Ce travail de recherche vise à évaluer les enjeux éthiques posés par le recours au diagnostic anténatal dans le cadre des maladies génétiques à révélation tardive.Notre première étude a été d’analyser les décisions prises en réunions de Centres Pluridisciplinaires de Diagnostic Prénatal (CPDPN) et, à travers des situations réelles et singulières, relever les éléments de discussion et plus particulièrement ceux pouvant influencer la décision. Nous avons, ensuite réalisé deux enquêtes par questionnaires qui ont permis: 1) D’explorer le point de vue des personnes directement concernées par une telle pathologie (porteurs du gène responsable - malades ou asymptomatiques -, conjoints et/ou parents d’une personne porteuse du gène) ; 2) D’étudier la position des professionnels de la parentalité, travaillant en lien avec un CPDPN, et qui sont décideurs de la recevabilité ou non d’une demande de diagnostic anténatal dans ce contexte.Ce travail a ainsi contribué à faire émerger des questionnements pertinents sur le plan éthique et une réflexion sur de possibles évolutions législatives et sociétales dans ce domaine. / This research carries out with the aim of evaluating the ethical challenges faced by the use of antenatal diagnosis in late-onset genetic diseases.In a first study, we analysed the decisions of Multidisciplinary Centres for Prenatal Diagnosis (MCPD) and, through real and specific situations, we identified the elements for discussion and more particularly the ones that could influence the decision-making process. Then, we conducted two questionnaire surveys that allowed to: 1) Explore the viewpoints of people directly affected by this type of pathology (responsible gene carriers - ill or asymptomatic individuals -, partners and/or parents of gene carriers); 2) Examinate the opinions of professionals, working in association with a CPDPN and who are decision-makers of the acceptability or not for an antenatal diagnosis request in this context.This work has therefore brought out questions on ethics, and views on the potential legal and social developments in this area.

No description available.

La neuropathie sensitive et motrice héréditaire avec agénésie du corps calleux (NSMH/ACC) se traduit par une atteinte neurodégénérative sévère associée à des anomalies développementales dans le système nerveux central et du retard mental. Bien que rare dans le monde, ce désordre autosomique récessif est particulièrement fréquent dans la population Québécoise du Canada Français du fait d’un effet fondateur. L’unique étude réalisée sur la mutation québécoise du gène qui code pour le co-transporteur de potassiumchlore 3 (KCC3) a montré qu’il y a une perte de fonction de la protéine. Cependant, la maladie est également retrouvée hors du Québec et il reste encore à élucider les pathomécanismes mis en jeu. Nous avons donc séquencé les 26 exons du gène KCC3 chez des individus recrutés dans le monde entier et suspectés d’être atteints de la maladie. Nous avons ainsi identifié trois nouvelles mutations. L’étude fonctionnelle de ces mutations nous a confirmé la perte de fonction systématique des co-transporteurs mutés. Puisque l’inactivation de KCC3 se produit majoritairement via l’élimination de segments peptidiques en C-terminus, nous avons concentré notre attention sur l’identification des interactions qui s’y produisent. À l’aide d’approches double hybride, pull-down et immunomarquage, nous avons déterminé que KCC3 interagit avec la créatine kinase CK-B et que cette interaction est perturbée par les mutations tronquantes. De plus, l’utilisation d’un inhibiteur de créatine kinase inactive KCC3, ce qui démontre qu’il existe bien un lien fonctionnel et pathologique entre KCC3 et ses partenaires C-terminaux. Nous avons aussi identifié des anomalies majeures de localisation membranaire des KCC3 mutés. Que KCC3 soit tronqué ou pleine longueur, sa distribution subcellulaire est affectée dans des cellules en culture, dans les ovocytes de Xenopes et dans des échantillons de cerveau de patients. La perte d’interaction entre KCC3 et CK-B et/ou les défauts de transit intracellulaire de KCC3 sont donc les mécanismes pathologiques majeurs de la NSMH/ACC. / Heredirary motor and sensory neuropathy with agenesis of the corpus callosum (HMSN/ACC) is a severe neurodegenerative disease associated with developmental anomalies in the central nervous system and mental retardation. Although rare worldwide, this autosomal-recessive disorder is frequent in the French-Canadian population of Quebec because of a founder effect. Different mutations in the gene coding for the potassiumchloride co-transporter 3 (KCC3) have been associated with the disease; however, little is known about the mechanisms leading to the inactivation of the co-transporter. We sequenced 26 exons of the KCC3 gene in individuals recruited worldwide and suspected to be affected by the disease. We identified three new mutations. The functional study of these mutations gave confirmation of a systematic loss-of-function of the mutant co-transporters. As the loss of function occurs mainly via the elimination of C-terminal peptide fragments, we focused on the identification of C-terminal interacting partners. Using different biochemical approaches, such as yeast two-hydbrid, pull-down, and immunostaining, we established that KCC3 interacts with the brain-type creatine kinase CK-B and that this interaction is disrupted by the HMSN/ACC truncation mutations. In addition, a specific creatine kinase inhibitor inactivates KCC3 and shows for the first time the functional link between KCC3 and its C-terminal partners. In addition, we found that anomalies in KCC3 transit—as seen in cultured cells, in Xenopus oocytes, and in human brain samples—is a major pathogenic mechanism that also leads to the disease manifestations.

