Approximately one third of children with steroid-resistant nephrotic syndrome (SRNS) carry pathogenic variants in one of the many associated genes. The WT1 gene coding for the WT1 transcription factor is among the most frequently affected genes. Cases from the Czech national SRNS database were sequenced for exons 8 and 9 of the WT1 gene. Eight distinct exonic WT1 variants in nine children were found. Three children presented with isolated SRNS, while the other six manifested with additional features. To analyze the impact of WT1 genetic variants, wild type and mutant WT1 proteins were prepared and the DNA-binding affinity of these proteins to the target EGR1 sequence was measured by microscale thermophoresis. Three WT1 mutants showed significantly decreased DNA-binding affinity (p.Arg439Pro, p.His450Arg and p.Arg463Ter), another three mutants showed significantly increased binding affinity (p.Gln447Pro, p.Asp469Asn and p.His474Arg), and the two remaining mutants (p.Cys433Tyr and p.Arg467Trp) showed no change of DNA-binding affinity. The protein products of WT1 pathogenic variants had variable DNA-binding affinity, and no clear correlation with the clinical symptoms of the patients. Further research is needed to clarify the mechanisms of action of the distinct WT1 mutants; this could potentially lead to individualized treatment of a so far unfavourable disease.

• MeSH
• dítě MeSH
• DNA terapeutické užití   MeSH
• léková rezistence MeSH
• lidé MeSH
• mutace MeSH
• nefrotický syndrom * farmakoterapie   genetika   metabolismus   MeSH
• proteiny WT1 * genetika   metabolismus   MeSH
• steroidy farmakologie   MeSH
• Check Tag
• dítě MeSH
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Recently, an increasing incidence of HPV-induced oropharyngeal squamous cell carcinoma (OPSCC) has been observed. Moreover, locoregionally advanced stages require a combined modal approach, and the prognosis is poor. Therefore, it is essential to find early diagnostic and prognostic biomarkers. DNA methylation changes play a crucial role in the process of carcinogenesis and are often investigated as promising biomarkers in many types of cancer. For analysis of DNA methylation levels of selected tumour suppressor genes in HPV-positive and HPV-negative samples (including primary tumours and corresponding metastases of metastasizing OPSCCs, primary tumours of non-metastasizing OPSCCs, and control samples), methylation-specific MLPA and methylation-specific high-resolution melting analyses were used. A significant difference in methylation between OPSCCs and the control group was observed in WT1, PAX6 (P < 0.01) and CADM1, RARβ (P < 0.05) genes. CADM1 and WT1 hypermethylation was detected mostly in HPV-positive samples; all but one HPV-negative samples were unmethylated. Moreover, hypermethylation of PAX5 gene was observed in metastases compared with control samples and was also associated with shorter overall survival of all patients (P < 0.05). Associations described herein between promoter methylation of selected genes and clinicopathological data could benefit OPSCC patients in the future by improvement in screening, early detection, and prognosis of the disease.

• MeSH
• aktivátorový protein specifický pro B-buňky genetika   MeSH
• Alphapapillomavirus * MeSH
• buněčná adhezní molekula 1 genetika   metabolismus   MeSH
• dlaždicobuněčné karcinomy hlavy a krku genetika   MeSH
• DNA metabolismus   MeSH
• infekce papilomavirem * komplikace   MeSH
• lidé MeSH
• metylace DNA MeSH
• nádory hlavy a krku * genetika   MeSH
• nádory orofaryngu * patologie   MeSH
• Papillomaviridae MeSH
• prognóza MeSH
• proteiny WT1 genetika   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

