Prognostic significance of VEGF and components of the plasminogen activator system in endometrial cancer.

OBJECTIVE: The plasminogen activator system (PAS) and vascular endothelial growth factor (VEGF) are important in the carcinogenesis and play a key role in cancer invasion and mediating metastasis of carcinomas. The aim of the study was to evaluate the correlation of serum levels of VEGF and components of the PAS with clinicopathological risk factors and outcome in patients with endometrial cancer (EC). METHODS: Preoperative blood was collected from 173 patients treated for EC between 1999 and 2009. Serum concentrations of VEGF, urokinase plasminogen activator (uPA) tissue plasminogen activator (tPA), plasminogen activator inhibitor type-1 (PAI-1) and -2 (PAI-2) were assessed by enzyme-linked immunosorbent assays (ELISA). RESULTS: Serum levels of VEGF and components of the PAS were significantly associated with stage of the disease, tumor histology, tumor grade, myometrial invasion (MI), presence of lymphovascular space invasion (LVSI) and lymph node metastases (LNM). Preoperative serum levels of PAI-1 and -2 and tPA were higher in patients who experienced a recurrence than in patients who remained disease free (p < 0.01). PAI-1 and -2 and tPA were significantly independent prognostic factors for DFS with a HR of 3.85 (95% CI 1.84-8.07), 3.90 (95% CI 1.75-8.66) and 2.53 (95% CI 1.16-5.55), respectively. PAI-1 and tPA turned out to be independent prognostic factors for OS, with a HR of 2.09 (95% CI 1.08-4.05) and 2.16 (95% CI 1.06-4.44), respectively. CONCLUSION: Serum levels of VEGF and components of the PAS at primary diagnosis were associated with well-known clinicopathological risk factors such as; FIGO stage, tumor histology, tumor grade, MI, LVSI and LNM. High concentrations of PAI-1 and-2 and tPA are independent factors for poor prognosis in patients with endometrial cancer.

Malaria early in the first pregnancy: Potential impact of iron status.

BACKGROUND & AIMS: Low iron stores may protect from malaria infection, therefore improving iron stores in early pregnancy in line with current recommendations could increase malaria susceptibility. To test this hypothesis we compared iron biomarkers and red cell indices in nulliparae and primigravidae who participated in a randomized controlled trial of long-term weekly iron supplementation. METHODS: Cross-sectional and longitudinal data analysis from a randomized controlled trial of long-term weekly iron supplementation in rural Burkina Faso. Malaria parasitaemia was monitored and biomarkers and red cell indices measured at study end-points: plasma ferritin, transferrin receptor (sTfR), zinc protoporphyrin, hepcidin, sTfR/log10 ferritin ratio, body iron, haemoglobin, red cell distribution width; mean corpuscular haemoglobin concentration/volume, and C-reactive protein. Correlation coefficients between biomarkers and red cell indices were determined. A regression correction approach based on ferritin was used to estimate iron body stores, allowing for inflammation. Body iron differences were compared between nulliparae and primigravidae, and the association determined of iron biomarkers and body iron stores with malaria. RESULTS: Iron and haematological indices of 972 nulliparae (mean age 16.5 years) and 314 primigravidae (median gestation 18 weeks) were available. Malaria prevalence was 54.0% in primigravidae and 41.8% in nulliparae (relative risk 1.28, 95% CI 1.13-1.45, P < 0.001), anaemia prevalence 69.7% and 43.4% (P < 0.001), and iron deficient erythropoiesis (low body iron) 8.0% and 11.7% (P = 0.088) respectively. Unlike other biomarkers the sTfR/log10 ferritin ratio showed no correlation with inflammation as measured by CRP. Most biomarkers indicated reduced iron deficiency in early pregnancy, with the exception of haemoglobin. Body iron increased by 0.6-1.2 mg/kg in early gestation, did not differ by malaria status in nulliparae, but was higher in primigravidae with malaria (6.5 mg/kg versus 5.0 mg/kg; relative risk 1.53, 95% CI 0.67-2.38, P < 0.001). CONCLUSION: In primigravidae, early pregnancy haemoglobin was not a good indicator of requirement for iron supplementation, which could be detrimental given the association of better iron status with increased malaria infection. TRIAL REGISTRATION: clinicaltrials.gov:NCT01210040. Until placed in a public repository, data relating to the current study can be requested from the corresponding author and will be made available following an end user data agreement and sponsor approval.

