Breast Cancer and the Other Non-Coding RNAs.

Breast cancer is very heterogenous and the most common gynaecological cancer, with various factors affecting its development. While its impact on human lives and national health budgets is still rising in almost all global areas, many molecular mechanisms affecting its onset and development remain unclear. Conventional treatments still prove inadequate in some aspects, and appropriate molecular therapeutic targets are required for improved outcomes. Recent scientific interest has therefore focused on the non-coding RNAs roles in tumour development and their potential as therapeutic targets. These RNAs comprise the majority of the human transcript and their broad action mechanisms range from gene silencing to chromatin remodelling. Many non-coding RNAs also have altered expression in breast cancer cell lines and tissues, and this is often connected with increased proliferation, a degraded extracellular environment, and higher endothelial to mesenchymal transition. Herein, we summarise the known abnormalities in the function and expression of long non-coding RNAs, Piwi interacting RNAs, small nucleolar RNAs and small nuclear RNAs in breast cancer, and how these abnormalities affect the development of this deadly disease. Finally, the use of RNA interference to suppress breast cancer growth is summarised.

Colposcopic scoring indexes in the evaluation of cervical lesions with the cytological result of atypical squamous cells, cannot exclude high-grade lesion.

AIM: Colposcopic indexes including Reid index and Swede score were developed to make the colposcopy more objective. The aim of our study was to evaluate the significance of colposcopic indexes in the evaluation of ASC-H cervical lesions. METHODS: We carried out a cross-sectional study in the Clinic of Obstetrics and Gynecology between January 2013 and December 2018. The study included 535 women, from which 66 women had a cytological result ASC-H. Scoring of all colposcopic findings was assessed according to Reid modified index and Swede score and a composite score was determined. Frequency distributions were compared using χ2 /Fisher exact test. Spearman rank correlation coefficient was computed between RCI and Swede score. RESULTS: Sensitivity, specificity, positive and negative predictive value and positive likelihood ratio of modified Reid colposcopic index at a cutoff of ≥4 for the detection of HSIL+ lesions were: 86.11% (95% CI: 70.5-95.3), 83.33% (95% CI: 65.3-94.4), 86.11% (95% CI: 69.7-94.8), 83.33% (95% CI: 64.5-93.7) and 5.17 (95% CI: 2.3-11.6). Swede score with the cutoff value ≥5 showed comparable results to modified Reid index with the increased sensitivity: 94.44% (95% CI: 81.3-99.3). CONCLUSION: ASC-H category represents the trickiest cytological diagnosis as it is underlined with the high risk of severe cervical dysplasia. Evaluating the cervical lesion by the use of colposcopic indices helps the gynecologist to objectively evaluate all the pathologies of uterine cervix. Swede score with the cutoff value 8 also enables a 'see and treat' option in management of atypical squamous cells, cannot exclude high-grade lesions.

miRNA in a multiomic context for diagnosis, treatment monitoring and personalized management of metastatic breast cancer.

Metastatic breast cancer is characterized by aggressive spreading to distant organs. Despite huge multilevel research, there are still several important challenges that have to be clarified in the management of this disease. Therefore, recent investigations have implemented a modern, multiomic approach with the aim of identifying specific biomarkers for not only early detection but also to predict treatment responses and metastatic spread. Specific attention is paid to short miRNAs, which regulate gene expression at the post-transcriptional level. Aberrant miRNA expression could initiate cancer development, cell proliferation, invasion, migration, metastatic spread or drug resistance. An miRNA signature is, therefore, believed to be a promising biomarker and prediction tool that could be utilized in all phases of carcinogenesis. This article offers comprehensive information about miRNA profiles useful for diagnostic and treatment purposes that may sufficiently advance breast cancer management and improve individual outcomes in the near future.

DNA methylation and detection of cervical cancer and precancerous lesions using molecular methods.