Introdução: A Hipercolesterolemia Familiar (HF) é uma doença genética caracterizada clinicamente por elevados níveis de lipoproteína de baixa densidade (LDL-C) na corrente sanguínea desde a infância. Indivíduos que apresentam HF podem desenvolver doença aterosclerótica ainda em idade jovem. Os principais preditores de risco no desenvolvimento da doença cardiovascular (DCV) nesses indivíduos após entrarem em um programa de rastreamento genético não são conhecidos na nossa população. Além disso, a HF é subdiagnosticada e subtratada mundialmente e o rastreamento genético em cascata dos familiares tem sido mundialmente avaliado como o método diagnóstico mais custo. Contudo, a efetividade do rastreamento genético em cascata é dependente dos critérios clínicos de entrada do primeiro indivíduo da família e não há um consenso de qual critério apresenta a melhor acurácia para detecção de uma mutação. Objetivos: Identificar os fatores determinantes para ocorrência de eventos cardiovasculares (CV) em todos os indivíduos da coorte e avaliar o critério clínico para detecção de uma variante genética patogênica para HF, no primeiro indivíduo da família, após serem inseridos em um programa de rastreamento genético em cascata.Métodos: Estudo de coorte prospectiva aberta dos pacientes que foram inseridos no programa de rastreamento genético em cascata para HF. A população do estudo é definida como caso índice (CI), o primeiro da família a ser identificado clinicamente e encaminhado para o teste genético, e os familiares, que são os parentes de 1º grau do CI em que foi encontrada uma alteração genética. Todos os indivíduos são inseridos na coorte no momento em que recebem o laudo genético (tempo zero, T0). Um ano depois do T0 é realizado o primeiro contato telefônico, ou seja, primeiro ano de seguimento (T1) Resultados: No T1, o total de 818 indivíduos foi incluído, sendo verificados 47 eventos CV, sendo 14 (29,7%) fatais. Para o CI, o único fator independente associado ao aumento do risco de eventos CV no T1 foi a presença de arco corneano (OR: 9,39; IC 95%: 2,46-35,82). Para os familiares com uma mutação positiva os fatores associados ao aumento do risco de eventos CV foram diabetes mellitus (OR: 7,97; IC 95%: 2,07-30,66) e consumo de tabaco (OR: 3,70; IC 95%: 1,09-12,50). Na análise do melhor critério clínico para detecção de uma mutação patogênica no CI os valores de LDL-C >= 230 mg/dL tiveram a melhor relação entre sensibilidade e especificidade. Na análise da curva ROC o escore Dutch Lipid Clinic Network (DLCN) apresentou melhor desempenho do que o LDL-C para identificar uma mutação, a área sob a curva ROC foi 0,744 (IC 95%: 0,704-0,784) e 0,730 (IC 95%: 0,687-0,774), respectivamente, p = 0, 014. Conclusão: Em um ano de seguimento essa coorte identificou uma alta incidência de eventos CV após a entrada em um programa de rastreamento genético em cascata e os preditores dos eventos CV diferem entre CI e familiares. Esses resultados podem contribuir para o desenvolvimento de ações preventivas nesse grupo altamente susceptível de indivíduos. Além disso, devido a importância da detecção da mutação para um diagnóstico definitivo de HF e a importância da cascata ser custo efetiva o estudo identificou que o critério único do LDL-C >= 230 mg/dl é viável para indicar o CI para o teste genético / Introduction: Familial Hypercholesterolemia (FH) is a genetic disease characterized clinically by high levels of low density lipoprotein (LDL-C) in the bloodstream since childhood. Individuals with FH can develop atherosclerotic disease at a young age. The main predictors of cardiovascular disease (CVD) risk in these individuals after entering a genetic screening program are not known in our population. In addition, FH is underdiagnosed and undertreated worldwide and cascaded genetic screening of family members has been evaluated globally as the most cost effective for the diagnosis of FH. However, the effectiveness of cascading genetic screening is dependent on the clinical entry criteria of the first individual in the family and there is no consensus as to which criterion shows the best accuracy for detecting a mutation. Objectives: To identify the determinant factors for cardiovascular (CV) events in all individuals in the cohort and to evaluate the clinical criteria for detecting a genetic variant pathogenic to FH in the first individual of the family after being inserted into a genetic screening program in cascade. Methods: Open prospective cohort study of patients who were enrolled in the cascade genetic screening program for FH. The study population is defined as index case (IC), the first of the family to be clinically identified and referred to the genetic test, and relatives, who are the first-degree relatives of the IC in which a genetic alteration was found. All individuals are inserted into the cohort at the moment they receive the genetic report (time zero, T0). The first follow-up telephone contact is made one year after T0 (first year of follow-up, T1). Results: In T1, a total of 818 subjects were included, and 47 CV events