RESUMO
Dyskeratosis congenita (DC) is a rare inherited bone marrow failure syndrome, caused by genetic mutations that principally affect telomere biology. Approximately 35% of cases remain uncharacterised at the genetic level. To explore the genetic landscape, we conducted genetic studies on a large collection of clinically diagnosed cases of DC as well as cases exhibiting features resembling DC, referred to as 'DC-like' (DCL). This led us to identify several novel pathogenic variants within known genetic loci and in the novel X-linked gene, POLA1. In addition, we have also identified several novel variants in POT1 and ZCCHC8 in multiple cases from different families expanding the allelic series of DC and DCL phenotypes. Functional characterisation of novel POLA1 and POT1 variants, revealed pathogenic effects on protein-protein interactions with primase, CTC1-STN1-TEN1 (CST) and shelterin subunit complexes, that are critical for telomere maintenance. ZCCHC8 variants demonstrated ZCCHC8 deficiency and signs of pervasive transcription, triggering inflammation in patients' blood. In conclusion, our studies expand the current genetic architecture and broaden our understanding of disease mechanisms underlying DC and DCL disorders.
Assuntos
Contusões , Púrpura , Contusões/diagnóstico , Humanos , Lactente , Masculino , Púrpura/diagnóstico , Púrpura/etiologiaRESUMO
The inclusion of familial myeloid malignancies as a separate disease entity in the revised WHO classification has renewed efforts to improve the recognition and management of this group of at risk individuals. Here we report a cohort of 86 acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) families with 49 harboring germline variants in 16 previously defined loci (57%). Whole exome sequencing in a further 37 uncharacterized families (43%) allowed us to rationalize 65 new candidate loci, including genes mutated in rare hematological syndromes (ADA, GP6, IL17RA, PRF1 and SEC23B), reported in prior MDS/AML or inherited bone marrow failure series (DNAH9, NAPRT1 and SH2B3) or variants at novel loci (DHX34) that appear specific to inherited forms of myeloid malignancies. Altogether, our series of MDS/AML families offer novel insights into the etiology of myeloid malignancies and provide a framework to prioritize variants for inclusion into routine diagnostics and patient management.
Assuntos
Mutação em Linhagem Germinativa , Leucemia Mieloide Aguda/genética , Síndromes Mielodisplásicas/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Adenosina Desaminase/genética , Dineínas do Axonema/genética , Estudos de Coortes , Humanos , Degradação do RNAm Mediada por Códon sem Sentido , Linhagem , Perforina/genética , Glicoproteínas da Membrana de Plaquetas/genética , RNA Helicases/genética , Receptores de Interleucina-17/genética , Proteínas de Transporte Vesicular/genética , Sequenciamento do ExomaRESUMO
An asymptomatic 36-year-old woman presented with significantly elevated white blood cells (165.9 × 109/L) at antenatal booking, in her first pregnancy. Further investigations revealed the diagnosis of chronic lymphocytic leukaemia with monoallelic deletion of chromosome 13q14. She was supported and monitored through out pregnancy, without treatment, and delivered a healthy baby boy at term with no complications and is currently being followed up by the haem-oncology team.
RESUMO
Exome sequencing was performed in three index cases with bone marrow failure and neurological dysfunction and whose parents are first-degree cousins. Homozygous truncating mutations were identified in ERCC6L2 in two of the individuals. Both of these mutations affect the subcellular localization and stability of ERCC6L2. We show here that knockdown of ERCC6L2 in human A549 cells significantly reduced their viability upon exposure to the DNA-damaging agents mitomycin C and Irofulven, but not etoposide and camptothecin, suggesting a role in nucleotide excision repair. ERCC6L2-knockdown cells also displayed H2AX phosphorylation, which significantly increased upon genotoxic stress, suggesting an early DNA-damage response. Intriguingly, ERCC6L2 was seen to translocate to the mitochondria and the nucleus in response to DNA damage, and ERCC6L2 knockdown induced intracellular reactive oxygen species (ROS). Treatment with the ROS scavenger N-acetyl cysteine attenuated the Irofulven-induced cytotoxicity in ERCC6L2-knockdown cells and abolished ERCCGL2 traffic to the mitochondria and nucleus in response to this DNA-damaging agent. Collectively, these observations identify a distinct bone-marrow-failure syndrome due to mutations in ERCC6L2, a gene implicated in DNA repair and mitochondrial function.
