Molecular dissection of structural variations involved in antithrombin deficiency

Inherited antithrombin deficiency, the most severe form of thrombophilia, is predominantly caused by variants in SERPINC1. Few causal structural variants have been described, usually detected by multiplex ligation-dependent probe amplification or cytogenetic arrays, which only define the gain or loss and the approximate size and location. This study has done a complete dissection of the structural variants affecting SERPINC1 of 39 unrelated patients with antithrombin deficiency using multiplex ligation-dependent probe amplification, comparative genome hybridization array, long-range PCR, and whole genome nanopore sequencing. Structural variants, in all cases only affecting one allele, were deleterious and caused a severe type I deficiency. Most defects were deletions affecting exons of SERPINC1 (82.1%), but the whole cohort was heterogeneous, as tandem duplications, deletion of introns, or retrotransposon insertions were also detected. Their size was also variable, ranging from 193 bp to 8 Mb, and in 54% of the cases involved neighboring genes. All but two structural variants had repetitive elements and/or microhomologies in their breakpoints, suggesting a common mechanism of formation. This study also suggested regions recurrently involved in structural variants causing antithrombin deficiency and found three structural variants with a founder effect: the insertion of a retrotransposon, duplication of exon 6, and a 20-gene deletion. Finally, nanopore sequencing was determined to be the most appropriate method to identify and characterize all structural variants at nucleotide level, independently of their size or type.Supported by the National Institute for Health Research (NIHR) for the NIHR BioResource project (grant numbers RG65966 and RG94028), by the Instituto de Salud Carlos III grant; Fondo Europeo de Desarrollo Regional (FEDER) grant PI18/00598; and Fundación Séneca 19873/GERM/15. M.E.d.l.M.-B. has a postdoctoral contract from University of Murcia, Murcia, Spain. C.B.-P. has a Río Hortega fellowship. B.d.l.M.-B. has a postdoctoral fellowship from Fundación Séneca. J.C.-G. has a predoctoral fellowship from the Ministry of Universities FPU19/03662

The FXII c.-4T > C Polymorphism as a Disease Modifier in Patients With Hereditary Angioedema Due to the FXII p.Thr328Lys Variant

Background: Hereditary angioedema due to the Thr328Lys variant in the coagulation factor XII (HAE-FXII) affects mainly women in whom the symptomatology is dependent on high estrogen levels. Clinical variability and incomplete penetrance are challenging features that hinder the diagnosis and management of HAE-FXII. The c.-4T>C Kozak polymorphism is the only common variation accounting for FXII plasma levels and was previously shown to modify the course of HAE due to C1-Inhibitor deficiency. Objectives: To assess the influence of the c.-4T>C polymorphism on disease expression in 39 Spanish HAE-FXII index patients. Methods: The c.-4T>C polymorphism was sequenced by the standard Sanger method, and HAE severity was calculated according to the score by Cumming et al. (2003) The activation of the contact system was quantified by the kallikrein-like activity of plasma in chromogenic assays upon activation with high-molecular-weight dextran sulfate. Results: The c.-4CC genotype was overrepresented in the studied cohort: 82% were CC-homozygous (expected frequency = 59%) and 18% were CT-heterozygous (expected frequency = 39%) (p = 0.001). Patients with a c.-4CC genotype exhibited higher kallikrein-like activity (0.9659 +/- 0.1136) than those with a c.-4TC genotype (0.7645 +/- 0.1235) (p = 0.024) or healthy donors. Moreover, the polymorphism influenced HAE-FXII severity score (c.-4CC = 4.43 +/- 2.28 vs c.-4TC = 2.0 +/- 1.15; p = 0.006) but not the degree of estrogen dependence or time until remission. Conclusion: The c.-4T>C polymorphism is overrepresented in a Spanish HAE-FXII cohort and significantly influences the degree of contact system activation and the clinical severity of the disease

