Monogenic gene variants in lung transplant recipients with usual interstitial pneumonia.

AIM: The prevalence of monogenic disease-causing gene variants in lung transplant recipients with idiopathic pulmonary fibrosis is not fully known. Their impact on clinical outcomes before and after transplantation requires more evidence. PATIENTS AND METHODS: We retrospectively performed sequence analysis of genes associated with pulmonary fibrosis in a cohort of 23 patients with histologically confirmed usual interstitial pneumonia that had previously undergone double lung transplantation. We evaluated the impact of confirmed molecular diagnoses on disease progression, clinical outcomes and incidence of acute rejection or chronic lung allograft dysfunction after transplantation. RESULTS: 15 patients out of 23 (65%) had a variant in a gene associated with interstitial lung disease. 11 patients (48%) received a molecular diagnosis, of which nine involved genes for telomerase function. Five diagnostic variants were found in the gene for Telomerase reverse transcriptase. Two of these variants, p.(Asp684Gly) and p.(Arg774*), seemed to be enriched in Finnish lung transplant recipients. Disease progression and the incidence of acute rejection and chronic lung allograft dysfunction was similar between patients with telomere-related disease and the rest of the study population. The incidence of renal or bone marrow insufficiency or skin malignancies did not differ between the groups. CONCLUSION: Genetic variants are common in lung transplant recipients with pulmonary fibrosis and are most often related to telomerase function. A molecular diagnosis for telomeropathy does not seem to impact disease progression or the risk of complications or allograft dysfunction after transplantation.

Next-generation sequencing in childhood-onset epilepsies: Diagnostic yield and impact on neuronal ceroid lipofuscinosis type 2 (CLN2) disease diagnosis.

Epilepsy is one of the most common childhood-onset neurological conditions with a genetic etiology. Genetic diagnosis provides potential for etiologically-based management and treatment. Existing research has focused on early-onset (<24 months) epilepsies; data regarding later-onset epilepsies is limited. The goal of this study was to determine the diagnostic yield of a clinically available epilepsy panel in a selected pediatric epilepsy cohort with epilepsy onset between 24-60 months of life and evaluate whether this approach decreases the age of diagnosis of neuronal ceroid lipofuscinosis type 2 (CLN2). Next-generation sequencing (NGS)-based epilepsy panels, including genes associated with epileptic encephalopathies and inborn errors of metabolism (IEMs) that present with epilepsy, were used. Copy-number variant (CNV) detection from NGS data was included. Variant interpretation was performed per American College of Medical Genetics and Genomics (ACMG) guidelines. Results are reported from 211 consecutive patients with the following inclusion criteria: 24-60 months of age at the time of enrollment, first unprovoked seizure at/after 24 months, and at least one additional finding such as EEG/MRI abnormalities, speech delay, or motor symptoms. Median age was 42 months at testing and 30 months at first seizure onset; the mean delay from first seizure to comprehensive genetic testing was 10.3 months. A genetic diagnosis was established in 43 patients (20.4%). CNVs were reported in 25.6% diagnosed patients; 27.3% of CNVs identified were intragenic. Within the diagnosed cohort, 11 (25.6%) patients were diagnosed with an IEM. The predominant molecular diagnosis was CLN2 (14% of diagnosed patients). For these patients, diagnosis was achieved 12-24 months earlier than reported by natural history of the disease. This study supports comprehensive genetic testing for patients whose first seizure occurs ≥ 24 months of age. It also supports early application of testing in this age group, as the identified diagnoses can have significant impact on patient management and outcome.

Diagnostic yield of genetic testing in a heterogeneous cohort of 1376 HCM patients.

