VEGFs) And Their Receptors Research Articles

Structural analysis of vascular endothelial growth factor receptor‐2/ligand complexes by small‐angle X‐ray solution scattering

Receptor tyrosine kinases play essential roles in tissue development and homeostasis, and aberrant signaling by these molecules is the basis of many diseases. Understanding the activation mechanism of these receptors is thus of high clinical relevance. We investigated vascular endothelial growth factors (VEGFs) and their receptors (VEGFRs), which regulate blood and lymph vessel formation. We analyzed the structural changes in the extracellular receptor domain that were induced by ligand binding and that represent the initial step in transmembrane signaling, culminating in the activation of the intracellular receptor kinase domain. High-resolution structural information for the ligand binding domain became available recently, but the flexibility of the extracellular domain and inhomogeneous glycosylation of VEGFRs have prevented the production of highly diffracting crystals of the entire extracellular domain so far. Therefore, we chose to further investigate VEGFR structure by small-angle X-ray scattering in solution (SAXS). SAXS data were combined with independent distance restraint determination obtained by mass spectrometric analysis of chemically cross-linked ligand/receptor complexes. With these data, we constructed a structural model of the entire extracellular receptor domain in the unbound form and in complex with VEGF.

Aurora-A as an independent molecular prognostic marker in gastric cancer

Aurora-A, encoding serine/threonine kinases with a key role in mitosis has been demonstrated to be involved in tumor progression. Hematogenous and lymphatic metastasis are also involved in cancer progression. Vascular endothelial growth factors (VEGFs) and their receptors (VEGFRs) have been implicated in tumor-related angiogenesis and lymphagenesis. The purpose of this study was to analyze the expression and prognostic significance of Aurora-A, VEGFs and VEGFRs in gastric cancer. The expression profiles of Aurora-A, VEGF-A and VEGF-D in gastric cancer cell lines were detected employing real-time reverse transcription polymerase chain reaction and Western blot analysis. The expression levels of Aurora-A, VEGF-A/VEGFR-2, VEGF-D/VEGFR-3 and clinicopathological characteristics were analyzed in 89 gastric cancer patients treated with curative surgery. Univariate analysis demonstrated that histological grade (P=0.052), TNM stage (P<0.001), lymphovascular involvement (P<0.001), Aurora-A (P<0.001) and VEGF-D (P=0.048) were prognostic factors. The presence of Aurora-A was correlated with tumor progression (P=0.053) and shorter survival (P=0.001). Cox multivariate regression analysis demonstrated that Aurora-A positive expression, stage III-IV and lymphovascular involvement were independent unfavorable prognostic factors in gastric cancer. Aurora-A positive expression was predictive for worse outcome in patients without lymph node metastasis (P<0.05) and in patients with stage III-IV (P<0.001). Aurora-A could serve as an independent prognostic marker in gastric cancer and could identify patients with worse outcome even in a relatively early and local disease, thus offering valuable information for administering individualized treatment and/or surveillance for these patients.

Cancers du rein avancés en situations particulières. Comment les traiter?

Advanced or metastatic renal carcinoma represents a frequent disease in oncologic practice. Few years ago, in immunotherapy era, treatments had quickly reached deadlock. New therapies targeting vascular endothelial growth factors and their receptors (VEGF-R), sorafenib, sunitinib and bevacizumab, and the mammalian target of rapamycin (mTOR), temsirolimus and everolimus, have modified these patients prognosis and their quality of life in a few years. Nevertheless, patients included in randomized trials presented severe inclusion criteria. Then in the daily practice, patients have distinctive characteristics which were not evaluated in large pivotal studies: poor performance status, older patients, renal dysfunction, cerebral metastases or non clear cell renal cancer. In published trials, a few data concerning these situations are reported, and these studies have often included small samples, were retrospective or not randomised. However compared to global population, tolerance have not been very different in geriatric patients, or patients with poor performance status, or with central neurological metastases, or with papillary and chromophobe sub-types. On the contrary progression free or overall survivals increases are more difficult to confirm. Also before starting treatment, ratio between potential benefit and possible toxicities have to be evaluated. In patients with renal insufficiency, VEGF receptor inhibitors seem to be cautiously initiated at reduced doses, and to be increased according to tolerance. Due to these poor proof levels, clinical trials are needed for these specific populations.

