Heterozygous c.175C>T variant in PURA gene causes severe developmental delay

Abstract

Key Clinical Message

This case report presents a child with PURA‐related neurodevelopmental disorder, caused by the heterozygous pathogenic variant c.175C>T (p.Gln59*). The clinical symptoms included microcephaly, brachygnathia, central and peripheral hypotonia, and developmental delay (non‐verbal), among others. On comparison with published literature, even patients with the same mutation present different clinical symptoms.

Abstract

This case report presents a child with PURA‐related neurodevelopmental disorder, caused by the heterozygous pathogenic variant c.175C>T (p.Gln59*), whose symptoms included microcephaly, brachygnathia, the development of a high anterior hairline, hip dysplasia, strabismus, severe hypotonia, developmental delay (non‐meaningful verbal), feeding difficulties, and respiratory difficulties. His development ceased with age, such that his development at 10 years corresponded to an infant of 6 months. Moreover, even patients with the same variant can have different clinical symptoms, such as the presence or absence of epilepsy or congenital malformations. Therefore, we should follow his long‐term clinical course and provide medical support as necessary.

1. INTRODUCTION

The purine‐rich element‐binding protein A (PURA) gene, located on chromosome 5, encodes Pur‐α. ¹ PURA is unique, in that it lack introns; only 3% of human genes lack introns. This suggests that evolutionary pressure has prevented genomic rearrangements in PURA. ² Pur‐α plays a critical role in normal postnatal brain development enabling the proliferation of neuronal cells and synapse formation. ³ Pur‐α, a DNA‐, and RNA‐binding protein, is thought to play roles in neurodegenerative disorders and may function as a neuroprotective factor. ²

PURA syndrome (MIM number: 616158) was initially thought to be caused by a single chromosomal deletion, the 5q31.3 microdeletion, which results in the deletion of a copy of PURA and neighboring genes. However, recent research shows that it can also be caused by pathogenic de novo variants in PURA. ⁴ , ⁵ , ⁶ Consistently, PURA‐related neurodevelopmental disorders (PURA‐NDDs) have been defined, that is, neurodevelopmental disorders associated with variants and deletions encompassing PURA. ⁷ This classification also includes the PURA syndrome, which can be caused by pathogenic variants of PURA in addition to the 5q31.3 microdeletion syndrome. ⁸

Symptoms of PURA‐NDDs include dysmorphic facial features, hypotonia, movement disorders, impaired intelligence, epilepsy, and brain abnormalities. ⁶ , ⁸ Most patients are unable to independently walk or speak, and frequently present with feeding difficulties and respiratory problems during the neonatal period, which leads to severe hypotonia. ⁶

Pur‐α knock‐out mouse models present neurological disorders, including spontaneous seizures, tremors, and early death. ⁹ Moreover, even Pur‐α (+/−) mice exhibit hypotonia, gait defects, and memory dysfunction, which is attributed to neuronal loss in the cerebellum. ¹⁰ Here, we report a case of PURA‐NDD associated with a heterozygous pathogenic variant in PURA, c.175C>T (p.Gln59*). Although this variant and the associated clinical manifetation was reported previously by Johannesen et al., ¹¹ there is no case report describing the clinical course associated with this variant. Herein, we present the clinical symptoms of the patient and discuss the relationship between the clinical manifestations and this pathogenic variant.

2. METHODS

Whole exome sequencing was performed for the patient described in this report. The process involved several steps, including library preparation, sequencing, data acquisition, processing, variant calling, annotation, and filtering for rare variants, which were performed as described previously. ¹² The candidate variant was confirmed by Sanger sequencing. Polymerase chain reaction (PCR) primer for the single nucleotide variant was designed using Primer3 software. PCR products amplified using LA Taq HS with GC buffer I (Takara Bio Inc.) were sequenced using BigDye Terminator v3.1 Cycle Sequencing kit (Thermo Fisher Scientific) using the 3500xL genetic analyzer (Applied Biosystems).

3. CASE REPORT

A six‐year‐old boy with microcephaly, hypotonia, and developmental delay visited our institution. He was born via caesarean section after 42 weeks and 0 days of gestation due to abnormalities in fetal heart rate. At birth, his height, weight, and Apgar score were 54.0 cm (+2.1 standard deviation [SD]), 4258 g (+2.6 SD), and 9/10, respectively. The patient's family had no history of hereditary disorders.

