Combination of Dll4/Notch and Ephrin-B2/EphB4 targeted therapy is highly effective in disrupting tumor angiogenesis

Background: Dll4/Notch and Ephrin-B2/EphB4 pathways play critical roles in tumor vessel development and maturation. This study evaluates the efficacy of the inhibition of both signaling pathways, alone and in combination, in reducing the growth of an autochthonous mouse tumor and assesses potential adverse effects. Methods: We used the transgenic RIP1-Tag2 tumor model to study the effects of 1) inhibition of Dll4/Notch by either Dll4 allelic deletion or use of a soluble extracellular Dll4 (sDll4), 2) inhibition of Ephrin-B2/EphB4 signaling by a soluble extracellular EphB4 fused to albumin (sEphB4-Alb), and 3) inhibition of both pathways by sEphB4-Alb combined with either Dll4 allelic deletion or sDll4. To investigate adverse effects, we used inducible endothelial-specific Dll4 knock-out mice, treated with sEphB4-Alb, and carried out histopathological analysis. Results: Dll4 allele deletion or soluble Dll4 treatment resulted in increased tumor vessel density, reduced mural cell recruitment and vessel perfusion which resulted in reduced tumor size. The soluble EphB4 instead reduced vessel density and vessel perfusion, leading to reduction of tumor size. Greater efficacy was observed when sEphB4-Alb was combined with either Dll4 allele deletion or sDll4 in regards to tumor size, vessel perfusion and mural cell recruitment. Induced endothelial specific Dll4 loss-of-function caused hepatic vascular alterations, which were prevented by concomitant sEphB4-Alb treatment. Conclusion: Combination targeting of Dll4/Notch and Ephrin-B2/EphB4 has potential for clinical investigation, providing cumulative efficacy and increased safety over Dll4/Notch inhibition alone.