A splice site variant in INPP5E causes diffuse cystic renal dysplasia and hepatic fibrosis in dogs

Ciliopathies presenting as inherited hepatorenal fibrocystic disorders are rare in humans and in dogs. We describe here a novel lethal ciliopathy in Norwich Terrier puppies that was diagnosed at necropsy and characterized as diffuse cystic renal disease and hepatic fibrosis. The histopathological findings were typical for cystic renal dysplasia in which the cysts were located in the straight portion of the proximal tubule, and thin descending and ascending limbs of Henle's loop. The pedigree of the affected puppies was suggestive of an autosomal recessive inheritance and therefore, whole exome sequencing and homozygosity mapping were used for identification of the causative variant. The analyses revealed a case-specific homozygous splice donor site variant in a cilia related gene, INPP5E: c.1572+5G>A. Association of the variant with the defect was validated in a large cohort of Norwich Terriers with 3 cases and 480 controls, the carrier frequency being 6%. We observed that the identified variant introduces a novel splice site in INPP5E causing a frameshift and formation of a premature stop codon. In conclusion, our results suggest that the INPP5E: c.1572+5G>A variant is causal for the ciliopathy in Norwich Terriers. Therefore, genetic testing can be carried out in the future for the eradication of the disease from the breed

Appropriate calibration curve fitting in ligand binding assays

Calibration curves for ligand binding assays are generally characterized by a nonlinear relationship between the mean response and the analyte concentration. Typically, the response exhibits a sigmoidal relationship with concentration. The currently accepted reference model for these calibration curves is the 4-parameter logistic (4-PL) model, which optimizes accuracy and precision over the maximum usable calibration range. Incorporation of weighting into the model requires additional effort but generally results in improved calibration curve performance. For calibration curves with some asymmetry, introduction of a fifth parameter (5-PL) may further improve the goodness of fit of the experimental data to the algorithm. Alternative models should be used with caution and with knowledge of the accuracy and precision performance of the model across the entire calibration range, but particularly at upper and lower analyte concentration areas, where the 4-and 5-PL algorithms generally outperform alternative models. Several assay design parameters, such as placement of calibrator concentrations across the selected range and assay layout on multiwell plates, should be considered, to enable optimal application of the 4- or 5-PL model. The fit of the experimental data to the model should be evaluated by assessment of agreement of nominal and model-predicted data for calibrators