S41 Characterisation of mutations in the gene encoding growth and differentiation factor 2 (GDF2) in patients with pulmonary arterial hypertension

Most heritable cases of pulmonary arterial hypertension (PAH) are explained by mutations in BMPR2, which encodes the bone morphogenetic protein type 2 receptor (BMPR-II). Additionally, mutations in the endothelial type 1 receptor, ALK1, can cause PAH. Growth and differentiation factor 2 (GDF2; also known as BMP9) is a circulating vascular quiescence factor that possesses high affinity for the BMPR-II/ALK1 complex. Furthermore, exogenous GDF2 administration reverses PAH in animal models. Recently, an unbiased case-control analysis revealed rare missense mutations in GDF2 in PAH cases.1 Here we determined the functional impact and pathogenicity of disease-associated mutations in GDF2. Sixteen GDF2 missense variants were generated in an overexpression system and characterised by ELISAs, western blotting, signalling and apoptosis assays. BMP9 and BMP10 levels were assayed in plasma samples from PAH patients harbouring GDF2 missense variants and deletions at the GDF2 locus, and age/sex matched healthy controls. Missense variants predicted to be pathogenic by in silico analysis exhibited severely reduced secretion and activity of GDF2 in vitro. For most variants, reduced secretion was associated with disruption of the interaction between the prodomain and growth factor domain of GDF2, leading to destabilisation of the molecule. In contrast, missense variants predicted to be benign exhibited normal signalling and functional responses. Similarly, plasma from pathogenic GDF2 mutation carriers exhibited greatly reduced levels of the GDF2 growth factor domain compared to healthy controls and benign missense carriers. Intriguingly, plasma levels of BMP10, the other circulating ligand for ALK1/BMPR-II, were also markedly reduced in patients with pathogenic GDF2 variants, suggesting co-regulation of these circulating BMPs. These studies provide evidence for the pathogenicity of rare missense mutations in GDF2 and demonstrate that the major mechanism involves destabilisation of the GDF2 molecule leading to reduced secretion of the growth factor domain. These data provide support for the central role of the GDF2/BMPR-II/ALK1 axis in PAH and suggest that GDF2 replacement may be an effective therapeutic strategy in these individuals. This study was produced by the Morrell group on behalf of the UK PAH Cohort Study. Reference Nat Commun 12 April 2018;9(1):1416. doi:10.1038/s41467-018-03672-4