RESUMO
Fanconi anemia (FA) is an autosomal recessive disease characterized by pancitopenia, congenital malformations, predisposition to cancers and chromosomal instability. We report the clinical and molecular features of a patient initially identified as a potential FA case only because of chemotherapy toxicity during the treatment of a T-lineage acute lymphoblastic leukemia (ALL). Cells from this patient showed a moderate chromosomal instability, increasing sensitivity to DNA crosslinking agents but normal response to ionizing radiation. The analysis of FA proteins demonstrated a marked reduction of FANCD2 (>95%), but normal levels of FANCA or FANCG. Interestingly, this defect was associated with a homozygous missense mutation of FANCD2, resulting in a novel amino-acid substitution (Leu153Ser) at residue Leu153, which is highly conserved through evolution. The FANCD2(L153S) protein, whose reduced expression was not due to impaired transcription, was detected also in its monoubiquitinated form in the nucleus, suggesting that the mutation does not affect post-translation modifications or subcellular localization but rather the stability of FANCD2. Therefore, the hypomorphic Leu153Ser mutation represents the first example of a FANCD2 defect that might promote clonal progression of tumors, such as T-ALL, and severe chemotherapy toxicity in patients without any clinical manifestations typical of FA.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/efeitos adversos , Proteína do Grupo de Complementação D2 da Anemia de Fanconi/genética , Leucemia-Linfoma de Células T do Adulto/tratamento farmacológico , Leucemia-Linfoma de Células T do Adulto/genética , Mutação , Substituição de Aminoácidos , Antígenos CD , Antígenos de Diferenciação Mielomonocítica , Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Antígenos CD13 , Criança , Instabilidade Cromossômica , Progressão da Doença , Anemia de Fanconi/genética , Humanos , Infecções/etiologia , Infecções/genética , Leucemia-Linfoma de Células T do Adulto/fisiopatologia , Masculino , Pancitopenia/induzido quimicamente , Pancitopenia/genética , Indução de Remissão , Lectina 3 Semelhante a Ig de Ligação ao Ácido SiálicoRESUMO
Fanconi anemia (FA) is a clinically and genetically heterogeneous disorder. Clinical care is complicated by variable age at onset and severity of hematologic symptoms. Recent advances in the molecular biology of FA have allowed us to investigate the relationship between FA genotype and the nature and severity of the clinical phenotype. Two hundred forty-five patients from all 7 known complementation groups (FA-A to FA-G) were studied. Mutations were detected in one of the cloned FANC genes in 169 patients; in the remainder the complementation group was assigned by cell fusion or Western blotting. A range of qualitative and quantitative clinical parameters was compared for each complementation group and for different classes of mutation. Significant phenotypic differences were found. FA-G patients had more severe cytopenia and a higher incidence of leukemia. Somatic abnormalities were less prevalent in FA-C, but more common in the rare groups FA-D, FA-E, and FA-F. In FA-A, patients homozygous for null mutations had an earlier onset of anemia and a higher incidence of leukemia than those with mutations producing an altered protein. In FA-C, there was a later age of onset of aplastic anemia and fewer somatic abnormalities in patients with the 322delG mutation, but there were more somatic abnormalities in patients with IVS4 + 4A --> T. This study indicates that FA patients with mutations in the FANCG gene and patients homozygous for null mutations in FANCA are high-risk groups with a poor hematologic outcome and should be considered as candidates both for frequent monitoring and early therapeutic intervention. (Blood. 2000;96:4064-4070)
Assuntos
Proteínas de Ciclo Celular , Proteínas de Ligação a DNA/genética , Anemia de Fanconi/genética , Heterogeneidade Genética , Proteínas Nucleares/genética , Proteínas/genética , Proteínas de Ligação a RNA/genética , Anormalidades Múltiplas/epidemiologia , Anormalidades Múltiplas/genética , Doença Aguda , Adolescente , Adulto , Idade de Início , Substituição de Aminoácidos , Anemia Aplástica/genética , Anemia Aplástica/mortalidade , Criança , Pré-Escolar , Análise Mutacional de DNA , Anemia de Fanconi/classificação , Anemia de Fanconi/mortalidade , Proteína do Grupo de Complementação A da Anemia de Fanconi , Proteína do Grupo de Complementação E da Anemia de Fanconi , Proteína do Grupo de Complementação F da Anemia de Fanconi , Proteína do Grupo de Complementação G da Anemia de Fanconi , Proteínas de Grupos de Complementação da Anemia de Fanconi , Deleção de Genes , Frequência do Gene , Teste de Complementação Genética , Genótipo , Humanos , Lactente , Leucemia Mieloide/epidemiologia , Leucemia Mieloide/genética , Pessoa de Meia-Idade , Síndromes Mielodisplásicas/epidemiologia , Síndromes Mielodisplásicas/genética , Fenótipo , Mutação Puntual , Risco , Deleção de Sequência , Análise de SobrevidaRESUMO
Fanconi's anemia (FA) is a very rare genetically heterogeneous disease which has been hypothesized to be defective in the detoxification of reactive oxygen species. In this work we report the results obtained by morphometric and biochemical analyses on the red blood cells (RBCs) from FA patients. With respect to RBCs from healthy donors the following changes have been detected: (i) a variety of ultrastructural alterations, mainly surface blebbing typical of acanthocytes and stomatocytes; (ii) a significant quantitative increase of these altered forms; (iii) modifications of spectrin cytoskeleton network; (iv) an altered redox balance, e.g. a decreased catalase activity and significant variations in the GSSG/GSH ratio. We hypothesize that remodeling of the redox state occurring in FA patients results in cytoskeleton-associated alterations of red blood cell integrity and function.
