DataSheet1_A genotype–phenotype correlation in split-hand/foot malformation type 1: further refinement of the phenotypic subregions within the 7q21.3 locus.PDF

Background: Split-hand/foot malformation type 1 (SHFM1) refers to the group of rare congenital limb disorders defined by the absence or hypoplasia of the central rays of the autopods with or without accompanying anomalies, such as hearing loss, craniofacial malformation, and ectodermal dysplasia. Consequently, the condition is characterized by clinical variability that hinders diagnostic and counseling procedures. SHFM1 is caused by pathogenic variants affecting the DLX5/6 genes and/or their tissue-specific enhancers at the 7q21.3 locus. Herein, we report on seven patients from five unrelated Polish families affected by variable symptoms of the SHFM1 spectrum, all harboring 7q21.3 or 7q21.2-q21.3 rearrangements, and provide a genotype–phenotype correlation in the studied cohort.Methods: We applied GTG banding, array-based comparative genomic hybridization (aCGH), and whole-genome sequencing (WGS) in order to identify the causative aberrations in all affected patients.Results: The identified pathogenic structural variants included deletions and/or translocations involving the 7q21.3 locus, i.e., t(7;10)(q21.3;q22.2) and t(7;12)(q21.3;q21.2) in all affected individuals. Interestingly, a sporadic carrier of the latter aberration presented the SHFM1 phenotype with additional features overlapping with Baker–Gordon syndrome (BAGOS), which resulted from the translocation breakpoint at chromosome 12 within the SYT1 gene.Conclusion: Clinical variability of the studied cohort reflects the composition of the DLX5/6 regulatory elements that were dislocated from their target genes by chromosomal rearrangements. The correlation of our data with the previously published observations enabled us to update the phenotypic subregions and regulatory units within the SHFM1 locus. In addition, we present the first case of SHFM1 and BAGOS-like phenotype that resulted from translocation breakpoints at chromosomes 7 and 12, both of which were pathogenic, and consequently, we show the first evidence that BAGOS can also result from the regulatory loss-of-function SYT1 mutations. In this paper, we emphasize the utility of sequence-based approaches in molecular diagnostics of disorders caused by regulatory structural variants.</p

DataSheet2_A genotype–phenotype correlation in split-hand/foot malformation type 1: further refinement of the phenotypic subregions within the 7q21.3 locus.pdf

Background: Split-hand/foot malformation type 1 (SHFM1) refers to the group of rare congenital limb disorders defined by the absence or hypoplasia of the central rays of the autopods with or without accompanying anomalies, such as hearing loss, craniofacial malformation, and ectodermal dysplasia. Consequently, the condition is characterized by clinical variability that hinders diagnostic and counseling procedures. SHFM1 is caused by pathogenic variants affecting the DLX5/6 genes and/or their tissue-specific enhancers at the 7q21.3 locus. Herein, we report on seven patients from five unrelated Polish families affected by variable symptoms of the SHFM1 spectrum, all harboring 7q21.3 or 7q21.2-q21.3 rearrangements, and provide a genotype–phenotype correlation in the studied cohort.Methods: We applied GTG banding, array-based comparative genomic hybridization (aCGH), and whole-genome sequencing (WGS) in order to identify the causative aberrations in all affected patients.Results: The identified pathogenic structural variants included deletions and/or translocations involving the 7q21.3 locus, i.e., t(7;10)(q21.3;q22.2) and t(7;12)(q21.3;q21.2) in all affected individuals. Interestingly, a sporadic carrier of the latter aberration presented the SHFM1 phenotype with additional features overlapping with Baker–Gordon syndrome (BAGOS), which resulted from the translocation breakpoint at chromosome 12 within the SYT1 gene.Conclusion: Clinical variability of the studied cohort reflects the composition of the DLX5/6 regulatory elements that were dislocated from their target genes by chromosomal rearrangements. The correlation of our data with the previously published observations enabled us to update the phenotypic subregions and regulatory units within the SHFM1 locus. In addition, we present the first case of SHFM1 and BAGOS-like phenotype that resulted from translocation breakpoints at chromosomes 7 and 12, both of which were pathogenic, and consequently, we show the first evidence that BAGOS can also result from the regulatory loss-of-function SYT1 mutations. In this paper, we emphasize the utility of sequence-based approaches in molecular diagnostics of disorders caused by regulatory structural variants.</p

