Congenital nephrotic syndrome with diffuse mesangial sclerosis caused by compound heterozygous mutation in LAMA5 gene.

A two-and-a-half-month-old female infant presented with generalized edema for 10 days. At presentation, she had periorbital puffiness, moderate ascites, and pedal edema. Laboratory investigations revealed serum albumin 1.3 g/dL, spot urine protein to creatinine ratio (Up:Uc) 20.87 mg/mg, total cholesterol 380 mg/dL, and serum creatinine 0.31 mg/dL. Exome sequencing revealed compound heterozygous variants in LAMA5 gene (NM_005560.6). There was a heterozygous likely pathogenic missense variant in exon 2: LAMA5: c.385C > A (depth 195 ×) and another heterozygous pathogenic variant in exon 31: LAMA5: c.3932_3936dup; parental segregation by Sanger sequencing proved that the variants were in trans. Kidney biopsy showed diffuse mesangial sclerosis (DMS). Our case adds LAMA5 gene to the constellation of genes causing DMS, in addition to the classically described WT1, LAMB2, and PLCE1 genes and to the list of genes causing congenital nephrotic syndrome (CNS).

Clinical characteristics, genetic profile and short-term outcomes of children with primary hyperoxaluria type 2: a nationwide experience.

BACKGROUND: Three types of primary hyperoxaluria (PH) are recognized. However, data on PH type 2 (PH2), caused by defects in the GRHPR gene, are limited. METHODS: We reviewed the medical records of patients < 18 years of age with genetically-proven PH2 from seven centres across India to identify the age of onset, patterns of clinical presentation, short-term outcomes and genetic profile, and to determine if genotype-phenotype correlation exists. RESULTS: We report 20 patients (all with nephrolithiasis or nephrocalcinosis) diagnosed to have PH2 at a median (IQR) age of 21.5 (7, 60) months. Consanguinity and family history of kidney stones were elicited in nine (45%) and eight (40%) patients, respectively. The median (IQR) serum creatinine at PH2 diagnosis was 0.45 (0.29, 0.56) mg/dL with the corresponding estimated glomerular filtration rate being 83 (60, 96) mL/1.73 m2/min. A mutational hotspot (c.494 G > A), rare in Caucasians, was identified in 12 (60%) patients. An intronic splice site variant (c.735-1G > A) was noted in five (25%) patients. Four (20%) patients required surgical intervention for stone removal. Major adverse kidney events (mortality or chronic kidney disease (CKD) stages 3-5) were noted in six (30%) patients at a median (IQR) follow-up of 12 (6, 27) months. Risk factors for CKD progression and genotype-phenotype correlation could not be established. CONCLUSIONS: PH2 should no longer be considered an innocuous disease, but rather a potentially aggressive disease with early age of presentation, and possible rapid progression to CKD stages 3-5 in childhood in some patients. A mutational hotspot (c.494 G > A variant) was identified in 60% of cases, but needs further exploration to decipher the genotype-phenotype correlation.

Neurodegeneration with brain iron accumulation: a case series highlighting phenotypic and genotypic diversity in 20 Indian families.

Neurodegeneration with brain iron accumulation (NBIA) is an umbrella term encompassing various inherited neurological disorders characterised by abnormal iron accumulation in basal ganglia. We aimed to study the clinical, radiological and molecular spectrum of disorders with NBIA. All molecular-proven cases of NBIA presented in the last 5 years at 2 tertiary care genetic centres were compiled. Demographic details and clinical and neuroimaging findings were collated. We describe 27 individuals from 20 unrelated Indian families with causative variants in 5 NBIA-associated genes. PLA2G6-associated neurodegeneration (PLAN) was the most common, observed in 13 individuals from 9 families. They mainly presented in infancy with neuroregression and hypotonia. A recurrent pathogenic variant in COASY was observed in two neonates with prenatal-onset severe neurodegeneration. Pathogenic bi-allelic variants in PANK2, FA2H and C19ORF12 genes were observed in the rest, and these individuals presented in late childhood and adolescence with gait abnormalities and extrapyramidal symptoms. No intrafamilial and interfamilial variability were observed. Iron deposition on neuroimaging was seen in only 6/17 (35.3%) patients. A total of 22 causative variants across 5 genes were detected including a multiexonic duplication in PLA2G6. The variants c.1799G > A and c.2370 T > G in PLA2G6 were observed in three unrelated families. In silico assessments of 8 amongst 9 novel variants were also performed. We present a comprehensive compilation of the phenotypic and genotypic spectrum of various subtypes of NBIA from the Indian subcontinent. Clinical presentation of NBIAs is varied and not restricted to extrapyramidal symptoms or iron accumulation on neuroimaging.

