This retrospective, single-center study of the largest reported cohort of Chinese patients with SSADH deficiency showed that the phenotypes and genotypes of the patients were complex. 12 novel variants on SSADH gene were identified. The onset age of Patient 1 was as early as 1 months old. Patient 2 and Patient 9 were lost of follow-up in the clinical course. And Patient 11 died of status epilepticus. The clinical-genetic correlations of our patients allow to speculate that the missense variants of Patient 2, Patient 9 and Patient 11 (c.637C > T, c.515G > A, c.1501G > C and c.1274T > C variants) are all dramatically reduce SSADH enzyme activity and crucial for the disease’s severity. Drugs such as L-carnitine, vigabatrin, taurine and rehabilitation training are all crucial to SSADH deficient patients.
SSADH deficiency is a rare disorder of organic acid metabolism. SSADH participates in the metabolic processes of the inhibitory neurotransmitter GABA in the brain and catalyzes the conversion of succinic semialdehyde into succinic acid, which enters the tricarboxylic acid cycle and eventually forms glutamate to maintain the balance of excitatory and inhibitory neurotransmitters in the brain. Consequently, SSADH enzyme deficiency blocks the transformation of succinic semialdehyde to succinic acid, causing the accumulation and significant increase of the intermediate metabolite 4-hydroxybutyric acid in the blood, urine, and cerebrospinal fluid, resulting in neurological damage [9]. Elevated GABA is predicted to disrupt the GABA shunt linking GABA transamination to the Krebs cycle and maintaining the balance of excitatory-inhibitory neurotransmitters. 4-hydroxybutyric acid is predicted to impact β-oxidation flux [9–11].
The clinical features of SSADH deficiency are highly heterogeneous, with neurological and psychiatric symptoms being the prominent manifestations of the disease [12–14]. Most patients experience mental retardation, hypotonia, hyporeflexia, and expressive language impairment in the first 2 years of life. Approximately 10% of patients present with progressive extrapyramidal symptoms and also show neurological symptoms such as ataxia and dyskinesia [12]. The onset age was within 1 year for all 13 patients in this study, with developmental delay appearing as the initial symptom. The children exhibited varied intellectual and motor retardation, and showed major motor, fine motor skills, and language development severely behind their peers [12].
Epileptic seizures are the main neurological complications of SSADH deficiency. Approximately 50% of patients show epileptic seizures. Tokatly Latzer et al. [14] reported that among 61 patients with SSADH deficiency, 30 (49%) cases showed epilepsy with the onset age usually within 10 year (4 mo to 19 year). The seizure types included absence seizures, generalized tonic-clonic seizures, myoclonic seizures, dystonic seizures, focal seizures, and epileptic spasms. A small number of patients also reported status epilepticus. The onset and severity of seizures correlate with an age-related decline in GABA and GABA-related metabolites in the cerebrospinal fluid. The older the patients and the lower the concentration of metabolites, the more serious the epileptic seizures. In this study, seven children (53.8%) experienced epileptic seizures in various forms, including absence seizures, generalized tonic-clonic seizures, focal seizures, and myoclonic seizures. One patient died of status epilepticus, consistent with the findings of previous literature reviews. The onset of epileptic seizures in Patient 1 occurred at 1 month of age, which was younger than the age of onset reported in the previous literature. Thus, this study further broadened the age spectrum of the onset of seizures in patients with SSADH deficiency and provided more clinical evidence for epilepsy research in relation to this disease in the future.
Neuropsychiatric symptoms were also common manifestations in patients with SSADH deficiency [12–13], with approximately 5–15% of adolescence and young adulthood. Moreover, 63–70% of the patients have been reported to show attention deficit hyperactivity disorder, obsessive compulsive disorder, anxiety, autism-like behaviors, aggression, and sleep disturbances. In our study, 76.9% of the patients showed sleep disturbances and 23.1% showed psychiatric disorders. One patient had autism spectrum disorder, one showed schizophrenia, and 15.4% of the patients showed emotional disorders. These finding are consistent with the results of previous studies. Among these patients, Patient 13 was diagnosed as showing schizophrenia at the age of 14 years, indicating that hallucinations, delusions, and schizophrenic behaviors were more common in elder children and adolescents. The results suggest that inherited metabolic disorders such as SSADH deficiency should be considered in the differential diagnosis of children with autism or schizophrenia accompanied by epileptic seizures and psychomotor retardation.
