Vascular Endothelial Growth factor-A Inhibitor Research Articles

Identification of Phytochemical Inhibitors Targeting VEGFA and TGFBR2 of Cervical Cancer: Insights from Virtual Screening, ADMET and DFT Studies

AbstractThe vascular endothelial growth factor A (VEGFA) and Transforming Growth Factor Beta type II receptor (TGFBR2) serves as a promising candidate in the treatment of cervical cancer, playing a crucial role in angiogenesis and tumorigenesis. Its involvement in cervical cancer has been well documented. The pursuit of phytochemical inhibitors to counteract the cancer promoting effects of VEGFA and TGFBR2 has gained considerable momentum as very few drugs are available. In this study, we employed computational techniques to identify potential VEGFA and TGFBR2 inhibitors from 101 phytochemicals. The Spirosolane, Withaferin A & Silybin B for VEGFA and Ginkgetin, Hesperedin for TGFBR2 were identified as promising candidates as indicated by their favorable docking scores and robust energy profiles. Furthermore, Absorption, Distribution, Metabolism, Excretion, and Toxicity (ADMET) profile indicating that Spirosolane and Ginkgetin exhibit favorable drug‐like properties and medicinal chemistry characteristics, suggesting their potential as novel drug candidates. The Density Functional Theory (DFT) studies revealed that these compounds exhibited the highest electrophilicity index for occupied molecular orbitals, basicity, and dipole moment, all contributing to their stability and strong binding affinity at the active site. To advance this line of research, additional experimental validation is warranted to facilitate the development of highly selective and effective VEGFA and TGFBR2 inhibitors.

Efficacy and Safety of Faricimab for Macular Edema due to Retinal Vein Occlusion: 24-Week Results from the BALATON and COMINO Trials

To evaluate the 24-week efficacy and safety of the dual angiopoietin-2 (Ang-2) and vascular endothelial growth factor (VEGF)-A inhibitor faricimab versus aflibercept in patients with vein occlusion. Phase 3, global, randomized, double-masked, active comparator-controlled trials: BALATON/COMINO (ClincalTrials.gov identifiers: NCT04740905/NCT04740931; sites: 149/192). Patients with treatment-naïve foveal center-involved macular edema resulting from branch (BALATON) or central or hemiretinal (COMINO) RVO. Patients were randomized 1:1 to faricimab 6.0 mg or aflibercept 2.0 mg every 4 weeks for 24weeks. Primary end point: change in best-corrected visual acuity (BCVA) from baseline to week 24. Efficacy analyses included patients in the intention-to-treat population. Safety analyses included patients who received ≥ 1 doses of study drug. Enrollment: BALATON, n = 553; COMINO, n = 729. The BCVA gains from the baseline to week 24 with faricimab were noninferior versus aflibercept in BALATON (adjusted mean change,+16.9 letters [95.03% confidence interval (CI), 15.7-18.1 letters] vs.+17.5 letters [95.03% CI, 16.3-18.6 letters]) and COMINO (+16.9 letters [95.03% CI, 15.4-18.3 letters] vs.+17.3 letters [95.03% CI, 15.9-18.8 letters]). Adjusted mean central subfield thickness reductions from the baseline were comparable for faricimab and aflibercept at week 24 in BALATON (-311.4 μm [95.03% CI,-316.4 to-306.4 μm] and-304.4 μm [95.03% CI,-309.3 to-299.4 μm]) and COMINO (-461.6 μm [95.03% CI,-471.4 to-451.9 μm] and-448.8 μm [95.03% CI,-458.6 to-439.0 μm]). A greater proportion of patients in the faricimab versus aflibercept arm achieved absence of fluorescein angiography-based macular leakage at week 24 in BALATON (33.6% vs. 21.0%; nominal P= 0.0023) and COMINO (44.4% vs. 30.0%; nominal P= 0.0002). Faricimab was well tolerated, with an acceptable safety profile comparable with aflibercept. The incidence of ocular adverse events was similar between patients receiving faricimab (16.3% [n= 45] and 23.0% [n= 84] in BALATON and COMINO, respectively) and aflibercept (20.4% [n= 56] and 27.7% [n= 100], respectively). These findings demonstrate the efficacy and safety of faricimab, a dual Ang-2/VEGF-A inhibitor, in patients with macular edema secondary to retinal vein occlusion. Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.