BACKGROUND: This work summarizes our experience with WT1 monitoring before and after allogeneic hematopoietic stem cell transplantation (allo-HSCT). PATIENTS AND METHODS: The expression of WT1 gene was measured by real-time polymerase chain reaction in peripheral blood according to the European Leukemia Net recommendations. Between May 2005 and August 2019, we analyzed 147 consecutive patients with acute myeloid leukemia with high WT1 expression at diagnosis, transplanted in first (CR1) or second (CR2) complete remission. RESULTS: At the time of allo-HSCT, 107 patients had WT1-normal expression (WT1 ≤ 50 copies), and 40 patients had WT1-high expression. The median follow-up was 21 months. The estimated 5-year overall survival and event-free survival was significantly better in the WT1-normal cohort (65% and 57% vs. 37% and 25%; P = .0003 and P < .0001, respectively) and 5-year cumulative incidence of relapse was significantly lower in the WT1-normal group (19% vs. 53%; P < .0001). Five-year non-relapse mortality was not significantly different (20% and 23%). Multivariate analysis revealed WT1-high expression and acute graft-versus-host disease grade 3/4 as significantly negative prognostic factors for OS. Overall, 49 patients developed WT1 molecular relapse in the post-transplant period; in 14 cases, the therapeutic intervention was done. In all but 1 relapsed patient where WT1 minimal residual disease (MRD) was monitored (38 patients), we detected WT1-high levels (sensitivity of 97%). CONCLUSION: The results of the analysis confirmed our previous experience that WT1 status before allo-HSCT is a strong prognostic factor for both OS and relapse risk. In addition, we confirmed the usefulness of this marker for MRD monitoring after allo-HSCT. The main advantage is the possibility of frequent MRD monitoring in peripheral blood and early bone marrow examination based on WT1-high expression.

• MeSH
• akutní myeloidní leukemie krev   genetika   mortalita   terapie   MeSH
• dospělí MeSH
• hodnocení rizik metody   MeSH
• incidence MeSH
• lidé středního věku MeSH
• lidé MeSH
• lokální recidiva nádoru epidemiologie   genetika   MeSH
• mladý dospělý MeSH
• nádorové biomarkery krev   metabolismus   MeSH
• následné studie MeSH
• nemoc štěpu proti hostiteli diagnóza   epidemiologie   MeSH
• přežití bez známek nemoci MeSH
• prognóza MeSH
• proteiny WT1 krev   metabolismus   MeSH
• regulace genové exprese u leukemie MeSH
• reziduální nádor MeSH
• rizikové faktory MeSH
• senioři MeSH
• stanovení celkové genové exprese MeSH
• studie proveditelnosti MeSH
• stupeň závažnosti nemoci MeSH
• transplantace hematopoetických kmenových buněk škodlivé účinky   MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

• MeSH
• biologické markery MeSH
• diagnostické zobrazování metody   využití   MeSH
• dvojčata monozygotní MeSH
• farmakoterapie MeSH
• fatální výsledek MeSH
• genetické testování * MeSH
• klinický obraz nemoci MeSH
• kojenec MeSH
• lidé MeSH
• nefrektomie metody   využití   MeSH
• nefrologie MeSH
• nefrotický syndrom * diagnóza   etiologie   terapie   MeSH
• proteiny WT1 izolace a purifikace   MeSH
• renální insuficience farmakoterapie   terapie   MeSH
• statistika jako téma MeSH
• ultrasonografie prenatální metody   využití   MeSH
• vrozené, dědičné a novorozenecké nemoci a abnormality diagnóza   etiologie   terapie   MeSH
• Wilmsův nádor * diagnóza   etiologie   terapie   MeSH
• Check Tag
• kojenec MeSH
• lidé MeSH
• ženské pohlaví MeSH
• Publikační typ
• kazuistiky MeSH

BACKGROUND: Up to 55% of non-APL acute myeloid leukemias (AML) lack a molecular target suitable for standardized disease monitoring. We aimed to evaluate the prognostic significance of WT1 gene expression at early stages of intensive treatment. PATIENTS AND METHODS: A total of 106 consecutive patients with intermediate and high-risk AML who had WT1 expression at diagnosis >500 copies/104ABL and who achieved remission after 1 to 2 cycles of induction treatment were analyzed. WT1 expression was measured in peripheral blood using a standardized European LeukemiaNet method. Overexpression was defined as >50 copies/104ABL. The median follow-up was 30 months. RESULTS: Patients with normal versus high WT1 expression after 2 cycles of chemotherapy had overall survival (OS) at 3 years of 66% versus 41% (P = .01); event-free survival (EFS) 45% versus 22% (P = .01). Prognostic significance of WT1 expression after 2 cycles of treatment was maintained in the group of patients treated with chemotherapy alone without hematopoietic stem cell transplantation in first line treatment (OS 70% vs. 36%, P = .02; EFS 35% vs. 0%, P = .03). Significant prognostic factors for EFS on multivariate analysis were the achievement of molecular remission (<50 copies of WT1) at any time during treatment (hazard ratio [HR] 0.47, P = .04) and increased WT1 expression after 2 cycles of chemotherapy (HR 2.0, P = .03). CONCLUSION: Increased WT1 expression after 2 cycles of chemotherapy is a negative prognostic factor for survival. WT1 remains a valuable molecular marker in AML without any leukemia-specific mutation, especially if next generation sequencing and/or digital polymerase chain reaction are not routinely available.

