以血管内皮生长因子及其受体为靶向的肿瘤治疗

实体瘤的生长和转移有赖于新生血管生成。在血管生成的过程中,血管内皮生长因子（vascular endothelial growth factor,VEGF）是作用最强的促血管生成因子之一,它主要通过与血管内皮细胞上的特异性受体-2（VEGFreceptor-2,VEGFR-2）结合促进内皮细胞增殖、迁移,引发新生血管生成。近10年来,随着分子生物学及相关技术的发展与应用,以VEGF/VEGFR为靶向的抗肿瘤血管生成治疗策略,成为肿瘤治疗研究的热点。     

VEGF/VEGFR2信号转导通路在抗肿瘤血管生成中的作用

以肿瘤血管内皮细胞为靶点抑制肿瘤血管生成从而阻断肿瘤血液供应已成为当前抗肿瘤生长和转移的研究热点.其中,VEGF/VEGFR2信号转导通路在肿瘤周围血管生成起主要作用.阻断该信号通路,能够抑制实体瘤的生长和转移.本文就VEGF/VEGFR2信号通路在抗肿瘤生长和转移中的研究进展作一概述.     

血管内皮生长因子受体与肝细胞癌侵袭转移的关系

血管内皮生长因子受体(VEGFR)含3种受体酪氨酸激酶(RTK)超家族成员和2种非RTK超家族成员,其与配体以非一一对应形式相互结合,通过细胞表面受体内化作用介导Raf1→MAP2K1/2→ERK1/2等多条信号传导通路,引起肝癌细胞增殖和血管、淋巴管生成。VEGFR在肝癌患者局部癌灶高表达,是介导肝细胞癌恶性增生、侵袭转移的关键因素,与患者无进展生存期呈负相关。基质金属蛋白酶-9、热休克蛋白90β可上调VEGFR促进肝癌细胞增殖转移,而miR-203a、miR-378a、miR-199a-3p等可下调VEGFR表达抑制肝癌细胞浸润。基于VEGFR靶向药物治疗可诱导肝癌细胞凋亡,阻断肿瘤血管新生,延缓患者病情进展。     

血管内皮生长因子及其受体的生物学特点

<正>血管内皮生长因子(vascular endothelial growth factor,VEGF)是胚胎形成、骨骼生长和生殖功能过程中血管生成的重要调节因子,与肿瘤、眼内新生血管性等疾病有关[1]。VEGF的生物学活性受2种酪氨酸激酶受体(tyrosine kinases)的调节:VEGFR-1和VEGF-2,这2种受体的信号学活性差异非常大。目前,多种VEGF抑制因子在进行恶性肿瘤临床实验,人们试图通过抑制VEGF降低血管生成、血管渗漏等。VEGF和VEGFR     

血管内皮生长因子受体信号转导通路与生物学效应

血管生成是指从已存在的血管中以出芽的方式生成新血管的过程.血管内皮细胞生长因子(VEGF)能促进生理和病理的血管生成.VEGF有两个主要的酪氨酸激酶受体VEG-FR-1(Flt-1)和VEGFR-2(KDR/Flk-1).VEGF的生物学效应主要是通过VEGFR-2实现的,多种信号蛋白的激活均与VEGF的信号转导有关,涉及促内皮细胞生存、增殖、迁移,促进血管渗透增加,促进NO合成及释放等多项生物学功能.     

血管内皮生长因子和肺癌

肿瘤血管生成在肺癌的生长和转移中发挥着重要的作用,众多肿瘤血管生成因子和抑制因子参与了肿瘤血管生成的调控,其中,血管内皮生长因子(VEGF)是己知最重要的血管内皮细胞有丝分裂素.本文就肺癌血管生成中血管内皮生长因子的作用机制、诱导因素及其抗血管生成治疗作一简要综述.     

大黄素对结肠癌血管内皮生长因子受体的影响及对结肠癌抑制作用的研究

[目的]研究大黄素对血管内皮生长因子(VEGF)受体(VEGFR)阻断作用及对结肠癌细胞抑制作用的机制.[方法]软琼脂集落实验法,流式细胞术检测大黄素对结肠癌细胞增殖和凋亡的影响;酪氨酸激酶活性分析,Western blot方法检测大黄素对VEGFR的抑制作用.[结果]大黄素抑制结肠癌细胞生长,引起细胞凋亡,凋亡率由对照组(0 μmol/L)的(8.1±2.7)%上升至大黄素40 μmol/L时的(27.8±10.9)%(P＜0.01),呈浓度依赖性.大黄素抑制VEGFR酪氨酸激酶活性,VEGFR-1相对活性由对照组的100%降至大黄素40μmol/L时的22.4%(P＜0.01),VEGFR-2降至58.5%(P＜0.01),VEGFR-3降至31.6%(P＜0.01),大黄素作用后,VEGFR酪氨酸磷酸化状态蛋白量减少.[结论]大黄素能够通过抑制VEGFR酪氨酸激酶活性而抑制结肠癌生长,可作为一种有效的肿瘤血管生成抑制剂.     

