Heterozygous NM Research Articles

A Rare Case of TP63-Associated Lymphopenia Revealed by Newborn Screening Using TREC

The expanded newborn screening (NBS) program in the Russian Federation was initiated in 2023, among which severe combined immunodeficiency (SCID) is screened using TREC/KREC assays. Here, we report a rare case of a TP63-associated disease identified through this NBS program. Dried blood spots from newborns were initially screened for TREC/KREC levels, and those with values below the cut-off underwent confirmatory testing and further genetic analysis, including whole-exome sequencing (WES). A male newborn was identified with significantly reduced TREC values, indicative of T cell lymphopenia. Genetic analysis revealed a heterozygous NM_003722.5:c.1027C>T variant in TP63, leading to the p.(Arg343Trp) substitution within the DNA binding domain. This mutation has been previously associated with Ectrodactyly–Ectodermal Dysplasia–Cleft lip/palate syndrome (EEC) syndrome and shown to reduce the transactivation activity of TP63 in a dominant-negative manner. This case represents one of the few instances of immune system involvement in a patient with a TP63 mutation, highlighting the need for further investigation into the immunological aspects of TP63-associated disorders. Our findings suggest that comprehensive immunological evaluation should be considered for patients with TP63 mutations to better understand and manage potential immune dysfunctions.

Integration of validated functional evidence to support the pathogenicity of KCNH2 variants

Functional investigation of genetic variants found in long QT syndrome can provide evidence that is needed to confirm the genetic diagnosis and establish the cause of the condition. We performed functional assessment to determine the z-score, using a clinically calibrated automated patch clamp assay, for 2 missense KCNH2 variants found in 2 families that have been diagnosed with long QT syndrome. These variants are currently classified as variant of uncertain significance in ClinVar. The z-scores for homozygous and heterozygous NM_000238.4:c.1819A>T p.(Ile607Phe) from family 1 were –5.16 and –3.97, respectively, and for heterozygous NM_000238.4:c.1832A>G p.(Tyr611Cys) from family 2 was –6.63. The z-scores for these variants are consistent with severe loss-of-function phenotypes. We have established the genetic cause of the long QT syndrome in these 2 families by providing validated functional evidence that supports the pathogenicity of p.(Ile607Phe) and p.(Tyr611Cys). Clinical diagnosis of long QT syndrome has been very successful in providing adequate clinical management for patients. However, functional assessment of variants found in these patients by using variant-specific z-scores can further enhance the current clinical management of patients with long QT syndrome through shared decision making based on validated experimental evidence.

Coexistence of Neonatal Bartter Syndrome and Congenital Cutis Laxa, in which a new mutation in SLC12A1 was identified

Introduction: Neonatal Bartter Syndrome (NBS) is an extremely rare congenital chronic renal tubular transport disorder characterized by preterm birth, polyhydramnios, polyuria, salt wasting, and severe dehydration. Congenital Cutis Laxa (CCL) is a rare disease characterized by loss of skin flexibility and laxity, and atypical facial appearance.
 Case Presentation: A 1290-gram female baby at 30+6 weeks of gestation was hospitalized due to premature birth and respiratory distress. During physical examination, an atypical facial appearance, wrinkles on the forehead, skin laxity, and philtrum were visible as well as a small chin and arachnodactyly in the fingers and toes. The patient displayed metabolic alkalosis in her blood gases. The patient, with an atypical face and current findings submitted her exome for genetic diagnosis. Genetic analysis revealed the presence of homozygous NM_001184832.2:c.2485+5G>A (rs749269268) in SLC12A1, and NBS Type 1 was diagnosed. A novel heterozygous NM_000501.4:c.352G>T, p.Val118Phe (rs781922544) variant was detected in ELN, and CCL syndrome with autosomal dominant inheritance was diagnosed.
 Conclusions: The rare co-occurrence of NBS and CCL syndrome in our patient was seen as a mere coincidence. Since this is the first reported case in the literature, it was deemed appropriate to present this case.

