Identification of Variants Underlying Phenylalanine Hydroxylase Deficiency in Saudi Arabia.

Background: Deleterious mutations in the human gene phenylalanine hydroxylase (PAH) encoding the phenylalanine hydroxylase enzyme give rise to classic phenylketonuria and hyperphenylalaninemia. Our study was designed to characterize the spectrum of variants in the PAH gene in Saudi patients. Materials and Methods: We screened a cohort of 72 Saudi patients with clinical and biochemical diagnoses of hyperphenylalaninemia at the largest tertiary care center in Saudi Arabia; the King Faisal Specialist Hospital and Research Center (KFSH&RC), Riyadh. All patient's charts were reviewed under an approved study by Institutional Review Board. Results: Twenty-one different PAH variants were identified among the 144 PAH alleles assessed by targeted gene sequencing. Within the studied cohort, 60 of 72 patients had homozygous mutations with the the remaining 12 being compound heterozygotes. The most prevalent of the disease alleles identified in this study was the p.(Arg252Trp) mutation, which accounted for 38 of 144 alleles (26.4%). With the high incidence of genetic disorders in the population, religiously permissible preventive reproductive measures are a priority in our practice. Prenatal diagnoses carried out on four fetuses revealed two that were homozygous for PAH pathogenic variants. In addition, pre-implantation genetic diagnoses were initiated for 19 families. Eight of these families completed more than one full cycle of treatment, from which one healthy newborn was delivered. Conclusions: This study describes the spectrum of PAH variants in the Saudi population and highlights the molecular heterogeneity underlying phenylketonuria and hyperphenylalaninemia. These results add to the existing knowledge about PAH variants in Middle Eastern Countries. These results can be further translated to provide: informed counseling; cascade carrier testing in extended family members; and pre-marital screening.

Knowledge and attitudes regarding non-invasive prenatal testing among women in Saudi Arabia.

OBJECTIVES: To explore women's knowledge and attitudes regarding NIPT, its implications, the factors affecting their decision to undergo the test and actions taken following a positive result. METHODS: In this descriptive study, women who were offered NIPT through the foetal maternal clinic, were asked to complete an anonymous questionnaire about NIPT. The questionnaire consisted of 29 statements and covered four areas: demographics, knowledge, attitudes and decision-making. RESULTS: A total of 150 women who were offered NIPT participated in this study. The results showed that generally women had poor knowledge of critical aspects of NIPT. This included the conditions tested for, the implications of the test and its limitations. Over 90% of women were in favour of NIPT and it being offered to all women of advanced maternal age while 66% of the tested women agreed to having confirmatory invasive testing in the case of a positive result. CONCLUSION: This study shows that the acceptance rate for NIPT is high despite incomplete understanding of the benefits and limitations of the test. The study findings support the need for education regarding this test through dedicated genetic counselling sessions in order to ensure that an informed decision can be made.

Autozygome and high throughput confirmation of disease genes candidacy.

PURPOSE: Establishing links between Mendelian phenotypes and genes enables the proper interpretation of variants therein. Autozygome, a rich source of homozygous variants, has been successfully utilized for the high throughput identification of novel autosomal recessive disease genes. Here, we highlight the utility of the autozygome for the high throughput confirmation of previously published tentative links to diseases. METHODS: Autozygome and exome analysis of patients with suspected Mendelian phenotypes. All variants were classified according to the American College of Medical Genetics and Genomics guidelines. RESULTS: We highlight 30 published candidate genes (ACTL6B, ADAM22, AGTPBP1, APC, C12orf4, C3orf17 (NEPRO), CENPF, CNPY3, COL27A1, DMBX1, FUT8, GOLGA2, KIAA0556, LENG8, MCIDAS, MTMR9, MYH11, QRSL1, RUBCN, SLC25A42, SLC9A1, TBXT, TFG, THUMPD1, TRAF3IP2, UFC1, UFM1, WDR81, XRCC2, ZAK) in which we identified homozygous likely deleterious variants in patients with compatible phenotypes. We also identified homozygous likely deleterious variants in 18 published candidate genes (ABCA2, ARL6IP1, ATP8A2, CDK9, CNKSR1, DGAT1, DMXL2, GEMIN4, HCN2, HCRT, MYO9A, PARS2, PLOD3, PREPL, SCLT1, STX3, TXNRD2, WIPI2) although the associated phenotypes are sufficiently different from the original reports that they represent phenotypic expansion or potentially distinct allelic disorders. CONCLUSIONS: Our results should facilitate the timely relabeling of these candidate disease genes in relevant databases to improve the yield of clinical genomic sequencing.

