<p><strong>ABSTRACT</strong></p>
<p><strong>Introduction</strong>. To date abnormalities in more than 20 genes have been associated with SRNS. Sequencing of all SRNS genes requires ~ 600 PCR amplicons, rendering conventional mutation testing unfeasible due to financial and time constraints. Hence, current screening algorithms usually include only the most common disease genes and/or use preselection according to additional phenotypic criteria. This practice typically allows for mutation detection in ~15% of patients. We have evaluated targeted NGS screening of SRNS patients enrolled in the PodoNet registry who were found negative for mutations in the first-line SRNS-associated genes.</p>
<p><strong>Material and Methods</strong>. Molecular analysis of 31 known or plausible SDNS disease genes was performed by NGS using a custom-designed multiplex PCR kit (MASTR FSGS, Multiplicom). The pilot group consisted of 22 patients with 4.7 years median age at disease onset (range 0.5-20 years), positive family history in 68%, parental consanguinity in 18% and chronic renal insufficiency at last observation in 59%.</p>
<p><strong>Results.</strong> Mean coverage was 1269x (median 1286, range 929-1826). 18/22 runs had at least 15x read depth covering 99% of the target sequences. One patient was diagnosed with hereditary SRNS due to a previousy described homozygous pathogenic mutation in SMARCAL1 gene. In addition, three novel sequence variants in the genes PLCE1 (homozygous), LAMB2 (homozygous) and WT1 (heterozygous) were detected. In silico studies support their classification as pathogenic, even though the patients do not present the characteristic clinical and/or histopathological features typically reported for patients with mutations in these genes.</p>
<p><strong>Conclusions</strong>. Our detection of pathogenic mutations in 4 out of 22 SRNS patients screened negative by conventional selective screening approaches support targeted NGS testing in all SRNS patients, regardless of age at diagnosis, absence of extrarenal manifestations or histological subtype. We anticipate that systematic NGS screening of the SRNS cohorts collected in EURenOmics will allow re-evaluation of mutation incidence rates in SRNS and become the new standard of genetic diagnostics in this condition.</p