Cervical cancer is the third most common cancer disease affecting the female population, and a key factor in the development of the disease is the human papillomavirus infection (HPV). The disease is also impacted by epigenetic changes such as DNA methylation, which causes activation or exclusion of certain genes. The aim of our review is to summarize and compare the most common molecular methods for detection of methylated promoter regions in biomarkers occurring in cervical carcinoma and also show the importance of connections of HR-HPV testing with methylation analysis in patients with cervical intraepithelial neoplasia. Insight into genetic and epigenetic alterations associated with cervical cancer development can offer opportunities for the molecular biomarkers that can be useful for screening, diagnosis, and also as new ways of treatment of cervical cancer precursor lesions.

New molecular aspects of endometrial carcinoma.

Endometrial cancer is the most commonly diagnosed gynecological cancer and its incidence is increasing worldwide. The number of patients with this disease is likely to continue to grow, including younger patients. It is a complex disease driven by abnormal genetic and epigenetic alterations, as well as environmental factors. Many endometrial cancers show estrogen-dependent proliferation. The carcinogenic mechanisms are unknown or not completely explained beyond mutations of single oncogenes and tumor suppressor genes. Possible carcinogenic mechanisms include imbalance between endometrial proliferation by unopposed estrogen and the mismatch repair (MMR) system; rmethylation changes and mutation of genes. Epigenetic changes resulting in aberrant gene expression are dynamic and modifiable features of many cancer types. A significant epigenetic change is aberrant DNA methylation. In this review, we review evidence on the role of different changes in relation to endometrial carcinogenesis. Carcinogenic mechanisms of endometrial cancer involve both genetic and epigenetic changes. Determination of the detailed carcinogenic mechanisms will be useful for prevention and diagnosis of endometrial cancer, risk assessment, and development of new treatment strategies targeting genes.

Detection of methylation of the promoter region of the MAL and CADM1 genes by pyrosequencing in cervical carcinoma.

OBJECTIVE: Cervical cancer is the second most common cancer disease affecting the female population. A key factor in development of the disease is the human papillomavirus infection (HPV). The disease is also impacted by epigenetic changes such as DNA methylation, which causes activation or exclusion of certain genes, and simultaneously the hypermethylation of cytosines in the promoters and turn-off of previously active genes occur. In this study, we focused on the introduction of pyrosequencing for the detection of DNA methylation of the selected CADM1 and MAL genes. METHODS: DNA was isolated from cytological cervical smear of patients with different types of dysplasia [L-SIL (n=14), ASC-US (n=15), H-SIL (n=1)] and four control samples from healthy women. Prepared samples were further analyzed by bisulfite conversion and subsequent pyrosequencing (Pyromark Q96 ID, Qiagen, Germany). We examined the extent of methylation of CpG islands and as control samples of this method we used a fully methylated and unmethylated DNA. Methylation level (Met level) from each sample was quantified as the mean value [sum of all methylated CpG islands in %/total number of CpG islands (MAL n=4; CADM1 n=3)]. RESULTS: In total, 30 clinical samples and 4 control samples from healthy women were analyzed. By means of the analysis of the CADM1promoter region, the values of the Met level were obtained [fully methylated DNA (94.83 and 88); completely unmethylated DNA (0 and 0); and control samples from healthy patients (6.825 and 0.825), L-SIL (2.107 and 2.778), ASC-US (7.313 and 3.626), H-SIL (0 and 0)]. By means of the analysis of the MAL promoter region, the values of Met level were obtained [fully methylated DNA (53.25); completely unmethylated DNA (0.875); and control samples from healthy patients (2.925), L-SIL (1.517), ASC-US (2.833), and H-SIL (4)]. CONCLUSION: We introduced a pyrosequencing method for quantification of methylation of CADM1, MAL promoter regions, and detected methylations in clinical samples and also some basal methylation in healthy women.

Association of Circulating miRNA Expression with Preeclampsia, Its Onset, and Severity.

