Deregulation of Selected MicroRNAs in Sinonasal Squamous Cell Carcinoma: Searching for Potential Prognostic Biomarkers.

Sinonasal carcinomas are head and neck tumours arising from the nasal cavity and paranasal sinuses characterized by unfavourable outcome, difficult treatment, diagnosis and prognosis. MicroRNAs are key molecules in the regulation of development and progression of cancer and their expression profiles could be used as prognostic biomarkers, to predict the patients' survival and response to treatment. In this study, we used quantitative real­time PCR with TaqMan® Advanced miRNA Assays to investigate the relative expression values of selected micro- RNAs in a unique set of formalin-fixed paraffin-embedded tissue samples obtained from 46 patients with sinonasal squamous cell carcinoma. Our results showed statistically significant up-regulation of three mature microRNAs: miR-9-5p (fold change: 6.80), miR-9-3p (fold change: 3.07) and let-7d (fold change: 3.93) in sinonasal carcinoma patients. Kaplan-Meier survival analysis and logrank test identified association between higher expression of miR-9-5p and longer survival of the patients (P = 0.0264). Lower expression of let-7d was detected in the patients with impaired survival, and higher expression of miR-137 was linked to shorter survival of the patients. We also identified several correlations between expression of the studied microRNAs and recorded clinicopathological data. Higher expression of miR-137 and lower expression of let-7d correlated with local recurrence (P = 0.045 and P = 0.025); lower expression of miR-9-5p and higher expression of miR-155-5p correlated with regional recurrence (P = 0.045 and P = 0.036). Higher expression of miR-9-3p correlated with occupational risk (P = 0.031), presence of vascular invasion (P = 0.013) and perineural invasion (P = 0.031). Higher expression of miR-155-5p was present in the samples originating from maxillary sinus (P = 0.011), cN1-3 classified tumours (P = 0.009) and G2-3 classified tumours (P = 0.017). In conclusion, our study supports the hypothesis of future prospect to use expression of miRNAs as prognostic biomarkers of squamous cell sinonasal carcinoma. In particular, miR-9-5p and miR-9-3p seem to be important members of the sinonasal cancer pathogenesis.

Gynecological and urological aspects of pelvic vasculitis.

OBJECTIVE: To review contemporary knowledge of isolated vasculitis in urology and gynaecology. DESIGN: A review. SETTING: Department of Obstetrics and Gynaecology, University Hospital Hradec Kralove. METHODS: To present own experience and an overview of recent literature. CONCLUSION: Vascular system inflammation is a very important and broadly studied medical condition. It can affect either veins or arteries. In pelvic veins it can significantly increase the risk of thromboembolic complications, while in the case of arteries; the clinical significance is still unknown. We still do not know what does the histopathological proven isolated vasculitis in this area mean. Is it just a local finding, or should we look for systemic vasculitis? Unlike most of non-symptomatic gynecological vasculitis, urological cases are often accompanied by severe symptoms depending on the anatomical location of the process. This work presents a basic overview and includes our experience with this issue. Our thesis does not include the vasculitis in pregnancy.

Analysis of DNA methylation and microRNA expression in NUT (nuclear protein in testis) midline carcinoma of the sinonasal tract: a clinicopathological, immunohistochemical and molecular genetic study.

The aim of this study was a detailed clinicopathological investigation of sinonasal NUT midline carcinoma (NMC), including analysis of DNA methylation and microRNA (miRNA) expression. Three (5%) cases of NMC were detected among 56 sinonasal carcinomas using immunohistochemical screening and confirmed by fluorescence in situ hybridization. The series comprised 2 males and 1 female, aged 46, 60, and 65 years. Two tumors arose in the nasal cavity and one in the maxillary sinus. The neoplasms were staged pT1, pT3, and pT4a (all cN0M0). All patients were treated by radical resection with adjuvant radiotherapy. Two patients died 3 and 8 months after operation, but one patient (pT1 stage; R0 resection) experienced no evidence of disease at 108 months. Microscopically, all tumors consisted of infiltrating nests of polygonal cells with vesicular nuclei, prominent nucleoli and basophilic cytoplasm. Abrupt keratinization was present in only one case. Immunohistochemically, there was a diffuse expression of cytokeratin (CK) cocktail, CK7, p40, p63, and SMARCB1/INI1. All NMCs tested negative for EBV and HPV infection. Two NMCs showed methylation of RASSF1 gene. All other genes (APC, ATM, BRCA1, BRCA2, CADM1, CASP8, CD44, CDH13, CDKN1B, CDKN2A, CDKN2B, CHFR, DAPK1, ESR1, FHIT, GSTP1, HIC1, KLLN, MLH1a, MLH1b, RARB, TIMP3, and VHL) were unmethylated. All NMCs showed upregulation of miR-9 and downregulation of miR-99a and miR-145 and two cases featured also upregulation of miR-21, miR-143, and miR-484. In summary, we described three cases of sinonasal NMCs with novel findings on DNA methylation and miRNA expression, which might be important for new therapeutic strategies in the future.