Retrospective diagnosis by parental testing in the next generation sequencing era and utility of reanalysis of exome data.

OBJECTIVE: Genetic diseases are an important cause of neonatal and childhood mortality. For couples with a history of demise of previous children, screening for carrier status can be done by exome sequencing (ES) of the parents. Our aim was to describe the clinical utility of "targeted parental ES" in such couples and to assess the utility of reanalysis of parental ES data. METHOD: We analyzed previous records, including ES reports, of 52 families with demise of previous offspring with a suspected genetic disorder. We also retrieved and reanalyzed raw data of parental ES in FASTQ format from the testing lab. RESULTS: A potential diagnosis was obtained in 30/52 (57.7%) of couples. We found 38/70 (54.3%) novel variants in this cohort. Shared carrier status for more than one autosomal or X-linked recessive disorder was identified in 18% of couples. Reanalysis of raw data resulted in a reclassification of variants in 15% of cases. CONCLUSION: Targeted parental ES can be helpful for families with demise of previous offspring with a suspected genetic disorder.

COASY related pontocerebellar hypoplasia type 12: A common Indian mutation with expansion of the phenotypic spectrum.

Pontocerebellar hypoplasia (PCH) type 12 is a rare, perinatal lethal neurodegenerative genetic disorder caused by biallelic mutations in the COASY gene. Herein, we describe the clinical and neuroradiological profile of nine affected fetuses/neonates from five families identified with a common COASY: c.1486-3C>G biallelic variant. Four of the five families were identified after data reanalysis of unresolved, severe PCH like phenotype and the fifth family through collaboration. The common antenatal phenotype was cerebellar hypoplasia. Microcephaly, arthrogryposis, and intrauterine growth restriction were the shared postnatal findings. The neurological manifestations included seizures, poor sucking, and spasticity. Novel findings of corpus callosum agenesis, simplified gyral pattern, normal sized pons, optic neuropathy, and a small thorax are reported in this series. The allele frequency of the COASY: c.1486-3C>G variant was 0.62% in the available Asian Indian database. We describe this as a possible common Indian origin variant. To the best of our knowledge, this is the largest PCH12 series reported.

Whole-exome sequencing and variant spectrum in children with suspected inherited renal tubular disorder: the East India Tubulopathy Gene Study.

BACKGROUND: Inherited tubulopathies are a heterogeneous group of genetic disorders making whole-exome sequencing (WES) the preferred diagnostic methodology. METHODS: This was a multicenter descriptive study wherein children (< 18 years) with clinically suspected tubular disorders were recruited for molecular testing through WES. Multiplex ligation-dependent probe amplification (MLPA) and Sanger sequencing were done when required. Variants were classified as per American College of Medical Genetics 2015 guidelines and pathogenic (P)/likely pathogenic (LP) variants were considered causative. RESULTS: There were 77 index cases (male =73%). Median age at diagnosis was 48 months (IQR 18.5 to 108 months). At recruitment, the number of children in each clinical group was as follows: distal renal tubular acidosis (dRTA) = 25; Bartter syndrome = 18; isolated hypophosphatemic rickets (HP) = 6; proximal tubular dysfunction (pTD) = 12; nephrogenic diabetes insipidus (NDI) = 6; kidney stone/nephrocalcinosis (NC) = 6; others = 4. We detected 55 (24 novel) P/LP variants, providing genetic diagnoses in 54 children (70%). The diagnostic yield of WES was highest for NDI (100%), followed by HP (83%; all X-linked HP), Bartter syndrome (78%), pTD (75%), dRTA (64%), and NC (33%). Molecular testing had a definite impact on clinical management in 24 (31%) children. This included revising clinical diagnosis among 14 children (26% of those with a confirmed genetic diagnosis and 18% of the overall cohort), detection of previously unrecognized co-morbidities among 8 children (sensorineural deafness n = 5, hemolytic anemia n = 2, and dental changes n = 1) and facilitating specific medical treatment for 7 children (primary hyperoxaluria n = 1, cystinosis n = 4, tyrosinemia n = 2). CONCLUSION: WES is a powerful tool in the diagnosis and management of children with inherited tubulopathies in the Indian population. A higher resolution version of the Graphical abstract is available as Supplementary information.

