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

Abstract

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.

Keywords: genetics and development; human genetics; pediatric nephrology; vesico-ureteral reflux.

Copyright © 2021 by the American Society of Nephrology.

Figures

Graphical abstract

Figure 1.

Excess burden of large, rare CNVs in VUR. VUR patients are enriched in large (size ≥100 kb), rare (frequency in PCA-defined populations ≤0.1%), gene-intersecting CNVs. (A) Frequency of subjects with their largest CNV size at or above the indicated size thresholds (kb). (B) Proportion of CNV (size ≥100 kb) within the indicated size intervals (kb). The 100–500 kb interval is not shown. Blue bars represent VUR patients and black bars, controls.

Figure 2.

Regional association plots for loci associated with VUR. The calculated −log10 P values are shown according to genomic position (hg19). The most significant SNP is colored purple. Other colors indicate LD (r2; see inset) to top SNP, based on the 1000 Genomes European sample set.

Figure 3.

Wnt5a expression and its role in lower urinary tract morphogenesis. (A–E) In situ hybridization analysis. (A) Wnt5a expression at E9 in a whole mount embryo. (B) Section through a whole mount embryo showing Wnt5a expression surrounding the cloaca. (C–E) Wnt5a expression in sagittal sections from embryos at E11 (C), E12 (D), and E15 (E). (F and N) Whole-mount urogenital tracts from a control (F) and Wnt5a a mutant (N). (G and O) Hematoxylin and eosin–stained sections through the bladder/urethra and hindgut of controls (G) and Wnt5a mutants (O). Green arrowheads point to the nephric ducts. Yellow arrowhead in (O) points to the distal ureter. (H and P) Schematic representation of histology shown in (G and O). (I and Q) Sagittal sections through Wnt5a mutants at E12. (J and R) Sagittal sections through a control (J) and Wnt5a mutant embryo (R). (K, L, S, and T) Ecad staining (green) reveals nephric duct and cloacal epithelia at E9, in controls (K and L) and Wnt5a mutants (L and T). (M and U) Schematic showing nephric duct insertion into the cloaca in a wild-type embryo (M) and in a Wnt5a mutant embryo (U). bl, bladder; cl, cloaca; cl* truncated cloaca in Wnt5a mutant; ki, kidney; nd, nephric duct; ur, ureter; wd, Wolffian duct.