were verified, of which 14 (29.7%) were fatal. For IC, the only factor independently associated with the increased risk of CV events in T1 was the presence of a corneal arch (OR: 9.39; 95% CI: 2.46-35.82). For relatives with positive mutation, factors associated with increased risk of CV events were diabetes mellitus (OR: 7.97; 95% CI: 2.07-30.66) and tobacco consumption (OR: 3.70; 95% CI: 1.09-12.50). In the analysis of the best clinical criteria for the detection of a pathogenic mutation in the IC, the LDL-C values >= 230 mg/dL had the best relationship between sensitivity and specificity. In the ROC curve analysis, the Dutch Lipid Clinic Network (DLCN) score performed better than LDL-C to identify a mutation, the area under the ROC curve was 0.744 (95% CI: 0.704-0.784) and 0.730 (CI 95 %: 0.687-0.774), respectively, p = 0.014. Conclusion: At one year follow-up this cohort identified a high incidence of CV events following entry into a cascade genetic screening program and the predictors of CV events differ between IC and family members. These results may contribute to the development of preventive actions in this group highly susceptible to individuals. In addition, because of the importance of detecting the mutation for a definitive diagnosis of HF and the importance of the cascade being cost effective, the study identified that the single LDL-C criterion >= 230 mg / dl is feasible to indicate IC for the genetic test

Made available in DSpace on 2016-01-26T12:51:35Z (GMT). No. of bitstreams: 1 helenicebianchibolini_tese.pdf: 2235516 bytes, checksum: 0e1ddb1957f1b10e0d67a793c7e4a245 (MD5) Previous issue date: 2010-10-27 / The practical problem observed referred to the clinical symptoms of NF1, involving intellectual performance and psychosocial characteristics of their carriers. This sends us to the NF1-intelligence interface. Objective: To identify and compare indices of intelligence and their frequencies in patients with NF1, attended at CEPAN. Methods and Casuistry: Medical records were used in research, semi-structured interview, the Wechsler Scales and Test Progressive Matrices Scale-General. Were applied indidually, to the 77 subjects, of which 30 patients with NF1, 17 family members, and 30 non- carriers between 2006 and 2010. The data were treated qualitative-quantitative. Results: The socioeconomic and cultural rights did not differ between subjects. Minors (<20 ) and larger (>20 ) time spent in the execution of Test Progressive Matrices Sacale-General were relatives of patients with NF1 and most were using medication. Mean correct responses were lower in patients with NF1, they had one and two symptoms that bothered when they were diagnosed, currently, the troubled portability three, four and two symptoms. Patients with NF1 had their first symptoms identified with more than five years age, they had relatives suffering from NF1 1st and/or 2nd degrees of relatedness (vertical transmission). They had mild MR and moderate, and learning disabilities. The subjects of this investigation showed TIQ; VIQ; TIQ/Gc; IOP/Gv; IVP/Gt average and below average; although the ability Reasoning/ Fluid Intelligence Gf category V. Conclusions: The subjects of this investigation have average and average lower, with limited sustainability. There are differences in intellectual performance among them: relatives of patients with NF1 are superior to the carriers and no- carriers are superior to both. There are mental retardation, learning disabilities, difficulties visual-perceptual-motor, memory impairments, and speech in written and spoken language in patients with NF1. There are no correlations between TIQ/Gc; IMO/Gsm; ICV/Gc; IOP/Gv; IVP/Gt and number of symptoms. There is no correlation between IQ, intellectual level, types, numbers, uncomfortable symptoms and age of onset of symptoms. / O problema prático observado, referiu-se aos sintomas clínicos de NF1, que envolviam o desempenho intelectual e as características psicossociais de seu portador, remetendo-nos à interface inteligência Neurofibromatose 1-NF1. Objetivo: Identificar e comparar os índices de inteligência e suas frequências em portadores de NF 1 atendidos no CEPAN. Casuística e Métodos: Utilizou-se pesquisa em prontuários; entrevista semi-estruturada; as Escalas de Wechsler e Testes de Matrizes Progressivas - Escala Geral, aplicados individualmente a 77 sujeitos, dos quais 30 portadores de NF1, 17 seus familiares e 30 não-portadores, entre 2006 e 2010. Os dados receberam tratamento quali-quantitativo. Resultados: As características socioeconômicas e culturais não diferiram entre os sujeitos Os tempos menores (<20 ) e maiores (>20 ) gastos na execução do Raven - Escala Geral foram de familiares e portadores de NF1, que mais faziam uso de medicação. As médias de acertos de portadores de NF1 foram as menores; possuíam 1 e 2 sintomas que os incomodavam quando foram diagnosticados; atualmente, era a portabilidade de 3, 4 e 2 sintomas que os incomodavam; primeiros sintomas identificados com mais de 5 anos; possuíam parentes portadores de NF1 de 1º e/ou 2º graus