Generation of an induced pluripotent stem cell line XJHi001-A from a fibronectin glomerulopathy patient carrying a heterozygous NM_212482.2(C.5888-1G > C) mutation in the FN1 gene

Fibronectin glomerulopathy (FNG) is an autosomal dominant disease, and nearly half of patients are involved in a mutation of the fibronectin gene (FN1). By using electrotransformation, we generated an induced pluripotent stem cell (iPSC) line from peripheral blood mononuclear cells (PBMCs) of a 32-years-old female patient who carried a heterozygous NM_212482.2(C.5888-1G > C) mutation in the FN1 gene. This cellular model is a useful tool to investigate pathogenic mechanisms in FNG and to develop potential treatments for FNG.

Curious cases of GATA2 deficiency: clonal evolution or dual diagnoses?

GATA2 deficiency can often predispose individuals to hematologic malignancies. While many pathogenic/likely pathogenic germline variants have been characterized in GATA2 deficiency, less is known about other variants that arise during clonal evolution of hematologic malignancies or as potential dual diagnoses in these patients. Here, we present two cases enrolled in the NIAID Centralized Sequencing Program with GATA2 variants and additional pathogenic/likely pathogenic variants in WT1 and KRAS detected via genome sequencing.The first case is a 9-year-old female with myelodysplastic syndrome with monosomy 7, pulmonary alveolar proteinosis, and sepsis. Genome sequencing in the proband detected 3 rare variants in GATA2: one pathogenic heterozygous NM_032638.5:c.202del (p.Ala68ArgfsTer12) frameshift variant, one heterozygous c.1031_1087dup (p. Arg344_Arg362dup) duplication variant of uncertain significance (VUS), and one apparently somatic mosaic c.1306_1307insTCCA p. (His436LeufsTer101) frameshift VUS. Two variants were also detected in WT1: a likely pathogenic, apparently somatic mosaic, NM_024426.6: c.1159_1163del (p.Ala387Ter) nonsense variant and a heterozygous c.887G>A (p.Ser296Asn) missense VUS. The WT1 p.Ser296Asn variant was previously published in a female affected with Wilms tumor who inherited the variant from her unaffected father, however our participant does not have a personal history of Wilms tumor.The second case is a 6-year-old male diagnosed with acute myeloid leukemia (AML). Genome sequencing detected a heterozygous GATA2 NM_032638.5(GATA2):c.575C>T (p.Ser192Phe) missense VUS that has not been previously published in individuals with GATA2 deficiency. Additionally, a pathogenic, apparently somatic mosaic, NM_004985.5: c.173C>T (p.Thr58Ile) missense variant in KRAS was detected, which has been reported in the literature as a germline variant in individuals with Noonan syndrome and cardiofaciocutaneous syndrome. The variant allele fraction of the KRAS p.Thr58Ile variant was 27%, however it is unknown if this variant could be somatic mosaic in association with the subject's AML clone, or if the subject has a genetic diagnosis of RAS-associated autoimmune lymphoproliferative syndrome (RALD). Future studies include segregation analysis in family members.These cases highlight the importance of additional variants found in patients with GATA2 variants that may be contributing to dual diagnoses or acting as secondary mutations via clonal evolution of myeloid malignancies.

PIGN-Related Disease in Two Lithuanian Families: A Report of Two Novel Pathogenic Variants, Molecular and Clinical Characterisation.

Background and Objectives: Pathogenic variants of PIGN are a known cause of multiple congenital anomalies-hypotonia-seizures syndrome 1 (MCAHS1). Many affected individuals have clinical features overlapping with Fryns syndrome and are mainly characterised by developmental delay, congenital anomalies, hypotonia, seizures, and specific minor facial anomalies. This study investigates the clinical and molecular data of three individuals from two unrelated families, the clinical features of which were consistent with a diagnosis of MCAHS1. Materials and Methods: Next-generation sequencing (NGS) technology was used to identify the changes in the DNA sequence. Sanger sequencing of gDNA of probands and their parents was used for validation and segregation analysis. Bioinformatics tools were used to investigate the consequences of pathogenic or likely pathogenic PIGN variants at the protein sequence and structure level. Results: The analysis of NGS data and segregation analysis revealed a compound heterozygous NM_176787.5:c.[1942G>T];[1247_1251del] PIGN genotype in family 1 and NG_033144.1(NM_176787.5):c.[932T>G];[1674+1G>C] PIGN genotype in family 2. In silico, c.1942G>T (p.(Glu648Ter)), c.1247_1251del (p.(Glu416GlyfsTer22)), and c.1674+1G>C (p.(Glu525AspfsTer68)) variants are predicted to result in a premature termination codon that leads to truncated and functionally disrupted protein causing the phenotype of MCAHS1 in the affected individuals. Conclusions: PIGN-related disease represents a wide spectrum of phenotypic features, making clinical diagnosis inaccurate and complicated. The genetic testing of every individual with this phenotype provides new insights into the origin and development of the disease.