Analysis of the mechanism of Sophorae Flavescentis Radix in the treatment of intractable itching based on network pharmacology and molecular docking.

Sophorae Flavescentis Radix (Kuh-seng, SFR), a Traditional Chinese Medicine (TCM), is widely used alone or within a TCM formula to treat pruritus, especially histamine-independent intractable itching. In the previous study, potential antipruritic active components of the SFR were screened based on cell membrane immobilized chromatography (CMIC), revealing oxymatrine (OMT) as an antipruritic agent. However, the low oral bioavailability (OB) of OMT cannot explain the antipruritic effect of SFR when administered orally in clinic. In this study, we investigated the antipruritic effects and underlying mechanisms of orally administered SFR. A network pharmacology and molecular docking were employed to screen the active components of SFR and predict their binding to disease-related target proteins, while the potential mechanisms were explored with Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. The binding energy between components and target proteins was calculated by molecular docking. The SFR-components-targets-intractable itching Protein-Protein Interactions (PPI) network was established, and 22 active components and 42 targets were screened. The GO enrichment analysis showed that the key target genes of SFR were related to nuclear receptors, transcription factors, and steroid hormone receptors. The results of the KEGG enrichment pathway analysis include Hepatitis B, epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor resistance, advanced glycation end product (AGE)-receptor for AGE (RAGE) signaling pathway in diabetic complications, etc. Molecular docking showed that three key target proteins in the network, the vascular endothelial growth factor A (VEGFA), epidermal growth factor receptor (EGFR) and caspase-3 (CASP3), have higher binding activities with inermine, phaseolin and kushenol O, respectively; the binding energy of each pair is stronger than that of the target protein-corresponding inhibitors. The complexity of the SFR-components-targets-intractable itching network demonstrated the holistic treatment effect of SFR on intractable itching. The partial coherence between results screened by CMIC in the previous study and network pharmacology demonstrated the potential of network pharmacology in active component screening. Inermine screened from both CMIC and network pharmacology is a VEGFA inhibitor, which possibly accounts for the antipruritic effect of orally administered SFR.

Variation in VEGFA and risk of cardiovascular disease in the UK Biobank

BackgroundCardiovascular disease (CVD) is an escalating global health crisis, contributing significantly to worldwide mortality and morbidity. Dyslipidemia stands as a critical risk factor for CVD. Vascular endothelial growth factor A (VEGFA) is pivotal in angiogenesis and represents a clinical target for CVD intervention. However, the impact of genetic modulation of VEGFA on lipid levels and the subsequent risk of cardiovascular events remains unclear.MethodsWe used LDpred2 to calculate genetic scores for lipid levels based on VEGFA variation, serving as instrumental variables to simulate the effect of VEGFA inhibitors. We then assessed the associations between genetic risk for lipid levels and CVD risk by conducting One-sample Mendelian randomization.ResultsOur results indicated that low-density lipoprotein cholesterol [LDL-C; odds ratio (OR) = 1.09, 95% CI: 1.06–1.11], remnant cholesterol (RC; OR = 1.24, 95% CI: 1.13–1.36), and triglycerides (TG; OR = 1.14, 95% CI: 1.07–1.22) were positively associated with the incidence of CVD. In contrast, high-density lipoprotein cholesterol (HDL-C) was inversely associated with the incidence of CVD (OR = 0.80, 95% CI: 0.76–0.86). When considering the genetic score for LDL-C constructed based on VEGFA, the group with a high genetic score demonstrated an elevated CVD risk (OR = 1.11, 95% CI: 1.04–1.19) compared to those with a low genetic score. Notably, One-sample Mendelian randomization results provided evidence of a causal relationship between LDL-C and CVD (p = 8.4×10−3) when using genetic variation in VEGFA as an instrumental variable.ConclusionsGenetic variation mimicking the effect of VEGFA inhibition, which lowers LDL-C levels, was causally associated with a reduced risk of cardiovascular events. These findings offer insight into the potential therapeutic relevance of modulating VEGFA-mediated lipid changes in the prevention and management of CVD.