不同转移潜能肝癌细胞株血管内皮生长因子及其受体的表达和意义

目的 探讨血管内皮生长因子受体-1(VEGFR-1)在不同转移潜能肝癌细胞株中的表达及其意义.方法 应用半定量RT-PCR、酶联免疫吸附试验和(或)Western blot检测4株不同转移潜能的肝癌细胞株及一株正常肝细胞中VEGFR-1、VEGF-A、VEGF-B及VEGFR-2的mRNA和(或)蛋白质表达.结果 MHCC97-H、MHCC97-L和SMMC-7721细胞均有VEGFR-1 mRNA和蛋白质表达,且VEGFR-1 mRNA和蛋白质在MHCC97-H中的表达高于MHCC97-L、SMMC-7721的表达,两者比较,差异有统计学意义(P＜0.05),而其配体VEG-A和VEGF-B在检测的4种肝癌细胞株和正常肝细胞株L-02中均有表达.同时在检测的四种肝癌细胞株和正常肝细胞株L-02中均能检测到VEGFR-2 mRNA和蛋白质表达,但各组间表达差异无统计学意义(P＞0.05).结论 具有转移潜能的肝癌细胞株有VEGFR-1表达,而且其表达强弱与肝癌细胞株的转移潜能呈正相关,VEGFR-1的表达可能促进了肝细胞癌的侵袭转移.     

肿瘤血管生成及其抗血管生成治疗的研究进展

肿瘤的生长和转移依赖于新生血管形成，血管内皮生长因子及其受体是目前发现的最重要的促肿瘤血管生长因子，在肿瘤血管生成过程中发挥关键作用。鉴于肿瘤血管生成在肿瘤生长、浸润和转移中的重要作用，近年来开始了抗血管生成治疗肿瘤的新方法——抗血管生成疗法。本文着重综述了肿瘤血管生成及其抗肿瘤血管生成治疗的研究进展。     

乳腺浸润性导管癌中T淋巴瘤侵袭转移诱导基因对淋巴管生成的作用

目的检测乳腺浸润性导管癌中T淋巴瘤侵袭转移诱导基因(Tiam)1与血管内皮生长因子受体(VEGFR)-3、血管内皮生长因子(VEGF)-D表达的关系,探讨Tiam1对肿瘤淋巴管生成的作用。方法确诊为乳腺浸润性导管癌新鲜肿瘤标本70例为观察组,收集正常乳腺新鲜组织46例为对照组,采用流式细胞术检测Tiam1与VEGFR-3、VEGF-D表达,分析三者在不同临床病理特征中表达差别。结果观察组Tiam1、VEGFR-3和VEGF-D表达量明显高于对照组,观察组Tiam1、VEGFR-3和VEGF-D表达量与肿瘤体积、脉管浸润、雌激素受体(ER)、孕激素受体(PR)、Ki67表达密切相关,观察组Tiam1、VEGFR-3和VEGF-D表达有正相关性,生存分析显示Tiam1、VEGFR-3和VEGF-D表达与预后相关。结论Tiam1可促进VEGFR-3和VEGF-D表达,进而促进肿瘤淋巴管生成,三者与肿瘤进展密切相关,术后联合检测Tiam1、VEGFR-3和VEGF-D可能对判断乳腺浸润性导管癌预后有一定价值。     

Inhibition of both paracrine and autocrine VEGF/ VEGFR-2 signaling pathways is essential to induce long-term remission of xenotransplanted human leukemias