BACKGROUND: Genetic testing in hypertrophic cardiomyopathy (HCM) is a published guideline-based recommendation. The diagnostic yield of genetic testing and corresponding HCM-associated genes have been largely documented by single center studies and carefully selected patient cohorts. Our goal was to evaluate the diagnostic yield of genetic testing in a heterogeneous cohort of patients with a clinical suspicion of HCM, referred for genetic testing from multiple centers around the world. METHODS: A retrospective review of patients with a suspected clinical diagnosis of HCM referred for genetic testing at Blueprint Genetics was undertaken. The analysis included syndromic, myopathic and metabolic etiologies. Genetic test results and variant classifications were extracted from the database. Variants classified as pathogenic (P) or likely pathogenic (LP) were considered diagnostic. RESULTS: A total of 1376 samples were analyzed. Three hundred and sixty-nine tests were diagnostic (26.8%); 373 P or LP variants were identified. Only one copy number variant was identified. The majority of diagnostic variants involved genes encoding the sarcomere (85.0%) followed by 4.3% of diagnostic variants identified in the RASopathy genes. Two percent of diagnostic variants were in genes associated with a cardiomyopathy other than HCM or an inherited arrhythmia. Clinical variables that increased the likelihood of identifying a diagnostic variant included: an earlier age at diagnosis (p < 0.0001), a higher maximum wall thickness (MWT) (p < 0.0001), a positive family history (p < 0.0001), the absence of hypertension (p = 0.0002), and the presence of an implantable cardioverter-defibrillator (ICD) (p = 0.0004). CONCLUSION: The diagnostic yield of genetic testing in this heterogeneous cohort of patients with a clinical suspicion of HCM is lower than what has been reported in well-characterized patient cohorts. We report the highest yield of diagnostic variants in the RASopathy genes identified in a laboratory cohort of HCM patients to date. The spectrum of genes implicated in this unselected cohort highlights the importance of pre-and post-test counseling when offering genetic testing to the broad HCM population.

Biallelic loss-of-function in NRAP is a cause of recessive dilated cardiomyopathy.

BACKGROUND: Familial dilated cardiomyopathy (DCM) is typically a monogenic disorder with dominant inheritance. Although over 40 genes have been linked to DCM, more than half of the patients undergoing comprehensive genetic testing are left without molecular diagnosis. Recently, biallelic protein-truncating variants (PTVs) in the nebulin-related anchoring protein gene (NRAP) were identified in a few patients with sporadic DCM. METHODS AND RESULTS: We determined the frequency of rare NRAP variants in a cohort of DCM patients and control patients to further evaluate role of this gene in cardiomyopathies. A retrospective analysis of our internal variant database consisting of 31,639 individuals who underwent genetic testing (either panel or direct exome sequencing) was performed. The DCM group included 577 patients with either a confirmed or suspected DCM diagnosis. A control cohort of 31,062 individuals, including 25,912 individuals with non-cardiac (control group) and 5,150 with non-DCM cardiac indications (Non-DCM cardiac group). Biallelic (n = 6) or two (n = 5) NRAP variants (two PTVs or PTV+missense) were identified in 11 unrelated probands with DCM (1.9%) but none of the controls. None of the 11 probands had an alternative molecular diagnosis. Family member testing supports co-segregation. Biallelic or potentially biallelic NRAP variants were enriched in DCM vs. controls (OR 1052, p<0.0001). Based on the frequency of NRAP PTVs in the gnomAD reference population, and predicting full penetrance, biallelic NRAP variants could explain 0.25%-2.46% of all DCM cases. CONCLUSION: Loss-of-function in NRAP is a cause for autosomal recessive dilated cardiomyopathy, supporting its inclusion in comprehensive genetic testing.

Genome wide association study of 40 clinical measurements in eight dog breeds.

