Genetic evaluation of patients with Alström syndrome in the Polish population

Alström syndrome (AS) is a rare syndromic form of obesity and type 2 diabetes (T2D) in children coexisting with retinal dystrophy and disorders of many organs caused by the mutations in ALMS1 gene. Aim of this study was to identify the causative mutations in ALMS1 in a group of 12 patients of Polish origin with clinical symptoms of AS, and their 21 first‐degree relatives. Using DNA sequencing, nine different mutations including three novel were identified. These mutations were not present in 212 Polish individuals with no symptoms of AS, subjected to whole‐exome sequencing and collected in a national registry. Looking for genotype–phenotype relationships, we confirmed a severe phenotype in a boy with homozygous mutation in exon 16, and a relationship between a presence of T2D and mutations in exon 19. Evaluation of the type of mutation and its clinical effects gives hope for earlier diagnosis of AS in future patients and more advanced therapeutic approaches for patients with already diagnosed AS.     

Homozygous loss-of-function mutation in ALMS1 causes the lethal disorder mitogenic cardiomyopathy in two siblings

BackgroundTwo siblings from consanguineous parents of Turkish descent presented with isolated dilated cardiomyopathy, leading to early death in infancy. The diagnosis of mitogenic cardiomyopathy was made histologically.Methods and resultsLinkage analysis combined with exome sequencing identified a homozygous deleterious mutation in the ALMS1 gene as the cause of this phenotype.ConclusionsAlström syndrome is characterized by a typically transient dilating cardiomyopathy in infancy, suggesting that mitogenic cardiomyopathy represents the extreme phenotype, resulting in demise before the other clinical symptoms become evident. This observation further illustrates the role of ALMS1 and cell cycle regulation.     

A Next Generation Sequencing custom gene panel as first line diagnostic tool for atypical cases of syndromic obesity: Application in a case of Alström syndrome

Obesity phenotype can be manifested as an isolated trait or accompanied by multisystem disorders as part of a syndromic picture. In both situations, same molecular pathways may be involved to different degrees. This evidence is stronger in syndromic obesity, in which phenotypes of different syndromes may overlap. In these cases, genetic testing can unequivocally provide a final diagnosis. Here we describe a patient who met the diagnostic criteria for Alström syndrome only during adolescence. Genetic testing was requested at 25 years of age for a final confirmation of the diagnosis. The genetic diagnosis of Alström syndrome was obtained through a Next Generation Sequencing genetic test approach using a custom-designed gene panel of 47 genes associated with syndromic and non-syndromic obesity. Genetic analysis revealed a novel homozygous frameshift variant p.(Arg1550Lysfs*10) on exon 8 of the ALMS1 gene.This case shows the need for a revision of the diagnostic criteria guidelines, as a consequence of the recent advent of massive parallel sequencing technology. Indications for genetic testing reported in these currently accepted diagnostic criteria for Alström syndrome, were drafted when sequencing was expensive and time consuming. Nowadays, Next Generation Sequencing testing could be considered as first line diagnostic tool not only for Alström syndrome but, more generally, for all those atypical or not clearly distinguishable cases of syndromic obesity, thus avoiding delayed diagnosis and treatments. Early diagnosis permits a better follow-up and pre-symptomatic interventions.     

ALMS1 null mutations: a common cause of Leber congenital amaurosis and early‐onset severe cone–rod dystrophy

In our previous studies, mutations in known candidate genes were detected in approximately 50% of Chinese patients with various forms of retinal degeneration. The next stage, identifying additional causative mutations in patients with various forms of genetic eye diseases based on whole exome sequencing of 1220 samples, revealed frequent homozygous or compound heterozygous null mutations in ALMS1, which are known to associate with Alström syndrome as well as individuals diagnosed with Leber congenital amaurosis (LCA) or early‐onset severe cone–rod dystrophy (CORD) without signs of systemic phenotypes except that one had a congenital heart abnormity. Sanger sequencing, co‐segregation analysis and analysis of normal individuals identified a total of 13 null mutations in ALMS1 in 11 probands, including 4 probands with homozygous mutations and 7 with compound heterozygous mutations. Follow‐up examinations revealed absent or mild systemic manifestations of Alström syndrome in those available: 9 of 15 patients in 11 families. These findings not only expand the spectrum of phenotypes associated with ALMS1 mutations but also suggest that ALMS1 should be regarded as a candidate causative gene in patients diagnosed with isolated LCA and early‐onset severe CORD.     

