Choosing the Best Tissue and Technique to Detect Mosaicism in Fibrous Dysplasia/McCune-Albright Syndrome (FD/MAS).

GNAS-activating somatic mutations give rise to Fibrous Dysplasia/McCune-Albright syndrome (FD/MAS). The low specificity of extra-skeletal signs of MAS and the mosaic status of the mutations generate some difficulties for a proper diagnosis. We studied the clinical and molecular statuses of 40 patients referred with a clinical suspicion of FD/MAS to provide some clues. GNAS was sequenced using both Sanger and Next-Generation Sequencing (NGS). We were able to identify the pathogenic variants in 25% of the patients. Most of them were identified in the affected tissue, but not in blood. Additionally, NGS demonstrated the ability to detect more patients with mosaicism (8/34) than Sanger sequencing (4/39). Even if in some cases, the clinical information was not complete, we confirmed that, as in previous works, when the patients were young children with a single manifestation, such as hyperpigmented skin macules or precocious puberty, the molecular diagnosis was usually negative. In conclusion, as FD/MAS is caused by mosaic variants, it is essential to use sensitive techniques that allow for the detection of low percentages and to choose the right tissue to study. When not possible, and due to the low positive genetic rate, patients with FD/MAS should only be genetically tested when the clinical diagnosis is really uncertain.

Deficiencia de vitamina D en la edad adulta: presentación de 2 casos familiares de seudohipoparatiroidismo / Vitamin D deficiency in adulthood: Presentation of 2familial cases simulating pseudohypoparathyroidism

Antecedentes y objetivo El solapamiento clínico y bioquímico de diversas enfermedades del metabolismo fosfocálcico puede conllevar un erróneo diagnóstico y su consecuente abordaje clínico. Un ejemplo es el seudohipoparatiroidismo, que puede confundirse con el raquitismo dependiente de vitamina D (VDDR1) si no se hacen las determinaciones bioquímicas adecuadas. Pacientes y métodos Dos parejas de hermanos, de familias independientes, fueron diagnosticados clínicamente en la adolescencia de seudohipoparatiroidismo al presentar hipocalcemia, niveles elevados de hormona paratiroidea y valores normales o elevados de fósforo. Tras descartar alteraciones en GNAS, se realizó un estudio, mediante secuenciación masiva, de genes asociados a otros diagnósticos diferenciales. Resultados Se identificaron 2variantes genéticas en el gen CYP27B1 potencialmente asociadas con el fenotipo. Variantes patogénicas en este gen se asocian con VDDR1A. La reevaluación clínica-bioquímica de los pacientes confirmó dicho diagnóstico y se adecuó el tratamiento. Conclusiones Si bien la VDDR1A es un trastorno del metabolismo de diagnóstico infrecuente en la edad adulta, en casos de hipocalcemia con valores elevados de PTH es relevante la determinación de las formas 1,25(OH)2D3 y 25(OH)D3 de la vitamina D para alcanzar un diagnóstico correcto (AU)

Background and objective The clinical and biochemical overlap of various pathologies of phosphocalcic metabolism can lead to misdiagnosis and consequent clinical management. One example is pseudohypoparathyroidism, which can be confused with vitamin D-dependent rickets (VDDR1) if appropriate biochemical determinations are not performed. Patients and methods Two pairs of siblings, from independent families, were clinically diagnosed in adolescence with pseudohypoparathyroidism due to hypocalcaemia, elevated parathyroid hormone levels and normal or elevated phosphorus values. After ruling out alterations in GNAS, a massive sequencing study of genes associated with other differential diagnoses was carried out. Results Two genetic variants in the CYP27B1 gene potentially associated with the phenotype were identified. Pathogenic variants in this gene are associated with VDDR1A. Clinical-biochemical re-evaluation of the patients confirmed this diagnosis and treatment was adapted. Conclusions Although VDDR1A is an infrequently diagnosed pathology in adulthood, in cases of hypocalcaemia with elevated PTH values, determination of the 1,25(OH)2D3 and 25(OH)D3 forms of vitamin D is relevant to reach a correct diagnosis (AU)

Incidental finding at methylation-specific multiplex ligation-dependent probe amplification (MS-MLPA): how to proceed?

Introduction: Since the advent of new generation sequencing, professionals are aware of the possibility of obtaining findings unrelated to the pathology under study. However, this possibility is usually forgotten in the case of studies aimed at a single gene or region. We report a case of a 16-month-old girl with clinical suspicion of Silver-Russell syndrome (SRS). Methods: Following the international SRS consensus, methylation alterations and copy number variations (CNVs) at 11p15 region and maternal uniparental disomy of chromosome 7 were analysed and discarded by MS-MLPA. Results: Unexpectedly, the 11p15 region MS-MLPA showed a decrease in the signal of a copy number reference probe. Deletions affecting a single probe are inconclusive. So, we faced the ethical dilemma of whether it was appropriate to confirm this alteration with independent techniques and to offer a diagnostic possibility that was in no way related to clinical suspicion. Fortunately, in this particular case, the informed consent had not been specific to a particular pathology but to any disorder associated with growth failure. Performed alternative studies allowed the final diagnosis of 22q deletion syndrome. Conclusion: We demonstrate the importance of informing patients about the possibility of obtaining incidental findings in genetic techniques (not only in next generation sequencing) during pre-test genetic counselling consultations. In addition, we highlight the relevance of including in the informed consent the option of knowing these unexpected incidental findings as in some cases, this will help to elucidate the definitive diagnosis and provide the correct follow-up and treatment.