The domestic dog represents an ideal model for identifying susceptibility genes, many of which are shared with humans. In this study, we investigated the genetic contribution to individual differences in 40 clinically important measurements by a genome-wide association study (GWAS) in a multinational cohort of 472 healthy dogs from eight breeds. Meta-analysis using the binary effects model after breed-specific GWAS, identified 13 genome-wide significant associations, three of them showed experimental-wide significant associations. We detected a signal at chromosome 13 for the serum concentration of alanine aminotransferase (ALT) in which we detected four breed-specific signals. A large proportion of the variance of ALT (18.1-47.7%) was explained by this locus. Similarly, a single SNP was also responsible for a large proportion of the variance (6.8-78.4%) for other measurements such as fructosamine, stress during physical exam, glucose, and morphometric measurements. The genetic contribution of single variant was much larger than in humans. These findings illustrate the importance of performing meta-analysis after breed-specific GWAS to reveal the genetic contribution to individual differences in clinically important measurements, which would lead to improvement of veterinary medicine.

Interbreed variation of biomarkers of lipid and glucose metabolism in dogs.

BACKGROUND: Markers of lipid and glucose metabolism are used in both clinical practice and research. Detection of abnormal laboratory results often relies on species-specific reference intervals, but interbreed variation can also affect data interpretation. OBJECTIVES: The purpose of the present study was to compare concentrations of selected biochemical variables among different dog breeds. METHODS: We analyzed a database containing information on biochemical variables from 534 dogs belonging to nine different breeds. All dogs were confirmed to be healthy based on history, physical examination, and ancillary tests. Concentrations of glucose, fructosamine, insulin, cholesterol, triglycerides, fatty acids, and C-reactive protein were compared using the nonparametric Kruskal-Wallis and Dunn's tests. RESULTS: All variables tested showed significant interbreed differences, although all breeds remained within the previously established RIs for dogs. Fructosamine, insulin, and cholesterol showed a wide interbreed variation that could affect the interpretation of results. CONCLUSIONS: Breed is an important factor to consider when assessing energy metabolism in dogs, especially for markers like fructosamine, insulin, and cholesterol, which vary considerably among breeds.

Scaling resolution of variant classification differences in ClinVar between 41 clinical laboratories through an outlier approach.

ClinVar provides open access to variant classifications shared from many clinical laboratories. Although most classifications are consistent across laboratories, classification differences exist. To facilitate resolution of classification differences on a large scale, clinical laboratories were encouraged to reassess outlier classifications of variants with medically significant differences (MSDs). Outliers were identified by first comparing ClinVar submissions from 41 clinical laboratories to detect variants with MSDs between the laboratories (650 variants). Next, MSDs were filtered for variants with ≥3 classifications (244 variants), of which 87.6% (213 variants) had a majority consensus in ClinVar, thus allowing for identification of outlier classifications in need of reassessment. Laboratories with outlier classifications were sent a custom report and encouraged to reassess variants. Results were returned for 204 (96%) variants, of which 62.3% (127) were resolved. Of those 127, 64.6% (82) were resolved due to reassessment prompted by this study and 35.4% (45) resolved by a previously completed reassessment. This study demonstrates a scalable approach to classification resolution and capitalizes on the value of data sharing within ClinVar. These activities will help the community move toward more consistent variant classifications, which will improve the care of patients with, or at risk for, genetic disorders.

Heterozygous junctophilin-2 (JPH2) p.(Thr161Lys) is a monogenic cause for HCM with heart failure.

During the last two decades, mutations in sarcomere genes have found to comprise the most common cause for hypertrophic cardiomyopathy (HCM), but still significant number of patients with dominant HCM in the family are left without molecular genetic diagnosis. Next generation sequencing (NGS) does not only enable evaluation of established HCM genes but also candidate genes for cardiomyopathy are frequently tested which may lead to a situation where conclusive interpretation of the variant requires extensive family studies. We aimed to characterize the phenotype related to a variant in the junctophilin-2 (JPH2) gene, which is less known non-sarcomeric candidate gene. In addition, we did extensive review of the literature and databases about JPH2 variation in association with cardiac disease. We characterize nine Finnish index patients with HCM and heterozygous for JPH2 c.482C>A, p.(Thr161Lys) variant were included and segregation studies were performed. We identified 20 individuals affected with HCM with or without systolic heart failure and conduction abnormalities in the nine Finnish families with JPH2 p.(Thr161Lys) variant. We found 26 heterozygotes with the variant and penetrance was 71% by age 60 and 100% by age 80. Co-segregation of the variant with HCM phenotype was observed in six families. Main clinical features were left ventricular hypertrophy, arrhythmia vulnerability and conduction abnormalities including third degree AV-block. In some patients end-stage severe left ventricular heart failure with normal or mildly enlarged diastolic dimensions was detected. In conclusion, we propose that the heterozygous JPH2 p.(Thr161Lys) variant is a new Finnish mutation causing atypical HCM.