• MeSH
• akutní myeloidní leukemie genetika   mortalita   MeSH
• analýza přežití MeSH
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• proteiny WT1 metabolismus   MeSH
• rizikové faktory MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

:Background: Current diagnosis and staging of advanced epithelial ovarian cancer (aEOC) has important limitations and better biomarkers are needed. We investigate the performance of non-haematopoietic circulating cells (CCs) at the time of disease presentation and relapse. Methods: Venous blood was collected prospectively from 37 aEOC patients and 39 volunteers. CCs were evaluated using ImageStream TechnologyTM and specific antibodies to differentiate epithelial cells from haematopoetic cells. qRT-PCR from whole blood of relapsed aEOC patients was carried out for biomarker discovery. Results: Significant numbers of CCs (CK+/WT1+/CD45-) were identified, quantified and characterised from aEOC patients compared to volunteers. CCs are abundant in women with newly diagnosed aEOC, prior to any treatment. Evaluation of RNA from the CCs in relapsed aEOC patients (n = 5) against a 79-gene panel revealed several differentially expressed genes compared to volunteers (n = 14). Size differentiation of CCs versus CD45+ haematopoietic cells was not reliable. Conclusion: CCs of non-haematopoetic origin are prevalent, particularly in patients with newly diagnosed aEOC. Exploiting a CC-rich population in aEOC patients offers insights into a part of the circulating microenvironment.

• MeSH
• antigeny CD45 metabolismus   MeSH
• karcinom krev   MeSH
• lidé středního věku MeSH
• lidé MeSH
• nádorové biomarkery metabolismus   MeSH
• nádorové buněčné linie MeSH
• nádorové cirkulující buňky metabolismus   patologie   MeSH
• nádory vaječníků krev   MeSH
• proteiny WT1 metabolismus   MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Renal mass lesions with a follicular architecture resembling atrophic kidney have been described, but their distinction from thyroid-like follicular carcinoma of the kidney remains controversial. We collected 8 cases of this purported "atrophic kidney"-like lesion to fully describe their clinical and histologic spectrum, their possible etiology, and to discuss their distinction from other renal neoplasms. Eight total cases were identified with patient ages ranging from 9 to 48 years (mean: 29 y; median: 28.5 y). Four patients were female and 4 were male. The tumors were unifocal and size ranged from 1.6 to 4.9 cm (mean: 3.4 cm; median: 3.4 cm). All 8 tumors had a remarkably similar histology. Each was enveloped by a smooth muscle rich capsule and had an overall low power "follicular" architecture. The luminal spaces of the "follicles" (or cysts) contained eosinophilic secretions and the lining epithelium was often flattened and atrophic, but some had more rounded cells with a distinctive hobnail arrangement. Many cysts contained discohesive round cells floating within the eosinophilic material, and some contained small intraluminal tufts with features of markedly atrophic glomeruli. Periodic acid-Schiff stains highlighted basement membrane material extending into these glomerular-like tufts, and some contained small distinct capillaries surrounded by endothelial cells, interspersed mesangial-like cells, and rare surrounding podocyte-like cells, providing additional evidence for glomerulocystic structures. Scattered calcifications were present within cysts (or within cyst walls) in varying numbers and were characterized by 2 types: psammoma body-like or more amorphous deposits. The tissue between cystic glomeruli contained predominantly small atrophic tubular structures, but collagenized stroma and smaller collapsed glomeruli were also present. The 2 tumors from the oldest 2 patients (48 and 39 y) had a more striking degree of stromal hyalinization. Immunohistochemically, the cyst lining cells had a predominant WT-positive/PAX-8 negative/CK7-negative phenotype, while tubules were typically WT-1 negative/PAX-8 positive/CK7-positive. Upon comparison to a control group of 10 kidneys containing incidental non-mass-forming glomerulocystic change, the morphologic features and immunophenotype were identical. To date, no patient has had any recurrence or aggressive clinical behavior based on follow status in 7 of 8 cases (follow-up range: 9 to 168 mo; median: 24 mo; mean: 40 mo). In summary, we describe the clinicopathologic features of 8 unique, benign "atrophic kidney"-like lesions that may simply represent a non-neoplastic form of organizing tubular atrophy and glomerulocystic change, and emphasize their distinction from thyroid-like follicular carcinoma of the kidney.

