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PubMed2005

Angiogenesis in malignant lymphoma.

PURPOSE OF REVIEW: Angiogenesis plays an important role in the pathophysiology of both solid tumors and hematologic malignancies. Angiogenesis-associated parameters are important prognosticators, and tumor blood vessels are an emerging target for therapy. This review addresses the evidence of the role of angiogenesis in malignant lymphoma and discusses some therapeutic implications. RECENT FINDINGS: In angiogenesis assays, lymphoma cells show angiogenic properties. Tumor vascularization is higher in lymphoma tissue than in reactive lymph nodes and increases in step with clinically more aggressive lymphoma subtypes and advanced-stage disease. High levels of vascular endothelial growth factor in blood and tissue are associated with adverse prognosis. Vascular endothelial growth factor and vascular endothelial growth factor receptors are also present in lymphoma cells. Therapy against vascular endothelial growth factor in animal models is effective and points to both the tumor cell and the host endothelium as targets. Structural microvessel abnormalities are present in some lymphoma subtypes. The role of angiogenesis might vary in lymphoma subtypes because the prognostic value of microvessel density and the expression of angiogenesis-related molecules differ between lymphoma subtypes. There are also differences in blood vessel phenotype between lymphoma subtypes. This heterogeneity may have implications for antiangiogenic therapies. Antiangiogenic therapy in human lymphoma is still in its infancy. SUMMARY: The role of angiogenesis in malignant lymphoma is evident. Tumor vasculature is an attractive target for lymphoma therapy. Differences between lymphoma subtypes must be taken into account in the selection of the most suitable patients for trials with antiangiogenic agents.

Angiogenesis Inhibitors↗
PubMed2003

Temporal profile of angiogenesis and expression of related genes in the brain after ischemia.

Angiogenesis is an intricately regulated phenomenon. Its mechanisms in the ischemic brain have not been clearly elucidated. The authors investigated expression of angiogenesis-related genes using a complementary DNA (cDNA) array method as well as Western blotting and immunohistochemistry, and compared these studies with a temporal profile of angiogenesis in mouse brains after ischemia. The number of vessels significantly increased 3 days after injury, and proliferating endothelial cells increased as early as 1 day. This means that angiogenesis occurs immediately after the injury. Ninety-six genes implicated in angiogenesis were investigated with a cDNA array study. It was found that 42, 29, and 13 genes were increased at 1 hour, 1 day, and 21 days, respectively. Most of the well-known angiogenic factors increased as early as 1 hour. Vessel-stabilizing factors such as thrombospondins also increased. At 1 day, however, thrombospondins decreased to lower levels than in the control, indicating a shift from vascular protection to angiogenesis. At 21 days, many genes were decreased, but some involved in tissue repair were newly increased. Western blotting and immunohistochemistry showed findings compatible with the cDNA array study. Many molecules act in an orchestrated fashion in the brain after ischemia and should be taken into account for therapeutic angiogenesis for stroke.

Angiogenesis Inducing Agents↗
PubMed2005

Imiquimod is a strong inhibitor of tumor cell-induced angiogenesis.

BACKGROUND: Imiquimod, a potent immunomodulator, not having a direct antiproliferative activity, was found to be effective in genital and cutaneous premalignancies and malignancies. As tumor development depends on blood vessel supply, the inhibition of angiogenesis could be responsible for the antitumor activity. OBJECTIVE: To find in a murine model whether imiquimod has antiangiogenic activity and whether this activity is mediated by locally induced cytokines. METHODS: The study was performed in two cell lines: Skv human keratinocytes containing multiple integrated copies of HPV16 derived from bowenoid papulosis, and murine L1 lung sarcoma cells of Balb/c mice. The murine model of cutaneous angiogenesis was used to assess and count the new blood vessel formation. The mice were immunosuppressed by a total body X-ray irradiation and treated with 5% or 2.5% imiquimod cream before or after induction of angiogenesis with intradermally injected tumor cell suspension. In some experiments the mice were, in addition, treated intraperitoneally with monoclonal antibodies against murine IFNalpha, TNFgamma or IL-18. RESULTS: Topical application of imiquimod on the murine skin resulted in reduction of angiogenesis (P < 0.001) induced by intradermal injection of both human and mouse tumor cells, more pronounced when 5% cream was applied on three consecutive days. Antibodies against murine IFNgamma, TNFalpha and IL-18 completely abolished the inhibitory effect of imiquimod on angiogenesis induced by murine L1 sacroma cells. When human Skv cells were used in angiogenesis assay, the effect of imiquimod was abolished by antibodies against IL-18 but not against TNFalpha, which may be due to overproduction of TNFalpha by Skv cells. CONCLUSIONS: Antiangiogenic effect of imiquimod was found to be mediated by IL-18, probably through promoting production of INFgamma, the most important inhibitor of angiogenesis.

