Hisham Alkhalidi

Genomic analysis of mitochondrial diseases in a consanguineous population reveals novel candidate disease genes

Objective To investigate the utility of autozygome analysis and exome sequencing in a cohort of patients with suspected or confirmed mitochondrial encephalomyopathy. Methods Autozygome was used to highlight candidate genes for direct sequencing in 10 probands, all born to consanguineous parents. Autozygome was also used to filter the variants from exome sequencing of four probands. Results In addition to revealing mutations in known mitochondrial genes, the analysis revealed the identification of two novel candidate disease genes: MFF and FARS2, encoding the mitochondrial fission factor and phenylalanyl-tRNA synthetase, respectively. Interpretation These findings expand the repertoire of genes that are mutated in patients with mitochondrial disorders and highlight the value of integrating genomic approaches in the evaluation of these patients.

Metformin Rescues the Myocardium from Doxorubicin-Induced Energy Starvation and Mitochondrial Damage in Rats

Clinical use of doxorubicin (DOX) is limited by its cardiotoxic side effects. Recent studies established that metformin (MET), an oral antidiabetic drug, possesses an antioxidant activity. However, whether it can protect against DOX-induced energy starvation and mitochondrial damage has not been reported. Our results, in a rat model of DOX-induced cardiotoxicity, show that DOX treatment significantly increased serum levels of LDH and CK-MB, indicators of cardiac injury, and induced expression of hypertrophic gene markers. DOX also caused marked decreases in the cardiac levels of glutathione, CoA-SH and ATP, and mRNA expression of catalase and NQO-1. These biochemical changes were associated with myocardial histopathological and ultrastructural deteriorations, as observed by light and electron microscopy, respectively. Cotreatment with MET (500 mg/kg) eliminated all DOX-induced biochemical, histopathological, and ultrastructural changes. These findings demonstrate that MET successfully prevents DOX-induced cardiotoxicity in vivo by inhibiting DOX-induced oxidative stress, energy starvation, and depletion of intramitochondrial CoA-SH.

New Findings in a Global Approach to Dissect the Whole Phenotype of PLA2G6 Gene Mutations

Mutations in PLA2G6 gene have variable phenotypic outcome including infantile neuroaxonal dystrophy, atypical neuroaxonal dystrophy, idiopathic neurodegeneration with brain iron accumulation and Karak syndrome. The cause of this phenotypic variation is so far unknown which impairs both genetic diagnosis and appropriate family counseling. We report detailed clinical, electrophysiological, neuroimaging, histologic, biochemical and genetic characterization of 11 patients, from 6 consanguineous families, who were followed for a period of up to 17 years. Cerebellar atrophy was constant and the earliest feature of the disease preceding brain iron accumulation, leading to the provisional diagnosis of a recessive progressive ataxia in these patients. Ultrastructural characterization of patients’ muscle biopsies revealed focal accumulation of granular and membranous material possibly resulting from defective membrane homeostasis caused by disrupted PLA2G6 function. Enzyme studies in one of these muscle biopsies provided evidence for a relatively low mitochondrial content, which is compatible with the structural mitochondrial alterations seen by electron microscopy. Genetic characterization of 11 patients led to the identification of six underlying PLA2G6 gene mutations, five of which are novel. Importantly, by combining clinical and genetic data we have observed that while the phenotype of neurodegeneration associated with PLA2G6 mutations is variable in this cohort of patients belonging to the same ethnic background, it is partially influenced by the genotype, considering the age at onset and the functional disability criteria. Molecular testing for PLA2G6 mutations is, therefore, indicated in childhood-onset ataxia syndromes, if neuroimaging shows cerebellar atrophy with or without evidence of iron accumulation.

A novel syndrome of hypohidrosis and intellectual disability is linked to COG6 deficiency.