de parentesco (transmissão vertical); apresentaram RM leve e moderada; e distúrbio de aprendizagem (QI>70) em portadores de NF1. Os sujeitos dessa investigação apresentaram quocientes de inteligência e índices fatoriais médios e médio-inferiores; a capacidade Raciocínio/ Inteligência Fluida Gf encontrou-se comprometida-categoria V. Conclusões: Os sujeitos da investigação possuem inteligência média e média inferior, porém com dificuldade de sustentabilidade. Há diferenças de desempenho intelectual: familiares de portadores de NF1 são superiores aos portadores de NF1; e não portadores são superiores a ambos. Há retardo mental, distúrbio de aprendizagem, disfunção no desenvolvimento da linguagem dificuldades viso-motoras e perceptuais, deficiências de memória e de expressão na linguagem escrita e verbal em portadores de NF1; ausência de correlação entre QIT/Gc; IMO/Gsm; ICV/Gc; IOP/ Gv; IVP/ Gt e número de sintomas; e ausência de relação entre QI; nível intelectual; tipos; números; incômodos de sintomas e faixa etária do aparecimento dos primeiros sintomas.

Rare diseases, when considered as a whole, affect up to 7% of the population, which would represent 3.5 million individuals in the United Kingdom alone. However, while 'personalised medicine' is now yielding remarkable results using recent sequencing technologies in terms of diagnosing genetic conditions, we have made much less headway in translating this patient information into therapies and effective treatments. Even with recent calls for greater research into personalised treatments for those affected by a rare disease, progress in this area is still severely lacking, in part due to the astronomical cost and time involved in bringing treatments to the clinic. Gene correction using the recently-described genome editing technology CRISPR/Cas9, which allows precise editing of DNA, offers an exciting new avenue of treatment, if not cure, for rare diseases; up to 80% of which have a genetic component. This system allows the researcher to target any locus in the genome for cleavage with a short guide-RNA, as long as it precedes a highly ubiquitous NGG sequence motif. If a repair sequence is then also provided, such as a wild-type copy of the mutated gene, it can be incorporated by homology-directed repair (HDR), leading to gene correction. As both guide-RNA and repair template are easily generated, whilst the machinery for editing and delivery remain the same, this system could usher in the era of 'personalised medicine' and offer hope to those with rare genetic diseases. However, currently it is difficult to test the efficacy of CRISPR/Cas9 for gene correction, especially in vivo. Therefore, in my PhD I have developed a novel fluorescent reporter system which provides a rapid, visual read-out of both non-homologous end joining (NHEJ) and homology-directed repair (HDR) driven by CRISPR/Cas9. This system is built upon a cassette which is stably and heterozygously integrated into a ubiquitously expressed locus in the mouse genome. This cassette contains a strong hybrid promoter driving expression of membrane-tagged tdTomato, followed by a strong stop sequence, and then membrane-tagged EGFP. Unedited, this system drives strong expression of membrane-tdTomato in all cell types in the embryo and adult mouse. However, following the addition of CRISPR/Cas9 components, and upon cleavage, the tdTomato is rapidly excised, resulting via NHEJ either in cells without fluorescence (due to imperfect deletions) or with membrane-EGFP. If a repair template containing nuclear tagged-EGFP is also supplied, the editing machinery may then use the precise HDR pathway, which results in a rapid transition from membrane-tdTomato to nuclear- EGFP. Thereby this system allows the kinetics of editing to be visualised in real time and allows simple scoring of the proportion of cells which have been edited by NHEJ or corrected by HDR. It therefore provides a simple, fast and scalable manner to optimise reagents and protocols for gene correction by CRISPR/Cas9, especially compared to sequencing approaches, and will prove broadly useful to many researchers in the field. Further to this, I have shown that methods which lead to gene correction in our reporter system are also able to partially repair mutations found in the disease-causing gene, Zmynd10; which is implicated in the respiratory disorder primary ciliary dyskinesia (PCD), for which there is no effective treatment. PCD is an autosomal-recessive rare disorder affecting motile cilia (MIM:244400), which results in impaired mucociliary clearance leading to neonatal respiratory distress and recurrent airway infections, often progressing to lung failure. Clinically, PCD is a chronic airway disease, similar to CF, with progressive deterioration of lung function and lower airway bacterial colonization. However, unlike CF which is monogenic, over 40 genes are known to cause PCD. The high genetic heterogeneity of this rare disease makes it well suited to such a genome editing strategy, which can be tailored for the correction of any mutated locus.