The effects of VEGF-R1 and VEGF-R2 ligands on angiogenic responses and left ventricular function in mice

Vascular endothelial growth factors (VEGFs) and their receptors (VEGF-Rs) are among the most powerful factors regulating vascular growth. However, it has remained unknown whether stimulation of VEGF-R1, VEGF-R2 or both of the receptors produces the best angiogenic responses in myocardium. The aim of this study was to compare the VEGF-R1-specific ligand VEGF-B(186), VEGF-R2-specific ligand VEGF-E and VEGF-A(165,) which stimulates both receptors, regarding their effects on angiogenesis and left ventricular function in mice. High-resolution echocardiography was used to guide the closed-chest injections of adenoviral (Ad) vectors expressing VEGF-B(186,) VEGF-E, and VEGF-A(165) into the anterior wall of the left ventricle in C57Bl/6J mice. Angiogenic and functional effects were analysed using histology, ultrasound and perfusion analyses 6 (D6) and 14 (D14) days after the Ad injection. AdVEGF-A(165) induced a strong angiogenic response seen as an enlargement of myocardial capillaries whereas angiogenesis induced by AdVEGF-B(186) and AdVEGF-E seemed more physiological. The increase in the capillary area was accompanied with an increase in myocardial perfusion at D6 after the gene injection. AdVEGF-A(165) and AdVEGF-E induced endothelial-specific proliferation whereas AdVEGF-B(186) mostly induced proliferation of cardiomyocytes. AdVEGF-A(165) induced more pronounced tissue damage than AdVEGF-B(186) and AdVEGF-E. Left ventricular function measured as ejection fraction did not change during the follow-up. AdVEGF-A(165) increased both VEGF-R1 and VEGF-R2 protein expression whereas AdVEGF-B(186) and AdVEGF-E did not affect endogenous receptor expression levels. AdVEGF-B(186) and AdVEGF-E are equally potent in inducing therapeutic angiogenesis in mouse myocardium and produce less side effects than AdVEGF-A(165).

Stratégie de la prise en charge médicale dans les stades métastatiques

For a long time, advanced or metastatic renal carcinoma has presented a difficult therapeutic challenge in oncology practice. Only a few years ago, treatment mainly involved immunotherapy, which frequently led to stalemate. In just a few years, new therapies targeting vascular endothelial growth factors and their receptors (VEGF-R): sorafenib, sunitinib and bevacizumab, or the mammalian target of rapamycin (mTOR): temsirolimus and everolimus, have changed these patients’ prognosis and also their quality of life. With these new treatments, median overall survival is more than 26 months, when it was from 10 to 12 months with immunotherapy. Published trial data mean that in 2009 we have decision-making algorithms enabling us to offer patients optimal treatment. However, patients with distinctive characteristics (cerebral metastases, papillary or chromophobe histological subtype, older patients, etc) have rarely been evaluated with respect to the effect of these molecules. A few data are reported, often concerning small samples, in retrospective or not randomised trials. For these situations, tolerance and therapeutic impact of targeted therapies will be studied in future clinical trials.

Prognostic significance of the expression of vascular endothelial growth factors A, B, C, and D and their receptors R1, R2, and R3 in patients with nonsmall cell lung cancer

Lung cancer is the leading cause of cancer death in the world. The objective of this study was to investigate the expression of vascular endothelial growth factors (VEGFs) and their receptors (VEGFRs) in patients with nonsmall cell lung cancer (NSCLC) and its correlation with the prognosis for patients with lung cancer. The expression status of VEGFs and VEGFRs was examined in 48 nonconsecutive specimens of primary lung cancer by immunohistochemistry. Correlations between the expression of VEGFs and VEGFRs and clinicopathologic parameters were analyzed. Nineteen of 48 samples (39.6%) were moderately/highly immunoreactive for VEGF-A, 6 samples (12.5%) were reactive for VEGF-B, 14 samples (29.2%) were reactive for VEGF-C, 11 samples (22.9%) were reactive for VEGF-D, 20 samples (41.7%) were reactive for VEGFR1, 26 samples (54.2%) were reactive for VEGFR2, 20 samples (41.7%) were reactive for VEGFR3, and 19 samples (39.6%) were reactive for nuclear expression of VEGFR3. Patients with moderate/high VEGF-C, VEGFR1, and VEGFR2 expression had worse survival, whereas patients with moderate/high VEGF-D and nuclear VEGFR3 expression had better survival. After adjusting according to tumor stage, VEGF-B and VEGF-D expression had a significant correlation with worse survival in patients with stage I and II disease. Patients with stage III and IV disease who had VEGFR1 and VEGFR2 expression had worse survival, whereas the expression of VEGF-D was correlated significantly with better survival. Finally, stage, VEGF-D expression, and VEGFR1 expression were significantly independent prognostic predictors. The results of the current study indicated that the over-expression of VEGFs and VEGFRs plays an important role in the survival of patients with NSCLC. The inclusion of angiogenic factors in the standard pathologic study of lung cancer may improve the clinical evaluation of patients with NSCLC.