Clinical, Biochemical, Tumoural and Mutation Profile of VHL- and MEN2A-Associated Pheochromocytoma: A Comparative Study.

OBJECTIVE: To compare clinical, biochemical, tumoural and mutational characteristics of Von Hippel Lindau Syndrome (VHL)-associated pheochromocytoma (PCC) to multiple endocrine neoplasia 2A (MEN2A)-associated pheochromocytoma. DESIGN: Retrospective study design in a tertiary health care centre in Northern India. METHODS: A total of 47 patients with biochemical and histologically proven pheochromocytoma/paraganglioma (PCC/PGL): 29 associated with VHL and 18 with MEN2A, were divided in two cohorts, respectively. Analysis of their medical records along with a prospective follow-up was done. RESULTS: There were more children <19 years in VHL group (13 vs 1). Despite majority of VHL-PCC showing elevation of normetanephrine (NMN) (93%) as compared to MEN2A-PCC (22.2%), 75.8% presented with hypertension as compared to MEN2A (33.3%). The average size of VHL-PCC tumours was 5.66 cm. VHL-PCC as compared to MEN2A-PCC were multifocal (75% vs 61.1%), bilateral synchronous (72.4 vs 61.1%) and extra-adrenal (17.2% vs 0%). Both VHL (24%) and MEN2A-PCC (27.7%) showed multiple nodules, but more MEN2A PCC showed extra-tumoural hyperplasia (44.4% vs. 6.8%). In VHL, the commonest mutation (n = 17) was missense mutation with a hot spot on exon 3, while in MEN2A-PCC majority (66.6%) had 634 mutation in exon 11 and only 2 patients had the rare 611 mutation in exon 10. CONCLUSION: In contrast to world literature, our study suggests Indian VHL-PCC can be symptomatic in spite of noradrenergic phenotype, large in size and multifocal. Multiple nodules in VHL-PCC could increase risk of recurrence following subtotal adrenalectomy.

Meckel Gruber and Joubert Syndrome Diagnosed Prenatally: Allelism between the Two Ciliopathies, Complexities of Mutation Types and Digenic Inheritance.

Background: Antenatally detected occipital encephalocele and polycystic kidneys are a common presentation of ciliopathies like Joubert syndrome and Meckel Gruber syndrome which have considerable genetic and phenotypic overlap. Case reports: We describe 3 cases of antenatally diagnosed occipital encephalocele and enlarged kidneys with fetal autopsy, histopathology & exome sequencing results. A novel nonsense variant in the CEP290 gene was reported in first case (Meckel syndrome). The second case shows the importance of fetal exome where the parents were carriers for 2 ciliopathy genes (TMEM138 & SDCCAG8). Diagnosis in this case was confirmed by fetal exome sequencing (Joubert syndrome). Multiexon deletion in TMEM67 and KIF14 present in trans was identified in the third case (Meckel syndrome), likely resulting in digenic inheritance. Conclusion: We report 2 cases of Meckel syndrome with a novel variant and multiexon deletion, and 1 case of Joubert syndrome which depicts the limitations of preconceptional carrier screening in ciliopathies due to overlapping phenotypes.

A data set of variants derived from 1455 clinical and research exomes is efficient in variant prioritization for early-onset monogenic disorders in Indians.

Given the genomic uniqueness, a local data set is most desired for Indians, who are underrepresented in existing public databases. We hypothesize patients with rare monogenic disorders and their family members can provide a reliable source of common variants in the population. Exome sequencing (ES) data from families with rare Mendelian disorders was aggregated from five centers in India. The dataset was refined by excluding related individuals and removing the disease-causing variants (refined cohort). The efficiency of these data sets was assessed in a new set of 50 exomes against gnomAD and GenomeAsia. Our original cohort comprised 1455 individuals from 1203 families. The refined cohort had 836 unrelated individuals that retained 1,251,064 variants with 181,125 population-specific and 489,618 common variants. The allele frequencies from our cohort helped to define 97,609 rare variants in gnomAD and 44,520 rare variants in GenomeAsia as common variants in our population. Our variant dataset provided an additional 1.7% and 0.1% efficiency for prioritizing heterozygous and homozygous variants respectively for rare monogenic disorders. We observed additional 19 genes/human knockouts. We list carrier frequency for 142 recessive disorders. This is a large and useful resource of exonic variants for Indians. Despite limitations, datasets from patients are efficient tools for variant prioritization in a resource-limited setting.

