Salma M. Wakil

The Affymetrix DMET Plus Platform Reveals Unique Distribution of ADME-Related Variants in Ethnic Arabs

Background. The Affymetrix Drug Metabolizing Enzymes and Transporters (DMET) Plus Premier Pack has been designed to genotype 1936 gene variants thought to be essential for screening patients in personalized drug therapy. These variants include the cytochrome P450s (CYP450s), the key metabolizing enzymes, many other enzymes involved in phase I and phase II pharmacokinetic reactions, and signaling mediators associated with variability in clinical response to numerous drugs not only among individuals, but also between ethnic populations. Materials and Methods. We genotyped 600 Saudi individuals for 1936 variants on the DMET platform to evaluate their clinical potential in personalized medicine in ethnic Arabs. Results. Approximately 49% each of the 437 CYP450 variants, 56% of the 581 transporters, 56% of 419 transferases, 48% of the 104 dehydrogenases, and 58% of the remaining 390 variants were detected. Several variants, such as rs3740071, rs6193, rs258751, rs6199, rs11568421, and rs8187797, exhibited significantly either higher or lower minor allele frequencies (MAFs) than those in other ethnic groups. Discussion. The present study revealed some unique distribution trends for several variants in Arabs, which displayed partly inverse allelic prevalence compared to other ethnic populations. The results point therefore to the need to verify and ascertain the prevalence of a variant as a prerequisite for engaging it in clinical routine screening in personalized medicine in any given population.

Mutation of the slow myosin heavy chain rod domain underlies hyaline body myopathy

Objective: To identify the gene and specific mutation underlying hyaline body myopathy in the family studied. Methods: A microsatellite-based whole genome scan was performed. Linkage analysis assumed autosomal dominant inheritance and equal allele frequencies. A candidate gene approach within the linked interval and direct sequencing were used for mutation detection. Results: Initial analysis indicated a maximum lod score of 3.01 at D14S1280. High-density mapping surrounding the linked locus was performed. Multipoint analysis showed that the linked region with a maximum lod score of 3.01 extended from D14S742 to D14S608 with a peak non-parametric linkage (NPL) score of 3.75 at D14S608. The myosin heavy chain genes MYH6 and MYH7 map to the region between D14S742 and D14S1280. Sequence analysis of the coding regions of MYH7 revealed an A→T transversion at nucleotide position 25596 (M57965) resulting in a histidine-to-leucine amino acid change at residue 1904 (H1904L). Conclusion: Pathogenicity of the MYH7 H1904L mutation most likely results from disruption of myosin heavy chain assembly or stability of the sarcomeric protein. The MYH7 tail domain mutation results in an inclusion body myopathy with an apparent absence of hypertrophic cardiomyopathy usually associated with mutations of this gene.

A new GWAS and meta-analysis with 1000Genomes imputation identifies novel risk variants for colorectal cancer

Genome-wide association studies (GWAS) of colorectal cancer (CRC) have identified 23 susceptibility loci thus far. Analyses of previously conducted GWAS indicate additional risk loci are yet to be discovered. To identify novel CRC susceptibility loci, we conducted a new GWAS and performed a meta-analysis with five published GWAS (totalling 7,577 cases and 9,979 controls of European ancestry), imputing genotypes utilising the 1000 Genomes Project. The combined analysis identified new, significant associations with CRC at 1p36.2 marked by rs72647484 (minor allele frequency [MAF] = 0.09) near CDC42 and WNT4 (P = 1.21 × 10−8, odds ratio [OR] = 1.21 ) and at 16q24.1 marked by rs16941835 (MAF = 0.21, P = 5.06 × 10−8; OR = 1.15) within the long non-coding RNA (lncRNA) RP11-58A18.1 and ~500 kb from the nearest coding gene FOXL1. Additionally we identified a promising association at 10p13 with rs10904849 intronic to CUBN (MAF = 0.32, P = 7.01 × 10-8; OR = 1.14). These findings provide further insights into the genetic and biological basis of inherited genetic susceptibility to CRC. Additionally, our analysis further demonstrates that imputation can be used to exploit GWAS data to identify novel disease-causing variants.

Clinical genomics expands the morbid genome of intellectual disability and offers a high diagnostic yield

