Next-Generation Sequencing in the Field of Primary Immunodeficiencies: Current Yield, Challenges, and Future Perspectives

Abstract

Primary immunodeficiencies comprise a group of inborn errors of immunity that display significant clinical and genetic heterogeneity. Next-generation sequencing techniques and predominantly whole exome sequencing have revolutionized the understanding of the genetic and molecular basis of genetic diseases, thereby also leading to a sharp increase in the discovery of new genes associated with primary immunodeficiencies. In this review, we discuss the current diagnostic yield of this generic diagnostic approach by evaluating the studies that have employed next-generation sequencing techniques in cohorts of patients with primary immunodeficiencies. The average diagnostic yield for primary immunodeficiencies is determined to be 29% (range 10-79%) and 38% specifically for whole-exome sequencing (range 15-70%). The significant variation between studies is mainly the result of differences in clinical characteristics of the studied cohorts but is also influenced by varying sequencing approaches and (in silico) gene panel selection. We further discuss other factors contributing to the relatively low yield, including the inherent limitations of whole-exome sequencing, challenges in the interpretation of novel candidate genetic variants, and promises of exploring the non-coding part of the genome. We propose strategies to improve the diagnostic yield leading the way towards expanded personalized treatment in PIDs.

Keywords: Diagnostic yield; Next-generation sequencing; Precision medicine; Primary immunodeficiencies; Whole exome sequencing.

Fig. 1

Schematic overview of the mutational mechanisms and effects on gene, protein, and pathway level with the phenotypic manifestations that result from various forms of PID. For autosomal recessive, X-linked and autosomal dominant forms of PID, the effects on the gene level and subsequently on the protein level are indicated that lead to either deficiency or hyperactivation of the immunological pathways involved. These effects at the pathway level can either result in a phenotype manifesting with symptoms of immunodeficiency, autoimmunity, or autoinflammation. Pathogen exposure can be a prerequisite for the phenotype to develop, which contributes to the incomplete penetrance observed in PIDs.

Fig. 2

The general steps used to filter variants from WES. The approximate number of variants in each step is indicated. All filtering steps can be applied in silico. The number of variants that remain after filtering depends on the cut-off values used for filtering based on allele frequency and on algorithms used to call CNVs [3, 19, 26]. CNV numbers are highly dependent on the algorithm used; therefore, the number of CNVs is not indicated here. For the analysis of de novo variants, sequencing of a patient’s parents as a trio is required, after which all variants present in the parents can be filtered [3].

Fig. 3

The relationship between variant effect size, allele frequency, and the diagnostic success rate in the field of PIDs. The triangles indicate variant effect, allele frequency, and diagnostic yield, ranging from highly impacting to weakly impacting, from extremely rare to common and from high to low, respectively. The characteristics of (severe) combined immunodeficiency ((S)CID), common variable immunodeficiency (CVID), and common disease are indicated in their approximate location within these three indicators