Influence of Levamisole and Other Angiogenesis Inhibitors on Angiogenesis and Endothelial Cell Morphology in Vitro

Abstract

Angiogenesis, the formation of new blood vessels from existing vessels is required for many physiological processes and for growth of solid tumors. Initiated by hypoxia, angiogenesis involves binding of angiogenic factors to endothelial cell (EC) receptors and activation of cellular signaling, differentiation, migration, proliferation, interconnection and canalization of ECs, remodeling of the extracellular matrix and stabilization of newly formed vessels. Experimentally, these processes can be studied by several in vitro and in vivo assays focusing on different steps in the process. In vitro, ECs form networks of capillary-like tubes when propagated for three days in coculture with fibroblasts. The tube formation is dependent on vascular endothelial growth factor (VEGF) and omission of VEGF from the culture medium results in the formation of clusters of undifferentiated ECs. Addition of angiogenesis inhibitors to the coculture system disrupts endothelial network formation and influences EC morphology in two distinct ways. Treatment with antibodies to VEGF, soluble VEGF receptor, the VEGF receptor tyrosine kinase inhibitor SU5614, protein tyrosine phosphatase inhibitor (PTPI) IV or levamisole results in the formation of EC clusters of variable size. This cluster morphology is a result of inhibited EC differentiation and levamisole can be inferred to influence and block VEGF signaling. Treatment with platelet factor 4, thrombospondin, rapamycin, suramin, TNP-470, salubrinal, PTPI I, PTPI II, clodronate, NSC87877 or non-steriodal anti-inflammatory drugs (NSAIDs) results in the formation of short cords of ECs, which suggests that these inhibitors have an influence on later steps in the angiogenic process, such as EC proliferation and migration. A humanized antibody to VEGF is one of a few angiogenesis inhibitors used clinically for treatment of cancer. Levamisole is approved for clinical treatment of cancer and is interesting with respect to anti-angiogenic activity in vivo since it inhibits ECs in vitro with a morphology resembling that obtained with antibodies to VEGF.

Interleukin-8: A potent promoter of angiogenesis in gastric cancer

“…Currently, angiogenesis is acknowledged as a key and validated cancer therapeutic target due to its pivotal role in the progression and metastasis of malignancies (26,27). Worldwide, gastric cancer is one of the leading causes of cancer-associated mortality (2).…”

Physicochemical and Biological Activity Analysis of Low-Density Polyethylene Substrate Modified by Multi-Layer Coatings Based on DLC Structures, Obtained Using RF CVD Method

“…Treated and untreated cells were incubated for 72 h. Every 24 h, the medium was changed for a fresh one. After that time, cytotoxicity was determined by Alamar Blue assay (Sigma-Aldrich, Bornem, Belgium) according to the well-described protocol in [24]. In brief, cells were washed with phosphate-buffered saline (PBS) and incubated with resazurin sodium salt solution (25 µM in PBS) for 4 h at 37 • C in the dark.…”

The anti-angiogenic potential of (±) gossypol in comparison to suramin

“…In this study, suramin was used as the positive control because it inhibits angiogenesis by targeting the receptor signals of fibroblast growth factor and vascular endothelial growth factor in cultured endothelial cells (Khoo et al 2011;Friis et al 2013). Different suramin doses were chosen in the literature (Garrido et al 1995;Firsching et al 1995;Kolmakova et al 2009;Khoo et al 2011;Minamikawa-Tachino et al 2013;Friis et al 2013, Prigozhina et al 2013.…”

“…(±)-gossypol on various cancer cell lines and tumors has been used in cancer therapy (Jaroszewski et al 1990). Suramin inhibits angiogenesis by targeting the receptor signals of fibroblast growth factor and vascular endothelial growth factor (Khoo et al 2011;Friis et al 2013). Despite their different targets, these substances appear to affect the pathways leading to angiogenesis inhibition.…”