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1.
Presse Med ; 48(7-8 Pt 1): 816-824, 2019.
Artigo em Francês | MEDLINE | ID: mdl-31439443

RESUMO

Diagnosis of mature B cell malignancies is highly multidisciplinary. Biological tools provide diagnostic, prognostic and theranostic information. Biological hematology allows considering mature B cell diseases from two perspectives : cellular and molecular approaches. Cytomorphology and flow cytometry are tools from cell hematology. Conventional cytogenetics, FISH and molecular biology are tools from molecular hematology. NGS is a new technique that could dramatically change diagnostic and therapeutic management of B cell malignancies in the near future. Integration of clinical, pathological and biological data allows for personalized management of these diseases.


Assuntos
Técnicas de Laboratório Clínico/métodos , Leucemia de Células B/diagnóstico , Linfoma de Células B/diagnóstico , Integração de Sistemas , Hibridização Genômica Comparativa/métodos , Citodiagnóstico/métodos , Análise Citogenética/métodos , Análise Mutacional de DNA/métodos , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Imunofenotipagem/métodos , Hibridização in Situ Fluorescente/métodos , Leucemia de Células B/genética , Leucemia de Células B/patologia , Linfoma de Células B/genética , Linfoma de Células B/patologia , Técnicas de Diagnóstico Molecular/métodos , Imagem Multimodal/métodos , Imagem Multimodal/tendências
2.
Presse Med ; 48(7-8 Pt 1): 832-841, 2019.
Artigo em Francês | MEDLINE | ID: mdl-31444019

RESUMO

Lymphoplasmocytic lymphona with monoclonal lgM, rare. Median age at diagnosis 70 years old, frail population. Heterogenous clinic presentation. Molecular diagnosis with MYD88. Treatment required for symptomatic WM patients only. 1st line therapy: DRC. Input of targeted therapies (ibrutinib) for frail patients, maintenance effect.


Assuntos
Macroglobulinemia de Waldenstrom , Idade de Início , Idoso , Análise Mutacional de DNA/métodos , Idoso Fragilizado , Humanos , Técnicas de Diagnóstico Molecular/métodos , Terapia de Alvo Molecular/métodos , Terapia de Alvo Molecular/tendências , Fator 88 de Diferenciação Mieloide/genética , Pirazóis/uso terapêutico , Pirimidinas/uso terapêutico , Macroglobulinemia de Waldenstrom/diagnóstico , Macroglobulinemia de Waldenstrom/epidemiologia , Macroglobulinemia de Waldenstrom/genética , Macroglobulinemia de Waldenstrom/terapia
3.
J Cancer Res Clin Oncol ; 145(9): 2325-2333, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31317326

RESUMO

PURPOSE: Nodal positive lung adenocarcinoma includes wide range of survival. Several methods for the classification of nodal-positive lung cancer have been proposed. However, classification considering the impact of targetable genetic variants are lacking. The possibility of genetic variants for the better stratification of nodal positive lung adenocarcinoma was estimated. METHODS: Mutations of 36 genes between primary sites and metastatic lymph nodes (LNs) were compared using next-generation sequencing. Subsequently, mutations in EGFR and BRAF, rearrangements in ALK and ROS1 were evaluated in 69 resected pN1-2M0 adenocarcinoma cases. Recurrence-free survival (RFS), post-recurrence survival (PRS), and overall survival (OS) were evaluated with respect to targetable variants and tyrosine kinase inhibitor (TKI) therapy after recurrence. RESULTS: About 90% of variants were shared and allele frequencies were similar between primary and metastatic sites. In 69 pN1-2M0 cases, EGFR/ALK were positive in primary sites of 39 cases and same EGFR/ALK variants were confirmed in metastatic LNs of 96.7% tissue-available cases. Multivariate analyses indicated positive EGFR/ALK status was associated with worse RFS (HR 2.366; 95% CI 1.244-4.500; P = 0.009), and PRS was prolonged in cases receiving TKI therapy (no post-recurrence TKI therapies, HR 3.740; 95% CI 1.449-9.650; P = 0.006). OS did not differ with respect to targetable variants or TKI therapy. CONCLUSIONS: Cases harbouring targetable genetic variants had a higher risk of recurrence, but PRS was prolonged by TKI therapy. Classification according to the targetable genetic status provides a basis for predicting recurrence and determining treatment strategies after recurrence.


