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Background: Behçet's disease (BD) is a chronic inflammatory vasculitis affecting multiple organs. Uveitis is frequently seen in patients with BD, especially in Turkish population., Objectives: To investigate vascular endothelial growth factor (VEGF) gene polymorphisms along with the levels of VEGF and VEGF receptors in patients with Behçet's uveitis (BU)., Material and Methods: Fifty-five BD-associated uveitis patients and 30 ageand sex-matched controls were included in this case-control study. The genotypes of the single nucleotide poymorphisms (SNPs): rs2010963 (+405G), rs3025039 (+936T) and rs699947 (-2598A) of the VEGF-A gene were determined using real-time polymerase chain reaction (RT-PCR) and serum levels of VEGF and VEGF receptors were measured using enzyme-linked immunosorbent assay (ELISA)., Results: No associations of the VEGF gene polymorphisms were observed in BD uveitis patients, but arthritis was present in 53.3% of patients not possessing CT genotype in C3025039→T polymorphism (p = 0.024). Although there were no statistically significant differences in serum VEGF-A, VEGF-C and soluble vascular endothelial growth factor receptor-3 (sVEGFR-3) levels (p < 0.05), serum vascular endothelial growth factor receptor-1 (VEGFR-1) and sVEGFR-3 levels were significantly higher in the BD group (p < 0.001 and p = 0.001, respectively). In addition, VEGF-C/soluble vascular endothelial growth factor receptor-2 (sVEGFR-2) ratio was significantly higher (p < 0.001), while VEGF-A/VEGFR-1 and VEGF-C/sVEGFR-3 ratios were significantly lower (p < 0.001 and p = 0.033, respectively) in BD patients compared to controls. Also, VEGF-C/sVEGFR-3 (p = 0.024, r = 0.37) and VEGF-C/sVEGFR-2 (p = 0.020, r = 0.38) ratios were positively correlated with disease duration., Conclusions: The significant changes in sVEGFR-3 levels and VEGF-C/sVEGFR-3 ratio has shown that lymphangiogenesis processes might take place in the pathogenesis of BD uveitis, and these parameters can be important indicators of evaluation of BD patients with uveitis together with disease duration.

Objective: Pulmonary arterial hypertension (PAH) is a major complication in systemic sclerosis (SSc), a disease marked by vascular and lymphatic vessel abnormalities. This study was undertaken to assess the role of the lymphangiogenic factors vascular endothelial growth factor C (VEGF-C) and angiopoietin 2 (Ang-2) and the soluble forms of their respective cognate receptors, soluble VEGF receptor 3 (sVEGFR-3) and soluble TIE-2, in patients with SSc, and to evaluate their predictive ability as markers for PAH development in SSc., Methods: In this cohort study, we used multiplex bead assays to assess serum levels of lymphangiogenic factors in 2 well-characterized SSc cohorts: an unselected identification cohort of SSc patients from Oslo University Hospital (n = 371), and a PAH-enriched validation cohort of SSc patients from Zurich University Hospital and Oslo University Hospital (n = 149). As controls for the identification and validation cohorts, we obtained serum samples from 100 healthy individuals and 68 healthy individuals, respectively. Patients in whom SSc-related PAH was identified by right-sided heart catheterization (RHC) in both cohorts were studied in prediction analyses. PAH was defined according to the European Society of Cardiology/European Respiratory Society 2015 guidelines for the diagnosis and treatment of PAH. Associations of serum levels of lymphangiogenic factors with the risk of PAH development were assessed in logistic regression and Cox regression analyses. Associations in Cox regression analyses were expressed as the hazard ratio (HR) with 95% confidence interval (95% CI)., Results: In the identification cohort, SSc patients had lower mean serum levels of VEGF-C and higher mean serum levels of Ang-2 compared to healthy controls (for VEGF-C, mean ± SD 2.1 ± 0.5 ng/ml in patients versus 2.5 ± 0.4 ng/ml in controls; for Ang-2, mean ± SD 6.1 ± 7.6 ng/ml in patients versus 2.8 ± 1.8 ng/ml in controls; each P < 0.001); these same trends were observed in SSc patients with PAH compared to those without PAH. The association of serum VEGF-C levels with SSc-PAH was confirmed in the PAH-enriched RHC validation cohort. For prediction analyses, we assembled all 251 cases of SSc-PAH identified by RHC from the identification and validation cohorts. In multivariable Cox regression analyses adjusted for age and sex, the mean serum levels of VEGF-C and sVEGFR-3 were predictive of PAH development in patients with SSc (for VEGF-C, HR 0.53 [95% CI 0.29-0.97], P = 0.04; for sVEGFR-3, HR 1.21 [95% CI 1.01-1.45], P = 0.042)., Conclusion: These findings support the notion that lymphangiogenesis is deregulated during PAH development in SSc, and indicate that VEGF-C could be a promising marker for early PAH detection in patients with SSc., (© 2021 The Authors. Arthritis & Rheumatology published by Wiley Periodicals LLC on behalf of American College of Rheumatology.)

Significance: A differential outcome in randomized controlled trials of anti-vascular endothelial growth factor (anti-VEGF) therapy, including ranibizumab, for diabetic macular edema is a major dilemma for planning, optimizing, and managing clinical usage. The variable outcome of the therapeutics necessitates the importance of finding a predictive biomarker for anti-VEGF therapy to improve subject selection., Purpose: Our study correlates the baseline pro- and anti-VEGF isoforms and its three receptors (VEGFReceptor1, VEGFReceptor2, and VEGFReceptor3) for circulatory candidate protein molecules among diabetic patients with macular edema, with the clinical outcome of ranibizumab therapy., Methods: This study included 86 individuals who were anti-VEGF naive at the time of ascertainment but have completed the standardized therapy regimen of the clinic. Plasma proteins for pro- and anti-VEGF isoforms and its three receptors were determined in replicate by an enzyme-linked immunosorbent assay., Results: The study demonstrated that 56 (65.12%) individuals benefited from the therapy in terms of letter gain (Snellen chart). Baseline plasma soluble VEGF receptor 2 (sVEGFR-2) was significantly higher among responders (65.10 pg/mL; 95% confidence interval, 55.41 to 74.80 pg/mL) compared with nonresponders (46.38 pg/mL; 95% confidence interval, 38.69 to 54.07 pg/mL; PFDR = .03). Diffuse diabetic macular edema with proliferative diabetic retinopathy increases the risk of nonresponse to the therapy by 3.03-fold (PFDR = .04)., Conclusions: The present study postulates that diffuse diabetic macular edema with proliferative diabetic retinopathy and baseline circulatory soluble VEGF receptor 2 may be potential candidates as therapy-stratifying markers for ranibizumab treatment among patients with diabetic macular edema., Competing Interests: Conflict of Interest Disclosure: None of the authors have reported a financial conflict of interest., (Copyright © 2020 American Academy of Optometry.)

