(In)Distinctive Role of Long Non-Coding RNAs in Common and Rare Ovarian Cancers

Simple Summary:& nbsp;Ovarian cancers (OCs) are the most lethal form of gynecological tumors. The commonest are high-grade serous OCs, while rare OCs originate from many different cell types, such as epithelial, germ cell, sex cord-stromal, or mixed types. Rare OCs have distinct molecular characteristics, prognosis, and therapeutic approaches. However, all ovarian malignancies mostly share the same problem: late diagnosis due to the lack of specific symptoms. Therefore, there is a perpetual need to discover better diagnostic, prognostic, and predictive biomarkers, as well as new therapeutic approaches. In recent years, long non-coding RNAs (lncRNAs) have gained widespread attention because of their important role in various biological pathways. They have multiple mechanisms of action with an important role in many cellular processes related to OCs development and progression. This review will focus on the different aspects of lncRNAs in OCs and attempt to highlight the distinctive role of lncRNAs in common and rare OCs

Could MicroRNAs be Useful Tools to Improve the Diagnosis and Treatment of Rare Gynecological Cancers? A Brief Overview

Gynecological cancers pose an important public health issue, with a high incidence among women of all ages. Gynecological cancers such as malignant germ-cell tumors, sex-cord-stromal tumors, uterine sarcomas and carcinosarcomas, gestational trophoblastic neoplasia, vulvar carcinoma and melanoma of the female genital tract, are defined as rare with an annual incidence of <6 per 100,000 women. Rare gynecological cancers (RGCs) are associated with poor prognosis, and given the low incidence of each entity, there is the risk of delayed diagnosis due to clinical inexperience and limited therapeutic options. There has been a growing interest in the field of microRNAs (miRNAs), a class of small non-coding RNAs of 22 nucleotides in length, because of their potential to regulate diverse biological processes. miRNAs usually induce mRNA degradation and translational repression by interacting with the 30 untranslated region (30-UTR) of target mRNAs, as well as other regions and gene promoters, as well as activating translation or regulating transcription under certain conditions. Recent research has revealed the enormous promise of miRNAs for improving the diagnosis, therapy and prognosis of all major gynecological cancers. However, to date, only a few studies have been performed on RGCs. In this review, we summarize the data currently available regarding RGCs.peer-reviewe

Effects of cyclopamine (A) and tamoxifen (B) on Hh-Gli pathway gene expression in MCF-7 and SkBr-3 cells.

<p>The Hh-Gli pathway is upregulated after short-term combined treatment in MCF-7, but the effect is negated after longer treatment. On the Western blot image, band quantification relative to actin and non-treated cells is denoted below the bands. (C). The effect of combined treatment on SkBr-3 cell line is weak (D). Gene expression levels are shown on graph as relative fold change relative to non-treated conditions with reference value 1 pointed out with emboldened bar. Only combined cyclopamine and tamoxifen treatment induces migration in MCF-7 cells. Representative images of the wound healing assay at 0 and 26 h (after processing with TScratch software <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0114510#pone.0114510-Gebck1" target="_blank">[23]</a>) are shown for non-treated conditions (NT; N = 16), cyclopamine treatment (CYC; N = 16), tamoxifen treatment (TAM; N = 14) and combined treatment with cyclopamine and tamoxifen (C+T; N = 12) (E). Quantitative analysis of the percentage of open wound areas is shown on the graph, (*) P<0.05 (F). Transwell migration assay confirmed increased migration capacity of cells after combined cyclopamine and tamoxifen treatment. Representative images of migrated cells after 48 h are shown for non-treated conditions (NT; N = 15), cyclopamine treatment (CYC; N = 15), tamoxifen treatment (TAM; N = 15) and combined treatment (C+T; N = 15) (G). Quantitative analysis of the relative number of migrated cells (analyzed relative to non-treated cells) is shown on graph, (*) P<0.0001 (H).</p

Differentially expressed proteins in MCF-7 cells treated with cyclopamine and tamoxifen compared with non-treated control cells.

