BACKGROUND: The detection of the EGFR T790M (T790M) mutation in non-small cell lung cancer (NSCLC) patients who progressed under treatment with first- or second-generation EGFR-tyrosine kinase inhibitors (TKIs) is important to offer a subsequent therapy with a third-generation EGFR-TKI. Liquid biopsy is a powerful tool to determine the T790M mutation status. Several liquid biopsy platforms with varying degrees of accuracy are available to test for T790M mutations, and sensitivities may differ among these methods. METHODS: As no standard exists for the testing of T790M mutation in liquid biopsy, we performed a collaborative study to describe and compare the sensitivity of different in-house liquid biopsy platforms for the detection of the T790M mutation, EGFR exon 19 deletion (del19) and EGFR L858R mutation (L858R) across multiple participating laboratories in seven Central and Eastern European countries. RESULTS: Of the 25 invited laboratories across Central and Eastern Europe, 21 centers participated and received 10 plasma samples spiked with cell-line DNA containing the T790M, del19, or L858R mutation in different concentrations. In-house PCR-based and NGS-based methods were used accordingly, and results were reported as in routine clinical practice. Two laboratories, which used the AmoyDx® EGFR 29 Mutations Detection Kit (AmoyDx) with Cobas® cfDNA Sample Preparation Kit and QX200 Droplet Digital PCR (ddPCR) with the QIAamp Circulating Nucleic Acid Kit identified all ten samples correctly. Cobas® EGFR Mutation Test v2 (Cobas), the NGS methods, and the IdyllaTM detection method used in this study performed within the known sensitivity range of each detection method. CONCLUSIONS: If a negative result was obtained from methods with lower sensitivity (e.g., Cobas), repeated liquid biopsy testing and/or tissue biopsy analysis should be performed whenever possible, to identify T790M-positive patients to allow them to receive the optimal second-line treatment with a third-generation EGFR TKI.

• Klíčová slova
• EGFR, NGS, NSCLC, PCR, T790M mutation, liquid biopsy,
• Publikační typ
• časopisecké články MeSH

Pheochromocytomas (PHEOs) and paragangliomas (PGLs) are rare, catecholamine-producing tumors that are usually sporadic. However, about 30% of these tumors have been identified as being of inherited origin. To date, nine genes have been confirmed as participating in PHEO or PGL tumorigenesis. Germline mutations were found in 100% of syndromic cases and in about 90% of patients with positive familial history. In nonsyndromic patients with apparently sporadic tumors, genetic mutations have been found in up to 27%, and genetic testing is now recommended for all patients with PHEOs and PGLs. Patients with syndromic lesions, a positive family history, or both should be tested for the appertaining gene. Recent discoveries have shown that the order of tested genes in nonsyndromic, nonfamilial cases can be based on histologic evaluation, location, and the biochemical phenotype of PHEOs and PGLs--the "rule of three." Identification of a gene mutation may lead to early diagnosis and treatment, regular surveillance, and a better prognosis for patients and their relatives.

• MeSH
• asymptomatické nemoci mortalita   MeSH
• časná detekce nádoru MeSH
• dospělí MeSH
• feochromocytom * diagnóza   genetika   mortalita   terapie   MeSH
• genetické testování * MeSH
• katecholaminy genetika   MeSH
• lidé středního věku MeSH
• lidé MeSH
• míra přežití MeSH
• mnohočetná endokrinní neoplazie typ 2A * diagnóza   genetika   mortalita   terapie   MeSH
• nádory nadledvin * diagnóza   genetika   mortalita   terapie   MeSH
• sukcinátdehydrogenasa genetika   MeSH
• výsledek terapie MeSH
• zárodečné mutace MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• katecholaminy MeSH
• sukcinátdehydrogenasa MeSH

Biallelic pathogenic GJB2 gene mutations cause pre-lingual genetic hearing loss in up to 50% of individuals with bilateral sensorineural hearing loss worldwide. Sequencing of the entire GJB2 gene-coding region in Czech patients with pre-lingual bilateral hearing loss revealed that 10.3% of Czech patients carry only one monoallelic pathogenic mutation in the coding region of the GJB2 gene, which is significantly more than the population frequency of 3.4%. The 309-kb GJB6 deletion, frequent in Spain and France, is very rare in the Czech population. In order to evaluate the impact of the IVS1 + 1 G to A splice site mutation in the non-coding part of the GJB2 gene among Czech patients, we tested all available patients with pre-lingual hearing loss with only one monoallelic mutation in the coding part of GJB2. By sequencing of the exon 1 region of the GJB2 gene and HphI restriction analysis in 20 Czech patients we identified nine patients carrying IVS1 + 1 G to A. Testing for this mutation explained deafness in 45% of Czech GJB2 monoallelic patients. This mutation represents now 4% of GJB2 pathogenic mutations in Czech patients and is the third most common GJB2 mutation found in our cohort of 242 unrelated Czech patients with prelingual hearing loss. A similar frequency may also be expected in other Central European or Slavic populations.