Assuntos
Citoesqueleto/ultraestrutura , Membrana Eritrocítica/ultraestrutura , Eritrócitos/metabolismo , Eritrócitos/ultraestrutura , Anemia de Fanconi/sangue , Adolescente , Adulto , Catalase/sangue , Criança , Pré-Escolar , Glutationa/sangue , Dissulfeto de Glutationa/sangue , Humanos , Microscopia Eletrônica de Varredura , Valores de Referência , Espectrina/ultraestrutura , Superóxido Dismutase/sangue , Superóxidos/sangue , Zinco/sangueRESUMO
A number of congenital conditions include oxidative stress as a phenotypic hallmark, in spite of very distinct genotypic assets. These disorders include some rare chromosomal instability syndromes and two frequent congenital conditions, Down's syndrome and cystic fibrosis. All of these disorders generate severe biomedical and social handicaps in their progression, resulting in neurological symptoms or malformations, early ageing and cancer proneness. The onset of oxidative stress has been related to excess formation or defective detoxification of reactive oxygen species. Investigations conducted so far have focussed on individual disorders and have utilised a range of different methodologies, so leading to a number of unanswered questions regarding the role(s) for oxidative stress in these disorders. Comparative studies are required, based on homogeneous methods, to provide insights into the pathogenesis of each individual condition, which may open up new avenues for studying ageing and cancer.
Assuntos
Aberrações Cromossômicas/genética , Fibrose Cística/genética , Síndrome de Down/genética , Estresse Oxidativo/fisiologia , Aberrações Cromossômicas/classificação , Transtornos Cromossômicos , HumanosRESUMO
In spite of very distinct genotypic assets, a number of congenital conditions include oxidative stress as a phenotypic hallmark. These disorders include Fanconi's anaemia, ataxia telangiectasia, xeroderma pigmentosum and Bloom's syndrome, as well as two frequent congenital conditions: Down's syndrome and cystic fibrosis. Cancer proneness is a clinical feature shared by these disorders, while other manifestations include early ageing, neurological symptoms or congenital malformations. The onset of oxidative stress has been related to excess formation, or defective detoxification, of reactive oxygen species (ROS). This can arise from either the abnormal expression or inducibility of ROS-detoxifying enzymes, or by defective absorption of nutrient antioxidants. Resulting oxidative injury has been characterized through: (i) DNA, protein or lipid oxidative damage; (ii) excess ROS formation (in vitro and ex vivo); (iii) sensitivity to oxygen-related toxicity; (iv) improvement of cellular defects by either hypoxia or antioxidants; and (v) circumstantial evidence for in vivo oxidative stress (as e.g. clastogenic factors). Investigations conducted so far have been confined to individual disorders. Comparative studies of selected indicators for oxidative stress could provide further insights into the pathogenesis of each individual condition. Such a unified approach may have wide-ranging consequences for studies of ageing and cancer.
Assuntos
Suscetibilidade a Doenças , Doenças Genéticas Inatas , Neoplasias/etiologia , Estresse Oxidativo , Envelhecimento , Animais , Apoptose , Doenças Genéticas Inatas/genética , Humanos , Neoplasias/genética , Estresse Oxidativo/genética , FenótipoRESUMO
Fanconi's anaemia (FA) is a rare autosomal recessive disease characterised by progressive pancytopoenia, a diverse assortment of congenital malformations, an increased sensitivity to reactive oxygen species and a predisposition to the development of malignancies. In the present study, we assessed the propensity to undergo apoptosis of peripheral blood mononuclear cells (PBMC) from Italian FA patients. Cells were challenged by 2-deoxy-D-ribose (dRib) or TNF-alpha plus cycloheximide as agents that induce apoptosis by interfering with cell redox status and mitochondrial membrane potential (MMP), and PBMC from FA patients resulted to be less prone to die than those from healthy subjects. The decreased susceptibility of FA cells to undergo apoptosis was also evident when another parameter highly correlated with the apoptotic process, i.e. MMP, was measured. Moreover, when N-acetylcysteine was added to dRib-treated PBMC, a strong protection was evident either in PBMC from control subjects or from FA patients. These data indicate that an alteration of unknown nature of the mechanisms favouring apoptosis is present in freshly collected cells from FA patients, and that such alteration could contribute to the pathogenesis of the disease, and particularly to the increased susceptibility to cancer.