Discovery novel VEGFA inhibitors through structure-based virtual screening and verify the ability to inhibit the proliferation, invasion and migration of gastric cancer

This paper aims to screen small molecule inhibitors 1,2,3,4,6-penta-O-galloyl-beta-D-glucose (PGG) targeting vascular endothelial growth factor A (VEGFA) through structure based virtual screening and molecular dynamics simulation, and verify the effect of anti gastric cancer.First, Based on Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform, the candidate small-molecule compounds targeting VEGFA were screened by a molecular docking method using Computer-Aided Drug Design. Second, CCK8 was used to determine the effect of three commercially available candidate drugs on the proliferation activity of HGC27 and AGS. PGG was selected for further cell cloning, invasion, migration and apoptosis experiments. Finally, the complex system of three compounds and VEGFA was analyzed by molecular dynamics simulation.According to the ranking of the scoring function, the selected small molecular compounds are PGG, 2,3,4,6-tetra-O-galactosyl-D-glucopyranoside, rutin, quercetin-5,3-d-galactoside and 1F-Fructofuranosylnystose (1FF). CCK8 showed that PGG had the best inhibitory effect on the proliferation of AGS and HGC27 cells, and it was concentration and time dependent. Treatment of AGS and HGC27 with IC50 PGG can significantly inhibit the cloning of HGC27 and AGS, block their invasion and migration, and induce their apoptosis. Molecular dynamics simulation experiments showed that the binding of PGG to VEGFA target protein was better than that of other two small molecular compounds, which was consistent with the results of molecular docking and biological activity experiments.

TJP1 promotes vascular mimicry in bladder cancer by facilitating VEGFA expression and transcriptional activity through TWIST1

Tight junction protein 1 (TJP1) is a recently identified prominent regulator of bladder cancer (BLCA) angiogenesis and tumorigenesis. Vascular mimicry (VM) is a newly described tumor feature and is correlated with an increased risk of tumor metastasis. However, the relationship between TJP1 expression and VM in bladder cancer remains elusive. In the present study, we report a novel function for TJP1 in accommodating VM to promote tumor progression. We found that the elevated TJP1 expression was positively related to VM in patients and xenograft tumor models in bladder cancer. Enforced expression of TJP1 increased VM of BLCA cells in vitro and in vivo by elevating Vascular endothelial growth factor A (VEGFA) levels. Furthermore, VM induced by TJP1 overexpression was significantly blocked by the VEGFA and VEGFR inhibitors (Bevacizumab and Sunitinib). Mechanistically, TJP1 promoted VEGFA transcriptional and protein level in a TWIST1-dependent manner. Taken together, our study reveals that TJP1-regulated VEGFA overexpression may indicate a potential therapeutic target for clinical intervention in the early tumor neovascularization of bladder cancer.

VEGFA promotes the occurrence of PLA2R-associated idiopathic membranous nephropathy by angiogenesis via the PI3K/AKT signalling pathway