Delineating the phenotypic spectrum of hyperphosphatasia with mental retardation syndrome 4 in 14 patients of Middle-Eastern origin.

Hyperphosphatasia with mental retardation syndrome 4 (HPMRS4) is a rare autosomal recessive condition caused by an impairment of glycosylphophatidylinositol biosynthesis. The cardinal features of HPMRS4 include; characteristic facial features, severe intellectual disability and various neurologic abnormalities. We report here detailed clinical, biochemical, and molecular findings of 14 patients clinically suspected to have HPMRS4, from three Middle-Eastern Countries; Saudi Arabia, Qatar, and Oman. All patients in our series presented with the cardinal features pointing to HPMRS4 and with an elevated alkaline phosphatase level. Five patients had megalocornea, which have been reported recently in an Arab patient. Additionally, fracture, bilateral coxa valga, camptodactyly, truncal obesity, and hyperpigmented macules of the upper thigh, each was seen once and was not described before with HPMRS4. Additional clinical and radiological findings are described, supporting the novel clinical and radiological findings recently described in Egyptian patients. The utilization of homozygosity mapping coupled with PGAP3 sequencing and whole exome sequencing facilitated the mutation detection in these patients. These missense mutations include c.320C > T (p.S107 L), c.850C > T (p.H284Y), and c.851A > G (p.H284R) in the PGAP3 gene. We believe that the recurrent mutations identified in our cohort may represent founder mutations in big tribes from a certain geographical region of Saudi Arabia, Qatar, and Oman. Therefore, in case of a clinical suspicion of HPMRS4 in these populations, targeted genetic testing for the identified mutations should be performed first to expedite the genetic diagnosis.

Lethal digenic mutations in the K+ channels Kir4.1 (KCNJ10) and SLACK (KCNT1) associated with severe-disabling seizures and neurodevelopmental delay.

A 2-yr-old boy presented profound developmental delay, failure to thrive, ataxia, hypotonia, and tonic-clonic seizures that caused the death of the patient. Targeted and whole exome sequencing revealed two heterozygous missense variants: a novel mutation in the KCNJ10 gene that encodes for the inward-rectifying K+ channel Kir4.1 and another previously characterized mutation in KCNT1 that encodes for the Na+-activated K+ channel known as Slo2.2 or SLACK. The objectives of this study were to perform the clinical and genetic characterization of the proband and his family and to examine the functional consequence of the Kir4.1 mutation. The mutant and wild-type KCNJ10 constructs were generated and heterologously expressed in Xenopus laevis oocytes, and whole cell K+ currents were measured using the two-electrode voltage-clamp technique. The KCNJ10 mutation c.652C>T resulted in a p.L218F substitution at a highly conserved residue site. Wild-type KCNJ10 expression yielded robust Kir current, whereas currents from oocytes expressing the mutation were reduced, remarkably. Western Blot analysis revealed reduced protein expression by the mutation. Kir5.1 subunits display selective heteromultimerization with Kir4.1 constituting channels with unique kinetics. The effect of the mutation on Kir4.1/5.1 channel activity was twofold: a reduction in current amplitudes and an increase in the pH-dependent inhibition. We thus report a novel loss-of-function mutation in Kir4.1 found in a patient with a coexisting mutation in SLACK channels that results in a fatal disease.NEW & NOTEWORTHY We present and characterize a novel mutation in KCNJ10 Unlike previously reported EAST/SeSAME patients, our patient was heterozygous, and contrary to previous studies, mimicking the heterozygous state by coexpression resulted in loss of channel function. We report in the same patient co-occurrence of a KCNT1 mutation resulting in a more severe phenotype. This study provides new insights into the phenotypic spectrum and to the genotype-phenotype correlations associated with EAST/SeSAME and MMFSI.

Twenty novel mutations in BCKDHA, BCKDHB and DBT genes in a cohort of 52 Saudi Arabian patients with maple syrup urine disease.

Maple syrup urine disease (MSUD), an autosomal recessive inborn error of metabolism due to defects in the branched-chain α-ketoacid dehydrogenase (BCKD) complex, is commonly observed among other inherited metabolic disorders in the kingdom of Saudi Arabia. This report presents the results of mutation analysis of three of the four genes encoding the BCKD complex in 52 biochemically diagnosed MSUD patients originating from Saudi Arabia. The 25 mutations (20 novel) detected spanned across the entire coding regions of the BCKHDA, BCKDHB and DBT genes. There were no mutations found in the DLD gene in this cohort of patients. Prediction effects, conservation and modelling of novel mutations demonstrated that all were predicted to be disease-causing. All mutations presented in a homozygous form and we did not detect the presence of a "founder" mutation in any of three genes. In addition, prenatal molecular genetic testing was successfully carried out on chorionic villus samples or amniocenteses in 10 expectant mothers with affected children with MSUD, molecularly characterized by this study.

