Rolling the Dice on Crohn’s Disease

Researchers worldwide use genomic tools to gain ground in the fight against Crohn’s Disease and other common human diseases.

click to enlarge   Shown here is a typical readout from the Illumina genotyping array used in some of the Crohn’s Disease genome association studies. (Source: Struan Grant, PhD)

Crohn’s Disease is one of many inflammatory bowel diseases and is characterized by abdominal pain, diarrhea, rectal bleeding, weight loss, arthritis, skin problems, and fever. Inflammation in the intestines and bowel are responsible for the swelling, abdominal pain, and rectal bleeding that are hallmarks of Crohn’s. Anti-inflammatory agents are prescribed to treat the inflammation-associated symptoms of the disease.

Crohn’s disease is typically diagnosed in persons 20 to 30 years of age, with one-quarter of new diagnoses occurring in persons under the age of 20 (www.genome.gov/pfv.cfm?pageID=25521854). Although the pathogenesis of Crohn’s disease is not completely understood, it is believed to have both genetic and environmental factors, which seem to synergize in the pathogenic process.

One environmental factor currently under investigation is the association between gut bacteria and Crohn’s disease. Marcel A. Behr, MD, M.Sc., FRCPC, associate professor, McGill University, Montreal, Canada, is studying this association. “That these bacteria are part of the etiology of Crohn’s is the driving hypothesis of my research,” says Behr. Behr has performed bacterial microarrays to do comparative genomics, determine the genomic differences between Mycobacterium avium-paratuberculosis and Mycobacterium avium, and to determine the genomic identity of particular bacteria.

“So in that sense, when I have been approaching Crohn’s disease, I have not been saying I am going to throw Crohn’s tissue on an array. I am going to try to find out what is the bug and what are the sequences unique to the bug that we could use as either molecular or immunologic targets,” says Behr. In fact, Behr is not performing human arrays to study Crohn’s at all. He only sees the data output.

“Where I think the biggest advances in Crohn’s disease have been is in diagnostic genome association studies of Crohn’s patients,” says Behr. The studies Behr is referring to were reported in a now-famous Nature Genetics article (Nat Genet. 2007 Nov; 39(11):1329-37). Behr points out that some of the genes found through these studies to be associated with Crohn’s Disease, including NOD2, ATG-16L1, IL-23, and LRG-47, suggest defective innate immune responses to intracellular bacterial products in the Crohn’s host. Behr is trying to identify the nature of this suggested defect, i.e., whether it is a generic defect in the innate immunity against all bacteria or just a few specific bacteria.

Genetics of Crohn’s
Crohn’s Disease seems to run in families, especially in those with Jewish ancestry. The lifetime risk for the likelihood of Crohn’s disease in children with at least one affected parent is 7% to 9% (www.genome.gov/pfv.cfm?pageID=25521854). There is also a 10% chance that these children will develop an additional inflammatory bowel disease. For children of two parents affected by inflammatory bowel disease, the lifetime risk of developing Crohn’s Disease is 35%.

Researchers worldwide are gaining a better understanding of the genetic component of Crohn’s Disease. For example, the aforementioned Nature Genetics paper was the result of a multinational study aimed at identifying single nucleotide polymorhisms (SNPs) associated with a number of autoimmune diseases, including Crohn’s disease. One of the authors on that paper was Miles Parkes, MD, consultant gastroenterologist, Addenbrooke’s Hospital, Cambridge, UK. To find polymorphisms associated with Crohn’s, Parkes, in conjunction with the Wellcome Trust in the UK (as well as other groups in Europe and North America) performed genome-wide association studies. Parkes published his groups’ results as part of the Wellcome Trust Case Control Consortium (WTCCC). “The study looked at not just Crohn’s disease but also six other common complex diseases. But I would say Crohn’s disease was, by some distance, the winner in terms of producing interesting data from that,” says Parkes.

Two and a half years ago, when this study began, it was an enormous undertaking. Roughly 17,000 DNA samples, isolated from blood samples of populations of patients stratified into “control” and “case” groups, had to be analyzed. In terms of experimental design, the WTCCC study was performed using 2,000 cases and 3,000 controls. Parkes used the Affymetrix 500K platform to perform the genotyping, whereas the North Americans and Europeans used the Illumina 300K chip. “And while that looks fairly tractable by today’s standard, do bear in mind that this was being set up 2.5 years ago, at a time when the technology was much less certain, a lot of it had not been tested at that point,” says Parkes. “There was a lot of debate about the design of the chips in terms of the coverage of the genome. Obviously, there are advantages of the Illumina chip in terms of its design with HapMap SNPs and its deliberate design with a tagging strategy compared to the Affymetrix 500 chip, which did not follow that design. For some reason, we were able to get around that design by the shear size of the experiment we were doing with the WTCCC.”

Parkes is now conducting a meta-analysis of the Wellcome Trust data alongside the European and North American data. For this study, the number of cases and controls increased to 3200 and 4800, respectively. The meta-analysis has eliminated another 20 new Crohn’s disease susceptibility genes. Some of the major new Crohn’s-associated genes that have come out over the last year to 18 months are the IL-23R, IL-12B, IL-23, IL-12, and some autophagy genes (ATG-16L1 and IRGM). “I think [the autophagy] genes are intriguing in the sense that there are two separate genes within the same pathway, previously unsuspected as playing a role in Crohn’s disease pathogenesis,” says Parkes.

As a result of the all of the WTCCC study and the subsequent meta-analysis, “there are now over 30 Crohn’s disease susceptibility disease genes and genetic loci with confirmed association with Crohn’s disease at unequivocal levels of statistical significance,” says Parkes. This study represents a great degree of progress for Crohn’s disease because there were only two known Crohn’s susceptibility genes 18 months ago.

“The genome-wide association approach has been a revolution since 2006,” says Struan Grant, PhD, associate director of the Center for Applied Genomics, Children’s Hospital of Philadelphia. Like Parkes, Grant is also involved in an inflammatory bowel disease study using Illumina SNP arrays to identify SNPs associated with these diseases.

“Before these chips came onto the market in mid-2006, you could either carry out candidate gene studies where you just took a set of your favorite genes and you studied them intensely, or you could do family/linkage studies,” says Grant. However, linkage data has been confusing and has only contributed to a minor degree to Crohn’s research. “It’s really only in the last 18 months that real genetic underpinnings of complex disease have been found, including major breakthroughs in inflammatory bowel disease.”

This article was published in Drug Discovery & Development magazine: Vol. 11, No. 3, March, 2008, pp. 44-45.

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Filed Under: Genomics/Proteomics

 

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