La neuropathie sensitive et motrice héréditaire avec agénésie du corps calleux (NSMH/ACC) se traduit par une atteinte neurodégénérative sévère associée à des anomalies développementales dans le système nerveux central et du retard mental. Bien que rare dans le monde, ce désordre autosomique récessif est particulièrement fréquent dans la population Québécoise du Canada Français du fait d’un effet fondateur. L’unique étude réalisée sur la mutation québécoise du gène qui code pour le co-transporteur de potassiumchlore 3 (KCC3) a montré qu’il y a une perte de fonction de la protéine. Cependant, la maladie est également retrouvée hors du Québec et il reste encore à élucider les pathomécanismes mis en jeu. Nous avons donc séquencé les 26 exons du gène KCC3 chez des individus recrutés dans le monde entier et suspectés d’être atteints de la maladie. Nous avons ainsi identifié trois nouvelles mutations. L’étude fonctionnelle de ces mutations nous a confirmé la perte de fonction systématique des co-transporteurs mutés. Puisque l’inactivation de KCC3 se produit majoritairement via l’élimination de segments peptidiques en C-terminus, nous avons concentré notre attention sur l’identification des interactions qui s’y produisent. À l’aide d’approches double hybride, pull-down et immunomarquage, nous avons déterminé que KCC3 interagit avec la créatine kinase CK-B et que cette interaction est perturbée par les mutations tronquantes. De plus, l’utilisation d’un inhibiteur de créatine kinase inactive KCC3, ce qui démontre qu’il existe bien un lien fonctionnel et pathologique entre KCC3 et ses partenaires C-terminaux. Nous avons aussi identifié des anomalies majeures de localisation membranaire des KCC3 mutés. Que KCC3 soit tronqué ou pleine longueur, sa distribution subcellulaire est affectée dans des cellules en culture, dans les ovocytes de Xenopes et dans des échantillons de cerveau de patients. La perte d’interaction entre KCC3 et CK-B et/ou les défauts de transit intracellulaire de KCC3 sont donc les mécanismes pathologiques majeurs de la NSMH/ACC. / Heredirary motor and sensory neuropathy with agenesis of the corpus callosum (HMSN/ACC) is a severe neurodegenerative disease associated with developmental anomalies in the central nervous system and mental retardation. Although rare worldwide, this autosomal-recessive disorder is frequent in the French-Canadian population of Quebec because of a founder effect. Different mutations in the gene coding for the potassiumchloride co-transporter 3 (KCC3) have been associated with the disease; however, little is known about the mechanisms leading to the inactivation of the co-transporter. We sequenced 26 exons of the KCC3 gene in individuals recruited worldwide and suspected to be affected by the disease. We identified three new mutations. The functional study of these mutations gave confirmation of a systematic loss-of-function of the mutant co-transporters. As the loss of function occurs mainly via the elimination of C-terminal peptide fragments, we focused on the identification of C-terminal interacting partners. Using different biochemical approaches, such as yeast two-hydbrid, pull-down, and immunostaining, we established that KCC3 interacts with the brain-type creatine kinase CK-B and that this interaction is disrupted by the HMSN/ACC truncation mutations. In addition, a specific creatine kinase inhibitor inactivates KCC3 and shows for the first time the functional link between KCC3 and its C-terminal partners. In addition, we found that anomalies in KCC3 transit—as seen in cultured cells, in Xenopus oocytes, and in human brain samples—is a major pathogenic mechanism that also leads to the disease manifestations.