Antiangiogenic agents block the effects of tumor-derived angiogenic factors (paracrine factors), such as vascular endothelial growth factor (VEGF), on endothelial cells (EC), inhibiting the growth of solid tumors. However, whether inhibition of angiogenesis also may play a role in liquid tumors is not well established. We recently have shown that certain leukemias not only produce VEGF but also selectively express functional VEGF receptors (VEGFRs), such as VEGFR-2 (Flk-1, KDR) and VEGFR1 (Flt1), resulting in the generation of an autocrine loop. Here, we examined the relative contribution of paracrine (EC-dependent) and autocrine (EC-independent) VEGF/VEGFR signaling pathways, by using a human leukemia model, where autocrine and paracrine VEGF/VEGFR loops could be selectively inhibited by neutralizing mAbs specific for murine EC (paracrine pathway) or human tumor (autocrine) VEGFRs. Blocking either the paracrine or the autocrine VEGF/VEGFR-2 pathway delayed leukemic growth and engraftment in vivo, but failed to cure inoculated mice. Long-term remission with no evidence of disease was achieved only if mice were treated with mAbs against both murine and human VEGFR-2, whereas mAbs against human or murine VEGFR-1 had no effect on mice survival. Therefore, effective antiangiogenic therapies to treat VEGF-producing, VEGFR-expressing leukemias may require blocking both paracrine and autocrine VEGF/VEGFR-2 angiogenic loops to achieve remission and long-term cure.     

Placental growth factor neutralising antibodies give limited anti-angiogenic effects in an in vitro organotypic angiogenesis model

Vascular Endothelial Growth Factor Receptor (VEGFR) mediated signalling drives angiogenesis. This is predominantly attributed to the activity of VEGFR-2 following binding of VEGF-A. Whether other members of the VEGFR and ligand families such as VEGFR-1 and its ligand Placental Growth Factor (PlGF) can also contribute to developmental and pathological angiogenesis is less clear. We explored the function of PlGF in VEGF-A dependent angiogenesis using an in vitro co-culture assay in which endothelial cells are cultured on a fibroblast feeder layer. In the presence of 2% FS MCDB media (containing limited growth factors) in vitro endothelial tube formation is driven by endogenous angiogenic stimuli which are produced by the fibroblast and endothelial cells. Under these conditions independent sequestration of either free VEGF-A or PlGF with polyclonal and monoclonal antibodies inhibited tube formation suggesting that both ligands are required to drive an angiogenic response. Endothelial tube formation could only be driven within this assay by the addition of exogenous VEGF-A, VEGF-E or VEGF-A/PlGF heterodimer, but not by PlGF alone, implying that activation of either VEGFR-2/VEGFR-1 heterodimers or VEGFR-2 homodimers were responsible for eliciting an angiogenic response directly, but not VEGFR-1 homodimers. In contrast to results obtained with an endogenous angiogenic drive, sequestration of PlGF did not affect endothelial tube formation when the assay was driven by 1 ng/ml exogenous VEGF-A. These data suggest that although neutralising PlGF can be shown to reduce endothelial tube formation in vitro, this effect is only observed under restricted culture conditions and is influenced by VEGF-A. Such data questions whether neutralising PlGF would have a therapeutic benefit in vivo in the presence of pathological concentrations of VEGF-A.     

Adenoviral expression of vascular endothelial growth factor-C induces lymphangiogenesis in the skin

The growth of blood and lymphatic vasculature is mediated in part by secreted polypeptides of the vascular endothelial growth factor (VEGF) family. The prototype VEGF binds VEGF receptor (VEGFR)-1 and VEGFR-2 and is angiogenic, whereas VEGF-C, which binds to VEGFR-2 and VEGFR-3, is either angiogenic or lymphangiogenic in different assays. We used an adenoviral gene transfer approach to compare the effects of these growth factors in adult mice. Recombinant adenoviruses encoding human VEGF-C or VEGF were injected subcutaneously into C57Bl6 mice or into the ears of nude mice. Immunohistochemical analysis showed that VEGF-C upregulated VEGFR-2 and VEGFR-3 expression and VEGF upregulated VEGFR-2 expression at 4 days after injection. After 2 weeks, histochemical and immunohistochemical analysis, including staining for the lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1), the vascular endothelial marker platelet-endothelial cell adhesion molecule-1 (PECAM-1), and the proliferating cell nuclear antigen (PCNA) revealed that VEGF-C induced mainly lymphangiogenesis in contrast to VEGF, which induced only angiogenesis. These results have significant implications in the planning of gene therapy using these growth factors.     