Vascular endothelial growth factors synthesized by human lung mast cells exert angiogenic effects

Angiogenesis and lymphangiogenesis are critical for several allergic, inflammatory, and neoplastic disorders. Mast cells infiltrate the sites of inflammation and tumors. We sought to characterize the expression and functions of vascular endothelial growth factors (VEGFs) and their receptors (VEGFRs) in human mast cells. VEGF expression was evaluated by means of RT-PCR and Western blotting in primary human lung mast cells and in the mast cell lines LAD-2 and HMC-1. Angiogenic activity of mast cell supernatants was determined by using the chick embryo chorioallantoic membrane assay. VEGFR expression was assessed by means of RT-PCR and flow cytometry. Modified Boyden chambers were used for chemotaxis assay. Human mast cells express VEGF-A, VEGF-B, VEGF-C, and VEGF-D at both the mRNA and protein level. Prostaglandin E(2) (PGE(2)) enhanced the expression of VEGFA, VEGFB, and VEGFC, whereas an adenosine analog (5'-[N-ethylcarboxamido] adenosine [NECA]) increased VEGFA, VEGFC, and VEGFD expression. In addition, PGE(2) and NECA enhanced VEGF-A release, and supernatants of PGE(2)- and NECA-activated human lung mast cells induced angiogenic responses in the chorioallantoic membrane assay that were inhibited by an anti-VEGF-A antibody. Mast cells expressed mRNA for VEGFR1 and VEGFR2. These receptors were present on the mast cell surface. VEGF-A(165), VEGF-B(167), VEGF-C, VEGF-D, and placental growth factor 1 induced mast cell chemotaxis. These chemotactic effects were mediated by the activation of both VEGFR-1 and VEGFR-2. Our data indicate that human mast cells are both a source and a target of angiogenic and lymphangiogenic factors and therefore might play a role in inflammatory and neoplastic angiogenesis through the expression of several forms of VEGFs and their receptors.

Differential Expression of VEGF Receptors on Human T Cell Subsets

Vascular endothelial growth factors (VEGF) and their receptors (VEGFR) deliver important signals that controlling the angiogenesis of growing, injured, and malignant tissues. Expression of VEGF receptors has been described on a variety of normal and malignant leukocytes, and presumably plays a role in their trafficking to injured tissue. Differential expression of neuropilin-1 (NRP-1) has been described as a marker of regulatory T cells (Treg) in mice, and may be expressed on human thymocytes, but expression on human Tregs has not been confirmed. Moreover, earlier reports of Treg-specific markers in humans were based on the CD4+CD25+ subset, which is not as reliable for Treg identification as it is in mice. We used cell sorting to obtain pure populations of human Tregs (using the more stringent criteria of CD4+CD25+CD125−) and T helper cells (Th) (CD4+CD25−CD127+). We compared expression in these subsets of VEGFR-1, -2, -3, and NRP-1 and -2 using flow cytometry and quantitative RT-PCR, and NRP-1 splice isoforms by RT-PCR using isoform-specific primers. We were not able to identify any NRP-1 expression in Tregs by any of these methods. In contrast, there was marked differential expression of VEGFR-2 (Flk-1) and -3 (Flt-4), both of which exhibited up to 16-fold higher expression in Tregs than in Th. These findings suggest a role for VEGF in recruiting Tregs to sites of tissue growth as a mechanism for blunting potential autoimmunity to neoexpressed antigens, and thus may also explain the accumulation of Tregs in tumors.

Prognostic relevance of pretreatment soluble vascular endothelial growth factors (A,C,D) and their receptors (R1, R2, and R3) in advanced melanoma patients

