Rare variants found in clinical gene panels illuminate the genetic and allelic architecture of orofacial clefting

Purpose Orofacial clefts (OFCs) are common birth defects including cleft lip (CL), cleft lip and palate (CLP), and cleft palate (CP). OFCs have heterogeneous etiologies, complicating clinical diagnostics as it is not always apparent if the cause is Mendelian, environmental, or multifactorial. Sequencing is not currently performed for isolated or sporadic OFCs, so we estimated the diagnostic yield for 418 genes in 841 cases and 294 controls. Methods We evaluated 418 genes using genome sequencing and curated variants to assess their pathogenicity using American College of Medical Genetics criteria. Results 9.04% of cases and 1.02% of controls had ‘likely pathogenic’ (LP) variants (p<0.0001), which was almost exclusively driven by heterozygous variants in autosomal genes. CP (17.6%) and CLP (9.09%) cases had the highest yield while CL cases had a 2.80% yield. Out of 39 genes with LP variants, nine genes, including CTNND1 and IRF6 , accounted for more than half of the yield (4.64% of cases). Most variants (61.8%) were ‘variants of uncertain significance’ (VUSs), occurring more frequently in cases (p=0.004), but no individual gene showed a significant excess of VUSs. Conclusion These results underscore the etiological heterogeneity of OFCs and suggest sequencing could reduce the diagnostic gap in OFCs. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement These studies are part of the Gabriella Miller Kids First Pediatric Research Program, supported by the Common Fund of the Office of the Director of the National Institutes of Health (NIH). Sequencing of the European trios was completed at Washington University McDonnell Genome Institute (3U54HG003079-12S1 and X01-HL132363 [M.L.M., E.F.]) and the Colombian and Taiwanese trios were sequenced at the Broad Institute Sequencing Center (U24-HD090743, X01-HL136465 [M.L.M., E.F.], X01-HL140516 [T.H.B.]). The sequencing centers plus the Kids First Data Resource Center, supported by the NIH Common Fund through grant U2CHL138346, provided technical and analytical support of this project. The assembling of the sample of child-parent trios, collection of the phenotypic data and samples, and data analysis were supported by NIH grants: R01-DE016148 [M.L.M. and S.M.W.], R03-DE026469 [E.F. and M.L.M.], R03-DE027193 [E.J.L.], R03-DE027103 [E.J.L], R00-DE025060 [E.J.L.], R01-DE027983 [E.J.L.], R01-DE028342 [E.J.L], R01-DE030342 [E.J.L], R01-DE011931 [J.T.H.], U01-DD000295 [G.W.], R03-DE027121 [T.H.B], and T32-GM008490 [K.D.P], R01-DE031261 [H.B.], and R00-DE026824 [H.B.]. This work was supported in part by a grant to Emory University from the Howard Hughes Medical Institute through the James H. Gilliam Fellowships for Advanced Study program [K.D.P]. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: IRBs at the University of Pittsburgh, University of Iowa, and Emory University gave ethical approval for this work. I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines and uploaded the relevant EQUATOR Network research reporting checklist(s) and other pertinent material as supplementary files, if applicable. Yes The case data analyzed and reported in this manuscript were accessed from the database of Genotypes and Phenotypes (dbGaP; European trios, dbGaP: phs001168.v2.p2; Colombian trios, dbGaP: phs001420.v1.p1; Taiwanese trios, dbGaP: phs000094.v1.p1) and from the Kids First Data Resource Center. The control data is available from public data repositories as described in .