![]() ![]() CMTPX-labeled tumor cells (TCs) were then dispensed into inserts, and the amount of TC fluorescence emitting from the underside of insert membranes after 20 h (B–C) or 28 h (E) reflects the efficiency of trans-endothelial TC migration. Another 30mins later, the amount of FITC-dextran diffusion into the lower chambers (A, D) provides a measure of endothelial permeability. Thirty minutes later, ECs were stimulated with thrombin, 0.1% BSA (vehicle/negative control), or EDTA (positive control). The concentrations used, 10–20 ng/mL VT and 200–400 ng/mL Ang1, are estimated molar equivalents (0.71–1.43 nM). ECs were first treated with Vasculotide (VT), PBS (vehicle/negative control), Ang1 (positive control), PEG-Cys (polyethylene glycol backbone), T7c (non-PEGylated CHHHRHSF peptides), or VT in the presence of 100-fold molar excess of T7c. Vasculotide and Ang1 treatments counteract thrombin induction of trans-endothelium macromolecular permeability and tumor cell migration in vitroĪ–E Microvascular leak and tumor cell extravasation were modeled in vitro using modified Boyden chamber experiments where lung HMVECs (A–C) and dermal HMVECs (D–E) were grown to 100% confluence over 8-μm-pore insert membranes. Published under the terms of the CC BY 4.0 license. Tie2 angiopoietin metastasis tumor cell extravasation vascular permeability. Unexpectedly, detailed investigations into the putative mechanism of action of VT revealed no evidence of Tie2 agonism or Tie2 binding alternative mechanisms have yet to be determined. In the post-surgical adjuvant treatment setting, VT therapeutically complemented sunitinib therapy, an anti-angiogenic tyrosine kinase inhibitor which limited the local growth of residual disease. In the in vivo LM2-4 model, VT monotherapy had no effect on primary tumors, but significantly delayed distant metastatic dissemination to the lungs. ![]() ![]() In mice, VT treatment inhibited the transit of TCs through the pulmonary endothelium, but not the hepatic or lymphatic endothelium. In vitro, VT and Ang1 treatments impeded endothelial hypermeability and the transendothelial migration of MDA-MB-231∙LM2-4 (breast), HT29 (colon), or SN12 (renal) cancer cells to varying degrees. We hypothesized that systemic therapy with Vasculotide (VT)-a purported Ang1 mimetic, Tie2 agonist-can reduce the extravasation of potentially metastatic circulating TCs by similarly stabilizing the host vasculature. Angiopoietin-1 (Ang1) activation of Tie2 receptors on endothelial cells (ECs) reduces adhesion by tumor cells (TCs) and limits junctional permeability to TC diapedesis. ![]()
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