ABSTRACT
In order to clarify the role of TGF-beta in mammary development and tumorigenesis, we investigated the efficacy of full- or partial-length TbetaRII antisense RNA specifically to reduce TbetaRII levels in both in vitro and in vivo model systems. Here we show that the expression of TbetaRII antisense RNA in vitro reduced TbetaRII cell surface expression and inhibited the antiproliferative and transcriptional responses to exogenous TGF-beta. Expression of full-length TbetaRII antisense RNA in a transgenic mouse model under control of the mouse mammary tumor virus promotor resulted in precocious lobuloalveolar development of the mammary gland, a phenotype that resembles that of early pregnancy. These data demonstrate that TbetaRII plays a critical role in maintaining the nondifferentiated character of virgin mammary gland epithelium.
Subject(s)
Epithelial Cells/cytology , Mammary Glands, Animal/cytology , RNA, Antisense/genetics , Receptors, Transforming Growth Factor beta/genetics , Signal Transduction/physiology , Transforming Growth Factor beta/physiology , Animals , COS Cells , Cell Differentiation , Cell Line , Chlorocebus aethiops , Female , Genes, Reporter , Humans , In Situ Hybridization , Luciferases/genetics , Mice , Mice, Transgenic , Mink , Protein Serine-Threonine Kinases , Receptor, Transforming Growth Factor-beta Type II , Transforming Growth Factor beta/antagonists & inhibitorsABSTRACT
A novel naturally occurring antiangiogenic agent isolated from cartilage, referred to as Neovastat (AE-941), was examined for its efficacy against tumor neovascularization and progression. Exposure to Neovastat results in ex ovo antiangiogenic properties in the chorioallantoid membrane of chicken embryo (71% decrease in the angiogenic index as compared to the basic fibroblast growth factor (bFGF) treated control embryos, P < 0.0001). Oral administration of Neovastat inhibits bFGF-induced angiogenesis in the Matrigel mouse model (87.5% decrease in hemoglobin as compared to the bFGF-treated control implants, P < 0.0001). Neovastat also induces a dose response decrease of lung metastases in the Lewis lung carcinoma model (oral administration; 69.1% of inhibition obtained at the maximal dose of 0.5 ml/day, P < 0.0001). Combined with a sub-optimal dose of cisplatinum (2 mg/kg, i.p.), Neovastat (0.5 ml/day) improved the therapeutic index by increasing the antimetastatic efficacy and by exerting a protective activity against cisplatinum-induced body weight loss and myelosuppression. In summary, our experimental data provide evidence of antiangiogenic and antimetastatic properties of Neovastat, following oral administration.
Subject(s)
Angiogenesis Inhibitors/pharmacology , Antineoplastic Agents/pharmacology , Blood Vessels/drug effects , Neovascularization, Pathologic/prevention & control , Tissue Extracts/pharmacology , Administration, Oral , Angiogenesis Inhibitors/isolation & purification , Animals , Antineoplastic Agents/isolation & purification , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Body Weight/drug effects , Carcinoma, Lewis Lung/blood supply , Carcinoma, Lewis Lung/drug therapy , Carcinoma, Lewis Lung/pathology , Cartilage/chemistry , Chick Embryo , Cisplatin/administration & dosage , Collagen , Dose-Response Relationship, Drug , Drug Combinations , Female , Fibroblast Growth Factor 2/toxicity , Humans , Laminin , Mice , Mice, Inbred BALB C , Proteoglycans , Tissue Extracts/isolation & purificationABSTRACT
PURPOSE: Vascular endothelial growth factor (VEGF) is a potent regulator of angiogenesis, which exerts direct effects on vascular endothelial cells, including endothelial cell proliferation and survival, tubulogenesis, and vascular permeability. In this study, we examined whether Neovastat, a naturally occurring multifunctional antiangiogenic drug, could inhibit the endothelial cell response to VEGF stimulation. RESULTS: We demonstrated that Neovastat was able to block the VEGF-dependent microvessel sprouting from Matrigel-embedded rat aortic rings, and it also blocked the VEGF-induced endothelial cell tubulogenesis in vitro. In vivo studies showed that Neovastat was able to specifically inhibit VEGF-induced plasma extravasation in numerous tissues, including pancreas and skin. The mechanism of action of Neovastat on VEGF-mediated effects was also evaluated at the molecular level. Neovastat was shown to compete against the binding of VEGF to its receptor in endothelial cells and significantly inhibited the VEGF-dependent tyrosine phosphorylation of VEGF receptor-2, whereas it had no significant effect on VEGF receptor-1 activity. Moreover, the inhibition of receptor phosphorylation was correlated with a marked decrease in the ability of VEGF to induce pERK activation. Neovastat does not compete against the binding of basic fibroblast growth factor, indicating a preferential inhibitory effect on the VEGF receptor. CONCLUSIONS: Because Neovastat was shown previously to inhibit metalloproteinase activities, these results suggest that Neovastat is able to target multiple steps in tumor neovascularization, further emphasizing its use as a pleiotropic, multifunctional antiangiogenic drug.
