RT Journal Article SR Electronic T1 Characterization of ANGPT2 mutations associated with primary lymphedema JF Science Translational Medicine FD American Association for the Advancement of Science SP eaax8013 DO 10.1126/scitranslmed.aax8013 VO 12 IS 560 A1 Leppänen, Veli-Matti A1 Brouillard, Pascal A1 Korhonen, Emilia A. A1 Sipilä, Tuomas A1 Jha, Sawan Kumar A1 Revencu, Nicole A1 Labarque, Veerle A1 Fastré, Elodie A1 Schlögel, Matthieu A1 Ravoet, Marie A1 Singer, Amihood A1 Luzzatto, Claudia A1 Angelone, Donatella A1 Crichiutti, Giovanni A1 D’Elia, Angela A1 Kuurne, Jaakko A1 Elamaa, Harri A1 Koh, Gou Young A1 Saharinen, Pipsa A1 Vikkula, Miikka A1 Alitalo, Kari YR 2020 UL http://stm.sciencemag.org/content/12/560/eaax8013.abstract AB Lymphedema can occur when tissue fluid cannot enter or leaks from the lymphatic system into surrounding tissues. Some genetic causes of primary lymphedema are known, but these currently explain a minority of cases. Previous studies have shown that dominant-negative mutations in angiopoietin 2 (ANGPT2), which is involved in lymphatic vessel formation and maturation, promote lymphangiogenesis in mice. Leppänen et al. now show that inactivating mutations in angiopoietin 2 associate with primary lymphedema in humans.Primary lymphedema is caused by developmental and functional defects of the lymphatic vascular system that result in accumulation of protein-rich fluid in tissues, resulting in edema. The 28 currently known genes causing primary lymphedema can explain <30% of cases. Angiopoietin 1 (ANGPT1) and ANGPT2 function via the TIE1-TIE2 (tyrosine kinase with immunoglobulin-like and epidermal growth factor–like domains 1 and 2) receptor complex and α5β1 integrin to form an endothelial cell signaling pathway that is critical for blood and lymphatic vessel formation and remodeling during embryonic development, as well as for homeostasis of the mature vasculature. By screening a cohort of 543 individuals affected by primary lymphedema, we identified one heterozygous de novo ANGPT2 whole-gene deletion and four heterozygous ANGPT2 missense mutations. Functional analyses revealed three missense mutations that resulted in decreased ANGPT2 secretion and inhibited the secretion of wild-type (WT)–ANGPT2, suggesting that they have a dominant-negative effect on ANGPT2 signaling. WT-ANGPT2 and soluble mutants T299M and N304K activated TIE1 and TIE2 in an autocrine assay in human lymphatic endothelial cells. Molecular modeling and biophysical studies showed that amino-terminally truncated ANGPT subunits formed asymmetrical homodimers that bound TIE2 in a 2:1 ratio. The T299M mutant, located in the dimerization interphase, showed reduced integrin α5 binding, and its expression in mouse skin promoted hyperplasia and dilation of cutaneous lymphatic vessels. These results demonstrate that primary lymphedema can be associated with ANGPT2 mutations and provide insights into TIE1 and TIE2 activation mechanisms.