Comprehensive Characterization and Effective Combinatorial Targeting of Epithelial Ovarian Cancer and High-Grade Serous Ovarian Cancer via Single-cell Analysis.

Ovarian cancer kills more than 40 000 women in Europe and more than 150 000 women globally each year. The epithelial ovarian cancer (EOC) is the most common subtype and encompasses a collection of neoplasms with distinct clinical-pathological, molecular features and prognosis; among them the most common are the clear cells (CCOC), the mucinous (MOC), the endometrioid (ENOC), the low grade serous (LGSOC) and the high grade serous ovarian cancer (HGSOC). The HGSOC, in particular, is the most deadly form of EOC due to its high intratumor heterogeneity and lack of early diagnosis. The mechanical properties can be useful as new possible biomarkers of EOC subtypes in association to the classic ones used in clinical routine. In this thesis, the mechanical characterization of 9 ovarian cancer cell lines with different histological and morphological classification was carried out by AFM. Then, the achieved mechanical properties were evaluated on the same cell lines as markers of metastatic potential and drug sensitivity against 2C, a compound synthesized by the group of prof. Benedetti at the University of Trieste with anti-tumoral and cytoskeleton depolymerizing activity. Finally, an assessment of the effects of the culture medium composition on mechanical properties was performed. The mechanical characterization showed that the HGSOC cell lines had a high variability in the average Young’s modulus and resulted stiffer than the other cell lines with different histological classification. Moreover, the cell lines displayed two distinct single cell Young’s moduli distribution patterns: unimodal and bimodal. The cell lines with bimodal pattern showed two different populations with distinct mechanical behaviors. The invasion assay indicated a correlation between the stiffness decrease and the increase of invasion capacity. Accordingly, in cell lines with bimodal pattern only the “softer” population showed eventually the metastatic potential to invade. The cell lines with bimodal pattern were more resistant to 2C than the ones with unimodal pattern. For cell lines with bimodal pattern, the “stiffer” population had tendentially a higher resistance to 2C than the “softer” one. The F-actin network organization could influence the 2C resistance and the stiffness of cell lines: the HEY and OVCAR4 (high 2C resistance and high average Young’s modulus) had an Actin cytoskeleton more distributed over the cell than the TYKNU (low 2C resistance and low average Young’s modulus). Finally, variations in the culture medium components had an impact on the achieved Young’s moduli. This highlighted the need to develop optimized culture protocols for elasticity measurements, able to overcome the effects of different media on the mechanical properties.