• MeSH
• alely MeSH
• genetické testování MeSH
• kohortové studie MeSH
• konexin 26 MeSH
• konexiny genetika   MeSH
• lidé MeSH
• místa sestřihu RNA genetika   MeSH
• mutace * MeSH
• mutační analýza DNA MeSH
• oboustranná nedoslýchavost diagnóza   epidemiologie   genetika   MeSH
• restrikční mapování MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika MeSH
• Názvy látek
• GJB2 protein, human MeSH Prohlížeč
• konexin 26 MeSH
• konexiny MeSH
• místa sestřihu RNA MeSH

Hypertrophic cardiomyopathy (HCM) is a common inherited heart disease with an estimated prevalence of up to 1 in 200 individuals. In the majority of cases, HCM is considered a Mendelian disease, with mainly autosomal dominant inheritance. Most pathogenic variants are usually detected in genes for sarcomeric proteins. Nowadays, the genetic basis of HCM is believed to be rather complex. Thousands of mutations in more than 60 genes have been described in association with HCM. Nevertheless, screening large numbers of genes results in the identification of many genetic variants of uncertain significance and makes the interpretation of the results difficult. Patients lacking a pathogenic variant are now believed to have non-Mendelian HCM and probably have a better prognosis than patients with sarcomeric pathogenic mutations. Identifying the genetic basis of HCM creates remarkable opportunities to understand how the disease develops, and by extension, how to disrupt the disease progression in the future. The aim of this review is to discuss the brief history and recent advances in the genetics of HCM and the application of molecular genetic testing into common clinical practice.

• Klíčová slova
• genetics, hypertrophic cardiomyopathy, molecular genetic testing, next-generation sequencing, pathogenic mutations, variants of uncertain significance,
• MeSH
• genetické testování * MeSH
• hypertrofická kardiomyopatie diagnóza   genetika   MeSH
• lidé MeSH
• mutace * MeSH
• sarkomery genetika   MeSH
• svalové proteiny genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• svalové proteiny MeSH

BACKGROUND: Hereditary cancer syndromes are an important subset of malignant cancers caused by pathogenic variants in one of many known cancer predisposition genes. Diagnosis of cancer predisposition is based on genetic testing using next-generation sequencing. This allows many genes to be analysed at once, increasing the number of variants identified. The correct classification of the variants found is essential for the clinical interpretation of genetic test results. PURPOSE: The aim of this study is to summarise the rules for classifying identified variants within individual laboratories and to present the process for creating a common classification. In the Czech Republic, the sharing of identified genetic variants and the development of their consensus classification among national laboratory diagnostic communities is carried out within the Czech Cancer Panel for Clinical Application (CZECANCA) consortium of scientific and diagnostic oncogenetic laboratories. Consensus for variant classification follows a defined protocol. Sharing the results and consensus classification accelerates and refines the release of genetic test results, harmonises results between laboratories and thus contributes to improving the care of individuals at high risk of cancer and their relatives.

• Klíčová slova
• clinical relevance, genetic testing, hereditary neoplastic syndromes, massively parallel sequencing, massively-parallel sequencing, national consensus, variant classification,
• MeSH
• dědičné nádorové syndromy * genetika   MeSH
• genetická predispozice k nemoci * MeSH
• genetické testování metody   MeSH
• konsensus MeSH
• lidé MeSH
• zárodečné buňky MeSH
• zárodečné mutace MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