C9ORF72 Gene GGGGCC Hexanucleotide Expansion: A High Clinical Variability from Amyotrophic Lateral Sclerosis to Frontotemporal Dementia.

The expanded GGGGCC hexanucleotide repeat (HRE) in the non-coding region of the C9ORF72 gene (C9ORF72-HRE) is the most common genetic cause of familial forms of amyotrophic lateral sclerosis (ALS), FTD, and concurrent ALS and FTD (ALS-FTD), in addition to contributing to the sporadic forms of these diseases. Both syndromes overlap not only genetically, but also sharing similar clinical and neuropathological findings, being considered as a spectrum. In this paper we describe the clinical-genetic findings in a Basque family with different manifestations within the spectrum, our difficulties in reaching the diagnosis, and a narrative review, carried out as a consequence, of the main features associated with C9ORF72-HRE. Family members underwent a detailed clinical assessment, neurological examination, and genetic analysis by repeat-primed PCR. We studied 10 relatives of a symptomatic carrier of the C9ORF72-HRE expansion. Two of them presented the expansion in the pathological range, one of them was symptomatic whereas the other one remained asymptomatic at 72 years. Given the great intrafamilial clinical variability of C9ORF72-HRE, the characterization of patients and family members with particular clinical and genetic subgroups within ALS and FTD becomes a bottleneck for medication development, in particular for genetically focused medicines for ALS and FTD.

[Vitamin D deficiency in adulthood: Presentation of 2familial cases simulating pseudohypoparathyroidism]. / Deficiencia de vitamina D en la edad adulta: presentación de 2 casos familiares de seudohipoparatiroidismo.

BACKGROUND AND OBJECTIVE: The clinical and biochemical overlap of various pathologies of phosphocalcic metabolism can lead to misdiagnosis and consequent clinical management. One example is pseudohypoparathyroidism, which can be confused with vitamin D-dependent rickets (VDDR1) if appropriate biochemical determinations are not performed. PATIENTS AND METHODS: Two pairs of siblings, from independent families, were clinically diagnosed in adolescence with pseudohypoparathyroidism due to hypocalcaemia, elevated parathyroid hormone levels and normal or elevated phosphorus values. After ruling out alterations in GNAS, a massive sequencing study of genes associated with other differential diagnoses was carried out. RESULTS: Two genetic variants in the CYP27B1 gene potentially associated with the phenotype were identified. Pathogenic variants in this gene are associated with VDDR1A. Clinical-biochemical re-evaluation of the patients confirmed this diagnosis and treatment was adapted. CONCLUSIONS: Although VDDR1A is an infrequently diagnosed pathology in adulthood, in cases of hypocalcaemia with elevated PTH values, determination of the 1,25(OH)2D3 and 25(OH)D3 forms of vitamin D is relevant to reach a correct diagnosis.

Analysis of a large case series of fatal familial insomnia to determine tests with the highest diagnostic value.

Fatal familial insomnia (FFI) is a rare prionopathy with unusually high incidence in the Basque Country. We report detailed data on clinical, diagnostic, histopathological, and biochemical characteristics of a recent FFI case series. The Basque Brain Bank database was screened for patients diagnosed from 2010 to 2021 with standard genetic and/or neuropathological criteria. This series includes 16 patients, 25% without family history, with 12 cases from 9 unrelated (but geographically-linked, Basque country) kindreds, onset ranging from 36 to 70 years, and disease course from 7 to 11.5 months. Insomnia was the initial symptom in most cases, with consistent polysomnography in 92% of the cases. In contrast, 14-3-3 and RT-QuIC from cerebrospinal fluid were negative. Most patients were homozygous for methionine. Gliosis and neuronal loss in basal ganglia and thalamus were the main histopathological findings; Western blotting identified preferentially the protease-resistant prion protein (PrPres) type 2, although detection of the scrapie isoform of the prion protein (PrPSc) identified using brain tissue RT-QuIC was more successful. This is one of the largest current studies on FFI patients performed to provide improvements in diagnostic reliability. Among the analyzed tests, polysomnography and the genetic study show the highest diagnostic value in FFI.

Frequency of de novo variants and parental mosaicism in families with inactivating PTH/PTHrP signaling disorder type 2.

Objective: iPPSD2 (which includes PHP1A and PPHP/POH) is a rare inherited autosomal dominant endocrine disorder caused by inactivating GNAS pathogenic variants. A high percentage of de novo cases has been suggested. In rare cases, parental mosaicism has been described, but its real frequency is unknown. Design: A retrospective study including a series of 95 genetically confirmed iPPSD2 probands. Methods: The frequency of de novo cases was evaluated and the distribution of the type of variants was compared according to the type of inheritance. The putative involved allele was determined by reverse transcriptase PCR (RT-PCR) or allele specific oligonucleotide RT-PCR (ASO-RT-PCR). The possibility of GNAS mosaicism was studied by next-generation sequencing (NGS) on the corresponding parental DNA. Results: In 41 patients the variant was of de novo origin and in 24 the origin could not be established. In both cases 66.67% of variants generated a truncated or absent protein whereas the rest of the variants were missense or in-frame deletion/duplication. Parental origin was studied in 45 of those patients and determined in 35. Curiously, the percentage of de novo variants at the paternal allele was higher than when paternally inherited (31.1% vs 6.67%). NGS detected mosaicism in three independent families: one from paternal DNA (allelic ratio 10%) and two from maternal DNA (allelic ratio 10% and 2%). Conclusion: De novo pathogenic variants are frequent in iPPSD2 (around 45%). Parental mosaicism is infrequent (8.11%) but should be analyzed with NGS, taking into account its importance in genetic counselling.