Background Numerous syndromic forms of intellectual disability have been described including those with abnormal sweating pattern. Purpose To describe the clinical and molecular analysis of a large multiplex consanguineous Saudi family with an unusual constellation of severe intellectual disability, hypohidrosis, abnormal teeth, and acquired microcephaly. Methods Clinical evaluation, autozygosity mapping, exome sequencing, and expression analysis. Results Autozygosity mapping revealed a single critical locus corresponding to chr13:39 338 062–40 857 430. Exome sequencing uncovered a deep intronic (NM_020751.2:c.1167–24A>G) variant in COG6 that largely replaces the consensus acceptor site, resulting in pronounced reduction of the normal transcript and consequent deficiency of COG6 protein. Patient cells also exhibited pronounced deficiency of STX6, consistent with the established stabilising effect of COG6 on STX6. Four additional patients representing two families of the same tribal origin as the original family were found to have the same mutation, confirming a founder effect. Remarkably, none of the patients displayed any detectable abnormality in the glycosylation pattern of transferrin, which contradicts a previously published report of a patient whose abnormal glycosylation pattern was presumed to be caused by a missense variant in COG6. Conclusions Our data implicate COG6 in the pathogenesis of a novel hypohidrotic disorder in humans that is distinct from congenital disorders of glycosylation.

A novel syndrome of Klippel-Feil anomaly, myopathy, and characteristic facies is linked to a null mutation in MYO18B

Background Klippel-Feil anomaly (KFA) can be seen in a number of syndromes. We describe an apparently novel syndromic association with KFA. Methods Clinical phenotyping of two consanguineous families followed by combined autozygome/exome analysis. Results Two patients from two apparently unrelated families shared a strikingly similar phenotype characterised by KFA, myopathy, mild short stature, microcephaly, and distinctive facies. They shared a single founder autozygous interval in which whole exome sequencing revealed a truncating mutation in MYO18B. There was virtually complete loss of the transcript in peripheral blood, indicative of nonsense-mediated decay. Electron microscopy of muscle confirms abnormal myosin filaments with accompanying myopathic changes. Conclusions Deficiency of MYO18B is linked to a novel developmental disorder which combines KFA with myopathy. This suggests a widespread developmental role for this gene in humans, as observed for its murine ortholog.

Efficient identification of novel mutations in patients with limb girdle muscular dystrophy

Limb girdle muscular dystrophy type 2 (LGMD2) is a genetically heterogeneous autosomal recessive disorder caused by mutations in 15 known genes. DNA sequencing of all candidate genes can be expensive and laborious, whereas a selective sequencing approach often fails to provide a molecular diagnosis. We aimed to efficiently identify pathogenic mutations via homozygosity mapping in a population in which the genetics of LGMD2 has not been well characterized. Thirteen consanguineous families containing a proband with LGMD2 were recruited from Saudi Arabia, and for 11 of these families, selected individuals were genotyped at 10,204 single nucleotide polymorphisms. Linkage analysis excluded all but one or two known genes in ten of 11 genotyped families, and haplotype comparisons between families allowed further reduction in the number of candidate genes that were screened. Mutations were identified by DNA sequencing in all 13 families, including five novel mutations in four genes, by sequencing at most two genes per family. One family was reclassified as having a different myopathy based on genetic and clinical data after linkage analysis excluded all known LGMD2 genes. LGMD2 subtypes A and B were notably absent from our sample of patients, indicating that the distribution of LGMD2 mutations in Saudi Arabian families may be different than in other populations. Our data demonstrate that homozygosity mapping in consanguineous pedigrees offers a more efficient means of discovering mutations that cause heterogeneous disorders than comprehensive sequencing of known candidate genes.

Autozygosity reveals recessive mutations and novel mechanisms in dominant genes: implications in variant interpretation

Purpose:The purpose of this study is to describe recessive alleles in strictly dominant genes. Identifying recessive mutations in genes for which only dominant disease or risk alleles have been reported can expand our understanding of the medical relevance of these genes both phenotypically and mechanistically. The Saudi population is enriched for autozygosity, which enhances the homozygous occurrence of alleles, including pathogenic alleles in genes that have been associated only with a dominant inheritance pattern.Methods:Exome sequencing of patients from consanguineous families with likely recessive phenotypes was performed. In one family, the genotype of the deceased children was inferred from their parents due to lack of available samples.Results:We describe the identification of 11 recessive variants (5 of which are reported here for the first time) in 11 genes for which only dominant disease or risk alleles have been reported. The observed phenotypes for these recessive variants were novel (e.g., FBN2-related myopathy and CSF1R-related brain malformation and osteopetrosis), typical (e.g., ACTG2-related visceral myopathy), or an apparently healthy state (e.g., PDE11A), consistent with the corresponding mouse knockout phenotypes.Conclusion:Our results show that, in the era of genomic sequencing and “reverse phenotyping,” recessive variants in dominant genes should not be dismissed based on perceived “incompatibility” with the patient’s phenotype before careful consideration.Genet Med advance online publication 06 April 2017

