The purpose of this review is to provide an overview of the effect of (lymph)angiogenic cytokines on hematopoietic cells involved in acute myeloid leukemia (AML). Like angiogenesis, lymphangiogenesis occurs in pathophysiological conditions but not in healthy adults. AML is closely associated with the vasculature system, and the interplay between lymphangiogenic cytokines maintains leukemic blast survival in the bone marrow (BM). Once AML is induced, proangiogenic cytokines function as angiogenic or lymphangiogenic factors and affect hematopoietic cells, including BM-derived immune cells. Simultaneously, the representative cytokines, VEGFs and their receptors are expressed on AML blasts in vascular and osteoblast niches in both the BM and the peripheral circulation. After exposure to (lymph)angiogenic cytokines in leukemogenesis and infiltration, immune cell phenotypes and functions are affected. These dynamic behaviors in the BM reflect the clinical features of AML. In this review, we note the importance of lymphangiogenic factors and their receptors in hematopoietic cells in AML. Understanding the functional characterization of (lymph)angiogenic factors in the BM niche in AML will also be helpful in interrupting the engraftment of leukemic stem cells and for enhancing immune cell function by modulating the tumor microenvironment.
Animals ; Cytokines/*immunology ; Hematopoietic Stem Cells/immunology/*pathology ; Humans ; Immunity, Cellular ; Leukemia, Myeloid, Acute/immunology/*physiopathology ; *Lymphangiogenesis ; Lymphatic Vessels/immunology/*physiopathology ; Vascular Endothelial Growth Factor A/immunology

Bispecific antibodies have been exploited as both cancer immunodiagnostics and cancer therapeutics, which have shown promises in clinical trials in cancer imaging and therapy. To improve the anti-tumor effect, an scDb (bispecific single-chain diabody) was constructed from the variable domain genes of two scFvs (single-chain variable fragment antibodies) directed against human EGFR (epidermal growth factor receptor) and VEGFR2 (vascular endothelial growth factor receptor 2) extracellular domains. The anti-EGFR/ anti-KDR scDb was constructed into pHEN2 plasmid and expressed in Escherichia coli HB2151 host. After purification by one-step affinity chromatography of IMAC, scDb protein was characterized by Western blotting. The yield of scDb protein was 570 microg per liter medium. scDb bound to EGFR as efficiently as the parental antibody scFv-E10, while a little bit weaker than the parental antibody scFv-AK404R when bound to KDR. In conclusion, the scDb protein could bind both EGFR and KDR specifically and could be applied for further anti-tumor research.
Antibodies, Bispecific ; biosynthesis ; genetics ; Escherichia coli ; metabolism ; Humans ; Plasmids ; Protein Binding ; Receptor, Epidermal Growth Factor ; immunology ; Single-Chain Antibodies ; biosynthesis ; genetics ; Vascular Endothelial Growth Factor Receptor-2 ; immunology

Monoclonal antibodies (mAbs) have become a major class of therapeutic agents providing effective alternatives to treating various human diseases. To date, 15 mAbs have been approved by regulatory agencies in the world for clinical use in oncology indications. The selectivity and specificity, the unique pharmacokinetics, and the ability to engage and activate the host immune system differentiate these biologics from traditional small molecule anticancer drugs. mAb-based regimens have brought clinical benefits, including improvements in overall survival, to patients with a variety of cancers. Many challenges still remain, however, to fully realize the potential of these new medicines. With our further understanding of cancer biology, mechanism of antibody action, and advancement of antibody engineering technologies, many novel antibody formats or antibody-derived molecules are emerging as promising new generation therapeutics. Carefully designed and engineered, they retain the advantage of specificity and selectivity of original antibodies, but in the meantime acquire additional special features such as improved pharmacokinetics, increased selectivity, and enhanced anticancer efficacy. Promising clinical results are being generated with these newly improved antibody-based therapeutics.
Antibodies, Monoclonal ; therapeutic use ; Antibodies, Monoclonal, Humanized ; therapeutic use ; Antigens, CD20 ; immunology ; Humans ; Immunoconjugates ; therapeutic use ; Neoplasms ; drug therapy ; immunology ; Protein Engineering ; RANK Ligand ; immunology ; Receptor, Epidermal Growth Factor ; immunology ; Receptor, ErbB-2 ; immunology ; Vascular Endothelial Growth Factor A ; immunology

OBJECTIVETo prepare a neutralizing monoclonal antibody (McAb) against vascular endothelial growth factor receptor KDR and study its biological activity.

METHODSExtracellular immunoglobulin (Ig)-like domain III of KDR (KDR III) was expressed in E. coli and purified by affinity chromatograph. Monoclonal antibody against KDR III was prepared by hybridoma technique. ELISA and FACS analysis were used to identify its specificity. Immunoprecipitation and [(3)H]-TdR incorporation assay were also used to detect the activity of anti-KDR McAb blocking the phosphorylation of KDR tyrosine kinase receptor and the influence on VEGF-induced mitogenesis of human endothelial cells.

