολή, Ε.Κ.Π.Α
5. Ανδρέας Αγαθαγγελίδης, Επίκουρος Καθηγητής, Τμήμα Βιολογίας, Ε.Κ.Π.Α
6. Νικόλαος Δανιάς, Επίκουρος Καθηγητής, Ιατρική Σχολή, Ε.Κ.Π.Α
7. Χρήστος Κοντός, Επίκουρος Καθηγητής, Τμήμα Βιολογίας, Ε.Κ.Π.Α

ΤΑΥΤΟΠΟΙΗΣΗ ΝΕΩΝ ΜΗ ΚΩΔΙΚΩΝ RNAS (NCRNAS), ΜΕΣΩ ΑΛΛΗΛΟΥΧΗΣΗΣ ΝΕΑΣ ΓΕΝΙΑΣ (NGS), ΚΑΙ ΜΕΛΕΤΗ ΤΗΣ ΕΚΦΡΑΣΗΣ ΤΟΥΣ ΣΕ ΚΑΡΚΙΝΙΚΑ ΚΥΤΤΑΡΑ ΚΑΙ ΣΤΕΡΕΟΥΣ ΟΓΚΟΥΣ

Greek

IDENTIFICATION AND EXPRESSION ANALYSIS OF NON-CODING RNAS (NCRNAS) IN CANCER CELLS AND SOLID TUMORS, THROUGH NEXT-GENERATION SEQUENCING (NGS).

Colon cancer and prostate cancer constitute categories of solid malignancies. Chronic lymphocytic leukemia is also a major category of haematological malignancies. Colon cancer is the third most common cancer diagnosed in men and women, numbering hundreds of thousands of new cases worldwide, on an annual basis. This cancer is one of the leading (3rd in women and 2nd in men) causes of cancer death. At the same time, prostate cancer (PCa) is the perfect incidence of cancer in Europe and is the most common non-skin cancer among men. Hematologic malignancies are a heterogeneous group of diseases of the hematopoietic tissue and together, represent about 10% of all cancers. The discovery of new sensitive molecular biomarkers for the valid diagnosis, prognosis and prediction of treatment is also a major issue. Next-Generation Sequencing (NGS) is a state-of-the-art set of methodologies for large scale genome and transcript studies, as it allows the sequencing of an extremely large number of nucleic acid molecules without the need to design an equally large number of nucleotides. . For these reasons, NGS is a "revolution" in the fields of Molecular Biology. One of the most important applications of NGS technology is RNA sequencing (RNA-seq), a method for detailed transcript analysis. The application of RNA-seq and modern bioinformatics tools has revealed the existence of an abundance of ncRNAs that are transcribed from the genome. NcRNAs are divided by size into two broad categories, the short-lived ncRNA (sncRNA) and the large ncRNA (lncRNA) molecules. Data from research related to the sequencing of the human genome suggest that more than 80% of human DNA is transcriptionally active. However, less than 3% encode proteins. Non-coding RNAs (ncRNAs) are distinguished into regulatory ncRNAs as well as housekeeping ncRNAs. The latter include ribosomal RNAs (rRNAs) and transport RNAs (tRNAs). Regulatory ncRNAs depending on their size can be divided into two groups: small ncRNAs ( 200 nucleotides). Small RNAs contain microRNAs, small nuclear RNAs (snRNAs), small nuclear RNAs (snoRNAs), small interfering RNAs (siRNAs) and RNAs that interact with the PIWI protein (piRNAs). It is worth noting that the largest percentage of human cellular RNA consists of ribosomal rRNA. Other abundant species, such as mRNA, snoRNA and snRNA, are expressed at levels 1-2 times lower than rRNA and tRNA. MicroRNAs may also show high levels of expression, but this depends on the type and function of each cell (eg microRNA-92a-3p). In cancer samples they are found to have a deviant expression from normal samples. Altering the expression of microRNAs in turn affects the expression of a few tens-hundreds of genes. The expression of microRNAs is also affected by the permethylation and hypomethylation of CpG islets, specifically in colorectal cancer. As for lncRNAs, overall they are relatively low. Finally, it is worth mentioning that more and more types of non-coding RNAs are being discovered, such as enhancer RNAs (eRNAs), which are thought to act by regulating the three-dimensional structure of chromosomes near the transcription sites. Another example is cyclic RNAs that act as sponge microRNAs, as they homologize with them and thus prevent them from interacting with other molecules. The present study aims at the molecular identification, study and evaluation of new biomolecules that can be used as molecular biomarkers, mainly in colon and prostate cancer. In the present dissertation, cutting-edge molecular methodologies - such as Next-Generation Sequencing (NGS) - have been applied to identify, develop assay protocols, and study the expression of new small non-coding RNA (sncRNA) molecules. With this approach, 10 novel small non-coding RNA molecules were identified and registered in the corresponding database (GenBank of NCBI). This was followed by the development and validation of molecular methodologies for their determination in cancer cells and tissues, and then their expression in a number of cancer lines and in colon and prostate cancer tissues was studied. At the same time, the effect of chemotherapeutic drugs on colon cancer cell lines was studied while an in vitro study of cell viability and proliferation rate was performed, as well as a study of the cytotoxicity of chemotherapeutic drugs. The results showed that a) In the cells of the human colon cancer cell line Caco-2, incubated with the anti-cancer drug Irinotecan, there is a marked increase in the expression levels of MT815884 after 24 hours which decreases to 48 and 72 hours of incubation. Regarding the effect of Elotaxin, a gradual increase in incubation time-related effects was observed that was not observed with methotrexate. b) The expression of the non-coding RNA 28S rRF molecule (MT815881) in prostate cancer patients was found to be a new, candidate molecular biomarker with an unfavorable prognosis of the disease. c) Expression of the small 8 non-coding miR-16-5p molecule was found to be a new, candidate molecular biomarker of adverse prognosis in colon adenocarcinoma. d) Expression of miR-92a-3p in peripheral blood mononuclear cells has been found to be a new, independent molecular biomarker with a favorable prognosis in chronic lymphocytic leukemia. In addition, a low-risk, high-risk patient molecular classification system was developed to address the risk of recurrence of the studied non-coding RNA molecules under study

Science

Non-coding RNAs (ncRNAs) , Next-Generation Sequencing (NGS) , Quantitive Real – time PCR (qPCR) , miRDeep* ,microRNA-16-5p , microRNA-92a-3p , Cancer

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https://pergamos.lib.uoa.gr/uoa/dl/object/3216404