MicroRNAs (miRNAs) are one of the important regulators of cellular functions fundamental for healthy pregnancy processes, including angiogenesis and differentiation of trophoblast cells, and their deregulation could be implicated in the pathogenesis of pregnancy complications, including preeclampsia (PE). The aim of this study was to assess the association of miRNA expression in plasma samples with PE, its onset, and severity. Our study enrolled 59 pregnant women, 27 in the preeclamptic study group and 32 in the control group with physiological pregnancy. Preeclamptic pregnancies were divided into subgroups based on the severity and onset of disease. Relative expression of miR-21-5p, miR-155-5p, miR-210-5p, miR-16-5p, and miR-650 isolated from plasma samples was analysed by quantitative real-time PCR and normalised to experimentally established reference genes. Our results revealed upregulation of miR-21-5p (1.16-fold change, p = 0.0015), miR-155-5p (1.62-fold change, p = 0.0005) in preeclamptic pregnancies, compared to controls. Overexpression of these two miRNAs was observed, especially in subgroups of severe and late-onset PE compared to healthy pregnancies. Although we hypothesised that the expression level of studied miRNAs could vary between PE subtypes (mild vs. severe, early onset vs. late-onset), no obvious differences were detected. In conclusion, our study could contribute to the large-scale studies for the identification of non-invasive biomarkers for PE detection to improve outcomes for women and their new-borns.

Mullerian adenosarcomas of the uterine cervix with sarcomatous overgrowth.

Mullerian adenosarcoma with sarcomatous overgrowth (MASO) of the uterine cervix is an extremely rare variant of adenosarcoma of the genital tract associated with aggressive clinical course. We searched the PubMed and Medline databases for MASO of the cervix and we identified and reviewed eleven cases published between years 2004 and 2017. The most common clinical picture includes abnormal vaginal bleeding, postcoital bleeding, pelvic pain and foul-smelling vaginal discharge. Therapeutic options for MASO are still undefined. Radical hysterectomy with sufficient tumour-free margins combined with adjuvant chemotherapy and radiotherapy should serve as an effective treatment tool with favourable outcome.

Different methylation levels in the KLF4, ATF3 and DLEC1 genes in the myometrium and in corpus uteri mesenchymal tumours as assessed by MS-HRM.

Mesenchymal tumours of the corpus uteri comprise common benign lesions - leiomyomas and very rare malignant variants - sarcomas. It can be difficult to distinguish between the particular types of mesenchymal tumours pre-surgically. Primarily, leiomyomas and the very aggressive leiomyosarcomas can be easily misdiagnosed when using only imaging devices. Therefore, a reliable non-invasive marker for these tumour types would provide greater certitude for patients that the lesion remains benign. Our collection comprises 76 native leiomyomas, an equal number of healthy myometrium samples and 49 FFPE samples of various types of sarcomas. The methylation level was assessed by MS-HRM method and we observed differences in the methylation level between healthy, benign and (semi)malignant tissues in the KLF4 and DLEC1 genes. The mean methylation levels of leiomyomas compared to myometrium and leiomyosarcomas were 70.7% vs. 6.5% vs. 39.6 % (KLF4) and 66.1% vs. 14.08% vs. 37.5% (DLEC1). The ATF3 gene was differentially methylated in leiomyomatous and myometrial tissues with 98.1% compared to 76.6%. The AUC values of the predictive logistic regression model for discrimination between leiomyomas and leiomyosarcomas based on methylation levels were 0.7829 (KLF4) and 0.7719 (DLEC1). Finally, our results suggest that there should be distinct models for the methylation events in benign leiomyomas and sarcomas, and that the KLF4 and DLEC1 genes can be considered potential methylation biomarkers for uterine leiomyomas.

Breast Cancer in Young Women: Status Quo and Advanced Disease Management by a Predictive, Preventive, and Personalized Approach.