HPV Status and Mutation Analysis Using Multiparallel Sequencing in Distal Oesophageal and Gastro-oesophageal Junction Adenocarcinomas.

The incidence of adenocarcinoma of oesophagus or gastro-oesophageal junction is increasing in Europe and other regions of the Western world. Research of possible causes has shifted to the molecular level. This study evaluated human papillomavirus (HPV) using real-time PCR and mutational status of selected genes using the multiparallel sequencing method (NGS) in DNA extracted from paraffin-embedded tumour tissue of 56 patients with oesophageal or gastro-oesophageal junction adenocarcinoma. The genetic material was in sufficient quality for the analysis in 37 cases (66 %). No HPV-positive sample was found. NGS revealed higher frequency of mutations in TP53, ARID1A, PIK3CA, SMAD4, ERBB2, MSH6, BRCA2, and RET genes. Association between gene mutations and histological grade, subtype according to Lauren, or primary tumour site was not statistically significant. In conclusion, the study did not confirm any HPV-positive sample of oesophageal and gastro-oesophageal junction adenocarcinoma. The study confirmed the usefulness of NGS analysis of paraffin-embedded tissue of these tumours, and it could be used in clinical studies to evaluate the prognostic and/or predictive value of the tested mutations. The association between gene mutations and histological features should be tested in larger patient cohorts.

Discordance between clinical and pathological TNM classification: influence on results of treatment and prognosis in patients with laryngeal cancer.

The goals of this retrospective cohort study were to compare the results of clinical and pathological TNM staging in patients with laryngeal squamous cell carcinoma and to determine the impact of the discordance on prognosis and treatment results. A total of 124 patients with laryngeal cancer, primarily indicated for surgical treatment, were enrolled. The concordance or discordance between the clinical and pathological staging was compared with the frequency of cancer relapse and disease-specific survival. Other potential prognostic factors, like age, the stage and location of the primary tumor, the status of neck lymph nodes, histological margins, and an indication for postoperative radiotherapy, were also evaluated. A disparity in at least one component of TNM staging was found in 40 patients (32%). The discordance had significant negative influence on both disease-free survival (DSF) and disease-specific survival (DSS). Other significant negative prognostic factors were the stage of the primary tumor, nodal status and postoperative radiotherapy. Our results indicate that the discordance between clinical and pathological staging affects the results of cancer treatment significantly. Some improvement can be probably achieved with higher preoperative diagnostic method accuracy.

Human papillomavirus and Epstein-Barr virus in nasopharyngeal carcinoma in a non-endemic eastern european population.

Nasopharyngeal carcinoma (NPC) is a rare malignancy in the Czech Republic and Slovakia, with the standardized incidence rate of < 1:100000 person-years. Viral status of NPC in these non-endemic Eastern European regions is currently unknown. In a retrospective study, we evaluated the presence of EBV and HPV in 62 NPC cases. EBV status was determined by the use of in situ hybridization (ISH) for EBV encoded small RNA 1 (EBER1). HPV status was examined with p16 immunohistochemistry, DNA ISH and DNA polymerase chain reaction. Sixty-one studied cases showed non-keratinizing morphology and one was keratinizing squamous cell carcinoma. Only one NPC with non-keratinizing morphology was scored as p16-positive (nuclear and cytoplasmic staining ≥ 70% of tumor cells). This case was positive for high-risk HPV by ISH and the DNA PCR confirmed the presence of HPV18 type. At the same time, this case was found negative for EBV. Remaining sixty-one cases that were scored as p16-negative were all found HPV-negative by ISH and the DNA PCR. EBV was detected in 85.5% (53/62) of cases and 9 cases were EBV-negative, including the case of keratinizing NPC. In contrast with previous reports on the prevalence of EBV-positivity in Caucasian patients with NPC, the majority of patients coming from this non-endemic region show EBV-positivity; therefore, they may be candidates for novel EBV-targeting therapies. Conversely, HPV-positive NPC is very rare and HPV does not seem to play a significant role in the etiopathogenesis of NPC in these Eastern European populations.