Provisional standardization of hepcidin assays: creating a traceability chain with a primary reference material, candidate reference method and a commutable secondary reference material.

Background Hepcidin concentrations measured by various methods differ considerably, complicating interpretation. Here, a previously identified plasma-based candidate secondary reference material (csRM) was modified into a serum-based two-leveled sRM. We validated its functionality to increase the equivalence between methods for international standardization. Methods We applied technical procedures developed by the International Consortium for Harmonization of Clinical Laboratory Results. The sRM, consisting of lyophilized serum with cryolyoprotectant, appeared commutable among nine different measurement procedures using 16 native human serum samples in a first round robin (RR1). Harmonization potential of the sRM was simulated in RR1 and evaluated in practice in RR2 among 11 measurement procedures using three native human plasma samples. Comprehensive purity analysis of a candidate primary RM (cpRM) was performed by state of the art procedures. The sRM was value assigned with an isotope dilution mass spectrometry-based candidate reference method calibrated using the certified pRM. Results The inter-assay CV without harmonization was 42.1% and 52.8% in RR1 and RR2, respectively. In RR1, simulation of harmonization with sRM resulted in an inter-assay CV of 11.0%, whereas in RR2 calibration with the material resulted in an inter-assay CV of 19.1%. Both the sRM and pRM passed international homogeneity criteria and showed long-term stability. We assigned values to the low (0.95±0.11 nmol/L) and middle concentration (3.75±0.17 nmol/L) calibrators of the sRM. Conclusions Standardization of hepcidin is possible with our sRM, which value is assigned by a pRM. We propose the implementation of this material as an international calibrator for hepcidin.

The macrophage migration inhibitory factor pathway in human B cells is tightly controlled and dysregulated in multiple sclerosis.

In MS, B cells survive peripheral tolerance checkpoints to mediate local inflammation, but the underlying molecular mechanisms are relatively underexplored. In mice, the MIF pathway controls B-cell development and the induction of EAE. Here, we found that MIF and MIF receptor CD74 are downregulated, while MIF receptor CXCR4 is upregulated in B cells from early onset MS patients. B cells were identified as the main immune subset in blood expressing MIF. Blocking of MIF and CD74 signaling in B cells triggered CXCR4 expression, and vice versa, with separate effects on their proinflammatory activity, proliferation, and sensitivity to Fas-mediated apoptosis. This study reveals a new reciprocal negative regulation loop between CD74 and CXCR4 in human B cells. The disturbance of this loop during MS onset provides further insights into how pathogenic B cells survive peripheral tolerance checkpoints to mediate disease activity in MS.

Effects of Weekly Iron and Folic Acid Supplements on Malaria Risk in Nulliparous Women in Burkina Faso: A Periconceptional, Double-Blind, Randomized Controlled Noninferiority Trial.

Background: The safety of iron supplementation for young women is uncertain in malaria-endemic settings. Methods: This was a double-blind, randomized controlled noninferiority trial in rural Burkina Faso. Results: A total of 1959 nulliparae were assigned to weekly supplementation (60 mg iron and 2.8 mg folic acid) (n = 980) or 2.8 mg folic acid (n = 979) until first antenatal visit (ANC1), or 18 months if remaining nonpregnant. Three hundred fifteen women attended ANC1, and 916 remained nonpregnant. There was no difference at ANC1 in parasitemia prevalence (iron, 53.4% [95% confidence interval {CI}, 45.7%-61.0%]; control, 55.3% [95% CI, 47.3%-62.9%]; prevalence ratio, 0.97 [95% CI, .79-1.18]; P = .82), anemia (adjusted effect, 0.96 [95% CI, .83-1.10]; P = .52), iron deficiency (adjusted risk ratio [aRR], 0.84 [95% CI, .46-1.54]; P = .58), or plasma iron biomarkers. Outcomes in nonpregnant women were parasitemia (iron, 42.9% [95% CI, 38.3%-47.5%]; control, 39.2% [95% CI, 34.9%-43.7%]; prevalence ratio, 1.09 [95% CI, .93-1.28]; P = .282); anemia (aRR, 0.90 [95% CI, .78-1.05]; P = .17), and iron deficiency (aRR, 0.99 [95% CI, .77-1.28]; P = .96), with no iron biomarker differences. Conclusions: Weekly iron supplementation did not increase malaria risk, improve iron status, or reduce anemia in young, mostly adolescent menstruating women, nor in early pregnancy. World Health Organization Guidelines for universal supplementation for young nulliparous women may need reassessment. Clinical Trials Registration: NCT01210040.