Purpose: Various biomarkers have been proposed for sunitinib therapy in gastrointestinal stromal tumor (GIST). However, the lack of "real-life" comparative studies hampers the selection of the most appropriate one. We, therefore, set up a pharmacometric simulation framework to compare each proposed biomarker., Experimental Design: Models describing relations between sunitinib exposure, adverse events (hand-foot syndrome, fatigue, hypertension, and neutropenia), soluble VEGFR (sVEGFR)-3, and overall survival (OS) were connected to evaluate the differences in survival and adverse events under different dosing algorithms. Various fixed dosing regimens [4/2 (weeks on/weeks off) or 2/1 (50 mg), and continuous daily dosing (37.5 mg)] and individualization approaches [concentration-adjusted dosing (CAD), toxicity-adjusted dosing (TAD), and sVEGFR-3-adjusted dosing (VAD)] were explored following earlier suggested blood sampling schedules and dose-reduction criteria. Model-based forecasts of biomarker changes were evaluated for predictive accuracy and the advantage of a model-based dosing algorithm was evaluated for clinical implementation., Results: The continuous daily dosing regimen was predicted to result in the longest survival. TAD (24.5 months) and VAD (25.5 months) increased median OS as compared with a fixed dose schedule (19.9 and 21.5 months, respectively) and CAD (19.7 and 21.3 months, respectively), without markedly raising the risk of intolerable toxicities. Changes in neutrophil count and sVEGFR-3 were accurately forecasted in the majority of subjects (>65%), based on biweekly blood sampling., Conclusions: Dose adjustments based on the pharmacodynamic biomarkers neutrophil count and sVEGFR-3 can increase OS while retaining drug safety. Future efforts could explore the possibility of incorporating a model-based dose approach in clinical practice to increase dosing accuracy for these biomarkers., (©2020 American Association for Cancer Research.)

Vascular Endothelial Growth Factor Receptor 3 (VEGFR-3) is a very important factor which promotes lymphangiogenesis not only in physiological but also in pathological processes in which we can include neoplasia. The aim of this study was to analyze the plasma concentrations and diagnostic utility of this parameter in comparison and in combination with CA 15-3 in breast cancer (BC) patients and in relation to the control groups. The study included 120 breast cancer and 60 control patients (28 with benign breast tumors and 32 healthy women). Plasma levels of VEGFR-3 were determined by an Enzyme-Linked Immunosorbent Assay (ELISA), and those of CA 15-3 by a Chemiluminescent Microparticle Immuno Assay (CMIA). Differences in concentrations of both of the tested parameters were statistically significant when breast cancer patients were compared to the control groups. VEGFR-3 had higher values of sensitivity (SE), specificity (SP), predictive value of a positive (PPV) and negative test result (NPV) in the whole BC group (90%; 98.33%; 99.08%; 83.10%, respectively) and, more importantly, in the early stages of BC, than CA 15-3. VEGFR-3 was also a better parameter in terms of statistically significant Area Under Curve (0.9656) in the whole group and at all BC stages (I-IV), but a maximum range was obtained for the combination of VEGFR-3 and CA 15-3 (0.9710). The combined analysis of VEGFR-3 and CA 15-3 provides hope that a new BC diagnostic panel may be developed in the future.

Background: We have previously conducted the Sirolimus and Autophagy Inhibition in LAM (SAIL) trial, a phase 1 dose-escalation study of the combination of sirolimus and hydroxychloroquine in patients with lymphangioleiomyomatosis (LAM). The goal of the present study was to analyze sera from the SAIL trial to identify novel biomarkers that could shed light into disease pathogenesis and response to therapy., Methods: We used the DiscoveryMAP platform from Rules Based Medicine to simultaneously measure 279 analytes in sera collected at each visit from subjects enrolled in the SAIL trial. We used longitudinal regression and pathway analysis to examine analyte rate of change and corresponding effect on lung function and to identify networks and potential nodes of interest., Results: A total of 222 analytes were included in the analysis. We identified 32 analytes that changed over the treatment period of the study. Pathway analysis revealed enrichment in cytokine-receptor interaction and mechanistic/mammalian target of rapamycin-related pathways, in addition to seemingly unrelated processes such as rheumatoid arthritis. Search Tool for the Retrieval of Interacting Genes/Proteins analysis identified two hubs centered around acetyl-CoA carboxylase alpha and beta and coagulation factor II. In addition, we identified vascular endothelial growth factor receptor-3 and CCL21 as molecules significantly associated with changes in FEV 1 during the study period., Conclusions: We performed a large-scale analyte study in sera of women with LAM and identified potential markers that could be linked to disease pathogenesis, lung injury, and therapeutic response. These data will enable future investigation into the specific roles of these molecules in LAM., Trial Registry: ClinicalTrials.gov; No. NCT01687179; URL: www.clinicaltrials.gov)., (Copyright © 2018 American College of Chest Physicians. All rights reserved.)

To discover novel biomarkers of psoriasis, a target-specific antibody array screening of serum samples from psoriasis patients was initially performed. The results revealed that vascular endothelial growth factor receptor 3 (VEGFR-3) was significantly elevated in the sera of psoriasis patients, compared to healthy controls. Next, ELISA validation studies in a larger cohort of psoriasis patients (N = 73) were conducted, which confirmed that serum VEGFR-3 was indeed significantly increased in patients with psoriasis compared to healthy controls (P < 0.001). Furthermore, receiver operating characteristic curve analysis demonstrated that serum VEGFR-3 exhibited potential in distinguishing healthy controls from psoriasis patients: area under the curve = 0.85, P < 0.001. In addition, serum levels of VEGFR-3 were correlated with Psoriasis Area Severity Index scores (R = 0.32, P = 0.008) in psoriasis patients. Interestingly, serum VEGFR-3 levels were significantly elevated in psoriatic arthritis compared to non-psoriatic arthritis (P = 0.026). A pilot longitudinal study demonstrated that serum levels of VEGFR-3 could reflect disease progression in psoriasis. Collectively, serum VEGFR-3 may have a clinical value in monitoring disease activity of psoriasis., (© 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: Lymphedema (LE) is a chronic clinical manifestation of filarial nematode infections characterized by lymphatic dysfunction and subsequent accumulation of protein-rich fluid in the interstitial space-lymphatic filariasis. A number of studies have identified single nucleotide polymorphisms (SNPs) associated with primary and secondary LE. To assess SNPs associated with LE caused by lymphatic filariasis, a cross-sectional study of unrelated Ghanaian volunteers was designed to genotype SNPs in 285 LE patients as cases and 682 infected patients without pathology as controls. One hundred thirty-one SNPs in 64 genes were genotyped. The genes were selected based on their roles in inflammatory processes, angiogenesis/lymphangiogenesis, and cell differentiation during tumorigenesis., Results: Genetic associations with nominal significance were identified for five SNPs in three genes: vascular endothelial growth factor receptor-3 (VEGFR-3) rs75614493, two SNPs in matrix metalloprotease-2 (MMP-2) rs1030868 and rs2241145, and two SNPs in carcinoembryonic antigen-related cell adhesion molecule-1 (CEACAM-1) rs8110904 and rs8111171. Pathway analysis revealed an interplay of genes in the angiogenic/lymphangiogenic pathways. Plasma levels of both MMP-2 and CEACAM-1 were significantly higher in LE cases compared to controls. Functional characterization of the associated SNPs identified genotype GG of CEACAM-1 as the variant influencing the expression of plasma concentration, a novel finding observed in this study., Conclusion: The SNP associations found in the MMP-2, CEACAM-1, and VEGFR-3 genes indicate that angiogenic/lymphangiogenic pathways are important in LE clinical development.