Importance of Tumour Suppressor Gene Methylation in Sinonasal Carcinomas.

Epigenetic changes are considered to be a frequent event during tumour development. Hypermethylation of promoter CpG islands represents an alternative mechanism for inactivation of tumour suppressor genes, DNA repair genes, cell cycle regulators and transcription factors. The aim of this study was to investigate promoter methylation of specific genes in samples of sinonasal carcinoma by comparison with normal sinonasal tissue. To search for epigenetic events we used methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) to compare the methylation status of 64 tissue samples of sinonasal carcinomas with 19 control samples. We also compared the human papilloma virus (HPV) status with DNA methylation. Using a 20% cut-off for methylation, we observed significantly higher methylation in RASSF1, CDH13, ESR1 and TP73 genes in the sinonasal cancer group compared with the control group. HPV positivity was found in 15/64 (23.4 %) of all samples in the carcinoma group and in no sample in the control group. No correlation was found between DNA methylation and HPV status. In conclusion, our study showed that there are significant differences in promoter methylation in the RASSF1, ESR 1, TP73 and CDH13 genes between sinonasal carcinoma and normal sinonasal tissue, suggesting the importance of epigenetic changes in these genes in carcinogenesis of the sinonasal area. These findings could be used as prognostic factors and may have implications for future individualised therapies based on epigenetic changes.

[Expression of ATP-binding Cassette Proteins Pgp, MRP1, and MRP3 in Malignant and Benign Ovarian Lesions]. / Exprese ABC transportéru Pgp, MRP1 a MRP3 u maligních a benigních zmen vajecníku.

BACKGROUND: This study was designed to compare the expression of PgP (P-glycoprotein), MRP1 (multidrug related protein), and MRP3 in ovarian cancer patients, patients with benign ovarian tumors, and healthy women, and to evaluate the correlation between the expression of ATP-binding cassette proteins Pgp, MRP1, and MRP3 with stage, grade, and histological type. PATIENTS AND METHODS: Tissue specimens from 212 women who underwent surgery at the Department of Obstetrics and Gynecology at University Hospital Hradec Králové were subjected to immunohistochemical staining for Pgp, MRP1, and MRP3. RESULTS: The expression of Pgp and MRP1 was higher in ovarian tumor cells than in the cells lining the ovarian cyst. The lowest level of expression was found in normal ovarian tissue (p < 0.001). Histological subtype of epithelial ovarian cancer correlated with the expression of PgP, MRP1, and MRP3. The lowest level of Pgp and MRP1 expression was found in endometrioid ovarian cancers (p = 0.151; p = 0.013). Patients with advanced ovarian cancer (FIGO III + IV) had higher MRP1 expression than those with early stage ovarian cancer (median MRP1 FIGO I + II 80%; CI 60-100; FIGO III + IV 100%; CI 90-100; p = 0.100). An association was observed between MRP1 and tumor grade (p < 0.001). CONCLUSION: Pgp and MRP1 expression was higher in ovarian tumor cells than in cells lining the ovarian cyst. The lowest level of expression was found in normal ovarian tissue. ATP-binding cassette proteins play an important role in ovarian cancer pathogenesis.Key words: ATP-binding cassette proteins - ovarian cancer - P-glycoprotein (Pgp) - multidrug related protein 1 (MRP1) - multidrug related protein 3 (MRP3) - drug resistanceThis work was supported by the Czech Ministry of Health NT 14107-3/2013.The authors declare they have no potential confl icts of interest concerning drugs, products, or services used in the study.The Editorial Board declares that the manuscript met the ICMJE recommendation for biomedical papers.Submitted: 9. 11. 2015Accepted: 30. 8. 2016.

[Discordance between Clinical and Pathological TNM Classifications in Patients with Oropharyngeal Cancer - Influence on Treatment and Prognosis]. / Shoda a neshoda klinické a patologické TNM klasifikace u karcinomu orofaryngu - vliv na prognózu a výsledky lécby.