Adjuvants, Immunologic↗
PubMed1983

Quantitation of angiogenesis factor in bovine retina and tumour extracts by means of radioimmunoassay.

Using an antiserum raised against tumour angiogenesis factor (TAF) we have developed a radioimmunoassay for retina and tumour angiogenesis factor(s). This antiserum was previously shown to bind to both human and animal tumour extracts and to inhibit the angiogenesis induced by TAF in vivo. TAF from rat Walker tumour was used for iodination by the chloramine-T method. An excess of 125I-labelled TAF was incubated with TAF antibody in the absence (maximum binding) and presence (inhibition of maximum binding) of unlabelled tissue extract. A double antibody technique was used to separate free and bound TAF. Unlabelled human Wilms tumour TAF was used as a standard. The extent of inhibition of 125I-TAF-anti-TAF binding provided a measure of TAF in tissue extracts examined. Extracts of normal bovine retina, cornea, lung, aorta, lymph nodes, iris, vitreous humour, and human tumours and normal human pituitary and liver were assayed. Only bovine retina and human tumours were found to contain angiogenesis factor. These findings, together with our earlier results, suggest that angiogenesis factor from both bovine retina and human tumours induce angiogenesis in vivo and possess common antigenic determinants. The presence of angiogenesis factor in healthy retina and its relationship to neovascularisation in clinical conditions is discussed.

Angiogenesis Inducing Agents↗
PubMed2006

NAD(P)H oxidase modulates angiogenesis and the development of portosystemic collaterals and splanchnic hyperaemia in portal hypertensive rats.

BACKGROUND: Recent studies have shown the presence of vascular endothelial growth factor (VEGF)-dependent splanchnic angiogenesis in experimental models of portal hypertension, and the role of such neovascularisation on the development of both portosystemic collaterals and hyperdynamic splanchnic circulation. However, the mechanisms modulating angiogenesis in portal hypertension are unknown. Experimental evidence indicates that NAD(P)H oxidase is required for VEGF-induced angiogenesis. Interestingly, we have recently shown that splanchnic NAD(P)H oxidase activity is significantly increased in portal hypertensive rats. Therefore, it could be possible that activated NAD(P)H oxidases modulate angiogenesis in portal hypertension. AIM: To determine the effects of chronic NAD(P)H oxidase inhibition on angiogenesis and splanchnic haemodynamics in portal hypertensive rats. METHODS: Partial portal vein-ligated and sham-operated rats were treated with the NAD(P)H oxidase inhibitor apocynin, or with vehicle for 5 days. Then, the expression of angiogenesis markers (western blotting), the formation of portosystemic collaterals (radioactive microspheres) and the production of superoxide anion (lucigenin-enhanced chemiluminescence) were determined. Mean arterial pressure, portal pressure, and superior mesenteric arterial blood flow and resistance were also measured. RESULTS: In portal hypertensive rats, NAD(P)H oxidase blockade significantly decreased portosystemic collateral formation, and superior mesenteric arterial flow. It also reduced the splanchnic expression of VEGF, VEGF receptor-2 and CD31, and attenuated the increased production of superoxide, compared with vehicle. CONCLUSIONS: NAD(P)H oxidase plays an important role in experimental portal hypertension, modulating splanchnic angiogenesis, the formation of portosystemic collaterals and the development of splanchnic hyperdynamic circulation. These results suggest that NAD(P)H oxidase may represent a new target in the treatment of portal hypertension.

Acetophenones↗
PubMed2005

Angiogenesis: a curse or cure?