Functional characterization of novel variants in SMPD1 in Indian patients with acid sphingomyelinase deficiency.

Pathogenic variations in SMPD1 lead to acid sphingomyelinase deficiency (ASMD), that is, Niemann-Pick disease (NPD) type A and B (NPA, NPB), which is a recessive lysosomal storage disease. The knowledge of variant spectrum in Indian patients is crucial for early and accurate NPD diagnosis and genetic counseling of families. In this study, we recruited 40 unrelated pediatric patients manifesting symptoms of ASMD and subnormal ASM enzyme activity. Variations in SMPD1 were studied using Sanger sequencing for all exons, followed by interpretation of variants based on American College of Medical Genetics and Genomics & Association for Molecular Pathology (ACMG/AMP) criteria. We identified 18 previously unreported variants and 21 known variants, including missense, nonsense, deletions, duplications, and splice site variations with disease-causing potential. Eight missense variants were functionally characterized using in silico molecular dynamic simulation and in vitro transient transfection in HEK293T cells, followed by ASM enzyme assay, immunoblot, and immunofluorescence studies. All the variants showed reduced ASM activity in transfected cells confirming their disease-causing potential. The study provides data for efficient prenatal diagnosis and genetic counseling of families with NPD type A and B.

Molecular spectrum, family screening and genetic counselling of Spinocerebellar Ataxia (SCA) cases in an Indian scenario.

Spinocerebellar Ataxia (SCA) is a heterogeneous adult-onset disorder with an autosomal dominant inheritance pattern mainly caused by triplet repeat expansions. Clinical diagnosis of SCA is based on phenotypic features followed by confirmation through molecular diagnosis. To identify status of repeat range in Indian SCA cases and provide extended family screening, we enrolled 70 clinical SCA suspects. For molecular diagnosis, multiplex PCR (M-PCR) was used for common Indian SCA subtypes 1, 2, 3, 6, 7, 10, 12 and 17. TP-PCR was further used in SCA2, 7 and 10 to identify larger expansions. Eighteen out of 70 SCA suspects (25%) were found to be positive for various SCA subtypes- (5 SCA1 (28%), 6 SAC2 (34%), 2 SCA3 (12%), 3 SCA7 (16%) and one each for SCA6 (1%) and SCA17 (1%) subtypes). Genetic counselling and extended family screening were offered to all positive cases and yielded additional nine cases. We have established M-PCR and TP-PCR to detect the CAG repeat expansion in SCA suspects. This method can confirm SCA subtypes in a reliable, rapid and cost-effective way. Genetic characterization of SCA-related genes has great clinical relevance, as it could provide additional information and guidance to clinicians and family members regarding prognosis.

Carrier frequency of SMN1-related spinal muscular atrophy in north Indian population: The need for population based screening program.

Chromosome 5q related Spinal muscular atrophy (SMA) is an autosomal recessive, progressive, neuromuscular disorder most commonly caused by homozygous deletion of exon 7 or exon 7 and 8 of SMN1 gene. Being the leading genetic cause of infant mortality, studies of its prevalence and incidence are necessary. Carrier testing for the common pathogenic variant for SMA is offered to the couples visiting our tertiary care hospital in North India. Subjects were tested for SMA carrier status by Multiplex Ligation-dependent Probe amplification (MLPA) technique for deletion of exons 7 and 8 of SMN1 gene. The retrospective data of individuals tested for SMA carrier status in last 4 years (2016-2019) was evaluated. Six hundred and six individuals without family history of SMA or carrier of SMA who were subjected to MLPA based screening for SMA carrier status were included in the study. The carrier frequency of SMN1 deletion (deletion of exon 7 and/or exon 8) was found to be 1 in 38 (16 out of 606). The catchment area of our medical genetics clinic covering the state of Uttar Pradesh (16.5% of Indian population according to censusindia.gov.in, 2011) and neighboring states, showing SMA carrier frequency of 1:38 in a cohort with no prior positive family history has important significance for policy making.

