Effect of ovarian cancer ascites on SKOV-3 cells proteome: new proteins associated with aggressive phenotype in epithelial ovarian cancer

Abstract

BackgroundEpithelial ovarian cancer is the second most lethal gynecological cancer worldwide. Ascites can be found in all clinical stages, however in advanced disease stages IIIC and IV it is more frequent and could be massive, associated with worse prognosis. Due to the above, it was our interest to understanding how the ascites of ovarian cancer patients induces the mechanisms by which the cells present in it acquire a more aggressive phenotype and to know new proteins associated to this process.MethodsA proteomic analysis of SKOV-3 cells treated with five different EOC ascites was performed by two-dimensional electrophoresis coupled to MALDI-TOF. The level of expression of the proteins of interest was validated by RT-PCR because several of these proteins have only been reported at the messenger level.ResultsAmong the proteins identified that increased their expression in ascites-treated SKOV-3 cells, were Ran GTPase, ZNF268, and Synaptotagmin like-3. On the other hand, proteins that were negatively regulated by ascites were HLA-I, HSPB1, ARF1, Synaptotagmin 1, and hnRNPH1, among others. Furthermore, an interactome for every one of these proteins was done in order to identify biological processes, molecular actions, and cellular components in which they may participate.ConclusionsIdentified proteins participate in cellular processes highly relevant to the aggressive phenotype such as nuclear transport, regulation of gene expression, vesicular trafficking, evasion of the immune response, invasion, metastasis, and in resistance to chemotherapy. These proteins may represent a source of information which has the potential to be evaluated for the design of therapies directed against these malignant cells that reside on ovarian cancer ascites.

Ascites from Ovarian Cancer Induces Novel Fucosylated Proteins

“…On the other hand, ascites-treated cells showed a much more accentuated spindleshaped morphology (Fig. 2, ascites, light microscopy), as has been previously reported [24]. These morphological changes were observed with the 13 different ascites tested in this work.…”

“…To confirm this hypothesis, it will be necessary to determine the IL-6 levels in our ascites samples. As reported by Toledo-Leyva et al (2018) [24], the presence of ascites changes the phenotype of SKOV-3 cells, and this acquired phenotype (mesenchymal) is associated with increased invasion and migration properties [46]. Fucosylation is also modified, and this was demonstrated in this work, using cell-free ascites; therefore, we can affirm that the alterations observed in the fucosylation patterns are related to the soluble factors present in ascites and not to overgrowth of ascites tumor cells.…”

“…The SKOV-3 (ATCC HTB-77) and OVCAR3 (ATCC HTB 161) cell lines were used. The SKOV-3 model has been used previously in our laboratory as described [24]. Briefly, SKOV-3 cells were cultured in McCoy's 5A (Corning, 10-050-CVR) culture medium (15 mL) supplemented with 10% fetal bovine serum (Corning, 35-010-CV) and 1% penicillin/streptomycin (PAA, P11-010) at 37°C and 5% CO 2 .…”

“…The samples were processed as described previously [24] with some modifications. Briefly, peptide digestion was performed using mass grade trypsin (Sigma-Aldrich, T 6567).…”

“…It has been confirmed that ascites can change the cellular phenotype, and the ascites compounds that are produced by or in response to malignant cells may be a good alternative to the microenvironment in which malignant cells are immersed [24]; these components could participate in the altered regulation of different processes, including fucosylation. For this reason, the goal of our work was to determine the differentially fucosylated proteins in ovarian cancer cell lines cultured with ascites stimulation.…”