Role of pathogen-driven selection in shaping the predisposition to IBD:

identification of disease susceptibility alleles

Mario (Mago) Clerici, M.D.Chair of Immunology

Head, PhD School in Molecular Medicine, Universita' di Milano

Scientific Director, IRCCS SM Nascente,

Fondazione Don C Gnocchi, Milano

Bardolino, January 16th, 2013

The evolutionary perspective for Inflammatory Diseases

Inflammatory/autoimmune diseases can have early onset (i.e. before reproductive age)

Inflammatory/autoimmune diseases have a strong genetic component

Inflammatory/autoimmune diseases have a relatively high prevalence in human populations

Why has evolution failed to eliminate the risk alleles?

1) Susceptibility alleles increased in frequency by genetic drift

2) Susceptibility alleles increased in frequency as a result of natural selection (they confer a selective advantage to the carriers; e.g. protection from infection) [hygiene hypothesis]

3) Risk alleles were neutral under different environmental conditions (e.g. high prevalence of infections/worms) [hygiene hypothesis]

Aims

Application of population genetic approaches to study the evolutionary history of inflammatory disease risk alleles in human populations

Study the role of past infections in shaping the present-day distribution of inflammatory disease risk alleles

Use evolutionary information to identify novel risk variants for Crohn’s disease

An innovative approach

We developed a strategy to detect pathogen-driven selection

Pathogen-driven selection implies that allele frequencies at a locus are shaped by selective pressure imposed by one or more infectious agents

Strengths: 1) We test a specific hypothesis on the underlying selective pressure (can distinguish among different pathogen groups)

2) High power to detect selection on standing variation

Weaknesses: 1) Use of a relatively low-density SNP panel

2) Use of a population panel with uneven geographic representation

Pathogen-driven selection Identifies correlations between genetic variability and pathogen-driven selective pressure.

We need a measure of selective-pressure that reflects historical pressures (evolution acts over long time periods).

pathogen-driven selective pressure pathogen-driven selective pressure

neutral variant selected variantselected variant

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Pathogen diversity can be used as a measure of the selective pressure exerted by infectious agents on human populations.

Pathogen diversity more closely reflects historical pressures than other estimates such as the prevalence of specific infections

HLAA

HLAB

Over 650 000 SNPs genotyped in 52 populations (HGDP-CEPH panel).

Pathogen diversity: number of different pathogen species/families present in different geographical areas of the world from the Gideon database.

We calculate Kendall's rank correlation coefficient (tau) between allele frequencies in HGDP-CEPH populations and pathogen diversity.

A SNP was considered to be significantly associated with pathogen diversity if it displayed a significant correlation and a rank higher than 0.99

OUR APPROACH

99th percentile

tau

pathogen diversity

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Pathogen diversity:

Micro-pathogens: viruses, bacteria, fungi and protozoaMacro-pathogens: insects, arthropods and helminths

rs917997

Among variants subjected to pathogen driven selection we identified an IBD-associated SNP located in IL18RAP.

The risk allele for IBD correlates significantly with pathogen richness

Six out of 9 risk variants for CeD or IDB/Crohn's disease (CD) were associated with micro- and macro-pathogen richness

A specific measure

Quantifying selectionEstimate pathogen-driven selection (virus, protozoa, helminth, bacteria) for single SNPs in the HGDP-CEPH panel

Retrieve all GWAS SNPs associated to any trait or disease from the NHGRI Catalog of Published Genome-Wide Association Studies

Collapse SNPs in tight LD (r2 >0.8) into single loci and retain only variants genotyped in the HGDP-CEPH panel (n=2773)

Total SNPs for CD: 43, UC: 42

Count SNPs that significantly correlate with the diversity of each pathogen group (expected 5%; observed 18%)

Apply a re-sampling approach on the 2773 GWAS SNPs to assess significance and calculate the empirical probability (on MAF-matched variants)

GWAS SNPs for CD, UC and CeD that correlate with the diversity of different

pathogens

Exploiting selection signatures to identify novel risk variants that are not picked up by

GWASExtract all SNPs with 0.05<p value <5x10-5 from CD meta-

analysis (Bartett, 2008)

Identify those selected by protozoa

Rank them based on p value

Select genic SNPs

Discard SNPs close (less than 2 Mb) to previously associated CD loci

Analyse the top 5 SNPs [rs2364403 (ARHGEF2), rs3782567 (HEBP1), rs9636320 (ARID3B), rs199533 (NSF), rs1011312

(TPST2)] in an Italian cohort

Combine results with the partially independent 6-study meta-analysis (Franke, 2010)

Association analysis

ARHGEF2: a central component of pathogen recognition by NOD1

NSF: involved in autophagy

HEBP1: promotes calcium mobilization and chemotaxis in monocytes and dendritic cells

Interaction (Ingenuity model) network

known CD susceptibility gene

Disregulated in CD and IBD mouse models; increased in IBD-associated neoplastic transformation; underexpressed in Treg; regulator of FOXP3 expression

Increased activation in mucosa of CD patients; pharamacological inhibition of RhoA patway reduces colonic inflammation in rats

Conclusions

Adaptation to pathogen exposure results in the selection for alleles that confer increased protection against infections, but predispose to CD

This information can be exploited to identify novel risk variants for CD

These data suggest that infections (e.g. T. gondii) might interact with genotype to determine CD susceptibility

These observations help building an evolutionary framework for the development of novel treatment strategies

Lab 1Chair of ImmunologyUniversity of Milano

Lab Lab 2Don C Gnocchi Foundation

Milano

MANUELA SIRONI

Rachele CaglianiUberto PozzoliDiego ForniStefania RivaMatteo Fumagalli