Objective: The underlying problem in lymphatic filariasis is irreversible swelling of the limbs (lymphoedema), which is a unique feature of lymphatic insufficiency. It is still unclear whether the natural ability of lymphatics to form functional lymphatic vasculature is achieved or attenuated in the lymphoedemal pathology. Clinical studies have clearly shown that circulating lymphatic progenitors (CLPs), a subset of bone marrow-derived mononuclear cells (PBMCs), contribute to post-natal lymph vasculogenesis. CLP-based revascularisation could be a promising strategy to bypass the endothelial disruption and damage incurred by the filarial parasites. Thus our aim was to compare and characterise the functional prowess of PBMCs in physiological and lymphoedemal pathology., Methods: PBMCs were isolated from venous blood sample from drug-naive endemic normals (EN) and drug-deprived filarial lymphoedema (FL) individuals using density gradient centrifugation. Adhesion, transwell migration and in vitro matrigel assays were employed to characterise the lymphvasculogenic potential of PBMCs. CLPs were phenotypically characterised using flow cytometry; expression levels of lymphatic markers and inflammatory cytokines were quantified using qRT-PCR and ELISA, respectively., Results: PBMCs from FL group display poor adherence to fibronectin (P = 0.040), reduced migration towards SDF-1α (P = 0.035), impaired tubular network (P = 0.004) and branching point (P = 0.048) formation. The PBMC mRNA expression of VEGFR3 (P = 0.039) and podoplanin (P = 0.050) was elevated, whereas integrin α9 (P = 0.046) was inhibited in FL individuals; additionally, the surface expression of CD34 (P = 0.048) was significantly reduced in the FL group compared to the EN group., Conclusion: PBMCs from filarial lymphoedema show defective and dysregulated lymphvasculogenic function compared to endemic normals., (© 2017 John Wiley & Sons Ltd.)

The tyrosine kinase inhibitor sunitinib is used as first-line therapy in patients with metastasized renal cell carcinoma (mRCC), given in fixed-dose regimens despite its high variability in pharmacokinetics (PKs). Interindividual variability of drug exposure may be responsible for differences in response. Therefore, dosing strategies based on pharmacokinetic/pharmacodynamic (PK/PD) models may be useful to optimize treatment. Plasma concentrations of sunitinib, its active metabolite SU12662, and the soluble vascular endothelial growth factor receptors sVEGFR-2 and sVEGFR-3, were measured in 26 patients with mRCC within the EuroTARGET project and 21 patients with metastasized colorectal cancer (mCRC) from the C-II-005 study. Based on these observations, PK/PD models with potential influence of genetic predictors were developed and linked to time-to-event (TTE) models. Baseline sVEGFR-2 levels were associated with clinical outcome in patients with mRCC, whereas active drug PKs seemed to be more predictive in patients with mCRC. The models provide the basis of PK/PD-guided strategies for the individualization of anti-angiogenic therapies., (© 2017 The Authors CPT: Pharmacometrics & Systems Pharmacology published by Wiley Periodicals, Inc. on behalf of American Society for Clinical Pharmacology and Therapeutics.)

Aim: Esophageal cancer is one of the most aggressive tumors of the gastrointestinal tract. In this study, we quantified the serum vascular endothelial growth factor-3 (VEGFR-3) expression in patients with esophageal squamous cell carcinoma (ESCC) to evaluate the role of VEGFR-3 in ESCC., Materials and Methods: Ninety five patients with ESCC were studied. Pre-therapy and preoperative samples were stored and ELISA was used to designate the concentrations of VEGFR-3., Results: The serum values of VEGFR-3 were significantly higher in patients with ESCC than in healthy donors (p<0.0001)., Conclusions: The results imply a very good sensitivity of VEGFR-3 in ESCC. VEGFR-3 may be a good diagnostic biomarker for ESCC., Key Words: Biomarker, ESCC, VEGFR-3.

The purpose of this study was to evaluate the association of (18)F-fluorodeoxyglucose (FDG)-PET/CT findings with the vascular endothelial growth factor (VEGF) family and its receptor (VEGFR) levels in metastatic and nonmetastatic colorectal cancer (CRC). Fluorine-18 FDG-PET/CT scans were performed for initial staging and restaging of patients with CRC. FDG-PET/CT findings of tumor (such as the presence of a primary tumor, the lymphatic or distance metastases, and the maximum standardized uptake value (SUVmax) of the primary tumor), serum VEGF A-C-D-E levels, and serum VEGF receptor 1-2-3 levels were analyzed. A total of 63 patients were included into the study (35 males, mean age 61.3 ± 11.9 years). Patients were divided into two groups, based on positive and negative PET/CT findings. Patients were also categorized according to the presence of metastasis. All evaluated parameters were significantly higher in the PET/CT-positive group than the PET/CT-negative group (p < 0.001). All those parameters were also positively correlated with each other. The highest correlation for SUVmax of primary tumor was found with VEGFR-3 (p < 0.001, r = 0.665). Patients with metastases had high levels of VEGF-D, VEGF-A, VEGF-C, VEGF-E, and VEGFR-3 than those without metastases. These parameters had better specificity and sensitivity values than the SUVmax of the primary tumor for detection of metastases. However, VEGF-D was the best indicator of metastasis in all of those parameters (VEGF-D vs SUVmax; sensitivity 100 vs 100 %; specificity 76 vs 76 %; AUC 0.903 vs 0.835; p < 0.001, respectively). Vascular endothelial growth factor family and its receptors were significantly higher in metastatic CRC patients. VEGF-D was the best indicator of metastasis than all VEGF family, VEGFR-3, and primary tumor SUVmax. VEGF family (A-C-D-E) and VEGFR-3 may help to determine the prognosis and management of CRC.

Background: Angiogenesis and lymphangiogenesis are classical features of granuloma formation in pulmonary tuberculosis (PTB). In addition, the angiogenic factor--VEGF-A is a known biomarker for PTB., Aims/methodology: To examine the association of circulating angiogenic factors with PTB, we examined the systemic levels of VEGF-A, VEGF-C, VEGF-D, VEGF-R1, VEGF-R2 and VEGF-R3in individuals with PTB, latent TB (LTB) or no TB infection (NTB)., Results: Circulating levels of VEGF-A, VEGF-C andVEGF-R2 were significantly higher in PTB compared to LTB or NTB individuals. Moreover, the levels of VEGF-A, VEGF-C and VEGF-R2 were significantly higher in PTB with bilateral and/or cavitary disease. The levels of these factors also exhibited a significant positive relationship with bacterial burdens in PTB. ROC analysis revealed VEGF-A and VEGF-R2 as markers distinguishing PTB from LTB or NTB. Finally, the circulating levels of all the angiogenic factors examined were significantly reduced following successful chemotherapy., Conclusion: Therefore, our data demonstrate that PTB is associated with elevated levels of circulating angiogenic factors, possibly reflecting vascular and endothelial dysfunction. In addition, some of these circulating angiogenic factors could prove useful as biomarkers to monitor disease severity, bacterial burden and therapeutic responses.

Objective: To investigate serum vascular endothelial growth factor-C (VEGF-C), VEGF-D and VEGFR-3 levels in patients with papillary thyroid carcinoma (PTC) and analyze their relation with the clinicopathological and thyroid function of the patients., Methods: Serum samples and the data of thyroid function were collected from 55 patients with PTC and 24 with benign thyroid tumor (BT). ELISA was used to detect VEGF-C/D and VEGFR-3 concentration in the serum samples and their relation with the thyroid function was analyzed., Results: The VEGF-C and VEGFR-3 levels were significantly higher in PTC group than in BT group (P<0.05), but VEGF-D level was comparable between them (P>0.05). In PTC patients, the elevation of serum VEGF-C and VEGFR-3 levels was associated with an advanced clinical stage (III-IV), elevated thyroid-stimulating hormone (TSH) level, an age over 45 years, and a tumor diameter exceeding 2 cm (P<0.05 or P<0.01). Patients with lymph node metastasis had significantly higher VEGF-C level but lower VEGF-3 level than those without metastasis regardless of gender. Serum VEGF-D level was higher in PTC patients with lymph node metastasis (P<0.05) and elevated TSH level (P<0.01) without association with the clinical stage, tumor diameter, age, or gender. The area under ROC curve (AUC) of serum VEGF-C, VEGFR-3 and TSH was 0.803, 0.734 and 0.707 respectively (P<0.01), and that of VEGF-D was 0.556 (P>0.05); when combined, serum VEGF-C, VEGFR-3 and TSH showed an AUC of 0.862 (P<0.01)., Conclusion: Detecting serum VEGF-C and VEGFR-3 levels combined with TSH may enhance the early diagnosis rate of papillary thyroid carcinoma.

