Chronic Granulomatous Disease via the CYBB Gene

Chronic granulomatous disease (CGD) an inherited immunodeficiency characterized by repeated infections with bacterial and fungal pathogens and formation of granulomas. CGD immunodeficiency is due to an impairment of the NADPH oxidase complex resulting in an inability to generate superoxide in phagocytic cells to lyse pathogens (Song et al. 2011). Common pathogens include Staphylococcus aureus, Pseudomonas species, Candida albicans, Aspergillus species, and Nocardia species. Pneumonia, granuloma formation within gastrointestinal and genitourinary tracts, and failure to thrive are hallmark symptoms of the disorder. In severe cases, granulomas can lead to abscess formation and organ failure. Treatments include long course antimicrobials to ward off infections (Leiding and Holland 2012). Simultaneous administration of antimicrobials and corticosteroids may be used to resolve colitis associated with heightened inflammatory responses to infection (Leiding et al. 2012). Patients with CGD should avoid areas where fungal spores are common such as mulch, gardens, and yard waste. Approximately one in 200,000 newborns in the US are affected with CGD (Winkelstein et al 2000). Genetic testing can aid in differential diagnosis of CGD from other disorders associated with granuloma formation and hyperinflammation such as cystic fibrosis, hyper IgE syndrome, Crohn’s disease, allergic bronchopulmonary aspergillosis, and glucose 6-phosphate dehydrogenase deficiency (Leiding and Holland 2012).

CGD is inherited in an X-linked manner through mutations in the CYBB gene. Autosomal recessive forms of CGD also occur through mutations in the CYBA, NCF1, NCF2, and NCF4 genes (Roos and de Boer 2014). Disease onset with individuals with X-linked CGD is ~3 years with mortality occurring in 20% compared to autosomal recessive forms with onset ~7 years and mortality occurring in 8% of cases. The majority of CYBB mutations are truncating with nonsense (30%), deletions (22%) and splice site (19.5%) mutations being causative for CGD (Roos et al 2010; Piirilä et al. 2006). Missense mutations are found in 20% of cases with mutations residing in amino acids 1-309 minimally affecting superoxide production and associated with good prognosis compared to missense mutations in amino acid 310 affecting FAD and NAPDH binding domains and rendering NADPH complex nonfunctional leading to worse prognosis (Leiding and Holland 2012). Insertions, gross deletions, and promoter mutations are present in less than 5% of cases of X-linked CGD. Contiguous gene deletions have been reported to span several genes leading to associations of Kell phenotype/Mcleod syndrome (XK gene), Duchenne muscular dystrophy (DMD gene), and retinitis pigmentosa (RPGR gene) with X-linked CGD (Brown et al. 1996; Watkins et al. 2011). Adult-onset of CGD has been reported in females heterozygous for CYBB mutations and is thought to occur through skewed X-inactivation (Gono et al. 2008; Anderson-Cohen et al. 2003). The CYBB gene encodes the gp91phox subunit of the NADPH oxidase complex. This complex is responsible for transporting electrons from NAPDH to oxygen to generate superoxide within the phagolysosome to facilitate lysis of pathogens in phagocytic cells such as neutrophils and macrophages (Song et al. 2011).

Mutations in the CYBB gene account for ~70% of cases of CGD (Leiding and Holand 2012; Roos et al. 2010). Analytical sensitivity is >95% for detection of causative mutations within the CYBB gene. Deletions of one or more exons are common and also detectable by Sanger sequencing in males. In females, analytical sensitivity is lower as we are unable to detect large heterozygous deletions by this method. Gross deletions of the CYBB gene have been reported in less than 5% of cases (Roos et al. 2010). Deletions have been reported to span several genes leading to associations of Kell phenotype/Mcleod syndrome (XK gene), Duchenne muscular dystrophy (DMD gene), and retinitis pigmentosa (RPGR gene) with X-linked CGD (Brown et al. 1996; Watkins et al. 2011).

This test provides full coverage of all coding exons of the CYBB gene plus 10 bases of flanking noncoding DNA in all available transcripts along with other non-coding regions in which pathogenic variants have been identified at PreventionGenetics or reported elsewhere. We define full coverage as >20X NGS reads or Sanger sequencing. PGnome panels typically provide slightly increased coverage over the PGxome equivalent. PGnome sequencing panels have the added benefit of additional analysis and reporting of deep intronic regions (where applicable).

Dependent on the sequencing backbone selected for this testing, discounted reflex testing to any other similar backbone-based test is available (i.e., PGxome panel to whole PGxome; PGnome panel to whole PGnome).