Angiogenesis, the growth of new blood vessels is essential during fetal development, female reproductive cycle, and tissue repair. In contrast, uncontrolled angiogenesis promotes the neoplastic disease and retinopathies, while inadequate angiogenesis can lead to coronary artery disease. A balance between pro-angiogenic and antiangiogenic growth factors and cytokines tightly controls angiogenesis. Considerable progress has been made in identifying these molecular components to develop angiogenesis based treatments. One of the most specific and critical regulators of angiogenesis is vascular endothelial growth factor (VEGF), which regulates endothelial proliferation, permeability, and survival. Several VEGF based treatments including anti-VEGF and anti-VEGF receptor antibodies/agents are in clinical trials along with several other antiangiogenic treatments. While bevacizumab (anti-VEGF antibody) has been approved for clinical use in colorectal cancer, the side effects of antiangiogenic treatment still remain a challenge. The pros and cons of angiogenesis based treatment are discussed.

Angiogenesis Inhibitors↗
PubMed2005

Endogenous inhibitors of angiogenesis.

Angiogenesis, the formation of new blood vessels, is required for many pathologic processes, including invasive tumor growth as well as physiologic organ/tissue maintenance. Angiogenesis during development and adulthood is likely regulated by a balance between endogenous proangiogenic and antiangiogenic factors. It is speculated that tumor growth requires disruption of such balance; thus, the angiogenic switch must be turned "on" for cancer progression. If the angiogenic switch needs to be turned on to facilitate the tumor growth, the question remains as to what the physiologic status of this switch is in the adult human body; is it "off," with inhibitors outweighing the stimulators, or maintained at a fine "balance," keeping the proangiogenic properties of many factors at a delicate "activity" balance with endogenous inhibitors of angiogenesis. The physiologic status of this balance is important to understand as it might determine an individual's predisposition to turn the switch on during pathologic events dependent on angiogenesis. Conceivably, if the physiologic angiogenesis balance in human population exists somewhere between off and even balance, an individual's capacity and rate to turn the switch on might reflect their normal physiologic angiogenic status. In this regard, although extensive knowledge has been gained in our understanding of endogenous growth factors that stimulate angiogenesis, the activities associated with endogenous inhibitors are poorly understood. In this review, we will present an overview of the knowledge gained in studies related to the identification and characterization of 27 different endogenous inhibitors of angiogenesis.

Angiogenesis Inhibitors↗
PubMed2006

The vasohibin family: a negative regulatory system of angiogenesis genetically programmed in endothelial cells.

Biological phenomena are under the precise control by the genome. For the regulation of angiogenesis, proangiogenic genes such as VEGFs and angiopoietins are highly conserved, act specifically on endothelial cells, and play a fundamental role. In this sense, nature should prepare specific antiangiogenic genes as well. However, this counterpart of genomic regulation of angiogenesis remains to be established. We recently isolated a novel endothelium-derived angiogenesis inhibitor and named it vasohibin. Vasohibin is dominantly expressed in endothelial cells, induced by the stimulation with VEGF or FGF-2, and selectively affects on endothelial cells and inhibits angiogenesis. Although the mechanism of how vasohibin inhibits angiogenesis remains to be elucidated, our discovery of vasohibin as an endothelium-derived VEGF-inducible angiogenesis inhibitor should shed light on the genomic basis of the negative regulation of angiogenesis.

Alternative Splicing↗
PubMed2002

Inhibition of endothelial cell survival and angiogenesis by protein kinase A.

Receptors for the provisional ECM are important regulators of angiogenesis. One of these receptors, integrin alpha5beta1, plays a critical role in tumor- and growth factor-induced angiogenesis, because antagonists of this integrin potently inhibit angiogenesis and tumor growth. Here we show that the integrin alpha5beta1 promotes endothelial cell survival during angiogenesis in vivo by suppressing the activity of protein kinase A (PKA). Antagonists of integrin alpha5beta1 activate PKA, which then leads to the activation of caspase-8 and induction of apoptosis. Direct activation of PKA by cAMP or by expression of the PKA catalytic subunit also induces endothelial cell apoptosis, resulting in angiogenesis inhibition in vivo. Our studies indicate that ligation of integrin alpha5beta1 during angiogenesis suppresses an apoptotic program that is dependent on PKA. These studies also indicate that induction of endothelial cell apoptosis in vivo by genetic or pharmacological activation of PKA may be a useful strategy to inhibit angiogenesis.

