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Cortical atrophy and hypofibrinogenemia due to FGG and TBCD mutations in a single family: a case report
© The Author(s). 2018
Received: 29 November 2017
Accepted: 1 May 2018
Published: 16 May 2018
Blended phenotypes or co-occurrence of independent phenotypically distinct conditions are extremely rare and are due to coincidence of multiple pathogenic mutations, especially due to consanguinity. Hereditary fibrinogen deficiencies result from mutations in the genes FGA, FGB, and FGG, encoding the three different polypeptide chains that comprise fibrinogen. Neurodevelopmental abnormalities have not been associated with fibrinogen deficiencies. In this study, we report an unusual patient with a combination of two independently inherited genetic conditions; fibrinogen deficiency and early onset cortical atrophy.
The study describes a male child from consanguineous family presented with hypofibrinogenemia, diffuse cortical atrophy, microcephaly, hypertonia and axonal motor neuropathy. Through a combination of homozygosity mapping and exome sequencing, we identified bi-allelic pathogenic mutations in two genes: a homozygous novel truncating mutation in FGG (c.554del; p.Lys185Argfs*14) and a homozygous missense mutation in TBCD (c.1423G > A;p.Ala475Thr). Loss of function mutations in FGG have been associated with fibrinogen deficiency, while the c.1423G > A mutation in TBCD causes a novel syndrome of neurodegeneration and early onset encephalopathy.
Our study highlights the importance of homozygosity mapping and exome sequencing in molecular prenatal diagnosis, especially when multiple gene mutations are responsible for the phenotype.
The manifestation of two genetically and phenotypically distinct conditions in a single individual is rare and can be due to the co-occurrence of multiple inherited pathogenic loci. Although it is common on patients from consanguineous families due to higher chance of homozygosity of multiple recessively inherited genes, non-consanguineous families with such conditions have also been reported . Fibrinogen, a glycoprotein synthesized in hepatocytes, functions in the final steps of blood coagulation as a precursor monomer of the fibrin hemostatic plug. Fibrinogen deficiency (Factor I deficiency), is a rare inherited bleeding condition due to bi-allelic mutations in one of the three fibrinogen genes FGA, FGB and FGG; these encode α, β and γ fibrinogen polypeptides, respectively, which are folded together to form the mature fibrinogen hexameric structure . Mutations in the fibrinogen genes either affect the quantity of circulating fibrinogen (as in afibrinogenemia or hypofibrinogenemia) or the quality of fibrinogen (as in dysfibrinogenemia) . Symptoms of fibrinogen deficiency include bleeding of the umbilical cord or GI tract, oral and mucosal bleeding, and isolated intracranial bleeding due to traumatic injury; neurodevelopmental symptoms have not been documented [3, 4].
Microtubules are components of the cellular cytoskeleton and are involved in several cellular processes including the cell cycle, motility and intracellular trafficking. In eukaryotes, microtubules form by polymerization of α-β tubulin heterodimers in a head-to-tail fashion, using GTP hydrolysis as the fuel source . Proper polymerization and folding of tubulin monomers involves a series of molecular chaperones (TBCA-TBCE) that assist the formation of α-β tubulin heterodimer . Microtubule polymerization dynamics is crucial for cells, especially for the cellular differentiation and migration of neurons. A spectrum of neurological disorders have been characterized by abnormal neuronal migration and impaired axon guidance due to mutations in the genes that encode α and β tubulin subtypes . Recently, a group of patients was reported with early onset cortical atrophy, neurodegeneration and microcephaly due to bi-allelic mutations in TBCD, a tubulin folding chaperone encoding gene [8–10].
In this study, we present a consanguineous family whose proband presented with hypofibrinogenemia and cortical atrophy. Whole exome sequencing revealed that our proband’s blended phenotype is due to mutations in two unrelated genes from two different loci, TBCD and FGG.
At 24 months of age, the proband's weight was 11 kg (< 5th centile), height 81 cm (< 5th centile) and head circumference of 45.5 cm (< 3rd centile). He had microcephaly, deep set eyes, increased tone in all four limbs with exaggerated deep tendon reflexes and contracture of hamstrings muscles (Fig. 1b). He had a head lag on pulling to sitting position and, on axial suspension, scissored due to excessive axial tone. Visual tracking was absent. Brain MRI at 18 months revealed diffuse cortical atrophy with white matter volume loss and dysgenesis of the posterior corpus callosum (Fig. 1c and d). EEG showed left frontal epileptiform abnormalities during sleep. Nerve conduction velocity showed axonal motor neuropathy affecting bilateral peroneal and ulnar nerves. Fundus evaluation revealed marked temporal disc pallor bilaterally. Visual evoked potentials showed asymmetrically reduced amplitude from the left eye compared to the right eye. Brain stem auditory evoked response (BERA) showed normal latencies of all the waves, with the threshold estimated at 35 dB. Based on his predominant white matter neurodegeneration, enzyme analyses for Krabbe disease and neuronal ceroid lipofuscinosis were performed and found to be normal. At 30 months, the boy developed a huge hematoma involving the right upper eyelid following a trivial fall; the bleeding was controlled with infusion of two units of cryoprecipitate (Fig. 1b).
The Proband had three other siblings. His oldest sibling is a female with normal motor and mental development (II.1). Another sibling, the second child in the family, was a female with prolonged bleeding from the umbilical cord and documented hypofibrinogenemia (II.2). Her birth weight was 3000 g, and there was no birth asphyxia. She had global developmental delay, seizures by 6 months of age, and only partial head control by 15 months. There was no neuroregression. She expired at 15 months of age following an attack of pneumonia. The third child (II.3) in this family was a male with a birth weight of 3650 g. He had normal motor and mental development until 8 months of age and could sit when put in a sitting position. By 8 months, he developed seizures and lost all acquired milestones. He expired at 4 years of age with pneumonia. Brain MRI at 1 year of age showed areas of diffuse cortical atrophy with predominant white matter volume loss and atrophic corpus callosum (Fig. 1e and f). He did not have any bleeding manifestations. The CARE guidelines were followed in reporting this case.
