A Human Pleiotropic Multiorgan Condition Caused by Deficient Wnt Secretion.

BACKGROUND: Structural birth defects occur in approximately 3% of live births; most such defects lack defined genetic or environmental causes. Despite advances in surgical approaches, pharmacologic prevention remains largely out of reach. METHODS: We queried worldwide databases of 20,248 families that included children with neurodevelopmental disorders and that were enriched for parental consanguinity. Approximately one third of affected children in these families presented with structural birth defects or microcephaly. We performed exome or genome sequencing of samples obtained from the children, their parents, or both to identify genes with biallelic pathogenic or likely pathogenic mutations present in more than one family. After identifying disease-causing variants, we generated two mouse models, each with a pathogenic variant "knocked in," to study mechanisms and test candidate treatments. We administered a small-molecule Wnt agonist to pregnant animals and assessed their offspring. RESULTS: We identified homozygous mutations in WLS, which encodes the Wnt ligand secretion mediator (also known as Wntless or WLS) in 10 affected persons from 5 unrelated families. (The Wnt ligand secretion mediator is essential for the secretion of all Wnt proteins.) Patients had multiorgan defects, including microcephaly and facial dysmorphism as well as foot syndactyly, renal agenesis, alopecia, iris coloboma, and heart defects. The mutations affected WLS protein stability and Wnt signaling. Knock-in mice showed tissue and cell vulnerability consistent with Wnt-signaling intensity and individual and collective functions of Wnts in embryogenesis. Administration of a pharmacologic Wnt agonist partially restored embryonic development. CONCLUSIONS: Genetic variations affecting a central Wnt regulator caused syndromic structural birth defects. Results from mouse models suggest that what we have named Zaki syndrome is a potentially preventable disorder. (Funded by the National Institutes of Health and others.).

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.

Lysosomal storage disorders: Novel and frequent pathogenic variants in a large cohort of Indian patients of Pompe, Fabry, Gaucher and Hurler disease.

OBJECTIVES: Diagnosis of lysosomal storage disorders (LSDs) remains challenging due to wide clinical, biochemical and molecular heterogeneity. The study applies a combined biochemical and genetic approach to diagnose symptomatic Indian patients of Pompe, Fabry, Gaucher and Hurler disease to generate a comprehensive dataset of pathogenic variants for these disorders. DESIGN & METHODS: Symptomatic patients were biochemically diagnosed by fluorometric methods and molecular confirmation was carried out by gene sequencing. Genetic variants were analyzed according to the ACMG/AMP 2015 variant interpretation guidelines. RESULTS: Amongst the 2181 suspected patients, 285 (13%) were biochemically diagnosed. Of these, 22.5% (64/285) diagnosed with Pompe disease harboured c.1933G>A, c.1A>G, c.1927G>A and c.2783G>C as common and 10 novel pathogenic variants while 7.4% (21/285) patients diagnosed with Fabry disease carried c.851T>C, c.902G>A, c.905A>C and c.1212_1234del as frequent disease-causing variants along with 7 novel pathogenic variants. As many as 48.4% (138/285) patients were diagnosed with Gaucher disease and had c.1448T>C as the most common pathogenic variant followed by c.1342G>C and c.754T>C with 7 previously unreported disease-causing variants and in the 21.7% (62/285) diagnosed cases of Hurler disease, c.1469T>C, c.754delC c.568_581del and c.1898C>T were identified as the most common causative variants along with 21 novel pathogenic variants. CONCLUSION: This comprehensive data set of disease-causing frequent and novel pathogenic variants reported for the first time in such a large patient cohort for each of these four LSDs from the Indian sub-continent, along with their biochemical and clinical spectrum will contribute towards providing definitive diagnosis and treatment, identifying carrier status, as well as in counselling prenatal cases to reduce the morbidity and mortality associated with these disorders.

