Genome-Wide Association Study of Non-syndromic Orofacial Clefts in a Multiethnic Sample of Families and Controls Identifies Novel Regions

Abstract

Orofacial clefts (OFCs) are among the most prevalent craniofacial birth defects worldwide and create a significant public health burden. The majority of OFCs are non-syndromic and vary in prevalence by ethnicity. Africans have the lowest prevalence of OFCs (~ 1/2,500), Asians have the highest prevalence (~1/500), Europeans and Latin Americans lie somewhere in the middle (~1/800 and 1/900, respectively). Thus, ethnicity appears to be a major determinant of the risk of developing OFC. The Pittsburgh Orofacial Clefts Multiethnic study was designed to explore this ethnic variance, comprising a large number of families and individuals (~12,000 individuals) from multiple populations worldwide: US and Europe, Asians, mixed Native American/Caucasians, and Africans. In this current study, we analyzed 2,915 OFC cases, 6,044 unaffected individuals related to the OFC cases, and 2,685 controls with no personal or family history of OFC. Participants were grouped by their ancestry into African, Asian, European, and Central and South American subsets, and genome-wide association run on the combined sample as well as the four ancestry-based groups. We observed 22 associations to cleft lip with or without cleft palate at 18 distinct loci with p-values < 1e-06, including 10 with genome-wide significance (<5e-08), in the combined sample and within ancestry groups. Three loci - 2p12 (rs62164740, p = 6.27e-07), 10q22.2 (rs150952246, p = 3.14e-07), and 10q24.32 (rs118107597, p = 8.21e-07) are novel. Nine were in or near known OFC loci - PAX7, IRF6, FAM49A, DCAF4L2, 8q24.21, NTN1, WNT3-WNT9B, TANC2, and RHPN2. The majority of the associations were observed only in the combined sample, European, and Central and South American groups. We investigated whether the observed differences in association strength were (a) purely due to sample sizes, (b) due to systematic allele frequency difference at the population level, or (c) due to the fact certain OFC-causing variants confer different amounts of risk depending on ancestral origin, by comparing effect sizes to observed allele frequencies of the effect allele in our ancestry-based groups. While some of the associations differ due to systematic differences in allele frequencies between groups, others show variation in effect size despite similar frequencies across ancestry groups.

Genome‐wide association study of multiethnic nonsyndromic orofacial cleft families identifies novel loci specific to family and phenotypic subtypes

“…Finally, the observed EAF within cases from the two GWASs were examined to assess whether these differed significant between cleft subtypes. We have previously shown that ancestry impacts association to CL/P in our POFC sample (Mukhopadhyay et al, 2021); therefore, we examined the subtype-specific effect sizes within each ancestry group to assess whether the differences observed were similar to the those observed for the meta-analysis. EAFs within cases were also compared across the eight phenotypic subtypes within each ancestry group in addition to the cases pooled across ancestry groups for each phenotypic subset.…”

“…We have shown previously that the degree of OFC risk at certain susceptibility loci varies with ancestry of the sample participants (Mukhopadhyay et al, 2021). To control for this variance, we first classified subjects into four different genetically defined ancestry groups using the principal component analysis-based classification defined in a previous study using POFC subjects (Leslie et al, 2016).…”

“…Because the degree of OFC risk at certain susceptibility loci varies with ancestry (Mukhopadhyay et al, 2021), the effect of ancestry was incorporated into our analyses. The four ancestry groups used to classify study participants are AFR (African ancestry), ASIA (Asian ancestry), EUR (White, European ancestry) and CSA (Central and South American ancestry).…”

Decoding the Human Face: Progress and Challenges in Understanding the Genetics of Craniofacial Morphology

“…This was clearest for East African individuals, which is unsurprising due to the higher divergence in allele frequency and linkage disequilibrium between African and non-African populations ( 93 ). Furthermore, a recent GWAS of nsCL/P risk from multiple broad ancestry groups found substantial dependence on ancestry in terms of SNP effect size and significance ( 109 ).…”

Association of rs2013162 and rs2235375 Polymorphisms in IRF6 Gene with Susceptibility to Non-Syndromic Cleft Lip and Palate

“…Although many susceptibility loci have been suggested by linkage and candidate gene association studies, the vast majority were non replicable across studies 6 . A remarkable exception was IRF6 (interferon regulatory factor 6), in which heterozygous loss-of-function mutations lead to van der Woude syndrome and their association with NSCL/P was confirmed in several GWAS 7 . The IRF6 gene on chromosome 1q32.3-q41 encodes interferon regulatory factor 6, which is a key element in oral and maxillofacial problems and is one of the candidate genes associated with both syndromic and non-syndromic forms of clefts 8 .…”