Napoved rezistence na kemoterapijo na osnovi platine pri raku jajčnikov

Fortuna, Žan (author), Černe, Katarina (mentor)

Rak jajčnikov je izmed ginekoloških rakov najsmrtonosnejša oblika bolezni, kjer je najpogostejši serozni karcinom visokega gradusa (HGSC), ki izhaja iz epitelija na površini jajčnikov. Eden izmed glavnih razlogov za slabo prognozo je pozno odkrivanje bolezni, saj se simptomi največkrat pojavijo šele v napredovalnem stadiju. Zdravljenje se začne s citoreduktivno operacijo, ki ji sledi kemoterapija na osnovi platine (karboplatin), ta pa zaradi različnih genetskih in epigenetskih lastnosti celic istega tumorja ni vedno učinkovita. Približno tretjina bolnic se na primarno kemoterapijo ne odzove, pri 80 % tistih, ki se odzovejo pa pride do ponovitve bolezni, ki je v večini primerov rezistentna na platino in zato težko ozdravljiva. V želji po natančnejši diagnozi se išče nove prediktivne molekularne biooznačevalce, s katerimi bi lažje prilagodili zdravljenje in potencialno povečali verjetnost preživetja. Med njimi je tudi ATP7A, ATPaza tipa P, katere vloga v normalnih celicah je transport Cu+ v ekstracelular, pri rezistentnem HGSC pa podobno prenaša tudi Pt-BM (zdravilo na osnovi platine), s čimer prepreči toksične učinke zdravila. V okviru diplomskega dela smo želeli komercialno dostopnima celičnima linijama OAW42 in PEO1 določiti optimalne pogoje gojenja in morfološke lastnosti, po uspešnem gojenju pa tudi prisotnost transporterja ATP7A. Celice OAW42 so bolje rasle, ko so imele v mediju več natrijevega piruvata, celic PEO1 pa nam v mediju s pufrom MOPS ni uspelo uspešno gojiti. Ugotovili smo, da se morfologija celic ujema z literaturnimi podatki, saj so celice obeh celičnih linij zavzele značilno epitelijsko adherentno in enoplastno morfologijo, poleg tega pa imele tudi večje število jedrc, ki so posledica mutacij v genih in proteinih, ki ohranjajo integriteto DNK in uravnavajo celični cikel. S pretočnim citometrom smo celicam obeh celičnih linij potrdili prisotnost ATP7A. To predstavlja prvi korak pri ugotavljanju udeleženosti transporterja v mehanizem rezistence. V nadaljnjih študijah bo potrebno na podoben način s pomočjo drugih celičnih linij ta podatek ovrednotiti in narediti še teste citotoksičnosti s karboplatinom, s čimer se bo lažje povezalo ATP7A z rezistenco in ovrednotilo celični liniji OAW42 in PEO1 kot ustrezna modela za preučevanje rezistence.

• Language: Slovenian
• Keywords: rak jajčnikov, rezistenca, celični liniji OAW42 in PEO1, ATP7A, pretočna citometrija
• Work type: Bachelor thesis/paper
• Typology: 2.11 - Undergraduate Thesis
• Organization: FKKT - Faculty of Chemistry and Chemical Technology
• Year: 2022
• PID: 20.500.12556/RUL-138972
• COBISS.SI-ID: 120816899
• Publication date in RUL: 26.08.2022

Secondary language

• Language: English
• Title: Prediction of platinum-based chemotherapy resistance in ovarian cancer

Abstract

Ovarian cancer is the deadliest type of gynaecological cancer with high-grade serous carcinoma (HGSC) arising from the epithelium on the surface of the ovaries being the most common. One of the main reasons for poor prognosis is late detection of the disease, as symptoms most often appear only in the advanced stage. Treatment begins with cytoreductive surgery followed by platinum-based chemotherapy (carboplatin), which is not always effective due to the different genetic and epigenetic properties of cells from the same tumour. About one-third of patients do not respond to primary chemotherapy and 80 % of those who respond have a recurrence of the disease, which in most cases is resistant to platinum and therefore difficult to cure. In the desire for a more accurate diagnosis new predictive molecular biomarkers are being sought to help tailor treatment and potentially increase the likelihood of survival. Among them is ATP7A, P-type ATPase, whose role in normal cells is to transport Cu+ to extracellular and in resistant HGSC it similarly transmits Pt-BM (platinum-based medicine), thus preventing the toxic effects of the drug. In this work we wanted to determine the commercially available cell lines OAW42 and PEO1 optimal growing conditions and morphological properties. After successful cultivation we also wanted to determine the presence of the ATP7A transporter. OAW42 cells grew better when they had more sodium pyruvate in the medium, but we were unable to successfully grow PEO1 cells in medium with MOPS buffer. We found that the morphology of the cells matched the literature data, as the cells of both cell lines took on characteristic epithelial adherent and monolayer morphology and had a higher number of nuclei due to mutations in genes and proteins that maintain DNA integrity and regulate cell cycle. Using a flow cytometer, the presence of ATP7A was confirmed to the cells of both cell lines. This represents the first step in determining the involvement of the transporter in the resistance mechanism. Subsequent studies will need to evaluate this data in a similar way using other cell lines and perform carboplatin cytotoxicity tests to link ATP7A more easily to resistance and evaluate OAW42 and PEO1 cell lines as appropriate models for further resistance testing.

• Keywords: ovarian cancer, resistance, cell lines OAW42 and PEO1, ATP7A, flow cytometry