Inborn Errors of Ketogenesis: Novel Variants, Clinical Presentation, and Follow-Up in a Series of Four Patients.

Inborn errors of ketogenesis are rare disorders that result in acute and fulminant decompensation during lipolytic stress, particularly in infants and children. These include mitochondrial 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) synthase (HMGCS) deficiency and HMG-CoA lyase (HMGCL) deficiency. In this series, we describe the clinical, biochemical, and molecular profiles of four patients along with dietary interventions and their outcomes on a long-term follow-up. Two patients each of HMGCS and HMGCL deficiency were evaluated with clinical history, biochemical investigations, including tandem mass spectrometry (TMS) and urine gas chromatography-mass spectrometry (GCMS). Molecular analysis was performed by whole-exome sequencing, as well as exon array validated by long-range polymerase chain reaction. All individuals were diagnosed with acute metabolic decompensation in the early infancy period except one with HMGCL deficiency who had the first presentation at 5 years of age. Central nervous system manifestations, severe metabolic acidosis, hyperammonemia, hypoglycemia with a normal lactate, and absence of urinary ketones were observed in all the affected individuals. The disorder was life-threatening in three individuals and one succumbed to the illness. TMS was nonspecific and urine GCMS revealed dicarboxylic aciduria in HMGCS deficiency. Both the patients with HMGCL deficiency demonstrated elevated 3 hydroxyisovaleryl carnitine levels in TMS and metabolites of leucine degradation in urine GCMS. We identified five novel variants that included a large deletion involving exon 2 in HMGCL gene. There was no evidence of long-term neurological sequelae in the living individuals. Diet with moderation of fat intake was followed in two individuals with HMGCS deficiency. Low leucine and protein diet with moderation of fat intake was followed in the individual with HMGCL deficiency. All affected individuals are thriving well with no further major metabolic decompensation.

Anastrozole-mediated modulation of mitochondrial activity by inhibition of mitochondrial permeability transition pore opening: an initial perspective.

The mitochondrial permeability transition pore (mtPTP) plays a vital role in altering the structure and function of mitochondria. Cyclophilin D (CypD) is a mitochondrial protein that regulates mtPTP function and a known drug target for therapeutic studies involving mitochondria. While the effect of aromatase inhibition on the mtPTP has been studied previously, the effect of anastrozole on the mtPTP has not been completely elucidated. The role of anastrozole in modulating the mtPTP was evaluated by docking, molecular dynamics and network-guided studies using human CypD data. The peripheral blood mononuclear cells (PBMCs) of patients with mitochondrial disorders and healthy controls were treated with anastrozole and evaluated for mitochondrial permeability transition pore (mtPTP) function and apoptosis using a flow cytometer. Spectrophotometry was employed for estimating total ATP levels. The anastrozole-CypD complex is more stable than cyclosporin A (CsA)-CypD. Anastrozole performed better than cyclosporine in inhibiting mtPTP. Additional effects included inducing mitochondrial membrane depolarization and a reduction in mitochondrial swelling and superoxide generation, intrinsic caspase-3 activity and cellular apoptosis, along with an increase in ATP levels. Anastrozole may serve as a potential therapeutic agent for mitochondrial disorders and ameliorate the clinical phenotype by regulating the activity of mtPTP. However, further studies are required to substantiate our preliminary findings.Communicated by Ramaswamy H. Sarma.

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.

T2 olivary nuclei hyperintensities: A characteristic neuroimaging finding in FIG4-related leukoencephalopathy.