HE4 is superior to CA125 in the detection of recurrent disease in high-risk endometrial cancer patients.

OBJECTIVE: To date, biomarkers are not routinely used in endometrial cancer diagnosis, prognosis, and follow-up. The purpose of this study was to evaluate whether serum HE4 was related to clinicopathological risk factors and outcome. Second, the role of serum HE4 and CA125 was assessed as indicator for recurrent disease during follow-up. METHODS: A total of 174 patients with endometrial cancer between 1999 and 2009 were selected for this retrospective study. Serum HE4 and CA125 were analyzed at primary diagnosis, during follow-up, and at the time of recurrence. Correlations with clinicopathological factors were studied by univariate and multivariate survival analyses. Lead time was calculated in order to determine which serum marker was elevated prior to clinical detection of recurrent disease. RESULTS: Serum levels of HE4 and CA125 were significantly associated with high tumor grade, myometrial invasion, lymph node involvement, and advanced stage (p < 0.01). HE4 was an independent prognostic factor for reduced disease-free survival and overall survival with hazard ratios of 2.96 (95% confidence interval: 1.18-7.99) and 3.27 (95% confidence interval: 1.18-9.02), respectively. At recurrence, 75% of the patients had an elevated HE4 compared to 54% with an elevated CA125. HE4 levels were more frequently elevated in patients with distant metastasis compared to local recurrences, 67% and 37%, respectively. Serum HE4 detected a recurrence with a median of 126 days earlier than clinical confirmation. CONCLUSION: Elevated serum HE4 is an independent risk factor for reduced disease-free survival and overall survival. HE4 seems to be superior to CA125 in the detection of recurrent disease during follow-up, mainly in high-risk endometrial cancer patients who are more prone to distant metastasis.

Iron absorption from oral iron supplements given on consecutive versus alternate days and as single morning doses versus twice-daily split dosing in iron-depleted women: two open-label, randomised controlled trials.