Intellectual disability (ID) is a measurable phenotypic consequence of genetic and environmental factors. In this study, we prospectively assessed the diagnostic yield of genomic tools (molecular karyotyping, multi-gene panel and exome sequencing) in a cohort of 337 ID subjects as a first-tier test and compared it with a standard clinical evaluation performed in parallel. Standard clinical evaluation suggested a diagnosis in 16% of cases (54/337) but only 70% of these (38/54) were subsequently confirmed. On the other hand, the genomic approach revealed a likely diagnosis in 58% (n=196). These included copy number variants in 14% (n=54, 15% are novel), and point mutations revealed by multi-gene panel and exome sequencing in the remaining 43% (1% were found to have Fragile-X). The identified point mutations were mostly recessive (n=117, 81%), consistent with the high consanguinity of the study cohort, but also X-linked (n=8, 6%) and de novo dominant (n=19, 13%). When applied directly on all cases with negative molecular karyotyping, the diagnostic yield of exome sequencing was 60% (77/129). Exome sequencing also identified likely pathogenic variants in three novel candidate genes (DENND5A, NEMF and DNHD1) each of which harbored independent homozygous mutations in patients with overlapping phenotypes. In addition, exome sequencing revealed de novo and recessive variants in 32 genes (MAMDC2, TUBAL3, CPNE6, KLHL24, USP2, PIP5K1A, UBE4A, TP53TG5, ATOH1, C16ORF90, SLC39A14, TRERF1, RGL1, CDH11, SYDE2, HIRA, FEZF2, PROCA1, PIANP, PLK2, QRFPR, AP3B2, NUDT2, UFC1, BTN3A2, TADA1, ARFGEF3, FAM160B1, ZMYM5, SLC45A1, ARHGAP33 and CAPS2), which we highlight as potential candidates on the basis of several lines of evidence, and one of these genes (SLC39A14) was biallelically inactivated in a potentially treatable form of hypermanganesemia and neurodegeneration. Finally, likely causal variants in previously published candidate genes were identified (ASTN1, HELZ, THOC6, WDR45B, ADRA2B and CLIP1), thus supporting their involvement in ID pathogenesis. Our results expand the morbid genome of ID and support the adoption of genomics as a first-tier test for individuals with ID.

Unbiased targeted next-generation sequencing molecular approach for primary immunodeficiency diseases

BACKGROUND Molecular genetics techniques are an essential diagnostic tool for primary immunodeficiency diseases (PIDs). The use of next-generation sequencing (NGS) provides a comprehensive way of concurrently screening a large number of PID genes. However, its validity and cost-effectiveness require verification. OBJECTIVES We sought to identify and overcome complications associated with the use of NGS in a comprehensive gene panel incorporating 162 PID genes. We aimed to ascertain the specificity, sensitivity, and clinical sensitivity of the gene panel and its utility as a diagnostic tool for PIDs. METHODS A total of 162 PID genes were screened in 261 patients by using the Ion Torrent Proton NGS sequencing platform. Of the 261 patients, 122 had at least 1 known causal mutation at the onset of the study and were used to assess the specificity and sensitivity of the assay. The remaining samples were from unsolved cases that were biased toward more phenotypically and genotypically complicated cases. RESULTS The assay was able to detect the mutation in 117 (96%) of 122 positive control subjects with known causal mutations. For the unsolved cases, our assay resulted in a molecular genetic diagnosis for 35 of 139 patients. Interestingly, most of these cases represented atypical clinical presentations of known PIDs. CONCLUSIONS The targeted NGS PID gene panel is a sensitive and cost-effective diagnostic tool that can be used as a first-line molecular assay in patients with PIDs. The assay is an alternative choice to the complex and costly candidate gene approach, particularly for patients with atypical presentation of known PID genes.

A European spectrum of pharmacogenomic biomarkers: Implications for clinical pharmacogenomics

Pharmacogenomics aims to correlate inter-individual differences of drug efficacy and/or toxicity with the underlying genetic composition, particularly in genes encoding for protein factors and enzymes involved in drug metabolism and transport. In several European populations, particularly in countries with lower income, information related to the prevalence of pharmacogenomic biomarkers is incomplete or lacking. Here, we have implemented the microattribution approach to assess the pharmacogenomic biomarkers allelic spectrum in 18 European populations, mostly from developing European countries, by analyzing 1,931 pharmacogenomics biomarkers in 231 genes. Our data show significant inter-population pharmacogenomic biomarker allele frequency differences, particularly in 7 clinically actionable pharmacogenomic biomarkers in 7 European populations, affecting drug efficacy and/or toxicity of 51 medication treatment modalities. These data also reflect on the differences observed in the prevalence of high-risk genotypes in these populations, as far as common markers in the CYP2C9, CYP2C19, CYP3A5, VKORC1, SLCO1B1 and TPMT pharmacogenes are concerned. Also, our data demonstrate notable differences in predicted genotype-based warfarin dosing among these populations. Our findings can be exploited not only to develop guidelines for medical prioritization, but most importantly to facilitate integration of pharmacogenomics and to support pre-emptive pharmacogenomic testing. This may subsequently contribute towards significant cost-savings in the overall healthcare expenditure in the participating countries, where pharmacogenomics implementation proves to be cost-effective.

Syndromic congenital sensorineural deafness, microtia and microdontia resulting from a novel homoallelic mutation in fibroblast growth factor 3 (FGF3)

We identified a homozygous missense mutation (c.196G → T) in fibroblast growth factor 3 (FGF3) in 21 affected individuals from a large extended consanguineous Saudi family, phenotypically characterized by autosomal recessive syndromic congenital sensorineural deafness, microtia and microdontia. All affected family members are descendents of a common ancestor who had lived six generations ago in a geographically isolated small village. This is the second report of FGF3 involvement in syndromic deafness in humans, and independently confirms the genes positive role in inner ear development. The c.196G → T mutation results in substitution of glycine by cysteine at amino acid 66 (p.G66C). This residue is conserved in several species and across 18 FGF family members. Conserved glycine/proline residues are central to the ‘β-trefoil fold’ characteristic of the secondary structure of FGF family proteins and substitution of these residues is likely to disrupt structure and consequently function.

