ActivinA Induced SMAD1/5 Signaling in an iPSC Derived EC Model of Fibrodysplasia Ossificans Progressiva (FOP) Can Be Rescued by the Drug Candidate Saracatinib

Abstract

Balanced signal transduction is crucial in tissue patterning, particularly in the vasculature. Heterotopic ossification (HO) is tightly linked to vascularization with increased vessel number in hereditary forms of HO, such as Fibrodysplasia ossificans progressiva (FOP). FOP is caused by mutations in the BMP type I receptor ACVR1 leading to aberrant SMAD1/5 signaling in response to ActivinA. Whether observed vascular phenotype in human FOP lesions is connected to aberrant ActivinA signaling is unknown. Blocking of ActivinA prevents HO in FOP mice indicating a central role of the ligand in FOP. Here, we established a new FOP endothelial cell model generated from induced pluripotent stem cells (iECs) to study ActivinA signaling. FOP iECs recapitulate pathogenic ActivinA/SMAD1/5 signaling. Whole transcriptome analysis identified ActivinA mediated activation of the BMP/NOTCH pathway exclusively in FOP iECs, which was rescued to WT transcriptional levels by the drug candidate Saracatinib. We propose that ActivinA causes transcriptional pre-patterning of the FOP endothelium, which might contribute to differential vascularity in FOP lesions compared to non-hereditary HO. Graphical abstract

Fibrodysplasia Ossificans Progressiva: What Have We Achieved and Where Are We Now? Follow-up to the 2015 Lorentz Workshop

“…Upon ALK2 knockdown, hypersprouting was observed in in vitro EC models, whereas ALK3 knockdown appeared to have the opposite effect ( 28 ). Recent data showed that EC’s derived from human induced pluripotent stem cells (hiPSC) follow the same principle and hiPSCs derived from patients with FOP show activin A induced SMAD 1/5 signaling ( 29 ).…”

“…Periodontal ligament fibroblasts have also been isolated and induced to osteogenesis and osteoclastogenesis ( 34 ). hiPSCs obtained from patients with FOP are able to differentiate to ECs ( 29 , 35 , 36 ) and pericytes with increased mineralization, but did not develop into mature osteoblasts ( 36 ). Connective tissue progenitor cells from discarded primary teeth have been used to examine the effects of FOP mutations on BMP signaling and chondrogenic/osteogenic differentiation ( 19 , 24 , 37 , 38 ).…”

Single-Cell Integration Analysis of Heterotopic Ossification and Fibrocartilage Developmental Lineage: Endoplasmic Reticulum Stress Effector Xbp1 Transcriptionally Regulates the Notch Signaling Pathway to Mediate Fibrocartilage Differentiation

“…In GSE94683, GSE102929, GSE168153, and GSE138515, the upregulation of Xbp1 was found to be associated with the upregulation of WNT, Notch, Hh, and TGF beta signaling pathways, which may be due to the crosstalk of these pathways in ectopic ossified tissues. However, the specific regulatory mechanisms remain to be further investigated [ 7 , 115 , 116 ].…”

Contemporary perspectives on heterotopic ossification

“…187), and through introduction of FOP mutations through gene editing technologies including CRISPR/Cas9 (188), with phenotypic validation of increased endochondral ossification phenotypes in vitro (189). Notably, these technologies are being implemented into drug discovery/validation pipelines, as seen with rapamycin (190) and saracatinib (191). Other similar models incorporating primary tissues include excision of neurogenic HO and surrounding muscle with sequencing data uploaded to public repositories (88), and primary connective tissue cells harvested from patients with ossification of the posterior longitudinal ligament that subsequently are exposed to cyclical mechanical stresses to approximate HO (192).…”