BACKGROUND: Current guidelines to treat iron deficiency recommend daily provision of ferrous iron divided through the day to increase absorption. However, daily dosing and split dosing might increase serum hepcidin and decrease iron absorption from subsequent doses. Our study aim was to compare iron absorption from oral iron supplements given on consecutive versus alternate days and given as single morning doses versus twice-daily split dosing. METHODS: We did two prospective, open-label, randomised controlled trials assessing iron absorption using (54Fe)-labelled, (57Fe)-labelled, or (58Fe)-labelled ferrous sulfate in iron-depleted (serum ferritin ≤25 µg/L) women aged 18-40 years recruited from ETH Zurich and the University of Zurich, Switzerland. In study 1, women were randomly assigned (1:1) to two groups. One group was given 60 mg iron at 0800 h (±1 h) on consecutive days for 14 days, and the other group was given the same doses on alternate days for 28 days. In study 2, women were assigned to two groups, stratified by serum ferritin so that two groups with similar iron statuses could be formed. One group was given 120 mg iron at 0800 h (±1 h) and the other was given the dose split into two divided doses of 60 mg at 0800 h (±1 h) and 1700 h (±1 h) for three consecutive days. 14 days after the final dose, the groups were each crossed over to the other regimen. Within-individual comparisons were done. The co-primary outcomes in both studies were iron bioavailability (total and fractional iron absorption), assessed by measuring the isotopic label abundance in erythrocytes 14 days after administration, and serum hepcidin. Group allocations in both studies were not masked and primary and safety analyses were done on an intention-to-treat basis. The studies were registered at ClinicalTrials.gov, numbers NCT02175888 (study 1) and NCT02177851 (study 2) and are complete. FINDINGS: For study 1, 40 women were enrolled on Oct 15-29, 2015. 21 women were assigned to the consecutive-day group and 19 to the alternate-day group. At the end of treatment (14 days for the consecutive-day group and 28 days for the alternate-day group), geometric mean (-SD, +SD) cumulative fractional iron absorptions were 16·3% (9·3, 28·8) in the consecutive-day group versus 21·8% (13·7, 34·6) in the alternate-day group (p=0·0013), and cumulative total iron absorption was 131·0 mg (71·4, 240·5) versus 175·3 mg (110·3, 278·5; p=0·0010). During the first 14 days of supplementation in both groups, serum hepcidin was higher in the consecutive-day group than the alternate-day group (p=0·0031). In study 2, 20 women were enrolled between Aug 13 and 18, 2015. Ten women were assigned to receive once-daily dosing and ten were assigned to receive twice-daily divided dosing. No significant differences were seen in fractional (day 1-3 geometric mean: 11·8% [7·1, 19·4] once daily vs 13·1% [8·2, 20·7] twice daily; p=0·33) or total iron absorption (day 1-3: 44·3 mg [29·4, 66·7] once daily vs 49·4 [35·2, 69·4] twice daily; p=0·33) between the two dosing regimens. Twice-daily divided doses resulted in a higher serum hepcidin concentration than once-daily dosing (p=0·013). No grade 3 or 4 adverse events were reported in either study. INTERPRETATION: In iron-depleted women, providing iron supplements daily as divided doses increases serum hepcidin and reduces iron absorption. Providing iron supplements on alternate days and in single doses optimises iron absorption and might be a preferable dosing regimen. These findings should be confirmed in iron-deficient anaemic patients. FUNDING: Swiss National Science Foundation, Bern, Switzerland.

Meta-GWAS and Meta-Analysis of Exome Array Studies Do Not Reveal Genetic Determinants of Serum Hepcidin.

Serum hepcidin concentration is regulated by iron status, inflammation, erythropoiesis and numerous other factors, but underlying processes are incompletely understood. We studied the association of common and rare single nucleotide variants (SNVs) with serum hepcidin in one Italian study and two large Dutch population-based studies. We genotyped common SNVs with genome-wide association study (GWAS) arrays and subsequently performed imputation using the 1000 Genomes reference panel. Cohort-specific GWAS were performed for log-transformed serum hepcidin, adjusted for age and gender, and results were combined in a fixed-effects meta-analysis (total N 6,096). Six top SNVs (p<5x10-6) were genotyped in 3,821 additional samples, but associations were not replicated. Furthermore, we meta-analyzed cohort-specific exome array association results of rare SNVs with serum hepcidin that were available for two of the three cohorts (total N 3,226), but no exome-wide significant signal (p<1.4x10-6) was identified. Gene-based meta-analyses revealed 19 genes that showed significant association with hepcidin. Our results suggest the absence of common SNVs and rare exonic SNVs explaining a large proportion of phenotypic variation in serum hepcidin. We recommend extension of our study once additional substantial cohorts with hepcidin measurements, GWAS and/or exome array data become available in order to increase power to identify variants that explain a smaller proportion of hepcidin variation. In addition, we encourage follow-up of the potentially interesting genes that resulted from the gene-based analysis of low-frequency and rare variants.

Toward Worldwide Hepcidin Assay Harmonization: Identification of a Commutable Secondary Reference Material.