<p>General Functions are obtained from the UniProt and NCBI Gene databases. Protein numbers correspond to the numbers marked on the 2-D gels (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0114510#pone.0114510.s001" target="_blank">Figure S1</a>). Numbers in the table correspond to spot numbers denoted on the 2-D gel images; missing numbers in the table are unidentified proteins or proteins with score less than 39.</p><p>Differentially expressed proteins in MCF-7 cells treated with cyclopamine and tamoxifen compared with non-treated control cells.</p

Effect of cyclopamine and tamoxifen combination on MCF-7 cell proliferation.

<p>When tamoxifen is in higher concentrations, and cyclopamine in lower concentrations, MCF-7 cell viability is decreased. However, when cyclopamine concentration is increased (with tamoxifen concentration remaining constant) cell viability increases (A). Similar effect can be seen vice-versa, when cyclopamine concentration is constant and tamoxifen concentration is increased (B) as measured by MTT assay after 48 h.</p

Gene and protein expression levels after transfection with <i>GLI1</i> (GLI1) and additional stimulation with Shh protein (GLI1+SHH).

<p><i>ERα</i> gene expression increases in MCF-7 cells only after additional Shh stimulation (A) while <i>ERα</i> gene expression does not change in SkBr-3 cells (B). Gli1, Ptch1 and ERα protein levels in MCF-7 cells after <i>GLI1</i> transfection and additional Shh stimulation (C). Protein bands were quantified and normalized relative to actin and non-treated conditions and the relative values are denoted below each band. Relative gene expression of <i>PTCH1</i> (D, E) and <i>ERα</i> (F,G) after silencing of <i>PTCH1</i> gene in MCF-7 and SkBr-3 cell line. Efficient silencing (<30% of residual expression) was achieved 24 h post-transfection in MCF-7 cell line, and 48 h post-transfection in SkBr-3 cell line.</p

Cell viability after tamoxifen (A,B), cyclopamine (C,D) or combined treatment (E,F) in MCF-7 and SkBr-3 cell lines.

<p>Tamoxifen and cyclopamine each inhibit proliferation of MCF-7 cells in a dose dependent manner (A,C). When administered simultaneously, they cause a short term survival effect in MCF-7 cells (C+T 48 h) – pointed out with arrow, whereas long term simultaneous treatment induces strong cell death in these cells (C+T 96 h). Combination treatment of cyclopamine for 48 h followed by tamoxifen for 48 h (C 48 h →T 48 h) or vice versa (T 48 h →C 48 h) showed an effect similar to tamoxifen alone (E). Tamoxifen and cyclopamine show only a mild inhibitory effect on SkBr-3 cell proliferation at longest exposures (B,D) while combined treatment has no pronounced effect (F).</p

Immunofluorescent staining of MCF-7 cell line in non-treated cells (NT) and treated with Shh protein detected by confocal microscopy.

<p>ERα is stained green (column 1), Shh is stained red (column 2), nuclei are stained blue with DAPI (column 3), and the last column shows the overlay of signals. Yellow staining shows areas of green and red signal co-localization (A). Shh-treated cells show significantly decreased nuclear staining and increased co-localization of ERα and Shh compared to non-treated cells, as determined by ImageJ software, (*) P<0.05. (B). Shh protein co-immunoprecipitates with ERα protein in MCF-7 cells, both in non-treated conditions and after treatment with exogenous Shh protein for 48 h; NT = non-treated, neg.ctrl. = negative control. Western blot of input proteins is provided as control for presence of the proteins in cell lysates (C).</p

Effect of stimulation with Shh protein on pathway activity in MCF-7 (A,C) and SkBr-3 cells (B).

<p>Gene expression levels are shown on graph as relative fold change relative to non-treated conditions with reference value 1 pointed out with emboldened bar. Relative gene expression of <i>ERα</i> after treatment with Shh protein (D,E). Non-treated cells (NT) have a relative value 1. ERα protein expression in MCF-7 cells increases after treatment with Shh protein for 48 h (F) Protein bands were quantified and normalized relative to actin and non-treated conditions and the relative values are denoted below each band.</p