Genetic heterogeneity of disorders with overgrowth and intellectual disability: Experience from a center in North India.

Overgrowth, defined as height and/or OFC ≥ +2SD, characterizes a subset of patients with syndromic intellectual disability (ID). Many of the disorders with overgrowth and ID (OGID) are rare and the full phenotypic and genotypic spectra have not been unraveled. This study was undertaken to characterize the phenotypic and genotypic profile of patients with OGID. Patients with OGID were ascertained from the cohort of patients who underwent cytogenetic microarray (CMA) and/or exome sequencing (ES) at our center over a period of 6 years. Thirty-one subjects (six females) formed the study group with ages between 3.5 months and 13 years. CMA identified pathogenic deletions in two patients. In another 11 patients, a disease causing variant was detected by ES. The spectrum of disorders encompassed aberrations in genes involved in the two main pathways associated with OGID. These were genes involved in epigenetic regulation like NSD1, NFIX, FOXP1, and those in the PI3K-AKT pathway like PTEN, AKT3, TSC2, PPP2R5D. Five novel pathogenic variants were added by this study. NSD1-related Sotos syndrome was the most common disorder, seen in five patients. A causative variant was identified in 61.5% of patients who underwent only ES compared to the low yield of 11.1% in the CMA group. The molecular etiology could be confirmed in 13 subjects with OGID giving a diagnostic yield of 42%. The major burden was formed by autosomal dominant monogenic disorders. Hence, ES maybe a better first-tier genomic test rather than CMA in OGID.

Uptake of next-generation sequencing in children with end-stage renal disease secondary to focal segmental glomerulosclerosis and parental decision for kidney transplantation-Experience from a low resource setting: A Retrospective Cohort Study.

BACKGROUND: Transplantation is the optimal modality for children with ESRD. High risk of disease recurrence and graft loss with FSGS, and its financial implications, may result in families refusing transplantation. Deceased donation is often preferred for FSGS, but access is limited in many low- and middle-income group countries (LMIC; per capita gross national income between $1026 and $3995). As FSGS secondary to an underlying genetic etiology has low recurrence risk, we hypothesized that in LMIC such as India, families with children in ESRD secondary to FSGS with proven pathogenic mutation are more likely to consent for transplantation than those with unknown etiology. METHODS: Prospective cross-sectional study with retrospective chart review was undertaken (March 2011 and February 2019) to identify children with ESRD from FSGS. The objective was to ascertain NGS uptake and findings, parental decision for transplantation, and transplant outcome. RESULTS: 28 children with FSGS started transplant workup, and 15 (54%) families agreed for NGS testing. Pathogenic mutation (NPHS1 x 2, WT1 x 2, COL4A3 x 2, CD2AP, CRB2, COL4A5, INF 2, ACTN4, NPHP4: 1 each) was identified in 12 (80%). 92% (11/12) agreed to proceed with transplantation in contrast to 13% (2/16) who either did not undergo NGS testing or had no pathogenic mutation identified (P = .001). No disease recurrences were noted in those with a known pathogenic mutation. CONCLUSION: In LMIC, NGS results are useful in transplant discussions with families for children with ESRD secondary to FSGS.

Spondyloepiphyseal dysplasia Omani type: CHST3 mutation spectrum and phenotypes in three Indian families.

We describe three consanguineous Indian families with a distinct form of spondyloepiphyseal dysplasia (SED Omani type). It is an autosomal recessive disorder due to mutation in CHST3 gene. CHST3 gene encodes the enzyme chondroitin 6-O-sulfotransferase-1 (C6ST-1) which mediates the sulfation of proteoglycans, (chondroitin sulfate), in the extracellular matrix of cartilage. CHST3 gene was sequenced in probands from three different families with SED. In two families missense mutations (c.904G>C predicting the substitution D302H) and c.491C>T (P164L) were identified. A frameshift (insertion) mutation (c.533_534ins G predicting the substitution A179Rfs*) was found in the third family. SNP micrarray in the family 2 helped to localize the common areas of homozygosity and identified the candidate gene. The confirmation by molecular diagnosis will be useful in the management and in the counseling of affected patients and their families. The presence of sclerosis of cranial sutures adds to the phenotypic spectrum of the disorder. Severe cardiac valvular disease in a case and triangular epiphyses of knees are other features which are highlighted in this report. © 2016 Wiley Periodicals, Inc.