Clinical characteristics and proteome modifications in two Charcot-Marie-Tooth families with the AARS1 Arg326Trp mutation

BackgroundAminoacyl tRNA-synthetases are ubiquitously-expressed enzymes that attach amino acids to their cognate tRNA molecules. Mutations in several genes encoding aminoacyl tRNA-synthetases, have been associated with peripheral neuropathy, i.e. AARS1, GARS1, HARS1, YARS1 and WARS1. The pathogenic mechanism underlying AARS1-related neuropathy is not known.MethodsFrom 2012 onward, all probands presenting at Telemark Hospital (Skien, Norway) with peripheral neuropathy were screened for variants in AARS1 using an “in-house” next-generation sequencing panel. DNA from patient’s family members was examined by Sanger sequencing. Blood from affected family members and healthy controls were used for quantification of AARS1 mRNA and alanine. Proteomic analyses were conducted in peripheral blood mononuclear cells (PBMC) from four affected family members and five healthy controls.ResultsSeventeen individuals in two Norwegian families affected by Charcot-Marie-Tooth disease (CMT) were characterized in this study. The heterozygous NM_001605.2:c.976C > T p.(Arg326Trp) AARS1 mutation was identified in ten affected family members. All living carriers had a mild to severe length-dependent sensorimotor neuropathy. Three deceased obligate carriers aged 74–98 were reported to be unaffected, but were not examined in the clinic. Proteomic studies in PBMC from four affected individuals suggest an effect on the immune system mediated by components of a systemic response to chronic injury and inflammation. Furthermore, altered expression of proteins linked to mitochondrial function/dysfunction was observed. Proteomic data are available via ProteomeXchange using identifier PXD023842.ConclusionThis study describes clinical and neurophysiological features linked to the p.(Arg326Trp) variant of AARS1 in CMT-affected members of two Norwegian families. Proteomic analyses based on of PBMC from four CMT-affected individuals suggest that involvement of inflammation and mitochondrial dysfunction might contribute to AARS1 variant-associated peripheral neuropathy.

Compound heterozygous c.598_612del and c.1746-20C\u2009>\u2009G CAPN3 genotype cause autosomal recessive limb-girdle muscular dystrophy-1: a case report

BackgroundAutosomal recessive limb–girdle muscular dystrophy-1 (LGMDR1), also known as calpainopathy, is a genetically heterogeneous disorder characterised by progression of muscle weakness. Homozygous or compound heterozygous variants in the CAPN3 gene are known genetic causes of this condition. The aim of this study was to confirm the molecular consequences of the CAPN3 variant NG_008660.1(NM_000070.3):c.1746-20C > G of an individual with suspected LGMDR1 by extensive complementary DNA (cDNA) analysis.Case presentationIn the present study, we report on a male with proximal muscular weakness in his lower limbs. Compound heterozygous NM_000070.3:c.598_612del and NG_008660.1(NM_000070.3):c.1746-20C > G genotype was detected on the CAPN3 gene by targeted next-generation sequencing (NGS). To confirm the pathogenicity of the variant c.1746-20C > G, we conducted genetic analysis based on Sanger sequencing of the proband’s cDNA sample. The results revealed that this splicing variant disrupts the original 3′ splice site on intron 13, thus leading to the skipping of the DNA fragment involving exon 14 and possibly exon 15. However, the lack of exon 15 in the CAPN3 isoforms present in a blood sample was explained by cell-specific alternative splicing rather than an aberrant splicing mechanism. In silico the c.1746-20C > G splicing variant consequently resulted in frameshift and formation of a premature termination codon (NP_000061.1:p.(Glu582Aspfs*62)).ConclusionsBased on the results of our study and the literature we reviewed, both c.598_612del and c.1746-20C > G variants are pathogenic and together cause LGMDR1. Therefore, extensive mRNA and/or cDNA analysis of splicing variants is critical to understand the pathogenesis of the disease.