Molecular analysis and long-term clinical evaluation of three siblings with Alström syndrome

Alström syndrome is a rare, autosomal recessive disorder characterized by a wide spectrum of clinical features including early‐onset retinal degeneration leading to blindness, sensorineural hearing loss, short stature, obesity, type 2 diabetes, hyperlipidemia and dilated cardiomyopathy. Renal, hepatic and pulmonary dysfunction may occur in the later phases of the disease. The three affected sisters, from a consanguineous Turkish family, with the characteristic features of Alström syndrome, were clinically diagnosed in 1987 and followed for 20 years. DNA sequence analysis of ALMS1, the causative gene in Alström syndrome, identified a novel homozygous disease‐causing mutation, c.8164C>T, resulting in a premature termination codon in exon 10 in each of the three affected sisters. Furthermore, we describe the longitudinal disease progression in this family and report new clinical findings likely associated with Alström syndrome, such as pes planus and hyperthyroidism.     

Phenotypic and mutational spectrum of 21 Chinese patients with Alström syndrome

Alström syndrome (AS) is a monogenic syndromic ciliopathy caused by mutations in the ALMS1 (Alström Syndrome 1) gene. A total of 21 subjects with AS from 20 unrelated Chinese families were recruited. Our cohort consists of 9 females and 12 males, between 5 months and 20 years old. The first symptom(s) appeared between 3 and 24 months. They were recorded to be either visual impairments (83%) or dilated cardiomyopathy (17%). Median time from symptom onset to seeking medical attention was 6 months (3–36 months) and the median time needed to reach the final molecular diagnosis is 54 months (6–240 months). System involvement at the time of the survey was as follows: visual symptoms (100%), hearing Impairment (67%), endocrine symptoms (43%), neurological symptoms (19%), hepatic symptoms (14%), and renal Involvement (14%). These findings are comparable to data reported in the literature. However, the proportion of subjects with cognitive impairment (33%) and behavioral problems (19%) were higher. Thirty‐three unique mutations were identified in the ALMS1 gene, of which 18 are novel mutations classified as pathogenic/likely pathogenic according to the American College of Medical Genetics (ACMG) guideline. Four recurrent mutations were identified in the cohort, in particular; c.2084C>A, p. (Ser695Ter), is suggestive to be a founder mutation in people of Chinese ancestry. The participation of AS subjects of differing ethnicities is essential to improve the algorithm in facial recognition/phenotyping, as well as to understand the mutation spectrum beyond than just those of European ancestry.     

Alström Syndrome: Mutation Spectrum of ALMS1

Alström Syndrome (ALMS), a recessive, monogenic ciliopathy caused by mutations in ALMS1, is typically characterized by multisystem involvement including early cone‐rod retinal dystrophy and blindness, hearing loss, childhood obesity, type 2 diabetes mellitus, cardiomyopathy, fibrosis, and multiple organ failure. The precise function of ALMS1 remains elusive, but roles in endosomal and ciliary transport and cell cycle regulation have been shown. The aim of our study was to further define the spectrum of ALMS1 mutations in patients with clinical features of ALMS. Mutational analysis in a world‐wide cohort of 204 families identified 109 novel mutations, extending the number of known ALMS1 mutations to 239 and highlighting the allelic heterogeneity of this disorder. This study represents the most comprehensive mutation analysis in patients with ALMS, identifying the largest number of novel mutations in a single study worldwide. Here, we also provide an overview of all ALMS1 mutations identified to date.     