Case reports of two pedigrees with recessive arrhythmogenic right ventricular cardiomyopathy associated with homozygous Thr335Ala variant in DSG2.

BACKGROUND: Arrhythmogenic right ventricular cardiomyopathy (ARVC) is an inherited cardiac disease, involving changes in ventricular myocardial tissue and leading to fatal arrhythmias. Mutations in desmosomal genes are thought to be the main cause of ARVC. However, the exact molecular genetic etiology of the disease still remains largely inconclusive, and this along with large variabilities in clinical manifestations complicate clinical diagnostics. CASE PRESENTATION: We report two families (n = 20) in which a desmoglein-2 (DSG2) missense variant c.1003A > G, p.(Thr335Ala) was discovered in the index patients using next-generation sequencing panels. The presence of this variant in probands' siblings and children was studied by Sanger sequencing. Five homozygotes and nine heterozygotes were found with the mutation. Participants were evaluated clinically where possible, and available medical records were obtained. All patients homozygous for the variant fulfilled the current diagnostic criteria for ARVC, whereas none of the heterozygous subjects had symptoms suggestive of ARVC or other cardiomyopathies. CONCLUSIONS: The homozygous DSG2 variant c.1003A > G co-segregated with ARVC, indicating autosomal recessive inheritance and complete penetrance. More research is needed to establish a detailed understanding of the relevance of rare variants in ARVC associated genes, which is essential for informative genetic counseling and rational family member testing.

ADAM23 is a common risk gene for canine idiopathic epilepsy.

BACKGROUND: Idiopathic or genetic adult-onset epilepsy is a common neurological disorder in domestic dogs. Genetic association has been reported only with ADAM23 on CFA 37 in few breeds. To identify novel epilepsy genes, we performed genome-wide association (GWA) analyses in four new breeds, and investigated the association of the previously reported ADAM23 haplotype with the epilepsy phenotype in eight breeds. RESULTS: GWA analysis did not reveal new epilepsy loci. ADAM23 association (p < 0.05) was identified in five breeds. Combined analysis of all eight breeds showed significant association (p = 4.6e-6, OR 1.9). CONCLUSIONS: Our results further support the role of ADAM23 in multiple breeds as a common risk gene for epilepsy with low penetrance. The lack of findings in the GWA analyses points towards inefficient capture of genetic variation by the current SNP arrays, causal variant(s) with low penetrance and possible phenocopies. Future work will include studies on ADAM23 function and expression in canine neurons, as well as whole-genome sequencing in order to identify additional IE genes.

Multiple Changes of Gene Expression and Function Reveal Genomic and Phenotypic Complexity in SLE-like Disease.

The complexity of clinical manifestations commonly observed in autoimmune disorders poses a major challenge to genetic studies of such diseases. Systemic lupus erythematosus (SLE) affects humans as well as other mammals, and is characterized by the presence of antinuclear antibodies (ANA) in patients' sera and multiple disparate clinical features. Here we present evidence that particular sub-phenotypes of canine SLE-related disease, based on homogenous (ANA(H)) and speckled ANA (ANA(S)) staining pattern, and also steroid-responsive meningitis-arteritis (SRMA) are associated with different but overlapping sets of genes. In addition to association to certain MHC alleles and haplotypes, we identified 11 genes (WFDC3, HOMER2, VRK1, PTPN3, WHAMM, BANK1, AP3B2, DAPP1, LAMTOR3, DDIT4L and PPP3CA) located on five chromosomes that contain multiple risk haplotypes correlated with gene expression and disease sub-phenotypes in an intricate manner. Intriguingly, the association of BANK1 with both human and canine SLE appears to lead to similar changes in gene expression levels in both species. Our results suggest that molecular definition may help unravel the mechanisms of different clinical features common between and specific to various autoimmune disease phenotypes in dogs and humans.