Defects of splicing in antithrombin deficiency

Background: There is increasing evidence supporting the relevance of aberrant splicing in multiple disorders. In antithrombin deficiency only 22 intronic mutations affecting splicing sites (7% of SERPINC1 mutations) are considered as splicing mutations. Methods: SERPINC1 was analyzed by Sanger sequencing and MLPA in 141 unrelated cases with antithrombin deficiency. Plasma antithrombin was studied by functional and western blot assays, purified by FPLC and characterized by proteomic analysis. In silico predictions on splicing was done with the Human Splicing Finder software. Results: We detected 89 different SERPINC1 defects, 13 with potential effect on splicing. Ten cases presented 9 mutations disturbing splicing sites, 5 new. Three gross or small gene defects also disturbed a correct splicing. Interestingly, the first duplication of a single exon ever described (c.1154-13_1218+115dup), caused mild deficiency (75%). A deeper intronic mutation (c.1154-14G>A), identified in three unrelated patients with traces of disulphide dimers of antithrombin in plasma, created a cryptic splicing site that might generate a variant with 4 additional in frame residues according to in silico predictions. This aberrant splicing was confirmed by proteomic analysis of the dimer purified from plasma. Conclusions: A high proportion of cases with antithrombin deficiency (up to 13%) may be explained by an aberrant splicing. Up to 15% of mutations in SERPINC1: splicing site variations, gross gene defects and deep intronic mutations, may affect a correct splicing with three potential consequences type I, type II, and even moderate antithrombin deficiency

Congenital antithrombin deficiency in patients with splanchnic vein thrombosis

Splanchnic vein thromboses (SVT) are a rare condition that can be life-threatening. The most severe thrombophilia associated to SVT is antithrombin (AT) deficiency, usually caused by SERPINC1 mutations. Although transitory AT deficiencies and congenital disorders of the N-glycosylation pathways (CDG) have been recently reported as causes of AT deficiency, the current AT clinical screening still only includes anti-FXa activity. This study aims to 1) improve the detection of antithrombin deficiency in SVT and 2) characterize the features of antithrombin deficiency associated with SVT.The study was performed in 2 cohorts: 1) 89 SVT patients with different underlying etiologies but in whom AT deficiency had been ruled out by classical diagnostic methods; and 2) 271 unrelated patients with confirmed AT deficiency and venous thrombosis. Antithrombin was evaluated by functional (anti-FXa and anti-FIIa) and immunological methods (ELISA, crossed immunoelectrophoresis, western blot), and SERPINC1 sequencing was performed.In 4/89 patients (4.5%) additional alterations in AT were found (two had SERPINC1 mutations, one had a specific variant causing transient AT deficiency and one patient had CDG). In 11 of the 271 patients (4.1%) with AT deficiency and thrombosis, thrombosis was located at the splanchnic venous territory.AT deficiency may be underdiagnosed by current clinical screening techniques. Therefore, a comprehensive AT evaluation should be considered in cases of rethrombosis or doubtful interpretation of anti-FXa activity levels. SVT is a relatively common localization of the thrombotic event in patients with congenital AT deficiency.© 2019 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd

Expanding the genetic spectrum of TUBB1-related thrombocytopenia

β1-Tubulin plays a major role in proplatelet formation and platelet shape maintenance, and pathogenic variants in TUBB1 lead to thrombocytopenia and platelet anisocytosis (TUBB1-RT). To date, the reported number of pedigrees with TUBB1-RT and of rare TUBB1 variants with experimental demonstration of pathogenicity is limited. Here, we report 9 unrelated families presenting with thrombocytopenia carrying 6 β1-tubulin variants, p.Cys12LeufsTer12, p.Thr107Pro, p.Gln423*, p.Arg359Trp, p.Gly109Glu, and p.Gly269Asp, the last of which novel. Segregation studies showed incomplete penetrance of these variants for platelet traits. Indeed, most carriers showed macrothrombocytopenia, some only increased platelet size, and a minority had no abnormalities. Moreover, only homozygous carriers of the p.Gly109Glu variant displayed macrothrombocytopenia, highlighting the importance of allele burden in the phenotypic expression of TUBB1-RT. The p.Arg359Trp, p.Gly269Asp, and p.Gly109Glu variants deranged β1-tubulin incorporation into the microtubular marginal ring in platelets but had a negligible effect on platelet activation, secretion, or spreading, suggesting that β1-tubulin is dispensable for these processes. Transfection of TUBB1 missense variants in CHO cells altered β1-tubulin incorporation into the microtubular network. In addition, TUBB1 variants markedly impaired proplatelet formation from peripheral blood CD34+ cell-derived megakaryocytes. Our study, using in vitro modeling, molecular characterization, and clinical investigations provides a deeper insight into the pathogenicity of rare TUBB1 variants. These novel data expand the genetic spectrum of TUBB1-RT and highlight a remarkable heterogeneity in its clinical presentation, indicating that allelic burden or combination with other genetic or environmental factors modulate the phenotypic impact of rare TUBB1 variants.This work was partially supported by grants from Instituto de Salud Carlos III (ISCIII) and Feder (PI17/01311, PI17/01966, PI20/00926 and CB15/00055), Fundacion Séneca (19873/ GERM/15), Gerencia Regional de Salud (GRS 2061A/19 and 1647/A/17), Fundacion Mutua Madrile´ña (AP172142019), and ~ Sociedad Espanola de Trombosis y Hemostasia (Premio L ~ opez Borrasca 2019 and Ayuda a Grupos de Trabajo en Patologıa Hemorragica). The authors’ research on inherited platelet disorders is conducted in accordance with the aims of the Functional and Molecular Characterization of Patients with Inherited Platelet Disorders Project, from Grupo Espanol de Alteraciones Plaqueta- ~ rias Congenitas, which is supported by the Spanish Society of Thrombosis and Haemostasis. V.P.-B. has a predoctoral contract from CIBERER. L.B. was supported by a fellowship from Fondazione Umberto Veronesi. M.E.d.l.M.-B. holds a postdoctoral fellowship from the University of Murcia. A.M.-Q. holds a predoctoral grant from the Junta de Castilla y Leon