BACKGROUND: The aim of this study was to determine the percentage of discordance between clinical (c) and pathological (p) TNM classifications in cases of oropharyngeal carcinoma and whether it influences recurrence rate and prognosis of primary disease. MATERIALS AND METHODS: Fifty-one patients with oropharyngeal carcinoma who underwent primary surgical treatment were included in this retrospective study. Clinical TNM was determined on the basis of clinical examinations and imaging (US, CT, or MRI), and pathological TNM was determined by a histopathologist (analysis of the primary tumor and neck lymph nodes). Concordance and discordance were statistically evaluated. As potential prognostic factors, we statistically analyzed tumor recurrence, specific and nonspecific patient survival, patient age, extent of primary tumor, lymph node positivity, number of removed lymph nodes, and positive tumor margins. RESULTS: Discordance in the TNM classification was found in 27 cases. Disease-free survival was shorter in patients with discordance in T, and this was statistically significant (p = 0.034). Six patients died due to primary disease (11.8%). Disease-specific survival was at the limit of statistical significance (p = 0.069). CONCLUSIONS: Discordance between clinical and pathological TNM classifications was 52.9% patients with oropharyngeal carcinoma. Discordance in T is a potential prognostic factor. Improvement in cancer treatment to some extent relies on preoperative staging and should influence the decision about whether or not to administer adjuvant oncological treatment.

[Prognostic significance of Pgp, MRP1, and MRP3 in ovarian cancer patients]. / Prognostický význam Pgp, MRP1 a MRP3 u pacientek s karcinomem ovaria.

OBJECTIVE: To evaluate the correlation of resistance proteins Pgp (P-glycoprotein), MRP1 (Multidrug Related Protein, Multidrug Resistance-Associated Protein) and MRP3 with clinical - pathological factors and to find the clinical outcome of these data in ovarian cancer patients. DESIGN: Prospective study. SETTING: Department of Gynecology and Obstetrics, Charles University in Prague, Faculty of Medicine in Hradec Králové, University Hospital Hradec Králové. METHODS: 133 patients with epithelial ovarian cancer who underwent primary surgery from 2006-2010 had specimens stained with imunohistochemistry for Pgp, MRP1, MRP3. RESULTS: The histological subtype of epithelial ovarian cancer correlated with the expression of PgP, MRP1, and MRP3. The lowest incidence of Pgp and MRP1 expression was documented in endometrioid ovarian cancers (P = 0.151, P = 0.013). Patients with advanced ovarian cancer (FIGO III+IV) had higher MRP1 expression than those with early stage ovarian cancer (Med MRP1 FIGO I+II 80%; CI: 60-100; FIGO III+IV 100%; CI: 90-100; P = 0.100). An association was observed between MRP1 and tumor grade (Med MRP1 G1 80% (CI: 0-100), G2 80% (CI: 30-100), G3 100% (CI: 90-100); P < 0.001). There was no relationship between the size of the residual tumor after primary surgery and any resistance proteins. Patients with complete response after primary treatment had lower levels of LRP, Pgp, and MRP1 expression than other patients. Patients with higher Pgp and MRP1 expression had relapse of disease during the following 24 months more often than patients with lower Pgp and MRP1 expression. FIGO stage, histological type, debulking efficiency, and Pgp and MRP1 expression correlated with poor patient survival (P < 0.001, P < 0.001, P < 0.001, P = 0.040, P = 0.026). CONCLUSION: We found prognostic significance of Pgp, MRP1 and MRP3 expression in ovarian cancer patients. MRP1 have some additional prognostic value for the clinical outcome of patients with ovarian carcinoma.

[Tubulo-squamous polyp of the vagina]. / Tubulo-skvamózní polyp vaginy.

OBJECTIVE: To decribe a case of rare type of vaginal polyp. DESIGN: Case report. SETTING: Department of Pathological Anatomy, Regional hospital Nachod. CONCLUSION: Tubulo-squamous polyp is rare type of vaginal polyp first described in 2007, with only up to 20 cases have been reported in the literature so far. It affects mostly postmenopausal women and presents as a polypoid mass occurring in the upper part of vagina. Histologically, it is composed of squamous and glandular component within fibrous stroma. The etiopathogenesis of this lesion remains unclear, but it may arise from mesonephric remnants or Skene glands (so-called female prostate). This theory is supported by the fact that some cases show expression of prostate specific acid phosphatase (PSAP) and/or prostate specific antigen (PSA).

Promoter methylation of GATA4, WIF1, NTRK1 and other selected tumour suppressor genes in ovarian cancer.