The key methods to detect SSADH deficiency are urine organic acids assay and gene analysis, with patients showing abnormally high levels of 4-hydroxybutyric acid in the urine as well as elevated concentrations of 4-hydroxybutyric acid and GABA in the serum and cerebrospinal fluid. Malaspina et al. [10] reported that the concentration of 4-hydroxybutyric acid in the cerebrospinal fluid of patients with SSADH deficiency ranged from 116 to 1110 µmol/L (normal reference range, < 3 µmol/L), with the GABA concentration ranging from 13.6 to 22.4 µmol/L (normal reference range, < 12 µmol/L). The patients’ serum lactate levels were mildly increased, while the blood amino acid and acylcarnitine profiles showed no specific abnormalities except a mild decrease in free carnitine. The definite diagnosis of SSADH deficiency depends on urine organic acids analysis, SSADH activity test in lymphoblasts and skin fibroblasts or SSADH gene analysis [2, 3]. In this study, all patients showed an increase of urine 4-hydroxybutyric acid concentrations. Their serum lactic acid levels were also mildly elevated. These findings were consistent with the biochemical characteristics of SSADH deficiency.
The typical brain MRI abnormities of SSADH deficient patients include symmetric T2 signal prolongation on both sides of the globus pallidus, cerebellar dentate nucleus, subcortical white matter, and brainstem, and variable atrophy in the brain and cerebellum [16]. Afacan et al. [16] reported that in patients with SSADH deficiency, the GABA levels in the regions described above was higher; the ratio of glutamate and glutamine/Cr was higher; the Cho/Cr ratio was not significantly abnormal; and the NAA/Cr ratio in the posterior cingulate was slightly lower than the corresponding values in the control group. In this study, all patients' brain MRI scans showed different abnormality, of which symmetrical T2 hyperintensity in the bilateral globus pallidus and cerebellar dentate nucleus were the main changes followed by cerebral white matter lesions and brain atrophy, which was consistent with previous literature reports. Thus, these lesions may be related to the clinical symptoms of athetosis and ataxia in children. However, symmetrical T2 hyperintensities at these sites could also be seen in patients with glutaric aciduria type 1 and other organic acidemia; thus, finding was not a specific change associated with SSADH deficiency and had no diagnostic significance. Children with similar brain MRI changes should undergo assessment of clinical symptoms, biochemical metabolic tests, and genetic analysis for further differential diagnosis.
The ALDH5A1 gene is located on chromosome 6p22.3, contains 10 exons, and encodes a polypeptide of 535 amino acids [17] (Fig. 2). More than 70 variants have been reported in the HGMD, with missense, deletion, insertion, and frameshift variants accounting for 57%, 17%, 9%, and 8%, respectively. At present, no hotspot mutations, or prevalent mutations of the ALDH5A1 gene have been identified [4, 17, 18]. Four of the 13 patients in this study had c.617delT pathogenic variants, suggesting this variant may be common in the Chinese population. A total of 18 variants were detected in ALDH5A1 gene of these 13 patients, of which six were previously reported and 12 were novel. Apart from eight missense variations, one each of frameshift, nonsense, insertion, and deletion variants were detected. Based on ACMP/AMP pathogenicity grading, three variants were categorized as pathogenic, four as likely pathogenic, and five as variants of unclear significance. The novel 12 variants further enrich the ALDH5A1 pathogenic gene variation database. Notably, eight of the novel mutations are missense changes. In Patient 2, which showed a very severe disease, c.637C > T (p.Arg213Trp) was identified in those null frameshift variants. This case provided a convincing proof for the severe deficiency associated with the missense mutations.