Identification of a common risk haplotype for canine idiopathic epilepsy in the ADAM23 gene.

BACKGROUND: Idiopathic epilepsy is a common neurological disease in human and domestic dogs but relatively few risk genes have been identified to date. The seizure characteristics, including focal and generalised seizures, are similar between the two species, with gene discovery facilitated by the reduced genetic heterogeneity of purebred dogs. We have recently identified a risk locus for idiopathic epilepsy in the Belgian Shepherd breed on a 4.4 megabase region on CFA37. RESULTS: We have expanded a previous study replicating the association with a combined analysis of 157 cases and 179 controls in three additional breeds: Schipperke, Finnish Spitz and Beagle (p(c) = 2.9e-07, p(GWAS) = 1.74E-02). A targeted resequencing of the 4.4 megabase region in twelve Belgian Shepherd cases and twelve controls with opposite haplotypes identified 37 case-specific variants within the ADAM23 gene. Twenty-seven variants were validated in 285 cases and 355 controls from four breeds, resulting in a strong replication of the ADAM23 locus (p(raw) = 2.76e-15) and the identification of a common 28 kb-risk haplotype in all four breeds. Risk haplotype was present in frequencies of 0.49-0.7 in the breeds, suggesting that ADAM23 is a low penetrance risk gene for canine epilepsy. CONCLUSIONS: These results implicate ADAM23 in common canine idiopathic epilepsy, although the causative variant remains yet to be identified. ADAM23 plays a role in synaptic transmission and interacts with known epilepsy genes, LGI1 and LGI2, and should be considered as a candidate gene for human epilepsies.

The Shepherds' Tale: A Genome-Wide Study across 9 Dog Breeds Implicates Two Loci in the Regulation of Fructosamine Serum Concentration in Belgian Shepherds.

Diabetes mellitus is a serious health problem in both dogs and humans. Certain dog breeds show high prevalence of the disease, whereas other breeds are at low risk. Fructosamine and glycated haemoglobin (HbA1c) are two major biomarkers of glycaemia, where serum concentrations reflect glucose turnover over the past few weeks to months. In this study, we searched for genetic factors influencing variation in serum fructosamine concentration in healthy dogs using data from nine dog breeds. Considering all breeds together, we did not find any genome-wide significant associations to fructosamine serum concentration. However, by performing breed-specific analyses we revealed an association on chromosome 3 (pcorrected ≈ 1:68 × 10-6) in Belgian shepherd dogs of the Malinois subtype. The associated region and its close neighbourhood harbours interesting candidate genes such as LETM1 and GAPDH that are important in glucose metabolism and have previously been implicated in the aetiology of diabetes mellitus. To further explore the genetics of this breed specificity, we screened the genome for reduced heterozygosity stretches private to the Belgian shepherd breed. This revealed a region with reduced heterozygosity that shows a statistically significant interaction (p = 0.025) with the association region on chromosome 3. This region also harbours some interesting candidate genes and regulatory regions but the exact mechanisms underlying the interaction are still unknown. Nevertheless, this finding provides a plausible explanation for breed-specific genetic effects for complex traits in dogs. Shepherd breeds are at low risk of developing diabetes mellitus. The findings in Belgian shepherds could be connected to a protective mechanism against the disease. Further insight into the regulation of glucose metabolism could improve diagnostic and therapeutic methods for diabetes mellitus.

A missense change in the ATG4D gene links aberrant autophagy to a neurodegenerative vacuolar storage disease.