Análisis del sistema hemostático en neoplasias avanzadas

Introducción La relación entre sistema hemostático y cáncer es bidireccional. Los pacientes con cáncer tienen 4-7 veces más riesgo de desarrollar trombosis, especialmente en estadios avanzados de la enfermedad porque los tratamientos, las comorbilidades y las propias células tumorales, especialmente ciertos tipos, provocan diferentes alteraciones en el sistema hemostático que generan un estado de hipercoagulabilidad. Por otra parte, diferentes elementos del sistema hemostático juegan un papel importante en el crecimiento tumoral, progresión y metastatización. Hipótesis y objetivos La hipótesis que generó este trabajo fue que las distintas neoplasias, por el tipo de tumor o por los tratamientos recibidos, podrían provocar una alteración diferencial del sistema hemostático que podría tener relación con el desarrollo de trombosis y con la progresión tumoral. Por ello, el objetivo de nuestro estudio fue evaluar diferentes elementos del sistema hemostático en pacientes con 3 de las 4 neoplasias más frecuentes (cáncer de próstata, colorrectal y de pulmón). Los objetivos específicos fueron: 1. Estudiar si existe una activación diferente del sistema hemostático en distintas neoplasias avanzadas. 2. Analizar la relación entre alteraciones del sistema hemostático y el desarrollo de eventos trombóticos en pacientes con cáncer. 3. Estudiar el efecto de la quimioterapia basada en fluoropirimidinas sobre el sistema hemostático en pacientes con cáncer colorrectal. 4. Estudiar el valor pronóstico de las principales alteraciones del sistema hemostático en pacientes con cáncer. Material y métodos Se incluyeron muestras basales de 157 pacientes consecutivos; 32 con cáncer de próstata resistente a castración; 43 con cáncer colorrectal y 82 con carcinoma de pulmón. Se estudiaron: plaquetas y leucocitos; marcadores de muerte celular (lactato deshidrogenasa –LDH-); hipercoagulabilidad (Dímero D) y de daño endotelial (Factor von Willebrand –FvW-) y tiempos de coagulación (Tiempo de protrombina). Se analizaron la antitrombina y los dos elementos de la ruta de contacto (FXI y FXII) mediante métodos funcionales cromogénicos y/o western blot. Finalmente se estudiaron dos elementos recientemente implicados en trombosis: trastornos de glicosilación y ADN circulante (estudiados mediante electroforesis y HPLC; y por un método de cuantificación fluorescente, respectivamente). En casos excepcionales se realizaron estudios genéticos: secuenciación de genes (SERPINC1 y PMM2) o detección de grandes deleciones mediante MLPA. Resultados y Conclusiones 1. El sistema hemostático se encuentra afectado en pacientes con cáncer. Aunque hay elementos alterados comunes en las distintas neoplasias estudiadas, como son la elevación basal de LDH, DD y FvW, hay otros que varían entre ellas. En cáncer de próstata observamos una tendencia a una mayor activación de FXII que no se acompaña de activación de FXI. En cáncer colorrectal, se observó elevación de FvW y DD, sin activación de la ruta de contacto. En pacientes con CNMP, observamos una mayor incidencia de deficiencia de antitrombina que probablemente sea por consumo como lo sugiere la detección de complejos trombina-antitrombina. 2. La presencia de elevación del FvW en pacientes con cáncer de próstata se asocia con el desarrollo de eventos trombóticos en nuestra serie. No se ha encontrado relación entre el desarrollo de fenómenos trombóticos y otras alteraciones del sistema hemostático. 3. La quimioterapia basada en fluoropirimidinas no provoca alteraciones relevantes del sistema hemostático en pacientes con cáncer colorrectal. 