Mutation of the mitochondrial carrier SLC25A42 causes a novel form of mitochondrial myopathy in humans

Myopathies are heterogeneous disorders characterized clinically by weakness and hypotonia, usually in the absence of gross dystrophic changes. Mitochondrial dysfunction is a frequent cause of myopathy. We report a simplex case born to consanguineous parents who presented with muscle weakness, lactic acidosis, and muscle changes suggestive of mitochondrial dysfunction. Combined autozygome and exome analysis revealed a missense variant in the SLC25A42 gene, which encodes an inner mitochondrial membrane protein that imports coenzyme A into the mitochondrial matrix. Zebrafish slc25a42 knockdown morphants display severe muscle disorganization and weakness. Importantly, these features are rescued by normal human SLC25A42 RNA, but not by RNA harboring the patient’s variant. Our data support a potentially causal link between SLC25A42 mutation and mitochondrial myopathy in humans.

Choroid plexus papilloma metastases to both cerebellopontine angles mimicking neurofibromatosis type 2

Choroid plexus papilloma (CPP) is a benign tumor of neuroectodermal origin that occurs most commonly in the posterior fossa of adults [5]. Complete resection of the tumor typically results in a cure, but intracranial and spinal drop metastases occur [5, 8]. We present a patient with intracranial metastases from CPP involving both cerebellopontine angles (CPAs) and mimicking neurofibromatosis type 2 (NF2) [4]. When a 35 year old patient complained of headaches and episodes of blurred vision, an MRI scan was performed and revealed a well-circumscribed, contrast-enhancing, midline lesion in the fourth ventricle with no hydrocephalus. Suboccipital craniectomy resulted in macroscopically total excision of a well-circumscribed CPP. Sixteen years later, he developed transient visual obscurations in both eyes. Visual acuity was modestly reduced, left more than right, with a 2? afferent pupillary defect on the left and moderate atrophic papilledema bilaterally. Hearing was decreased on the left; he had no cafe au lait spots or other skin lesions. MRI revealed multiple enhancing extra-axial masses involving both CPAs, the foramina of Luschka and Magendie, the subfrontal region, and the middle and posterior cranial fossae (Fig. 1a, b, c). Masses had intermediate signal intensity on T1and T2-weighted images and multiple foci of punctuate calcification on CT. CPA masses extended into the internal auditory meatus, and the subfrontal mass measured 4 9 4.4 9 3 cm. Recurrence of CPP was considered, but NF2 was suspected both clinically and radiologically because of masses in both CPAs. Gross total resection of the large, symptomatic subfrontal mass was accomplished with no sequelae, and morphological and immunohistochemical features of the tumor were consistent with CPP (WHO grade I; Fig. 1d). The full coding region, exon–intron boundaries, and promoter region of the NF2 (neurofibromin 2) gene (NG_009057) were sequenced and no mutations were found. Benign CPP typically forms a well-defined, homogeneously enhancing mass on CT or MRI, occasionally in one or both CPAs at presentation [9]. Complete resection usually results in a cure, although recurrence and metastasis have been reported [5, 8]. Masses are usually isointense on T1-weighted images, showing a slightly high T2 signal intensity, and calcification is present in 4–20% of tumors. Various mechanisms have been hypothesized for dissemination, including iatrogenic seeding and local extension [4]. NF2 is a dominantly inherited, tumor-prone disorder characterized by the development of multiple schwannomas and meningiomas [1, 4]. The presence of bilateral vestibular schwannomas is considered pathognomonic [1]; however, bilateral CPA masses are not synonymous with bilateral vestibular schwannomas. Two previous reports describe patients with CPP metastatic to both CPAs [2, 3], leading to a challenging A. A. Al-Abdullah K. K. Abu-Amero (&) A. Hellani T. M. Bosley Department of Ophthalmology, College of Medicine, King Saud University, PO Box 245, Riyadh 11411, Saudi Arabia e-mail: abuamero@gmail.com

Cystic panfolliculoma of the scalp: report of a very rare case and brief review.

Panfolliculoma is an exceeding rare follicular benign neoplasm with differentiation toward both upper and lower segments of the hair follicle. In this report, we present a case of cystic panfolliculoma in the occipital region of the scalp of a 19-year-old female. We describe the histopathological and immunohistochemical features of the tumor and briefly discuss the differential diagnoses of this rare entity.