BackgroundThe M-type phospholipase A2 receptor (PLA2R)-associated idiopathic membranous nephropathy (IMN) is a common immune-related disease in adults. Vascular endothelial growth factor A (VEGFA) is the key mediator of angiogenesis, which leads to numerous kidney diseases. However, the role of VEGFA in IMN is poorly understood.MethodsIn the present study, we downloaded the microarray data GSE115857 from Gene Expression Omnibus (GEO). The differentially expressed genes (DEGs) were identified with R software. The cytoHubba plug-in were used to identify hub genes from the protein–protein interaction network. Gene set enrichment analysis (GSEA) was used to identify signalling pathway in IMN. CCK8 was performed to assess the cell viability in human vascular endothelial cells (HVECs). Then, passive Heymann nephritis (PHN) was induced in rats by a single tail vein injection of anti-Fx1A antiserum. Animals treated with VEGFA inhibitor bevacizumab (BV), with saline as a positive control. Proteinuria was evaluated by biochemical measurements. Immunohistochemistry and immunofluorescence was used to evaluate relative proteins expression. Electron microscopy was performed to observe the thickness of the glomerular basement membrane (GBM).ResultsWe revealed 3 hub genes, including one up-regulated gene VEGFA and two down-regulated genes JUN and FOS, which are closely related to the development of PLA2R-associated IMN. Pathway enrichment analysis found that the biological process induced by VEGFA is associated with PI3K/Akt signalling. GSEA showed that the signalling pathway of DEGs in GSE115857 was focused on angiogenesis, in which VEGFA acts as a core gene. We confirmed the high expression of VEGFA, PI3K, and AKT in IMN renal biopsy samples with immunohistochemistry. In HVECs, we found that BV suppresses cell viability in a time and dose dependent manner. In vivo, we found low dose of BV attenuates proteinuria via inhibiting VEGFA/PI3K/AKT signalling. Meanwhile, low dose of BV alleviates the thickening of the GBM.ConclusionVEGFA/PI3K/AKT signalling may play significant roles in the pathogenesis of IMN, which may provide new targets for the treatment of IMN.

OVARIO, A Phase 2 Study of Niraparib + Bevacizumab in Advanced Ovarian Cancer Following Front-Line Platinum-Based Chemotherapy with Bevacizumab: Updated Analysis (039)

OVARIO, A Phase 2 Study of Niraparib + Bevacizumab in Advanced Ovarian Cancer Following Front-Line Platinum-Based Chemotherapy with Bevacizumab: Updated Analysis (039)

In Silico Docking Studies of Vascular Endothelial Growth Factor-A (VEGFA): Possible Implications in Chronic Myeloid Leukemia (CML) Therapy

Background: Dysregulated angiogenesis resulting in neovascularization is a critical event in the expansion and progression of Chronic Myeloid Leukemia (CML), hematopoietic cancer. Vascular Endothelial Growth Factor- A (VEGFA), an important angiogenesis mediator, has been a target for treating cancer. Although several anti-VEGFA drugs are available, they are associated with adverse side effects, promoting the need to identify better drugs that may be less toxic. Objective: Our aim was to investigate whether Tyrosine Kinase Inhibitors (TKIs) could be repurposed for use as VEGFA inhibitors via in silico docking software. We also investigated the potential of phytochemicals as VEGFA inhibitors. Methods: We performed molecular docking using Schrödinger Maestro software suite 2014-3 to determine the most potent phytochemical and TKI VEGFA antagonists. Results: Among the TKIs investigated, Bosutinib had the best binding affinity and may be the most potent TKI against VEGFA. The order of binding affinities for the top ten docked ligands was: Ginsenoside Rg3> Bosutinib> Vitamin D> Paclitaxel> Dasatinib> Saponins> Ponatinib> Squalamine> Imatinib> Nilotinib. We found that Ginsenoside Rg3 had the highest binding affinity (MMGBSA score= -99.4 kcal/mol, glide Gscore = -9.16 kcal/mol) to VEGFA. Conclusion: Our study has shown for the first time the binding poses of these TKIs and phytochemicals to VEGFA, using computational methods. We propose that the use of the top scoring ligands, in isolation or a combination of phytochemicals plus TKI, could serve as potent angiogenesis inhibitors via their binding to and inhibiting VEGFA expression to prevent CML progression. We have also profiled the ligand binding residues, which may be explored in designing pharmacophores.