Assuntos
Adenocarcinoma de Pulmão/diagnóstico , Neoplasias Pulmonares/diagnóstico , Pulmão/metabolismo , Linfonodos/metabolismo , Mutação , Transcriptoma/genética , Adenocarcinoma de Pulmão/genética , Adenocarcinoma de Pulmão/patologia , Idoso , Idoso de 80 Anos ou mais , Análise Mutacional de DNA/métodos , Feminino , Regulação Neoplásica da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Pulmão/patologia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Linfonodos/patologia , Metástase Linfática , Masculino , Análise em Microsséries/métodos , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/genética , Prognóstico , Estudos Retrospectivos
4.
J Cancer Res Clin Oncol ; 145(8): 1977-1986, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31309300

RESUMO

CONTEXT: Parathyroid carcinoma (PC) is a rare endocrine malignancy with no approved systemic therapies for unresectable locally invasive or distant metastatic disease. Understanding the molecular changes in advanced PC can provide better understanding of this disease and potentially help directing targeted therapy. OBJECTIVE: To evaluate tumor-specific genetic changes using next-generation sequencing (NGS) panels. DESIGN: All patients with advanced PC were tested for hot-spot panels using NGS panels including a 50-gene panel, a 409-gene panel if the standard 50-gene panel (Ion Torrent, Life Technology) was negative or a FoundationOne panel. SETTING: The University of Texas MD Anderson Cancer Center, Houston, Texas, USA. PATIENTS OR OTHER PARTICIPANTS: 11 patients with advanced PC were selected to undergo molecular testing. MAIN OUTCOME MEASURE(S): Genetic profiles of advanced PC. RESULTS: Among the 11 patients, 4 patients had the 50-gene panel only, 6 had 409-gene panel after a negative 50-gene panel and 1 had FoundationOne. One patient who had 50-gene panel only also had his metastatic site (esophagus) of his tumor tested with FoundationOne. The most common mutations identified were in the PI3 K (PIK3CA, TSC1 and ATM) (4/11 patients) and TP53 (3/11) pathways. Genes not previously reported to be mutated in PC included: SDHA, TERT promoter and DICER1. Actionable mutations were found in 54% (6/11) of the patients. CONCLUSIONS: Mutational profiling using NGS panels in advanced PC has yielded important potentially targetable genetic alterations. Larger studies are needed to identify commonly mutated genes in advanced PC patients. Development of novel therapies targeting these cellular pathways should be considered.


Assuntos
Carcinoma/genética , Perfilação da Expressão Gênica , Técnicas de Diagnóstico Molecular/métodos , Monitorização Fisiológica/métodos , Neoplasias das Paratireoides/genética , Adulto , Idoso , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Carcinoma/diagnóstico , Carcinoma/patologia , Carcinoma/terapia , Análise Mutacional de DNA/métodos , Progressão da Doença , Feminino , Seguimentos , Perfilação da Expressão Gênica/métodos , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Masculino , Pessoa de Meia-Idade , Técnicas de Diagnóstico Molecular/tendências , Terapia de Alvo Molecular/métodos , Terapia de Alvo Molecular/tendências , Neoplasias das Paratireoides/diagnóstico , Neoplasias das Paratireoides/patologia , Neoplasias das Paratireoides/terapia
5.
Chem Commun (Camb) ; 55(58): 8466-8469, 2019 Jul 25.
Artigo em Inglês | MEDLINE | ID: mdl-31265022

RESUMO

We presented a branch migration based PCR in which a branch migration blocker was introduced to selectively reduce the amplification efficiency of the wild-type target and enrich the mutant-type target. The low-abundance mutations could be enriched and then detected by high resolution melting, Sanger sequencing or fluorescent DNA probe.


Assuntos
Análise Mutacional de DNA/métodos , DNA/análise , DNA/genética , Reação em Cadeia da Polimerase/métodos , Sequência de Bases , Sondas de DNA/química , Sondas de DNA/genética , DNA Polimerase Dirigida por DNA/química , Fluorescência , Corantes Fluorescentes/química , Genes , Humanos , Limite de Detecção , Mutação , Hibridização de Ácido Nucleico
6.
Nat Commun ; 10(1): 2969, 2019 07 05.
Artigo em Inglês | MEDLINE | ID: mdl-31278357

RESUMO

Analysis of mutational signatures is becoming routine in cancer genomics, with implications for pathogenesis, classification, prognosis, and even treatment decisions. However, the field lacks a consensus on analysis and result interpretation. Using whole-genome sequencing of multiple myeloma (MM), chronic lymphocytic leukemia (CLL) and acute myeloid leukemia, we compare the performance of public signature analysis tools. We describe caveats and pitfalls of de novo signature extraction and fitting approaches, reporting on common inaccuracies: erroneous signature assignment, identification of localized hyper-mutational processes, overcalling of signatures. We provide reproducible solutions to solve these issues and use orthogonal approaches to validate our results. We show how a comprehensive mutational signature analysis may provide relevant biological insights, reporting evidence of c-AID activity among unmutated CLL cases or the absence of BRCA1/BRCA2-mediated homologous recombination deficiency in a MM cohort. Finally, we propose a general analysis framework to ensure production of accurate and reproducible mutational signature data.