• MeSH
• atrofie MeSH
• biopsie MeSH
• buňky stromatu patologie   MeSH
• diferenciální diagnóza MeSH
• dítě MeSH
• dospělí MeSH
• eozinofily patologie   MeSH
• epitelové buňky patologie   MeSH
• folikulární adenokarcinom chemie   klasifikace   patologie   MeSH
• hladké svalstvo patologie   MeSH
• imunohistochemie MeSH
• keratin-7 analýza   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• nádorové biomarkery analýza   MeSH
• nádory ledvin chemie   klasifikace   patologie   MeSH
• prediktivní hodnota testů MeSH
• prospektivní studie MeSH
• proteiny WT1 analýza   MeSH
• studie případů a kontrol MeSH
• transkripční faktor PAX8 analýza   MeSH
• tumor burden MeSH
• Check Tag
• dítě MeSH
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

BACKGROUND: Steroid-resistant nephrotic syndrome (SRNS) has a heterogeneous spectrum of monogenic causes that substantially differ among populations. The aim of this study was to analyse the genetic aetiology of SRNS in Czech and Slovak paediatric patients. METHODS: We analysed clinical data from 74 patients (38 boys) with congenital (15%), infant (14%), and childhood-onset (71%) SRNS collected from the Czech Republic and Slovakia from 2000 to 2017 (inclusive). The DNA samples were first analysed by Sanger sequencing (genes NPHS2, NPHS1, and WT1) and then by next generation sequencing (NGS) using a targeted panel of 48 genes previously associated with SRNS. Family segregation of the causative variants was confirmed by Sanger sequencing when possible. RESULTS: Genetic diagnosis was established in 28/74 patients (38%) based on findings of pathogenic or likely pathogenic causative variants in genotypes conforming to the expected mode of inheritance. Sanger sequencing diagnosed 26% of patients, whereas second-tier testing by a targeted NGS panel diagnosed a further 12%. Frequent causative genes were NPHS2 (15%), WT1 (9.5%), and surprisingly NUP93 with four (5.4%) unrelated cases. Additional causative genes included COQ2 (two patients), NPHS1, INF2, DGKE, and LMX1B (one patient each). CONCLUSIONS: Compared with outright use of NGS, our tiered genetic testing strategy was considerably more rapid and marginally less expensive. Apart from a high aetiological fraction of NPHS2 and WT1 genes, our study has identified an unexpectedly high frequency of a limited set of presumably ancestral causative mutations in NUP93. The results may aid in tailoring testing strategies in Central European populations.

• MeSH
• dítě MeSH
• genetická predispozice k nemoci * MeSH
• genetické testování MeSH
• genotyp MeSH
• intracelulární signální peptidy a proteiny genetika   MeSH
• kojenec MeSH
• komplex proteinů jaderného póru genetika   MeSH
• lidé MeSH
• longitudinální studie MeSH
• membránové proteiny genetika   MeSH
• mladiství MeSH
• mutace MeSH
• mutační analýza DNA MeSH
• nefrotický syndrom genetika   MeSH
• novorozenec MeSH
• předškolní dítě MeSH
• prospektivní studie MeSH
• proteiny WT1 genetika   MeSH
• Check Tag
• dítě MeSH
• kojenec MeSH
• lidé MeSH
• mladiství MeSH
• mužské pohlaví MeSH
• novorozenec MeSH
• předškolní dítě MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika MeSH
• Slovenská republika MeSH