Angiogenesis Inhibitors↗
PubMedDate not supplied

Clinical experience with angiogenesis signaling inhibitors: focus on vascular endothelial growth factor (VEGF) blockers.

Angiogenesis is required for tumor growth and metastasis and, therefore, represents an exciting target for cancer treatment. Angiogenesis is a complex process that is tightly regulated by pro- and anti-angiogenic growth factors. Physiologic angiogenesis takes place during tissue growth and repair, during the female reproductive cycle, and during fetal development. Pathologic angiogenesis is characterized by either excessive (eg, cancer) or inadequate (eg, coronary artery disease) neovascularization. Angiogenesis occurs in a series of complex and interrelated steps that involve the release of pro-angiogenic growth factors, such as vascular endothelial growth factor (VEGF). VEGF regulates both vascular proliferation and permeability, and functions as an anti-apoptotic factor for newly formed blood vessels. The biological effects of VEGF are mediated by two receptors, VEGF-1 and VEGF-2, whose expression is largely limited to the vascular endothelium. VEGF is often expressed in tumors at substantially increased levels. It is expressed in response to hypoxia, oncogenes, and other cytokines, and its expression is associated with poor prognosis in several types of cancer. Several different strategies have been used to inhibit VEGF, including anti-VEGF monoclonal antibodies (eg, bevacizumab) and agents that inhibit the VEGF receptor (eg, SU5416). Both types of agents have tolerable side effects and have shown promise when evaluated in a wide range of tumor types. Angiogenesis, the role of VEGF in angiogenesis and malignancy, and strategies for cancer treatment with VEGF inhibitors are discussed.

Angiogenesis Inhibitors↗
PubMed2004

Therapeutic angiogenesis for myocardial ischemia.

Therapeutic angiogenesis offers promise as a novel treatment for ischemic heart disease, particularly for patients who are not candidates for current methods of revascularization. The goal of treatment is both relief of symptoms of coronary artery disease and improvement of cardiac function by increasing perfusion to the ischemic region. Protein-based therapy with cytokines including vascular endothelial growth factor and fibroblast growth factor demonstrated functionally significant angiogenesis in several animal models. However, clinical trials have yielded largely disappointing results. The attenuated angiogenic response seen in clinical trials of patients with coronary artery disease may be due to multiple factors including endothelial dysfunction, particularly in the context of advanced atherosclerotic disease and associated comorbid conditions, regimens of single agents, as well as inefficiencies of current delivery methods. Gene therapy has several advantages over protein therapy and recent advances in gene transfer techniques have improved the feasibility of this approach. The safety and tolerability of therapeutic angiogenesis by gene transfer has been demonstrated in phase I clinical trials. The utility of therapeutic angiogenesis by gene transfer as a treatment option for ischemic cardiovascular disease will be determined by adequately powered, randomized, placebo-controlled Phase II and III clinical trials. Cell-based therapies offer yet another approach to therapeutic angiogenesis. Although it is a promising therapeutic strategy, additional preclinical studies are warranted to determine the optimal cell type to be administered, as well as the optimal delivery method. It is likely the optimal treatment will involve multiple agents as angiogenesis is a complex process involving a large cascade of cytokines, as well as cells and extracellular matrix, and administration of a single factor may be insufficient. The promise of therapeutic angiogenesis as a novel treatment for no-option patients should be approached with cautious optimism as the field progresses.

Angiogenesis Inducing Agents↗
PubMed2006

Cancer gene therapy targeting angiogenesis: an updated review.

Since the relationship between angiogenesis and tumor growth was established by Folkman in 1971, scientists have made efforts exploring the possibilities in treating cancer by targeting angiogenesis. Inhibition of angiogenesis growth factors and administration of angiogenesis inhibitors are the basics of anti-angiogenesis therapy. Transfer of anti-angiogenesis genes has received attention recently not only because of the advancement of recombinant vectors, but also because of the localized and sustained expression of therapeutic gene product inside the tumor after gene transfer. This review provides the up-to-date information about the strategies and the vectors studied in the field of anti-angiogenesis cancer gene therapy.

Angiogenesis Inhibitors↗
PubMed2000

Angiogenesis: potentials for pharmacologic intervention in the treatment of cancer, cardiovascular diseases, and chronic inflammation.