FIG4 related leukoencephalopathy has recently been considered as an expanded spectrum of FIG4 related disorders characterized by upper and lower motor neuron involvement, dystonia, intellectual disability, bulbar symptoms with cerebellar atrophy. We report a 7-year-old girl who presented with classic clinical features of FIG4 related leukoencephalopathy and neuroimaging showed characteristic T2 olivary nuclei hyperintensities in addition to bilateral parietal lobe and thalamic hyperintensities and mild cerebellar atrophy. Trio exome sequencing with Sanger confirmation revealed a novel variant c.504C>G in the FIG4 gene. Phase contrast microscopy of skin fibroblast cultures detect enlarged vacuoles in 50% of patient's fibroblasts as opposed to 18.6% vacuolation in cultured control fibroblasts (p < 0.00001), a feature characteristic of fibroblasts with deleterious variants of FIG4. In addition, we have reviewed and compared the phenotypic features of published cases of FIG4 related leukoencephalopathy from literature. This case adds to the delineation of FIG4 related leukoencephalopathy phenotype. The radiological finding of T2 inferior olivary nuclei hyperintensities appear to be characteristic for the phenotype or at least for the cases due to variants in and around the 168th codon and active effort should be made to detect the same as it can add to the genotype phenotype spectrum.

Haemophilia management programme: Transformation during COVID-19.

Background & objectives: Haemophilia is a debilitating bleeding disorder with significant comorbidities affecting the quality of life. In India, the management of these individuals is still limited to on-demand institutional treatment with coagulant factors. In this study, we highlighted the problems faced by these patients in the COVID-19 period due to nationwide lockdown. Methods: A retrospective study was done to ascertain the trend in the number of patients with haemophilia A and B visiting the hospital, those succumbing to haemophilic complications and indications for factor requirement in the pre-COVID (October 2019-March 2020) and during the COVID-19 period (April-September 2020). Representative cases with unusual complications were described along with significant challenges faced in providing standard care of treatment to these individuals due to the COVID-19 pandemic. Results: A total of 818 and 162 individuals with haemophilia A and B, respectively, were registered with the department. The overall number of patient visits to the hospital significantly reduced from an average of 6.9 outpatient department (OPD) visits per patient in the pre-COVID-19 period to an average of 3.9 OPD visits per patient and admissions reduced to 50 per cent during the COVID-19 period. This led to a reduction in utilization of factors VIII and IX except VIIa for haemophilia with inhibitors. There was no factor utilization for elective surgeries during the COVID-19 period. A total of eight patients succumbed to haemophilia-related complications during the COVID-19 period due to delay in reaching the hospital. The challenges faced in the management of three cases with musculoskeletal bleeds, one case with scrotal haematoma and one with haemothorax during the COVID-19 period were also highlighted. Interpretation & conclusions: COVID-19 pandemic has unveiled the need for on-demand home treatment with coagulant factors and has also brought to light the existing need for primary prophylaxis, especially for younger individuals with haemophilia.

Integrated Management Strategies for Epidermolysis Bullosa: Current Insights.

Epidermolysis bullosa (EB) is a group of rare genodermatoses that is characterized by skin fragility resulting from minor trauma. There are four major subtypes, namely, EB simplex, junctional EB, dystrophic EB and Kindler EB, depending upon the localization of defective protein and resulting plane of blister formation. The phenotype is heterogeneous in terms of severity and majority of them present at birth or neonatal period. Currently, the treatment is mainly supportive and requires multidisciplinary care. The complex molecular pathology creates difficulty in discovering a unified curative treatment approach. But with arduous efforts, significant progress has been made in the development of treatment strategies in the last decade. The management strategies range from targeting the underlying causative factor to symptom-relieving approaches, and include gene, mRNA, protein, cell and combination therapies. In this review, we enumerate the promising approaches that are currently under various stages of investigation to provide effective treatment for patients with EB.

Deciphering the molecular landscape of microcephaly in 87 Indian families by exome sequencing.

Microcephaly is a frequent feature of neurodevelopmental disorders (NDDs). Our study presents the heterogeneous spectrum of genetic disorders in patients with microcephaly either in isolated form or in association with other neurological and extra-neural abnormalities. We present data of 91 patients from 87 unrelated families referred to our clinic during 2016-2020 and provide a comprehensive clinical and genetic landscape in the studied cohort. Molecular diagnosis using exome sequencing was made in 45 families giving a yield of 51.7%. In 9 additional families probable causative variants were detected. We identified disease causing variations in 49 genes that are involved in different functional pathways Among these, 36 had an autosomal recessive pattern, 8 had an autosomal dominant pattern (all inherited de novo), and 5 had an X-linked pattern. In 41 probands where sequence variations in autosomal recessive genes were identified 31 were homozygotes (including 16 from non-consanguineous families). The study added 28 novel pathogenic/likely pathogenic variations. The study also calls attention to phenotypic variability and expansion in spectrum as well as uncovers genes where microcephaly is not reported previously or is a rare finding. We here report phenotypes associated with the genes for ultra-rare NDDs with microcephaly namely ATRIP, MINPP1, PNPLA8, AIMP2, ANKLE2, NCAPD2 and TRIT1.