PURPOSE: Wilms tumor gene 1 (WT1), a zinc-finger transcription factor essential for testis development and function, along with other genes, was investigated for their role in the pathogenesis of testicular germ cell tumors (TGCT). METHODS: In total, 284 TGCT and 100 control samples were investigated, including qPCR for WT1 expression and BRAF mutation, p53 immunohistochemistry detection, and massively parallel amplicon sequencing. RESULTS: WT1 was significantly (p < 0.0001) under-expressed in TGCT, with an increased ratio of exon 5-lacking isoforms, reaching low levels in chemo-naïve relapsed TGCT patients vs. high levels in chemotherapy-pretreated relapsed patients. BRAF V600E mutation was identified in 1% of patients only. p53 protein was lowly expressed in TGCT metastases compared to the matched primary tumors. Of 9 selected TGCT-linked genes, RAS/BRAF and WT1 mutations were frequent while significant TP53 and KIT variants were not detected (p = 0.0003). CONCLUSIONS: WT1 has been identified as a novel factor involved in TGCT pathogenesis, with a potential prognostic impact. Distinct biologic nature of the two types of relapses occurring in TGCT has been demonstrated. Differential mutation rate of the key TGCT-related genes has been documented.

• MeSH
• down regulace MeSH
• fenotyp MeSH
• genetická predispozice k nemoci MeSH
• geny ras * MeSH
• germinální a embryonální nádory enzymologie   genetika   patologie   MeSH
• imunohistochemie MeSH
• kvantitativní polymerázová řetězová reakce MeSH
• lidé MeSH
• mutace * MeSH
• mutační analýza DNA metody   MeSH
• nádorové biomarkery genetika   MeSH
• nádorové buněčné linie MeSH
• nádorový supresorový protein p53 genetika   MeSH
• prospektivní studie MeSH
• proteiny WT1 genetika   MeSH
• protoonkogenní proteiny B-raf genetika   MeSH
• protoonkogenní proteiny c-kit genetika   MeSH
• retrospektivní studie MeSH
• studie proveditelnosti MeSH
• testikulární nádory enzymologie   genetika   patologie   MeSH
• vysoce účinné nukleotidové sekvenování MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The project deals with implementation of polychromatic flow cytometry and new molecular methods in diagnostic algorithm for most common hematological causes of bone marrow failure in childhood. By prospective follow-up and detailed analysis of patients with myelodysplastic syndrome and aplastic anemia we will define characteristic factors, which will contribute to classification of both diseases and distinguishing from reactive states in the future. The use of above listed techniques will lead to new information about immunopathology and pathogenesis of the diseases.

Projekt se zabývá začleněním polychromatické průtokové cytometrie a nových molekulárních metod do diagnostického algoritmu nejčastějších hematologických příčin selhání kostní dřeně v dětském věku. Prospektivním sledováním a detailní analýzou pacientů s myelodysplastickým syndromem a aplastickou anémií chceme nalézt charakteristické znaky, které do budoucna přispějí ke klasifikaci obou onemocnění a k rozlišení od reaktivních stavů. Využití výše uvedených technik povede k získání nových informací o imunopatologii a patogenezi nemoci.

• MeSH
• algoritmy MeSH
• aplastická anemie diagnóza   MeSH
• diagnostické techniky molekulární MeSH
• dítě MeSH
• imunofenotypizace MeSH
• myelodysplastické syndromy diagnóza   MeSH
• pancytopenie diagnóza   MeSH
• proteiny WT1 MeSH
• průtoková cytometrie MeSH
• vysoce účinné nukleotidové sekvenování MeSH
• vzácné nemoci diagnóza   MeSH
• Check Tag
• dítě MeSH

• Konspekt
• Biochemie. Molekulární biologie. Biofyzika
• NLK Obory
• molekulární biologie, molekulární medicína
• pediatrie
• hematologie a transfuzní lékařství
• onkologie
• vzácné nemoci
• NLK Publikační typ
• závěrečné zprávy o řešení grantu IGA MZ ČR