SEMA3A partially reverses VEGF effects through binding to neuropilin-1

Cross-talk between hematopoietic stem cells (HSCs) and bone marrow stromal cells (BMSCs) is essential for HSCs regulation and leukemogenesis. Studying bone marrow of myelodysplasia patients, a pre-leukemic condition, we found mRNA overexpression of vascular endothelial growth factor A (VEGFA) in CD34+ HSCs and semaphorin 3A (SEMA3A) in BMSCs. To better understand the role of VEGFA and SEMA3A in leukemogenesis, we recruited 30 myelodysplastic syndrome (MDS) patients, 29 acute myeloid leukemia (6 secondary to MDS) patients and 12 controls. We found higher VEGFA expression in de novo AML patients (without prior MDS) group (p=0.0073) and higher SEMA3A expression in all BMSCs patient's samples compared to control group. We then overexpressed VEGFA in an acute myelogenous leukemia cell line, KG1 cells, and in normal CD34+ cells. This overexpression increased KG1 (p=0.045) and CD34+ cell (p=0.042) viability and KG1 (p=0.042) and CD34+ cell (p=0.047) proliferation. Moreover, KG1 and CD34+ cells overexpressing VEGFA also had increased proliferation when co-cultured with human marrow stromal HS5 cells (p=0.045 and p=0.02, respectively). However, co-culture of these transformed cells with HS5 cells overexpressing SEMA3A reduced KG1 (p=0.004) and CD34+ (p=0.009) proliferation. Co-culture of KG1 transformed cells with HS27 cells overexpressing SEMA3A reduced KG1 proliferation as well (p=0.01). To investigate whether the dominant SEMA3A effect over VEGFA could be due to competition for neuropilin1 receptor (NRP1), we performed immunoprecipitation with anti-NRP1 antibody of cell extracts of co-cultured KG1 and HS5 cells, induced or not by VEGFA and SEMA3A recombinant proteins. Results showed a preferential association of NRP1 with SEMA3A, suggesting that SEMA3A can partially reverse the effects caused by the VEGFA preventing its binding with the NRP1 receptor. Since both hematopoietic cells, leukemic and normal, showed similar behavior, we suppose that the attempt to reversion of VEGF effects by SEMA3A is a homeostatic phenomenon in the hematopoietic niche. Finally, we conclude that VEGFA overexpression confers AML cell advantages and SEMA3A may partially reverse this effect; thus, SEMA3A protein combined with VEGFA inhibitors could be beneficial for AML treatment.

Ten years of anti-vascular endothelial growth factor therapy.

The targeting of vascular endothelial growth factor A (VEGFA), a crucial regulator of both normal and pathological angiogenesis, has revealed innovative therapeutic approaches in oncology and ophthalmology. The first VEGFA inhibitor, bevacizumab, was approved by the US Food and Drug Administration in 2004 for the first-line treatment of metastatic colorectal cancer, and the first VEGFA inhibitors in ophthalmology, pegaptanib and ranibizumab, were approved in 2004 and 2006, respectively. To mark this tenth anniversary of anti-VEGFA therapy, we discuss the discovery of VEGFA, the successes and challenges in the development of VEGFA inhibitors and the impact of these agents on the treatment of cancers and ophthalmic diseases.

Vascular endothelial growth factor-A inhibitors

Vascular endothelial growth factor-A inhibitors

Thyroid Complications of Cancer Therapy: An Increasing Challenge

Thyroid Complications of Cancer Therapy: An Increasing Challenge

Overexpression of Cardiomyocyte α1A-Adrenergic Receptors Attenuates Postinfarct Remodeling by Inducing Angiogenesis Through Heterocellular Signaling.