Autosomal recessive spinocerebellar ataxia-20 due to a novel SNX14 variant in an Indian girl.

Autosomal recessive spinocerebellar ataxia-20 is a rare disorder having distinctive coarse facies in addition to intellectual disability and cerebellar ataxia, with less than 35 cases reported worldwide. It is caused by biallelic variants in the SNX14 gene and is classified under the group of autophagy disorders. We report a 9-year-old girl who presented with classic clinical features of autosomal recessive spinocerebellar ataxia-20 and cerebellar atrophy on magnetic resonance imaging of brain. Trio exome sequencing with Sanger confirmation revealed a novel splice site variant, c.140 + 3A > T in the SNX14 gene. The variant pathogenicity established by mRNA expression study showed a significant reduction in the expression levels of SNX14 gene in proband and her parents on comparison to the control. The electron microscopy of the skin fibroblasts of proband depicted numerous cytoplasmic vacuoles with variable degrees of dense staining material. In addition, we have briefly reviewed and compared the phenotypic features of published cases of autosomal recessive spinocerebellar ataxia-20 in the literature. Coarse facies, intellectual disability with severe speech delay, hypotonia, and cerebellar atrophy were universal findings in the published cases. This is the second reported case from the Indian subcontinent.

Congenital Hyperinsulinemia of Infancy: Role of Molecular Testing in Management and Genetic Counseling.

Congenital hyperinsulinemia (CHI) is a genetically and clinically heterogenous disorder. In addition to the standard care of management of the proband, genetic counseling regarding the risk of recurrence in the future siblings is an important part in the management of the disorder. The counseling needs identification of accurate etiology and is challenging due to the complexity of the molecular mechanisms of CHI. This case highlights the importance of molecular testing which not only helped in planning the management of the proband with CHI but also helped in providing genetic counseling for which the family had consulted the medical genetics department.

Biallelic cGMP-dependent type II protein kinase gene (PRKG2) variants cause a novel acromesomelic dysplasia.

BACKGROUND: C-type natriuretic peptide (CNP), its endogenous receptor, natriuretic peptide receptor-B (NPR-B), as well as its downstream mediator, cyclic guanosine monophosphate (cGMP) dependent protein kinase II (cGKII), have been shown to play a pivotal role in chondrogenic differentiation and endochondral bone growth. In humans, biallelic variants in NPR2, encoding NPR-B, cause acromesomelic dysplasia, type Maroteaux, while heterozygous variants in NPR2 (natriuretic peptide receptor 2) and NPPC (natriuretic peptide precursor C), encoding CNP, cause milder phenotypes. In contrast, no variants in cGKII, encoded by the protein kinase cGMP-dependent type II gene (PRKG2), have been reported in humans to date, although its role in longitudinal growth has been clearly demonstrated in several animal models. METHODS: Exome sequencing was performed in two girls with severe short stature due to acromesomelic limb shortening, brachydactyly, mild to moderate platyspondyly and progressively increasing metaphyseal alterations of the long bones. Functional characterisation was undertaken for the identified variants. RESULTS: Two homozygous PRKG2 variants, a nonsense and a frameshift, were identified. The mutant transcripts are exposed to nonsense-mediated decay and the truncated mutant cGKII proteins, partially or completely lacking the kinase domain, alter the downstream mitogen activation protein kinase signalling pathway by failing to phosphorylate c-Raf 1 at Ser43 and subsequently reduce ERK1/2 activation in response to fibroblast growth factor 2. They also downregulate COL10A1 and upregulate COL2A1 expression through SOX9. CONCLUSION: In conclusion, we have clinically and molecularly characterised a new acromesomelic dysplasia, acromesomelic dysplasia, PRKG2 type (AMDP).

