Angiopoietin-2 Signaling Targeted Therapeutics for Arteriovenous Malformations

PROJECT SUMMARY Hereditary Hemorrhagic Telangiectasia (HHT) is a vascular genetic disorder characterized by enlarged, leaky small vessels (telangiectasias) and inappropriate, fragile connections between arteries and veins called arteriovenous malformations (AVMs). HHT patients develop AVMs in a specific subset of major organs, which can rupture causing severe hemorrhage and anemia, as well as aneurysms, stroke and even death. Causes of HHT are linked to the Transforming Growth Factor-beta (TGF-beta) signaling pathway, with over 90% of patients exhibiting heterozygous loss-of-function mutations in the Activin receptor-like kinase 1 (Acvrl1/Alk1) or Endoglin (Eng) co-receptors, or the downstream transcription factor, Smad-related protein 4 (Smad4). Despite knowing the causative mutations, a significant gap remains in our understanding of the immediate TGF-beta downstream signaling components responsible for HHT pathologies. Furthermore, no cure is currently available for HHT. We have found that directly downstream of Alk1, Eng and Smad4 loss-of-function, the angiogenic factor and antagonistic ligand to the Tie2 receptor, angiopoietin-2 (Angpt2/ANG2), is transcriptionally elevated to trigger HHT vascular pathologies; ANG2 neutralization efficiently reduced AVM pathology in two HHT mouse models. In addition, loss of Alk1 signaling led to a robust and consistent transcriptional and signaling inhibition of Tek/Tie2. Together, these observations strongly support the working model that ANG2 is elevated and Tie2 signaling is repressed during the pathogenic process of AVM development in HHT. Moreover, the metabolic PI3K/Akt/mTOR pathway is deregulated in HHT to sustain AVM development, however its connection to ANG2 pro-AVM signals is unclear. Using mouse models of the different genetic forms of HHT and primary endothelial cells (ECs), we have obtained strong pilot data indicating that ANG2-Tie2 deregulations, working in concert with enhanced FoxO1 transcriptional activity and sequential overactivation of the mTOR pathway via CXCR4 chemokine signaling, drive HHT phenotypes. The central objective of this application is to answer 3 fundamental questions: 1) how does ANG2 elevation and Tie2 signaling repression direct AVM pathogenesis, 2) what is the mechanism by which mTOR overactivation is controlled by ANG2-Tie2 signaling deregulations and 3) are approaches targeting ANG2 and the newly identified ANG2-regulated pathogenic signaling cascade universally effective in treating HHT vascular pathologies? We will address these topics by testing the following specific aims: 1) Assess ANG2 and Tie2 signaling deregulations in the most physiologically affected organs and determine if ANG2 inhibition universally blocks vascular pathologies in HHT mouse models; 2) Determine FoxO1 contributions to ANG2 and CXCR4 elevations and HHT vascular pathologies; and 3) Test if TGF-beta-Tie2- FoxO1-mediated CXCR4 elevations drive mTOR activation. These studies will advance our mechanistic understanding of AVM pathogenesis and uncover new potential targets for treating of HHT.