Spectrum of mutations in genes associated with familial colorectal cancer syndrome (MLH1, MSH2, PMS2, MSH6, and APC): A not so common hereditary cancer syndrome in Indian population.

Incidence of colorectal cancer (CRC) is lower in India than in other parts of the world. Approximately 5% to 10% of CRC is inherited. Hereditary non-polyposis colorectal cancer (HNPCC) syndrome and familial adenomatous polyposis (FAP) syndrome are the two known familial cancer syndromes of gastrointestinal tract, which occur due to inherited genetic predisposition. Not much is known about the molecular profile of families with inherited CRC syndromes seen in Indian population. At our institute, we have been providing genetic testing and counseling service to all the families referred to us with suspicion of inherited cancer predisposition syndrome. We analyzed 36 suspected families at our clinic. Personal and family history of cancer was obtained from the proband and appropriate genetic testing was performed in 19 patients (13 with HNPCC, 5 with FAP, and 1 with Cowden syndrome). We present here our experience and spectrum of pathogenic variants observed in this patient cohort and review on published studies describing molecular profile of Indian patients with CRC syndromes.

Expanding the phenotypic and genotypic spectrum of Wiedemann-Steiner syndrome: First patient from India.

Wiedemann-Steiner syndrome (WWS) is a rare disorder characterized by hypotonia, postnatal growth restriction, striking facial dysmorphism, and hirsutism. It is caused by heterozygous pathogenic variants in KMT2A. This gene has an established role in histone methylation, which explains the overlap of WWS with syndromes caused by genes involved in chromatin remodeling. We describe an infant with a novel single base pair deletion in KMT2A with features consistent with WWS, as well as additional features of stenosis of aqueduct of Sylvius and broad toes. The usefulness of Face2Gene as a tool for identification of dysmorphology syndromes is discussed, as in this patient, it suggested WWS as the top candidate disorder. To the best of our knowledge, this is the first patient of WWS reported from India, with a novel genotype and expanded phenotype.

Genetic analysis of familial hypercholesterolemia in Asian Indians: A single-center study.

BACKGROUND: Familial hypercholesterolemia (FH), an autosomal codominant disorder characterized by very high low-density lipoprotein cholesterol, is strongly associated with premature coronary artery disease. OBJECTIVES: Molecular landscape of FH in Asian Indians is not well studied, although this ethnic group comprises a large proportion of the world population. Knowledge of mutations in these groups is useful for identifying persons affected with FH, saving their lives, and cascade screening in their relatives. METHODS: Potential cases of FH (n = 100) were identified by criteria adapted for the Indian population from Dutch Lipid Clinic Network criteria. Pathogenic variants were analyzed in LDLR, APOB 100 (exons 26 and 29), PCSK9, and APOE genes using Sanger sequencing and multiplex ligation-dependent probe amplification technique. Cases in whom there were no pathogenic variants were tested by next-generation sequencing using a targeted panel of genes. RESULTS: Thirty-eight pathogenic variants were identified in 47 of 100 unrelated probands. Of these variants, 33 were identified in LDLR, 3 in APOB, and 2 in PCSK9 genes. Ten pathogenic variants were novel. Mutations were detected in 91.4% of those subjects classified as definite, 40% as probable, and in 18.8% as possible FH cases based on modified Dutch Lipid Clinic Network criteria. A likely founder mutation in intron 10 (c.1587-1G>A) of LDLR gene was observed in 6 North Indian families. The conventional pathogenic variants in APOB and PCSK9 genes and those previously reported in LDLR gene among Asian Indians were not detected in this cohort. CONCLUSION: This study demonstrates genetic heterogeneity of FH in India. The variants observed were different from those described in Western populations. Next-generation sequencing technology helped identify new mutations in APOB gene, suggesting that in less-studied populations, it is better to sequence the whole gene rather than test for specific mutations.

Co-inheritance of pathogenic variants in PKD1 and PKD2 genes presenting as severe antenatal phenotype of autosomal dominant polycystic kidney disease.