Assuntos
Análise Mutacional de DNA/normas , Leucemia Linfocítica Crônica de Células B/genética , Leucemia Mieloide Aguda/genética , Mieloma Múltiplo/genética , Proteína BRCA1/genética , Proteína BRCA2/genética , Biologia Computacional/métodos , Biologia Computacional/normas , Análise Mutacional de DNA/métodos , Conjuntos de Dados como Assunto , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Sequenciamento de Nucleotídeos em Larga Escala/normas , Humanos , Mutação , Guias de Prática Clínica como Assunto , Sequenciamento Completo do Genoma/métodos , Sequenciamento Completo do Genoma/normas
7.
J Clin Pathol ; 72(11): 748-754, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31262952

RESUMO

AIMS: Hereditary leiomyomatosis and renal cell carcinoma (HLRCC) is a newly recognised entity in the WHO 2016 classification defined as the germline mutation of FH gene. Fumaratehydratase-deficient renal cell carcinoma (FH-deficient RCC) is recommended for tumours with FH deficiency but lacking of genetic evidences of FH germline mutation. In this study, we described the clinicopathological and molecular changes of 13 FH-deficient RCCs. METHODS AND RESULTS: Histology features, clinicopathological data, radiology performance and outcomes were collected for each patient. Next-generation sequencing and DNA sequencing of FH gene were performed to examine FH mutations. The patient group included five females and eight males. Different morphological patterns of papillary, nested, adenoid, foam adenoid, cribriform, tubular, tubulocystic, cystic and loose oedema stroma were observed. Except typical big nuclei with or without eosinophilic nucleoli and perinucleolar halos, raisin-like, hobnail-like and even low-grade nuclei were also observed in these tumours. Eleven cases with high-grade nuclei showed disease progression or death, but no disease progression was detected in two cases with low-grade nuclei and eosinophilic cytoplasm. FH expression was absent in tumour cells except for case 11. Next-generation sequencing and DNA sequencing verified seven FH germline mutations and four somatic mutations out of 13 cases. CONCLUSIONS: FH-deficient RCC is a rare renal tumour and has a wide morphological spectrum. Most of the tumours had high-grade nuclei and were aggressive. However, we observed a morphological subtype of FH-deficient RCC with low-grade nuclei and eosinophilic cytoplasm, which might mainly occur in young women and show a relatively good prognosis.


Assuntos
Biomarcadores Tumorais/genética , Carcinoma de Células Renais/genética , Fumarato Hidratase/genética , Mutação em Linhagem Germinativa , Neoplasias Renais/genética , Adolescente , Adulto , Idoso , Biomarcadores Tumorais/deficiência , Biópsia , Carcinoma de Células Renais/enzimologia , Carcinoma de Células Renais/patologia , Carcinoma de Células Renais/terapia , Núcleo Celular/patologia , Citoplasma/patologia , Análise Mutacional de DNA/métodos , Feminino , Fumarato Hidratase/deficiência , Predisposição Genética para Doença , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Imuno-Histoquímica , Neoplasias Renais/enzimologia , Neoplasias Renais/patologia , Neoplasias Renais/terapia , Masculino , Pessoa de Meia-Idade , Gradação de Tumores , Fenótipo
8.
Crit Rev Oncol Hematol ; 141: 146-152, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31301542

RESUMO

Clinical response to checkpoint inhibitors-based (CPIs) therapies can vary among tumor types and between patients. This led to a significant amount of pre-clinical and clinical research into biomarker identification. Biomarkers have been found to cover both the tumor itself and the tumor microenvironment. Entering host-related parameters into the equation should provide a valuable strategy for identifying not only factors predictive of treatment efficacy but also of treatment-related toxicity. It is clear that germline variants can offer efficient and easily-assessable indicators (blood DNA) to enlarge the spectrum of predictive markers for CPI-based treatment. A major issue concerns the real functional significance of the reported single-nucleotide polymorphisms (SNPs) linked to CPI-treatment outcome. Powered calculations should lead to an optimal trade-off between sample size and allele frequency. New molecular technologies and new analytical methods should provide opportunities to bridge the knowledge gap between SNP-CPI treatment associations and the functional impact of these SNPs.


Assuntos
Resistencia a Medicamentos Antineoplásicos/genética , Mutação em Linhagem Germinativa/fisiologia , Imunogenética/métodos , Imunoterapia , Neoplasias/diagnóstico , Neoplasias/terapia , Análise Mutacional de DNA/métodos , Resistencia a Medicamentos Antineoplásicos/imunologia , Humanos , Fatores Imunológicos/uso terapêutico , Imunoterapia/métodos , Neoplasias/imunologia , Testes Farmacogenômicos/métodos , Prognóstico , Resultado do Tratamento , Microambiente Tumoral/genética , Microambiente Tumoral/imunologia
9.
Zhonghua Er Ke Za Zhi ; 57(6): 477-482, 2019 Jun 02.
Artigo em Chinês | MEDLINE | ID: mdl-31216807