Loes Score: Clinical and Radiological Profile of 22 Patients of X-Linked Adrenoleukodystrophy: Case Series from a Single Center.

Introduction X-linked adrenoleukodystrophy (X-ALD) is a devastating disease with a wide spectrum of presentation ranging from asymptomatic to a rapidly progressive childhood cerebral form. The gene responsible for adrenoleukodystrophy is ABCD1 gene, required for ß oxidation of fatty acids in various tissues. While biochemical and molecular techniques are available to confirm the diagnosis, brain magnetic resonance imaging (MRI) utilizing Loes score has been used for both prognosis and timely direction of hematopoietic stem cell therapy. Materials and Methods During the study period of 2016 to 2020, 22 individuals including 19 individuals with features suggestive of X-ALD and 3 asymptomatic siblings were evaluated from a single center in North India. After biochemical and molecular confirmation of the disease, detailed clinical and radiological findings using MRI brain were documented. A radiological scoring pattern proposed by Loes was employed to identify the severity of the disorder. Results The most common clinical presentations were visual difficulty and muscular weakness (58%). All symptomatic individuals had classic neuroimaging findings in the form of hyperintensities involving the parieto-occipital area and splenium of corpus callosum. Severe involvement in the form of global atrophy was observed in 52.6% of individuals. Asymptomatic siblings also showed neurological involvement based on MRI with highest Loes score of 9 in one individual. Conclusion This case series describes the clinical and radiological profile and employment of Loes score in individuals with X-ALD. Early identification of asymptomatic individuals by neuroimaging and use of Loes severity score for monitoring and disease progression will help in making therapeutic decisions in a timely manner.

Phenotypic and genotypic spectrum of CTSK variants in a cohort of twenty-five Indian patients with pycnodysostosis.

BACKGROUND: Pycnodysostosis is an autosomal recessive skeletal dysplasia with easily recognizable clinical features and marked molecular heterogeneity. In this study, we explored the clinical and molecular spectrum of 25 Indian patients with pycnodysostosis from 20 families. METHODS: Clinical information was collected on a predesigned clinical proforma. Sanger method was employed to sequence all the exons and exon/intron boundaries of the CTSK gene. Novel variants were systematically assessed by prediction softwares and protein modelling. The pathogenicity of variant was established based on ACMG-AMP criteria. An attempt was also made to establish a genotype-phenotype correlation and devise a diagnostic scoring system based on clinical and radiological findings. RESULTS: Consanguinity and positive family history were present in 65% (13/20) and 45% (9/20) of the families respectively. Short stature and fractures were the predominant presenting complaints and was evident in 96% (24/25) and 32% (8/25) of affected individuals respectively. Gestalt facial phenotype and acro-osteolysis were present in 76% (19/25) and 82.6% (19/23) of the individuals respectively. Hepatosplenomegaly was present in 15% (3/20) of the individuals with one of them having severe anaemia. Causative sequence variations were identified in all of them. A total of 19 variants were identified from 20 families amongst which 10 were novel. Homozygous variants were identified in 90% (18/20) families. Amongst the novel variants, there was a considerable proportion (40%) of frameshift variants (4/10). No significant genotype-phenotype correlation was noted. Scoring based on clinical and radiological findings led to the proposal that a minimum of 2 scores in each category is required in addition to high bone density to diagnose pycnodysostosis with certainty. CONCLUSION: This study delineated the genotypic and phenotypic characterisation of Indian patients with pycnodysostosis with identification of 10 novel variants. We also attempted to develop a clinically useful diagnostic scoring system which requires further validation.

Monogenic Lupus with IgA Nephropathy Caused by Spondyloenchondrodysplasia with Immune Dysregulation.

