Next-generation sequencing (NGS) is a powerful tool for the diagnosis of inborn errors of immunity (IEIs). However, one of the main limitations is the risk of identifying inconclusive results. In this context, we estimated the overall diagnostic rate of different NGS techniques, comparing the rate of targeted panels (TP), mendeliome trio, and whole-exome sequencing (WES). Moreover, we evaluated the ability of the recently developed PIDCAP score to predict the diagnosis of IEIs by comparing the diagnostic rate of NGS among different categories of risk.
198 patients with suspected IEIs were genetically investigated using either TP (73%), mendeliome trio (10%), or WES (17%). The genetic variants were classified according to 2015 ACMG standard guidelines and filtered based on frequency and category. The analysis was limited to coding and splicing variants with a minor allele frequency (MAF) or equal to 1% in internal and public databases. The PIDCAP scoring system was used to stratify the cohort in risk categories: >75 high, 35-75 moderate, and <35 low risk.
192 variants in 136 genes were identified, including 67 (52%) VUS, 28 (22%) likely pathogenic, 26 (20%) pathogenic, 4 (3%) reclassified as benign or likely benign, and 4 (3%) with conflicting interpretation. The remaining 63 have not been previously reported in the databases. The number of reported VUS was significantly lower for mendeliome trio compared with TP (p 0.03) and WES (p 0.00004). The diagnostic rate considering all techniques is 17%, and it was significantly higher for WES (34%) and mendeliome (38%) compared with TP (12.5%). In cases with high suspicion, the diagnostic rate rose to 34% in the high-risk group and to 21% in the medium-risk group vs 1.3% in the low-risk group, whereas when the clinical picture met ESID diagnostic criteria up to 78%. Interestingly, the analysis of the patients with medium risk revealed alterations in non-immune genes in 5/7 cases.
The analysis of many genes leads to the identification of a large number of variants, and the trio analysis facilitates the attribution of pathogenicity. The diagnostic rate is higher when the analysis is limited to patients with high suspicion.