Table2_Case report: The cardio-facio-cutaneous syndrome due to a novel germline mutation in MAP2K1: A multifaceted disease with immunodeficiency and short stature.docx

Cardio-facio-cutaneous syndrome (CFCS) belongs to the group of RASopathies, clinical disorders defined by disruptions in the RAS/MAPK signaling pathway. It is caused by heterozygous gain-of-function germline mutations in genes encoding protein kinases: BRAF, MAP2K1 (MEK1), MAP2K2 (MEK2), and in the GTPase-encoding gene KRAS. CFCS is characterized by craniofacial dysmorphic features, congenital heart defects, severe malnutrition, proportionate short stature, anomalies within the structure of skin and hair, and psychomotor disability. The pathophysiology of growth impairment is multifactorial with feeding difficulties, growth hormone deficiency, and insensitivity. Immunodeficiency has not been hitherto reported as an integral part of CFCS yet an increased activation of the RAS/MAPK signaling pathway may contribute to explaining the causal relationship between RASopathy and the dysfunctions within the B and T lymph cell compartments resulting in a deficiency in T cell costimulation and B cell maturation with impaired class switch recombination, somatic hypermutation, and high-affinity antibody production. We report on a boy born prematurely at 32 WGA, with the perinatal period complicated by pneumonia, respiratory distress syndrome, and valvular pulmonary stenosis. The boy suffered from recurrent pneumonia, obstructive bronchitis, sepsis, urinary tract infection, and recurrent fevers. He presented with severe hypotrophy, psychomotor disability, short stature, craniofacial dysmorphism, dental hypoplasia, sparse hair, and cryptorchidism. Whole genome sequencing showed a novel heterozygous pathogenic germline missense variant: c.364A > G; p.Asn122Asp in the MAP2K1 gene, supporting the diagnosis of CFCS. The immunological workup revealed hypogammaglobulinemia, IgG subclass, and specific antibody deficiency accompanied by decreased numbers of T helper cells and naive and memory B cells. Replacement immunoglobulin therapy with timely antibiotic prophylaxis were instituted. At the age of six years, growth hormone deficiency was diagnosed and the rGH therapy was started. The ever-increasing progress in genetic studies contributes to establishing the definitive CFCS diagnosis and sheds the light on the interrelated genotype-phenotype heterogeneity of RASopathies. Herein, we add new phenotypic features of predominating humoral immunodeficiency to the symptomatology of CFCS with a novel mutation in MAP2K1. While CFCS is a multifaceted disease, increased pediatricians’ awareness is needed to prevent the delay in diagnostics and therapeutic interventions.</p

Table1_Case report: The cardio-facio-cutaneous syndrome due to a novel germline mutation in MAP2K1: A multifaceted disease with immunodeficiency and short stature.docx

