Study of alternatively spliced variants of estrogen receptor alpha in breast cancer cell lines

Filip Lhota: Study of alternatively spliced variants of estrogen receptor alpha in breast cancer cell lines Abstract: Estrogen receptor α (ER-α) is a transcription factor responsible for mediation of the activities of its natural ligand 17-β-estradiol (E2), the hormone that together with progesterone belongs to the key regulators of mammary epithelial as well as breast cancer cells proliferation. Except to the major gene product consisting of all eight coding exons of ER-α, numerous qualitatively and quantitatively different spliced variants originated from primary transcript by activity of alternative splicing is expressed. Despite that some of these spliced variants have been functionally characterized, their precise role on final ER-α cellular activity remains to be elucidated. The functional characterization of individual alternative forms of ER-α and description of its participation on the overall ER-α activity is important for our understanding of their biogenesis and is also critical for the delineation of molecular bases for ER-α regulation during anti cancer chemotherapy. This work aimed to study the influence of alternatively spliced ER-α variants on the growth characteristics of clones constructed from stable mammary tissue cell lines in regulation to cultivation conditions and cellular..

Identification of hereditary alterations predisposing to breast cancer development using "next-gen" sequencing

Summary: Breast cancer (BC) is the most frequent cancer type in female population of Europe. Approximately 5 - 10 % accounts for its hereditary form which is characterized by high penetrance, early onset, risen recurrence risk and development of other cancers. Mutational analyses of high risk patients identify a predisposing mutation in one of the most studied genes (BRCA1, BRCA2, TP53, ATM, CHEK2, NBS1, PALB2) only in less than one third of tested breast cancer patients. Lately, with the use of new methods of next-generation sequencing, a number of other susceptibility or candidate genes were characterized, but the incidence of their pathogenic alteration is often geographically different. A notable proportion of high risk patients from families with hereditary BC can represent carriers of population-specific, or private mutations. Most of the to date identified BC susceptibility genes codes for proteins involved in DNA repair, especially repair of double strand break DNA repair. Nevertheless the mutation analysis was conducted only on a small fraction of these DNA repair genes. We can expect that in the group of yet nontested genes coding for DNA repair proteins a rare, but clinically important genetic alterations predisposing to BC in affected families can be discovered. This work describes a..

Proposal for the corporate design of FTVS UK

Title of study: Proposal for the corporate design of FTVS UK Study aim: An analysis of the present situation in the area of the faculty's visual style and proposals for its amelioration by means of a graphic manual. Method: Analysis of internal and external documents and a semi-structured interview are used in this Master's Thesis. Results: A complete graphic manual of FTVS will be presented as a final proposal for an amelioration of the present state. Key words: company communication, company identity, corporate design, logo, graphic manua

Marketing research of value of the brand Puma

My thesis introduces a problem of value identification concerning the brand Puma in the Czech Republic. The main objective of this thesis is to find out how is Puma perceived by the Czech population as well as how satisfied and familiar is the Czech population with its products. The theoretical part of the thesis will deal with obtaining information and defining terms in field of marketing research, brand value and propagation. The practical part will include data collection and its processing, which will enable this research to present concrete results. The techniques I will use are electronic questionnaire and depth interview. Keywords: brand, promotion, sponsorship, marketing researc

Germline alterations of the <i>CHEK2</i> gene changing the CHK2 protein structure identified in NHL patients and controls with their frequencies and related odds ratios (OR).

<p>^a New alterations</p><p>^b Two alterations of the <i>CHEK2</i> coding sequence were identified in one patient (c.470T>C and c.1259+1G>C); <i>OR</i>–odds ratio; <i>CI</i>–confidence interval.</p><p>Note: The nomenclature of <i>CHEK2</i> alterations was based on NCBI <i>CHEK2</i> Reference Sequences NG_008150.1 (gene) and NM_007194.3 (mRNA).</p><p>Germline alterations of the <i>CHEK2</i> gene changing the CHK2 protein structure identified in NHL patients and controls with their frequencies and related odds ratios (OR).</p

Association of Germline <i>CHEK2</i> Gene Variants with Risk and Prognosis of Non-Hodgkin Lymphoma