Spectrum of SMPD1 mutations in Asian-Indian patients with acid sphingomyelinase (ASM)-deficient Niemann-Pick disease.

Acid sphingomyelinase (ASM)-deficient Niemann-Pick disease is an autosomal recessive lysosomal storage disorder caused by biallelic mutations in the SMPD1 gene. To date, around 185 mutations have been reported in patients with ASM-deficient NPD world-wide, but the mutation spectrum of this disease in India has not yet been reported. The aim of this study was to ascertain the mutation profile in Indian patients with ASM-deficient NPD. We sequenced SMPD1 in 60 unrelated families affected with ASM-deficient NPD. A total of 45 distinct pathogenic sequence variants were found, of which 14 were known and 31 were novel. The variants included 30 missense, 4 nonsense, and 9 frameshift (7 single base deletions and 2 single base insertions) mutations, 1 indel, and 1 intronic duplication. The pathogenicity of the novel mutations was inferred with the help of the mutation prediction software MutationTaster, SIFT, Polyphen-2, PROVEAN, and HANSA. The effects of the identified sequence variants on the protein structure were studied using the structure modeled with the help of the SWISS-MODEL workspace program. The p. (Arg542*) (c.1624C>T) mutation was the most commonly identified mutation, found in 22% (26 out of 120) of the alleles tested, but haplotype analysis for this mutation did not identify a founder effect for the Indian population. To the best of our knowledge, this is the largest study on mutation analysis of patients with ASM-deficient Niemann-Pick disease reported in literature and also the first study on the SMPD1 gene mutation spectrum in India. © 2016 Wiley Periodicals, Inc.

Incessant left ventricular tachycardia of unusual etiology.

Apart from Coronary artery disease, left ventricular tachycardia may result from cardiac sarcoidosis, left ventricular tumor, chagas disease and idiopathic left ventricular tachycardia. We report a rare case of incessant left ventricular tachycardia resulting from left dominant arrhythmogenic cardiomyopathy.

A Defective Oxidative Burst and Impaired Antigen Presentation are Hallmarks of Human Visceral Leishmaniasis.

PURPOSE: Survival of the Leishmania parasite within monocytes hinges on its ability to effectively nullify their microbicidal effector mechanisms. Accordingly, this study aimed to delineate this biological niche in patients with visceral leishmaniasis (VL). METHODS: In monocytes, the redox status, antigen presenting capacity, expression of Toll-like receptors (TLRs), co-stimulatory molecules (CD80/86) and generation of intracellular cytokines (IL-8, IL-1ß, IL-10 and LAP-TGF-ß1) was measured by flow cytometry, levels of circulating cytokines (IL-1ß, IL-6, TNF-α, IL-8, IL-4, IL-13, IL-10 and GM-CSF) by ELISA and arginase activity by spectrophotometry. RESULTS: Within monocytes, generation of an oxidative burst was markedly attenuated as evident by decreased generation of nitric oxide and reactive oxygen species, concomitant with raised levels of thiols. This was accompanied by lowered frequency of TLR4(+) monocytes, but the arginase activity remained unaltered. Pathogen persistence was enhanced by the predominance of anti-inflammatory cytokines within monocytes, notably IL-10. Alongside, development of adaptive immunity was severely attenuated as manifested by a pronounced impairment of antigen presentation and co-stimulation evident by down regulation of CD54, HLA-DR and CD86. Treatment corrected the redox imbalance and reversed the impaired antigen presentation. CONCLUSIONS: In VL, monocyte functions were severely impaired facilitating parasite persistence; anti-leishmanial chemotherapy mediated parasite elimination through modulation of the macrophage microenvironment by restoring its redox status and antigen presenting capacity.

Co-occurrence of a de novo Williams and 22q11.2 microdeletion syndromes.

We report on a child with de novo deletions involving the 7q11.23 (Williams syndrome) and 22q11.2 (Velocardiofacial/DiGeorge syndrome) regions. We describe the clinical features of this rare double microdeletion syndrome reported here for the first time.