Background: A comprehensive characterization of the effects of cigarette smoke on systemic soluble immune/inflammatory markers may provide insight into the mechanisms through which smoking causes disease., Methods: Levels of 78 inflammation, immune, and metabolic markers were measured using multiplex immune assays in 1819 Prostate, Lung, Colorectal and Ovarian Cancer Screening Trial (PLCO) participants aged 55 to 74 years from three existing nested case-control studies. These data were made representative of the entire PLCO screening arm through reweighting with weights estimated in logistic regression models. We assessed associations between smoking status, cigarettes smoked per day, and time since quitting with dichotomized marker levels using adjusted weighted logistic regression models., Results: Current smoking was associated with 10 inflammation markers after correcting for multiple testing, encompassing several components of the immune/inflammation response. Levels of seven of these markers (interleukin [IL]-15, IL-1RA, IL-1β, IL-16, stem cell factor, soluble interleukin 6 receptor, and soluble vascular endothelial growth factor receptor 3) were lower among current smokers (n = 414) when compared with never smokers (n = 548), with odds ratios (ORs) ranging from 0.44 to 0.27, while levels of CC motif ligand (CCL)/thymus and activation regulated chemokine (CCL17/TARC) (OR = 4.08, 95% confidence interval [CI] = 2.01 to 8.25), CCL11/EOTAXIN (OR = 2.57, 95% CI = 1.45 to 4.55), and C-reactive protein (CRP) (OR = 2.54, 95% CI = 1.29 to 4.98) were elevated. These markers were not associated with cigarettes per day among current smokers, but there were trends in IL-15, IL-1RA, IL-1β, CCL17/TARC, CCL11/EOTAXIN, and CRP levels across categories of years since quitting smoking., Conclusions: Smoking is associated with a broad range of alterations in systemic immune and inflammation marker levels among older, long-term smokers. Smoking cessation may result in marker levels reverting back to those of never smokers over time., (Published by Oxford University Press 2014.)

Background: The aim of this study was the evaluation of pharmacokinetic parameters, biomarkers, clinical outcome, and imaging parameters in metastatic colorectal cancer (mCRC) patients treated with FOLFIRI plus sunitinib., Methods: mCRC patients with liver metastases were treated with FOLFIRI and sunitinib as 1st line therapy. At protocol-defined time points, multicontrast magnetic resonance imaging (MRI)measurements, computed tomography (CT) scans, pharmacokinetics (PK), and biomarker analyses were performed during the first and second treatment cycle. Thereafter, patients were treated until tumor progression, investigator’s decision due to toxicity, or patient withdrawal., Results: 28 patients were screened, 26 were included, and 23 received at least one study medication. Full safety analysis was performed in 23 patients. Full PK and biomarker analyses were performed in 21 patients. Strong responses in tumor size reduction forced a change from the original imaging timing scheme. This unforeseen change in the timing scheme resulted in subgroups too small for meaningful statistical analysis of most imaging parameters. Thus, only a descriptive analysis of the MRI data was possible. In 21/22 patients, MRI showeda decrease of the liver metastases. Best response was partial remission (PR) in 8/17 patients. Plasma concentrations of sVEGFR-2 and sVEGFR-3 decreased in all patients. The majority of the patients developed some kind of toxicity not always deducible to FOLFIRI or sunitinib., Conclusions: Due to the observed side effect profile, FOLFIRI plus sunitinib 37.5 mg per day cannot be recommended for previously untreated mCRC.

Background/aims: Despite the presence of many diagnostic methods, the differential diagnosis between benign and malignant biliary obstructions is still not easy. We aimed to evaluate the role of serum/biliary carcinoembryonic antigen (CEA), carbohydrate antigen 19-9 (CA 19-9), vascular endothelial growth factor receptor-3(VEGFR-3), and total antioxidant capacity (TAC) tests in this differential diagnosis., Materials and Methods: Patients (n:225; 110♂, 115♀) with diagnosis of malignant (n:96) or benign (n:129) biliary obstruction were included in this cross-sectional study. Serum and biliary CEA, CA 19-9, VEGFR-3, and TAC tests were analyzed, statistics were obtained, and significance was defined as p<0.05., Results: Mean age was 54.9±16.4 for the benign and 54.2±19.6 for the malignant group (p=0.89). Head of pancreas cancer (18.2%), cholangiocarcinoma (11.4%) and choledochal stone (48%) were the most common etiologies. The area under the curve (AUC)s by ROC analysis of serum/biliary CA 19-9, VEGFR-3, and TAC and serum CEA were 0.701/0.616, 0.622/0.663, 0.602/0.581, and 0713, respectively. Serum TAC had higher sensitivity (61.1%) and CEA had lower sensitivity (42.7%), whereas CEA had higher specificity (89.9%) and TAC had lower specificity (60.5%). In biliary tumor markers, CA 19-9 had higher sensitivity (74%) and VEGFR-3 had lower sensitivity (56.2%); however, VEGFR-3 had higher specificity (79.1%) and CA 19-9 had lower specificity (34.1%). Additionally, combination of serum CEA (p<0.001), CA 19-9 (p<0.001), VEGFR-3 (p<0.001), and biliary CA 19-9 (p=0.028) markers achieved 95% estimation probability, and the sensitivity, specificity, and accuracy were 88.5%, 45.7%, and 64%, respectively., Conclusion: Serum and biliary CEA, CA 19-9, VEGFR-3, and TAC tests would not be useful in the differentiation between malignant and benign biliary obstructions.

Purpose: We investigated potential biomarkers of efficacy in a phase III trial of sunitinib versus interferon-alpha (IFN-α), first-line in metastatic renal cell carcinoma (mRCC), by analyzing plasma levels of vascular endothelial growth factor (VEGF)-A, VEGF-C, soluble VEGF receptor-3 (sVEGFR-3) and interleukin (IL)-8., Methods: Seven hundred and fifty mRCC patients were randomized to oral sunitinib 50 mg/day in repeated cycles of a 4-week on/2-week off schedule or IFN-α 9 million units subcutaneously thrice weekly. Plasma samples collected from a subset of 63 patients on days 1 and 28 of cycles 1-4 and at end of treatment were analyzed by ELISA., Results: Baseline characteristics of biomarker-evaluated patients in sunitinib (N = 33) and IFN-α (N = 30) arms were comparable to their respective intent-to-treat populations. By univariate Cox regression analysis, low baseline soluble protein levels were associated with lower risk of progression/death (all P < 0.05): in both treatment arms, baseline VEGF-A and IL-8 were associated with overall survival (OS) and baseline VEGF-C with progression-free survival (PFS); in the sunitinib arm, baseline VEGF-A was associated with PFS and baseline sVEGFR-3 with PFS and OS; in the IFN-α arm, baseline IL-8 was associated with PFS. In multivariate analysis, baseline sVEGFR-3 and IL-8 remained independent predictors of OS in the sunitinib arm, while no independent predictors of outcome remained in the IFN-α arm. Pharmacodynamic changes were not associated with PFS or OS for any plasma protein investigated., Conclusions: Our findings suggest that, in mRCC, baseline VEGF-A and IL-8 may have prognostic value, while baseline sVEGFR-3 may predict sunitinib efficacy.