4. Determinadas alteraciones del sistema hemostático, como el aumento de FvW, DD y ADN circulante, se asocian con peor pronóstico en las neoplasias estudiadas. La elevación de FvW se asocia a peor pronóstico en tres de los cuatro tumores estudiados: cáncer de próstata, cáncer colorrectal y CNMP. La elevación de DD se asocia con peor pronóstico en cáncer de próstata. Finalmente, la presencia de concentraciones elevadas de ADN circulante se asocia con peor supervivencia en pacientes con cáncer de próstata y CNMP avanzado. ABSTRACT Introduction There is a strong relationship between the hemostatic system and cancer. Patients with cancer have increased risk (4-7-fold) to develop thrombosis, particularly in advanced stages of the disease, because the treatments, the associated co-morbidities and the features of cancer cells can modify the hemostatic balance, triggering a prothrombotic state. But also the different hemostatic elements play a key role in tumor growth, tumor progression and metastasis. Hypothesis and objectives We propose that the disturbance of the hemostatic system may be different depending of the type of cancer. The type of tumor cell or the treatment used in each tumor may have distinct consequences on the hemostatic balance with potential and different consequences on the risk of thrombosis and the progression of the tumor. Accordingly, the main objective of our study was to evaluate different elements of the haemostatic system in patients with three out of four more prevalent cancer (prostate, colorectal, and lung cancer). The specific objectives were: 1. To study a potential activation of the hemostatic system in different advanced tumors. 2. To analyse the relationship between changes in the hemostatic system and the development of thrombotic events in patients with cancer. 3. To evaluate the effect of chemotherapy based on fluoropirimidines on the hemostatic system in patients with colorectal cancer. 4. To study the prognostic value of the main variations of the hemostatic system identified in patients with cancer. Material and methods Our study included basal samples of 157 consecutive patients with cancer: 32 with prostate cancer resistant to castration; 43 with colorectal cancer; and 82 with lung cancer. These were the parameters analyzed: platelets and leukocites; markers of cell death (Lactato de-hidrogenase –LDH-); hypercoagulability (D Dimer) and vascular damage (von Willebrand Factor –vWF-). Clotting times (prothrombin time and activated partial thromboplastin time) were also evaluated. Additionally, antithrombin and two elements of the contact pathway of coagulation (FXI and FXII) were also evaluated by functional chromogenic methods and/or Western blot. Finally, two elements recently associated with thrombosis: disorders of glycosylation and circulating DNA, were also analyzed in this study by electrophoresis and HPLC and by a fluorogenic method, respectively. Exceptional cases required genetic analysis that included senquencing of genes (SERPINC1 or PMM2) or detection of gross gene defects by MLPA. Results and conclusions 1. The haemostatic system is altered in cancer patients. Several common alterations are observed, including high levels of LDH, D-dimers and von Willebrand Factor (vWF). However, several distinct alterations are observed: in prostate cancer we observe activation of Factor XII; in colo-rectal cancer no activation of the contact route is observed and n non-small cell lung cáncer we observed lower levels of antitrombin factor probably caused by consumption, as these patients also had remarkable levels of thrombin-antithrombin complexes. 2. The presence of higher levels of vWF is associated with the development of thromboembolic events in prostate cancer. We did not observe other associations between thrombosis and alteration of the haemostatic system. 3. Fluoropirimidine-based chemotherapy did not cause relevant disturbances on the hemostatic system in patients with colorectal cancer. 4. Some variations of the hemostatic system such as the increase of vWF, D dimer or circulating DNA, associated with a poor prognosis in cancer. Thus, increased levels of vWF associated with a poor prognosis in three out of four tumors evaluated in this study: prostate, colorectal and non microcytic lung cancer. Rise level of D dimer was also a marker of bad prognosis in prostate cancer. Finally, high values of circulating DNA associated with low survival in patients with prostate and non microcytic lung cancer