PURPOSE: Preimplantation genetic testing for monogenic disorders (PGT-M) allows early diagnosis in embryos conceived in vitro. PGT-M helps to prevent known genetic disorders in affected families and ensures that pathogenic variants in the male or female partner are not passed on to offspring. The trend in genetic testing of embryos is to provide a comprehensive platform that enables robust and reliable testing for the causal pathogenic variant(s), as well as chromosomal abnormalities that commonly occur in embryos. In this study, we describe PGT protocol that allows direct mutation testing, haplotyping, and aneuploidy screening. METHODS: Described PGT protocol called OneGene PGT allows direct mutation testing, haplotyping, and aneuploidy screening using next-generation sequencing (NGS). Whole genome amplification product is combined with multiplex PCR used for SNP enrichment. Dedicated bioinformatic tool enables mapping, genotype calling, and haplotyping of informative SNP markers. A commercial software was used for aneuploidy calling. RESULTS: OneGenePGT has been implemented for seven of the most common monogenic disorders, representing approximately 30% of all PGT-M indications at our IVF centre. The technique has been thoroughly validated, focusing on direct pathogenic variant testing, haplotype identification, and chromosome abnormality detection. Validation results show full concordance with Sanger sequencing and karyomapping, which were used as reference methods. CONCLUSION: OneGene PGT is a comprehensive, robust, and cost-effective method that can be established for any gene of interest. The technique is particularly suitable for common monogenic diseases, which can be performed based on a universal laboratory protocol without the need for set-up or pre-testing.

• Klíčová slova
• Aneuploidy, Monogenic disorders, Next-generation sequencing, Preimplantation genetic testing,
• MeSH
• aneuploidie MeSH
• blastocysta patologie   MeSH
• genetické testování metody   MeSH
• lidé MeSH
• mutace genetika   MeSH
• preimplantační diagnóza * metody   MeSH
• těhotenství MeSH
• vysoce účinné nukleotidové sekvenování metody   MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• těhotenství MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH

Mutations in IDH1/2 genes are a marker of good prognosis for glioma patients, associated with low grade gliomas and secondary glioblastomas. Immunohistochemistry and Sanger sequencing are current standards for IDH1/2 genotyping while many other methods exist. The aim of this study was to validate Competitive amplification of differentially melting amplicons (CADMA) PCR for IDH genotyping by comparison with SNaPshot assay and two immunohistochemical methods. In our study, 87 glioma patients (46 from Olomouc and 41 from Ostrava) were analyzed. IDH1/2 mutations in native bioptical samples were analyzed at DNA level by CADMA and SNaPshot while IDH1 mutations in FFPE samples were analyzed at protein level by two IHC methods. CADMA PCR sensitivity for IDH1 was 96.4% and specificity 100% for 86 concluded samples. SNaPshot assay sensitivity was 92.9% and specificity of 100% for 85 concluded samples. IHC in the laboratory no. 2 reached sensitivity 85.7% and specificity 100% for 86 concluded samples. IHC in the laboratory no. 4 reached sensitivity of 96.4% and specificity of 79.7% in 74 concluded samples. Only one IDH2 mutation was found by SNaPshot while CADMA yielded false negative result. In conclusion, CADMA is a valid method for IDH1 p.(R132H) testing with higher sensitivity than SNaPshot assay. Also, molecular genetic methods of IDH1 testing from native samples were more robust than IHC from FFPE.

• Klíčová slova
• CADMA, Glioma, IDH1, IDH2, Mutation testing,
• MeSH
• glioblastom genetika   MeSH
• gliom genetika   MeSH
• imunohistochemie metody   MeSH
• isocitrátdehydrogenasa genetika   MeSH
• lidé MeSH
• mutace genetika   MeSH
• mutační analýza DNA metody   MeSH
• nádorové biomarkery genetika   MeSH
• nádory mozku genetika   MeSH
• senzitivita a specificita MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• isocitrátdehydrogenasa MeSH
• nádorové biomarkery MeSH

BACKGROUND: Molecular classification has brought significant changes in the management of endometrial cancer (EC). In this article, we aim to analyze our first experience with an implementation of molecular testing into daily clinical practice. MATERIALS AND METHODS: In all newly diagnosed EC, the status of mismatch repair (MMR) and p53 proteins has been evaluated immunohistochemically as a part of the routine histopathological examination since May 2021. In tumors that do not meet clinical criteria for a low risk and those with MMR deficiency or p53 mutation, the molecular genetic testing of the POLE gene is performed as well. Recommendations for adjuvant treatment or follow-up are subsequently made based on the risk of recurrence. Genetic counselling is proposed to all patients with MMR-deficient tumors or family history of cancer. RESULTS: A total of 85 patients with newly diagnosed EC between May 2021 and May 2022 were enrolled in the analysis. The median age was 66 years. The results of molecular testing were as follows: 22 (26%) MMR-deficient, 8 (9%) p53-mutated and none POLE-ultramutated of those 40 tumors with performed POLE sequencing. A total of 46 (51%) patient had a low risk, 2 (2%) intermediate, 14 (16%) high-intermediate and 20 (24%) patients had a high risk of recurrence. Advanced or metastatic diseases were diagnosed in 6 (7%) patients. The median time between surgery and multidisciplinary tumor board decision was 21 days (8-36). A total of 76 (90%) patients underwent the whole treatment plan according to the recurrence risk. At the time of analysis, the results of genetic testing were available in 18 patients and revealed 4 (22%) carriers of a pathogenic variant in any of the genes associated with Lynch syndrome. CONCLUSION: Molecular testing combining immunohistochemical analyses of MMR and p53 proteins in all newly diagnosed EC patients with sequencing analysis of POLE in those with non-low-risk disease is feasible and does not prolong the time needed for treatment decision.