BACKGROUND: Absolute plasma hepcidin concentrations measured by various procedures differ substantially, complicating interpretation of results and rendering reference intervals method dependent. We investigated the degree of equivalence achievable by harmonization and the identification of a commutable secondary reference material to accomplish this goal. METHODS: We applied technical procedures to achieve harmonization developed by the Consortium for Harmonization of Clinical Laboratory Results. Eleven plasma hepcidin measurement procedures (5 mass spectrometry based and 6 immunochemical based) quantified native individual plasma samples (n = 32) and native plasma pools (n = 8) to assess analytical performance and current and achievable equivalence. In addition, 8 types of candidate reference materials (3 concentrations each, n = 24) were assessed for their suitability, most notably in terms of commutability, to serve as secondary reference material. RESULTS: Absolute hepcidin values and reproducibility (intrameasurement procedure CVs 2.9%-8.7%) differed substantially between measurement procedures, but all were linear and correlated well. The current equivalence (intermeasurement procedure CV 28.6%) between the methods was mainly attributable to differences in calibration and could thus be improved by harmonization with a common calibrator. Linear regression analysis and standardized residuals showed that a candidate reference material consisting of native lyophilized plasma with cryolyoprotectant was commutable for all measurement procedures. Mathematically simulated harmonization with this calibrator resulted in a maximum achievable equivalence of 7.7%. CONCLUSIONS: The secondary reference material identified in this study has the potential to substantially improve equivalence between hepcidin measurement procedures and contributes to the establishment of a traceability chain that will ultimately allow standardization of hepcidin measurement results.

Therapeutic targeting of autophagy in cancer. Part I: molecular pathways controlling autophagy.

Autophagy is a process in which cells can generate energy and building materials, by degradation of redundant and/or damaged organelles and proteins. Especially during conditions of stress, autophagy helps to maintain homeostasis. In addition, autophagy has been shown to influence malignant transformation and cancer progression. The precise molecular events in autophagy are complex and the core autophagic machinery described to date consists of nearly thirty proteins. Apart from these factors that execute the process of autophagy, several signalling pathways are involved in converting internal and external stimuli into an autophagic response. In this review we provide an overview of the signalling pathways that influence autophagy, particularly in cancer cells. We will illustrate that interference with multiple of these signalling pathways can have significant effects on cancer cell survival.

Therapeutic targeting of autophagy in cancer. Part II: pharmacological modulation of treatment-induced autophagy.

Autophagy, the catabolic pathway in which cells recycle organelles and other parts of their own cytoplasm, is increasingly recognised as an important cytoprotective mechanism in cancer cells. Several cancer treatments stimulate the autophagic process and when autophagy is inhibited, cancer cells show an enhanced response to multiple treatments. These findings have nourished the theory that autophagy provides cancer cells with a survival advantage during stressful conditions, including exposure to therapeutics. Therefore, interference with the autophagic response can potentially enhance the efficacy of cancer therapy. In this review we examine two approaches to modulate autophagy as complementary cancer treatment: inhibition and induction. Inhibition of autophagy during cancer treatment eliminates its cytoprotective effects. Conversely, induction of autophagy combined with conventional cancer therapy exerts severe cytoplasmic degradation that can ultimately lead to cell death. We will discuss how autophagy can be therapeutically manipulated in cancer cells and how interactions between the conventional cancer therapies and autophagy modulation influence treatment outcome.

Cetuximab reduces the accumulation of radiolabeled bevacizumab in cancer xenografts without decreasing VEGF expression.

Bevacizumab and cetuximab are approved for the treatment of cancer. However, in advanced colorectal cancer, addition of cetuximab to chemotherapy with bevacizumab did not improve survival. The reason for the lack of activity remains unclear. The aim of this study was to determine the effect of cetuximab on VEGF expression and targeting of bevacizumab to the tumor. Mice with subcutaneous SUM149 or WiDr xenografts were treated with cetuximab, bevacizumab, or a combination of the two. Before the start of cetuximab treatment and after 7 and 21 days of treatment, the uptake of radiolabeled bevacizumab in the tumor was measured by immunoSPECT/CT. Tumor growth of SUM149 xenografts was significantly inhibited by cetuximab, bevacizumab, or their combination, whereas growth of WiDr xenografts was not affected. Cetuximab caused a significant reduction of bevacizumab uptake in SUM149 xenografts, whereas tumor-to-blood ratios in mice with WiDr xenografts did not change. Biodistribution studies with an irrelevant antibody in the SUM149 model also showed significantly reduced tumor-to-blood ratios. Cetuximab treatment did not decrease VEGF expression. Without decreasing VEGF levels, cetuximab reduces tumor targeting of bevacizumab. This could, at least partly, explain why the combination of bevacizumab and cetuximab does not result in improved therapeutic efficacy.