Angiogenesis, or the formation of new blood vessels out of pre-existing capillaries, is a sequence of events that is fundamental to many physiologic and pathologic processes such as cancer, ischemic diseases, and chronic inflammation. With the identification of several proangiogenic molecules such as the vascular endothelial cell growth factor, the fibroblast growth factors (like in FGFs), and the angiopoietins, and the recent description of specific inhibitors of angiogenesis such as platelet factor-4, angiostatin, endostatin, and vasostatin, it is recognized that therapeutic interference with vasculature formation offers a tool for clinical applications in various pathologies. Whereas inhibition of angiogenesis can prevent diseases with excessive vessel growth such as cancer, diabetes retinopathy, and arthritis, stimulation of angiogenesis would be beneficial in the treatment of diseases such as coronary artery disease and critical limb ischemia in diabetes. In this review we highlight the current knowledge on angiogenesis regulation and report on the recent findings in angiogenesis research and clinical studies. We also discuss the potentials, limitations, and challenges within this field of research, in light of the development of new therapeutic strategies for diseases in which angiogenesis plays an important role.

Angiogenesis Inhibitors↗
PubMed2000

The N-myc oncogene in human neuroblastoma cells: down-regulation of an angiogenesis inhibitor identified as activin A.

Angiogenesis, the formation of new blood vessels, is seen during embryonic development and tumor progression, but the mechanisms have remained unclear. Recent data indicate that developmental and tumor angiogenesis can be induced by cellular oncogenes, leading to the enhanced activity of molecules stimulating angiogenesis. However, activated oncogenes might also facilitate angiogenesis by down-regulating endogenous inhibitors of angiogenesis. We report here that enhanced expression of the N-myc oncogene in human neuroblastoma cells down-regulates an inhibitor of endothelial cell proliferation, identified by amino acid sequencing as being identical with activin A, a developmentally regulated protein. Down-regulation appears to involve interaction of the N-Myc protein with the activin A promoter. In addition, activin A inhibits both endothelial cell proliferation in vitro and angiogenesis in vivo, and it induces hemorrhage in vivo. We suggest that the N-myc-induced down-regulation of activin A could contribute to developmental and tumor angiogenesis.

Activins↗
PubMed2001

Pharmacodynamic-mediated reduction of temozolomide tumor concentrations by the angiogenesis inhibitor TNP-470.

The angiogenic phenotype is associated with increased tumor neovascularization and a state of vascular hyperpermeability to macromolecules. Angiogenesis inhibitors could reverse these processes, resulting in tumor capillaries that have normal membrane permeability. It was proposed that the switch from a hyperpermeable to a normal permeable state could have the untoward effect of decreasing tumor concentrations of anticancer drugs coadministered with angiogenesis inhibitors. The current investigation evaluated a potential drug interaction between the angiogenesis inhibitor O-(N-chloroacetyl-carbamoyl)-fumagillol (TNP-470) and the alkylating agent temozolomide (TMZ), in xenograft models that differentially expressed vascular endothelial growth factor (VEGF), a driving force for angiogenesis. Nude rats bearing either s.c. low VEGF (V-) or high VEGF (V+) or intracerebral V+ gliomas were administered either a multiple-dose regimen of TNP-470 or vehicle control. One day after the last dose of vehicle or TNP-470, a steady-state dosing regimen of TMZ was administered with subsequent collection and high-performance liquid chromatography analysis of plasma and either tumor homogenate or tumor microdialysis steady-state TMZ concentrations, and in some cases [5-(3-methyltriazen-1-yl)imidazole-4-carboximide] MTIC, its active metabolite. Microvessel density (MVD) was quantitated by image analysis using an anti-CD31 method. Statistical analyses of pharmacokinetic and pharmacodynamic end points in the control and TNP-470 treatment groups were completed by nonparametric tests. In both the s.c. and intracerebral V+ models, TNP-470 treatment produced significant reductions in TMZ tumor concentrations and tumor:plasma concentration ratios compared with control, being reduced an average of 25% and 50% in the s.c. and intracerebral tumors, respectively. MTIC concentrations in V+ s.c. tumors also were reduced by 50% in the presence of TNP-470. Consistent with the lower extent of neovascularization in the V- tumors, TMZ and MTIC tumor concentrations were not different in TNP-470 and control treatment groups in s.c. tumors. MVD was reduced by TNP-470 compared with vehicle control in the V+ tumors, but was unaltered in V- tumors, attesting to the use of MVD as a pharmacodynamic end point and the effectiveness of TNP-470 as an angiogenesis inhibitor. Angiogenesis inhibitor's pharmacodynamic actions on tumor angiogenesis can produce a reduction in tumor concentrations of coadministered anticancer agents. It is increasingly important to understand the pharmacokinetic and pharmacodynamic behavior of each class of drug so that optimal dosing regimens can be designed.