RESUMO

Objective: To analyze the clinical manifestations and gene variations of tumor necrosis factor receptor-associated periodic syndrome (TRAPS). Methods: Clinical data and gene testing of four children and three adult relatives in a family from Puning, Guangdong were retrospectively analyzed. CD4(+)T cells, CD8(+)T cells, B cells, monocytes and NK cells were assessed by flow cytometry. Plasma level of TNFR receptors were assessed by enzyme linked immunosorbent assay (ELISA). TNFRSF1A gene variation was identified by second generation sequencing. Swiss-Model was used to analyze the potential impact of TNFRSF1A gene variation on its protein tertiary structure. Results: For all the patients,periodic fever was the main clinical feature,combined with arthralgia,myalgia,multiple serositis,periorbital edema and migratory cutaneous rash,accompanied with elevated level of acute-phase reactants and increased white blood cell counts during each episode. This disease was found in both gender and every generation in this family. The median age of onset was 2 years, ranging from 6 months to 30 years. The plasma level of TNFR1 of the patients range from 0 to 12.4 ng/L,which was lower than that of the normal controls range from 18.0~22.2 ng/L,while the level of TNFR2 was normal. Also, the numbers of T cells, B cells and monocytes were within normal range; however,number of NK cells in the patients (0.070±0.034) was lower than that in the normal controls (0.152±0.122). The TNFRSF1A variation,located in exon 3: c.295T>A (p.C99S),was found in the proband as well as the other 6 family members,which could induce change of the side chain of amino acid according to the prediction of the three-dimensional structure,subsequently affecting the binding to the receptor. Conclusions: TRAPS is characterized by periodic fever,arthralgia,myalgia,multiple serositis,periorbital edema and migratory cutaneous rash,with a significant decrease in plasma level of TNFR1 and NK cells. The gene sequencing analysis revealed a pathogenic variation in TNFRSF1A gene.


Assuntos
Análise Mutacional de DNA/métodos , Febre Familiar do Mediterrâneo/genética , Receptores Tipo I de Fatores de Necrose Tumoral/genética , Receptores do Fator de Necrose Tumoral/genética , Adulto , Criança , Pré-Escolar , Ensaio de Imunoadsorção Enzimática , Éxons , Febre Familiar do Mediterrâneo/diagnóstico , Febre , Citometria de Fluxo , Doenças Hereditárias Autoinflamatórias , Humanos , Mutação , Receptores Tipo I de Fatores de Necrose Tumoral/sangue , Estudos Retrospectivos , Análise de Sequência de DNA
10.
Nat Commun ; 10(1): 2750, 2019 06 21.
Artigo em Inglês | MEDLINE | ID: mdl-31227714

RESUMO

Understanding the clonal architecture and evolutionary history of a tumour poses one of the key challenges to overcome treatment failure due to resistant cell populations. Previously, studies on subclonal tumour evolution have been primarily based on bulk sequencing and in some recent cases on single-cell sequencing data. Either data type alone has shortcomings with regard to this task, but methods integrating both data types have been lacking. Here, we present B-SCITE, the first computational approach that infers tumour phylogenies from combined single-cell and bulk sequencing data. Using a comprehensive set of simulated data, we show that B-SCITE systematically outperforms existing methods with respect to tree reconstruction accuracy and subclone identification. B-SCITE provides high-fidelity reconstructions even with a modest number of single cells and in cases where bulk allele frequencies are affected by copy number changes. On real tumour data, B-SCITE generated mutation histories show high concordance with expert generated trees.


Assuntos
Evolução Clonal/genética , Biologia Computacional/métodos , Análise Mutacional de DNA/métodos , Modelos Genéticos , Neoplasias/genética , Algoritmos , Conjuntos de Dados como Assunto , Feminino , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Mutação , Filogenia , Análise de Célula Única/métodos , Software
11.
BMC Med Genet ; 20(1): 106, 2019 06 13.
Artigo em Inglês | MEDLINE | ID: mdl-31196117

RESUMO

BACKGROUND: Oculocutaneous albinism (OCA) is a human autosomal-recessive hypopigmentation disorder with hypopigmentation in the skin, hair, and eyes. OCA1 and OCA2 are caused by mutations of the TYR and OCA2 genes, respectively, which are responsible for most oculocutaneous albinism. However, the incidence of oculocutaneous albinism patients in Guangxi remains unclear. METHODS: To evaluate the molecular basis of oculocutaneous albinism in thirty-six patients in Guangxi, China. Peripheral venous blood samples were collected from these unrelated patients. The TYR and OCA2 genes of all individuals were analyzed by direct DNA sequencing and the sequences compared with are reference database and bioinformatics analysis. RESULTS: Among the 36 OCA patients, 8(22.2%) were found mutations on TYR gene, 28 (77.8%) on OCA2. And we identified Twenty-seven different TYR and OCA2 mutations in these patients, including one novel TYR framshift mutation c.561_562insTTATTATGTGTCAAATTATCCCCCA, three novel OCA2 mutations: one nonsense mutation c.2195C > G(p.S732X), one deletation mutation(c.1139-1141delTGG), one missense mutations c.2495A > C(p.H832P). The population screening and the bioinformatic analysis to determined the effects of the mutations, which revealed these four novel mutations were pathogenic. CONCLUSIONS: This study expands the mutation spectrum of oculocutaneous albinism. Four novel mutational alleles c.1139-1141delTGG, c.1832 T > C and c.2195C > G and of the OCA2 gene and c.561_562insTTATTATGTGTCAAATTATCCCCCA of TYR were associated with OCA. The genotype-phenotype correlations suggest that molecular diagnosis is more accurate and important in OCA.