Stimulation of cardiac α1A-adrenergic receptors (α1A-AR) has been proposed for treatment of heart failure, since it increases myocardial contractility. We investigated a different mechanism, induction of angiogenesis. Four to 6 weeks after permanent coronary artery occlusion, transgenic rats with cardiomyocyte-specific α1A-adrenergic receptor overexpression had less remodeling than their nontransgenic littermates, with less fibrosis, hypertrophy and lung weight, and preserved left ventricular ejection fraction and wall stress (all P<0.05). Coronary blood flow, measured with microspheres, increased in the infarct zone in transgenic rats compared with nontransgenic littermates (1.4±0.2 versus 0.5±0.08 mL min(-1) g(-1); P<0.05), which is consistent with angiogenesis, as reflected by a 20% increase in capillary density in the zone adjacent to the infarct. The question arose, how does transgenic overexpression of a gene in cardiomyocytes induce angiogenesis? We identified a paracrine mechanism, whereby vascular endothelial growth factor-A mRNA and protein were increased in isolated transgenic cardiomyocytes and also by nontransgenic littermate cardiomyocytes treated with an α1A-agonist, resulting in angiogenesis. Conditioned medium from cultured cardiomyocytes treated with an α1A agonist enhanced human umbilical vein endothelial cell tubule formation, which was blocked by an anti-vascular endothelial growth factor-A antibody. Moreover, improved cardiac function, blood flow, and increased capillary density after chronic coronary artery occlusion in transgenic rats were blocked by either a mitogen ERK kinase (MEK) or a vascular endothelial growth factor-A inhibitor. Cardiomyocyte-specific overexpression of the α1A-adrenergic receptors resulted in enhanced MEK-dependent cardiomyocyte vascular endothelial growth factor-A expression, which stimulates angiogenesis via a paracrine mechanism involving heterocellular cardiomyocyte/endothelial cell signaling, protecting against remodeling and heart failure after chronic coronary artery occlusion.

VEGF-A Inhibition Ameliorates Podocyte Apoptosis via Repression of Activating Protein 1 in Diabetes

Background/Aims: Vascular endothelial growth factor-A (VEGF-A) upregulation and podocyte apoptosis have been documented in diabetes. This study was designed to investigate whether inhibiting VEGF-A could ameliorate podocyte apoptosis in diabetes and the underlying mechanisms. Methods: In vitro, small interfering RNAs (siRNAs) of VEGF-A and activator protein 1 (AP-1, c-fos and c-jun), bevacizumab (VEGF-A inhibitor) and SP600125 (AP-1 inhibitor) were added to high glucose (30 m<smlcap>M</smlcap>) induced podocytes. Luciferase reporter assay was used to investigate whether AP-1 was a direct target of VEGF-A. In vivo, bevacizumab and SP600125 were administered to 12-week-old streptozotocin-induced male Sprague Dawley rats. The level of VEGF-A, c-fos, c-jun and bcl-2 were examined using immunostaining and Western blot analysis. Podocyte apoptosis was detected using the terminal deoxynucleotidyl transferase-mediated uridine 5′-triphosphate-biotin nick end labeling (TUNEL) assay, electron microscopy and flow cytometry. Results: Silencing VEGF-A or AP-1 upregulated bcl-2 and ameliorated podocyte apoptosis. Silencing VEGF-A decreased the level of c-fos and c-jun and bevacizumab and SP600125 treatment attenuated podocyte apoptosis. Luciferase reporter activity of VEGF-A-3′-UTR constructs was significantly provoked when stimulated with TGF-β1. In diabetic rat kidneys, VEGF-A co-localized with bcl-2 in podocytes. With bevacizumab and SP600125 treatment, the level of VEGF-A and AP-1 decreased while bcl-2 increased. Podocyte apoptotic rate was reduced with condensed podocyte nuclei less frequently observed. The urine albumin excretion rate (UAER) and albumin/creatinine were improved. Conclusion: This study demonstrates VEGF-A inhibition ameliorates podocyte apoptosis by regulating AP-1 and bcl-2 signaling. AP-1 is a direct target of VEGF-A and a novel player in podocyte apoptosis.