Autosomal dominant polycystic kidney disease (ADPKD) is caused by pathogenic variants in either PKD1 or PKD2 genes. Disease severity is dependent on various factors including the presence of modifier genes. We describe a family with recurrent foetal presentation of ADPKD due to co-inheritance of pathogenic variants in both PKD1 [c.3860T > C; p.(Leu1287Pro)] and PKD2 [(c.1000C > A; p.(Pro334Thr)] genes. Familial segregation studies revealed the mother and the father to be heterozygous for the same variants in the PKD1 and PKD2 genes, respectively, as found in the foetus. Renal ultrasonography detected evidence of cystic disease in the mother and two of her family members. No cysts were detected in the father, however the paternal grandfather died of renal cystic disease. The absence of disease in the father can be explained by the phenomenon of incomplete penetrance, or Knudson's two-hit hypothesis of cystogenesis in the grandfather. This case underscores the importance of sequencing PKD2 gene even in the presence of a familial PKD1 variant, as well as genetic testing of the cysts for evidence of the second hit.

Indian National Association for the Study of the Liver-Federation of Obstetric and Gynaecological Societies of India Position Statement on Management of Liver Diseases in Pregnancy.

Liver diseases occurring during pregnancy can be serious and can progress rapidly, affecting outcomes for both the mother and fetus. They are a common cause of concern to an obstetrician and an important reason for referral to a hepatologist, gastroenterologist, or physician. Liver diseases during pregnancy can be divided into disorders unique to pregnancy, those coincidental with pregnancy, and preexisting liver diseases exacerbated by pregnancy. A rapid differential diagnosis between liver diseases related or unrelated to pregnancy is required so that specialist and urgent management of these conditions can be carried out. Specific Indian guidelines for the management of these patients are lacking. The Indian National Association for the Study of the Liver (INASL) in association with the Federation of Obstetric and Gynaecological Societies of India (FOGSI) had set up a taskforce for development of consensus guidelines for management of patients with liver diseases during pregnancy, relevant to India. For development of these guidelines, a two-day roundtable meeting was held on 26-27 May 2018 in New Delhi, to discuss, debate, and finalize the consensus statements. Only those statements that were unanimously approved by most members of the taskforce were accepted. The primary objective of this review is to present the consensus statements approved jointly by the INASL and FOGSI for diagnosing and managing pregnant women with liver diseases. This article provides an overview of liver diseases occurring in pregnancy, an update on the key mechanisms involved in its pathogenesis, and the recommended treatment options.

Multi-gene testing in neurological disorders showed an improved diagnostic yield: data from over 1000 Indian patients.

BACKGROUND: Neurological disorders are clinically heterogeneous group of disorders and are major causes of disability and death. Several of these disorders are caused due to genetic aberration. A precise and confirmatory diagnosis in the patients in a timely manner is essential for appropriate therapeutic and management strategies. Due to the complexity of the clinical presentations across various neurological disorders, arriving at an accurate diagnosis remains a challenge. METHODS: We sequenced 1012 unrelated patients from India with suspected neurological disorders, using TruSight One panel. Genetic variations were identified using the Strand NGS software and interpreted using the StrandOmics platform. RESULTS: We were able to detect mutations in 197 genes in 405 (40%) cases and 178 mutations were novel. The highest diagnostic rate was observed among patients with muscular dystrophy (64%) followed by leukodystrophy and ataxia (43%, each). In our cohort, 26% of the patients who received definitive diagnosis were primarily referred with complex neurological phenotypes with no suggestive diagnosis. In terms of mutations types, 62.8% were truncating and in addition, 13.4% were structural variants, which are also likely to cause loss of function. CONCLUSION: In our study, we observed an improved performance of multi-gene panel testing, with an overall diagnostic yield of 40%. Furthermore, we show that NGS (next-generation sequencing)-based testing is comprehensive and can detect all types of variants including structural variants. It can be considered as a single-platform genetic test for neurological disorders that can provide a swift and definitive diagnosis in a cost-effective manner.