Assuntos
DNA Helicases/genética , Reparo do DNA/genética , Hemoglobinúria Paroxística/genética , Mitocôndrias/genética , Acetilcisteína/metabolismo , Anemia Aplástica , Doenças da Medula Óssea , Transtornos da Insuficiência da Medula Óssea , Linhagem Celular Tumoral , Núcleo Celular/genética , Núcleo Celular/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Dano ao DNA/efeitos dos fármacos , Feminino , Técnicas de Silenciamento de Genes , Células HEK293 , Células HeLa , Histonas/genética , Histonas/metabolismo , Humanos , Masculino , Mitocôndrias/efeitos dos fármacos , Mitomicina/toxicidade , Mutação , Linhagem , Fosforilação , Espécies Reativas de Oxigênio , Sesquiterpenos/toxicidadeRESUMO
Aplastic anemia (AA) and myelodysplasia (MDS) are forms of bone marrow failure that are often part of the same progressive underlying disorder. While most cases are simplex and idiopathic, some show a clear pattern of inheritance; therefore, elucidating the underlying genetic cause could lead to a greater understanding of this spectrum of disorders. We used a combination of exome sequencing and SNP haplotype analysis to identify causative mutations in a family with a history of autosomal-dominant AA/MDS. We identified a heterozygous mutation in SRP72, a component of the signal recognition particle (SRP) that is responsible for the translocation of nascent membrane-bound and excreted proteins to the endoplasmic reticulum. A subsequent screen revealed another autosomal-dominant family with an inherited heterozygous SRP72 mutation. Transfection of these sequences into mammalian cells suggested that these proteins localize incorrectly within the cell. Furthermore, coimmunoprecipitation of epitope-tagged SRP72 indicated that the essential RNA component of the SRP did not fully associate with one of the SRP72 variants. These results suggest that inherited mutations in a component of the SRP have a role in the pathophysiology of AA/MDS, identifying a third pathway for developing these disorders alongside transcription factor and telomerase mutations.
Assuntos
Anemia Aplástica/genética , Exoma , Mutação , Síndromes Mielodisplásicas/genética , Partícula de Reconhecimento de Sinal/genética , Anemia Aplástica/fisiopatologia , Análise Mutacional de DNA , Bases de Dados Genéticas , Feminino , Biblioteca Gênica , Heterozigoto , Humanos , Masculino , Síndromes Mielodisplásicas/fisiopatologia , Linhagem , RNA/genética , Partícula de Reconhecimento de Sinal/metabolismo , Telomerase/genética , Telomerase/metabolismo , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
The myelodysplastic syndromes (MDS) are heterogeneous and can evolve into acute myeloid leukaemia (AML). Rare familial cases are reported in which five disease genes have been identified to date (RUNX1, CEBPA, TERC, TERT and GATA2). Here we report the genetic categorization of 27 families with familial MDS/AML. All of these families were screened for RUNX1, CEBPA, TERC, TERT and GATA2 as well as TET2 and NPM1. Five of the 27 families had telomerase mutations; one had a RUNX1 mutation, while none were found to have TET2, CEBPA or NPM1 mutations. We identified four families with heterozygous GATA2 mutations, each associated with a different phenotype. While one of these mutations is novel, three have been previously reported: one has been described in dendritic cell, monocyte, B and NK lymphoid (DCML) deficiency and one is in a family that has been reported in a series with primary lymphoedema with a predisposition to AML (Emberger syndrome). In summary, genetic characterization was shown in 10 (four GATA2, three TERT, two TERC, one RUNX1) of these families; however 17 remain uncharacterized, highlighting marked genetic heterogeneity in familial MDS/AML and the scope for further functional pathways that could give rise to this group of disorders.
