Copy Number Variant Analysis and Genome-wide Association Study Identify Loci with Large Effect for Vesicoureteral Reflux.

BACKGROUND: Vesicoureteral reflux (VUR) is a common, familial genitourinary disorder, and a major cause of pediatric urinary tract infection (UTI) and kidney failure. The genetic basis of VUR is not well understood. METHODS: A diagnostic analysis sought rare, pathogenic copy number variant (CNV) disorders among 1737 patients with VUR. A GWAS was performed in 1395 patients and 5366 controls, of European ancestry. RESULTS: Altogether, 3% of VUR patients harbored an undiagnosed rare CNV disorder, such as the 1q21.1, 16p11.2, 22q11.21, and triple X syndromes ((OR, 3.12; 95% CI, 2.10 to 4.54; P=6.35×10-8) The GWAS identified three study-wide significant and five suggestive loci with large effects (ORs, 1.41-6.9), containing canonical developmental genes expressed in the developing urinary tract (WDPCP, OTX1, BMP5, VANGL1, and WNT5A). In particular, 3.3% of VUR patients were homozygous for an intronic variant in WDPCP (rs13013890; OR, 3.65; 95% CI, 2.39 to 5.56; P=1.86×10-9). This locus was associated with multiple genitourinary phenotypes in the UK Biobank and eMERGE studies. Analysis of Wnt5a mutant mice confirmed the role of Wnt5a signaling in bladder and ureteric morphogenesis. CONCLUSIONS: These data demonstrate the genetic heterogeneity of VUR. Altogether, 6% of patients with VUR harbored a rare CNV or a common variant genotype conferring an OR >3. Identification of these genetic risk factors has multiple implications for clinical care and for analysis of outcomes in VUR.

Urinary tract effects of HPSE2 mutations.

Urofacial syndrome (UFS) is an autosomal recessive congenital disease featuring grimacing and incomplete bladder emptying. Mutations of HPSE2, encoding heparanase 2, a heparanase 1 inhibitor, occur in UFS, but knowledge about the HPSE2 mutation spectrum is limited. Here, seven UFS kindreds with HPSE2 mutations are presented, including one with deleted asparagine 254, suggesting a role for this amino acid, which is conserved in vertebrate orthologs. HPSE2 mutations were absent in 23 non-neurogenic neurogenic bladder probands and, of 439 families with nonsyndromic vesicoureteric reflux, only one carried a putative pathogenic HPSE2 variant. Homozygous Hpse2 mutant mouse bladders contained urine more often than did wild-type organs, phenocopying human UFS. Pelvic ganglia neural cell bodies contained heparanase 1, heparanase 2, and leucine-rich repeats and immunoglobulin-like domains-2 (LRIG2), which is mutated in certain UFS families. In conclusion, heparanase 2 is an autonomic neural protein implicated in bladder emptying, but HPSE2 variants are uncommon in urinary diseases resembling UFS.

Heterozygous non-synonymous ROBO2 variants are unlikely to be sufficient to cause familial vesicoureteric reflux.

ROBO2, the receptor of SLIT2, is one of many genes/proteins that regulate the outgrowth of the ureteric bud, which is the first step in the development of the metanephric urinary system. Non-synonymous variants in ROBO2 have been found in a small proportion of patients with primary vesicoureteric reflux (VUR) in various countries. Here we sequenced 1 kb of promoter and all exons of ROBO2b with intronic margins in 227 index cases with primary VUR in an Irish population and found 55 variants, of which 20 were novel. We assessed the variants for evolutionary conservation and investigated novel and uncommon known conserved variants in 23 further index cases and family members of all index cases (to check for segregation with VUR), and then in healthy controls if we found segregation of the variants with VUR. Apart from one non-synonymous variant that was previously found in controls, we did not find any of the six other previously reported non-synonymous variants, but found four new non-synonymous variants. Of those, only two segregated with the disorder (p.Pro522Thr and p.Val799Ile). The former was not present in any of 592 healthy controls; the latter was present in one control. There are now 35 reported non-synonymous coding variants of ROBO2b. The predicted pathogenicity of those that have so far been found exclusively in VUR patients does not differ from that predicted for those variants also found in controls. Thus, our finding does not completely rule out that some variants may be the sole cause of VUR, but it is clear from the overall frequency that most of them cannot be. However, it is possible that some of these variants may cause VUR in combination with a mutation in another gene.

Investigation of DNA variants specific to ROBO2 Isoform 'a' in Irish vesicoureteric reflux patients reveals marked CpG island variation.

ROBO2 gene disruption causes vesicoureteric reflux (VUR) amongst other congenital anomalies. Several VUR patient cohorts have been screened for variants in the ubiquitously expressed transcript, ROBO2b, but, apart from low levels in a few adult tissues, ROBO2a expression is confined to the embryo, and might be more relevant to VUR, a developmental disorder. ROBO2a has an alternative promoter and two alternative exons which replace the first exon of ROBO2b. We screened probands from 251 Irish VUR families for DNA variants in these. The CpG island of ROBO2a, which includes the non-coding first exon, was found to contain a run of six variants abolishing/creating CpG dinucleotides, including a novel variant, present in the VUR cases in one family, that was not present in 592 healthy Irish controls. In three of these positions, the CpG was created by the non-reference allele, and the reference allele was not the nucleotide that would result from spontaneous deamination of methylcytosine to thymine, suggesting that there might have been selection for variability in number of CpGs in this island. This is in marked contrast to the CpG island at the start of ROBO2b, which only contained a single variant that abolishes a CpG.

Publisher Correction: Genome-wide linkage and association study implicates the 10q26 region as a major genetic contributor to primary nonsyndromic vesicoureteric reflux.

A correction to this article has been published and is linked from the HTML version of this paper. The error has been fixed in the paper.

Genome-wide linkage and association study implicates the 10q26 region as a major genetic contributor to primary nonsyndromic vesicoureteric reflux.

Vesicoureteric reflux (VUR) is the commonest urological anomaly in children. Despite treatment improvements, associated renal lesions - congenital dysplasia, acquired scarring or both - are a common cause of childhood hypertension and renal failure. Primary VUR is familial, with transmission rate and sibling risk both approaching 50%, and appears highly genetically heterogeneous. It is often associated with other developmental anomalies of the urinary tract, emphasising its etiology as a disorder of urogenital tract development. We conducted a genome-wide linkage and association study in three European populations to search for loci predisposing to VUR. Family-based association analysis of 1098 parent-affected-child trios and case/control association analysis of 1147 cases and 3789 controls did not reveal any compelling associations, but parametric linkage analysis of 460 families (1062 affected individuals) under a dominant model identified a single region, on 10q26, that showed strong linkage (HLOD = 4.90; ZLRLOD = 4.39) to VUR. The ~9Mb region contains 69 genes, including some good biological candidates. Resequencing this region in selected individuals did not clearly implicate any gene but FOXI2, FANK1 and GLRX3 remain candidates for further investigation. This, the largest genetic study of VUR to date, highlights the 10q26 region as a major genetic contributor to VUR in European populations.

Emerging technologies for point-of-care genetic testing.

In the coming years, genetic test results will be increasingly used as indicators that influence medical decision making. Novel instrumentation that is able to detect relevant mutations in a point-of-care setting is being developed to facilitate this increase, frequently as a spin-off from recent research in the area of biothreat monitoring. This market review will describe the current generation of instrumentation that is most suitable for use in a point-of-care setting; it will also try to identify some of the technologies that will make-up the next generation of instrumentation currently being prepared for the market.