8540 Background and aims: Circulating vascular endothelial growth factors (VEGFs) and their receptors (VEGFRs) were described as prognostic factors in several cancers. The aim of this study is to evaluate the prognostic values of these parameters in metastatic malignant melanoma diseases. Methods and Patients: Using enzyme-linked immunosorbent assays, VEGFs (A, C, D) and their receptors (1, 2, and 3) were measured in sera of 75 patients with metastatic malignant melanoma in comparison to 30 healthy controls. Disease free survival (DFS) and overall survival (OS) were calculated from the beginning of the treatment until either the progression or the death and analyzed using the Kaplan-Meier method. Results: Pretreatment median sVEGF-A, sVEGF-C as well as VEGFR-3 levels were significantly higher in MMM patients as compared to healthy ones p=0.0017, p=0.005, p&lt;0.00001 respectively). None of the studied markers correlated with gender, age or LDH levels. An inverse correlation between soluble VEGFR-2 and VEGF-D levels was observed (r=−0.33 p=0.040). High soluble VEGF-C and VEGFR-3 were correlated to high tumor burden (p=0.02, p=0.045). However, a relationship with lymph node metastasis was observed with sVEGF-A but not with sVEGFR-3. As shown by univariate analysis, only elevated levels of sVEGF-R1 concentration were found to exert a significantly unfavorable impact on both disease-free (χ2=6.64, p= 0.014) and overall survival (χ2=8.03, p= 0.0046). Conclusions: Our results suggest that, in metastatic malignant melanoma patients, soluble VEGF-A and VEGF-R3 pretreatment levels may prospectively identify high-risk patients with a worse prognosis; serum VEGFR-1 level may be a predictive factor of time to progression and overall survival. A multivariate analysis (Cox test) is ongoing in order to confirm if this parameter is an independent prognostic factor. No significant financial relationships to disclose.

EphB4 controls blood vascular morphogenesis during postnatal angiogenesis

Guidance molecules have attracted interest by demonstration that they regulate patterning of the blood vascular system during development. However, their significance during postnatal angiogenesis has remained unknown. Here, we demonstrate that endothelial cells of human malignant brain tumors also express guidance molecules, such as EphB4 and its ligand ephrinB2. To study their function, EphB4 variants were overexpressed in blood vessels of tumor xenografts. Our studies revealed that EphB4 acts as a negative regulator of blood vessel branching and vascular network formation, switching the vascularization program from sprouting angiogenesis to circumferential vessel growth. In parallel, EphB4 reduces the permeability of the tumor vascular system via activation of the angiopoietin-1/Tie2 system at the endothelium/pericyte interface. Furthermore, overexpression of EphB4 variants in blood vessels during (i) vascularization of non-neoplastic cell grafts and (ii) retinal vascularization revealed that these functions of EphB4 apply to postnatal, non-neoplastic angiogenesis in general. This implies that both neoplastic and non-neoplastic vascularization is driven not only by a vascular initiation program but also by a vascular patterning program mediated by guidance molecules.

Vascular Endothelial Growth Factor-C and Its Receptor VEGFR-3 in the Nasal Mucosa and in Nasopharyngeal Tumors

Vascular endothelial growth factors (VEGFs) and their receptors (VEGFRs) are important regulators of blood and lymphatic vessel growth and vascular permeability. Both blood and lymphatic vessels of the upper respiratory tract play important roles in pathological conditions, such as infections and tumors. Here we have studied the expression of VEGF-C and its receptor VEGFR-3 in the upper respiratory system by Northern blot analysis and immunohistochemistry of human tissues, and in situ mRNA hybridization of developing mouse embryos and beta-galactosidase staining of mouse embryos having a LacZ marker gene in the VEGFR-3 gene locus. The results demonstrate expression of VEGF-C and VEGFR-3 in the developing and adult nasal respiratory epithelium and in the nasal vascular plexus, respectively. Unlike in most other tissues, in the nasal mucosa VEGFR-3 is expressed in both blood and lymphatic vessels. Expression of VEGF-C was also detected in nasal and nasopharyngeal tumor islands, which were surrounded by VEGFR-3-positive angiogenic blood vessels. These results suggest that VEGF-C and VEGFR-3 have a role in the development of the nasal submucosal vascular plexus and in its normal function and that they are associated with angiogenesis in nasal and nasopharyngeal tumors.

VEGFc and VEGFR3 expression in human thyroid pathologies.

In vertebrates, vascular endothelial growth factors (VEGFs) and their receptors (VEGFRs) are major determinants of angiogenesis. In adults, the interaction between VEGFc and VEGFR3 (previously FLT4) is more specifically involved in the biology of lymphatics. Using PCR amplification of reverse-transcribed mRNA, we studied the expression of the VEGFR3 (including its short and long forms) and VEGFc genes in 38 samples of various human thyroid pathologies. VEGFR3 mRNA was detected in all samples of adenomas, nodular goiters and focal goitrogenic alterations; in all samples of thyroid tissue from patients with auto-immune diseases; and in some samples of adenocarcinomas. VEGFc mRNA was detected in most samples. We studied expression of the VEGFR3 and VEGFc proteins in thyroid tumors using appropriate antibodies. Co-expression of VEGFR3 and VEGFc was observed in most samples.