Biallelic intragenic deletion in MASP1 in an adult female with 3MC syndrome

3MC syndrome is a rare autosomal recessive disorder with characteristic craniofacial dysmorphism and multiple anomalies. It is caused by biallelic mutations in one of three genes, MASP1, COLEC11 and COLEC10, all encoding factors of the lectin complement pathway. In MASP1, either truncating mutations or missense variants in exon 12 encoding the C-terminal serine protease domain specific for isoform MASP-3 are causative.By trio exome sequencing we now identified a novel, homozygous 2kb deletion, partially affecting exon 12 in an adult female with the typical facial gestalt of 3MC syndrome and hearing loss, but without the main feature cleft lip/palate, and without intellectual disability, or short stature. We therefore expand the MASP1 associated mutational and clinical spectrum and describe the development of her clinical presentation over a period of 21 years. As the homozygous deletion in our patient was only found by thorough and visual evaluation of the whole exome sequencing data, such deletions might escape detection in some routine diagnostic workflows and might explain a few of the so far molecularly unconfirmed cases of 3MC syndrome.     

Sedaghatian-type spondylometaphyseal dysplasia: Whole exome sequencing in neonatal dry blood spots enabled identification of a novel variant in GPX4

Accumulation of lipid peroxides causes membrane damage and cell death. Glutathione peroxidase 4 (GPX4) acts as a hydroperoxidase which prevents accumulation of toxic oxidized lipids and blocks ferroptosis, an iron-dependent, non-apoptotic mode of cell death. GPX4 deficiency causes Sedaghatian-type spondylo-metaphyseal dysplasia (SSMD), a lethal autosomal recessive disorder, featuring skeletal dysplasia, cardiac arrhythmia and brain anomalies with only three pathogenic GPX4 variants reported in two SSMD patients. Our objective was to identify the underlying genetic cause of neonatal death of two siblings presenting with hypotonia, cardiorespiratory failure and SSMD. Whole exome sequencing (WES) was performed in DNA samples from two siblings and their parents. Since “critical samples” were not available from the patients, DNA was extracted from dry blood spots (DBS) retrieved from the Israeli newborn-screening center. Sanger sequencing and segregation analysis followed the WES. Homozygous novel GPX4 variant, c.153_160del; p.His52fs*1 causing premature truncation of GPX4 was detected in both siblings; their parents were heterozygotes. Segregation analysis confirmed autosomal recessive inheritance. This report underscores the importance of DBS WES in identifying the genes and mutations causing devastating rare diseases. Obtaining critical samples from a dying patient is crucial for enabling genetic diagnosis.     

Homozygous c.359del variant in MGME1 is associated with early onset cerebellar ataxia

We ascertained a child with early onset cerebellar ataxia and identified a novel frameshift deletion, c.359del [p. (Pro120Leufs*2), NM_052865.2] in exon 2 of MGME1 (mitochondrial genome maintenance exonuclease 1) by exome sequencing. Variations in MGME1 have been reported to cause mitochondrial DNA (mtDNA) depletion syndrome 11 (MIM #615084) in an earlier work. The phenotype included progressive external ophthalmoplegia, emaciation, respiratory failure and late onset progressive ataxia. However, the child presented here has early onset progressive ataxia, speech delay, microcephaly, cerebellar atrophy and fundus albipunctatus. This is the second report of a mutation in MGME1 and describes a more severe phenotype.     

Identification of Two Homozygous Sequence Variants in the COL7A1 Gene Underlying Dystrophic Epidermolysis Bullosa by Whole‐Exome Analysis in a Consanguineous Family