Why does healthcare of breast cancer (BC) patients, especially in a young population, matter and why are innovative strategies by predictive, preventive, and personalized medicine (PPPM) strongly recommended to replace current reactive medical approach in BC management? Permanent increase in annual numbers of new BC cases with particularly quick growth of premenopausal BC patients, an absence of clearly described risk factors for those patients, as well as established screening tools and programs represent important reasons to focus on BC in young women. Moreover, "young" BC cases are frequently "asymptomatic", difficult to diagnose, and to treat effectively on time. The objective of this article is to update the knowledge on BC in young females, its unique molecular signature, newest concepts in diagnostics and therapy, and to highlight the concepts of predictive, preventive, and personalized medicine with a well-acknowledged potential to advance the overall disease management.

Bilateral ovarian angiosarcoma arising from the mature cystic teratomas - A case report and review of the literature.

INTRODUCTION: Ovarian teratomas undergo the malignant transformation in 0.2-2% of cases. The behavior of malignancies in mature cystic teratomas (MCT) is determined by their phenotype and not their derivation from germ cells. We can recognize pure angiosarcomas or as a part of other tumors like malignant mixed Mullerian tumors and adenosarcomas. PRESENTATION OF CASE: We present the first case of bilateral ovarian angiosarcoma arising from the mature teratomas. Due to widespread disease, we performed limited surgical procedure consisting of bilateral adnexectomy and omentectomy. Exploratory laparotomy in 44-year old patient showed massive ascites, necrotic tissue of omentum and bilateral tumors originating from both ovaries measuring 8 and 6cm with necrotic surface. Immunohistochemistry of the tumors showed positive staining for CD31, vimentin, desmin and focal positivity for CD34. DISCUSSION: Sarcomas of gynecologic origin are extremely rare tumors. They present with unspecified symptoms and are diagnosed in late stages of the disease. The appropriate management of angiosarcomas is difficult due to the rarity of disease and late stage of the diseases. Surgical therapy should contain the hysterectomy with bilateral salpingo-oophorectomy and omentectomy. Pelvic lymphadenectomy was not performed in published cases with no effect on patient survival. CONCLUSION: This work summarizes the current knowledge in the diagnosis and treatment of angiosarcomas arising in the mature teratomas. Promising results are expected from the trials devoted to antiangiogenic strategies in treatment of aggressive sarcomas.

Methylation status of KLF4 and HS3ST2 genes as predictors of endometrial cancer and hyperplastic endometrial lesions.

Endometrial carcinoma is one of the most common tumours in developed countries. In addition to the active role of genetic factors, epigenetic changes also have an important effect. The present study analysed the methylation status of kruppel like factor 4 (KLF4) and heparan sulfate­glucosamine 3­sulfotransferase 2 (HS3ST2) genes in three endometrial tissue types for carcinoma prediction. The sample comprised 91 women with histologically­confirmed endometrial carcinoma (64.16±9.64 years old), 36 women with hyperplasia (53.39±9.64 years old) and 45 with no signs or symptoms of malignancy (48.53±11.11 years old). The CpG dinucleotide methylation levels were examined by quantitative pyrosequencing, and the discrimination accuracy of the model was calculated using the Random Forest classification algorithm of the area under the ROC curve (AUC). The mean values of KLF4 and HS3ST2 methylation indices were 23.83±11.39 and 8.52±2.57 in the control samples; 30.40±8.52 and 33.76±20.66 in hyperplasia and 34.72±10.79 and 34.49±18.39 in the cancerous tissues. Multinomial logistic regression indicated that the HS3ST2 CpG1 methylation status is a predictor of hyperplasia (P<0.05) and that the KLF4 CpG2 dinucleotide can predict carcinoma formation (P<0.001). The AUC value of 0.95 indicates high discrimination accuracy of the CpG nucleotides methylation status model between the controls and the two other diagnoses. The results of the present study establish the likelihood that aberrations in KLF4 and HS3ST2 gene methylation levels are important in the development of endometrial hyperplasia and carcinoma, with hyperplasia an intermediate step between healthy and tumour tissues.