Monogenic disorders causing systemic lupus erythematosus represent a small subset of cases. Type-1 interferonopathies, like spondyloenchondrodysplasia with immune dysregulation constitute an important functional category of monogenic lupus. Apart from autoimmune disorders, neurological and skeletal abnormalities are additional manifestations observed in this disorder. A young female presented with seizures due to acute hemorrhagic stroke secondary to malignant hypertension. On evaluating the cause for hypertension, there was evidence of glomerulonephritis and multiple autoantibodies positivity including dsDNA. A diagnosis of lupus was made based on clinical and laboratory findings. Kidney biopsy revealed mesangial proliferative glomerulonephritis with predominant IgA deposits favouring IgA nephropathy. Additional features in the form of short stature with vertebral abnormalities and bilateral basal ganglia calcification led to evaluation of Type-1 interferonopathies. Sanger sequencing identified a novel compound heterozygous variants c.550C>T (p.Q184*) in exon 3 and c.740T>G (p.L247R) in exon 4 of ACP5 gene. Parents were found to be carriers of the variants in ACP5 gene. Management included antihypertensive agents and symptomatic therapy. On follow-up, there was complete resolution of glomerulonephritis and normalization of blood pressure. This case report documents the classic phenotype comprising autoimmune, skeletal, and neurological abnormalities in spondyloenchondrodysplasia with immune dysregulation with a novel variant on Sanger sequencing in an Indian patient. This report also highlights the rare coexistence of IgA nephropathy in monogenic lupus.

Kallmann Syndrome and X-linked Ichthyosis Caused by Translocation Between Chromosomes X and Y: A Case Report.

BACKGROUND: Xp22.3 region is characterized by low frequency of interspersed repeats and low GC content. Several clinically important genes including ANOS1 (KAL1) reside in this region. This gene was first identified due to translocation between chromosomes X and Y in a patient with Kallmann syndrome. CASE PRESENTATION: A 20 year old male presented with complaints of delayed secondary sexual characteristics, impaired sense of smell, and poor scholastic performance. On examination, he had short stature (151 cm; <3rd centile). His sexual maturity corresponded to Tanner stage 3. Stretched penile length was 3.6 cm (<3rd centile). Right testis was undescended with low left testicular volume (12 ml). There was mild ichthyosis over abdomen and back. He had hyposmia, hoarse voice, and synkinesia. Investigations were suggestive of hypogonadotrophic hypogonadism. Karyotype revealed an extra chromosomal material on p arm of chromosome X (46,Xp+,Y). On cytogenetic microarray, deletion of 8.3 Mb on Xp22.33 region and duplication of 12.8 Mb on Yq11.22 region were identified. The breakpoint on X chromosome resulted in deletion of exons 7-14 of ANOS1 gene and complete STS, NLGN4X, ARSL (ARSE), SHOX, and VCX genes. CONCLUSION: Patients diagnosed with Kallmann syndrome should receive careful clinical evaluation to detect presence of a contiguous gene syndrome.

Association Between Neonatal Thyroid Stimulating Hormone Status and Maternal Urinary Iodine Status.

BACKGROUND: Maternal urinary iodine concentration (MUIC) and percentage of neonates with Thyroid stimulating hormone (TSH) >5 mIU/L are amongst the parameters suggested for assessing adequate iodine status. OBJECTIVE: To assess the correlation between MUIC and neonatal TSH levels. STUDY DESIGN: Cross-sectional. SETTINGS: Tertiary care center in Delhi, India, between November 2015 to November 2016. PARTICIPANTS: Postnatal mother-neonate dyads. METHODS: TSH levels assessed among neonatal samples were stratified as below and above 5 mIU/L. MUIC was measured in 544 mothers, 400 mother-neonate dyads with neonatal TSH levels >5 mIU/L (cases) and 144 mother-neonate newborn mother dyads with neonatal TSH <5 mIU/L (controls). RESULTS: Results: The percentage of mothers with iodine insufficiency (9.8% vs 5.6%) as well as iodine excess (54.3% vs 41.7%) were significant higher in cases than controls. Mean TSH was also higher (P=0.0002) in both the iodine deficient and iodine excess group. There was no correlation between neonatal TSH values and MUIC. CONCLUSION: Lack of correlation between neonatal TSH and MUIC is due to iodine excess together with iodine deficiency.