Genetic counselors' scope of practice and challenges in genetic counseling services in Saudi Arabia.

Genetic counseling is an evolving field in Saudi Arabia. In 2015, genetic counseling was recognized as a Master's program by the Saudi Commission for Health Specialties. Our genetic counselors combine their knowledge of genetics, counseling theory and interpersonal communication to serve Saudi and non-Saudi patients affected with a range of genetic conditions and/or birth defects. Most patients are referred to the clinic from different clinics at King Faisal Specialist Hospital and Research Centre (KFSHRC) and outside of KFSHRC for various indications. Carrier testing and preventative reproduction options rank highly on the reasons for referral to our clinics. The Saudi population has unique customs and beliefs, such as consanguinity and the evil eye. Challenges that are routinely encountered in our genetic counseling clinics include, but are not limited to, preventative reproductive options and termination of pregnancy, manifesting carriers, stigmatization of women and approaches to complex molecular findings. Working with families from different backgrounds and beliefs undoubtedly requires professionals with a distinctive set of skills and a structured clinical setting. This review article presents the scope of genetic counseling practice and tackles some of the challenges faced in providing genetic counseling in Saudi Arabia.

Combined TSC1 and LMX1B mutations in a single patient.

Tuberous sclerosis complex (TSC) and nail-patella syndrome (NPS) are autosomal dominant pleiotropic disorders with full penetrance that can both involve kidneys. TSC1 and NPS genes are located on chromosome 9q3. In a large family with the two disorders with two novel frameshift TSC1 and LMX1B mutations, we describe the phenotypes. The father, who has both disorders, has passed on TSC to three of his children, NPS to another three, and both TSC and NPS to one child. Patients carrying both mutations appear to show an additive phenotype and no obvious epistatic effects. The segregation of two dominant disorders in this family poses a challenge for genetic counseling and indicates the importance of a careful clinical and molecular evaluation for accurate risk assessment.

The development of genetic counseling services and training program in Saudi Arabia.

In 2005 the first Saudi genetic counseling training program was established by the Department of Medical Genetics at King Faisal Specialist Hospital and Research Center (KFSH&RC) in the Kingdom of Saudi Arabia. The program has graduated five genetic counselors with high diploma-level degree. This brief report describes the development of the genetic counseling training program and the factors that led to its establishment. Special emphasis is made to unique cultural practices including consanguinity, religious influence, and termination of pregnancy. This report also describes the current status of the genetic counseling services offered by KFSH&RC and availability of genetic testing.

Novel FBP1 gene mutations in Arab patients with fructose-1,6-bisphosphatase deficiency.

UNLABELLED: Deficiency of fructose-1,6-bisphosphatase (FBP) results in impaired gluconeogenesis, which is characterized by episodes of hyperventilation, apnea, hypoglycemia, and metabolic and lactic acidosis. This autosomal recessive disorder is caused by mutations in the FBP1 gene, which encodes for fructose-1,6-bisphosphatase 1 (FBP1). Although FBP1 gene mutations have been described in FBP-deficient individuals of various ethnicities, there has been limited investigation into the genetics of this disorder in Arab patients. This study employed five consanguineous Arab families, in which 17 patients were clinically diagnosed with FBP deficiency. Seven patients and six carrier parents were analyzed for mutations in the FBP1 gene. DNA sequencing of the FBP1 gene identified two novel mutations in these families. A novel six nucleotide repetitive insertion, c114_119dupCTGCAC, was identified in patients from three families. This mutation encodes for a duplication of two amino acids (p.Cys39_Thr40dup) in the N-terminal domain of FBP1. A novel nonsense c.841G>T mutation encoding for a p.Glu281X truncation in the active site of FBP1 was discovered in patients from two families. The newly identified mutations in the FBP1 gene are predicted to produce FBP1 deficiency. These mutations are the only known genetic causes of FBP deficiency in Arab patients. The p.Cys39_Thr40dup is the first reported amino acid duplication in FBP deficiency patients. CONCLUSION: This study provides a strong rationale for genetic testing of FBP deficient patients of Arab ethnicity for recurrent or novel mutations in the FBP1 gene.