Background: The present study investigated whether serum levels of soluble vascular endothelial growth factor receptor (sVEGFR)-1, -2 and -3 are related to poor coronary collateralization in patients with stable coronary artery disease (CAD)., Methods and Results: Serum levels of sVEGFR-1, -2, -3, VEGF, and placental growth factor (PLGF) were determined in 403 consecutive patients with angiographic total or subtotal occlusion of at least 1 major coronary artery. The degree of collateralization was graded according to the Rentrop scoring system. Low (Rentrop score of 0 or 1) and high (Rentrop score of 2 or 3) coronary collateralization occurred in 161 and 242 patients, respectively. Serum levels of sVEGFR-1 and -2 were significantly elevated, in contrast, VEGF and PLGF levels were remarkably decreased in patients with low collateralization than in those with high collateralization (all P<0.05). Significant differences in sVEGFR-1, VEGF and PLGF levels was consistently detected between the low and high collateralization subgroups for patients with and without type 2 diabetes mellitus (DM) (for all comparisons, P<0.01). Multivariable regression analysis revealed that DM, dyslipidemia, elevated sVEGFR-1, and reduced VEGF and PLGF in serum were independently associated with a low degree of coronary collateralization., Conclusions: Increased serum sVEGFR-1 level is associated with poor coronary collateralization in patients with stable CAD. Type 2 DM is a predominant factor affecting collateral growth in these patients.

Aim: Lymph node metastasis is one of the predictive factors associated with poor prognosis of epithelial ovarian cancer. To clarify the role of CD34 and vascular endothelial growth factor receptor-3-positive (CD34+/VEGFR3+) lymphatic/vascular endothelial progenitor cells (LVEPC) in patients with lymph node metastasis and epithelial ovarian cancer progression, the levels of circulating CD34+/VEGFR3+ LVEPC in epithelial ovarian cancer patients were detected. We also tested the plasma protein levels of VEGF and stromal cell-derived factor to find out their possible relationships with lymph node metastasis in our epithelial ovarian cancer cohort., Material and Methods: Peripheral blood samples were collected from 54 patients diagnosed as epithelial ovarian cancer, and 31 normal samples as control. The circulating levels of LVEPC were carried out by flow cytometry, and blood protein levels of biomarkers were measured by enzyme-linked immunosorbent assay (ELISA)., Results: The level of circulating LVEPC was significantly higher in patients with ovarian cancer compared with that of healthy controls. There was also a statistically significant correlation between LVEPC levels and surgical staging of epithelial ovarian cancer (P < 0.01)., Conclusion: The circulating levels of bone marrow-derived LVEPC are significantly increased in epithelial ovarian cancer patients and these levels correlate with lymph node metastasis too., (© 2013 The Authors. Journal of Obstetrics and Gynaecology Research © 2013 Japan Society of Obstetrics and Gynecology.)

Background: Metronomic chemotherapy (MCT) with cyclophosphamide (Cy) and celecoxib (Cel) has therapeutic efficacy and low toxicity profile in advanced breast cancer patients (ABCP), but no reliable biomarkers of response have been found yet that allow patient selection for treatment., Aim: To investigate the potential role as biomarkers of pro- and antiangiogenic parameters and evaluate their response in ABCP receiving metronomic Cy 50 mg p.o./day + Cel 400 mg p.o./day., Materials and Methods: Serum levels of vascular endothelial growth factor-C (VEGF-C), soluble VEGF receptors 2 and 3 (sVEGFR-2, sVEGFR-3), were measured at different time points in 13/15 patients included in a phase II trial of MCT with Cy+Cel., Results: Serum levels of sVEGFR-2 and sVEGFR-3 increased significantly during treatment (P = 0.0392; P = 0.0066, respectively). VEGF-C showed no significant modifications. Previous determinations of VEGF and TSP-1 in the same patients were utilized. VEGF/sVEGFR-2, VEGF/TSP-1, and VEGF-C/sVEGFR-3 ratios decreased significantly along the treatment (P = 0.0092; P = 0.0072; P = 0.0141, respectively). Nonsignificant variations were observed for VEGF-C/sVEGFR-2 ratio. Baseline values of VEGF/sVEGFR-2 and VEGF/TSP-1 ratios were associated with time to progression (TTP) (P = 0.0407; P = 0.0394, respectively) meanwhile baseline VEGF was marginally significant (P = 0.0716). Patients with values lower than the 50th percentile for both ratios showed longer TTP., Conclusions: We have identified the baseline VEGF/sVEGFR-2 and VEGF/TSP-1 ratios as potential biomarkers of response in ABCP treated metronomically with Cy+Cel. This finding warrants its confirmation in a higher number of patients.

Abstract Antiangiogenic and antilymphangiogenic gene therapy with soluble vascular endothelial growth factor receptor-2 (VEGFR-2) and soluble VEGFR-3 in combination with chemotherapy is a potential new treatment for ovarian carcinoma. We evaluated the safety, toxicology, and biodistribution of intravenous AdsVEGFR-2 and AdsVEGFR-3 combined with chemotherapy in healthy rats (n=90) before entering a clinical setting. The study groups were: AdLacZ and AdLacZ with chemotherapy as control groups, low dose AdsVEGFR-2 and AdsVEGFR-3, high dose AdsVEGFR-2 and AdsVEGFR-3, combination of low dose AdsVEGFR-2 and AdsVEGFR-3 with chemotherapy, combination of high dose AdsVEGFR-2 and AdVEGFR-3 with chemotherapy, and chemotherapy only. The follow-up time was 4 weeks. Safety and toxicology were assessed by monitoring the clinical status of the animals and by histological, hematological, and clinical chemistry parameters. For the biodistribution studies, quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR) and enzyme-linked immunosorbent assay (ELISA) were used. Low dose (2×10(10) vp) AdsVEGFR-2 and AdsVEGFR-3 gene therapy was well tolerated, even when gene therapy was combined with chemotherapy. Notably, only transient elevation of liver enzymes and mild regenerative changes were seen in liver after the gene transfer in the groups that received high doses (2×10(11) vp) of AdsVEGFR-2 and AdsVEGFR-3 with or without chemotherapy. No life-threatening adverse effects were noticed in any of the treatment groups. The highest protein concentration of soluble VEGFR-2 (sVEGFR-2) in circulation was seen 1 week after the gene transfer. The combination of chemotherapy to gene therapy seemed to prolong the time of detectable transgene protein at least 1 week in the circulation. The expression of AdsVEGFR-2 and AdsVEGFR-3 transgenes was mainly seen in the liver and spleen as detected by qRT-PCR. According to these results, AdsVEGFR-2 and AdsVEGFR-3 gene therapy combined with chemotherapy is safe and can be brought to clinical testing in ovarian cancer patients.