Nuevas alteraciones genéticas y mecanismos implicados en la deficiencia de antitrombina

INTRODUCCIÓN La antitrombina es un anticoagulante natural que juega un papel crucial en la hemostasia. Las diferentes proteasas procoagulantes diana y el potente mecanismo inhibidor de esta serpina explican que incluso reducciones moderadas de antitrombina aumenten significativamente el riesgo de trombosis venosa. Así, la deficiencia de antitrombina es la primera y más grave trombofilia, estudiada ampliamente durante más de 50 años. Sin embargo, aún existen retos en torno a este anticoagulante, como conocer los factores implicados en la heterogeneidad clínica, caracterizar las variantes estructurales implicadas y describir la base molecular del 25% de casos sin defecto genético conocido. En esta tesis hemos intentado correlacionar datos genéticos y clínicos en una gran cohorte de pacientes con deficiencia de antitrombina. OBJETIVO El objetivo de esta tesis es aportar información nueva y original sobre este trastorno. Específicamente, pretendíamos describir el impacto clínico de la deficiencia de antitrombina. Además, quisimos caracterizar en profundidad las variantes estructurales implicadas en la deficiencia de antitrombina, e identificar nuevos defectos moleculares responsables de este trastorno. MÉTODOS Hemos estudiado una de las mayores cohortes de pacientes con esta enfermedad rara reclutada durante más de 20 años. En todos los pacientes, se ha realizado estudios funcionales, bioquímicos y moleculares y se han recopilado datos clínicos. Para estudios específicos se empleó expresión recombinante y una novedosa técnica para el análisis de haplotipos. Además, se han utilizado diferentes tecnologías para la caracterización de los defectos moleculares en SERPINC1, incluyendo la secuenciación de tercera generación y la PCR larga. RESULTADOS La deficiencia de antitrombina incrementa notablemente el riesgo de trombosis pediátrica (300 veces mayor que el de la población general). Por el contrario, hemos caracterizado una deficiencia leve de antitrombina de difícil diagnóstico funcional causada por p.Thr147Ala, una mutación con efecto fundador en la población africana. El estudio de la mayor cohorte de variantes estructurales que causan deficiencia de antitrombina empleando diferentes métodos, incluido la secuenciación del genoma completo con tecnología de nanoporos, mostró que son heterogéneas en tamaño y tipo e identificó por primera vez una variante estructural compleja y la inserción de un nuevo retrotransposon. Describimos un mapa detallado de variantes estructurales con zonas calientes intra e intergénicas, y encontramos un mecanismo común que involucra elementos repetitivos, para la formación de variantes estructurales causantes de este desorden. CONCLUSIONES Nuestro estudio proporciona información nueva y original sobre la trombofilia más grave. El alto riesgo de trombosis pediátrica apoya el cribado de ciertos tipos de deficiencia de antitrombina en niños de familias afectadas para beneficiarse de estrategias preventivas. Las mutaciones fundadoras expandidas en diferentes poblaciones, como la descrita en nuestro estudio en población africana, causan deficiencias leves de difícil diagnóstico funcional. Identificamos y caracterizamos un nuevo mecanismo que causa deficiencia de antitrombina: la inserción de un retrotransposón en un intrón. Además, diseccionamos las variantes estructurales implicadas en deficiencia de antitrombina. Nuestro estudio revela que la combinación de diferentes métodos es útil para la caracterización de estos grandes defectos genéticos, pero la secuenciación de nanoporos es el método más adecuado para su completa caracterización.INTRODUCTION Antithrombin is a natural anticoagulant that plays a crucial role in hemostasis. The wide range of target procoagulant proteases and the potent inhibitory mechanism of this serpin explain that even moderate reductions of antithrombin significantly increase the risk of venous thrombosis. Thus, antithrombin deficiency is the first and the strongest thrombophilia that has been widely studied during more than 50 years. However, there are still some challenges concerning this anticoagulant, such as the factors involved in the clinical heterogeneity, to do a better characterization of structural variants involved in antithrombin deficiency, and to describe the molecular base of up to 25% of cases with no genetic defect. In this thesis we have tried to correlate genetic and clinical data in a large cohort of patients with antithrombin deficiency. OBJECTIVE The objective of this thesis is to supply new and original information of this rare disorder. Specifically, we aimed to describe the clinical impact of antithrombin deficiency. Moreover, we also want to deeply characterize structural variants involved in antithrombin deficiency, and to identify new molecular defects involved in this disorder. METHODS We have studied one of the largest cohorts of patients with this rare disorder, recruited during more than 20 years. For all patients, a complete set of functional, biochemical and molecular studies have been done and clinical data has been collected. Specific studies included recombinant expression and a novel approach for haplotype analysis. New molecular technologies, including third generation sequencing and long range PCR, have been used for characterization of molecular defects in SERPINC1. RESULTS Antithrombin deficiency causes very high risk of pediatric thrombosis (300-fold compared with the general population). In contrast, we have characterized a mild antithrombin deficiency of difficult functional diagnosis caused by p.Thr147Ala, a mutation with founder effect in African population. The study by different methods, including whole genome sequencing with nanopore technology, of the largest cohort of structural variants causing antithrombin deficiency showed that they are heterogeneous in size and type, and identified for the first time a complex structural variant and an insertion of a new SVA element. We described a detailed map of structural variants with intra and intergenic hotspots, and we found a common mechanism for formation of structural variants causing antithrombin deficiency that involved repetitive elements. CONCLUSIONS Our study supplies new and original information on the strongest thrombophilia. The high risk of pediatric thrombosis supports the screening of certain types of antithrombin deficiency in children of affected families to benefit from preventive strategies. In contrast, founder mutations expanded in different populations like the first one described by our study in African population will probably cause a mild deficiency of difficult functional diagnosis. We identified and characterized a new mechanism causing antithrombin deficiency: the insertion of a retrotransposon in an intron. Moreover, we dissected the structural variants involved in antithrombin deficiency. Our study also reveals that the combination of different methods is useful for the characterization of these gross gene defects, but nanopore sequencing is the most suitable method to fully characterize them