<div><p>The checkpoint kinase 2 gene (<i>CHEK2</i>) codes for the CHK2 protein, an important mediator of the DNA damage response pathway. The <i>CHEK2</i> gene has been recognized as a multi-cancer susceptibility gene; however, its role in non-Hodgkin lymphoma (NHL) remains unclear. We performed mutation analysis of the entire <i>CHEK2</i> coding sequence in 340 NHL patients using denaturing high-performance liquid chromatography (DHPLC) and multiplex ligation-dependent probe amplification (MLPA). Identified hereditary variants were genotyped in 445 non-cancer controls. The influence of <i>CHEK2</i> variants on disease risk was statistically evaluated. Identified <i>CHEK2</i> germline variants included four truncating mutations (found in five patients and no control; P = 0.02) and nine missense variants (found in 21 patients and 12 controls; P = 0.02). Carriers of non-synonymous variants had an increased risk of NHL development [odds ratio (OR) 2.86; 95% confidence interval (CI) 1.42–5.79] and an unfavorable prognosis [hazard ratio (HR) of progression-free survival (PFS) 2.1; 95% CI 1.12–4.05]. In contrast, the most frequent intronic variant c.319+43dupA (identified in 22% of patients and 31% of controls) was associated with a decreased NHL risk (OR = 0.62; 95% CI 0.45–0.86), but its positive prognostic effect was limited to NHL patients with diffuse large B-cell lymphoma (DLBCL) treated by conventional chemotherapy without rituximab (HR-PFS 0.4; 94% CI 0.17–0.74). Our results show that germ-line <i>CHEK2</i> mutations affecting protein coding sequence confer a moderately-increased risk of NHL, they are associated with an unfavorable NHL prognosis, and they may represent a valuable predictive biomarker for patients with DLBCL.</p></div

Germline intronic and silent alterations in the <i>CHEK2</i> gene in NHL patients and controls with their frequencies and related odds ratios (OR).

<p>^a New alterations</p><p>^bThe c.319+43dupA alteration also did not show a statistically significant deviation from the Hardy-Weinberg equilibrium in any of the analyzed groups (all p > 0.05).</p><p>Germline intronic and silent alterations in the <i>CHEK2</i> gene in NHL patients and controls with their frequencies and related odds ratios (OR).</p

Validation of CZECANCA (CZEch CAncer paNel for Clinical Application) for targeted NGS-based analysis of hereditary cancer syndromes

<div><p>Background</p><p>Carriers of mutations in hereditary cancer predisposition genes represent a small but clinically important subgroup of oncology patients. The identification of causal germline mutations determines follow-up management, treatment options and genetic counselling in patients’ families. Targeted next-generation sequencing-based analyses using cancer-specific panels in high-risk individuals have been rapidly adopted by diagnostic laboratories. While the use of diagnosis-specific panels is straightforward in typical cases, individuals with unusual phenotypes from families with overlapping criteria require multiple panel testing. Moreover, narrow gene panels are limited by our currently incomplete knowledge about possible genetic dispositions.</p><p>Methods</p><p>We have designed a multi-gene panel called CZECANCA (CZEch CAncer paNel for Clinical Application) for a sequencing analysis of 219 cancer-susceptibility and candidate predisposition genes associated with frequent hereditary cancers.</p><p>Results</p><p>The bioanalytical and bioinformatics pipeline was validated on a set of internal and commercially available DNA controls showing high coverage uniformity, sensitivity, specificity and accuracy. The panel demonstrates a reliable detection of both single nucleotide and copy number variants. Inter-laboratory, intra- and inter-run replicates confirmed the robustness of our approach.</p><p>Conclusion</p><p>The objective of CZECANCA is a nationwide consolidation of cancer-predisposition genetic testing across various clinical indications with savings in costs, human labor and turnaround time. Moreover, the unified diagnostics will enable the integration and analysis of genotypes with associated phenotypes in a national database improving the clinical interpretation of variants.</p></div

Identification and Functional Testing of ERCC2 Mutations in a Multi-national Cohort of Patients with Familial Breast- and Ovarian Cancer

The increasing application of gene panels for familial cancer susceptibility disorders will probably lead to an increased proposal of susceptibility gene candidates. Using ERCC2 DNA repair gene as an example, we show that proof of a possible role in cancer susceptibility requires a detailed dissection and characterization of the underlying mutations for genes with diverse cellular functions (in this case mainly DNA repair and basic cellular transcription). In case of ERCC2, panel sequencing of 1345 index cases from 587 German, 405 Lithuanian and 353 Czech families with breast and ovarian cancer (BC/OC) predisposition revealed 25 mutations (3 frameshift, 2 splice-affecting, 20 missense), all absent or very rare in the ExAC database. While 16 mutations were unique, 9 mutations showed up repeatedly with population-specific appearance. Ten out of eleven mutations that were tested exemplarily in cell-based functional assays exert diminished excision repair efficiency and/or decreased transcriptional activation capability. In order to provide evidence for BC/OC predisposition, we performed familial segregation analyses and screened ethnically matching controls. However, unlike the recently published RECQL example, none of our recurrent ERCC2 mutations showed convincing co-segregation with BC/OC or significan