Cytotoxicity of VEGF121/rGel on vascular endothelial cells resulting in inhibition of angiogenesis is mediated via VEGFR-2

Khalid A. Mohamedali, Sophia Ran, Candelaria Gomez-Manzano, Latha Ramdas, Jing Xu, Sehoon Kim, Lawrence H. Cheung, Walter N. Hittelman, Wei Zhang, Johannes Waltenberger, Philip E. Thorpe, Michael G. Rosenblum

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Abstract

Background: The fusion protein VEGF121/rGel composed of the growth factor VEGF121 and the plant toxin gelonin targets the tumor neovasculature and exerts impressive anti-vascular effects. We have previously shown that VEGF121/rGel is cytotoxic to endothelial cells overexpressing VEGFR-2 but not to endothelial cells overexpressing VEGFR-1. In this study, we examined the basis for the specific toxicity of this construct and assessed its intracellular effects in vitro and in vivo.Methods: We investigated the binding, cytotoxicity and internalization profile of VEGF121/rGel on endothelial cells expressing VEGFR-1 or VEGFR-2, identified its effects on angiogenesis models in vitro and ex vivo, and explored its intracellular effects on a number of molecular pathways using microarray analysis.Results: Incubation of PAE/VEGFR-2 and PAE/VEGFR-1 cells with 125I-VEGF121/rGel demonstrated binding specificity that was competed with unlabeled VEGF121/rGel but not with unlabeled gelonin. Assessment of the effect of VEGF121/rGel on blocking tube formation in vitro revealed a 100-fold difference in IC50 levels between PAE/VEGFR-2 (1 nM) and PAE/VEGFR-1 (100 nM) cells. VEGF121/rGel entered PAE/VEGFR-2 cells within one hour of treatment but was not detected in PAE/VEGFR-1 cells up to 24 hours after treatment. In vascularization studies using chicken chorioallantoic membranes, 1 nM VEGF121/rGel completely inhibited bFGF-stimulated neovascular growth. The cytotoxic effects of VEGF121/rGel were not apoptotic since treated cells were TUNEL-negative with no evidence of PARP cleavage or alteration in the protein levels of select apoptotic markers. Microarray analysis of VEGF121/rGel-treated HUVECs revealed the upregulation of a unique "fingerprint" profile of 22 genes that control cell adhesion, apoptosis, transcription regulation, chemotaxis, and inflammatory response.Conclusions: Taken together, these data confirm the selectivity of VEGF121/rGel for VEGFR-2-overexpressing endothelial cells and represent the first analysis of genes governing intoxication of mammalian endothelial cells by a gelonin-based targeted therapeutic agent.

Original languageEnglish (US)
Article number358
JournalBMC Cancer
Volume11
DOIs
Publication statusPublished - Aug 17 2011

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ASJC Scopus subject areas

  • Oncology
  • Cancer Research
  • Genetics

Cite this

Mohamedali, K. A., Ran, S., Gomez-Manzano, C., Ramdas, L., Xu, J., Kim, S., ... Rosenblum, M. G. (2011). Cytotoxicity of VEGF121/rGel on vascular endothelial cells resulting in inhibition of angiogenesis is mediated via VEGFR-2. BMC Cancer, 11, [358]. https://doi.org/10.1186/1471-2407-11-358