Glucocorticoids: Structure, Signaling and Molecular Mechanisms in the Treatment of Diabetic Retinopathy and Diabetic Macular Edema

Diabetic retinopathy (DR) is one of the leading causes of blindness in the working population worldwide. Vascular leakage, angiogenesis and neuronal degeneration are key features of DR. Current effective interventions for DR include treatment of systemic risk factors such as elevated blood glucose, blood pressure and dyslipidemia. Ocular treatments include vascular endothelial growth factor A (VEGF-A) inhibitors, laser photocoagulation and surgery. While anti-VEGF therapy has become as first-line treatment for diabetic macular edema (DME) that causes reduced vision, intravitreal glucocorticoids also have been shown to be efficacious in this situation. It has been reported that all the major pathological processes of DR are susceptible to glucocorticoid treatment. The effects of glucocorticoids on vascular leakage and angiogenesis may be mediated through their well established anti-inflammatory role. Alternatively, glucocorticoids may affect other mechanisms known to be activated in DR. Potential mechanisms for the anti-inflammatory effects of glucocorticoids include blockage of cytokine production and inhibition of leukocyte adhesion induced by VEGF-A. Glucocorticoids decrease the expression of VEGF-A directly, and increase the production, or decrease phosphorylation, of tight junction-associated proteins. Glucocorticoids have also been shown to be neuroprotective, in contrast to VEGF-A inhibitors which animal studies suggest may be neurotoxic. This review outlines the biological properties of synthetic glucocorticoids, with particular emphasis on the potential beneficial effect of combining glucocorticoids with anti-VEGF treatment for DME and DR.

P16INK4A Represses Breast Stromal Fibroblasts Migration/Invasion and Their VEGF-A-dependent Promotion of Angiogenesis through Akt Inhibition

Stromal fibroblasts, the most abundant and probably the most active cellular component of breast cancer-associated stroma, become active and promote angiogenesis through paracrine effects. However, it still unclear how these processes are regulated. Here, we have shown that down-regulation of the tumor suppressor p16(INK4A) protein enhances the migration/invasion abilities of breast stromal fibroblasts, which form dendritic network of extensions into matrigel. Furthermore, we present clear evidence that p16(INK4A) represses the expression/secretion of the proangiogenesis protein vascular endothelial growth factor A (VEGF-A). Consequently, p16(INK4A)-deficient breast stromal fibroblasts and mouse embryonic fibroblasts enhanced endothelial cell differentiation into capillary-like structures in a paracrine manner. This effect was suppressed by adding bevacizumab, a specific VEGF-A inhibitor. Additionally, p16(INK4A)-defective mouse embryonic fibroblasts enhanced angiogenesis in breast cancer xenografts in mice. Furthermore, we have shown that p16(INK4A) suppresses the Akt/mammalian target of rapamycin (mTOR) signaling pathway and its downstream effector hypoxia-inducible factor 1-alpha (HIF-1α), which transactivates VEGF-A. Consequently, Akt inactivation suppressed both the p16(INK4A)-dependent autocrine effect on fibroblast migration/invasion and the paracrine effect on angiogenesis, showing the important role of this protein kinase in mediating the various effects related to p16(INK4A) deficiency. These results indicate that p16(INK4A) is an efficient inhibitor of the migration/invasion abilities of breast stromal fibroblasts and also their paracrine proangiogenic effects, through inhibition of Akt. Therefore, pharmacologic restoration of p16(INK4A) level in stromal fibroblasts may be exploited as therapeutic strategy to help eradicate tumor cells and/or prevent their recurrence, through suppressing cell non-autonomous procarcinogenic mediators.

P.3.c.027 The effect of antipsychotic drug treatment on serum levels of vascular endothelial growth factor (VEGF), sVEGFR-1, and sVEGFR-2 in schizophrenia

