In this study, we investigated a proband with ISOD from a nonconsanguineous Han Chinese family (Figure 1A). Proband’s younger sister was died and her younger brother was phenotypically normal. Proband’s parents were also phenotypically normal and asymptomatic.
In 2009, after a successful and uneventful pregnancy, proband’s mother (I-2) gave birth to a baby girl (II-1). One hour after birth, she was identified with "general cyanosis" and consecutively transferred to the department of neonatology and pediatrics of our hospital for treatment. After admission, we found that the baby was unable for sucking milk and high tension of her quadrilateral muscles with spasmodic twitch. Then, we performed CT and found subarachnoid haemorrhage. Hence, we clinically diagnosed the baby girl with "neonatal hypoxic-ischemic encephalopathy with subarachnoid hemorrhage". The child died on the same day.
Two years later, in 2011, proband’s mother (I-2) gave birth to a baby boy (II-2). During pregnancy, the color ultrasound was performed and showed that the posterior fossa of the fetus was 1.03cm. After birth, the baby had repeated intussusception. At the age of 2 months, the baby had convulsions once. The baby's head was found stable at the age of 5 months. At the age of 18 months, the baby could walk alone. We performed CT and found that the bilateral frontotemporal lobe sulcus and fissure were widened, and the brain was not fully developed. The rehabilitation treatment was carried out in the hospital. Presently, the psychomotor development of the boy is become normal.
In 2019, proband’s mother (I-2) achieved her third pregnancy. Prenatal examination identified no abnormalities. At 31 weeks of gestation, B-ultrasound was performed and found no abnormalities. However, at 37+4 weeks of gestation, B-ultrasound was performed again and showed that the left lateral ventricle was 1.12 cm wide (Figure 2.A), the depth of cisterna magna was 1.44 cm (Figure 2.B), there was a cystic area about 1.61cm × 0.91cm in size located in the midline of the brain above the anterior thalamus (Figure 2.C), and thin corpus callosum (Figure 2.D). At 38 weeks of gestation, MRI was performed and found that the posterior horn of bilateral ventricles was slightly widened, the third ventricles were enlarged and uplifted, connected with the longitudinal fissure cistern of the brain, and the possible absence of the corpus callosum was considered. There were many small and flaky abnormal signal shadows were identified in the white matter of bilateral frontotemporal lobe and bilateral basal ganglia, as well as many softened foci were found (Figure 2.E-H).
At 38+4 weeks of gestation, a live baby girl, the proband was born through a cesarean section. The birth weight of the proband was 3.17 kg, and the Apgar score was 10. Soon after delivery, the proband was identified with intermittent tremor or seizures in limb. Five days after birth, we also found that the proband was presented with intermittent tremor or seizures in limb, neonatal encephalopathy, subarachnoid haemorrhage, subarachnoid cyst, dysplasia of corpus callosum, neonatal convulsion, hypotension, respiratory failure, heart failure, patent foramen ovale, hyperlactatemia, severe metabolic acidosis, hyperglycemia, hyperkalemia, moderate anemia, high atrioventricular block and complete right bundle branch block.
Combined with the prenatal MRI result and based on the clinical symptoms, the proband was immediately transferred to the neonatological department of our hospital. Emergency CT scan was performed and showed multiple hypodense lesions in bilateral, frontal and parietal cortex, symmetrical hyperdense shadows were identified in bilateral basal ganglia, dysplasia of corpus callosum, a small amount of subarachnoid hemorrhage, enlargement of cisterna magna, possible subarachnoid abscess and compression of cerebellar hemisphere (Figure 2. I-M).
After admission, the patient still had intermittent scream, obvious tremor or seizures in limb, decrease in oxygen saturation in blood and blood pressure under the sedative state with chloral hydrate. The proband was also sedated by the treatment with phenobarbital sodium, and the convulsion still occasionally occurred. Dopamine and dobutamine were continuously pumped to maintain normal blood pressure, while continuous positive airway pressure ventilation was used to maintain the normal blood oxygen.