Ovarian cancer is the leading cause of death from gynaecologic tumours, but the molecular and especially epigenetic events underlying the transformation are poorly understood. Various methylation changes have been identified and show promise as potential cancer biomarkers. The aim of this study was to investigate promoter methylation of selected tumour suppressor genes in ovarian cancer by comparison with normal ovarian tissue. To search for epigenetic events we used methylation-specific multiplex ligation-dependent probe amplification to compare the methylation status of 44 tissue samples of ovarian cancer with 30 control samples. Using a 20% cut-off for methylation, we observed significantly higher methylation in genes NTKR1, GATA4 and WIF1 in the ovarian cancer group compared with the control group. These findings could potentially be used in screening of ovarian cancer, and may have implications for future chemotherapy based on epigenetic changes.

[Solitary fibrous tumor of endometrium--a case report]. / Solitární fibrózní tumor endometria - kazuistika.

OBJECTIVE: Solitary fibrous tumor (SFT) is an uncommon mesenchymal tumor. We present a case of SFT occurring in endometrium. DESIGN: Case report. SETTING: Department of Obstetrics and Gynecology, The Fingerland Department of Pathology, Medical Faculty of Charles University and Faculty Hospital Hradec Králové. CASE REPORT: We report a case of 57 years old woman with SFT arising from the endometrium, which was diagnosed and treated at our department. Histological finding was supported by typical immunohistochemical profile of the tumor. Aggressive nature of the tumor wasnt showed. The patient underwent abdominal hysterectomy with bilateral adnexectomy and is followed up in regular periods. CONCLUSION: Occurence of solitary fibrous tumor (SFT) in the female genital tract is extremely rare. To the best of our knowledge, we report the first case of SFT occurring in endometrium. Because of potencial aggressive behaviour of the tumor complete surgical excision and close follow-up is highly recommended.

[Can be considered the primary fallopian tube cancer an enigma at the beginning of the third millenium?]. / Lze povazovat i na zacátku tretího tisíciletí primární karcinom vejcovodu za enigma?

OBJECTIVE: To analyze differences between primary fallopian tube cancer and ovarian cancer. DESIGN: Overview study. SETTING: Department of Obstetrics and Gynecology, The Fingerland Department of Pathology, Department of Oncology and Radiotherapy, Faculty of Medicine and University Hospital Hradec Kralove. METHODS: Overview study focused on analysis of data of primary fallopian tube cancer. CONCLUSION: The incidence of primary fallopian tube cancer was thought to be very low in the past but it is very difficult to assess the primary origin in the case of advanced disease (ovary versus fallopian tube). Tumorogenic potential of endosalpinx in relation to epithelial tumours is much more bigger. According to the current knowledge, the vast majority of high-grade serous carcinomas of the "ovary" in fact arise in the mucosa of fimbrial portion of fallopian tube.

[Drug resistance proteins LRP, Pgp, MRP1, MRP3 and MRP5 in ovarian cancer patients]. / Proteiny rezistence LRP, Pgp,MRP1, MRP3 a MRP5 u pacientek s karcinomem ovaria.

OBJECTIVE: To evaluate the correlation of drug resistance proteins LRP (Lung Resistance Protein), Pgp (P-glycoprotein), MRP1 (Multidrug Related Protein, Multidrug Resistance-Associated Protein), MRP3 a MRP5 with clinical - pathological factors and to find the clinical outcome of these data in ovarian cancer patients. DESIGN: Prospective study. SETTING: Department of Obstetrics and Gynecology, Charles University in Prague, Faculty of Medicine and University Hospital Hradec Kralove. METHODS: 111 patients with epithelial ovarian cancer who underwent primary surgery from 2006-2010 had specimens stained with imunohistochemistry for LRP, Pgp, MRP1, MRP3, MRP5. RESULTS: The histological subtype of epithelial ovarian cancer correlated to the LRP, Pgp, MRP1 and MRP3 expression. Patients with late ovarian cancer had a higher MRP1 compared to early stage ovarian cancer (I+II 71.6% (CI 60-100), III+IV 83.6% (CI 100-100),p = 0.03). Correlation of MRP1 with grading was found(G1 60.83% (CI 10-100), G2 36.80% (CI 20-100),G3 88.87% (CI 100-100), p = 0.039). Patients with high Pgp, MRP1 and MRP3 expression had significantly shorter progression-free survival. (Kaplan-Meier test - PFS, Pgp < 85% Med PFS 23 months (CI 8-37) vs > 85% Med PFS 11 months (CI 7-17), p = 0.054), (MRP1 < 85% Med PFS 33 months (CI 11-49) vs > 85% Med PFS 11 months(CI 7-16), p = 0.046). CONCLUSION: We found clinical significance of LRP, Pgp, MRP1 and MRP3 expression in ovarian cancer patients. MRP5 expression did not correlate with neither histo-pathological parameters nor progression free survival. MRP1 have some additional predictive and prognostic value for the clinical outcome of patients with ovarian carcinoma.

