title: Context- and cell-dependent effects of Delta-like 4 targeting in the bone marrow microenvironment.
abstract: Delta-like 4 (Dll4) is a ligand of the Notch pathway family which has been widely studied in the context of tumor angiogenesis, its blockade shown to result in non-productive angiogenesis and halted tumor growth. As Dll4 inhibitors enter the clinic, there is an emerging need to understand their side effects, namely the systemic consequences of Dll4:Notch blockade in tissues other than tumors. The present study focused on the effects of systemic anti-Dll4 targeting in the bone marrow (BM) microenvironment. Here we show that Dll4 blockade with monoclonal antibodies perturbs the BM vascular niche of sub-lethally irradiated mice, resulting in increased CD31(+), VE-Cadherin(+) and c-kit(+) vessel density, and also increased megakaryocytes, whereas CD105(+), VEGFR3(+), SMA(+) and lectin(+) vessel density remained unaltered. We investigated also the expression of angiocrine genes upon Dll4 treatment in vivo, and demonstrate that IGFbp2, IGFbp3, Angpt2, Dll4, DHH and VEGF-A are upregulated, while FGF1 and CSF2 are reduced. In vitro treatment of endothelial cells with anti-Dll4 reduced Akt phosphorylation while maintaining similar levels of Erk 1/2 phosphorylation. Besides its effects in the BM vascular niche, anti-Dll4 treatment perturbed hematopoiesis, as evidenced by increased myeloid (CD11b(+)), decreased B (B220(+)) and T (CD3(+)) lymphoid BM content of treated mice, with a corresponding increase in myeloid circulating cells. Moreover, anti-Dll4 treatment also increased the number of CFU-M and -G colonies in methylcellulose assays, independently of Notch1. Finally, anti-Dll4 treatment of donor BM improved the hematopoietic recovery of lethally irradiated recipients in a transplant setting. Together, our data reveals the hematopoietic (BM) effects of systemic anti-Dll4 treatment result from qualitative vascular changes and also direct hematopoietic cell modulation, which may be favorable in a transplant setting.
authors.affiliation: Angiogenesis Lab, CIPM, Portuguese Institute of Oncology, IPOLFG, EPE, Lisbon, Portugal.
authors.forename: Leonor
authors.initials: L
authors.lastname: Remédio
authors.forename: Tânia
authors.initials: T
authors.lastname: Carvalho
authors.forename: Francisco
authors.initials: F
authors.lastname: Caiado
authors.forename: Ana
authors.initials: A
authors.lastname: Bastos-Carvalho
authors.forename: Diana
authors.initials: D
authors.lastname: Martins
authors.forename: António
authors.initials: A
authors.lastname: Duarte
authors.forename: Hideo
authors.initials: H
authors.lastname: Yagita
authors.forename: Sergio
authors.initials: S
authors.lastname: Dias
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D057134:Antibodies, Neutralizing / Q000494:pharmacology; D001853:Bone Marrow / Q000098:blood supply / Q000187:drug effects / Q000378:metabolism*; D001854:Bone Marrow Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism*; D016026:Bone Marrow Transplantation; D002135:Calcium-Binding Proteins; D060833:Cellular Microenvironment / Q000187:drug effects; D006410:Hematopoiesis / Q000187:drug effects; D061307:Human Umbilical Vein Endothelial Cells / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D018345:Mice, Knockout; D014916:Whole-Body Irradiation
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D057134:Antibodies, Neutralizing; D002135:Calcium-Binding Proteins; C519665:DLL4 protein, mouse; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins