Confirmación molecular de caso clínico diagnosticado en 1984 con atrofia muscular espinal tipo II (AME tipo II) y análisis familiar de portadores
Spinal muscular atrophy (SMA) is a neurodegenerative disease considered one of the most common and most devastating ?rare? diseases known, and one of the main inherited causes of infant mortality. It has an approximate incidence of 1/6000 to 1/10000 births, and a carrier frequency of 1/40 -1/60 in t...
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| Formato: | Tesis de maestría acceptedVersion |
| Lenguaje: | Español |
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Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica
2022
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| Acceso en línea: | http://repositoriouba.sisbi.uba.ar/gsdl/cgi-bin/library.cgi?a=d&c=afamaster&cl=CL1&d=HWA_6943 https://repositoriouba.sisbi.uba.ar/gsdl/collect/afamaster/index/assoc/HWA_6943.dir/6943.PDF |
| Aporte de: |
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I28-R145-HWA_6943 |
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dspace |
| institution |
Universidad de Buenos Aires |
| institution_str |
I-28 |
| repository_str |
R-145 |
| collection |
Repositorio Digital de la Universidad de Buenos Aires (UBA) |
| language |
Español |
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spa |
| topic |
AME Diagnostico molecular SMN1 SMN2 PCR-RFLP MLPA secuenciación Real Time PCR SMA Molecular Diagnosis SMN1 SMN2 PCR-RFLP MLPA Sequencing Real Time PCR Ciencias de la vida |
| spellingShingle |
AME Diagnostico molecular SMN1 SMN2 PCR-RFLP MLPA secuenciación Real Time PCR SMA Molecular Diagnosis SMN1 SMN2 PCR-RFLP MLPA Sequencing Real Time PCR Ciencias de la vida Ousset, María Julia Confirmación molecular de caso clínico diagnosticado en 1984 con atrofia muscular espinal tipo II (AME tipo II) y análisis familiar de portadores |
| topic_facet |
AME Diagnostico molecular SMN1 SMN2 PCR-RFLP MLPA secuenciación Real Time PCR SMA Molecular Diagnosis SMN1 SMN2 PCR-RFLP MLPA Sequencing Real Time PCR Ciencias de la vida |
| description |
Spinal muscular atrophy (SMA) is a neurodegenerative disease considered one of the most common and most devastating ?rare? diseases known, and one of the main inherited causes of infant mortality. It has an approximate incidence of 1/6000 to 1/10000 births, and a carrier frequency of 1/40 -1/60 in the general population. The gene that has been associated as a determinant of the disease is called SMN1 and codes for the SMN protein. This disease most affects the cells of the anterior horn of the spinal cord (motor neurons) and causes symmetric proximal weakness and progressive atrophy of muscle groups. The disease is classified into 4 groups according to the clinical picture based on the severity of the symptoms, the age of onset and its evolution.\nThe disease phenotypes range from severe neonatal manifestation to adult form with minimal weakness. The severity of the disease has been correlated with the number of copies of the SMN2 gene, a sequence homologous to SMN1, whose coding sequence differs by one nucleotide in exon 7 (c.840C> T), respect to SMN1. Both, SMN1 and SMN2, consist of nine exons that code for the 294 amino acid SMN protein. The SMN2 gene, unlike SMN1, produces only 10-25% active SMN protein. Currently, clinical research studies are being carried out with drugs that stimulate a greater synthesis of active protein from SMN2, these therapies range from antisense oligo therapies (Spinraza) to gene therapy (Zolgensma).\nThe most widely methodologies used for the molecular diagnosis of the disease are PCR-RFLP (restriction fragment length polymorphism), sequencing and MLPA (multiplex ligation dependent probe amplification) and for genotype-phenotype correlation studies, quantitative molecular techniques are used that allow performing gene dosing of SMN2 as real-time PCR or MLPA. Carriers of the disease can be identified by indirect methods by performing linkage analysis, or by the aforementioned quantitative methodologies.\nIn 2007 the Senate and the Chamber of Deputies of the Province of Buenos Aires passed a law (Law 13682) that declares the investigation and treatment of AME of provincial interest. In addition, this initiative establishes that the Executive Branch, through the Ministry of Health, implement specific programs aimed at disseminating the characteristics of the disease, promotes\nknowledge of its antecedents and effects, its methods of diagnosis and detection, informing about currently existing treatments and organizing regional health responses for patients affected by the disease. This information supports the need in the country for statistics and information on patients and carriers in the population and adequate genetic counseling for families with affected people to better understand the probabilities and risks of disease in their offspring.\nThe aim of this project was to study and confirm the clinical diagnosis of SMA in a patient using molecular techniques, and to identify carriers in the family. This was one of the first cases clinically diagnosed with SMA in Argentina in 1984, when not even the determining gene for the disease was yet known. During the development of this work, by means of different techniques, the SMA diagnosis of the patient was confirmed and two carriers were identified among their relatives. In contrast to what has been reported in gene dosing by real-time PCR for SMA, in this work the technique was developed with two reference genes instead of one. This minimizes false results due to polymorphisms in the primer binding sequences and gives a higher degree of certainty to the result obtained. |
| author2 |
Bilen, Marcos |
| author_facet |
Bilen, Marcos Ousset, María Julia |
| format |
Tesis de maestría Tesis de maestría acceptedVersion |
| author |
Ousset, María Julia |
| author_sort |
Ousset, María Julia |
| title |
Confirmación molecular de caso clínico diagnosticado en 1984 con atrofia muscular espinal tipo II (AME tipo II) y análisis familiar de portadores |
| title_short |
Confirmación molecular de caso clínico diagnosticado en 1984 con atrofia muscular espinal tipo II (AME tipo II) y análisis familiar de portadores |
| title_full |
Confirmación molecular de caso clínico diagnosticado en 1984 con atrofia muscular espinal tipo II (AME tipo II) y análisis familiar de portadores |
| title_fullStr |
Confirmación molecular de caso clínico diagnosticado en 1984 con atrofia muscular espinal tipo II (AME tipo II) y análisis familiar de portadores |
| title_full_unstemmed |
Confirmación molecular de caso clínico diagnosticado en 1984 con atrofia muscular espinal tipo II (AME tipo II) y análisis familiar de portadores |
| title_sort |
confirmación molecular de caso clínico diagnosticado en 1984 con atrofia muscular espinal tipo ii (ame tipo ii) y análisis familiar de portadores |
| publisher |
Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica |
| publishDate |
2022 |
| url |
http://repositoriouba.sisbi.uba.ar/gsdl/cgi-bin/library.cgi?a=d&c=afamaster&cl=CL1&d=HWA_6943 https://repositoriouba.sisbi.uba.ar/gsdl/collect/afamaster/index/assoc/HWA_6943.dir/6943.PDF |
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AT oussetmariajulia confirmacionmoleculardecasoclinicodiagnosticadoen1984conatrofiamuscularespinaltipoiiametipoiiyanalisisfamiliardeportadores |
| _version_ |
1840330186044735488 |
| spelling |
I28-R145-HWA_69432025-07-31 Spinal muscular atrophy (SMA) is a neurodegenerative disease considered one of the most common and most devastating ?rare? diseases known, and one of the main inherited causes of infant mortality. It has an approximate incidence of 1/6000 to 1/10000 births, and a carrier frequency of 1/40 -1/60 in the general population. The gene that has been associated as a determinant of the disease is called SMN1 and codes for the SMN protein. This disease most affects the cells of the anterior horn of the spinal cord (motor neurons) and causes symmetric proximal weakness and progressive atrophy of muscle groups. The disease is classified into 4 groups according to the clinical picture based on the severity of the symptoms, the age of onset and its evolution.\nThe disease phenotypes range from severe neonatal manifestation to adult form with minimal weakness. The severity of the disease has been correlated with the number of copies of the SMN2 gene, a sequence homologous to SMN1, whose coding sequence differs by one nucleotide in exon 7 (c.840C> T), respect to SMN1. Both, SMN1 and SMN2, consist of nine exons that code for the 294 amino acid SMN protein. The SMN2 gene, unlike SMN1, produces only 10-25% active SMN protein. Currently, clinical research studies are being carried out with drugs that stimulate a greater synthesis of active protein from SMN2, these therapies range from antisense oligo therapies (Spinraza) to gene therapy (Zolgensma).\nThe most widely methodologies used for the molecular diagnosis of the disease are PCR-RFLP (restriction fragment length polymorphism), sequencing and MLPA (multiplex ligation dependent probe amplification) and for genotype-phenotype correlation studies, quantitative molecular techniques are used that allow performing gene dosing of SMN2 as real-time PCR or MLPA. Carriers of the disease can be identified by indirect methods by performing linkage analysis, or by the aforementioned quantitative methodologies.\nIn 2007 the Senate and the Chamber of Deputies of the Province of Buenos Aires passed a law (Law 13682) that declares the investigation and treatment of AME of provincial interest. In addition, this initiative establishes that the Executive Branch, through the Ministry of Health, implement specific programs aimed at disseminating the characteristics of the disease, promotes\nknowledge of its antecedents and effects, its methods of diagnosis and detection, informing about currently existing treatments and organizing regional health responses for patients affected by the disease. This information supports the need in the country for statistics and information on patients and carriers in the population and adequate genetic counseling for families with affected people to better understand the probabilities and risks of disease in their offspring.\nThe aim of this project was to study and confirm the clinical diagnosis of SMA in a patient using molecular techniques, and to identify carriers in the family. This was one of the first cases clinically diagnosed with SMA in Argentina in 1984, when not even the determining gene for the disease was yet known. During the development of this work, by means of different techniques, the SMA diagnosis of the patient was confirmed and two carriers were identified among their relatives. In contrast to what has been reported in gene dosing by real-time PCR for SMA, in this work the technique was developed with two reference genes instead of one. This minimizes false results due to polymorphisms in the primer binding sequences and gives a higher degree of certainty to the result obtained. Fil: Ousset, María Julia. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Buenos Aires, Argentina Bilen, Marcos Giliberto, Florencia Ousset, María Julia 2022-03-28 La atrofia muscular espinal (AME) es una enfermedad neurodegenerativa considerada una de las enfermedades ?raras? más frecuentes y más devastadoras que se conocen, y una de las principales causas hereditarias de mortalidad infantil. Tiene una incidencia aproximada de 1/6000 a 1/10000 nacimientos, y una frecuencia de portadores de 1/40-1/60 en la población en general. El gen que se ha asociado como determinante de la enfermedad, se denomina SMN1 y codifica para la proteína SMN.\nEsta enfermedad afecta en mayor medida a las células del asta anterior de la médula espinal (neuronas motoras) y causa debilidad proximal simétrica y atrofia progresiva de los grupos musculares. La enfermedad se clasifica en 4 grupos según el cuadro clínico basado en la gravedad de los síntomas, la edad de aparición y la evolución de la misma. Los fenotipos de la enfermedad van desde la manifestación neonatal severa a la forma del adulto con mínima debilidad. La severidad de la enfermedad se ha correlacionado con el número de copias del gen SMN2, una secuencia homóloga a SMN1, cuya secuencia codificante difiere en un nucleótido en el exón 7 (c.840C>T), respecto a SMN1. Tanto SMN1 como SMN2 consisten en nueve exones que codifican para la proteína SMN de 294 amino ácidos. El gen SMN2, a diferencia de SMN1, produce sólo entre un 10%-25% de proteína SMN activa. Actualmente se están desarrollando estudios de investigación clínicos con medicamentos que estimulan una mayor síntesis de proteína activa a partir de SMN2, estas terapias van desde terapias con oligonucleótidos antisentido (Spinraza) hasta terapia génica (Zolgensma).\nLas metodologías más utilizadas para el diagnóstico molecular de la enfermedad son PCR-RFLP (restriction fragment length polymorphism), secuenciación y MLPA (múltiplex ligation dependent probe amplification) y para estudios de correlación genotipo-fenotipo se utilizan técnicas moleculares cuantitativas que permitan realizar dosaje génico de SMN2 como PCR en tiempo real o MLPA. Los portadores de la enfermedad se pueden identificar por métodos indirectos realizando análisis de ligamiento, o mediante las metodologías cuantitativas anteriormente mencionadas.\nEn el año 2007 el Senado y la Cámara de Diputados de la Provincia de Buenos Aires sancionó una ley (ley 13682) que declara de interés provincial la investigación y tratamiento de la AME, y además dicha iniciativa establece que el poder ejecutivo por intermedio del Ministerio de Salud, implemente programas específicos tendientes a difundir las características de la enfermedad, promover el conocimiento de sus antecedentes y efectos, sus métodos de diagnóstico y detección, informando sobre los tratamientos existentes en la actualidad y organizando respuestas sanitarias regionales para los pacientes afectados por la enfermedad.\nEsta información avala la necesidad en el país de estadísticas e información de enfermos y portadores en la población y el asesoramiento genético adecuado a familias con afectados para entender mejor las probabilidades y riesgos de enfermedad en su descendencia.\nEl objetivo del presente trabajo fue estudiar y confirmar mediante técnicas moleculares, el diagnóstico clínico de AME realizado a una paciente, e identificar los portadores en la familia. Este fue uno de los primeros casos diagnosticados clínicamente con AME en Argentina en 1984, cuando aún no se conocía ni siquiera el gen determinante de la enfermedad.\nDurante el desarrollo de este trabajo, mediante diferentes técnicas, se logró confirmar el diagnóstico AME de la paciente y se lograron identificar dos portadores entre sus familiares. En contraste con lo reportado en dosaje génico por PCR en tiempo real para AME, en este trabajo se puso a punto la técnica con dos genes de referencia en lugar de uno. Esto minimiza falsos resultados debidos a polimorfismos en las secuencias de unión de primers y da un grado de certeza mayor al resultado obtenido. application/pdf Cotignola, Javier Rossetti, Liliana de la Mata, Manuel AME Diagnostico molecular SMN1 SMN2 PCR-RFLP MLPA secuenciación Real Time PCR SMA Molecular Diagnosis SMN1 SMN2 PCR-RFLP MLPA Sequencing Real Time PCR spa Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica info:eu-repo/semantics/openAccess http://creativecommons.org/licenses/by-nc-nd/2.5/ar/ Ciencias de la vida Magíster de la Universidad de Buenos Aires en Biología Molecular Médica Confirmación molecular de caso clínico diagnosticado en 1984 con atrofia muscular espinal tipo II (AME tipo II) y análisis familiar de portadores info:eu-repo/semantics/masterThesis info:ar-repo/semantics/tesis de maestría info:eu-repo/semantics/acceptedVersion http://repositoriouba.sisbi.uba.ar/gsdl/cgi-bin/library.cgi?a=d&c=afamaster&cl=CL1&d=HWA_6943 https://repositoriouba.sisbi.uba.ar/gsdl/collect/afamaster/index/assoc/HWA_6943.dir/6943.PDF |