Angiogenesis Inhibitors↗
PubMedDate not supplied

A quantitative angiogenesis model for efficacy testing of chemopreventive agents.

One of the approaches in chemoprevention to prevent or delay the progression of precancerous lesions, is to apply chemopreventive agents that can potentially block angiogenesis. A quantitative in vivo angiogenesis inhibition assay was developed to test the efficacy of twelve chemopreventive agents that represent different chemical classes and multiple biological activities, using the chick chorioallantoic membrane (CAM) model and an oncogene-transfected angiogenic cell line (6 Ti ras/SV myc # 4). These tumorigenic cells held by a primary agarose pellet, were placed alone or with a secondary pellet incorporating five concentrations of the test agent, on an exposed CAM of 7-day-old chick embryo for 72 hours in a humidified chamber at 35 degrees C. The cell-induced angiogenic blood vessels, including the microvessels radiating from the cell pellet focal area, were scored using a computerized custom image analysis system. The results show that nonsteroidal antiinflammatory drugs (NSAIDS); aspirin, sulindac, sulindac sulfide and sulindac sulfone, were effective inhibitors of cell-induced angiogenesis (23-66%). Aspirin displayed a dose-dependent response with the highest inhibition at 300 microM and an EC50 (the effective molar concentration that inhibits angiogenesis by 50%) of 26 microM. Sulindac sulfone was more effective than sulindac with an EC50 of 5 microM versus 85 microM. However, sulindac sulfide showed an intermediate response with an EC50 of 41 microM. The retinoids; all-trans-retinoic acid (ATRA), 9-cis-retinoic acid (9-cis-RA), and 13-cis-retinoic acid (13-cis-RA) were also highly effective inhibitors of cell-mediated CAM-angiogenesis. 13-cis-RA with an EC50 of 3.6 nM, has been the most efficacious test agent. > 400-fold more effective than 9-cis-RA (1.5 microM). ATRA exhibited an intermediate response between 9-cis-RA and 13-cis-RA with an EC50 of 0.3 microM, and was 100-fold more efficacious than 9-cis-RA. However, the synthetic retinoid, N-(4-hydroxyphenyl) retinamide (4-HPR), was not an effective inhibitor of CAM angiogenesis. Thalidomide, a compound with multiple biological activities, exhibited dose-dependent inhibition ranging from 10-1000 microM with an EC50 of 19 microM. Other agents that exhibited dose-dependent inhibition included Bowman-Birk inhibitor (BBI), EC50: 10 microg/ml, tamoxifen, EC50, 0.05 microM and difluoromethyl omithine (DFMO), with an EC50 of 13 microM. These results suggest that tumor-associated angiogenesis can be modulated by non-toxic concentrations of chemopreventive agents representing multiple biological activities and multiple targets.

Angiogenesis Inhibitors↗
PubMed2003

Angiopoietin-1 inhibits vascular permeability, angiogenesis, and growth of hepatic colon cancer tumors.