The proband was tested for genetic metabolic diseases and found no abnormalities (Supplementary Table S2). The proband was identified with high level of mean corpulscular volume (MCV), mean corpuscular hemoglobin (MCH), neutrophilic granulocyte percentage (NEUT%), immature cell percentage (IG%), NRBC%, NEUT, MONO#, IG#, NRBC#, RDW-SD, RDW-CV and IRF as well as low level of hemoglobin concentration (HGB), lymphocytes percentage (LYMPH%) and LFR (Supplementary Table S3). Liver function, renal function, electrolyte and blood ammonia test of the proband showed extremely high level of GGT, high level of Aspartic transaminase (AST), AST/ALT and low level of Total protein (TP) and Albumin (ALB) (Supplementary Table S4). We also performed TroPI, myocardial enzyme and AMON test for the proband and found extremely high level of Creatine Kinase (CK), Creatine kinase MB isoenzyme (CK-MB), Troponin (TroPI) and Ammonia (AMON) (Supplementary Table S5).
On the 6th day of hospitalization, the proband was identified with a sudden weak breath, the heart rate dropped to about 80 times/ minute, the heart sound was low and blunt, the blood oxygen saturation dropped to about 70%, the blood pressure was 47/12 mmHg. The patient was immediately intubated with mechanical ventilation, the resuscitation capsule was pressurized with oxygen, and no improvement was found after external chest compression, and finally the proband died.
Karyotype and chromosomal microarray analyses
Karyotyping found no abnormalities in structure of all the chromosomes in the proband (46, XX) (Figure 3.A). No pathogenic copy number variations (CNVs) were identified in all the chromosomes of the proband by CAN (Figure 3.B).
WES and Sanger sequencing identified a novel variant in SUOX gene
WES was performed for the proband. WES identified a novel 16 nucleotide deletion (c.1406_1421delCCTGGCAGGTGGCTAA) and a previously reported substitution (c.1200C>G) in exon 6 of the SUOX gene in the proband (Figure 4). The novel 16 nucleotide deletion (c.1406_1421delCCTGGCAGGTGGCTAA) leads to a frameshift (p.Thr469Serfs*20) followed by premature termination of translation which finally results into the formation of a truncated sulfite oxidase of 488 amino acids instead of the wild type sulfite oxidase of 545 amino acids. The substitution (c.1200C>G) also leads to a premature stop codon (p.Y400*) followed by the formation of a truncated sulfite oxidase of 399 amino acids instead of the wild type sulfite oxidase of 545 amino acids. Hence, both the variants are loss-of-function variants. The proband’s father and mother is carrying the substitution (c.1200C>G) and deletion (c.1406_1421delCCTGGCAGGTGGCTAA) in a heterozygous state respectively (Figure 4). These two variants were not identified in the elder brother of the proband as well as in the 100 healthy individuals. This heterozygous novel 16 nucleotide deletion (c.1406_1421delCCTGGCAGGTGGCTAA) also not present in the Human Gene Variant database (HGMD, www.hgmd.cf.ac.uk/), Online Mendelian Inheritance in Man (MIM, (https://www.omim.org). These two heterozygous variants are not found in BGI’s database, consisting of ~ 50,000 Chinese Han samples. We also did not find these two variants in ExAC, gnomAD, dbSNP and 1000 Genome Database. Our present study identified the first variant in SUOX gene associated with ISOD in Chinese population. In this study, we also described the importance of WES as a potential sequencing technology for identifying candidate variant in the SUOX associated ISOD patients.
In silico Analysis
These two variants (c.1406_1421delCCTGGCAGGTGGCTAA, p.Thr469Serfs*20; c.1200C>G, p.Tyr400*) were predicted as “disease causing” by Mutation Taster (http://mutationtaster.org/) .