title: Screening assay for blood vessel maturation inhibitors.
abstract: In cancer patients, the development of resistance to anti-angiogenic agents targeting the VEGF pathway is common. Increased pericyte coverage of the tumor vasculature undergoing VEGF targeted therapy has been suggested to play an important role in resistance. Therefore, reducing the pericytes coverage of the tumor vasculature has been suggested to be a therapeutic approach in breaking the resistance to and increasing the efficacy of anti-angiogenic therapies. To screen compound libraries, a simple in vitro assay of blood vessel maturation demonstrating endothelial cells and pericytes association while forming lumenized vascular structures is needed. Unfortunately, previously described 3-dimensional, matrix based assays are laborious and challenging from an image and data acquisition perspective. For these reasons they generally lack the scalability needed to perform in a high-throughput environment. With this work, we have developed a novel in vitro blood vessel maturation assay, in which lumenized, vascular structures form in one optical plane and mesenchymal progenitor cells (10T1/2) differentiate into pericyte-like cells, which associate with the endothelial vessels (HUVECs). The differentiation of the 10T1/2 cells into pericyte-like cells is visualized using a GFP reporter controlled by the alpha smooth muscle actin promoter (SMP-8). The organization of these vascular structures and their recruited mural cells in one optical plane allows for automated data capture and subsequent image analysis. The ability of this assay to screen for inhibitors of pericytes recruitment was validated. In summary, this novel assay of in vitro blood vessel maturation provides a valuable tool to screen for new agents with therapeutic potential.
authors.affiliation: Department of Oncology, Johns Hopkins University, Baltimore, MD, USA.
authors.forename: Chenglai
authors.initials: C
authors.lastname: Fu
authors.forename: Anita
authors.initials: A
authors.lastname: van der Zwan
authors.forename: Stephanie
authors.initials: S
authors.lastname: Gerber
authors.forename: Susan
authors.initials: S
authors.lastname: Van Den Berg
authors.forename: Elisa
authors.initials: E
authors.lastname: No
authors.forename: Wayne C H
authors.initials: WC
authors.lastname: Wang
authors.forename: Nader
authors.initials: N
authors.lastname: Sheibani
authors.forename: Michael A
authors.initials: MA
authors.lastname: Carducci
authors.forename: Sushant
authors.initials: S
authors.lastname: Kachhap
authors.forename: Hans J
authors.initials: HJ
authors.lastname: Hammers
mesh_terms: D000199:Actins / Q000378:metabolism; D020533:Angiogenesis Inhibitors / Q000494:pharmacology*; D000818:Animals; D001549:Benzamides / Q000494:pharmacology; D001808:Blood Vessels / Q000187:drug effects* / Q000473:pathology*; D002460:Cell Line; D018920:Coculture Techniques; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D042783:Endothelial Cells / Q000166:cytology; D005347:Fibroblasts / Q000166:cytology; D049452:Green Fluorescent Proteins / Q000378:metabolism; D057809:HEK293 Cells; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D000068877:Imatinib Mesylate; D007211:Indoles / Q000494:pharmacology; D016086:Lentivirus / Q000378:metabolism; D051379:Mice; D009130:Muscle, Smooth / Q000378:metabolism; D018919:Neovascularization, Physiologic; D020286:Pericytes / Q000166:cytology; D010879:Piperazines / Q000494:pharmacology; D011401:Promoter Regions, Genetic; D011743:Pyrimidines / Q000494:pharmacology; D011758:Pyrroles / Q000494:pharmacology; D013234:Stem Cells / Q000166:cytology; D000077210:Sunitinib
chemical_list: D000199:Actins; D020533:Angiogenesis Inhibitors; D001549:Benzamides; D007211:Indoles; D010879:Piperazines; D011743:Pyrimidines; D011758:Pyrroles; D049452:Green Fluorescent Proteins; D000068877:Imatinib Mesylate; D000077210:Sunitinib
keywords: Angiogenesis model; Blood vessel maturation; Drug screening; Endoglin; Imatinib; Pericyte; Sunitinib
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