Dystrophic epidermolysis bullosa (DEB) is an inherited skin disorder with variable severity and heterogeneous genetic involvement. Diagnostic approaches for this condition include clinical evaluations and electron microscopy of patients’ skin biopsies, followed by Sanger sequencing (SS) of a large gene (118 exons) that encodes the alpha chain of type VII collagen (COL7A1) located on Chromosome 3p21.1. However, the use of SS may hinder diagnostic efficiency and lead to delays because it is costly and time‐consuming. We evaluated a 5‐generation consanguineous family with 3 affected individuals presenting the severe generalised DEB phenotype. Human whole‐exome sequencing (WES) revealed 2 homozygous sequence variants: the previously reported variant p.Arg578* in exon 13 and a novel variant p.Arg2063Gln in exon 74 of the COL7A1 gene. Validation by SS, performed on all family members, confirmed the cosegregation of the 2 variants with the disease phenotype. To the best of our knowledge, 2 homozygous COL7A1 variants have never been simultaneously reported in DEB patients; however, the upstream protein truncation variant is more likely to be disease‐causing than the novel missense variant. WES can be used as an efficient molecular diagnostic tool for evaluating autosomal recessive forms of DEB.     

Novel truncating and missense variants extending the spectrum of EMC1-related phenotypes,causing autism spectrum disorder,severe global development delay and visual impairment

The EMC1 gene, located on 1p36.13, encodes the subunit 1 of the endoplasmic reticulum-membrane protein complex, a highly conserved and ubiquitous multiprotein transmembrane complex. Pathogenic monoallelic and biallelic variants in EMC1 in humans have been reported only in six families, causing isolated visual impairment or in association with psychomotor retardation and cerebellar atrophy. We report a ten-year-old boy, born to unrelated parents, with early-onset severe global development delay due to novel EMC1 biallelic pathogenic variants. A truncating variant, p.(Tyr378*) and a missense variant, p.(Phe953Ser), located in exon 11 and 23 of EMC1 gene respectively, have been found by reanalysis of exome sequencing data. The proband's phenotype included several signs that overlap with the phenotype of previously reported patients, associating severe global developmental delay, abnormal ophthalmological examination, and postnatal slow-down of the head circumference growth. Some distinguishing clinical signs were observed in comparison to patients from literature, such as autism spectrum disorder, absence of seizures, scoliosis or facial dysmorphic features, thus extending the spectrum of EMC1-related phenotypes. Similarly, brain MRI, performed at 2 years, showed normal cerebellar volume and structure, whereas cerebellar atrophy was described in literature. Moreover, difficulties of clinical differential diagnosis between EMC1-associated disease and other etiologies of global development delay support the importance of large-scale genetic investigations. Our diagnostic approach, through reanalysis of exome sequencing data, highlights the importance of reconsidering initial negative results for patients with a strong suspicion of genetic disease, and to update analytic pipelines in order to improve the diagnostic yield of exome sequencing.     

Intrafamilial variability of XYLT2-related spondyloocular syndrome

Spondyloocular syndrome is characterized by generalized osteoporosis, multiple fractures and severe ocular findings. The causative XYLT2 mutations have recently been identified with the use of whole exome sequencing. We report on two siblings with spondyloocular syndrome who presented with varying clinical severity. A novel XYLT2 missense mutation was detected in a region evolutionary conserved across the species. This report along with the previous reports demonstrates that variable expressivity may be possible even within the same family. These two siblings with a novel mutation further expand the clinical and mutational spectrum of spondyloocular syndrome.     

Clinical exome sequencing for cerebellar ataxia and spastic paraplegia uncovers novel gene–disease associations and unanticipated rare disorders

Cerebellar ataxia (CA) and hereditary spastic paraplegia (HSP) are two of the most prevalent motor disorders with extensive locus and allelic heterogeneity. We implemented clinical exome sequencing, followed by filtering data for a ‘movement disorders'' gene panel, as a generic test to increase variant detection in 76 patients with these disorders. Segregation analysis or phenotypic re-evaluation was utilized to substantiate findings. Disease-causing variants were identified in 9 of 28 CA patients, and 8 of 48 HSP patients. In addition, possibly disease-causing variants were identified in 1 and 8 of the remaining CA and HSP patients, respectively. In 10 patients with CA, the total disease-causing or possibly disease-causing variants were detected in 8 different genes, whereas 16 HSP patients had such variants in 12 different genes. In the majority of cases, the identified variants were compatible with the patient phenotype. Interestingly, in some patients variants were identified in genes hitherto related to other movement disorders, such as TH variants in two siblings with HSP. In addition, rare disorders were uncovered, for example, a second case of HSP caused by a VCP variant. For some patients, exome sequencing results had implications for treatment, exemplified by the favorable L-DOPA treatment in a patient with HSP due to ATP13A2 variants (Parkinson type 9). Thus, clinical exome sequencing in this cohort of CA and HSP patients suggests broadening of disease spectra, revealed novel gene–disease associations, and uncovered unanticipated rare disorders. In addition, clinical exome sequencing results have shown their value in guiding practical patient management.     

