The 50 monogenic Selleckchem RG7422 defects associated with IBD provide an initial filter to identify patients with monogenic disorders. Because of the greatly reduced costs of next-generation sequencing, it is probably cost effective in many cases to perform multiplex gene sequencing, WES, or whole-genome sequencing rather than sequential Sanger sequencing of multiple genes. A big advantage of WES is the potential to identify novel causal genetic variants once the initial candidate filter list of known disease-causing candidates has been analyzed. The number of gene variants associated with VEOIBD is indeed constantly increasing, largely

due to the new sequencing technologies, so data sets derived from WES allow updated analysis of candidates as well as novel genes. Because multiple genetic defects can lead to spontaneous or induced colitis in mice,1 and 139 assuming homology, it is likely that many additional human gene variants will be associated with IBD. Targeted sequencing of genes of interest is an alternative approach to exome-targeted sequencing. Initial studies to perform targeted next-generation parallel sequencing showed the potential power of this approach.140 Targeted next-generation sequencing of the 170 primary immunodeficiency (PID)-related genes accurately detected point mutations and exonic deletions.140 Only 9 of 170 PID-related

genes analyzed showed inadequate coverage. Four of 26 patients with PID without an established prescreening genetic diagnosis, despite routine Sorafenib nmr functional and genetic testing, were diagnosed, RG7204 order indicating the advantage of parallel genetic screening. Because a major group of VEOIBD-causing variants is associated with PID-related genes, it is obvious how this approach can be adapted and extended to monogenic IBD genes. Genetic approaches also offer practical advantages. Specialized functional immune assays are often only available in research laboratories and are not necessarily validated; functional tests often require rapid processing of peripheral blood mononuclear cells or biopsy specimens

in specialized laboratories. This means that handling of DNA and sequencing seems far less prone to error or variation. However, relying solely on genetic screening can be misleading, because computational mutation prediction can fail to detect functional damaging variants. For example, variants in the protein-coding region of the IL10RA gene were misclassified as “tolerated” by certain prediction tools, whereas other prediction tools and functional analysis reported defects in IL-10 signaling. 30 Although most studies report variants in protein-coding regions in monogenic diseases, there could be selection bias. It is indeed far more difficult to establish the biological effects of variants that affect processes such as splicing, gene expression, or messenger RNA stability. It should go without saying that novel genetic variants require appropriate functional validation.