Submitted by Ana Maria Fiscina Sampaio (fiscina@bahia.fiocruz.br) on 2012-08-29T21:44:32Z No. of bitstreams: 1 Taisa Manuela Bonfim Machado. Migração estrutura populacional Tese 2012.pdf: 1095441 bytes, checksum: 16e3cea3a6b286c226470a459e5720fb (MD5) / Made available in DSpace on 2012-08-29T21:44:32Z (GMT). No. of bitstreams: 1 Taisa Manuela Bonfim Machado. Migração estrutura populacional Tese 2012.pdf: 1095441 bytes, checksum: 16e3cea3a6b286c226470a459e5720fb (MD5) Previous issue date: 2012 / Fundação Oswaldo Cruz. Centro de Pesquisas Gonçalo Moniz. Salvador, Bahia, Brasil / A migração é o fator evolutivo capaz de dispersar a diversidade genética entre populações, inserindo novas características fenotípicas e genotípicas. A dinâmica matrimonial, juntamente como a estrutura da população são fatores que podem alterar a frequência destas características. Exemplo dessas características são as doenças genéticas, onde a frequência e distribuição destas auxilia na compreensão da influência de fatores evolutivos em uma população. No município de Monte Santo, localizado no interior da Bahia, foram encontradas doenças genéticas com elevada frequência, como mucopolissacaridose do tipo VI e fenilcetonúria. Existem evidências que algumas doenças mostram associação entre a raça e o risco de sua ocorrência. Dados moleculares mostraram que na Bahia a contribuição africana é de 47,2%, entretanto, dados baseados em classificação fenotípica apontam para o aumento da contribuição europeia com o afastamento do litoral. Para inferir a origem de algumas doenças genéticas em Monte Santo foram analisados marcadores informativos de ancestralidade autossômicos (AT3-I/D, APO, PV92 e SB19.3 genotipados por PCR; GC*1F e GC*1S por PCR/RFLP; e os marcadores FYnull, CKMM e LPL por PCR em tempo real) e marcadores uniparentais do mtDNA (sequenciamento da região HVS-I) e do cromossomo Y (marcador YAP por PCR; DYS 199, 92R7 e M207 por PCR/RFLP; e M60, PN2, PN3, M34, M89, M9 por sequenciamento). Assim, através da identificação da origem desses marcadores foi possível inferir a contribuição das populações que formaram a população de Monte Santo, e a origem de algumas das alterações gênicas responsáveis pelas doenças genéticas aqui estudadas (síndrome de Treacher Collins, hipotireoidismo congênito, fenilcetonúria, mucopolissacaridose tipo VI, surdez hereditária não sindrômica e osteogênese imperfeita). Os dados do cromossomo Y e dos autossômicos apontam para maior contribuição europeia, e os resultados dos marcadores mitocondriais para elevada contribuição africana e ameríndia. A elevada contribuição europeia tanto paterna quanto autossômica sugere origem europeia para as mutações c.35delG e R252W, responsáveis por aproximadamente 24% dos casos de surdez hereditária não sindrômica e por todos os casos de fenilcetonúria, respectivamente. A mucopolissacaridose do tipo VI tem como causa a mutação p.H178L, a presença desta alteração apenas em pacientes brasileiros, que compartilham o mesmo haplótipo intragênico sugere origem autóctone. Além de marcadores moleculares também foram analisados os tipos de casamentos (endogâmicos, exogâmicos e entre imigrantes) e sua frequência no município. Foi observada elevada frequência de casamentos endogâmicos e baixa taxa de migração, sugerindo crescimento populacional interno. Além disso, a maioria da população reside em povoados, cujo tamanho varia de 113 a 582 pessoas por povoado. Nesta cidade 80% da população tem renda mensal equivalente a meio salário mínimo, o que explica a baixa taxa de migração por ausência de atrativos econômicos. Avaliando os casamentos dentro das genealogias dos afetados é possível observar que a maioria deles é filho de pais consanguíneos. Estes resultados mostram que o elevado grau de endogamia e endocruzamento assim como possível efeito fundador e deriva genética estão associados ao aumento da frequência e manutenção das doenças genéticas neste município. / Migration is the evolutionary factor able to disperse the genetic diversity among populations, inserting new phenotypic and genotypic characteristics. The dynamic of marriage and population structure are factors that may maintain or eliminate these characteristics. Examples of these traits are genetic diseases, where the frequency of these helps in understanding the evolutionary factors influence in a population. In Monte Santo city, situated in county of Bahia, were found genetic diseases with high frequency such as mucopolysaccharidosis type VI and phenylketonuria. It has been shown that some diseases have an important racial factor in determining risk of its occurrence. Molecular results show that in Bahia the African contribution is 47.2%. However, phenotypic classification data show an increase of European contribution with the distance from the coast. To infer the origin of some genetic disease in Monte Santo were analyzed autosomal ancestry informative markers (AT3-I/D, APO, PV92 and SB19.3 genotyped by PCR, GC*1F and GC*1S by PCR/RFLP and FYnull, CKMM and LPL genotyped by real time PCR) and uniparental markers of mtDNA (sequencing of the HVS-I region) and the Y chromosome (YAP marker by PCR; DYS199, 92R7 and M207 by PCR/RFLP, and M60, PN2, PN3, M34, M89, M9 by sequencing). Thus, by identifying the origin of these markers was possible to infer the contribution of the populations that formed Monte Santo, and the origin of some genetic mutations responsible for genetic diseases studied here (Treacher-Collins syndrome, congenital hypothyroidism, phenylketonuria, mucopolysaccharidosis type VI, hereditary non-syndromic deafness and osteogenesis imperfecta). The Y chromosome and autosomal results indicate greater European contribution, and the results from mtDNA show high contribution of African and Amerindian contribution. The high European contribution both paternal and autosomal suggests European origin for the c.35delG and R252W mutations, responsible for approximately 24% of cases of hereditary non-syndromic deafness and all phenylketonuria cases, respectively. The mucopolysaccharidosis type VI is caused by p.H178L mutation, the presence of this mutation only in Brazilian patients, who share the same intragenic haplotype suggest an autochthonous origin. In addition to molecular markers were also analyzed the types of marriages (endogamic, exogamous and between immigrant) and how often they occur in the city. We observed a high frequency of endogamic marriages and low migration rates, suggesting internal population growth. The population of Monte Santo is characterized by division into villages, where the majority of the population, the number of inhabitants varies from 113 to 582 people per village. In this city 80% of the population has income equivalent to half the minimum wage, which reinforces the absence of compelling economic and low migration rate. Evaluating the marriages inside the genetic diseases pedigree families can be observed that most of those affected are children of consanguineous parents. These results suggest that the high degree of inbreeding as well as the occurrence of founder effect and genetic drift were associated with increased frequency and maintenance of genetic diseases in the city.

