Massive parallel sequencing as a new diagnostic approach for phenylketonuria and tetrahydrobiopterin-deficiency in Thailand

Abstract Background Hyperphenylalaninemia (HPA) can be classified into phenylketonuria (PKU) which is caused by mutations in the phenylalanine hydroxylase (PAH) gene, and BH4 deficiency caused by alterations in genes involved in tetrahydrobiopterin (BH4) biosynthesis pathway. Dietary restriction of phenylalanine is considered to be the main treatment of PKU to prevent irreversible intellectual disability. However, the same dietary intervention in BH4 deficiency patients is not as effective, as BH4 is also a cofactor in many neurotransmitter syntheses. Method We utilized next generation sequencing (NGS) technique to investigate four unrelated Thai patients with hyperphenylalaninemia. Result We successfully identified all eight mutant alleles in PKU or BH4-deficiency associated genes including three novel mutations, one in PAH and two in PTS, thus giving a definite diagnosis to these patients. Appropriate management can then be provided. Conclusion This study identified three novel mutations in either the PAH or PTS gene and supported the use of NGS as an alternative molecular genetic approach for definite diagnosis of hyperphenylalaninemia, thus leading to proper management of these patients in Thailand

The most 5′ truncating homozygous mutation of WNT1 in siblings with osteogenesis imperfecta with a variable degree of brain anomalies: a case report

Abstract Background WNT1 mutations cause bone fragility as well as brain anomalies. There are some reported cases of WNT1 mutations with normal cognition. Genotype and phenotype correlation of WNT1 mutations has not been established. Case presentation Here we present two female siblings with osteogenesis imperfecta (OI) born to a consanguineous couple. Both sustained severe bone deformities. However, only the younger had severe brain anomalies resulting in an early death from pneumonia, while the older had normal intellectual development. Next generation sequencing showed a homozygous mutation, c.6delG, p.Leu3Serfs*36 in WNT1. To our knowledge, it is the most 5′ truncating mutation to date. Conclusion This report emphasizes the intrafamilial variability of brain anomalies found in this OI type and suggests that WNT1 may not be necessary for normal human cognitive development

Pathogenic variant detection rate by whole exome sequencing in Thai patients with biopsy-proven focal segmental glomerulosclerosis

Abstract The spectra of underlying genetic variants for various clinical entities including focal segmental glomerulosclerosis (FSGS) vary among different populations. Here we described the clinical and genetic characteristics of biopsy-proven FSGS patients in Thailand. Patients with FSGS pathology, without secondary causes, were included in our study. Clinical laboratory and pathological data were collected. Whole-exome sequencing (WES) was subsequently performed. 53 unrelated FSGS patients were recruited. 35 patients were adults (66.0%), and 51 patients were sporadic cases (96.2%). Clinical diagnosis before kidney biopsy was steroid-resistant nephrotic syndrome (SRNS) in 58.5%, and proteinuric chronic kidney disease in 32.1%. Using WES, disease-associated pathogenic/likely pathogenic (P/LP) variants could be identified in six patients including the two familial cases, making the P/LP detection rate of 11.3% (6/53). Of these six patients, two patients harbored novel variants with one in the COL4A4 gene and one in the MAFB gene. Four other patients carried previously reported variants in the CLCN5, LMX1B, and COL4A4 genes. Four of these patients (4/6) received immunosuppressive medications as a treatment for primary FSGS before genetic diagnosis. All four did not respond to the medications, emphasizing the importance of genetic testing to avoid unnecessary treatment. Notably, the mutation detection rates in adult and pediatric patients were almost identical, at 11.4% and 11.1%, respectively. In conclusion, the overall P/LP variant detection rate by WES in biopsy-proven FSGS patients was 11.3%. The most identified variants were in COL4A4. In addition, three novel variants associated with FSGS were detected

Details of discordant variants from whole genome and exome sequencing data.

<p>Details of discordant variants from whole genome and exome sequencing data.</p

Number of discordant variants after applying different exclusion criteria for WGS and WES experiments.

<p>Number of discordant variants after applying different exclusion criteria for WGS and WES experiments.</p

Electropherograms of Sanger sequencing.

<p>Electropherograms of Sanger sequencing of the selected 15 possible discordant variants between the two monozygotic twins identified by either whole genome or exome sequencing experiments. Upper and lower electropherograms of each panel represented twin A and twin B, respectively.</p

Summary of sequencing results from whole genome and exome sequencing data.

<p>Summary of sequencing results from whole genome and exome sequencing data.</p