Clinical and molecular characterization of Indian patients with fructose-1, 6-bisphosphatase deficiency: Identification of a frequent variant (E281K).

Fructose-1, 6-bisphosphatase deficiency is an autosomal recessive disorder of gluconeogenesis caused by genetic defect in the FBP1 gene. It is characterized by episodic, often life-threatening metabolic acidosis, liver dysfunction, and hyperlactatemia. Without a high index of suspicion, it may remain undiagnosed with devastating consequences. Accurate diagnosis can be achieved either by enzyme assay or gene studies. Enzyme assay requires a liver biopsy and is tedious, invasive, expensive, and not easily available. Therefore, genetic testing is the most appropriate method to confirm the diagnosis. Molecular studies were performed on 18 suspected cases presenting with episodic symptoms. Seven different pathogenic variants were identified. Two common variants were noted in two subpopulations from the Indian subcontinent; p.Glu281Lys (E281K) occurred most frequently (in 10 patients) followed by p.Arg158Trp (R158W, in 4 patients). Molecular analysis confirmed the diagnosis and helped in managing these patients by providing appropriate genetic counseling. In conclusion, genetic studies identified two common variants in the Indian subcontinent, thus simplifying the diagnostic algorithm in this treatable disorder.

Expanding the Phenotype of the Founder South Asian Mutation in the Nuclear Encoding Mitochondrial RMND1 Gene.

BACKGROUND: Mitochondrial disorders have a wide variability in the phenotype. A 10-mo-old girl presented with a severe phenotype of multisystem involvement due to an uncommon mitochondrial disease. Mutations in the RMND1 gene of nuclear DNA were identified on next generation sequencing. This mutation results in combined oxidative phosphorylation deficiency -11 (OMIM #614922) of the respiratory chain complex. So far in South Asia, patients of this disorder have been reported only from Pakistan and Bangladesh. RESULTS: In addition to the features reported in other patients of South Asia with the same mutation at c.1349G>C, index patient from India had hyperaldosteronism, long QT interval but no deafness. CONCLUSIONS: Thus, to conclude, this report emphasizes the diagnostic value of FGF21 assay in this disorder. It extends the phenotype associated with the founder mutation in RMND1 gene in patients from South Asia.

Prenatal screening for genetic disorders: Suggested guidelines for the Indian Scenario.

Prenatal testing is the best strategy for reducing the burden of genetic disorders and congenital disabilities that cause significant postnatal functional impairment. Universal prenatal screening is advisable for common genetic disorders and congenital anomalies such as Down syndrome, beta-thalassaemia and neural tube defects. Several prenatal-screening tests are now available for Down syndrome, but knowledge about the appropriate timing of the test and the need for pre- and post-test counselling may not be updated among the primary care physicians. There is also a considerable degree of confusion regarding the prenatal screening test to be chosen in each case, due to the availability of a number of new and advanced screening techniques. At present, there is no nation-wide consensus regarding the nature and timing of these prenatal-screening protocols. Due to the absence of any definite guidelines and the additional lacunae in the awareness regarding the appropriate prenatal screening in the country, the optimum benefits of these screening protocols are not reaching the population. This review focuses on the various prenatal screening and diagnostic tests that are available for common genetic conditions and congenital disabilities and attempts to outline the most cost-effective and gestational age-appropriate strategies for prenatal screening for the Indian healthcare set-up. The recommendations suggested would serve as a source guide for formulating prenatal-screening guidelines for reducing the incidence of common genetic disorders and congenital disabilities in India.

Inherited Metabolic Disorders: Efficacy of Enzyme Assays on Dried Blood Spots for the Diagnosis of Lysosomal Storage Disorders.

