Variants in nicotinamide adenine dinucleotide phosphate oxidase complex components determine susceptibility to very early onset inflammatory bowel disease.

BACKGROUND & AIMS The colitis observed in patients with very early onset inflammatory bowel disease (VEOIBD; defined as onset of disease at younger than 6 years of age) often resembles that of chronic granulomatous disease (CGD) in extent and features of colonic inflammation observed by endoscopy and histology. CGD is a severe immunodeficiency caused by defects in the genes that encode components of the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase complex. We investigated whether variants in genes that encode NADPH oxidase components affect susceptibility to VEOIBD using independent approaches. METHODS We performed targeted exome sequencing of genes that encode components of NADPH oxidases (cytochrome b light chain and encodes p22(phox) protein; cytochrome b-245 or NADPH oxidase 2, and encodes Nox2 or gp91(phox); neutrophil cytosol factor 1 and encodes p47 (phox) protein; neutrophil cytosol factor 2 and encodes p67 (phox) protein; neutrophil cytosol factor 4 and encodes p40 (phox) protein; and Ras-related C3 botulinum toxin substrate 1 and 2) in 122 patients with VEOIBD diagnosed at The Hospital for Sick Children, University of Toronto, from 1994 through 2012. Gene variants were validated in an independent International Early Onset Pediatric IBD Cohort Study cohort of patients with VEOIBD. In a second approach, we examined Tag single nucleotide polymorphisms in a subset of patients with VEOIBD in which the NOX2 NADPH oxidase genes sequence had been previously analyzed. We then looked for single nucleotide polymorphisms associated with the disease in an independent International Early Onset Pediatric IBD Cohort Study cohort of patients. We analyzed the functional effects of variants associated with VEOIBD. RESULTS Targeted exome sequencing and Tag single nucleotide polymorphism genotyping identified 11 variants associated with VEOIBD; the majority of patients were heterozygous for these variants. Expression of these variants in cells either reduced oxidative burst or altered interactions among proteins in the NADPH oxidase complex. Variants in the noncoding regulatory and splicing elements resulted in reduced levels of proteins, or expression of altered forms of the proteins, in blood cells from VEOIBD patients. CONCLUSIONS We found that VEOIBD patients carry heterozygous functional hypomorphic variants in components of the NOX2 NADPH oxidase complex. These do not cause overt immunodeficiency, but instead determine susceptibility to VEOIBD. Specific approaches might be developed to treat individual patients based on their genetic variant.

[1]  E. Schadt,et al.  Mutations in tetratricopeptide repeat domain 7A result in a severe form of very early onset inflammatory bowel disease. , 2014, Gastroenterology.

[2]  C. Thoeni,et al.  A Novel Nonsense Mutation in the EpCAM Gene in a Patient With Congenital Tufting Enteropathy , 2014, Journal of pediatric gastroenterology and nutrition.

[3]  M. Silverberg,et al.  IL-10R Polymorphisms Are Associated with Very-early-onset Ulcerative Colitis , 2013, Inflammatory bowel diseases.

[4]  M. Behr,et al.  Crohn’s as an immune deficiency: from apparent paradox to evolving paradigm , 2013, Expert review of clinical immunology.

[5]  David C. Wilson,et al.  Host-microbe interactions have shaped the genetic architecture of inflammatory bowel disease , 2012, Nature.

[6]  M. Silverberg,et al.  The NOD2insC polymorphism is associated with worse outcome following ileal pouch-anal anastomosis for ulcerative colitis , 2012, Gut.

[7]  S. Snapper,et al.  The age of gene discovery in very early onset inflammatory bowel disease. , 2012, Gastroenterology.

[8]  R. Holmdahl,et al.  Copy number variation of the gene NCF1 is associated with rheumatoid arthritis. , 2012, Antioxidants & redox signaling.

[9]  Don L. Armstrong,et al.  Lupus-associated causal mutation in neutrophil cytosolic factor 2 (NCF2) brings unique insights to the structure and function of NADPH oxidase , 2011, Proceedings of the National Academy of Sciences.

[10]  Judy H. Cho,et al.  NADPH oxidase complex and IBD candidate gene studies: identification of a rare variant in NCF2 that results in reduced binding to RAC2 , 2011, Gut.

[11]  Judy H. Cho,et al.  Single nucleotide polymorphisms that increase expression of the guanosine triphosphatase RAC1 are associated with ulcerative colitis. , 2011, Gastroenterology.

[12]  R. Xavier,et al.  Genetics and pathogenesis of inflammatory bowel disease , 2011, Nature.

[13]  David C Wilson,et al.  Pediatric modification of the Montreal classification for inflammatory bowel disease: The Paris classification , 2011, Inflammatory bowel diseases.