Inherited neurodegenerative disorders are debilitating diseases that occur across different species. We have performed clinical, pathological and genetic studies to characterize a novel canine neurodegenerative disease present in the Lagotto Romagnolo dog breed. Affected dogs suffer from progressive cerebellar ataxia, sometimes accompanied by episodic nystagmus and behavioral changes. Histological examination revealed unique pathological changes, including profound neuronal cytoplasmic vacuolization in the nervous system, as well as spheroid formation and cytoplasmic aggregation of vacuoles in secretory epithelial tissues and mesenchymal cells. Genetic analyses uncovered a missense change, c.1288G>A; p.A430T, in the autophagy-related ATG4D gene on canine chromosome 20 with a highly significant disease association (p = 3.8 x 10-136) in a cohort of more than 2300 Lagotto Romagnolo dogs. ATG4D encodes a poorly characterized cysteine protease belonging to the macroautophagy pathway. Accordingly, our histological analyses indicated altered autophagic flux in affected tissues. The knockdown of the zebrafish homologue atg4da resulted in a widespread developmental disturbance and neurodegeneration in the central nervous system. Our study describes a previously unknown canine neurological disease with particular pathological features and implicates the ATG4D protein as an important autophagy mediator in neuronal homeostasis. The canine phenotype serves as a model to delineate the disease-causing pathological mechanism(s) and ATG4D function, and can also be used to explore treatment options. Furthermore, our results reveal a novel candidate gene for human neurodegeneration and enable the development of a genetic test for veterinary diagnostic and breeding purposes.

Increased expression of MERTK is associated with a unique form of canine retinopathy.

Progressive retinal degenerations are among the most common causes of blindness both in human and in dogs. Canine progressive retinal atrophy (PRA) resembles human retinitis pigmentosa (RP) and is typically characterized by a progressive loss of rod photoreceptors followed by a loss of cone function. The disease gradually progress from the loss of night and day vision to a complete blindness. We have recently described a unique form of retinopathy characterized by the multifocal gray/brown discoloration and thinning of the retina in the Swedish Vallhund (SV) breed. We aimed to identify the genetic cause by performing a genome wide association analysis in a cohort of 18 affected and 10 healthy control dogs using Illumina's canine 22k SNP array. We mapped the disease to canine chromosome 17 (p = 7.7×10(-5)) and found a 6.1 Mb shared homozygous region in the affected dogs. A combined analysis of the GWAS and replication data with additional 60 dogs confirmed the association (p = 4.3×10(-8), OR = 11.2 for homozygosity). A targeted resequencing of the entire associated region in four cases and four controls with opposite risk haplotypes identified several variants in the coding region of functional candidate genes, such as a known retinopathy gene, MERTK. However, none of the identified coding variants followed a compelling case- or breed-specific segregation pattern. The expression analyses of four candidate genes in the region, MERTK, NPHP1, ANAPC1 and KRCC1, revealed specific upregulation of MERTK in the retina of the affected dogs. Collectively, these results indicate that the retinopathy is associated with overexpression of MERTK, however further investigation is needed to discover the regulatory mutation for the better understanding of the disease pathogenesis. Our study establishes a novel gain-of-function model for the MERTK biology and provides a therapy model for retinopathy MERTK inhibitors. Meanwhile, a marker-based genetic counseling can be developed to revise breeding programs.

A novel form of progressive retinal atrophy in Swedish vallhund dogs.