Soluble VEGFR-3 (sVEGFR-3) is a potential biomarker for the anti-angiogenic activity of tyrosine kinase inhibitors. The aim of this investigation was the validation of an enzyme-linked immunosorbent assay (ELISA) to measure sVEGFR-3 in human plasma and the investigation of its applicability in clinical trials as first step of the biomarker validation process. General validation criteria were assessed based on current guidelines and recommendations for immunoassays. The ELISA was applied in two clinical trials including healthy volunteers and metastatic colorectal cancer (mCRC) patients receiving 50 or 37.5mg sunitinib per day, respectively. SVEGFR-3 was measured at predefined time points. Undiluted, inactivated fetal calf serum was identified as surrogate matrix to substitute for human plasma. Dilutional linearity and parallelism could be successfully confirmed. The analyte was measured in the study matrix with intra- and inter-run precision and accuracy ≤20%. Stability was proven over a period of at least 15 months as well as upon three freeze-thaw cycles. SVEGFR-3 concentrations decreased in response to sunitinib to 57% (IQR 50-88%) and 58% (IQR 47-80%) of the respective baseline concentrations in healthy volunteers and mCRC patients, respectively, with subsequent increase after stop of treatment. The ELISA for the quantification of sVEGFR-3 in human plasma was successfully validated. The applicability of the assay was demonstrated in two clinical trials., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Background: The aim of this study was to determine whether improvement of filarial lymphedema (LE) by doxycycline is restricted to patients with ongoing infection (positive for circulating filarial antigen [CFA]), or whether the majority of CFA-negative patients with LE would also show a reduction in LE severity., Methods: One hundred sixty-two Ghanaian participants with LE stage 1-5 (Dreyer) were randomized blockwise into 2 groups (CFA positive or negative) and allocated to 3 treatment arms of 6 weeks: (1) amoxicillin (1000 mg/d), (2) doxycycline (200 mg/d), or (3) placebo matching doxycycline. All groups received standard hygiene morbidity management. The primary outcome was reduction of LE stages. Secondary outcomes included frequency of acute attacks and ultrasonographic assessment of skin thickness at the ankles. Parameters were assessed before treatment and after 3, 12, and 24 months., Results: Doxycycline-treated patients with LE stage 2-3 showed significant reductions in LE severity after 12 and 24 months, regardless of CFA status. Improvement was observed in 43.9% of doxycycline-treated patients, compared with only 3.2% and 5.6% in the amoxicillin and placebo arms, respectively. Skin thickness was correlated with LE stage improvement. Both doxycycline and amoxicillin were able to reduce acute dermatolymphangioadenitis attacks., Conclusions: Doxycycline treatment improves mild to moderate LE independent of ongoing infection. This finding expands the benefits of doxycycline to the entire population of patients suffering from LE. Patients with LE stage 1-3 should benefit from a 6-week course of doxycycline every other year or yearly, which should be considered as an improved tool to manage morbidity in filarial LE. Clinical Trials Registration. ISRCTN 90861344.

Background: Axitinib is an oral, potent and selective inhibitor of vascular endothelial growth factor receptors (VEGFRs) 1, 2 and 3. We report on data obtained from 18 Japanese patients with advanced solid tumors in two phase I trials that evaluated the safety, pharmacokinetics and antitumor activity of axitinib and also examined potential biomarkers., Methods: Six patients received a single 5-mg dose of axitinib followed by 5 mg twice daily (BID), and an additional six patients received axitinib 5 mg BID only. Another six patients received axitinib at 5-mg, 7-mg and 10-mg single doses followed by 5 mg BID., Results: Plasma pharmacokinetics following single doses of axitinib was generally linear. Common treatment-related adverse events were fatigue (83%), anorexia (72%), diarrhea (67%), hand-foot syndrome (67%) and hypertension (61%). Sixteen patients (89%) experienced thyroid-stimulating hormone (TSH) elevation. Grade 3/4 toxicities included hypertension (33%) and fatigue (28%). No grade 3/4 fatigue occurred in patients who started thyroid hormone replacement therapy when TSH was elevated. Thyroglobulin elevation was observed in all patients who continued treatment with axitinib for ≥3 months. Abnormal TSH correlated with exposure to axitinib (r = 0.72). Decrease in soluble (s) VEGFR-2 levels significantly correlated with exposure to axitinib (r = -0.94). Axitinib showed antitumor activity across multiple tumor types., Conclusions: Axitinib-related thyroid dysfunction could be due to a direct effect on the thyroid gland. Grade 3/4 fatigue and hypothyroidism appear to be controllable with use of thyroid hormone replacement therapy. sVEGFR-2 and TSH may act as biomarkers of axitinib plasma exposure.

Background: Ovarian hyperstimulation syndrome (OHSS) is a potentially life-threatening condition associated with increased vascular permeability. The vascular endothelial growth factor (VEGF) system and its receptors have been identified as the main angiogenic factors responsible for increased capillary permeability and are therefore discussed as crucial for the occurrence of OHSS. Recently, a number of soluble receptors for the VEGFs have been detected (sVEGF-Rs) and it has been shown that these sVEGF-Rs compete with the membrane-standing VEGF-R to bind VEGFs., Methods: We analyzed the serum levels of soluble VEGF-R1, -R2 and -R3 in 34 patients suffering from OHSS and in 34 controls without this disease. In a subgroup analysis, we correlated the severity of the OHSS with the detected amounts of VEGF-R1, -R2 and -R3. In addition, we determined the amount of total VEGF-A in the samples., Results: All the three soluble VEGF receptors tended to be higher in the control group compared with that in the OHSS group but this difference only reached significance for sVEGF-R2 (mean ± SEM: 15.5 ± 0.6 versus 13.8 ± 0.5 ng/ml, respectively, P< 0.05). In the subgroup analysis, sVEGF-R2 levels decreased as the severity of OHSS increased (OHSS-I: 16.8 ± 1.9 ng/ml and OHSS-III: 12.7 ± 1.0 ng/ml, P< 0.05) Moreover, the serum levels of total VEGF-A were higher in the OHSS group than those in the controls (537.7 ± 38.9 versus 351 ± 53.4 pg/ml, respectively P< 0.05)., Conclusions: We propose that VEGF-A plays a role in the occurrence of OHSS, that the amount of biologically available VEGF-A is modulated by sVEGF-Rs and that different combinations of VEGF-A and sVEGF-R levels might contribute to the severity of OHSS.

The objective of this study was to detect the expression levels of VEGF-C, VEGF-D, VEGFR-2 and VEGFR-3 in plasma of newly diagnosed lymphoma patients, and analyze their possible relationships with clinicopathological characteristics and prognosis. The expression levels of VEGF-C, VEGF-D, VEGFR-2 and VEGFR-3 in plasma from 86 newly diagnosed lymphoma patients were detected by enzyme-linked immunosorbent assay (ELISA). As a results, the multivariate analysis showed that VEGF-C level in non-Hodgkin's lymphoma patients was low, but high in Hodgkin's lymphoma patients; VEGFR-2 level was higher in patients > 60 years, while VEGF-D level was lower in patients with IPI > 2. The univariate analysis showed that VEGF-D level was lower in patients with IPI > 2, while VEGF-D and VEGF-C levels were higher in patients without B symptoms. Relationship analysis between these factors indicated that the relation of VEGF-D expression level with VEGFR-2 and VEGFR-3 was positive. It is concluded that VEGF-C, VEGF-D, VEGFR-2 and VEGFR-3 play important roles in the pathogenesis of lymphoma, and may be used as indicators of prognosis evaluation or even guide for the antiangiogenesis treatment of lymphoma.

Aim: To explore if vascular endothelial growth factor receptor-3 (VEGFR-3) and carcinoembryonic antigen (CEA) can predict overall survival in advanced gastric cancer., Methods: VEGFR-3 level was assessed by enzyme-linked immunosorbent assay, and CEA was assessed by chemiluminescence immunoassay in the sera of 81 advanced gastric cancer patients before treatment with oxaliplatin plus 5-fluorouracil and folinic acid., Results: Median survival time in patients with a low serum VEGFR-3 level was significantly longer than in those with a higher VEGFR-3 level (15.4 mo vs 7.7 mo, P < 0.001). Patients with a low CEA level had a longer survival than those with a higher CEA level (15.8 mo vs 8.6 mo, P < 0.001). Thirty-nine patients with low VEGFR-3 and low CEA levels had a median survival of 19.7 mo (P = 0.0006). The hazard ratio for patients with a high VEGFR-3 level was 2.443 (P = 0.002)., Conclusion: High serum VEGFR-3 level is correlated significantly with poor survival. In patients with a high serum level of VEGFR-3, alternative chemotherapy regimens should be considered.

