Proposal submitted to the MRC centenary award (presided over by George Davey Smith):

Filaggrin (FLG) genotyping to enable FLG+/- stratified analysis to identify FLG*SNP interactions in eczema.

Lavinia Paternoster (PI)

Filaggrin null mutations have been established as a major risk factor for eczema. Several low frequency loss-of-function mutations have been identified which result in skin barrier defects and are associated with increased risk of ichthyosis, eczema and asthma. Originally two mutations (R501X and 2282del4) were identified and used in genetic association studies. However, recently published data on a European population shows that whilst using these two mutations will classify ~13% of the population as FLG mutation carriers, including a further two loss of function mutations (R2447X and S3247X) will classify an additional ~4% of the population as carriers.

We recently carried out a large-scale GWAS meta-analysis within the EAGLE consortium, where we identified three novel loci for atopic dermatitis. Two of these variants were located near genes that play a role in epidermal differentiation and proliferation (OVOL1 and ACTL9), whilst one was in KIF3A within a cytokine cluster of genes with previous evidence of immune-related and inflammatory functions (including associations with asthma and psoriasis). These findings highlight the importance of both skin-barrier and immune related genes in the aetiology of eczema. We hypothesise that there are likely to be important interactions between these two pathways, in that genetic variants within inflammatory/immune-related genes might only predispose to eczema if there is also dysfunction in skin barrier pathways. Due to the large number of tests carried out, analyses often lack power to detect interactions between genetic variants, but we plan to focus on interactions between FLG and known eczema and atopy variants.

Proposed Analysis: We propose genotyping the 4 FLG null mutations in the EAGLE cohorts and then carry out FLG+/- stratified analyses for the known eczema (and other atopy) hits to identify interactions between each SNP and FLG. Each cohort will carry out the analyses and provide summary data to Bristol for meta-analysis.

Pilot results: In ALSPAC we have already genotyped two of the four FLG variants (i.e. R501X and 2282del4). Preliminary analyses show evidence for an interaction between FLG and one of the known eczema hits (rs2897442, in KIF3A), showing an association only in those individuals with a FLG mutation (FLG+ OR=1.75, 95%CI 1.22 to 2.50, p=0.002 compared to FLG- OR=1.08, 95%CI 0.96 to 1.21, p=0.212; interaction p-value =0.014). We also find some evidence for an interaction between atopy SNP rs2252226 (FCER1A) and FLG, with this SNP also only showing association amongst individuals with a FLG mutation (FLG+ OR=1.50, 95%CI 1.11 to 1.98, p=0.008 compared to FLG- OR=0.96, 95%CI 0.87 to 1.07, p=0.475; interaction p-value=0.005). This gene also has a role in immune response.

It is expected that genotyping the additional two FLG variants in ALSPAC will increase power to detect interaction (by improving classification of people into FLG+ and FLG- strata). We also plan to use the other EAGLE cohorts to replicate this finding and carry out a meta-analysis of all FLG*known atopy hit interactions.

This resource will subsequently be used to carry out a genome-wide test for FLG interactions amongst the EAGLE cohorts and could be used in the analysis of other conditions, such as asthma. More thorough genotyping of the FLG mutations will also enable us to investigate properly whether the GWAS signal seen in this region can be completely explained by known FLG mutations.