[14]  J. Casanova,et al.  Germline CYBB mutations that selectively affect macrophages in kindreds with X-linked predisposition to tuberculous mycobacterial disease , 2011, Nature Immunology.

[15]  Tariq Ahmad,et al.  Meta-analysis identifies 29 additional ulcerative colitis risk loci, increasing the number of confirmed associations to 47 , 2011, Nature Genetics.

[16]  Mostafa Ronaghi,et al.  pfSNP: An integrated potentially functional SNP resource that facilitates hypotheses generation through knowledge syntheses , 2011, Human mutation.

[17]  S. McCartney,et al.  The Neutrophil Respiratory Burst and Bacterial Digestion in Crohn’s Disease , 2011, Digestive Diseases and Sciences.

[18]  Tariq Ahmad,et al.  Genome-wide meta-analysis increases to 71 the number of confirmed Crohn's disease susceptibility loci , 2010, Nature Genetics.

[19]  Athos Bousvaros,et al.  Innate and adaptive immune connections in inflammatory bowel diseases , 2010, Current opinion in gastroenterology.

[20]  P. Bork,et al.  A method and server for predicting damaging missense mutations , 2010, Nature Methods.

[21]  Douglas R. McDonald,et al.  Small-molecule screen identifies reactive oxygen species as key regulators of neutrophil chemotaxis , 2010, Proceedings of the National Academy of Sciences.

[22]  A. Schäffer,et al.  Inflammatory bowel disease and mutations affecting the interleukin-10 receptor. , 2009, The New England journal of medicine.

[23]  Xing Jun Li,et al.  A new genetic subgroup of chronic granulomatous disease with autosomal recessive mutations in p40 phox and selective defects in neutrophil NADPH oxidase activity. , 2009, Blood.

[24]  T. To,et al.  Increasing incidence of paediatric inflammatory bowel disease in Ontario, Canada: evidence from health administrative data , 2009, Gut.

[25]  C. Béroud,et al.  Human Splicing Finder: an online bioinformatics tool to predict splicing signals , 2009, Nucleic acids research.

[26]  M. Novelli,et al.  Inflammatory Bowel Disease in CGD Reproduces the Clinicopathological Features of Crohn's Disease , 2009, The American Journal of Gastroenterology.

[27]  S. Henikoff,et al.  Predicting the effects of coding non-synonymous variants on protein function using the SIFT algorithm , 2009, Nature Protocols.

[28]  T. Merriman,et al.  Confirmation of association of IRGM and NCF4 with ileal Crohn's disease in a population-based cohort , 2008, Genes and Immunity.

[29]  R. Shamir,et al.  Pediatric onset Crohn's colitis is characterized by genotype‐dependent age‐related susceptibility , 2007, Inflammatory bowel diseases.

[30]  Judy H Cho,et al.  Genome-wide association study identifies new susceptibility loci for Crohn disease and implicates autophagy in disease pathogenesis , 2007, Nature Genetics.

[31]  Chia-Hung Liu,et al.  FASTSNP: an always up-to-date and extendable service for SNP function analysis and prioritization , 2006, Nucleic Acids Res..

[32]  William D. Lees,et al.  Defective acute inflammation in Crohn's disease: a clinical investigation , 2006, The Lancet.

[33]  Jeannie S. Huang,et al.  Chronic granulomatous disease caused by a deficiency in p47(phox) mimicking Crohn's disease. , 2004, Clinical gastroenterology and hepatology : the official clinical practice journal of the American Gastroenterological Association.

[34]  Lin Li,et al.  Gene regulation by Sp1 and Sp3. , 2004, Biochemistry and cell biology = Biochimie et biologie cellulaire.

[35]  A. Griffiths.  Specificities of inflammatory bowel disease in childhood. , 2004, Best practice & research. Clinical gastroenterology.

[36]  Jeannie S. Huang,et al.  Chronic granulomatous disease caused by a deficiency in p47(phox) mimicking Crohn's disease. , 2004, Clinical gastroenterology and hepatology : the official clinical practice journal of the American Gastroenterological Association.

[37]  Jens Holmberg,et al.  Positional identification of Ncf1 as a gene that regulates arthritis severity in rats , 2003, Nature Genetics.

[38]  G. Bokoch,et al.  Molecular basis for Rac2 regulation of phagocyte NADPH oxidase , 2001, Nature Immunology.

[39]  D. Goldblatt,et al.  Colitis in chronic granulomatous disease , 2001, Archives of disease in childhood.

[40]  J. Lekstrom‐Himes,et al.  Immunodeficiency diseases caused by defects in phagocytes. , 2000, The New England journal of medicine.