Vascular endothelial growth factor-A (VEGF-A) is a crucial mediator of vascular hyperpermeability, angiogenesis, and inflammation, processes intimately involved in tissue repair. Although much emphasis has been placed on understanding the synthesis and stability of VEGF-A mRNA, relatively little attention has been given to the study of the stability and processing of VEGF-A proteins themselves. In recent years, several studies indicated that VEGF-A protein activity is highly controlled through interaction with angiogenic or non-angiogenic mediators. We analyzed mechanisms that might control extracellular VEGF-A processing during wound repair. First, our studies provide evidence that VEGF-A protein is a target of proteases present in the microenvironment of human chronic non-healing wounds. Serine proteases, in particular plasmin cleave VEGF165 and digested VEGF fragments, showed decreased mitogenic activity. Inactivation of the plasmin cleavage site Arg110/Ala111 preserved the structural integrity and increased the angiogenic potency of VEGF165 when tested in an impaired healing mouse model. Secondly, we identified significantly increased levels of the potent VEGF-A inhibitor, the soluble form of the VEGF receptor VEGFR-1 (sVEGFR-1) in non-healing wounds when compared to healing wounds. Wound closure in healing and non-healing wounds correlated significantly with a decline in sVEGFR-1 levels. These observations support the concept that VEGF-A and VEGF-A receptors are important mediators in tissue repair. Further, our data provide mechanisms how VEGF-A-mediated interactions are disturbed during impaired healing.

Combining Bevacizumab with Radiation or Chemoradiation for Solid Tumors: A Review of the Scientific Rationale, and Clinical Trials

Radiation therapy or the combination of radiation and chemotherapy is an important component in the local control of many tumor types including glioblastoma, rectal cancer, and pancreatic cancer. The addition of anti-angiogenic agents to chemotherapy is now standard treatment for a variety of metastatic cancers including colorectal cancer and non-squamous cell lung cancer. Anti-angiogenic agents can increase the efficacy of radiation or chemoradiation for primary tumors through mechanisms such as vascular normalization and augmentation of endothelial cell injury. The most commonly used anti-angiogenic drug, bevacizumab, is a humanized monoclonal antibody that binds and neutralizes vascular endothelial growth factor A (VEGF-A). Dozens of preclinical studies nearly uniformly demonstrate that inhibition of VEGF-A or its receptors potentiates the effects of radiation therapy against solid tumors, and this potentiation is generally independent of the type or schedule of radiation and timing of VEGF-A inhibitor delivery. There are now several clinical trials combining bevacizumab with radiation or chemoradiation for the local control of various primary, recurrent, and metastatic tumors, and many of these early trials show encouraging results. Some added toxicities occur with the delivery of bevacizumab but common toxicities such as hypertension and proteinuria are generally easily managed while severe toxicities are rare. In the future, bevacizumab and other anti-angiogenic agents may become common additions to radiation and chemoradiation regimens for tumors that are difficult to locally control.

Intersegmental vessel formation in zebrafish: requirement for VEGF but not BMP signalling revealed by selective and non-selective BMP antagonists.

Bone morphogenetic proteins (BMPs) were first identified through their role in inducing bone and cartilage formation, but many other important functions have since been ascribed to BMPs, including dorsoventral patterning, angiogenesis and tissue homeostasis. Using dorsomorphin and LDN193189, selective small molecule inhibitors of BMP signalling, we investigated the role of BMP signalling in early vascular patterning in zebrafish. The effects of dorsomorphin and LDN193189 on vascular endothelial growth factor-a (VEGF) and BMP signalling in developing zebrafish and in human pulmonary artery endothelial cells were determined using confocal microscopy, Western blotting and quantitative PCR. We showed that dorsomorphin, similar to the VEGF inhibitor SU5416, strongly inhibits intersegmental vessel formation in zebrafish and that this is due to inhibition of VEGF activation of VEGF receptor 2 (VEGFR2), leading to reduced VEGF-induced phospho-ERK (extracellular regulated kinase) 1/2 and VEGF target gene transcription. These effects occurred at concentrations of dorsomorphin that block BMP signalling. We also showed that LDN193189, an analogue of dorsomorphin, more potently blocks BMP signalling but has no effect on VEGF signalling in zebrafish and does not disrupt early vascular patterning. Dorsomorphin inhibits both BMP and VEGF signalling, whereas LDN193189 is a more selective BMP antagonist. Results obtained in cardiovascular studies using dorsomorphin need to be interpreted with caution, and use of LDN193189 would be preferable due to its selectivity. Our data also suggest that BMP signalling is dispensable for early patterning of intersegmental vessels in zebrafish.