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Selective and effective killing of angiogenic vascular endothelial cells and cancer cells by targeting tissue factor using a factor VII-targeted photodynamic therapy for breast cancer.

ABSTRACT: The cell surface receptor tissue factor (TF) is regarded as a common but specific target on angiogenic tumor vascular endothelial cells (VECs) and tumor cells in many types of cancer including breast cancer. The purpose of this study is to develop a selective and effective TF-targeting photodynamic therapy (PDT) by using its natural ligand factor VII (fVII)-conjugated Sn(IV) chlorin e6 (SnCe6) for the treatment of breast cancer. A cross linker EDC was used to covalently conjugate fVII protein to SnCe6, and the binding activity and phototoxicity was confirmed by ELISA and in vitro PDT. The efficacy of fVII-tPDT was assessed in vitro by crystal violet staining assay and in vivo by measuring tumor size in mice carrying murine or human breast cancer xenografts. We show that active site-mutated (K341A) fVII protein can be internalized into breast cancer cells and vascular endothelial growth factor (VEGF)-stimulated human umbilical vein endothelial cells (HUVECs) as angiogenic VECs. fVII-tPDT not only enhances 12-fold the in vitro efficacy but also selectively and effectively kills angiogenic HUVECs and breast cancer cells via specifically binding of fVII to TF and inducing apoptosis and necrosis as the underlying mechanism. Furthermore, fVII-tPDT can significantly inhibit the tumor growth of murine and human breast cancer without obvious toxicities in mice. We conclude that fVII-tPDT using fVII-SnCe6 conjugate can selectively and effectively kill angiogenic VECs and breast cancer cells in vitro and significantly inhibit the tumor growth of murine and human breast cancer in mice.

SUBMITTER: Hu Z

PROVIDER: S-EPMC3951776 | biostudies-literature | 2011 Apr

REPOSITORIES: biostudies-literature

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Selective and effective killing of angiogenic vascular endothelial cells and cancer cells by targeting tissue factor using a factor VII-targeted photodynamic therapy for breast cancer.

Hu Zhiwei Z Rao Benqiang B Chen Shimin S Duanmu Jinzhong J

Breast cancer research and treatment 20100601 3

The cell surface receptor tissue factor (TF) is regarded as a common but specific target on angiogenic tumor vascular endothelial cells (VECs) and tumor cells in many types of cancer including breast cancer. The purpose of this study is to develop a selective and effective TF-targeting photodynamic therapy (PDT) by using its natural ligand factor VII (fVII)-conjugated Sn(IV) chlorin e6 (SnCe6) for the treatment of breast cancer. A cross linker EDC was used to covalently conjugate fVII protein to ...[more]

PMID: 20514515

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Project description:The objective of this study was to develop a ligand-targeted photodynamic therapy (tPDT) by conjugating factor VII (fVII) protein with photosensitiser verteporfin in order to overcome the poor selectivity and enhance the effect of non-targeted PDT (ntPDT) for cancer. fVII is a natural ligand for receptor tissue factor (TF) with high affinity and specificity. The reason for targeting receptor TF for the development of tPDT is that TF is a common but specific target on angiogenic tumour vascular endothelial cells (VEC) and many types of tumour cells, including solid tumours and leukaemia.Murine factor VII protein (mfVII) containing a mutation (Lys341Ala) was covalently conjugated via a cross linker EDC with Veterporfin (VP) that was extracted from liposomal Visudyne, and then free VP was separated by Sephadex G50 spin columns. fVII-tPDT using mfVII-VP conjugate, compared to ntPDT, was tested in vitro for the killing of breast cancer cells and VEGF-stimulated VEC and in vivo for inhibiting the tumour growth of breast tumours in a mouse xenograft model.We showed that: (i) fVII protein could be conjugated with VP without affecting its binding activity; (ii) fVII-tPDT could selectively kill TF-expressing breast cancer cells and VEGF-stimulated angiogenic HUVECs but had no side effects on non-TF expressing unstimulated HUVEC, CHO-K1 and 293 cells; (iii) fVII targeting enhanced the effect of VP PDT by three to four fold; (iii) fVII-tPDT induced significantly stronger levels of apoptosis and necrosis than ntPDT; and (iv) fVII-tPDT had a significantly stronger effect on inhibiting breast tumour growth in mice than ntPDT.We conclude that the fVII-targeted VP PDT that we report here is a novel and effective therapeutic with improved selectivity for the treatment of breast cancer. Since TF is expressed on many types of cancer cells including leukaemic cells and selectively on angiogenic tumour VECs, fVII-tPDT could have broad therapeutic applications for other solid cancers and leukaemia.