Structural and mechanistic insights into VEGF receptor 3 ligand binding and activation

Vascular endothelial growth factors (VEGFs) and their receptors (VEGFRs) are key drivers of blood and lymph vessel formation in development, but also in several pathological processes. VEGF-C signaling through VEGFR-3 promotes lymphangiogenesis, which is a clinically relevant target for treating lymphatic insufficiency and for blocking tumor angiogenesis and metastasis. The extracellular domain of VEGFRs consists of seven Ig homology domains; domains 1–3 (D1-3) are responsible for ligand binding, and the membrane-proximal domains 4–7 (D4-7) are involved in structural rearrangements essential for receptor dimerization and activation. Here we analyzed the crystal structures of VEGF-C in complex with VEGFR-3 domains D1-2 and of the VEGFR-3 D4-5 homodimer. The structures revealed a conserved ligand-binding interface in D2 and a unique mechanism for VEGFR dimerization and activation, with homotypic interactions in D5. Mutation of the conserved residues mediating the D5 interaction (Thr446 and Lys516) and the D7 interaction (Arg737) compromised VEGF-C induced VEGFR-3 activation. A thermodynamic analysis of VEGFR-3 deletion mutants showed that D3, D4-5, and D6-7 all contribute to ligand binding. A structural model of the VEGF-C/VEGFR-3 D1-7 complex derived from small-angle X-ray scattering data is consistent with the homotypic interactions in D5 and D7. Taken together, our data show that ligand-dependent homotypic interactions in D5 and D7 are essential for VEGFR activation, opening promising possibilities for the design of VEGFR-specific drugs.     

Involvement of VEGFR-2 (kdr/flk-1) but not VEGFR-1 (flt-1) in VEGF-A and VEGF-C-induced tube formation by human microvascular endothelial cells in fibrin matrices in vitro

Different forms of vascular endothelial growth factor (VEGF) and their cellular receptors (VEGFR) are associated with angiogenesis, as demonstrated by the lethality of VEGF-A, VEGFR-1 or VEGFR-2 knockout mice. Here we have used an in vitro angiogenesis model, consisting of human microvascular endothelial cells (hMVEC) cultured on three-dimensional (3D) fibrin matrices to investigate the roles of VEGFR-1 and VEGFR-2 in the process of VEGF-A and VEGF-C-induced tube formation. Soluble VEGFR-1 completely inhibited the tube formation induced by the combination of VEGF-A and TNFα (VEGF-A/TNFα). This inhibition was not observed when tube formation was induced by VEGF-C/TNFα or bFGF/TNFα. Blocking monoclonal antibodies specific for VEGFR-2, but not antibodies specifically blocking VEGFR-1, were able to inhibit the VEGF-A/TNFα-induced as well as the VEGF-C/TNFα-induced tube formation in vitro. PlGF-2, which interacts only with VEGFR-1, neither induced tube formation in combination with TNFα, nor inhibited or stimulated by itself the VEGF-A/TNFα-induced tube formation in vitro. These data indicate that VEGF-A or VEGF-C activation of the VEGFR-2, and not of VEGFR-1, is involved in the formation of capillary-like tubular structures of hMVEC in 3D fibrin matrices used as a model of repair-associated or pathological angiogenesis in vitro. This revised version was published online in June 2006 with corrections to the Cover Date.     

Involvement of the vascular endothelial growth factor receptor-1 in murine hepatocellular carcinoma development

BACKGROUND/AIMS: The role of the vascular endothelial growth factor receptor-1 (VEGFR-1) in hepatocellular carcinoma (HCC) development has not been elucidated yet. The aim of this study was to examine the role of VEGFR-1 in VEGF-mediated HCC development and angiogenesis as compared to that of VEGFR-2. METHODS: We examined the effects of VEGFR-1, and VEGFR-2 neutralizing monoclonal antibodies (R-1mAb and R-2mAb, respectively) on VEGF-mediated HCC development both in an allograft and orthotopic models. RESULTS: In the allograft model, both R-1mAb and R-2mAb significantly attenuated the VEGF-mediated tumor development in a dose dependent manner with associated reduction of angiogenesis in the tumor. The inhibitory effect of R-2mAb was more potent than that of R-1mAb, and the combination treatment with both mAbs almost completely attenuated VEGF-mediated HCC development. Immunohistochemical analysis revealed that apoptosis increased markedly in the tumor. Furthermore, these inhibitory effects with both mAbs were achieved even on established tumors and orthotopic transplantation. CONCLUSIONS: In addition to VEGFR-2, VEGFR-1 also lies on the signal transduction pathway by which VEGF augments HCC development and angiogenesis not only at the initial stage but also in the established tumor.     