A heterozygous mutation in the CCDC88C gene likely causes early-onset pure hereditary spastic paraplegia: a case report

BackgroundCCDC88C is a ubiquitously expressed protein with multiple functions, including roles in cell polarity and the development of dendrites in the nervous system. Bi-allelic mutations in the CCDC88C gene cause autosomal recessive congenital hydrocephalus (OMIM #236600). Studies recently linked heterozygous mutations in CCDC88C to the development of the late-onset spinocerebellar ataxia type 40 (OMIM #616053).Case presentationA 48-year-old Sudanese female presented with pure early onset hereditary spastic paraplegia. Exome sequencing, in-silico analysis, and Sanger sequencing identified the heterozygous NM_001080414.4:c.1993G > A (p.E665K) variant in CCDC88C as a potential cause of her illness. To explore the pathogenicity of the NM_001080414.4:c.1993G > A (p.E665K) variant, we expressed it in human embryonic kidney 293 cells and assessed its effects on apoptosis. In our experiment, NM_001080414.4:c.1993G > A (p.E665K) induced JNK hyper-phosphorylation and enhanced apoptosis. In contrast to previous reports, our patient developed neurological symptoms in early childhood and showed neither features of cerebellar ataxia, extrapyramidal signs, nor evidence of intellectual involvement.ConclusionWe, herein, heighlighted the possibility of extending the phenotype associated with variants in CCDC88C to include early-onset pure hereditary spastic paraplegia.

Complement factor D haplodeficiency is associated with a reduced complement activation speed and diminished bacterial killing.

ObjectivesComplete deficiency of alternative pathway (AP) complement factors, explained by homozygous mutations, is a well‐known risk factor for invasive bacterial infections; however, this is less obvious for heterozygous mutations. We describe two siblings with a heterozygous NM_001928.3(CFD):c.125C>A p.(Ser42*) mutation in the complement factor D (fD) gene having a history of recurrent bacterial infections. We determined the effect of heterozygous fD deficiency on AP complement activity.MethodsWe determined the effect of fD levels on complement activation as measured by AP activity, complement C3 binding to the bacterial surface of Neisseria meningitidis (Nm), Streptococcus pneumoniae (Sp) and non‐typeable Haemophilus influenzae (NTHi), and complement‐mediated killing of Nm and NTHi. In addition, we measured the effect of vaccination of complement C3 binding to the bacterial surface and killing of Nm.ResultsReconstitution of fD‐deficient serum with fD increased AP activity in a dose‐ and time‐dependent way. Reconstitution of patient serum with fD to normal levels increased complement C3 binding to Sp, Nm and NTHi, as well as complement‐mediated killing of Nm and NTHi. Vaccination increased complement C3 binding and resulted in complete killing of Nm without fD reconstitution.ConclusionWe conclude that low fD serum levels (< 0.5 μg mL−1) lead to a reduced speed of complement activation, which results in diminished bacterial killing, consistent with recurrent bacterial infections observed in our index patients. Specific antibodies induced by vaccination are able to overcome the diminished bacterial killing capacity in patients with low fD levels.

Extending the phenotype of Xia-Gibbs syndrome in a two-year-old patient with craniosynostosis with a novel de novo AHDC1 missense mutation

Xia-Gibbs syndrome (Mental retardation, autosomal dominant 25; MRD25) [MIM 615829] is a rare autosomal dominant disease characterized by mental retardation, developmental delay, speech delay, structural brain anomalies, hypotonicity, protuberant eyes, visual problems, laryngomalacia and snoring. Since the first description in 2014, fewer than 50 patients with Xia-Gibbs syndrome have been noticed in the literature. We describe here 2 years 2 months old girl with developmental delay, brain anomalies, laryngomalacia and craniosynostosis. Whole-Exome Sequencing (WES) analyses in patient showed a heterozygous NM_001029882: c.4370 A>G; p. (Asp1457Gly) mutation in AHDC1. Craniosynostosis rarely observed in the patients described to date, and west syndrome-like EEG pattern, constipation and electrolyte imbalance observed for the first time were present in our patient. Further reports and in-vivo/in-vitro works will make possible knowing of the genetic and clinical background of this disease.