Aims: To assess the antitumour activity, safety, pharmacokinetics and pharmacodynamics of continuous daily sunitinib dosing in patients with imatinib-resistant/intolerant gastrointestinal stromal tumour (GIST) and to assess morning dosing versus evening dosing., Patients and Methods: In this open-label phase II study, patients were randomised to receive morning or evening dosing of sunitinib 37.5mg/day. The primary end-point was clinical benefit rate (CBR; percent complete responses+partial responses [PRs]+stable disease [SD] 24 weeks). Secondary end-points included progression-free survival (PFS), overall survival (OS), safety, pharmacokinetic parameters and plasma biomarker levels., Results: Sixty of 61 planned patients received treatment (30 per dosing group); 26 completed the study. Overall, the CBR was 53% (95% exact CI, 40-66): eight patients (13%) achieved objective PRs; 24 (40%) achieved SD 24 weeks. Median PFS was 34 weeks (95% CI, 24-49); median OS was 107 weeks (95% CI, 72 - not yet calculable). Most adverse events (AEs) were of grade 1 or 2 in severity, and were manageable through dose modification or standard interventions. No new AEs were apparent compared with the approved intermittent dosing schedule. Antitumour activity and safety were generally similar with morning and evening dosing. Continuous daily sunitinib dosing achieved and sustained effective drug concentrations without additional accumulation across cycles. Decreases from baseline in plasma levels of soluble KIT after 20 and 24 weeks of dosing correlated with longer OS., Conclusion: For patients with imatinib-resistant/intolerant GIST, continuous daily sunitinib dosing appears to be an active alternative dosing strategy with acceptable safety.

Purpose: The presence of metastases in regional lymph nodes is a strong indicator of poor patient survival in many types of cancer. It has recently been shown that vascular endothelial growth factor-C (VEGF-C), and its receptor VEGFR-3, may play a pivotal role in the promotion of metastasis to regional lymph nodes. This study was designed to detect and evaluate whether the expression of VEGFR-3 or its soluble form plays a role in metastatic malignant melanoma and to determine the relationship with clinicopathological parameters and patients outcome., Experimental Design: VEGFR-3 expression on melanoma tumour was evaluated by immunohistochemical study. Using a sensitive enzyme-linked immunosorbent assay, sVEGFR-3 was measured in sera of 60 metastatic melanoma patients in comparison with 30 healthy controls., Results: Immunohistochemical study demonstrated a high expression of VEGFR-3 in melanoma cells. Median level of pre-treatment sVEGFR-3 was significantly higher (p=0.00001) in melanoma patients as compared to healthy donors. No association was noted between VEGFR-3 in situ or in sera and gender, age or LDH level. Median serum VEGFR-3 levels were significantly higher in patients with high tumour burden as compared to those with low tumour burden (p=0.013) as well as in non-responding patients (n=33) as compared to responding ones (n=27). Finally, low level of VEGFR-3 was also related positively to disease free survival (X(2)=3.85, p=0.022)., Conclusion: These results suggest that the expression and high pre-treatment sVEGFR-3 level are significantly correlated to poorer prognosis, and may be promising targets for new therapeutic strategies in melanoma disease.

Purpose: The newly identified bone marrow-derived cell population, called lymphatic/vascular endothelial progenitor cells (LVEPC), has been shown to contribute to lymph capillary growth in experimental tumor systems. The clinical significance of these cells has not yet been investigated in a human malignancy. Our aim was to study whether peripheral blood circulating LVEPCs participate in the progression of human small cell lung cancer (SCLC)., Experimental Design: A total of 88 patients with limited-stage SCLC and 32 tumor-free control subjects were included. Peripheral blood circulating LVEPC labeled with CD34 and vascular endothelial growth factor receptor-3 (VEGFR3) antibodies and the serum levels of the key lymphangiogenic molecule VEGF-C were measured by flow cytometry and ELISA, respectively., Results: CD34-positive/VEGFR3-positive LVEPC levels were significantly increased in patients (versus controls; P<0.01), and there was also a significant relationship between LVEPC counts and lymph node metastasis (P<0.01). High pretreatment circulating LVEPC numbers correlated with poor overall survival (P<0.01). Although we observed significantly elevated VEGF-C concentrations in patients (versus controls; P<0.01), there was no significant correlation between VEGF-C and LVEPC levels. Moreover, no significant differences in peripheral blood VEGF-C levels were seen between patients subgrouped by clinicopathologic variables including tumor and lymph node stages and survival., Conclusions: Peripheral blood levels of bone marrow-derived LVEPCs are significantly increased in patients with SCLC and correlate with lymphatic involvement and prognosis. This is the first study that shows evidence of increased numbers of circulating LVEPC in patients with a malignant tumor.

Purpose: To assess the safety and efficacy of sunitinib in patients with bevacizumab-refractory metastatic renal cell carcinoma (mRCC) and explore biomarkers for sunitinib response., Patients and Methods: Patients with mRCC and disease progression after bevacizumab-based therapy received oral sunitinib 50 mg once daily in 6-week cycles on a 4/2 schedule (4 weeks with treatment followed by 2 weeks without treatment) in a phase II multicenter study. The primary end point was objective response rate (ORR). Secondary end points included progression-free survival (PFS), duration of response (DR), overall survival (OS), and safety. Plasma soluble proteins (vascular endothelial growth factor [VEGF]-A, VEGF-C, soluble VEGF receptor [sVEGFR]-3, and placental growth factor [PlGF]) levels were measured., Results: Sixty-one patients were enrolled. The ORR was 23.0% (95% CI, 13.2% to 35.5%), median PFS was 30.4 weeks (95% CI, 18.3 to 36.7 weeks), median DR was 44.1 weeks (95% CI, 25.0 to 102.7 weeks), and median OS was 47.1 weeks (95% CI, 36.9 to 79.4 weeks). Mean plasma VEGF-A and PlGF levels significantly increased whereas VEGF-C and sVEGFR-3 levels decreased with sunitinib treatment. Lower baseline levels of sVEGFR-3 and VEGF-C were associated with longer PFS and ORR. Most treatment-related adverse events were of mild-to-moderate intensity and included fatigue, hypertension, and hand-foot syndrome., Conclusion: Sunitinib has substantial antitumor activity in patients with bevacizumab-refractory mRCC and modulates circulating VEGF pathway biomarkers. These data support the hypothesis that sunitinib inhibits signaling pathways involved in bevacizumab resistance. Baseline levels of sVEGFR-3 and VEGF-C may have potential utility as biomarkers of clinical efficacy in this setting.