High consanguinity rates, poor access to accurate diagnostic tests, and costly therapies are the main causes of increased burden of lysosomal storage disorders (LSDs) in developing countries. Therefore, there is a major unmet need for accurate and economical diagnostic tests to facilitate diagnosis and consideration of therapies before irreversible complications occur. In cross-country study, we utilized dried blood spots (DBS) of 1,033 patients clinically suspected to harbor LSDs for enzymatic diagnosis using modified fluorometric assays from March 2013 through May 2015. Results were validated by demonstrating reproducibility, testing in different sample types (leukocytes/plasma/skin fibroblast), mutation study, or measuring specific biomarkers. Thirty percent (307/1,033) were confirmed to have one of the LSDs tested. Reference intervals established unambiguously identified affected patients. Correlation of DBS results with other biological samples (n = 172) and mutation studies (n = 74) demonstrated 100% concordance in Gaucher, Fabry, Tay Sachs, Sandhoff, Niemann-Pick, GM1, Neuronal ceroid lipofuscinosis (NCL), Fucosidosis, Mannosidosis, Mucopolysaccharidosis (MPS) II, IIIb, IVa, VI, VII, and I-Cell diseases, and 91.4% and 88% concordance in Pompe and MPS-I, respectively. Gaucher and Pompe are the most common LSDs in India and Pakistan, followed by MPS-I in both India and Sri Lanka. Study demonstrates utility of DBS for reliable diagnosis of LSDs. Diagnostic accuracy (97.6%) confirms veracity of enzyme assays. Adoption of DBS will overcome significant hurdles in blood sample transportation from remote regions. DBS enzymatic and molecular diagnosis should become the standard of care for LSDs to make timely diagnosis, develop personalized treatment/monitoring plan, and facilitate genetic counseling.

Infantile Systemic Hyalinosis: Novel Founder Mutation in the Initiation Codon among "Malis (Farmers)" in Jodhpur.

Infantile systemic hyalinosis (OMIM 236490) is a progressive autosomal recessive disorder characterized by widespread deposition of hyaline material in many tissues leading to multiple subcutaneous skin nodules, gingival hypertrophy and joint contractures. The authors describe five children from four unrelated families, from the "mali (farmer)" community in Jodhpur, with the disorder. All of them had classical clinical features, and four died from severe infections between age of 7 mo to 3 y. Two affected children had the same, but novel mutation in the initiation codon, in homozygous form c.1 A > G; p. M1? in capillary morphogenesis protein-2 (CMG2), or ANTXR2 gene on chromosome 4q21.21. The other two parents had the same mutation in heterozygous form. It is likely that this is a founder mutation in this community.

MoM cutoffs for variables, an important tool for multivariate analysis and accurate interpretation of preeclampsia risk in high-risk pregnancy at 11-13+6 weeks gestation.

OBJECTIVE: To determine the diagnostic accuracy of preeclampsia (PE) screening test offered in early pregnancy for the prediction of the risks for early-onset (requiring delivery <32 weeks gestation) and late-onset (requiring delivery ≥32 weeks gestation) disease. METHODS: In a retrospective study of 615 women with singleton pregnancy, the risk for PE was calculated by the combined effect of multiple variables: serum placental growth factor (PLGF) and pregnancy-associated plasma protein-A (PAPP-A), maternal age, parity, ethnicity, mean arterial pressure (MAP), body mass index (BMI), uterine artery-pulsatility index, and previous history of PE or hypertension (HT). The results of the screening test in three different groups of women were validated by pregnancy outcome: (i) control group - without any history of PE/HT; (ii) history of PE without HT; and (iii) history of HT without PE. The performance of the screening test was evaluated for early- and late-onset PE. RESULTS: The multivariate screening effectively identified cases of PE with >97% specificity. The detection rate (DR) was 93.8% for late-onset PE at a false positive rate (FPR) of 2.3% and 44.4% for early-onset PE at an FPR of 0.0%. The incidence of PE was 7% overall, with 1.52% and 5.43% for early- and late-onset PE, respectively. CONCLUSION: The study demonstrated 96.6% diagnostic accuracy of the multi-variable screening test to predict the risk of PE in the first trimester. The negative predictive value (>98%) reinforces the utility of cost-effective noninvasive screening test for the early detection of PE. ABBREVIATIONS: PLGF: Placental growth factor; PAPP-A: Pregnancy-associated plasma protein-A; free ß-HCG: Free beta-human chorionic gonadotropin; BMI: Body mass index; MAP: Mean arterial blood pressure; Ut-PI: Mean uterine artery pressure (left and right uterine artery)-pulsatility index; MoM: Multiple of median; NICE: National Institute for Health and Clinical Excellence.