To identify the genetic etiology, SNP genotyping and whole exome sequencing have been performed. SNP genotyping was done on genomic DNA from the proband and parents as previously described . Whole exome sequencing was performed on the proband using the Agilent SureSelect Target Enrichment Kit and the Illumina Hiseq 2000/2500 sequencer (Illumina, Inc., San Diego, CA). Reads were aligned with the human reference genome (hg19; NCBI build 37; Feb. 2009) using Burrows-Wheeler Alignment Tool . Variant calling was performed with GATK  and functionally annotated using SnpEff . Given the history of consanguinity in the pedigree, homozygous variants that are in the homozygous areas were filtered based on allele frequency less than 0.01 with no reported healthy homozygotes in online databases, dbSNP, 1000G, ESP6500, ExAC and gnomAD (Additional file 1: Table S1). Pathogenicity was deemed likely if the variant was truncating (splicing or non-sense) or missense and in-frame indels were predicted to be pathogenic using online prediction tools, Polyphen, SIFT, CADD and Mutation Taster. Confirmation and family screening of identified candidates were performed using direct Sanger sequencing (Applied Biosystems).
Homozygous areas identified in the proband (Fig. 1a-II.4) and the position of candidate genes
142,030,172 - 159,106,629
18,132,514 - 38,613,097
94,947,498 - 122,902,875
148,087,853 - 158,539,654
78,658,801 - 84,498,637
1 - 3,249,881
71,338,760 - 109,374,231
94,260,616 - 107,839,818
113,571,236 - 118,861,871
20,398,227 - 46,335,050
74,402,197 - 81,195,210
54,610,564 - 59,128,983
44,295,438 - 46,625,603
18,757,589 - 34,101,573
Discussion and conclusions
Recent advances in next generation sequencing have greatly advanced molecular diagnosis of monogenic diseases, as well as the identification of cases with blended phenotypes due to multiple gene effects. In this study, we describe the first example of a proband diagnosed with hypofibrinogenemia and a neurodevelopmental disorder associated with homozygous variants in two unrelated genes, FGG and TBCD. Exome sequencing revealed homozygous variants in four candidate genes including, GLMP, SYNPO2, FGG and TBCD (Additional file 1: Table S2). GLMP encodes glycosylated lysosomal membrane protein which has role in the metabolic regulation of liver . We excluded this variant since the patient did not have any liver disease. Synaptopodin-2 encoded by the SYNPO2 gene is implicated in the regulation of cell migration, muscle actin binding and actin bundling . There are no human disease associated with SYNPO2, but knockout mice are embryonic lethal and show pre-weaning lethality and abnormal morphology of joints and fingers (IMPC, International Mouse Phenotyping Consortium), hence we excluded this gene. FGG and TBCD mutations correlated well with the phenotype among the four candidates.
The single base pair deletion in FGG identified in our patient predicted a frameshift with protein truncation at position 199 to shift the frame and truncate protein at 199th position. This abolishes the C-terminal region of the fibrinogen gamma chain that contains glutamyl lysine intermolecular cross linking sites essential for the formation of gamma chain dimers . In addition, functional studies demonstrated that mutations affecting the C-terminal region showed either impaired assembly or secretion of the fibrinogen hexamer .
Comparison of clinical presentations of p.Ala475Thr patients
Edvardson et al., 2016 
Pode-Shakked et al., 2016 
Number of patients and gender
Age of onset (months)
Global developmental delay, intractable seizures, brain atrophy, dystonia
Microcephaly, seizures, developmental delay, hypotonia
Seizures, neuroregression, excessive bleeding from venipuncture sites
Diffuse cerebral and cerebellar atrophy, thin corpus callosum
Mild to severe cortical atrophy, thin corpus callosum
Diffuse cerebral atrophy, very thin corpus callosum
TBCD defects were initially studied as a possible contributor to the severe microcephaly phenotype in a 7-year-old girl , whereby the proband had a combination of a maternally inherited duplication and a missense mutation in TBCD, apart from harboring WDR62 mutations. Our case represents another example in which a TBCD mutation contributes a phenotype on top of that attributable to a deleterious FGG mutation.
In conclusion, we identified two unrelated homozygous mutations in a single proband, who manifested two distinctive phenotypes associated with the relevant genes. This study shows the importance of performing exome sequencing when patients present with divergent phenotypes.
We thank patients and families for participating in this study.
This work was supported by the Intramural Research Program of the National Human Genome Research Institute, and the Common Fund, Office of the Director, National Institutes of Health, USA.
Availability of data and materials
All data generated or analyzed during this study are included in this published article. Pathogenic mutations identified are submitted to public database LOVD (Leiden Open Variation Database) and now available online (https://databases.lovd.nl/shared/individuals/00143186).
JS performed experiments and bioinformatic analysis and wrote the manuscript. SN, VKP, PR and DY clinically evaluated the patients and helped to write the clinical summary. WAG and MCVM supervised the project and wrote the manuscript. All authors have read and approved the manuscript.
Ethics approval and consent to participate
Written informed consent for clinical details, photography and sample collection was obtained from parents under the protocol 76-HG-0238 (Diagnosis and treatment of patients with inborn errors of metabolism or other genetic disorders) approved by the Institutional Review Board of National Human Genome Research Institute.
Consent for publication
Written informed consent was obtained from the parents for publication of this case report and any accompanying images.
The authors declare that they have no competing interests.
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