Immediately after birth, the newborn exhibited microcephaly, a small jaw, cleft of the soft palate, and hypotonia. During the neonatal period, he was fed using a nasogastric tube because of feeding difficulty. Two months after birth, he was able to feed without a nasogastric tube, and was then discharged with home oxygen therapy due to unstable breathing. The developmental delay progressed with persistent hypotonia. The Enjoji Scale revealed severe developmental delay at the age of 1 year and 4 months, with a total developmental quotient (DQ) of 30, which is an average of the following DQ results: personal relation, 40; hand movement, 35; speech, 35; ability to understand language, 35; basic habits, 22; and locomotive movement, 10. The results of muscle biopsy at the age of 1 year and 6 months were not abnormal. At 2 years of age, the patient developed bilateral hip subluxation. At 6 years of age, he had acquired stable head control and was able to roll over, but was unable to independently stand or say meaningful words. Brain magnetic resonance imaging (MRI) at the ages 1, 4, and 10 did not reveal any brain abnormalities (Figure 1A–C). Numerous tests were conducted to assess possible causes of the condition. Initial laboratory tests did not reveal any abnormal findings. Moreover, abnormalities were not found after performing a G‐banded chromosome analysis and gene analysis for spinal muscular atrophy.

FIGURE 1.

Clinical manifestations of the patient. Brain magnetic resonance imaging at age (A) 1 year, (B) 4 years, and (C) 10 years. His brain MRI did not show brain abnormalities including myelination delay. (D) Results of Sanger sequencing. The c.175C>T (p.Gln59*)) variant in PURA was not detected in the patient's father and mother. (E) Electroencephalogram at 10 years. His electroencephalograms showed a normal high voltage slow wave during sleeping. (F) Image of the child at 10 years. His facial muscles exhibit hypotonia, and he remains seated on a wheelchair.

Hence, the patient was registered in the Initiative on Rare and Undiagnosed Diseases (IRUD), ¹³ led by the Japan Agency for Medical Research and Development. Subsequently, whole exome sequencing was performed, and a heterozygous nonsense variant (c.175C>T [p.Gln59*]) of PURA (NM_005859.5) was identified. Sanger sequencing confirmed that this variant was a de novo mutation (Figure 1D). In‐silico analysis was performed, and this variant was predicted to be disease causing in Mutation Taster, ¹⁴ scored 35 in CADD, ¹⁵ and the genomic evolutionary rate profiling (GERP) score ¹⁶ was 4.49, suggesting that the affected amino acid residue (Gln59) was highly conserved evolutionarily. Moreover, the variant is not registered in Genome Aggregation Database (gnomad, https://gnomad.broadinstitute.org) and Japanese database (jMorp 54KJPN, https://jmorp.megabank.tohoku.ac.jp/). Therefore, this variant was considered pathogenic according to the ACMG/AMP guidelines (PVS1, PM2, PM6, and PP3), ¹⁷ and is registered as pathogenic in ClinVar (https://www.ncbi.nlm.nih.gov/clinvar/, Accession: VCV000582890.6). ¹⁰

Since birth, the patient has received rehabilitation therapy, but the Enjoji developmental test at age 10 showed an increase in the severity of developmental delay with a total DQ of 6 (a higher value indicates more severe developmental delay). This value is an average of the following DQ results: personal relation, 8; basic habits; 8, hand movement, 7; locomotive movement, 6; ability to understand language, 6; and speech, 4. The patient has not experienced an epileptic episode and electroencephalograms exhibited a normal high voltage slow wave during sleep (Figure 1E). At present, he lives a stable life without medication, receiving rehabilitation and is enrolled in a social support program for developmental delay (Figure 1F).

4. DISCUSSION

In the present report, we describe a case of PURA‐NDD caused by the heterozygous pathogenic variant c.175C>T (p.Gln59*) in a six‐year‐old boy. This variant was previously identified by Johannesen et al. ¹¹ ; however, several clinical symptoms reported by the group were absent from our case, including epilepsy (Table 1). Majority of the symptoms noted in the present case were consistent with those reported in other cases. ³ , ⁴ , ⁶ , ¹¹ , ¹⁸ , ¹⁹ , ²⁰ , ²¹ , ²² , ²³ , ²⁴ Reijnders et al. ⁶ reported that even patients with the same mutation can present with different clinical symptoms, such as the presence or absence of epilepsy or congenital malformations. However, Class D mutations, such as c.847delG (p.Glu283Argfs*45), with localization at the very end of the C‐terminus but intact three PUR domains, may be correlated with a less severe phenotype. Therefore, these results suggest a considerable clinical variability that is associated with PURA mutations, which accounts for the clinical and genetic heterogeneity observed in patients. The phenotypic variability of PURA‐NDDs is most likely influenced by several other factors, including genetic factors. ⁶

TABLE 1.

Clinical phenotype in patients with the c.175C>T variant.