RESULTSMcAb Ycom1D3 against KDR III was prepared which bound specifically to both the soluble KDR III and the cell-surface expressed KDR. It effectively blocked VEGF/KDR interaction and inhibited VEGF-stimulated activation of KDR expression on human endothelial cells. Furthermore, Ycom1D3 efficiently neutralized VEGF-induced mitogenesis of human umbilical vascular endothelial cells.

CONCLUSIONMcAb Ycom1D3 against KDR III may suppress the action of VEGF by blocking native vascular endothelial growth factor receptor KDR. It has potential clinical applications in the treatment of cancers and other diseases where pathological angiogenesis is involved.

Antibodies, Monoclonal ; pharmacology ; Cell Proliferation ; drug effects ; Cells, Cultured ; Endothelial Cells ; cytology ; Humans ; Neovascularization, Physiologic ; Umbilical Veins ; cytology ; Vascular Endothelial Growth Factor A ; metabolism ; Vascular Endothelial Growth Factor Receptor-2 ; immunology

The objective of this study was to investigate the synergistic effect of soluble human recombinant tumor necrosis factor related apoptosis inducing ligand (TRAIL) protein combined with anti-vascular endothelial growth factor (anti-VEGF) antibody on inducing apoptosis of leukemia K562 cells. The inhibitory rates and apoptotic rates of K562 cells treated with TRAIL and anti-VEGF antibody alone and their combination for 48 hours were examined by CCK-8 assay and flow cytometry respectively. The results indicated that the apoptotic rates of K562 cells induced with 75, 100 and 150 ng/ml TRAIL after culture for 48 hours were (4.26±0.67)%, (8.91±0.55)% and (11.71±0.78)% respectively. The apoptotic rates of K562 cells induced with 2.5, 5 and 7.5 µg/ml anti-VEGF antibody after culture for 48 hours were (3.95±0.69)%, (7.98±0.74)% and (10.26±0.83)% respectively. The apoptotic rates of K562 cells treated with combination use of 2.5 µg/ml anti-VEGF antibody and 75 ng/ml TRAIL, 5 µg/ml anti-VEGF antibody and 100 ng/ml TRAIL, and 7.5 µg/ml anti-VEGF antibody and 150 ng/ml TRAIL for 48 hours were (22.16±0.93)%, (36.32±1.31)% and (49.19±0.71)% respectively. The combined use of above mentioned agents induced significantly higher apoptosis and cytotoxicity than that of TRAIL or anti-VEGF antibody alone (p<0.05). It is concluded that the combination use of TRAIL and anti-VEGF antibody can significantly increase the sensitivity of K562 cells to apoptosis.
Antibodies, Monoclonal ; pharmacology ; Apoptosis ; drug effects ; Humans ; K562 Cells ; TNF-Related Apoptosis-Inducing Ligand ; pharmacology ; Vascular Endothelial Growth Factor A ; immunology

To investigate the effects of DENV infection on the expression of Vascular endothelial growth factor (VEGF) in monocytes, and to explore which innate immune signaling pathway is responsible for VEGF induction. Real-time PCR was used to determine the expression levels of VEGF in DENV-infected THP-1. We found that different serotype viruses (DENV1, DENV2, DENV3) induced the VEGF expression. Moreover, VEGF expression was significantly increased in human primary monocytes infected with DENV 2. In addition, VEGF induction by DENV2 was significantly impaired by knockdown of TLR3 and interferon-beta promoter stimulator 1 (IPS-1), or by inhibition of ERK, JNK or NF-kappaB. These results demonstrated that DENV induced VEGF expression in monocytes, and the activation of TLR3, IPS-1 signal pathways were required for DENV2-triggered VEGF induction, suggesting that VEGF might be a promising therapeutic target for DHF.
Dengue ; genetics ; immunology ; virology ; Dengue Virus ; classification ; genetics ; physiology ; Humans ; MAP Kinase Signaling System ; Monocytes ; NF-kappa B ; genetics ; immunology ; Up-Regulation ; Vascular Endothelial Growth Factor A ; genetics ; immunology

OBJECTIVETo investigate whether fetal bone marrow stromal cells have hemangioblastic characteristics.

METHODSHuman fetal bone marrow stromal cells (hfMSCs) were isolated and cultured. Immunophenotypes of hfMSCs were tested by FACS. hfMSCs seeded in the matrigel were induced with vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) in vitro. Vascularization and hematopoiesis were detected with immunohistochemistry and electron microscope.

RESULTSThe typical properties of this CD34- stromal cell population were that 99% cells expressed Flk1 (vascular endothelial cell growth factor receptor 2) and tube structure was formed. In the process of induction, hfMSCs could give rise to CD34+ round cells.

CONCLUSIONSWe have demonstrated that fetal bone marrow stroma-derived Flk1+ CD34- cells could differentiate into vascular endothelial cells and hematopoietic cells, indicating that fetal bone marrow stroma-derived Flk1+ CD34- cells have hemangioblastic characteristics.

