Inhibition of the mammary carcinoma angiogenic switch in C3(1)/SV40 transgenic mice by a mutated form of human endostatin

Abstract

Cancer therapies based on the inhibition of angiogenesis by endostatin have recently been developed. We demonstrate that a mutated form of human endostatin (P125A) can inhibit the angiogenic switch in the C3(1)/Tag mammary cancer model. P125A has a stronger growth-inhibitory effect on endothelial cell proliferation than wild-type endostatin. We characterize the angiogenic switch, which occurs during the transition from preinvasive lesions to invasive carcinoma in this model, and which is accompanied by a significant increase in total protein levels of vascular endothelial growth factor (VEGF) and an invasion of blood vessels. Expression of the VEGF 188 mRNA isoform, however, is suppressed in invasive carcinomas. The VEGF receptors fetal liver kinase-1 (Flk-1) and Fms-like tyrosine kinase-1 (Flt-1) become highly expressed in epithelial tumor and endothelial cells in the mammary carcinomas, suggesting a potential autocrine effect for VEGF on tumor cell growth. Angiopoietin-2 mRNA levels are also increased during tumor progression. CD-31 Key words: angiogenesis; endostatin; vascular endothelial growth factor; mammary cancerThe growth and metastatic spread of solid tumors is critically dependent on the formation of new blood vessels through angiogenesis. 1,2 During this process, the effects of proangiogenic factors dominate over the antiangiogenic factors, resulting in the angiogenic switch. 2 A large variety of growth factors modulate tumor angiogenesis, including those that alter the extracellular matrix as well as those that promote endothelial cell proliferation and migration. 3 Among the proangiogenic factors described, vascular endothelial growth factor (VEGF) has been shown to be frequently upregulated in a variety of tumors 4 and has been well characterized. 4,5 The 2 high-affinity VEGF receptors fetal liver kinase-1 (Flk-1) and Fms-like tyrosine kinase-1 (Flt-1), members of the tyrosine kinase receptor family, transduce the activity of this growth factor. 5 The activation of intracellular signaling pathways by these receptors involves their autophosphorylation and, as a consequence, the stimulation of endothelial cell proliferation (especially in the case of the Flk-1 receptor), migration, tubule formation, and angiogenesis in vivo. [5][6][7] Although the expression of VEGF receptors was initially localized to endothelial cells, recent studies have shown that epithelial and tumor cells from various tissues also express VEGF receptors and respond to stimulation by VEGF. 8 -10 Another family of vascular growth factors, angiopoietins, has recently been characterized. 11 Angiopoietin-1 mediates cell-cell interactions between endothelial cells and other neighboring cells, such as smooth muscle cells. 12,13 In contrast to VEGF, angiopoietin-1 does not directly stimulate endothelial cell growth. Angiopoietin-2 is typically expressed at sites of vascular remodeling in the adult 14 and in tumors, where it stimulates endothelial cell growth and angiogenesis. 11,14 Moreover, angiopoietin-2 acts in a complementary a...

Macrophages Regulate the Angiogenic Switch in a Mouse Model of Breast Cancer

“…The angiogenic switch, identified by the formation of a dense vascular network, occurred exclusively in this malignant transition region. This is consistent with other studies using transgenic mouse models of cancers in which a transition stage, termed angiogenic dysplasia, occured before the malignant carcinoma was identified (24)(25)(26)(27)(28). However, the close association between the angiogenic switch and malignant transition was not reported in these studies.…”

Endostatin therapy reveals a U-shaped curve for antitumor activity

“…Adenoviruses are the most commonly used vectors for this strategy. [22][23][24][25][26][27][28][29] However, expression of antiangiogenic molecules mediated by adenovirus-based vectors is limited by an effective host immune response and transient nature of expression. Adeno-associated virus (AAV)-based vectors are emerging as potential alternative vectors for cancer therapy because AAV is a non-pathogenic vector with limited host immune response, and allows for long-term and stable transgene expression.…”

Adeno-associated virus-mediated delivery of a mutant endostatin suppresses ovarian carcinoma growth in mice

“…As a consequence, the mutant protein inhibited EC migration, proliferation and angiogenesis more effectively when compared to the native molecule. 15,21 Preliminary studies suggest that the mutation could alter binding profile of endostatin towards cell surface integrins (unpublished). Structural modifications altering the biological activity can be beneficial in improving therapeutic efficacy.…”

“…Mutated human endostatin inhibited the spontaneous development of mammary cancer in C3 (1)/Tag mice. 15 P125A-endostatin (human endostatin wherein proline 125 was substituted with alanine) inhibits endothelial cell (EC) proliferation and migration better than the native protein. 15,21 In the present study, we evaluated the effect of rAAV-mediated delivery of the mutated endostatin in a mouse model of ovarian cancer.…”

“…15 P125A-endostatin (human endostatin wherein proline 125 was substituted with alanine) inhibits endothelial cell (EC) proliferation and migration better than the native protein. 15,21 In the present study, we evaluated the effect of rAAV-mediated delivery of the mutated endostatin in a mouse model of ovarian cancer. P125A-endostatin cDNA was cloned in the rAAV vector and IgG kappa chain leader sequence was engineered as a secretory signal.…”