NKX2.5 mutation identification on exome sequencing in a patient with heterotaxy

Exome sequencing enables us to screen most of the protein coding genes in an unbiased way, this technique represents an ideal tool to identify previously under- or unappreciated phenotypes associated with known disease genes and genetic disorders. Here we present an illustrative case that required exome sequencing to identify a genetic alteration associated with the clinical features. The phenotype of the proband included heterotaxy, double outlet right ventricle, common atrioventricular canal, total anomalous pulmonary venous connection, asplenia, failure to thrive and short stature. Exome sequencing demonstrated a frameshift mutation c.397_400del (p.P133GfsTer 42) in NKX2.5. Although a single previous case of heterotaxy was reported in a large familial case of NKX2.5, heterotaxy is not clinically appreciated to be a part of the phenotypic spectrum associated with NKX2.5 mutations. This case report demonstrates the utility of exome sequencing in expanding a phenotypic spectrum of a known Mendelian disorder. We predict that this type of unexpected identification of mutations in known-disease associated genes in patients with atypical or expanded phenotypes will occur with increasing frequency as the use of exome and genome sequencing become more common tools in diagnosing patients with syndromic and non-syndromic foms of structural birth defects.     

Ligase IV syndrome can present with microcephaly and radial ray anomalies similar to Fanconi anaemia plus fatal kidney malformations

Ligase IV (LIG4) syndrome is a rare disorder of DNA damage repair caused by biallelic, pathogenic variants in LIG4. This is a phenotypically heterogeneous condition with clinical presentation varying from lymphoreticular malignancies in developmentally normal individuals to significant microcephaly, primordial dwarfism, radiation hypersensitivity, severe combined immunodeficiency and early mortality. Renal defects have only rarely been described as part of the ligase IV disease spectrum.We identified a consanguineous family where three siblings presenting with antenatal growth retardation, microcephaly, severe renal anomalies and skeletal abnormalities, including radial ray defects. Autozygosity mapping and exome sequencing identified a novel homozygous frameshift variant in LIG4, c.597_600delTCAG, p.(Gln200LysfsTer33), which segregated in the family. LIG4 is encoded by a single exon and so this frameshift variant is predicted to result in a protein truncated by 678 amino acids. This is the shortest predicted LIG4 protein product reported and correlates with the most severe clinical phenotype described to date. We note the clinical overlap with Fanconi anemia and suggest that LIG4 syndrome is considered in the differential diagnosis of this severe developmental disorder.     

BCAP31-related syndrome: The first de novo report

Pathogenic variants in the BCAP31 gene have recently been associated with a severe congenital neurological phenotype, named DDCH after its key features: deafness, dystonia and central hypomyelination. BCAP31 is located at the Xq28 chromosomal region and only male individuals are currently known to be affected, the pathogenic variant being usually transmitted by healthy mothers.Here, we describe a three-year-old male child referred for severe developmental delay, failure to thrive, hearing loss and dyskinetic movements. After a conventional diagnostic workflow, including a normal array-CGH, a tentative diagnosis of dyskinetic cerebral palsy was retained. Clinical exome sequencing in the trio identified a small intragenic deletion in exon 8 of BCAP31, c.709_721del (p.Val237Trpfs*69), originated de novo and not previously reported. Based on the ACMG variant classification, this variant is predicted to be ‘likely pathogenic’. Given the consistent phenotypical overlap with the subjects already ascertained with DDCH, we considered this variant to be clinically relevant for this child and causative of his condition.