Methylation of CADM1 and MAL together with HPV status in cytological cervical specimens serves an important role in the progression of cervical intraepithelial neoplasia.

Cervical cancer (CC) is the second most common type of cancer affecting the female population. The development of CC takes several years, and involves a precancerous stage known as cervical intraepithelial neoplasia (CIN). A key factor in the development of disease is the human papillomavirus (HPV) infection, which initiates carcinogenesis. Furthermore, CC is also impacted by epigenetic changes such as DNA methylation, which causes activation or exclusion of certain genes, and the hypermethylation of cytosines in promoters, thereby switching off previously active genes. The majority of DNA methylation events occur at cytosine-guanine nucleotides, which in the human genome are known as CpG islands. The aim of the present study was to investigate the methylation levels in intronic sequences of the two tumor suppressor genes cell adhesion molecule 1 (CADM1) and T-lymphocyte maturation associated protein (MAL) using cytological samples and to identify potential biomarkers involved in CIN by pyrosequencing. DNA was isolated from cervical smears from patients with CINs, with healthy patients serving as a control group. Samples were converted by treatment with sodium bisulfite and subsequent pyrosequencing to detect the methylation status of the selected genes. The presence of HPV DNA infection analyzed by the polymerase chain reaction, was detected in each sample. Of the total number of samples (n=91), the present study confirmed the presence of one or two high-risk subtypes of HPV in 39 cases (42.85%) and HPV infection was significantly associated with CIN2+ lesions. For the two genes (MAL and CADM1) the present study confirmed that the median methylation was significantly higher in HPV positive patients [P=0.0097, 95% confidence interval (CI): (-0.030, -0.003)/P=0.0024, 95% CI: (-0.06, -0.01)] when compared with patients negative for HPV DNA infection, and the average methylation was demonstrated to be increased with the degree of cervical lesion. The present study used logistical regression to model the dependence between the case/control statuses (control group vs. Dg. 1-4). The area under the curve values for MAL were: 84% for cervical inflammation, 71% for CIN1, 73.4% for CIN2+ and 77% for squamous cell carcinoma (SCC); and for CADM1 were: 88.6% for cervical inflammation, 68% for CIN1, 80% for CIN2+ and 89% for SCC. The present study confirmed that there were statistically significant differences between the methylation levels of individual CpGs and significantly higher median methylation in patients positive for HPV16/18. CADM1 exhibited higher levels of methylation in almost every study group when compared with MAL during the transition of CIN and appeared to be a promising biomarker for future study.

The examination of superior mesenteric artery circulation in fetus during pregnancy.

OBJECTIVE: The aim of the study was a comprehensive examination of the circulation of superior mesenteric artery in different weeks of pregnancy using Doppler ultrasound examination. METHODS: The study was conducted at the Clinic of Obstetrics and Gynecology, Jessenius Faculty of Medicine, Comenius University, Slovakia between the years 2008 and 2012. Dynamics and changes in superior mesenteric artery circulation were studied in 856 women. Results of Doppler examination were processed and subsequently evaluated for each gestational week separately. RESULTS: We found that changes in resistance index of superior mesenteric artery are slower and have a tendency to decrease as a consequence of a gradual decrease in vascular resistance. The changes in pulsatility index are different in nature; there is a tendency for there are to be repeating periods of higher and then lower values, which are subject to a more pronounced dependence on flow rate, especially end diastolic velocity, and Vmean. CONCLUSIONS: This article and its results bring a new, comprehensive view not only of physiological changes in the splanchnic circulation, but also of changes that may be subject to a certain extent, not only to the sex, but primarily to the weight of the fetus, which, of course, indirectly reflects its metabolic and respiratory demands.