Assuntos
Albinismo Oculocutâneo/genética , Análise Mutacional de DNA/métodos , Predisposição Genética para Doença/genética , Mutação , Adolescente , Adulto , Albinismo Oculocutâneo/diagnóstico , Albinismo Oculocutâneo/etnologia , Grupo com Ancestrais do Continente Asiático/genética , Sequência de Bases , Criança , Pré-Escolar , China , Feminino , Estudos de Associação Genética , Predisposição Genética para Doença/etnologia , Humanos , Lactente , Recém-Nascido , Masculino , Proteínas de Membrana Transportadoras/genética , Monofenol Mono-Oxigenase/genética
12.
J Cancer Res Clin Oncol ; 145(8): 2071-2082, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31154543

RESUMO

PURPOSE: Fluorescence in situ hybridization (FISH) using tumor tissue is the gold standard for detection of anaplastic lymphoma kinase (ALK) rearrangement in non-small cell lung cancer (NSCLC). However, this method often is not repeatable due to difficulties in the acquisition of tumor tissues. Blood-based liquid biopsy using reverse transcription polymerase chain reaction (RT-PCR) is expected to be useful to overcome this limitation. Here, we investigated the feasibility of liquid biopsy using plasma and platelets for detection of ALK rearrangement and prediction of ALK inhibitor treatment outcomes. METHODS: ALK-FISH assays were performed in 1128 tumor specimens of NSCLC between January 2015 and June 2018. We retrospectively analyzed formalin-fixed paraffin-embedded (FFPE) tissues from previously confirmed FISH-positive (n = 199) and -negative (n = 920) cases. We recruited patients who had available tissue specimens and agreed to venous sampling. RNA was extracted from FFPE blocks, plasma, and platelets. Fusion RNA of echinoderm microtubule-associated protein-like 4 (EML4)-ALK was detected by quantitative PCR. RESULTS: Thirty-three FISH-positive and 28 FISH-negative patients were enrolled. In validation, data compared with FISH, RT-PCR using FFPE tissues showed 54.5% sensitivity, 78.6% specificity, and 75.5% accuracy. Liquid biopsy had higher sensitivity (78.8%), specificity (89.3%) and accuracy (83.6%). Higher positivity for liquid biopsy was shown in subgroups with delayed (≥ 6 months from diagnosis) blood sampling (plasma, 85.7%; platelets, 87.0%). In 26 patients treated with crizotinib, the platelet-positive subgroup showed longer median duration of treatment (7.2 versus 1.5 months), longer median progression-free survival (5.7 months versus 1.7 months), a higher overall response rate (70.6% versus 11.1%), and a higher disease control rate (88.2% versus 44.4%) than the platelet-negative subgroup. CONCLUSION: Liquid biopsy could have applications in the diagnosis of ALK-positive NSCLC, even when using RT-PCR, and platelets can be useful for predicting treatment outcomes of ALK inhibitors.


Assuntos
Quinase do Linfoma Anaplásico/genética , Plaquetas/química , Carcinoma Pulmonar de Células não Pequenas/diagnóstico , Análise Mutacional de DNA/métodos , Neoplasias Pulmonares/diagnóstico , Plasma/química , Adulto , Idoso , Idoso de 80 Anos ou mais , Análise Química do Sangue/métodos , Plaquetas/patologia , Carcinoma Pulmonar de Células não Pequenas/sangue , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , DNA de Neoplasias/análise , Resistencia a Medicamentos Antineoplásicos/genética , Estudos de Viabilidade , Feminino , Humanos , Hibridização in Situ Fluorescente , Biópsia Líquida , Neoplasias Pulmonares/sangue , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Masculino , Pessoa de Meia-Idade , Mutação , Células Neoplásicas Circulantes/patologia , Plasma/citologia , Inibidores de Proteínas Quinases/uso terapêutico , Estudos Retrospectivos , Sensibilidade e Especificidade
13.
Crit Rev Oncol Hematol ; 141: 36-42, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31212145

RESUMO

Liquid biopsy can quantify and qualify cell-free (cfDNA) and tumour-derived (ctDNA) DNA fragments in the bloodstream. CfDNA quantification and mutation analysis can be applied to diagnosis, follow-up and therapeutic management as novel oncologic biomarkers. However, some tumor-types release a low amount of DNA into the bloodstream, hampering diagnosis through standard liquid biopsy procedures. Several tumors, as such as brain, kidney, prostate, and thyroid cancer, are in direct contact with other body fluids and may be alternative sources for cfDNA and ctDNA. Non-blood sources of cfDNA/ctDNA useful as novel oncologic biomarkers include cerebrospinal fluids, urine, sputum, saliva, pleural effusion, stool and seminal fluid. Seminal plasma cfDNA, which can be analyzed with cost-effective procedures, may provide powerful information capable to revolutionize prostate cancer (PCa) patient diagnosis and management. In the near future, cfDNA analysis from non-blood biological liquids will become routine clinical practice for cancer patient diagnosis and management.