Assuntos
Apoptose , Anemia de Fanconi/patologia , Leucócitos Mononucleares/patologia , Adolescente , Apoptose/efeitos dos fármacos , Células Cultivadas , Criança , Pré-Escolar , Cicloeximida/farmacologia , Desoxirribose/farmacologia , Feminino , Humanos , Hibridização In Situ , Leucócitos Mononucleares/efeitos dos fármacos , Masculino , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
The present study was conducted on 13 patients with Fanconi anemia. 25 parents and 12 siblings. The chromosomal instability characteristic of this congenital breakage syndrome was associated with the presence of transferable clastogenic material in the plasma, as also reported previously for ataxia telangiectasia and Bloom's syndrome. While all plasma ultrafiltrates from homozygotes had chromosome damaging properties, the clastogenic material had to be concentrated in most heterozygotes to reach detectable levels. The clastogenic effect was exerted via the intermediacy of superoxide radicals, since it was regularly inhibited by superoxide dismutase (SOD). This adds further evidence for a prooxidant state in this hereditary disease. The autosustained clastogenic activity possibly plays a role in the progressive impairment of blood cell-producing bone marrow and may predispose patients to develop cancer and leukemia. Prophylactic use of antioxidants may be recommended, using clastogenic plasma activity as a guide.
Assuntos
Aberrações Cromossômicas/genética , Anemia de Fanconi/sangue , Anemia de Fanconi/genética , Mutagênicos/metabolismo , Adolescente , Adulto , Criança , Pré-Escolar , Feminino , Heterozigoto , Homozigoto , Humanos , MasculinoRESUMO
The present study was aimed at verifying the occurrence, if any, of in vivo oxidative DNA damage in FA homozygotes, their parents and siblings. 8-Hydroxy-2'-deoxyguanosine (8-OHdG) was measured, by HPLC/EC, in DNA from circulating blood leucocytes from FA homozygotes and their relatives and compared with a group of paediatric and adult healthy subjects. The population studied consisted of: (i) 15 FA homozygotes; (ii) 24 FA heterozygotes; (iii) 11 siblings. The 8-OHdG level in FA homozygotes was significantly higher with respect to age-matched controls, with a mean level of 33.3 +/- 6.8 (mean +/- SE) and 3.9 +/- 0.26 8-OHdG/10(5) dG respectively. The FA parents (heterozygotes) also displayed higher 8-OHdG levels relative to controls. The release of hydroxyl (.OH) and .OH-like radicals from leucocytes was determined by luminol-dependent chemiluminescence (LDCL) in a subgroup of FA homo- and heterozygotes, showing a very large in vivo formation of non-superoxide radicals. Chromosomal instability was also measured in the FA population. When relating either 8-OHdG or LDCL levels to spontaneous or diepoxybutane-induced chromosomal instability (S-CI and DEB-CI respectively), a significant correlation was observed between the 8-OHdG, LDCL and S-CI data. Within families a positive association was found between 8-OHdG levels in homozygotes and their related heterozygotes, suggesting segregation of the genetic defect(s) underlying the abnormal oxidative metabolism. The present study provides evidence for an in vivo pro-oxidant state in FA, in terms of excess formation of .OH and .OH-like radicals, and of DNA hydroxyl adducts. This finding appears to be shared by homozygotes and, to a lesser extent, by heterozygotes.
Assuntos
Dano ao DNA , DNA/sangue , Desoxiguanosina/análogos & derivados , Anemia de Fanconi/sangue , Anemia de Fanconi/genética , Espécies Reativas de Oxigênio/metabolismo , 8-Hidroxi-2'-Desoxiguanosina , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , Pré-Escolar , Cromatografia Líquida de Alta Pressão , Cromossomos Humanos , Desoxiguanosina/análise , Desoxiguanosina/sangue , Compostos de Epóxi , Feminino , Heterozigoto , Homozigoto , Humanos , Leucócitos/metabolismo , Medições Luminescentes , Luminol , Masculino , Pessoa de Meia-Idade , Oxirredução , Explosão Respiratória/fisiologiaRESUMO
This report describes the clinical and cytogenetic analysis of a kindred with multiple endocrine neoplasia type 2 (MEN-2 or Sipple's syndrome) in two generations. Medullary thyroid carcinoma was present in five members either as a large or as an occult tumour. Phaeochromocytoma was demonstrated in one severely hypertensive relative and urine vanillylmandelic acid (VMA) was increased in one normotensive member. Serum parathormone (PTH) was normal in all but one normocalcaemic patient of this family who did not have a history of nephrolithiasis. Prometaphase banding failed to detect a 20p12.2 deletion or chromosome instability as observed in some MEN-2 families.