Introdução: A Hipercolesterolemia Familiar (HF) é uma doença genética caracterizada clinicamente por elevados níveis de lipoproteína de baixa densidade (LDL-C) na corrente sanguínea desde a infância. Indivíduos que apresentam HF podem desenvolver doença aterosclerótica ainda em idade jovem. Os principais preditores de risco no desenvolvimento da doença cardiovascular (DCV) nesses indivíduos após entrarem em um programa de rastreamento genético não são conhecidos na nossa população. Além disso, a HF é subdiagnosticada e subtratada mundialmente e o rastreamento genético em cascata dos familiares tem sido mundialmente avaliado como o método diagnóstico mais custo. Contudo, a efetividade do rastreamento genético em cascata é dependente dos critérios clínicos de entrada do primeiro indivíduo da família e não há um consenso de qual critério apresenta a melhor acurácia para detecção de uma mutação. Objetivos: Identificar os fatores determinantes para ocorrência de eventos cardiovasculares (CV) em todos os indivíduos da coorte e avaliar o critério clínico para detecção de uma variante genética patogênica para HF, no primeiro indivíduo da família, após serem inseridos em um programa de rastreamento genético em cascata.Métodos: Estudo de coorte prospectiva aberta dos pacientes que foram inseridos no programa de rastreamento genético em cascata para HF. A população do estudo é definida como caso índice (CI), o primeiro da família a ser identificado clinicamente e encaminhado para o teste genético, e os familiares, que são os parentes de 1º grau do CI em que foi encontrada uma alteração genética. Todos os indivíduos são inseridos na coorte no momento em que recebem o laudo genético (tempo zero, T0). Um ano depois do T0 é realizado o primeiro contato telefônico, ou seja, primeiro ano de seguimento (T1) Resultados: No T1, o total de 818 indivíduos foi incluído, sendo verificados 47 eventos CV, sendo 14 (29,7%) fatais. Para o CI, o único fator independente associado ao aumento do risco de eventos CV no T1 foi a presença de arco corneano (OR: 9,39; IC 95%: 2,46-35,82). Para os familiares com uma mutação positiva os fatores associados ao aumento do risco de eventos CV foram diabetes mellitus (OR: 7,97; IC 95%: 2,07-30,66) e consumo de tabaco (OR: 3,70; IC 95%: 1,09-12,50). Na análise do melhor critério clínico para detecção de uma mutação patogênica no CI os valores de LDL-C >= 230 mg/dL tiveram a melhor relação entre sensibilidade e especificidade. Na análise da curva ROC o escore Dutch Lipid Clinic Network (DLCN) apresentou melhor desempenho do que o LDL-C para identificar uma mutação, a área sob a curva ROC foi 0,744 (IC 95%: 0,704-0,784) e 0,730 (IC 95%: 0,687-0,774), respectivamente, p = 0, 014. Conclusão: Em um ano de seguimento essa coorte identificou uma alta incidência de eventos CV após a entrada em um programa de rastreamento genético em cascata e os preditores dos eventos CV diferem entre CI e familiares. Esses resultados podem contribuir para o desenvolvimento de ações preventivas nesse grupo altamente susceptível de indivíduos. Além disso, devido a importância da detecção da mutação para um diagnóstico definitivo de HF e a importância da cascata ser custo efetiva o estudo identificou que o critério único do LDL-C >= 230 mg/dl é viável para indicar o CI para o teste genético / Introduction: Familial Hypercholesterolemia (FH) is a genetic disease characterized clinically by high levels of low density lipoprotein (LDL-C) in the bloodstream since childhood. Individuals with FH can develop atherosclerotic disease at a young age. The main predictors of cardiovascular disease (CVD) risk in these individuals after entering a genetic screening program are not known in our population. In addition, FH is underdiagnosed and undertreated worldwide and cascaded genetic screening of family members has been evaluated globally as the most cost effective for the diagnosis of FH. However, the effectiveness of cascading genetic screening is dependent on the clinical entry criteria of the first individual in the family and there is no consensus as to which criterion shows the best accuracy for detecting a mutation. Objectives: To identify the determinant factors for cardiovascular (CV) events in all individuals in the cohort and to evaluate the clinical criteria for detecting a genetic variant pathogenic to FH in the first individual of the family after being inserted into a genetic screening program in cascade. Methods: Open prospective cohort study of patients who were enrolled in the cascade genetic screening program for FH. The study population is defined as index case (IC), the first of the family to be clinically identified and referred to the genetic test, and relatives, who are the first-degree relatives of the IC in which a genetic alteration was found. All individuals are inserted into the cohort at