Quantification of patient-specific assay interference in different formats of enzyme-linked immunoassays for therapeutic monoclonal antibodies.

BACKGROUND: The use of therapeutic monoclonal antibodies for clinical purposes has significantly increased in recent years, and so has the need to monitor antibody concentrations. This may be achieved using the well-established enzyme linked immunoassay (ELISA) methods; however, these assays are subject to a variety of interferences. METHODS: In the present study, the authors have tested the ELISA methods for quantifying bevacizumab (BVZ) to investigate this interference. Three different ELISA methods were used and exhibited similar characteristics. RESULTS: The detection limits of the ELISA methods varied from 0.05 to 0.07 ng/mL. To monitor assay performance, BVZ was measured in a control sample during each run. The BVZ concentration in the control sample was 15.4 µg/mL, the within-run imprecision (CV) and between-run CV were 4.3% and 10.4% (direct ELISA), 5.2% and 12.9% (indirect/Rabbit ELISA), and 3.9% and 9.1% (indirect/Chicken ELISA). The assays exhibited good precision and parallelism in serial dilutions of samples and a mean recovery of 98% (range, 78%-118%). CONCLUSIONS: The authors show that the degree of interference by using direct and indirect target immobilization depends heavily on the method of target immobilization on the surface of the ELISA plate, and is patient-specific. The results highlight pitfalls of potential relevance to sandwich-type assays, and an approach to rectify such problems. This approach will yield a valid assay protocol for the measurement of monoclonal therapeutic antibodies in case no target is available for direct immobilization.

Associations of common variants in HFE and TMPRSS6 with iron parameters are independent of serum hepcidin in a general population: a replication study.

BACKGROUND: Genome-wide association studies have convincingly shown that single nucleotide polymorphisms (SNPs) in HFE and TMPRSS6 are associated with iron parameters. It was commonly thought that these associations could be explained by the intermediate effect on hepcidin concentration. A recent study in an isolated Italian population, however, concluded that these associations were not exclusively dependent on hepcidin values. We report here the second study to investigate the role of hepcidin in the associations between common variants in HFE and TMPRSS6 with iron parameters. METHODS: We extracted 101 SNPs in HFE and TMPRSS6 from genome-wide imputed SNP data of 1832 individuals from the general population (Nijmegen Biomedical Study). Single locus and haplotype associations with serum iron parameters and hepcidin were studied using linear regression analyses. RESULTS: We found that HFE rs1800562 and TMPRSS6 rs855791 are the main determinants of HFE and TMPRSS6 related variation in serum iron, ferritin, transferrin saturation, and total iron binding capacity. These SNPs are associated with the ratios hepcidin/ferritin (p<1×10(-5)) and hepcidin/transferrin saturation (p<1×10(-3)), but not with serum hepcidin (p>0.2). Adjustment for hepcidin or the ratio hepcidin/ferritin did not decrease the strength of the SNP-iron parameter associations. CONCLUSIONS: Our results do not support an intermediate role for hepcidin in the SNP-iron parameter associations, which confirms previous findings, and indicate a pleiotropic SNP effect on the hepcidin ratios and the iron parameters. Taken together, this suggests that there might be other, yet unknown, serum hepcidin independent mechanisms which play a role in the association of HFE and TMPRSS6 variants with serum iron parameters.

Maternal soluble fms-like tyrosine kinase-1, placental growth factor, plasminogen activator inhibitor-2, and folate concentrations and early fetal size: the Generation R study.