Inherited retinal degenerations, such as retinitis pigmentosa (RP) and age-related macular degeneration (AMD), represent leading causes of incurable blindness in humans. This is also true in dogs, where the term progressive retinal atrophy (PRA) is used to describe inherited photoreceptor degeneration resulting in progressive vision loss. Because of the similarities in ocular anatomy, including the presence of a cone photoreceptor-rich central retinal region, and the close genotype-phenotype correlation, canine models contribute significantly to the understanding of retinal disease mechanisms and the development of new therapies. The screening of the pure-bred dog population for new forms of PRA represents an important strategy to establish new large animal models. By examining 324 dogs of the Swedish vallhund breed in seven countries and across three continents, we were able to describe a new and unique form of PRA characterized by the multifocal appearance of red and brown discoloration of the tapetal fundus followed over time by thinning of the retina. We propose three stages of the disease based on the appearance of the ocular fundus and associated visual deficits. Electroretinography revealed a gradual loss of both rod and cone photoreceptor-mediated function in Stages 2 and 3 of the disease. In the few dogs that suffered from pronounced vision loss, night-blindness occurred first in late Stage 2, followed by decreased day-vision in Stage 3. Histologic examinations confirmed the loss of photoreceptor cells at Stage 3, which was associated with the accumulation of autofluorescent material in the adjacent retinal pigment epithelium. Pedigree analysis was suggestive of an autosomal-recessive mode of inheritance. Mutations in six known canine retinal degeneration genes as well as hypovitaminosis E were excluded as causes of the disease. The observed variability in the age of disease onset and rate of progression suggest the presence of genetic and/or environmental disease modifiers.

Phenotype, inheritance characteristics, and risk factors for idiopathic epilepsy in Finnish Spitz dogs.

OBJECTIVE: To determine the phenotype, inheritance characteristics, and risk factors for idiopathic epilepsy (IE) in Finnish Spitz dogs (FSDs). DESIGN: Prospective epidemiological study. ANIMALS: 2,141 FSDs. PROCEDURES: From 2003 to 2004, questionnaires (n = 5,960) were sent to all owners of 1-to 10-year-old FSDs in Finland. Phone interviews were performed 1 to 2 years later. RESULTS: Estimated prevalence of IE was 5.36% (111/2,069 of FSDs that were still alive). Males were predisposed to IE. The median age of onset was 3 years (range, 0.6 to 10 years). The median seizure frequency was 2 seizures/y (range, 0.5 to 48 seizures/y), and the median duration of the seizure episode was 11.75 minutes (range, 1.5 to 90 minutes). The majority (85%) of the seizures had a focal onset, and 54% were characterized as generalized secondary. A generalized seizure phase was determined to be a risk factor for development of progressive disease. Factors associated with the occurrence of a generalized phase were the age of onset, duration of the seizure, number of feeding times per day, and whether the dog was used for hunting. The seizures were not progressing in 678% of the dogs and were easily controlled by antiepileptic treatment in 78.9% of the dogs. The heritability estimate of IE in FSDs was 0.22; IE was best explained as a polygenic trait. CONCLUSIONS AND CLINICAL RELEVANCE: In the present study conducted in Finland, complex focal seizures were the most common seizure type for FSDs with IE, and a generalized seizure phase was a risk factor for progression of the disease. Results suggested a benign course of epilepsy in FSDs.

A nonsense mutation in the acid α-glucosidase gene causes Pompe disease in Finnish and Swedish Lapphunds.

Pompe disease is a recessively inherited and often fatal disorder caused by the deficiency of acid α-glucosidase, an enzyme encoded by the GAA gene and needed to break down glycogen in lysosomes. This glycogen storage disease type II has been reported also in Swedish Lapphund dogs. Here we describe the genetic defect in canine Pompe disease and show that three related breeds from Scandinavia carry the same mutation. The affected dogs are homozygous for the GAA c.2237G>A mutation leading to a premature stop codon at amino acid position 746. The corresponding mutation has previously been reported in humans and causes infantile Pompe disease in combination with a second fully deleterious mutation. The affected dogs from both the Finnish as well as the Swedish breed mimic infantile-onset Pompe disease genetically, but also clinico-pathologically. Therefore this canine model provides a valuable tool for preclinical studies aimed at the development of gene therapy in Pompe disease.

A SEL1L mutation links a canine progressive early-onset cerebellar ataxia to the endoplasmic reticulum-associated protein degradation (ERAD) machinery.

