Development of novel in vitro 3D cell models of ovarian cancer for drug screening

Kokondoska Grgič, Vesna (avtor), Sinreih, Maša (mentor), Jovchevska, Ivana (komentor)

High grade serous ovarian cancer (HGSOC) is the most aggressive and lethal subtype of ovarian cancers (OC). Although patients initially respond to platinum-based chemotherapy, most eventually relapse due to the development of chemoresistance. One of the major limitations lies in the use of two-dimensional (2D) cultures, which do not accurately reflect the complex architecture and microenvironment of solid tumours. The aim of this doctoral research was to develop and characterise advanced three-dimensional (3D) in vitro models of HGSOC that more closely resemble ovarian tumour biology and improve the predictive value of preclinical drug testing. Four representative HGSOC cell lines (OVCAR-4, OVSAHO, COV362, and Kuramochi) were used to establish 3D spheroid models via ultra-low attachment culture and 3D bioprinting. These models were compared with 2D cultures through detailed morphological assessment, proliferation analysis, invasion assays, viability testing, gene expression profiling, immunocytochemistry, flow cytometry, and RNA sequencing. The findings demonstrate clear biological differences between 2D and 3D models. The 3D models developed compact spheroid structures with rim–core organization, reduced proliferation rates compared with 2D, and distinct epithelial–mesenchymal transition (EMT) and extracellular matrix (ECM) gene expression patterns. Importantly, carboplatin sensitivity was significantly lower in 3D models, reflected by higher IC₅₀ values, indicating increased resistance that more closely mirrors clinical observations. Integration with TCGA HGSOC transcriptomic data further supported the clinical relevance of the identified resistance-associated pathways. In addition, 3D bioprinting enabled the production of reproducible and scalable tumour models suitable for parallel drug testing. Overall, this work provides robust and biologically relevant 3D models for OC that bridge the gap between simplified cell culture systems and patient tumours.

• Jezik: Angleški jezik
• Ključne besede: ovarian cancer, 3D models, EMT, 3D bioprinting, chemoresistance
• Vrsta gradiva: Doktorsko delo/naloga
• Organizacija: BF - Biotehniška fakulteta
• Leto izida: 2026
• PID: 20.500.12556/RUL-184312
• Datum objave v RUL: 03.07.2026

Sekundarni jezik

• Jezik: Slovenski jezik
• Naslov: Razvoj novih in vitro 3D celičnih modelov raka jajčnika za testiranje zdravilnih učinkovin

Izvleček

Serozni rak jajčnikov visokega gradusa (HGSOC) je najagresivnejši in najsmrtonosnejši podtip raka jajčnikov (OC). Čeprav se bolnice na začetku dobro odzovejo na zdravljenje s platinastimi citostatiki, pri večini sčasoma pride do ponovitve bolezni zaradi česar se razvije kemorezistenca. Ena izmed ključnih omejitev pri raziskovanju teh mehanizmov je uporaba dvo-dimenzionalne (2D) celičnih kultur, ki ne odražajo kompleksne arhitekture in mikrookolja solidnih tumorjev. Namen doktorskega dela je bil razviti in karakterizirati napredne tri-dimenzionalne (3D) in vitro modele HGSOC, ki bolje posnemajo biologijo tumorja in izboljšujejo napovedno vrednost predkliničnega testiranja zdravil. Za vzpostavitev 3D sferoidnih modelov smo uporabili štiri reprezentativne celične linije HGSOC (OVCAR-4, OVSAHO, COV362 in Kuramochi), ki smo jih gojili v ploščah z ultra nizko adhezijo ali jih vzpostavili z uporabo 3D biotiskanja. Modele smo primerjali z ustreznimi 2D kulturami z natančno morfološko analizo, oceno proliferacije, invazijskimi testi, določitvijo viabilnosti, analizo izražanja genov, imunocitokemijo, pretočno citometrijo ter RNA sekvenciranjem. Rezultati so pokazali jasne biološke razlike med 2D in 3D modeli. 3D modeli so tvorili kompaktne sferoide z značilno organizacijo rob–jedro, nižjo stopnjo proliferacije v primerjavi z 2D modeli ter spremenjenimi vzorci izražanja genov, povezanih z EMT in ECM. Občutljivost na karboplatin je bila v 3D modelih značilno nižja, kar se je odražalo v višjih vrednostih IC₅₀ in povečani odpornosti, ki bolje odraža klinično stanje. Integracija rezultatov s transkriptomskimi podatki TCGA HGSOC je dodatno potrdila klinično relevantnost identificiranih poti, povezanih z rezistenco. 3D biotiskanje je omogočilo pripravo ponovljivih in skalabilnih tumorskih modelov, primernih za vzporedno testiranje učinkovin. Doktorsko delo predstavlja pomemben korak k vzpostavitvi biološko relevantnih 3D modelov OC, ki premoščajo vrzel med poenostavljenimi celičnimi sistemi in tumorji bolnic.

• Ključne besede: rak jajčnikov, 3D modeli, EMT, 3D biotiskanje, kemorezistenca