Project description:BackgroundThe adipocyte-derived hormone leptin influences the behaviour of a wide range of cell types and is now recognised as a pro-angiogenic and pro-inflammatory factor. In the vasculature, these effects are mediated in part through its direct leptin receptor (ObRb)-driven actions on endothelial cells (ECs) but the mechanisms responsible for these activities have not been established. In this study we sought to more fully define the molecular links between inflammatory and angiogenic responses of leptin-stimulated human ECs.Methodology/principal findingsImmunoblotting studies showed that leptin increased cyclo-oxygenase-2 (COX-2) expression (but not COX-1) in cultured human umbilical vein ECs (HUVEC) through pathways that depend upon activation of both p38 mitogen-activated protein kinase (p38(MAPK)) and Akt, and stimulated rapid phosphorylation of vascular endothelial growth factor receptor 2 (VEGFR2) on Tyr(1175). Phosphorylation of VEGFR2, p38(MAPK) and Akt, and COX-2 induction in cells challenged with leptin were blocked by a specific leptin peptide receptor antagonist. Pharmacological inhibitors of COX-2, the phosphatidylinositol 3-kinase (PI3K)/Akt pathway and p38(MAPK) abrogated leptin-induced EC proliferation (assessed by quantifying 5-bromo-2'-deoxyuridine incorporation, calcein fluorescence and propidium iodide staining), slowed the increased migration rate of leptin-stimulated cells (in vitro wound healing assay) and inhibited leptin-induced capillary-like tube formation by HUVEC on Matrigel. Inhibition of VEGFR2 tyrosine kinase activity reduced leptin-stimulated p38(MAPK) and Akt activation, COX-2 induction, and pro-angiogenic EC responses, and blockade of VEGFR2 or COX-2 activities abolished leptin-driven neo-angiogenesis in a chick chorioallantoic membrane vascularisation assay in vivo.Conclusions/significanceWe conclude that a functional endothelial p38(MAPK)/Akt/COX-2 signalling axis is required for leptin's pro-angiogenic actions and that this is regulated upstream by ObRb-dependent activation of VEGFR2. These studies identify a new function for VEGFR2 as a mediator of leptin-stimulated COX-2 expression and angiogenesis and have implications for understanding leptin's regulation of the vasculature in both non-obese and obese individuals.

Project description:BACKGROUND:Secretoneurin is a neuropeptide located in nerve fibers along blood vessels, is upregulated by hypoxia, and induces angiogenesis. We tested the hypothesis that secretoneurin gene therapy exerts beneficial effects in a rat model of myocardial infarction and evaluated the mechanism of action on coronary endothelial cells. METHODS AND RESULTS:In vivo secretoneurin improved left ventricular function, inhibited remodeling, and reduced scar formation. In the infarct border zone, secretoneurin induced coronary angiogenesis, as shown by increased density of capillaries and arteries. In vitro secretoneurin induced capillary tubes, stimulated proliferation, inhibited apoptosis, and activated Akt and extracellular signal-regulated kinase in coronary endothelial cells. Effects were abrogated by a vascular endothelial growth factor (VEGF) antibody, and secretoneurin stimulated VEGF receptors in these cells. Secretoneurin furthermore increased binding of VEGF to endothelial cells, and binding was blocked by heparinase, indicating that secretoneurin stimulates binding of VEGF to heparan sulfate proteoglycan binding sites. Additionally, secretoneurin increased binding of VEGF to its coreceptor neuropilin-1. In endothelial cells, secretoneurin also stimulated fibroblast growth factor receptor-3 and insulin-like growth factor-1 receptor, and in coronary vascular smooth muscle cells, we observed stimulation of VEGF receptor-1 and fibroblast growth factor receptor-3. Exposure of cardiac myocytes to hypoxia and ischemic heart after myocardial infarction revealed increased secretoneurin messenger RNA and protein. CONCLUSIONS:Our data show that secretoneurin acts as an endogenous stimulator of VEGF signaling in coronary endothelial cells by enhancing binding of VEGF to low-affinity binding sites and neuropilin-1 and stimulates further growth factor receptors like fibroblast growth factor receptor-3. Our in vivo findings indicate that secretoneurin may be a promising therapeutic tool in ischemic heart disease.

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Selective and effective killing of angiogenic vascular endothelial cells and cancer cells by targeting tissue factor using a factor VII-targeted photodynamic therapy for breast cancer.

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Selective and effective killing of angiogenic vascular endothelial cells and cancer cells by targeting tissue factor using a factor VII-targeted photodynamic therapy for breast cancer.

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