Abstract
With the onset of resistance, ovarian cancer cells display almost unpredictable adaptive potential. This may derive from the tumor genetic ancestry and can be additionally tailored by post translational protein modifications (PTMs). In this study, we took advantage of high-end (phospho)-proteome analysis combined with multiparametric morphometric profiling in high-grade serous (OVCAR-3) and non-serous (SKOV-3) ovarian carcinoma cells. For functional experiments, we applied two different protocols, representing typical conditions of the abdominal cavity and of the growing tumor tissue: on the one side hypoxia (oxygen 1%) which develops within the tumor mass or is experienced during migration/extravasation in non-vascularized areas. On the other hand, fluid shear stress (250 rpm, 2.8 dyn/cm2) which affects tumor surface in the peritoneum or metastases in the bloodstream. After 3 hours incubation, treatment groups were clearly distinguishable by PCA analysis. Whereas basal proteome profiles of OVCAR-3 and SKOV-3 cells appeared almost unchanged, phosphoproteome analysis revealed multiple regulatory events. These affected primarily cellular structure and proliferative potential and consolidated in the proteome signature after 24h treatment. Upon oxygen reduction, metabolism switched toward glycolysis (e.g. upregulation hexokinase-2; HK2) and cell size increased, in concerted regulation of pathways related to Rho-GTPases and/or cytoskeletal elements, resembling a vasculogenic mimicry response. Shear stress regulated proteins governing cell cycle and structure, as well as the lipid metabolism machinery including the delta(14)-sterol reductase, kinesin-like proteins (KIF-22/20A) and the actin-related protein 2/3 complex. Independent microscopy-based validation experiments confirmed cell-type specific morphometric responses. In conclusion, we established a robust workflow enabling the description of the adaptive potential of ovarian cancer cells to physical and chemical stressors typical for the abdominal cavity and supporting the identification of novel molecular mechanisms sustaining tumor plasticity and pharmacologic resistance.
Highlights
Ovarian cancers develop in rather aggressive phenotypes which reflects in the highest fatality rate among gynecological tumors [1]
Characterization of proteome signatures of SKOV-3 and OVCAR3 confirmed that the clones of our laboratories displayed the typical features representing their pathological model
Cultivated ovarian cancer cells SKOV-3 and OVCAR-3 differ clearly in structure and proteome (Figure 1). This reflects in vitro some crucial features of their pathological behavior in vivo: SKOV-3 are classified as non-serous ovarian cancer cells and are characterized by high motility [24]
Summary
Ovarian cancers develop in rather aggressive phenotypes which reflects in the highest fatality rate among gynecological tumors [1]. It is clear that physical cues can contribute to the regulation of multiple pathways, including the cellular metabolic state, such as glycolysis [9, 10] and lipid metabolism [11] These aspects are relevant for ovarian cancer [12] the development in the peritoneal cavity necessarily requires endurance to physical stressors. Physical stimuli may support the progression of the disease: even without colonizing other tissues, ovarian cancer can spread broadly in the abdominal cavity [13, 14]: this is eased by the formation of spheroids that are growing and disseminating with the ascitic fluids [15] In this respect, recent studies correlated ovarian cancer cell morphology/survival to the capability to cope with biomechanical stimulation [16,17,18]. Literature is still limited and the elucidation of the molecular mechanisms influencing these responses is far from being complete
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