Assuntos
Leucemia Mieloide Aguda/genética , Síndromes Mielodisplásicas/genética , Adolescente , Adulto , Criança , Pré-Escolar , Cromatografia Líquida de Alta Pressão/métodos , DNA de Neoplasias/genética , Fator de Transcrição GATA2/genética , Genes Neoplásicos , Predisposição Genética para Doença , Humanos , Pessoa de Meia-Idade , Mutação , Nucleofosmina , Linhagem , Fenótipo , Adulto JovemRESUMO
The bone marrow failure syndrome dyskeratosis congenita (DC) has been considered to be a disorder of telomere maintenance in which disease features arise due to accelerated shortening of telomeres. By screening core components of the telomerase and shelterin complexes in patients with DC and related bone marrow failure syndromes we have identified 24 novel mutations: 11 in the RNA component of telomerase (TERC), 8 in the reverse transcriptase component (TERT), 4 in dyskerin (DKC1) and 1 in TRF1-interacting nuclear factor 2 (TINF2). This has prompted us to review these genetic subtypes in terms of telomere length, telomerase activity and clinical presentation among 194 genetically characterised index cases recruited onto the registry in London. While those with DKC1 and TINF2 mutations present at a younger age and have more disease features than those with TERC or TERT mutations, there is no difference in telomere length between these groups. There is no difference in the age of onset and numbers of disease features seen in those with TERC and TERT mutations despite the fact that the latter show higher levels of telomerase activity in vitro. The incidence of aplastic anaemia is greater in patients with TERC or TINF2 mutations compared to patients with DKC1 mutations, and cancer incidence is highest in patients with TERC mutations. These data are the first to provide robust comparisons between different genetic subtypes of telomerase and shelterin mutations (the "telomereopathies") and clearly demonstrate that disease severity is not explained by telomere length alone.
Assuntos
Mutação/genética , Índice de Gravidade de Doença , Telomerase/genética , Homeostase do Telômero , Proteínas de Ligação a Telômeros/genética , Adolescente , Adulto , Criança , Pré-Escolar , Humanos , Lactente , Pessoa de Meia-Idade , Complexo Shelterina , Adulto JovemRESUMO
Dyskeratosis congenita (DC) is a heterogeneous bone marrow failure disorder with known mutations in components of telomerase and telomere shelterin. Recent work in a mouse model with a dyskerin mutation has implicated an increased DNA damage response as part of the cellular pathology, while mouse models with Terc and Tert mutations displayed a normal response. To clarify how these contradictory results might apply to DC pathology in humans, we studied the cellular phenotype in primary cells from DC patients of several genetic subtypes, focussing on T lymphocytes to remain close to the haematopoietic system. We observed novel cell cycle abnormalities in conjunction with impaired growth and an increase in apoptosis. Using flow cytometry and confocal microscopy we examined induction of the DNA damage proteins γ-H2AX and 53BP1 and the cell cycle protein TP53 (p53). We found an increase in damage foci at telomeres in lymphocytes and an increase in the basal level of DNA damage in fibroblasts, but crucially no increased response to DNA damaging agents in either cell type. As the response to induced DNA damage was normal and levels of global DNA damage were inconsistent between cell types, DNA damage may contribute differently to the pathology in different tissues.
Assuntos
Dano ao DNA , Disceratose Congênita/genética , Adolescente , Adulto , Apoptose/genética , Ciclo Celular/genética , Pré-Escolar , Disceratose Congênita/imunologia , Disceratose Congênita/metabolismo , Feminino , Fibroblastos/metabolismo , Histonas/metabolismo , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Masculino , Microscopia Confocal , Linfócitos T/metabolismo , Linfócitos T/patologia , Telômero/genética , Técnicas de Cultura de Tecidos , Proteína Supressora de Tumor p53/metabolismo , Proteína 1 de Ligação à Proteína Supressora de Tumor p53 , Regulação para Cima/genética , Adulto JovemRESUMO
To address the association between travel and deep vein thrombosis (DVT) we examined the risk factors for DVT in 568 consecutive patients with suspected DVT attending King's College Hospital in London. No significant link between DVT and long-haul travel was demonstrable in this cohort, with an odds ratio of 1.3 (CI 0.6-2.8). Risk of DVT was only increased in long-haul travellers if one or more additional risk factors were present, with an odds ratio of 3.0 (CI 1.1-8.2). Such individuals may benefit from prophylactic measures to minimize risk.