Assuntos
Biomarcadores Tumorais/genética , Ácidos Nucleicos Livres/isolamento & purificação , Perfilação da Expressão Gênica/métodos , Oncologia/métodos , Neoplasias/diagnóstico , Patologia Clínica/métodos , Biomarcadores Tumorais/análise , Biomarcadores Tumorais/isolamento & purificação , Ácidos Nucleicos Livres/análise , DNA Tumoral Circulante/análise , DNA Tumoral Circulante/isolamento & purificação , DNA Tumoral Circulante/urina , Análise Mutacional de DNA/métodos , Fezes/química , Feminino , Humanos , Biópsia Líquida , Masculino , Oncologia/tendências , Neoplasias/genética , Neoplasias/patologia , Neoplasias/urina , Neoplasias da Próstata/diagnóstico , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/terapia , Urinálise/métodos
14.
Genet Test Mol Biomarkers ; 23(6): 401-408, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31161821

RESUMO

Background and Aims: The genetic spectrum underlying familial hypercholesterolemia (FH) remains unclear, especially in northeastern China. The aim of this study was to delineate the FH genetic spectrum and identify specific characteristics of FH patients in this region. Materials and Methods: The family history, personal medical history, and lifestyle habits of two unrelated patients clinically diagnosed with homozygous FH were recorded. DNA samples of the patients and their relatives were subjected to a newly designed next-generation sequencing panel using an Illumina Miseq platform. Detected variants were annotated and functionally predicted with in silico algorithms, and protein structures were modeled. Results: The patients' cholesterol levels were effectively reduced to 33.8% and 17.2% of the original level under conventional ezetimibe and statin treatment. Two pathogenic mutations, W483X and the novel mutation W483G, in the low-density lipoprotein receptor (LDLR) gene were identified. Both patients were heterozygous for the respective mutations. Under a high cholesterol/carbohydrate diet, these mutations could trigger a severe FH phenotype, but both patients responded well to regular medical treatments and dietary control. The W483X mutation results in a premature stop codon, leading to incomplete protein formation. Although the W483G mutation results in translation of the complete protein with no apparent structural difference, it still led to a severe FH phenotype similar to W483X. Conclusions: Identification of the novel W483G mutation expands the genetic spectrum of FH. Both mutations cause a severe FH phenotype under certain conditions, suggesting that W483 is important for LDLR function, highlighting potential targets for genetic screening or drug development.


Assuntos
Hiperlipoproteinemia Tipo II/genética , Receptores de LDL/genética , Adulto , Grupo com Ancestrais do Continente Asiático/genética , Criança , Colesterol/genética , LDL-Colesterol/genética , Análise Mutacional de DNA/métodos , Erros de Diagnóstico , Família , Feminino , Heterozigoto , Homozigoto , Humanos , Masculino , Pessoa de Meia-Idade , Mutação , Linhagem
15.
BMC Med Genet ; 20(1): 88, 2019 05 22.
Artigo em Inglês | MEDLINE | ID: mdl-31117962

RESUMO

BACKGROUND: Succinic semialdehyde dehydrogenase (SSADH) deficiency is a rare autosomal recessively-inherited defect of γ-aminobutyric acid (GABA) metabolism. The absence of SSADH, which is encoded by aldehyde dehydrogenase family 5 member A1 (ALDH5A1) gene, leads to the accumulation of GABA and γ-hydroxybutyric acid (GHB). Few cases with SSADH deficiency were reported in China. CASE PRESENTATION: In this study, four Chinese patients were diagnosed with SSADH deficiency in Tianjin Children's Hospital. We conducted a multidimensional analysis with magnetic resonance imaging (MRI) of the head, semi quantitative detection of urine organic acid using gas chromatography-mass spectrometry, and analysis of ALDH5A1 gene mutations. Two of the patients were admitted to the hospital due to convulsions, and all patients were associated with developmental delay. Cerebral MRI showed symmetrical hyperintense signal of bilateral globus pallidus and basal ganglia in patient 1; hyperintensity of bilateral frontal-parietal lobe, widened ventricle and sulci in patient 2; and widened ventricle and sulci in patient 4. Electroencephalogram (EEG) revealed the background activity of epilepsy in patient 1 and the disappearance of sleep spindle in patient 2. Urine organic acid analysis revealed elevated GHB in all the patients. Mutational analysis, which was performed by sequencing the 10 exons and flanking the intronic regions of ALDH5A1 gene for all the patients, revealed mutations at five sites. Two cases had homozygous mutations with c.1529C > T and c.800 T > G respectively, whereas the remaining two had different compound heterozygous mutations including c.527G > A/c.691G > A and c.1344-2delA/c.1529C > T. Although these four mutations have been described previously, the homozygous mutation of c.800 T > G in ALDH5A1 gene is a novel discovery. CONCLUSION: SSADH deficiency is diagnosed based on the elevated GHB and 4, 5DHHA by urinary organic acid analysis. We describe a novel mutation p.V267G (c.800 T > G) located in the NAD binding domain, which is possibly crucial for this disease's severity. Our study expands the mutation spectrum of ALDH5A1 and highlights the importance of molecular genetic evaluation in patients with SSADH deficiency.