Cardio-facio-cutaneous syndrome (CFCS) belongs to the group of RASopathies, clinical disorders defined by disruptions in the RAS/MAPK signaling pathway. It is caused by heterozygous gain-of-function germline mutations in genes encoding protein kinases: BRAF, MAP2K1 (MEK1), MAP2K2 (MEK2), and in the GTPase-encoding gene KRAS. CFCS is characterized by craniofacial dysmorphic features, congenital heart defects, severe malnutrition, proportionate short stature, anomalies within the structure of skin and hair, and psychomotor disability. The pathophysiology of growth impairment is multifactorial with feeding difficulties, growth hormone deficiency, and insensitivity. Immunodeficiency has not been hitherto reported as an integral part of CFCS yet an increased activation of the RAS/MAPK signaling pathway may contribute to explaining the causal relationship between RASopathy and the dysfunctions within the B and T lymph cell compartments resulting in a deficiency in T cell costimulation and B cell maturation with impaired class switch recombination, somatic hypermutation, and high-affinity antibody production. We report on a boy born prematurely at 32 WGA, with the perinatal period complicated by pneumonia, respiratory distress syndrome, and valvular pulmonary stenosis. The boy suffered from recurrent pneumonia, obstructive bronchitis, sepsis, urinary tract infection, and recurrent fevers. He presented with severe hypotrophy, psychomotor disability, short stature, craniofacial dysmorphism, dental hypoplasia, sparse hair, and cryptorchidism. Whole genome sequencing showed a novel heterozygous pathogenic germline missense variant: c.364A > G; p.Asn122Asp in the MAP2K1 gene, supporting the diagnosis of CFCS. The immunological workup revealed hypogammaglobulinemia, IgG subclass, and specific antibody deficiency accompanied by decreased numbers of T helper cells and naive and memory B cells. Replacement immunoglobulin therapy with timely antibiotic prophylaxis were instituted. At the age of six years, growth hormone deficiency was diagnosed and the rGH therapy was started. The ever-increasing progress in genetic studies contributes to establishing the definitive CFCS diagnosis and sheds the light on the interrelated genotype-phenotype heterogeneity of RASopathies. Herein, we add new phenotypic features of predominating humoral immunodeficiency to the symptomatology of CFCS with a novel mutation in MAP2K1. While CFCS is a multifaceted disease, increased pediatricians’ awareness is needed to prevent the delay in diagnostics and therapeutic interventions.</p

Additional file 1 of Compound heterozygous IFT140 variants in two Polish families with Sensenbrenner syndrome and early onset end-stage renal disease

Additional file 1. List of genes and SNPs included in the NGS gene panel

Additional file 2 of Compound heterozygous IFT140 variants in two Polish families with Sensenbrenner syndrome and early onset end-stage renal disease

Additional file 2. List of primer sequences used for PCR, Sanger sequencing and qPCR

Table3_Results from Genetic Studies in Patients Affected with Craniosynostosis: Clinical and Molecular Aspects.xlsx

Background: Craniosynostosis (CS) represents a highly heterogeneous genetic condition whose genetic background has not been yet revealed. The abnormality occurs either in isolated form or syndromic, as an element of hundreds of different inborn syndromes. Consequently, CS may often represent a challenging diagnostic issue.Methods: We investigated a three-tiered approach (karyotyping, Sanger sequencing, followed by custom gene panel/chromosomal microarray analysis, and exome sequencing), coupled with prioritization of variants based on dysmorphological assessment and description in terms of human phenotype ontology. In addition, we have also performed a statistical analysis of the obtained clinical data using the nonparametric test χ2.Results: We achieved a 43% diagnostic success rate and have demonstrated the complexity of mutations’ type harbored by the patients, which were either chromosomal aberrations, copy number variations, or point mutations. The majority of pathogenic variants were found in the well-known CS genes, however, variants found in genes associated with chromatinopathies or RASopathies are of particular interest.Conclusion: We have critically summarized and then optimised a cost-effective diagnostic algorithm, which may be helpful in a daily diagnostic routine and future clinical research of various CS types. Moreover, we have pinpointed the possible underestimated co-occurrence of CS and intellectual disability, suggesting it may be overlooked when intellectual disability constitutes a primary clinical complaint. On the other hand, in any case of already detected syndromic CS and intellectual disability, the possible occurrence of clinical features suggestive for chromatinopathies or RASopathies should also be considered.</p

Table5_Results from Genetic Studies in Patients Affected with Craniosynostosis: Clinical and Molecular Aspects.xlsx