Mendelian randomisation analysis strongly implicates adiposity with risk of developing colorectal cancer

Background:Observational studies have associated adiposity with an increased risk of colorectal cancer (CRC). However, such studies do not establish a causal relationship. To minimise bias from confounding we performed a Mendelian randomisation (MR) analysis to examine the relationship between adiposity and CRC.Methods:We used SNPs associated with adult body mass index (BMI), waist-hip ratio (WHR), childhood obesity and birth weight as instrumental variables in a MR analysis of 9254 CRC cases and 18 386 controls.Results:In the MR analysis, the odds ratios (ORs) of CRC risk per unit increase in BMI, WHR and childhood obesity were 1.23 (95% CI: 1.02–1.49, P=0.033), 1.59 (95% CI: 1.08–2.34, P=0.019) and 1.07 (95% CI: 1.03–1.13, P=0.018), respectively. There was no evidence for association between birth weight and CRC (OR=1.22, 95% CI: 0.89–1.67, P=0.22). Combining these data with a concurrent MR-based analysis for BMI and WHR with CRC risk (totalling to 18 190 cases, 27 617 controls) provided increased support, ORs for BMI and WHR were 1.26 (95% CI: 1.10–1.44, P=7.7 × 10−4) and 1.40 (95% CI: 1.14–1.72, P=1.2 × 10−3), respectively.Conclusions:These data provide further evidence for a strong causal relationship between adiposity and the risk of developing CRC highlighting the urgent need for prevention and treatment of adiposity.

Accelerating matchmaking of novel dysmorphology syndromes through clinical and genomic characterization of a large cohort

Purpose:Dysmorphology syndromes are among the most common referrals to clinical genetics specialists. Inability to match the dysmorphology pattern to a known syndrome can pose a major diagnostic challenge. With an aim to accelerate the establishment of new syndromes and their genetic etiology, we describe our experience with multiplex consanguineous families that appeared to represent novel autosomal recessive dysmorphology syndromes at the time of evaluation.Methods:Combined autozygome/exome analysis of multiplex consanguineous families with apparently novel dysmorphology syndromes.Results:Consistent with the apparent novelty of the phenotypes, our analysis revealed a strong candidate variant in genes that were novel at the time of the analysis in the majority of cases, and 10 of these genes are published here for the first time as novel candidates (CDK9, NEK9, ZNF668, TTC28, MBL2, CADPS, CACNA1H, HYAL2, CTU2, and C3ORF17). A significant minority of the phenotypes (6/31, 19%), however, were caused by genes known to cause Mendelian phenotypes, thus expanding the phenotypic spectrum of the diseases linked to these genes. The conspicuous inheritance pattern and the highly specific phenotypes appear to have contributed to the high yield (90%) of plausible molecular diagnoses in our study cohort.Conclusion:Reporting detailed clinical and genomic analysis of a large series of apparently novel dysmorphology syndromes will likely lead to a trend to accelerate the establishment of novel syndromes and their underlying genes through open exchange of data for the benefit of patients, their families, health-care providers, and the research community.Genet Med 18 7, 686–695.

A genome-wide association study reveals susceptibility loci for myocardial infarction/coronary artery disease in Saudi Arabs

BACKGROUND Multiple loci have been identified for coronary artery disease (CAD) by genome-wide association studies (GWAS), but no such studies on CAD incidence has been reported yet for any Middle Eastern population. METHODS In this study, we performed a GWAS for CAD and myocardial infarction (MI) incidence in 5668 Saudis of Arab descent using the Affymetrix Axiom Genotyping platform. RESULTS We describe SNPs at 16 loci that showed significant (P < 5 × 10(-8)) or suggestive GWAS association (P < 1 × 10(-5)) with CAD or MI, in the ethnic Saudi Arab population. Among the four variants reaching GWAS significance in the present study, the rs10738607_G [0.78(0.71-0.85); p = 2.17E-08] in CDNK2A/B gene was associated with CAD. Two other SNPs on the same gene, rs10757274_G [0.79(0.73-0.86); p = 2.98E-08] and rs1333045_C [0.79(0.73-0.86); p = 1.15E-08] as well as the rs9982601_T [1.38(1.23-1.55); p = 3.49E-08] on KCNE2 were associated with MI. These variants have been previously described in other populations. Several SNPs, including the rs7421388 (PLCL1) and rs12541758 (TRPA1) displaying a suggestive GWAS association (P < 1 × 10(-5)) with CAD as well as rs41411047 (RNF13), rs32793 (PDZD2) and rs4739066 (YTHDF3), similarly showing weak association with MI, were confirmed in an independent dataset. Furthermore, our estimation of heritability of CAD and MI based on observed genome-wide sharing in unrelated Saudi Arabs was approximately 33% and 44%, respectively. CONCLUSIONS Our study has identified susceptibility variants for CAD/MI in ethnic Arabs. These findings provide further insights into pathways contributing to the susceptibility for CAD and will enable more comprehensive genetic studies of these diseases in Middle East populations.