Uloga proteina DNER u epitelno-mezenhimskoj tranziciji ljudskih stanica tumora jajnika otpornih na karboplatinu

master's thesis

• Title (english): Role of DNER protein in carboplatin induced epithelial-mesenchymal transition in human ovarian cancer cells
• Author: Dora Gjirlić
• Mentor: Anamaria Brozović (mentor)
• Mentor: Maja Matulić (mentor)
• Committee member: Maja Matulić (predsjednik povjerenstva)
• Committee member: Duje Lisičić (član povjerenstva)
• Committee member: Nada Oršolić (član povjerenstva)
• Committee member: Silvija Černi (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Biology); Zagreb
• Defense date and country: 2019-09-27, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Biology

Abstract

Serozni tumor jajnika najsmrtonosniji je od svih ginekoloških karcinoma. Poodmakli stadij bolesti i razvoj otpornosti na terapiju glavni su razlog neuspješnosti terapije. U potrazi za zajedničkim biljegom odgovornim za regulaciju molekularnih mehanizama otpornosti na lijek te pojave epitelno-mezenhimske tranzicije (EMT), u preliminarnom istraživanju provedena je analiza genske ekspresije u roditeljskoj liniji stanica tumora jajnika MES-OV i liniji stanica otpornoj na karboplatinu (CBP) MES-OV 8CBP metodom microarray. Nakon provedene bioinformatičke analize utvrđeno je da je ekspresija gena DNER, koji kodira za protein DNER, netipični aktivator signalnog puta Notch, bila značajno povećana u MES-OV 8CBP u odnosu na roditeljsku liniju stanica. Ovim radom istražena je moguća uloga DNER u otpornosti OVCAR-3 7CBP GFP stanica na CBP i u EMT. Pokazano je da je linija stanica OVCAR-3 7CBP GFP 2-3 puta otpornija na CBP u odnosu na roditeljsku liniju stanica OVCAR-3 GFP te da ima povećanu ekspresiju biljega mezenhimskog fenotipa, i smanjenu ekspresiju biljega epitelnog fenotipa. Metodom RT-qPCR potvrđena je povećana ekspresija DNER u liniji stanica MES-OV 8CBP u odnosnu na roditeljsku liniju te u dodatne dvije linije stanica raka jajnika otpornih na CBP, OVCAR-3 7CBP GFP i SK-OV-3 6CBP, u odnosu na njihove odgovarajuće roditeljske linije stanica. Važnost DNER u odgovoru stanica na CBP uočena je nakon kratkotrajnog izlaganja OVCAR-3 GFP i OVCAR-3 7CBP GFP kliničkoj dozi CBP kada je povećana ekspresija DNER izmjerena kod otporne linije stanica. Zbog nedostatka specifičnog anti-DNER protutijela nije bilo moguće utvrditi konstitutivnu ekspresiju proteina DNER i postići zadane ciljeve.

Abstract (english)

Serous ovarian tumour is the deadliest of all gynaecologic carcinomas. The advanced stage of the disease and the development of drug resistance are main reasons for unsuccesful therapy. While searching for a common marker(s) responsible for regulating molecular mechanisms of drug resistance and the occurrence of epithelial-mesenchymal transition (EMT), gene expression analysis of ovarian tumour cell lines, parental MES-OV and carboplatin (CBP) resistant MES-OV 8CBP by microarray assay was performed previously as part of preliminary research. Among others, the increased expression of gene DNER that encodes DNER protein, an alternative activator of Notch signalling pathway, was found. This work sought to determine whether DNER plays a role in the CBP resistance of OVCAR-3 7CBP GFP cells and in the development of a mesenchymal-like phenotype. The OVCAR-3 7CBP GFP cell line has been shown to be 2-3 times more resistant to CBP than the parent OVCAR-3 GFP cell line and has increased expression of mesenchymal-like phenotype and decreased expression of the epithelial-like phenotype markers. The RT-qPCR method confirmed the result of preliminary studies on increased DNER expression in the MES-OV 8CBP cell line relative to the parent cells, as well as in two additional human ovarian cancer cell lines resistant to CBP, OVCAR-3 7CBP GFP and SK-OV-3 6CBP. The importance of DNER in response to CBP was observed after short-term exposure of OVCAR-3 GFP and OVCAR-3 7CBP GFP to a clinical dose of CBP when DNER expression was increased only in the resistant cell line. At the moment the lack of specific anti-DNER antibody has made it impossible to determine the constitutive expression of DNER protein in experimental cell model and to achived the set goals.

• Keywords: progresija tumora; otpornost na kemoterapeutik; signalni put; utišavanje
• Keywords (english): tumour progression; chemoresistance; signal path; silencing
• Language: croatian
• URN:NBN: urn:nbn:hr:217:147900
• Study programme: Title: Molecular Biology; Study programme type: university; Study level: graduate; Academic / professional title: magistar/magistra molekularne biologije (magistar/magistra molekularne biologije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access; Embargo expiration date: 2021-09-27
• Created on: 2019-10-14 08:52:16