Background: Craniosynostosis (CS) represents a highly heterogeneous genetic condition whose genetic background has not been yet revealed. The abnormality occurs either in isolated form or syndromic, as an element of hundreds of different inborn syndromes. Consequently, CS may often represent a challenging diagnostic issue.Methods: We investigated a three-tiered approach (karyotyping, Sanger sequencing, followed by custom gene panel/chromosomal microarray analysis, and exome sequencing), coupled with prioritization of variants based on dysmorphological assessment and description in terms of human phenotype ontology. In addition, we have also performed a statistical analysis of the obtained clinical data using the nonparametric test χ2.Results: We achieved a 43% diagnostic success rate and have demonstrated the complexity of mutations’ type harbored by the patients, which were either chromosomal aberrations, copy number variations, or point mutations. The majority of pathogenic variants were found in the well-known CS genes, however, variants found in genes associated with chromatinopathies or RASopathies are of particular interest.Conclusion: We have critically summarized and then optimised a cost-effective diagnostic algorithm, which may be helpful in a daily diagnostic routine and future clinical research of various CS types. Moreover, we have pinpointed the possible underestimated co-occurrence of CS and intellectual disability, suggesting it may be overlooked when intellectual disability constitutes a primary clinical complaint. On the other hand, in any case of already detected syndromic CS and intellectual disability, the possible occurrence of clinical features suggestive for chromatinopathies or RASopathies should also be considered.</p

Table2_Results from Genetic Studies in Patients Affected with Craniosynostosis: Clinical and Molecular Aspects.xlsx

Background: Craniosynostosis (CS) represents a highly heterogeneous genetic condition whose genetic background has not been yet revealed. The abnormality occurs either in isolated form or syndromic, as an element of hundreds of different inborn syndromes. Consequently, CS may often represent a challenging diagnostic issue.Methods: We investigated a three-tiered approach (karyotyping, Sanger sequencing, followed by custom gene panel/chromosomal microarray analysis, and exome sequencing), coupled with prioritization of variants based on dysmorphological assessment and description in terms of human phenotype ontology. In addition, we have also performed a statistical analysis of the obtained clinical data using the nonparametric test χ2.Results: We achieved a 43% diagnostic success rate and have demonstrated the complexity of mutations’ type harbored by the patients, which were either chromosomal aberrations, copy number variations, or point mutations. The majority of pathogenic variants were found in the well-known CS genes, however, variants found in genes associated with chromatinopathies or RASopathies are of particular interest.Conclusion: We have critically summarized and then optimised a cost-effective diagnostic algorithm, which may be helpful in a daily diagnostic routine and future clinical research of various CS types. Moreover, we have pinpointed the possible underestimated co-occurrence of CS and intellectual disability, suggesting it may be overlooked when intellectual disability constitutes a primary clinical complaint. On the other hand, in any case of already detected syndromic CS and intellectual disability, the possible occurrence of clinical features suggestive for chromatinopathies or RASopathies should also be considered.</p

Table4_Results from Genetic Studies in Patients Affected with Craniosynostosis: Clinical and Molecular Aspects.pdf

Background: Craniosynostosis (CS) represents a highly heterogeneous genetic condition whose genetic background has not been yet revealed. The abnormality occurs either in isolated form or syndromic, as an element of hundreds of different inborn syndromes. Consequently, CS may often represent a challenging diagnostic issue.Methods: We investigated a three-tiered approach (karyotyping, Sanger sequencing, followed by custom gene panel/chromosomal microarray analysis, and exome sequencing), coupled with prioritization of variants based on dysmorphological assessment and description in terms of human phenotype ontology. In addition, we have also performed a statistical analysis of the obtained clinical data using the nonparametric test χ2.Results: We achieved a 43% diagnostic success rate and have demonstrated the complexity of mutations’ type harbored by the patients, which were either chromosomal aberrations, copy number variations, or point mutations. The majority of pathogenic variants were found in the well-known CS genes, however, variants found in genes associated with chromatinopathies or RASopathies are of particular interest.Conclusion: We have critically summarized and then optimised a cost-effective diagnostic algorithm, which may be helpful in a daily diagnostic routine and future clinical research of various CS types. Moreover, we have pinpointed the possible underestimated co-occurrence of CS and intellectual disability, suggesting it may be overlooked when intellectual disability constitutes a primary clinical complaint. On the other hand, in any case of already detected syndromic CS and intellectual disability, the possible occurrence of clinical features suggestive for chromatinopathies or RASopathies should also be considered.</p