Selection of endogenous control and identification of significant microRNA deregulations in cervical cancer.

Introduction: Cervical cancer causes approximately 350,000 deaths each year. The availability of sensitive and specific diagnostic tests to detect cervical cancer in its early stages is essential to improve survival rates. Methods: In this study, we compared two strategies for selecting endogenous controls: miRNA profiling by small-RNA sequencing and a commercially available microfluidic card with 30 recommended endogenous controls preloaded by the manufacturer. We used the RefFinder algorithm and coefficient of variation to select endogenous controls. We selected the combination of miR-181a-5p and miR-423-3p as the most optimal normalizer. In the second part of this study, we determined the differential expression (between tumor/non-tumor groups) of microRNA in cervical cancer FFPE tissue samples. We determined the comprehensive miRNA expression profile using small-RNA sequencing technology and verified the results by real-time PCR. We determined the relative expression of selected miRNAs using the 2-ΔΔCt method. Results: We detected statistically significant upregulation of miR-320a-3p, miR-7704, and downregulation of miR-26a-5p in the tumor group compared to the control group. The combination of these miRNAs may have the potential to be utilized as a diagnostic panel for cervical cancer. Using ROC curve analysis, the proposed panel showed 93.33% specificity and 96.97% sensitivity with AUC = 0.985. Conclusions: We proposed a combination of miR-181a-5p and miR-423-3p as optimal endogenous control and detected potentially significant miRNAs (miR-320a-3p, miR-7704, miR-26a-5p). After further validation of our results, these miRNAs could be used in a diagnostic panel for cervical cancer.

Biologic importance and prognostic significance of selected clinicopathological parameters in patients with oral and oropharyngeal squamous cell carcinoma, with emphasis on smoking, protein p16(INK4a) expression, and HPV status.

The aim of this study is to evaluate the biologic importance and prognostic significance of selected clinicopathological parameters in patients with oral (OSCC) and oropharyngeal (OPSCC) squamous cell carcinoma, with emphasis on smoking, protein p16(INK4a) (p16) expression, and human papillomavirus (HPV) status.The study sample consisted of 48 patients with OSCC and 44 patients with OPSCC. Half of the patients were nonsmokers and the other half were gender-, age- and tumor localization-matched smokers. p16 expression was detected in 17/48 (35 %) OSCCs and in 36/44 (82 %) OPSCCs and HPV DNA was present in 7/48 (15 %) OSCCs and in 35/44 (80 %) OPSCCs. The sensitivity and specificity of p16 expression for HPV DNA presence were 0.74 and 0.88, respectively. The OPSCCs were more frequently basaloid (p < 0.001) while the OSCCs were more frequently conventional (p < 0.000001). The OSCCs were more likely to recur locally and to be the cause of death (p = 0.009 in both parameters).The HPV-positive tumors were more frequently localized in oropharynx, were basaloid SCCs and were p16- and HPV-positive (p < 0.000001 in all 4 parameters). The HPV-negative tumors were more frequently localized in oral cavity (p < 0.000001), more frequently asociated with local, regional and locoregional recurence (p = 0.011, p = 0.019 and p = 0.030, respectively) and with tumor-related death (p = 0.003). There was no significant difference with regard to smoking history (p > 0.05). The survival of patients with HPV-positive tumors was significantly longer (median 112 months; 95% CI 54 - 112 months) than that of patients with HPV-negative tumors (median 17 months; 95% CI 12 - 39 months) (p < 0.001). The HPV status of OSCC/OPSCC is an important biological and prognostic parameter and should be examined in all cases, using PCR or immunohistochemical detection of surrogate marker p16. Smoking itself does not seem to be an important prognostic factor.

Cellular cohesiveness in benign and malignant thyroid follicular tumours varies significantly, but the difference is not useful in diagnosis of individual cases.