Assuntos
Erros Inatos do Metabolismo dos Aminoácidos/genética , Análise Mutacional de DNA/métodos , Deficiências do Desenvolvimento/genética , Mutação , Succinato-Semialdeído Desidrogenase/deficiência , Succinato-Semialdeído Desidrogenase/genética , Erros Inatos do Metabolismo dos Aminoácidos/diagnóstico por imagem , Erros Inatos do Metabolismo dos Aminoácidos/etnologia , Grupo com Ancestrais do Continente Asiático/genética , Pré-Escolar , China , Deficiências do Desenvolvimento/diagnóstico por imagem , Deficiências do Desenvolvimento/etnologia , Feminino , Humanos , Lactente , Imagem por Ressonância Magnética/métodos , Masculino , Succinato-Semialdeído Desidrogenase/metabolismo
16.
J Clin Pathol ; 72(9): 609-614, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31110050

RESUMO

AIM: The rapid and fully automated Idylla EGFR Mutation Assay has been specifically designed to process formalin-fixed, paraffin-embedded sections without requiring preliminary DNA extraction. This study evaluates whether this approach can also process archival smears from patients with non-small cell lung cancer (NSCLC) by scraping the stained cellular material directly into the cartridge. METHODS: The study was divided into two parts. In the first part, we carried out Idylla EGFR Mutation Assay on archival stained smears from 39 patients with NSCLC. Among these, 14 cases harboured a mutation in either exon 19 (n=11) or exon 21 (n=3), previously detected on DNA extracts by fragment length and TaqMan assays. In the second part, we evaluated whether de-staining of the smears could reduce background fluorescence. RESULTS: The Idylla EGFR Mutation Assay confirmed the presence of EGFR mutation in 11 instances (78.6%). However, concordance was higher for exon 19 deletions (10/11) than for exon 21 p.L858R assessments. Raw data showed a high background fluorescence in channel 2, where the EGFR exon 21 p.L858R mutation was detected. This interference, due to dye residues from the original staining, was partially reduced by de-staining the cytological material. CONCLUSIONS: Our data, although preliminary, show that the Idylla EGFR Mutation Assay can reliably process most archival smears without requiring preliminary DNA extraction. Results may be further improved by de-staining the cellular material before insertion into the cartridge.


Assuntos
Biomarcadores Tumorais/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Análise Mutacional de DNA/métodos , Neoplasias Pulmonares/genética , Mutação , Manejo de Espécimes/métodos , Automação Laboratorial , Carcinoma Pulmonar de Células não Pequenas/enzimologia , Carcinoma Pulmonar de Células não Pequenas/patologia , Receptores ErbB/genética , Éxons , Estudos de Viabilidade , Predisposição Genética para Doença , Humanos , Neoplasias Pulmonares/enzimologia , Neoplasias Pulmonares/patologia , Fenótipo , Projetos Piloto , Valor Preditivo dos Testes , Dados Preliminares , Espectrometria de Fluorescência
17.
J Clin Neurosci ; 66: 187-190, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31088771

RESUMO

Pantothenate kinase-associated neurodegeneration (PKAN) is linked to brain iron accumulation caused by PANK2 gene mutation. Despite the importance of genetic testing to confirm PKAN and identify at risk parents, genetic screening is financially burdensome for developing countries like Thailand. Because genetic screeners are expensive and not reimbursed by the universal health care coverage system, they are not typically performed. To investigate clinical symptoms, radiological findings and mutation analysis for patients based in Thailand with unknown genetic status but suspected PKAN based on clinical symptoms. Genetic testing was performed for cases suspected for PKAN and their biological parents by direct genomic sequencing of PANK2 at Maharat Nakhon Ratchasima Hospital during 2017-2018. Clinical evaluation and documentation were performed by pediatric neurologists. Five children had classical onset form of PKAN. Most presented with gait dystonia. Three patients diagnosed after 4 years showed the eye-of-the-tiger sign in their brain MRI, whereas two younger patients revealed only isolated hyperintensity bilateral globus pallidus. However, PANK2 mutations were identified in all cases: the most common mutation was c.982-1G>C. This mutation was detected in four unrelated individuals but not reported in other studies. Genetic testing is recommended to confirm diagnoses in cases with supporting clinical features of PKAN with or without the classical 'eye-of-the-tiger-sign'. A novel PANK2 mutation (c.982-1G>C) was identified in South East Asian populations based in Thailand, suggesting that this genetic variant is a founder genotype in this population. Moreover, genetic diagnosis is helpful to provide appropriate genetic counseling to families.


Assuntos
Grupo com Ancestrais do Continente Asiático/etnologia , Grupo com Ancestrais do Continente Asiático/genética , Mutação/genética , Neurodegeneração Associada a Pantotenato-Quinase/etnologia , Neurodegeneração Associada a Pantotenato-Quinase/genética , Fosfotransferases (Aceptor do Grupo Álcool)/genética , Criança , Pré-Escolar , Análise Mutacional de DNA/métodos , Feminino , Testes Genéticos/métodos , Globo Pálido/diagnóstico por imagem , Humanos , Imagem por Ressonância Magnética , Masculino , Neurodegeneração Associada a Pantotenato-Quinase/diagnóstico por imagem , Tailândia/etnologia
18.
BMC Cancer ; 19(1): 457, 2019 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-31092228