Phenotype guided characterization and molecular analysis of Indian patients with long QT syndromes.

BACKGROUND: Long QT syndromes (LQTS) are characterized by prolonged QTc interval on electrocardiogram (ECG) and manifest with syncope, seizures or sudden cardiac death. Long QT 1-3 constitute about 75% of all inherited LQTS. We classified a cohort of Indian patients for the common LQTS based on T wave morphology and triggering factors to prioritize the gene to be tested. We sought to identify the causative mutations and mutation spectrum, perform genotype-phenotype correlation and screen family members. METHODS: Thirty patients who fulfilled the criteria were enrolled. The most probable candidate gene among KCNQ1, KCNH2 and SCN5A were sequenced. RESULTS: Of the 30 patients, 22 were classified at LQT1, two as LQT2 and six as LQT3. Mutations in KCNQ1 were identified in 17 (77%) of 22 LQT1 patients, KCNH2 mutation in one of two LQT2 and SCN5A mutations in two of six LQT3 patients. We correlated the presence of the specific ECG morphology in all mutation positive cases. Eight mutations in KCNQ1 and one in SCN5A were novel and predicted to be pathogenic by in-silico analysis. Of all parents with heterozygous mutations, 24 (92%) of 26 were asymptomatic. Ten available siblings of nine probands were screened and three were homozygous and symptomatic, five heterozygous and asymptomatic. CONCLUSIONS: This study in a cohort of Asian Indian patients highlights the mutation spectrum of common Long QT syndromes. The clinical utility for prevention of unexplained sudden cardiac deaths is an important sequel to identification of the mutation in at-risk family members.

KCNQ1 mutations associated with Jervell and Lange-Nielsen syndrome and autosomal recessive Romano-Ward syndrome in India-expanding the spectrum of long QT syndrome type 1.

Long QT syndrome type 1 (LQT1) is the most common type of all Long QT syndromes (LQTS) and occurs due to mutations in KCNQ1. Biallelic mutations with deafness is called Jervell and Lange-Nielsen syndrome (JLNS) and without deafness is autosomal recessive Romano-Ward syndrome (AR RWS). In this prospective study, we report biallelic mutations in KCNQ1 in Indian patients with LQT1 syndrome. Forty patients with a clinical diagnosis of LQT1 syndrome were referred for molecular testing. Of these, 18 were excluded from the analysis as they did not fulfill the inclusion criteria of broad T wave ECG pattern of the study. Direct sequencing of KCNQ1 was performed in 22 unrelated probands, parents and at-risk family members. Mutations were identified in 17 patients, of which seven had heterozygous mutations and were excluded in this analysis. Biallelic mutations were identified in 10 patients. Five of 10 patients did not have deafness and were categorized as AR RWS, the rest being JLNS. Eight mutations identified in this study have not been reported in the literature and predicted to be pathogenic by in silico analysis. We hypothesize that the homozygous biallelic mutations identified in 67% of families was due to endogamous marriages in the absence of consanguinity. This study presents biallelic gene mutations in KCNQ1 in Asian Indian patients with AR JLNS and RWS. It adds to the scant worldwide literature of mutation studies in AR RWS. © 2016 Wiley Periodicals, Inc.