the moment they receive the genetic report (time zero, T0). The first follow-up telephone contact is made one year after T0 (first year of follow-up, T1). Results: In T1, a total of 818 subjects were included, and 47 CV events were verified, of which 14 (29.7%) were fatal. For IC, the only factor independently associated with the increased risk of CV events in T1 was the presence of a corneal arch (OR: 9.39; 95% CI: 2.46-35.82). For relatives with positive mutation, factors associated with increased risk of CV events were diabetes mellitus (OR: 7.97; 95% CI: 2.07-30.66) and tobacco consumption (OR: 3.70; 95% CI: 1.09-12.50). In the analysis of the best clinical criteria for the detection of a pathogenic mutation in the IC, the LDL-C values >= 230 mg/dL had the best relationship between sensitivity and specificity. In the ROC curve analysis, the Dutch Lipid Clinic Network (DLCN) score performed better than LDL-C to identify a mutation, the area under the ROC curve was 0.744 (95% CI: 0.704-0.784) and 0.730 (CI 95 %: 0.687-0.774), respectively, p = 0.014. Conclusion: At one year follow-up this cohort identified a high incidence of CV events following entry into a cascade genetic screening program and the predictors of CV events differ between IC and family members. These results may contribute to the development of preventive actions in this group highly susceptible to individuals. In addition, because of the importance of detecting the mutation for a definitive diagnosis of HF and the importance of the cascade being cost effective, the study identified that the single LDL-C criterion >= 230 mg / dl is feasible to indicate IC for the genetic test

Alors qu’il n’existe pas de définition consensuelle du concept de maladie génétique, ce concept s’est progressivement élargi pour désigner des maladies communes, non héréditaires, non mendéliennes et polygéniques, aboutissant à une généticisation des maladies. Pour résoudre ce paradoxe de la génétique médicale contemporaine, les philosophes réfutent généralement cette généticisation comme une extension génocentriste abusive du concept de maladie génétique et cherchent à redéfinir un concept plus strict de maladie génétique. Nous montrons que cette stratégie échoue et proposons au contraire d’abandonner le concept de maladie génétique et de supposer que la généticisation révèle l’élaboration d’une explication du rôle commun des gènes dans toutes les maladies, que nous appelons une « théorie génétique de la maladie ». Nous définissons les conditions de possibilité et les critères nécessaires d’une théorie génétique a minima et aboutissons à un spectre des théories génétiques possibles. Nous proposons alors de tester si la généticisation des maladies révèle plutôt une théorie génétique des maladies, c’est-à-dire un ensemble de théories génétiques spécifiques à chaque classe de maladie, ou une théorie génétique de la maladie, reposant sur une définition générale de la maladie qui unifie le rôle commun des gènes dans toutes les maladies. Pour ce faire, nous analysons deux exemples de théories génétiques contemporaines : la théorie génétique des maladies infectieuses et la théorie génétique de la médecine des réseaux. Nous concluons à la coexistence nécessaire de plusieurs formes de théories génétiques dans la littérature biomédicale contemporaine. / While there is no consensual definition of the concept of genetic disease, this concept has gradually extended to designate common, non-hereditary, non-Mendelian, polygenic diseases, leading to the geneticization of diseases. In order to solve this paradox of the contemporary medical genetics, philosophers usually discard geneticization as an inappropriate genocentrist extension of the concept of genetic disease and attempt to define a stricter concept of genetic disease. We demonstrate the failure of this strategy and argue on the contrary that we should give up the concept of genetic disease and understand geneticization as the elaboration of an explanation of the common role of genes in diseases, what we call “a genetic theory of disease”. We define the conditions of possibility and the necessary criteria for a genetic theory a minima and end up with describing the spectrum of potential genetic theories. We then suggest to test whether geneticization of diseases reveals rather a genetic theory of diseases, that is, a set of genetic theories specific to each class of disease, or a genetic theory of disease, that is, a general definition of disease unifying the common role of genes in disease explanations. In order to do so, we analyse two examples of contemporary genetic theories: the genetic theory of infectious diseases and the genetic theory of network medicine. We conclude that several forms of genetic theories coexist in the contemporary biomedical literature and that this coexistence is necessary.