Janus-faced EPHB4-associated disorders: novel pathogenic variants and unreported intrafamilial overlapping phenotypes

Silvia Martin-Almedina,Alexandra Robinson,Gunnar Houge,Giles Atton,Katherine S Josephs ,Mark D Kilby,Sarah Robart,Hallvard Reigstad,Maja Hempel,Mary Beth Dinulos,Wolf-Henning Becker,Rhiannon Mellis,Fanny Kortuem,Cathrine Ebbing,Ege Sackey,Steve Jeffery,A Von Der Wense ,Dionysios Grigoriadis,Cassandra Polun,Christina Karapouliou,Peter Mortimer ,Sherri J Bale ,Kristiana Gordon,Jenny Lord,Jerome L Gorski,Stephanie E Vallee ,Siren Berland,Kazim Ogmen,Sahar Mansour,Noeline Nadarajah,Pia Østergaard

doi:10.1038/s41436-021-01136-7

Abstract

PurposeSeveral clinical phenotypes including fetal hydrops, central conducting lymphatic anomaly or capillary malformations with arteriovenous malformations 2 (CM-AVM2) have been associated with EPHB4 (Ephrin type B receptor 4) variants, demanding new approaches for deciphering pathogenesis of novel variants of uncertain significance (VUS) identified in EPHB4, and for the identification of differentiated disease mechanisms at the molecular level. MethodsTen index cases with various phenotypes, either fetal hydrops, CM-AVM2, or peripheral lower limb lymphedema, whose distinct clinical phenotypes are described in detail in this study, presented with a variant in EPHB4. In vitro functional studies were performed to confirm pathogenicity. ResultsPathogenicity was demonstrated for six of the seven novel EPHB4 VUS investigated. A heterogeneity of molecular disease mechanisms was identified, from loss of protein production or aberrant subcellular localization to total reduction of the phosphorylation capability of the receptor. There was some phenotype–genotype correlation; however, previously unreported intrafamilial overlapping phenotypes such as lymphatic-related fetal hydrops (LRFH) and CM-AVM2 in the same family were observed. ConclusionThis study highlights the usefulness of protein expression and subcellular localization studies to predict EPHB4 variant pathogenesis. Our accurate clinical phenotyping expands our interpretation of the Janus-faced spectrum of EPHB4-related disorders, introducing the discovery of cases with overlapping phenotypes.

Highlights

The EPHB4 gene encodes a receptor tyrosine kinase protein, EPHB4, that binds to its ligand EphrinB2, to initiate complex contact-dependent bidirectional signaling cascades, controlling cellular fate during embryonic angiogenesis and essential cellular processes such as adhesion, migration, and proliferation, in both blood and lymphatic endothelial cells.[1]We previously reported that monoallelic missense variants in the intracellular tyrosine kinase domain of EPHB4 cause a form of in utero primary lymphatic anomaly coined lymphatic-related fetal hydrops with/without atrial septal defect (LRFH) (OMIM 617300).[2]
Novel EPHB4 variants related to fetal hydrops of unknown etiology Since the first report of two pathogenic EPHB4 missense variants associated with LRFH,[2] four additional cases of fetal hydrops (FH1: II.[2], FH2:II.[1], FH4:II.[1], and FH5:II.2) and one case presenting with bilateral pleural effusions (FH3:II.7) with variants in EPHB4 have come to our attention (Fig. 1a–c)
Including other family members, who carried an EPHB4 variant, a total of seven individuals are reported all presenting with lymphovenous problems and/or congenital heart defects (CHD); for full clinical details and pedigrees see the Supplementary Information, Supplementary Table 1, and Supplementary Figs. 1, 2

Summary

Introduction

We previously reported that monoallelic missense variants in the intracellular tyrosine kinase domain of EPHB4 cause a form of in utero primary lymphatic anomaly coined lymphatic-related fetal hydrops with/without atrial septal defect (LRFH) (OMIM 617300).[2] The study included two families (GLDUK and GLDNOR) with a primarily lymphatic and venous phenotype, but with several family members presenting with fetal hydrops and/or atrial septal defects (ASD). Li et al identified an in-frame insertion in EPHB4 in a family with a history of fetal hydrops and lymphovenous dysfunction characterized by edema of the lower extremities, venous stasis, and variable chylous effusions.[4] This work confirmed the important role of EPHB4 in the function and development of the lymphatic system and the association of EPHB4 variants with fetal hydrops of lymphovenous origin. Under the umbrella of capillary malformation– arteriovenous malformation 2 (CM-AVM2) (OMIM 618196) are included several vascular pathologies such as isolated multifocal

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Conclusion