Antigens, CD34 ; immunology ; Bone Marrow Cells ; cytology ; immunology ; metabolism ; Cell Differentiation ; Cells, Cultured ; Fetus ; Fibroblast Growth Factor 2 ; metabolism ; Hematopoiesis ; Hematopoietic Stem Cells ; cytology ; immunology ; metabolism ; Humans ; Immunophenotyping ; Stromal Cells ; cytology ; immunology ; metabolism ; Vascular Endothelial Growth Factor Receptor-2 ; physiology

OBJECTIVETo investigate the effect of vasoactive intestinal peptide (VIP) on angiogenesis after focal cerebral ischemia.

METHODSFocal cerebral ischemia was induced by middle cerebral artery occlusion (MCAO) for 120 min in adult SD rats with intracerebroventricular VIP administration at the beginning of reperfusion. Immunohistochemistry was performed to assay BrdU immunoreactive endothelial cells, expressions of VEGF, flt-1 and flk-1 in the ischemic zone, and the protein expressions of vascular endothelial growth factor (VEGF) in the brain was measured using Western blotting.

RESULTSImmunohistochemical staining revealed significantly increased BrdU immunoreactive endothelial cells on the margins of the ischemic lesion in rats treated with VIP as compared with that in the control rats (P<0.05). VIP significantly increased the number of VEGF immunoreactive cells and flt-1- and flk-1-positive endothelial cells in comparison with the control group (P<0.01). Western blotting showed that VIP treatment resulted in significantly increased VEGF protein level in the ipsilateral hemisphere (P<0.05).

CONCLUSIONSVIP enhances angiogenesis in the ischemic brain by increasing the expressions of VEGF in the brain tissue and its receptors flt-1 and flk-1 in the endothelial cells.

Animals ; Brain ; drug effects ; metabolism ; Brain Ischemia ; immunology ; metabolism ; pathology ; physiopathology ; Endothelial Cells ; drug effects ; metabolism ; Gene Expression Regulation ; drug effects ; Male ; Neovascularization, Physiologic ; drug effects ; Rats ; Rats, Sprague-Dawley ; Vascular Endothelial Growth Factor A ; metabolism ; Vascular Endothelial Growth Factor Receptor-1 ; metabolism ; Vascular Endothelial Growth Factor Receptor-2 ; metabolism ; Vasoactive Intestinal Peptide ; pharmacology

The suppressive effect of anti-KDR antibody against VEGF on proliferation of hemangioma-derived vascular endothelial cells (HVECs) was investigated. HVECs from one case of hemangioma in proliferative phase were cultured. Both primary culture and sub-culture were conducted in M199 medium. The HVECs of passage 3 were divided into 4 groups based on the concentrations of anti-KDR antibody. Cell count was performed and inhibitory rate of HVECs was measured before and 9 days after interference. The results showed that the number of HVECs in the anti-KDR antibody-treated groups was significantly decreased and the inhibitory rate of HVECs by anti-KDR antibody (50, 10 and 2 microg/mL) was 84%, 63% and 39% respectively at 9th day after interference, with the difference being significant. In the control group, the number of HVECs was increased significantly. In was concluded that the anti-KDR antibody could suppress the activity of VEGF through blocking the KDR, indicating the potential clinical applications of anti-KDR antibody in the treatment of hemangioma.
Antibodies/*pharmacology ; Cell Proliferation/drug effects ; Endothelium, Vascular/*pathology ; Hemangioma/*pathology ; Tumor Cells, Cultured ; Vascular Endothelial Growth Factor Receptor-2/*immunology

OBJECTIVETo investigate the effect of anti-VEGF antibody on angiogenesis induced by osteosarcoma OS-732 cell line and tumor growth.

METHODSWith a tumor model on the chick embryo chorioallantoic membrane (CAM), the inhibition of angiogenesis and tumor growth by anti-VEGF antibody were observed under a stero-microscope and a light microscope. Furthermore, the proliferation and apoptosis in tumor cells and endothelial cells (EC) were studied by TdT-mediated duTP nick and labeling (TUNEL) and immunohistochemical staining using proliferating cell nuclear antigen (PCNA) monoclonal antibody.

RESULTSVEGF polyclonal antibody administration in tumor-bearing chick embryo resulted in growth arrest of xenografts and a markedly reduction in the new capillaries which converged upon the tumor. The tumor cell apoptotic index was higher in the anti-VEGF antibody treated group than the negative control group, but the proliferation index was not significantly different between them. At the same time, increased apoptosis and decreased proliferation in EC were also noted.

CONCLUSIONVEGF polyclonal antibody is able to inhibit the angiogenesis induced by OS-732 obviously, probably by the mechanism of inhibition of EC proliferation and promotion of their apoptosis, furtherly, which may contribute to the apoptosis of tumor cells and result in suppression of tumor growth.

Animals ; Antibodies ; therapeutic use ; Apoptosis ; Bone Neoplasms ; therapy ; Cell Division ; drug effects ; Chick Embryo ; Disease Models, Animal ; Endothelial Growth Factors ; immunology ; Endothelium, Vascular ; cytology ; drug effects ; Immunotherapy ; Lymphokines ; immunology ; Microscopy ; Neovascularization, Pathologic ; prevention & control ; Osteosarcoma ; therapy ; Vascular Endothelial Growth Factor A ; Vascular Endothelial Growth Factors