Tumor cell-derived placental growth factor sensitizes antiangiogenic and antitumor effects of anti-VEGF drugs

The role of placental growth factor (PlGF) in modulation of tumor angiogenesis and tumor growth remains an enigma. Furthermore, anti-PlGF therapy in tumor angiogenesis and tumor growth remains controversial in preclinical tumor models. Here we show that in both human and mouse tumors, PlGF induced the formation of dilated and normalized vascular networks that were hypersensitive to anti-VEGF and anti–VEGFR-2 therapy, leading to dormancy of a substantial number of avascular tumors. Loss-of-function using plgf shRNA in a human choriocarcinoma significantly accelerated tumor growth rates and acquired resistance to anti-VEGF drugs, whereas gain-of-function of PlGF in a mouse tumor increased anti-VEGF sensitivity. Further, we show that VEGFR-2 and VEGFR-1 blocking antibodies displayed opposing effects on tumor angiogenesis. VEGFR-1 blockade and genetic deletion of the tyrosine kinase domain of VEGFR-1 resulted in enhanced tumor angiogenesis. These findings demonstrate that tumor-derived PlGF negatively modulates tumor angiogenesis and tumor growth and may potentially serve as a predictive marker of anti-VEGF cancer therapy.     

Vascular endothelial growth factor receptor-1 (VEGFR-1/Flt-1): a dual regulator for angiogenesis

Vascular endothelial growth factor receptor-1 (VEGFR-1) is a member of the VEGFR family, and binds VEGF-A, PlGF, and VEGF-B. An important feature of VEGFR-1 is that, unlike other VEGFR genes, it expresses two types of mRNA, one for a full-length receptor and another for a soluble short protein known as soluble VEGFR-1 (sFlt-1). The binding-affinity of VEGFR-1 for VEGF-A is one order of magnitude higher than that of VEGFR-2, whereas the kinase activity of VEGFR-1 is about 10-fold weaker than that of VEGFR-2. Through its ligand-binding region and by trapping ligands, VEGFR-1 plays a negative role in angiogenesis at embryogenesis. In adulthood, however, VEGFR-1 is expressed not only on endothelial cells but also on macrophages, and promotes the function of macrophages, inflammatory diseases, cancer metastasis, and atherosclerosis via its kinase activity. Soluble VEGFR-1 is abnormally overexpressed in the placenta of preeclamptic patients, and suggested to cause the major pathological symptoms on the maternal side such as hypertension and renal dysfunction, most likely by blocking the physiological VEGF-A. VEGFR-1 including its soluble form is involved in a variety of human illnesses, making it an important target in the development of new strategies to suppress disease.     

Expression of a functional VEGFR-1 in tumor cells is a major determinant of anti-PlGF antibodies efficacy

PlGF, one of the ligands for VEGFR-1, has been implicated in tumor angiogenesis. However, more recent studies indicate that genetic or pharmacological inhibition of PlGF signaling does not result in reduction of microvascular density in a variety of tumor models. Here we screened 12 human tumor cell lines and identified 3 that are growth inhibited by anti-PlGF antibodies in vivo. We found that efficacy of anti-PlGF treatment strongly correlates with VEGFR-1 expression in tumor cells, but not with antiangiogenesis. In addition, PlGF induced VEGFR-1 signaling and biological responses in tumor cell lines sensitive to anti-PlGF, but not in refractory tumor cell lines or in endothelial cells. Also, genetic ablation of VEGFR-1 signaling in the host did not affect the efficacy of PlGF blockade. Collectively, these findings suggest that the role of PlGF in tumorigenesis largely consists of promoting autocrine/paracrine growth of tumor cells expressing a functional VEGFR-1 rather than stimulation of angiogenesis.     

Structure and function of placental growth factor

Placental growth factor (PlGF) belongs to the same family as the vascular endothelial growth factor A (VEGF-A). Recent gene inactivation studies in mice have demonstrated that loss of PlGF does not affect development, reproduction, or normal postnatal life. However, the mice show significantly impaired angiogenesis and arteriogenesis during pathological conditions such as ischemia and tumor formation, conditions in which the expression of VEGF-A is normally increased. Mice expressing a truncated form of the specific receptor for PlGF, the VEGF receptor 1 (VEGFR-1), show impaired angiogenesis similar to that observed in Plgf(-/-)mice. These data suggest a pivotal role for PlGF and VEGFR-1 in regulating VEGF-A-dependent angiogenesis under pathological conditions. VEGF-A has been utilized for the therapeutic stimulation of new blood vessels in ischemic hearts and limbs, with controversial results from the initial clinical experience. This review discusses the possibility of using the PlGF/VEGFR-1 pathway as an alternative target for angiogenic therapy.