Angiopoietin (Ang)-1 and -2 are critical regulators of embryonic and postnatal neovascularization. Ang-1 activates the endothelial cell-specific tyrosine kinase receptor Tie-2, which in turn leads to enhanced endothelial cell survival and stabilization. The effects of Ang-1 on tumor angiogenesis remain controversial; although we have previously demonstrated that Ang-1 overexpression in colon cancer cells leads to a decrease in s.c. tumor growth, others have shown that Ang-1 may be proangiogenic. Few studies have addressed the role of the Angs in tumors growing in the organ of metastatic growth. We hypothesized that overexpression of Ang-1 may inhibit the growth of colon cancers growing in the liver by inhibition of angiogenesis. We also wanted to investigate the mechanisms by which Ang-1 affects angiogenesis in vivo. Human colon cancer cells (HT29) were stably transfected with an Ang-1 construct or an empty vector (pcDNA) and injected directly into the livers of nude mice. After 37 days, livers were harvested and weighed, and tumor sizes were measured. In an additional experiment, to validate the paracrine effect of Ang-1, various mixtures of control cells and Ang-1-transfected cells were injected into livers, and tumor growth was assessed. Direct effects of recombinant Ang-1 on angiogenesis were studied with an in vivo Gelfoam angiogenesis assay. The impact of Ang-1 on vascular permeability was investigated using an intradermal Miles assay with conditioned media from transfected cells. Liver weights (P < 0.05), tumor volumes (P < 0.05), vessel counts (P < 0.01), and tumor cell proliferation (P < 0.01) in the Ang-1 group were significantly lower than those in the control (pcDNA) group. Tumor vessels in the Ang-1 group developed a significantly higher degree of pericyte coverage (P < 0.02) than vessels in pcDNA tumors. In the cell mixture experiment, even as few as a 1:10 mixture of Ang-1-transfected cells/control cells resulted in a significant reduction of hepatic tumor volumes (P < 0.04). In the angiogenesis assay, vessel counts in Gelfoam implants were significantly decreased by the addition of Ang-1 (P < 0.01). Finally, conditioned medium from Ang-1-transfected cells decreased vascular permeability more than that from control cells (P < 0.05). Our results suggest that Ang-1 is an important regulator of angiogenesis and vascular permeability and that this effect may be secondary to increasing periendothelial support and vessel stabilization. Thus, Ang-1 could potentially serve as an antineoplastic or anti-permeability agent for patients with metastatic colorectal cancer.

Angiogenesis Inducing Agents↗
PubMed2001

Differential inhibition of tumor angiogenesis by tie2 and vascular endothelial growth factor receptor-2 dominant-negative receptor mutants.

Tumor growth is angiogenesis-dependent. Current evidence suggests that vascular endothelial growth factor (VEGF), a major regulator of embryonic and hypoxia-mediated angiogenesis, is necessary for tumor angiogenesis. VEGF is expressed in tumor cells in vivo, and its tyrosine kinase receptors VEGFR-1 and VEGFR-2 are up-regulated in the tumor endothelium. A second endothelial cell-specific ligand/receptor tyrosine kinase system, consisting of the tie2 receptor, its activating ligand angiopoietin-1 and the inhibitory ligand angiopoietin-2, has been characterized. We have examined 6 human primary breast-cancer samples and 4 murine breast-cancer cell lines (M6363, M6378, M6444, M6468), transplanted into nude mice, by in situ hybridization and/or Northern analysis. Expression of angiopoietin-1, angiopoietin-2 and tie2 was compared to VEGF and VEGFR-2 expression. Human tumors expressed VEGFR-2 and tie2 but varied considerably in VEGF and angiopoietin-1/-2 expression. In the murine tumor models, we observed high heterogeneity of receptor and ligand expression. M6363 and M6378 tumors were analyzed in detail because they showed different expression of components of the tie2/angiopoietin signaling system. M6363 tumors expressed VEGF, VEGFR-2 and angiopoietin-2 but not tie2 or angiopoietin-1, suggesting activation of VEGFR-2 and inhibition of tie2 signaling pathways, whereas M6378 tumors expressed VEGF, VEGFR-2, tie2 and angiopoietin-1 but little angiopoietin-2, suggesting activation of both VEGFR-2 and tie2 signaling pathways. In vivo studies using truncated dominant-negative tie2 and VEGFR-2 mutants revealed inhibition of M6363 tumor growth by 15% (truncated tie2) and 36% (truncated VEGFR-2), respectively. In contrast, M6378 tumor growth was inhibited by 57% (truncated tie2) and 47% (truncated VEGFR-2), respectively. These findings support the hypothesis that tumor angiogenesis is dependent on VEGFR-2 but suggest that, in addition, tie2-dependent pathways of tumor angiogenesis may exist. For adequate application of angiogenesis inhibitors in tumor patients, analysis of prevailing angiogenesis pathways may be a prerequisite.

Adenocarcinoma, Mucinous↗