Inherited ataxias are characterized by degeneration of the cerebellar structures, which results in progressive motor incoordination. Hereditary ataxias occur in many species, including humans and dogs. Several mutations have been found in humans, but the genetic background has remained elusive in dogs. The Finnish Hound suffers from an early-onset progressive cerebellar ataxia. We have performed clinical, pathological, and genetic studies to describe the disease phenotype and to identify its genetic cause. Neurological examinations on ten affected dogs revealed rapidly progressing generalized cerebellar ataxia, tremors, and failure to thrive. Clinical signs were present by the age of 3 months, and cerebellar shrinkage was detectable through MRI. Pathological and histological examinations indicated cerebellum-restricted neurodegeneration. Marked loss of Purkinje cells was detected in the cerebellar cortex with secondary changes in other cortical layers. A genome-wide association study in a cohort of 31 dogs mapped the ataxia gene to a 1.5 Mb locus on canine chromosome 8 (p(raw) = 1.1x10(-7), p(genome) = 7.5x10(-4)). Sequencing of a functional candidate gene, sel-1 suppressor of lin-12-like (SEL1L), revealed a homozygous missense mutation, c.1972T>C; p.Ser658Pro, in a highly conserved protein domain. The mutation segregated fully in the recessive pedigree, and a 10% carrier frequency was indicated in a population cohort. SEL1L is a component of the endoplasmic reticulum (ER)-associated protein degradation (ERAD) machinery and has not been previously associated to inherited ataxias. Dysfunctional protein degradation is known to cause ER stress, and we found a significant increase in expression of nine ER stress responsive genes in the cerebellar cortex of affected dogs, supporting the pathogenicity of the mutation. Our study describes the first early-onset neurodegenerative ataxia mutation in dogs, establishes an ERAD-mediated neurodegenerative disease model, and proposes SEL1L as a new candidate gene in progressive childhood ataxias. Furthermore, our results have enabled the development of a genetic test for breeders.

Identification of a novel idiopathic epilepsy locus in Belgian Shepherd dogs.

Epilepsy is the most common neurological disorder in dogs, with an incidence ranging from 0.5% to up to 20% in particular breeds. Canine epilepsy can be etiologically defined as idiopathic or symptomatic. Epileptic seizures may be classified as focal with or without secondary generalization, or as primary generalized. Nine genes have been identified for symptomatic (storage diseases) and one for idiopathic epilepsy in different breeds. However, the genetic background of common canine epilepsies remains unknown. We have studied the clinical and genetic background of epilepsy in Belgian Shepherds. We collected 159 cases and 148 controls and confirmed the presence of epilepsy through epilepsy questionnaires and clinical examinations. The MRI was normal while interictal EEG revealed abnormalities and variable foci in the clinically examined affected dogs. A genome-wide association study using Affymetrix 50K SNP arrays in 40 cases and 44 controls mapped the epilepsy locus on CFA37, which was replicated in an independent cohort (81 cases and 88 controls; combined p = 9.70×10⁻¹°, OR = 3.3). Fine mapping study defined a ∼1 Mb region including 12 genes of which none are known epilepsy genes or encode ion channels. Exonic sequencing was performed for two candidate genes, KLF7 and ADAM23. No variation was found in KLF7 but a highly-associated non-synonymous variant, G1203A (R387H) was present in the ADAM23 gene (p = 3.7×10⁻8, OR = 3.9 for homozygosity). Homozygosity for a two-SNP haplotype within the ADAM23 gene conferred the highest risk for epilepsy (p = 6.28×10⁻¹¹, OR = 7.4). ADAM23 interacts with known epilepsy proteins LGI1 and LGI2. However, our data suggests that the ADAM23 variant is a polymorphism and we have initiated a targeted re-sequencing study across the locus to identify the causative mutation. It would establish the affected breed as a novel therapeutic model, help to develop a DNA test for breeding purposes and introduce a novel candidate gene for human idiopathic epilepsies.