RESUMO

BACKGROUND: In the past decade, systematic and comprehensive analyses of cancer genomes have identified cancer driver genes and revealed unprecedented insight into the molecular mechanisms underlying the initiation and progression of cancer. These studies illustrate that although every cancer has a unique genetic make-up, there are only a limited number of mechanisms that shape the mutational landscapes of cancer genomes, as reflected by characteristic computationally-derived mutational signatures. Importantly, the molecular mechanisms underlying specific signatures can now be dissected and coupled to treatment strategies. Systematic characterization of mutational signatures in a cancer patient's genome may thus be a promising new tool for molecular tumor diagnosis and classification. RESULTS: In this review, we describe the status of mutational signature analysis in cancer genomes and discuss the opportunities and relevance, as well as future challenges, for further implementation of mutational signatures in clinical tumor diagnostics and therapy guidance. CONCLUSIONS: Scientific studies have illustrated the potential of mutational signature analysis in cancer research. As such, we believe that the implementation of mutational signature analysis within the diagnostic workflow will improve cancer diagnosis in the future.


Assuntos
Análise Mutacional de DNA/métodos , Neoplasias/diagnóstico , Humanos , Técnicas de Diagnóstico Molecular , Neoplasias/genética , Sequenciamento Completo do Genoma
19.
In Vivo ; 33(3): 945-954, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31028221

RESUMO

BACKGROUND/AIM: KRAS is one of the frequently mutated genes in human cancers and often relates with drug resistance and poor prognosis. PANAMutyper™ is a novel technology that integrates PNAClamp™ and PANA S-Melting™. In the present study, PANAMutyper™ and PNAClamp™ were compared for the detection of KRAS mutations using different samples of patients with malignant pleural effusion. PATIENTS AND METHODS: A total of 103 patients (including 56 lung adenocarcinoma, 10 lung squamous carcinoma, 17 small cell lung cancer, 3 large cell lung cancer, 3 stomach cancer, 2 ovarian cancer, and others) with malignant pleural effusion were investigated using matched tumor tissue, cell block, and pleural effusion samples. The diagnostic performance of these two methods was compared. RESULTS: KRAS mutations were detected in 18 (17.5%) of 103 patients using tissue, cell block, and pleural effusion samples. All 18 patients with KRAS mutations were detected by PANAMutyper™ using any sample type, however, only 7 cases were detected by PNAClamp™. Among the subtypes of KRAS mutations, substitution in codon 12, 35G>T was the most frequent, followed by substitution in codon 12, 35G>A and codon 12, 34G>A. In pleural effusion specimens, PANAMutyper™ showed a better diagnostic performance compared to PNAClamp™. CONCLUSION: PANAMutyper™ had a diagnostic superiority for the detection of KRAS mutations in patients with malignant pleural effusion compared to PNAClamp™, although there was a concordance between PANAMutyper™ and PNAClamp™ results. Therefore, PANAMutyper™ can be used for a more sensitive and accurate detection of KRAS mutations.


Assuntos
Análise Mutacional de DNA/métodos , Mutação , Derrame Pleural Maligno/diagnóstico , Derrame Pleural Maligno/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Alelos , Linhagem Celular Tumoral , Análise Mutacional de DNA/normas , Feminino , Frequência do Gene , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Sensibilidade e Especificidade
20.
BMC Cancer ; 19(1): 296, 2019 Apr 02.
Artigo em Inglês | MEDLINE | ID: mdl-30940100

RESUMO

BACKGROUND: High-grade serous ovarian cancer is a detrimental disease. Treatment options in patients with a recurrent disease are dependent on BRCA1/2 mutation status since only patients with known BRCA mutation are eligible for treatment with poly(ADP-ribose) polymerase inhibitors (PARPi). The aim of this study was to compare concordance of BRCA mutation analyses from cytological samples (CS) with BRCA mutation analyses from histological formalin fixed paraffin embedded (FFPE) samples. METHODS: Mutation analysis of BRCA1 and BRCA2 genes was performed in 44 women diagnosed with primary or recurrent high-grade ovarian cancer from three different samples: blood, cytological sample (ascites, pleural effusion and enlarged lymph nodes) and tumor tissue. Results from all three samples were compared. RESULTS: Among 44 patients, there were 15 germline mutations and two somatic mutations. A 100% concordance was found between cytological and histologic samples. CONCLUSION: There is a 100% concordance in BRCA mutation testing between cytological and histologic samples. BRCA mutation testing from CS could replace testing from FFPE tissue in clinical decision making in ovarian cancer patients. TRIAL REGISTRATION: The study was retrospectively registered at ISRCTN registry on 24/11/2015 - ISRCTN42408038 .


Assuntos
Proteína BRCA1/genética , Proteína BRCA2/genética , Análise Mutacional de DNA/métodos , Mutação , Neoplasias Ovarianas/patologia , Adulto , Idoso , Técnicas Citológicas/métodos , Feminino , Mutação em Linhagem Germinativa , Humanos , Pessoa de Meia-Idade , Neoplasias Ovarianas/diagnóstico , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/metabolismo
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