# Comparative Analysis of Primary Ovarian Cancer Cells and Established Cell Lines as a New Tool for Studies on Ovarian Cancer Cell Complexity

## TL;DR

This study compares primary ovarian cancer cells with immortalized versions to understand cancer cell complexity and develop better research tools.

## Contribution

The study introduces new immortalized ovarian cancer cell lines with distinct stem cell-like features for research and preclinical modeling.

## Key findings

- Immortalized OvCa3 A hTERT cells retained a mesenchymal stem cell phenotype with CD73/CD90/CD105 positivity.
- OvCa7 A hTERT cells showed CD133 expression in 90% of cells, indicating a cancer stem cell (CSC) phenotype.
- Both cell lines formed spheroids and exhibited differences in gene expression and metabolic/migratory activity.

## Abstract

Primary cancer cells reflect the genetic background and phenotype of a tumor. Immortalized cells with higher proliferation activity have an advantage over primary cells. The aim of the study was to immortalize the primary ovarian cancer (OvCa) cells using the plasmid-carrying human telomerase reverse transcriptase (hTERT) gene and compare their phenotype and biological activity with the primary cells. The primary OvCa3 A and OvCa7 A cells were isolated from the ascitic fluid of two high-grade serous ovarian cancer patients and were characterized using immunocytochemical methods, flow cytometry, real-time RT-PCR, Western blot, metabolic activity, and migratory potential. Both immortalized ovarian cancer cell lines mirrored the phenotype of primary cancer cells, albeit with modifications. The OvCa3 A hTERT cells kept the mesenchymal stem cell phenotype of CD73/CD90/CD105-positivity and were CD133-negative, whereas the cell population of OvCa7 A hTERT lost CD73 expression, but almost 90% of cells expressed the CD133 characteristic for the CSCs phenotype. Immortalized OvCa cells differed in gene expression level with respect to Sox2 and Oct4, which was associated with stemness properties. The OvCa7 A hTERT cells showed higher metabolic and migratory activity and ALDH1 expression than the corresponding primary OvCa cells. Both primary and immortalized cell lines were able to form spheroids. The newly established unique immortalized cell line OvCa7 A hTERT, with the characteristic of a serous ovarian cancer malignancy feature, and with the accumulation of the p53, Pax8, and overexpression of the CD133 and CD44 molecules, may be a useful tool for research on therapeutic approaches, especially those targeting CSCs in ovarian cancer and in preclinical 2D and 3D models.