Nucleic acid	Amino acid	sex	Age	Phenotype	Reference
c.175C>T	p.Gln59*	M	10y	Hypotonia, feeding difficulties, respiratory difficulties, developmental delays (non‐verbal), strabismus, hip dysplasia, high anterior hairline	This study
		NA	2y	Epilepsy, hypotonia, feeding difficulties, respiratory difficulties, hypothermia, basal ganglia calcifications, agenesis of the corpus callosum, Omphalocele, schisis	Johannesen (2021)

Although majority of the symptoms of PURA‐NDDs may result from central nervous system (CNS) abnormalities, impaired peripheral neuromuscular system (PMS) or a combination of both CNS and PMS abnormalities may also be associated with clinical manifestations of PURA‐NDDs, given that PURA is ubiquitously expressed. Johannesen et al. ¹¹ performed MRI scanning and reported that almost half of the patients with PURA‐NDDs showed delayed myelination and brain volume loss. However, in the present case, we did not observe delayed myelination or brain volume loss. Nonetheless, his intelligence level was significantly lower than that of unaffected children of the same age. Moreover, his development ceased with age, such that his development at 10 years corresponded to that of a 6 month‐old infant.

Pur‐α interacts with the metabotropic glutamate receptor 5 (mGluR5) in the brain, which is involved in the synaptic plasticity through oscillations in the long‐term potentiation. ²⁵ Impaired function of mGluR5 is associated with several neurodevelopmental and psychiatric disorders. ²⁶ Moreover, the role of PURA in transcriptional and translational regulation ²⁷ is also associated with neurodegenerative expansion disorders.

The patient reported herein presented with weakness and feeding and breathing difficulty during the neonatal period. Moreover, although definite muscle atrophy was not noted, the patient exhibited lack of antigravity movement, decreased deep tendon reflexes, and myopathic face. Over the years, the patient has developed both CNS and PMS abnormalities. Although he has received regular rehabilitation and training in a special support program, substantial improvement in intelligence is unlikely. Therefore, we recommend that his long‐term clinical course should be followed and medical support should be provided as necessary.

There is no effective FDA‐approved drug for PURA‐NDDs. However, pyridostigmine may be effective for the muscular/neuromuscular junction presentation observed in PURA‐NDDs because this drug inhibits the action of acetylcholinesterase and is generally used for symptomatic treatment in patients with myasthenia gravis. ⁷ , ²⁸ Moreover, salbutamol, fluoxetine, and 4‐aminopyridine should also be considered as a treatment for specific symptoms associated with PURA‐NDDs. ⁷

5. CONCLUSION

The clinical manifestations of PURA‐NDDs may result from a combination of both CNS and PMS impairment. Moreover, the clinical phenotypes are different for each patient, even in patients with the same pathogenic variant. Since the information on patients with PURA‐NDDs is limited, we should carefully follow the long‐term clinical course of patients with PURA‐NDDs to expand our knowledge on the topic.

AUTHOR CONTRIBUTIONS

Yusuke Noda: Conceptualization; data curation; writing – original draft. Jun Kido: Conceptualization; data curation; formal analysis; investigation; project administration; resources; supervision; visualization; writing – original draft. Yohei Misumi: Formal analysis; methodology; supervision. Keishin Sugawara: Conceptualization; data curation; formal analysis; investigation. Sachiko Ohori: Data curation; formal analysis; investigation. Atsushi Fujita: Conceptualization; investigation; methodology. Naomichi Matsumoto: Funding acquisition; investigation; project administration. Mitsuharu Ueda: Conceptualization; data curation; project administration; supervision. Kimitoshi Nakamura: Conceptualization; supervision.

FUNDING INFORMATION

This work was supported in part by the Japan Society for the Promotion of Science (JSPS), Grants‐in‐Aid for scientific research (KAKENHI: grant numbers JP20K08207 (J. Kido) and JP22K15901 (A. Fujita)), Japan Agency for Medical Research and Development (AMED: grant numbers JP22ek0109486, JP22ek0109549, and JP22ek010993 (N. Matsumoto)), and the Takeda Science Foundation (N. Matsumoto).

CONFLICT OF INTEREST STATEMENT

The authors declare that there are no conflicts of interest related to this study.

ETHICS STATEMENT

This study was approved by the Ethics Committee of the Faculty of Life Science, Kumamoto University (No. 1574 (Genome No. 382)). Written informed consent was obtained from the legal guardian of the patient for publication of the details of their medical case and any accompanying images.

CONSENT

Written informed consent was obtained from the patient's mother to publish this case report and any accompanying images. All authors have viewed and agreed to the submission.

Noda Y, Kido J, Misumi Y, et al. Heterozygous c.175C>T variant in PURA gene causes severe developmental delay Clin Case Rep. 2023;11:e7779. doi: 10.1002/ccr3.7779

DATA AVAILABILITY STATEMENT

All data generated or analyzed during this study are included in this article and its online supplementary material. Further inquiries can be directed to the corresponding author.