• Klíčová slova
• POLE, endometrial cancer, mismatch-repair system, molecular testing, p53,
• MeSH
• diagnostické techniky molekulární MeSH
• genetické testování MeSH
• lidé MeSH
• mutace MeSH
• nádorový supresorový protein p53 * metabolismus   MeSH
• nádory endometria * genetika   terapie   MeSH
• oprava chybného párování bází DNA genetika   MeSH
• senioři MeSH
• Check Tag
• lidé MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• nádorový supresorový protein p53 * MeSH

Heterozygous germline BRCA2 mutations predispose to breast, ovarian, pancreatic and other types of cancer. The presence of a pathogenic mutation in patients or their family members warrants close surveillance or prophylactic surgery. Besides clearly pathogenic mutations, variants leading only to a single amino acid substitution are often identified. The influence of such variants on cancer risk is often unknown, making their presence a major clinical problem. When genetic methods are insufficient to classify these variants, functional assays with various cellular models are performed. We developed and applied a new syngeneic model of human cancer cells to test all variants of unknown significance in exon 18 identified by genetic testing of high-risk cancer patients in the Czech Republic, via introduction of constructs containing each of these variants into the wild-type allele of BRCA2-heterozygous DLD1 cells (BRCA2wt/Δex11). We found unaffected DNA repair function of BRCA2 in cell lines BRCA27997G>C/Δex11, BRCA28111C>T/Δex11, BRCA28149G>T/Δex11, BRCA28182G>A/Δex11, and BRCA28182G>T/Δex11, whereas the cell line BRCA28168A>G/Δex11 and the nonsense mutation carrying line BRCA28305G>T/Δex11 did affect protein function. Targeting the BRCA2 wild-type allele with a construct carrying the variant c.7988A> G resulted in incorporation exclusively into the already defective allele in all viable clones, strongly suggesting a detrimental phenotype. Our model thus offers a valuable tool for the functional evaluation of unclassified variants in the BRCA2 gene and provides a stable and distributable cellular resource for further research.

• Klíčová slova
• BRCA2, cancer biology, cancer risk, functional analysis, missense, mutation, syngeneic, transcription/repair, variants,
• MeSH
• anamnéza MeSH
• biologické modely * MeSH
• dospělí MeSH
• exony genetika   MeSH
• genetická predispozice k nemoci * MeSH
• genetické testování metody   MeSH
• hodnocení rizik metody   MeSH
• lidé středního věku MeSH
• lidé MeSH
• Markovovy řetězce MeSH
• mutace MeSH
• mutační analýza DNA metody   MeSH
• nádorové buněčné linie MeSH
• nádory diagnóza   genetika   MeSH
• protein BRCA2 genetika   MeSH
• senioři MeSH
• studie proveditelnosti MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika MeSH
• Názvy látek
• BRCA2 protein, human MeSH Prohlížeč
• protein BRCA2 MeSH

We present a boy with mild hyperglycemia detected during an upper respiratory infection. Novel splicing mutation in the intron 1 of the GCK gene (c.45+1G>A) was detected, and was subsequently confirmed in his father. This is the first case of genetically confirmed Macedonian family with MODY.

• Klíčová slova
• GCK mutation, GCK-MODY, Mild hyperglycemia,
• MeSH
• diabetes mellitus 2. typu genetika   terapie   MeSH
• glukokinasa genetika   MeSH
• glukózový toleranční test MeSH
• hyperglykemie genetika   MeSH
• lidé MeSH
• mutace * MeSH
• předškolní dítě MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• předškolní dítě MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• kazuistiky MeSH
• Geografické názvy
• Řecko MeSH
• Názvy látek
• glukokinasa MeSH