Patient 9 also lost follow-up probably because the poor response to treatment. The c.515G > A (p.Arg172His) coexisted with a second allele (c.1597G > A), resulting in a conserved glycine residue was substituted by Arginine (p.Gly533Arg). This variant was estimated as likely pathogenic, which strongly suggested that the single amino acid variations would have severe effects. AS for Patient 11 who died of status epilepticus, the variant type of c.1501G > C (p.Glu501Gln) and c.1274T > C (p.Leu425Pro) also suggested that these variants had severe side effects for the enzyme activity. On the other side of the clinical spectrum, the variant types of patients who showed good response to treatment and clinical recovery course also provide good indication of lack of severity for the novel missense mutations, while still needed further functional studies of such variants. These 12 novel variants further enriched the ALDH5A1 pathogenic gene variation database.
Curative treatments for SSADH deficiency are unavailable at present, and the existing clinical treatment regimens mainly focus on ameliorating some of the symptoms [19, 20]. Vigabatrin is an irreversible inhibitor of GABA transaminase that can inhibit the conversion of GABA to 4-hydroxybutyric acid, increase the level of GABA in the serum and cerebrospinal fluid, and reduce the level of 4-hydroxybutyric acid in patients. High doses of vigabatrin can prolong the survival time of SSADH deficient model mouse [19–21]. Valproic acid may affect mitochondrial function and inhibit residual SSADH activity and should be avoided in the treatment of SSADH deficiency. Methylphenidate, risperidone, fluoxetine, and benzodiazepines can be used for children experiencing psychological symptoms such as anxiety, depression, and hallucinations [21]. In addition, a ketogenic diet [22], L-cycloserine, and taurine have also been applied to the clinical treatment of this disease [2, 4]. At present, clinical trials are underway for drugs such as NCS-382 (4-hydroxybutyrate receptor antagonist), SGS-742 (GABA-B receptor antagonist) [23], and other therapeutic options targeting GABA and 4-hydroxybutyrate receptors, mTOR inhibitors. Enzyme replacement therapy [24] and other therapeutic modalities are also shown some clinical effects [19]. These methods indicated their potential as long-term treatment options. An experimental study on mice model suggested that glutamine supplementation improved peripheral but not central glutamine deficiency [25]. All 13 patients in this study received symptomatic treatment, including l-carnitine and nutrition support therapy. Seven patients with epilepsy were all treated with vigabatrin. During the follow-up period, two patients showed remission of convulsions, one was lost to follow-up, one died of status epilepticus, and three continued to experience frequent seizures. The symptoms related to intellectual and motor development, epileptic seizures, language development, and sleep disorders in eight patients were alleviated after treatment. During the follow-up period, all patients showed a reduction in the urine 4-hydroxybutyric acid level, indicating the effectiveness of the treatment.
Our study has certain limitation. One is the relatively small number of patients enrolled, which is understandable considering the ultra-rare nature of the disease. Another is the genotype-phenotype correlation study is solely based on the clinical manifestations of the patients. The quantity and type of enzyme proteins, as well as their structural and functional impairments, contribute to phenotype severity in SSADH deficiency [5]. For the further studies on the pathomechanisms and treatment of SSADH deficiency, functional analysis and animal models investigation should be performed.
In conclusion, SSADH deficiency is a ultra-rare disease caused by an abnormal GABA metabolic pathway. The clinical symptoms of the patients are complex and diverse. Prominent manifestations include movement disorders, delayed language development, and neuropsychiatric symptoms. With advancements in urine organic acid analysis and genetic testing technology, the early diagnosis rate of SSADH deficiency has increased. For patients showing unexplained language retardation, epileptic seizures, and psychomotor symptoms, urine organic acid and genetic testing should be performed promptly to ensure early diagnosis and treatment and avoid misdiagnosis. As for the genotype-phenotype correlations of our patients, it allow to infer that the c.637C > T, c.515G > A, c.1501G > C and c.1274T > C are all dramatically reduce SSADH enzyme activity and crucial for the disease’s severity.