Background: Sunitinib malate (SUTENT) is an oral, multitargeted tyrosine kinase inhibitor, approved multinationally for the treatment of advanced RCC and of imatinib-resistant or - intolerant GIST. The purpose of this study was to explore potential biomarkers of sunitinib pharmacological activity via serial assessment of plasma levels of four soluble proteins from patients in a phase II study of advanced RCC: VEGF, soluble VEGFR-2 (sVEGFR-2), placenta growth factor (PlGF), and a novel soluble variant of VEGFR-3 (sVEGFR-3)., Methods: Sunitinib was administered at 50 mg/day on a 4/2 schedule (4 weeks on treatment, 2 weeks off treatment) to 63 patients with metastatic RCC after failure of first-line cytokine therapy. Predose plasma samples were collected on days 1 and 28 of each cycle and analyzed via ELISA., Results: At the end of cycle 1, VEGF and PlGF levels increased >3-fold (relative to baseline) in 24/54 (44%) and 22/55 (40%) cases, respectively (P < 0.001). sVEGFR-2 levels decreased >or= 30% in 50/55 (91%) cases and >or= 20% in all cases (P < 0.001) during cycle 1, while sVEGFR-3 levels were decreased >or= 30% in 48 of 55 cases (87%), and >or= 20% in all but 2 cases. These levels tended to return to near-baseline after 2 weeks off treatment, indicating that these effects were dependent on drug exposure. Overall, significantly larger changes in VEGF, sVEGFR-2, and sVEGFR-3 levels were observed in patients exhibiting objective tumor response compared with those exhibiting stable disease or disease progression (P < 0.05 for each analyte; analysis not done for PlGF)., Conclusion: Sunitinib treatment in advanced RCC patients leads to modulation of plasma levels of circulating proteins involved in VEGF signaling, including soluble forms of two VEGF receptors. This panel of proteins may be of value as biomarkers of the pharmacological and clinical activity of sunitinib in RCC, and of angiogenic processes in cancer and other diseases.

Lymphatic filariasis is a disease of considerable socioeconomic burden in the tropics. Presently used antifilarial drugs are able to strongly reduce transmission and will thus ultimately lower the burden of morbidity associated with the infection, however, a chemotherapeutic principle that directly induces a halt or improvement in the progression of the morbidity in already infected individuals would constitute a major lead. In search of such a more-effective drug to complement the existing ones, in an area endemic for bancroftian filariasis in Ghana, 33 microfilaremic and 18 lymphedema patients took part in a double-blind, placebo-controlled trial of a 6-wk regimen of 200 mg/day doxycycline. Four months after doxycycline treatment, all patients received 150-200 microg/kg ivermectin and 400 mg albendazole. Patients were monitored for Wolbachia and microfilaria loads, antigenemia, filarial dance sign (FDS), dilation of supratesticular lymphatic vessels, and plasma levels of lymphangiogenic factors (vascular endothelial growth factor-C [VEGF-C] and soluble vascular endothelial growth factor receptor-3 [(s)VEGFR-3]). Lymphedema patients were additionally monitored for stage (grade) of lymphedema and the circumferences of affected legs. Wolbachia load, microfilaremia, antigenemia, and frequency of FDS were significantly reduced in microfilaremic patients up to 24 mo in the doxycycline group compared to the placebo group. The mean dilation of supratesticular lymphatic vessels in doxycycline-treated patients was reduced significantly at 24 mo, whereas there was no improvement in the placebo group. Preceding clinical improvement, at 12 mo, the mean plasma levels of VEGF-C and sVEGFR-3 decreased significantly in the doxycycline-treated patients to a level close to that of endemic normal values, whereas there was no significant reduction in the placebo patients. The extent of disease in lymphedema patients significantly improved following doxycycline, with the mean stage of lymphedema in the doxycycline-treated patients being significantly lower compared to placebo patients 12 mo after treatment. The reduction in the stages manifested as better skin texture, a reduction of deep folds, and fewer deep skin folds. In conclusion, a 6-wk regimen of antifilarial treatment with doxycycline against W. bancrofti showed a strong macrofilaricidal activity and reduction in plasma levels of VEGF-C/sVEGFR-3, the latter being associated with amelioration of supratesticular dilated lymphatic vessels and with an improvement of pathology in lymphatic filariasis patients.

Lymphatic vessel plasticity and stability are of considerable importance when attempting to treat diseases associated with the lymphatic vasculature. Development of lymphatic vessels during embryogenesis is dependent on vascular endothelial growth factor (VEGF)-C but not VEGF-D. Using a recombinant adenovirus encoding a soluble form of their receptor VEGFR-3 (AdVEGFR-3-Ig), we studied lymphatic vessel dependency on VEGF-C and VEGF-D induced VEGFR-3 signaling in postnatal and adult mice. Transduction with AdVEGFR-3-Ig led to regression of lymphatic capillaries and medium-sized lymphatic vessels in mice under 2 weeks of age without affecting collecting lymphatic vessels or the blood vasculature. No effect was observed after this period. The lymphatic capillaries of neonatal mice also regressed partially in response to recombinant VEGFR-3-Ig or blocking antibodies against VEGFR-3, but not to adenovirus-encoded VEGFR-2-Ig. Despite sustained inhibitory VEGFR-3-Ig levels, lymphatic vessel regrowth was observed at 4 weeks of age. Interestingly, whereas transgenic expression of VEGF-C in the skin induced lymphatic hyperplasia even during embryogenesis, similar expression of VEGF-D resulted in lymphangiogenesis predominantly after birth. These results indicate considerable plasticity of lymphatic vessels during the early postnatal period but not thereafter, suggesting that anti-lymphangiogenic therapy can be safely applied in adults.

Aims: To investigate the immunocytochemical expression of vascular endothelial growth factor C (VEGF-C) and its receptors (VEGFR-2 and VEGFR-3) in childhood acute lymphoblastic leukaemia (ALL) blasts and to determine the possible role of this complex in the pathogenesis and prognosis of ALL., Methods and Results: Bone marrow samples were taken from 120 children diagnosed with ALL. An indirect immunocytochemical procedure was performed with the use of monoclonal mouse anti-human antibodies against VEGF-C, VEGFR-2 and VEGFR-3 (diluted 1 : 100). The immunocytochemical expression of VEGF-C was confirmed exclusively in the cytoplasm of ALL lymphoblasts (the mean percentage was 36.4 +/- 7.2). It was absent from the cytoplasm of normal haematopoietic cells in the control group. No VEGFR-2 or VEGFR-3 expression was detected in the children of either the study or control groups. The risk of induction failure or leukaemic relapse was found to be significant in all VEGF-C+ patients (P < 0.0001 and P < 0.02, respectively; Fisher's exact test)., Conclusions: The absence of VEGF-C in blast cells predicts long-lasting remission in all leukaemic children. Our findings also suggest that leukaemic cell invasion, following VEGF-C-driven lymphangiogenesis, could be related to a mediating role of this peptide produced by blast cells themselves.

The presence of metastases in regional lymph nodes is a strong indicator of poor patient survival in many types of cancer. It has recently been shown that the lymphangiogenic growth factor, vascular endothelial growth factor-C (VEGF-C), and its receptor, VEGF receptor-3 (VEGFR3), may play a pivotal role in the promotion of metastasis to regional lymph nodes. In this study, human prostate and melanoma tumor models that preferentially metastasize to the lymph nodes following s.c. tumor cell implantation were established from lymph node metastases via in vivo selection. Melanoma tumor cell sublines established from lymph node metastasis express higher amounts of VEGF-C than the parental tumor cells. The inhibition of tumor-derived VEGF-C with a soluble VEGFR3 decoy receptor, sVEGFR3-Fc, expressed via a recombinant adeno-associated viral vector, potently blocks tumor-associated lymphangiogenesis and tumor metastasis to the lymph nodes, when the treatment was initiated before the tumor implantation. In addition, sVEGFR3-Fc serum levels required for efficient blockade of lymph node metastases are strictly dependent on the VEGF-C levels generated by the primary tumor. Recombinant adeno-associated virus-mediated gene transfer of sVEGFR3-Fc may represent a feasible therapeutic strategy for blockade of lymphogenous metastasis.