INTRODUCTION: The aim of our study was to search for new, readily available and statistically reliable cytological markers for differentiating benign and malignant follicular thyroid neoplasms pre-operatively. METHODS: Cohesiveness of tumour cells in cytology slides from a series of 58 follicular tumours diagnosed between 1998 and 2004 inclusive was studied, including 48 follicular adenomas, and eight minimally invasive and two widely invasive follicular carcinomas. Photomicrographs of the cytology slides were taken and the digital images were analysed using computer image analysis software. We evaluated the relative proportions of cells arranged in groups of various sizes. The cohesiveness of the cells in cytological smears was then correlated with the immunohistochemical expression of E-cadherin in corresponding histological slides. RESULTS: Cases from 15 men (26%) and 43 women (74%) with a mean age of 50 years (range, 19-79) were analysed. In follicular adenomas and carcinomas, respectively, isolated cells were seen in 16.8% and 24.7% (P = 0.028), groups of two to five cells in 9.7% and 11.5% (P = 0.145) and groups of more than five cells in 73.5% and 63.8% (P = 0.041). The mean cell count in groups with more than five cells was 46.5 and 27.0 in adenomas and carcinomas, respectively (P < 0.001). Cell cohesiveness, either as percentage of cells in groups of more than five (R(2) = 0.026) or as mean cell count per group of more than five (R(2) = 0.005), was not found to be dependent on the expression of E-cadherin. Using a threshold of 13% isolated tumour cells in cytological smears, follicular adenomas and carcinomas could be distinguished with 90% sensitivity and 41% specificity. CONCLUSIONS: Although we demonstrated a statistically significant difference in cell cohesion between follicular adenomas and carcinomas, these could not be distinguished in the clinical setting by evaluation of the percentage of isolated cells in cytological smears because the specificity was too low. The absence of correlation of cellular cohesiveness with E-cadherin expression indicates that other factors are probably responsible for the loss of cohesiveness observed in follicular thyroid malignancy.

Methylation analysis of tumour suppressor genes in ovarian cancer using MS-MLPA.

Epigenetic changes are considered to be a frequent event during tumour development. Hypermethylation of promoter CpG islands represents an alternative mechanism for inactivation of tumour suppressor genes, DNA repair genes, cell cycle regulators and transcription factors. The aim of this study was to investigate promoter methylation of specific genes in ovarian cancer by comparison with normal ovarian tissue. To search for epigenetic events we used methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA) to compare the methylation status of 69 tissue samples of ovarian cancer with 40 control samples. Using a 15% cut-off for methylation, we observed significantly higher methylation in genes MGMT, PAX5, CDH13, WT1, THBS1, GATA5 in the ovarian cancer group, while in the ESR1 gene we observed significantly higher methylation in the control group compared with the ovarian cancer group. These findings could potentially be used in screening of ovarian cancer and may have implications for future chemotherapy based on epigenetic changes.

[What is the prognostic importance of molecular genetic factors in endometrial carcinoma?]. / Jaký je prognostický význam molekulárne genetických faktoru u karcinomu endometria?

BACKGROUND: Evaluation of the importance of molecular genetic factors in endometrial carcinoma based on our review of available literature, and in the case of K- ras mutation based on our own data. The aim of the original part of our study was to compare the presence of K- ras mutation in early stages of endometroid carcinoma with normal endometrium and evaluate the role of the mutation in endometrial carcinogenesis. MATERIAL AND METHODS: Molecular bio-logical analysis was performed to detect K- ras mutation in samples of endometrial tissue obtained from women treated in the past at the Department of Obstetrics and Gynecology, University Hospital Hradec Kralove. The detection was made from DNA isolated from paraffin embedded sections using K- ras -StripAssay, ViennaLab Diagnostics GmbH. RESULTS: K- ras mutation was found in 7 out of 30 specimens of endometroid carcinoma in stage I (23%) and in 3 of 20 specimens of normal endometrium in the control group (15%). K- ras mutations were more frequent in IA stage and grade 1 of endometroid carcinoma. CONCLUSION: The importance of molecular genetic factors in endometrial carcinoma differs depending on the type of carcinoma. In more common type 1 endometroid cancer, published data are not as clear as in type 2 carcinoma, in which prevalence of alteration of p53 reaches 90%. Results of our study performed on local population of women support the theory about the possible role of K- ras mutation as an early event in the process of endometrial carcinogenesis in type 1 tumors.