Identificación de nuevos elementos implicados en la regulación de antitrombina= Identification of new elements involved in antithrombin regulation.

Resúmenes de 150 palabras en Inglés Deficiency of antithrombin caused by mutations affecting the gene encoding this key anticoagulant (SERPINC1) significantly increases the risk of thrombosis. We aim to identify new mechanisms involved in antithrombin deficiency. Using molecular, cellular and biochemical methods, we studied 29 patients with antithrombin deficiency without SERPINC1 mutation, a family study, three case-control studies including 2,980 patients and 3,996 controls, and two patients with congenital disorder of glycosylation (CDG). We identified the first mutation affecting the SERPINC1 promoter causing antithrombin deficiency. We confirmed the low genetic variability of SERPINC1 and its minor role on the heritability of antithrombin. Genome wide association studies and silencing experiments identified the first modulating gene of antithrombin, LARGE. We diagnosed a patient with CDG based on his antithrombin deficiency. Finally, we described a new disorder with identical biochemical features than CDGs, but only thrombosis, which is caused by a single mutation in PMM2 and concomitant alcohol consumption. Key Words: Antithrombin, Thrombosis, CDG, Mutations, SERPINC1 Resumen de 150 palabras en Castellano. La deficiencia de antitrombina causada por mutaciones en el gen SERPINC1 incrementa el riesgo trombótico. Nuestro objetivo fue identificar nuevos mecanismos implicados en la deficiencia de este anticoagulante. Empleando metodología molecular, celular y bioquímica estudiamos 29 pacientes con deficiencia de antitrombina sin mutaciones en SERPINC1, un estudio familiar, tres estudios caso-control (2,980 pacientes/3,996 controles) y dos pacientes con trastornos congénitos de glicosilación (CDG). Identificamos la primera mutación en el promotor de SERPINC1 que causa deficiencia de antitrombina. Confirmamos la baja variabilidad genética en SERPINC1 y su escasa influencia en la heredabilidad de antitrombina. Un GWAS y experimentos de silenciamiento mostraron que LARGE es el primer gen modulador de antitrombina. Diagnosticamos un CDG por la deficiencia de antitrombina de un paciente con trombosis recurrente y descubrimos nuevo desorden con patrón bioquímico similar al CDG pero solo con trombosis que es causado por una sola mutación en PMM2 y consumo de alcohol. Palabras clave: Antitrombina, Trombosis, CDG, Mutaciones, SERPINC

HIDDEN VARIANT OF ANTITHROMBIN DETECTED THROUGH COMPLETE GENOME SEQUENCING THAT MODULATES THE EFFECTIVENESS OF N-GLYCOSYLATION. CLINICAL AND BASIC IMPLICATIONS

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