OBJECTIVE: Fetal growth is dependent on adequate development of the placenta. Impaired angiogenesis and vasculogenesis in early pregnancy compromises placental and embryonic development. The proteins soluble fms-like tyrosine kinase (sFlt)-1, placental growth factor (PlGF), and plasminogen activator inhibitor (PAI)-2, and the B vitamin folate are determinants of placental development. This study aims to identify associations between these maternal biomarkers and early fetal size. STUDY DESIGN: From a prospective birth cohort study in The Netherlands, 1491 pregnant women were selected for this study. At a mean gestational age (GA) of 12.4 weeks (SD 0.8) maternal venous blood samples were obtained to determine the concentrations of sFlt-1, PlGF, PAI-2, and folate. Early fetal size was assessed with measurement of the crown-to-rump length (CRL) at a mean of 12.4 weeks' GA (SD 0.8). Analyses were performed using multivariable linear regression analyses with the biomarkers (continuous, quintiles) as regressors and CRL as main outcome measure. RESULTS: Linear trend analysis showed positive associations between maternal sFlt-1 (P < .001), PlGF (P = .042), PAI-2 (P < .001), and folate (P = .039) and CRL. These associations were independent of GA, maternal age, height, body mass index, ethnicity, fetal sex, parity, educational level, smoking, and folic acid supplement use (folate not adjusted). CONCLUSION: sFlt-1, PlGF, PAI-2, and folate are positively associated with first-trimester fetal size.

Role of endogenous vascular endothelial growth factor in endothelium-dependent vasodilation in humans.

Angiogenesis inhibitors have remarkably improved the outcome of patients with several types of cancer. Hypertension is the most reported side effect of angiogenesis inhibitors interfering with vascular endothelial growth factor signaling. In this study, we test the hypothesis that circulating vascular endothelial growth factor at physiological concentrations is essential to preserve normal endothelial control of vasomotor tone. In 7 healthy male volunteers, infusion of bevacizumab (monoclonal vascular endothelial growth factor antibody) into the brachial artery for 15 minutes (144 µg/dL forearm volume per minute) did not affect forearm vasodilator tone as measured with venous occlusion strain gauge plethysmography. In a separate group of 12 male volunteers, a similar bevacizumab infusion reduced the vasodilator response to 2 dosages of acetylcholine from (mean ± SE) 440 ± 157% and 926 ± 252% to 169 ± 40% and 612 ± 154% (P<0.05). Finally, in a third group of 12 volunteers, bevacizumab did not alter the percentage increase in forearm blood flow during infusion of sodium nitroprusside at dosages equipotent to acetylcholine. Bevacizumab acutely and specifically reduced endothelium-mediated vasodilation at local concentrations that resemble plasma concentrations after systemic exposure to bevacizumab. This observation suggests a physiological role for vascular endothelial growth factor in maintaining normal endothelial control of vasomotor tone. The role of the endothelium in the mechanism of bevacizumab-induced hypertension deserves further exploration.

Angiogenic and fibrinolytic factors in blood during the first half of pregnancy and adverse pregnancy outcomes.

OBJECTIVE: To estimate whether the imbalance of angiogenic factors (soluble fms-like tyrosine kinase-1, placental growth factor) and fibrinolytic factors (plasminogen activator inhibitor-2 [PAI-2]) might affect placentation in early pregnancy. METHODS: We studied the associations of maternal soluble fms-like tyrosine kinase-1, placental growth factor, and PAI-2 concentrations in the first trimester (before 18 weeks of gestation) and soluble fms-like tyrosine kinase-1 and placental growth factor concentrations in the second trimester (18-25 weeks of gestation) with placental function and adverse pregnancy outcomes. This study was embedded in a population-based prospective cohort study. Data were used from 7,519 women. Biomarker concentrations were divided into deciles and evaluated in multivariable linear and logistic regression models. RESULTS: First-trimester high soluble fms-like tyrosine kinase-1 was associated with a 5.2% lower uterine artery index in the second-trimester and a 1.6% higher birth weight (55 g, confidence interval [CI] 15-95). Neither in the first nor in the second trimester were soluble fms-like tyrosine kinase-1 concentrations significantly associated with preeclampsia. First-trimester low placental growth factor was associated with a 6.1% higher uterine artery index and a 3.4% lower birth weight (-115 g, CI -157 to -74). First-trimester low placental growth factor was associated with fetal growth restriction (odds ratio [OR] 2.62, CI 1.68-4.08) and preeclampsia (OR 2.46, CI 1.49-4.08). First-trimester low PAI-2 was associated with a 1.9% higher uterine artery index and a 2.7% lower birth weight (-94 g, CI -136 to -51). First-trimester low PAI-2 was associated with a higher risk of fetal growth restriction (OR 2.22, CI 1.39-3.55). CONCLUSION: First-half-of-pregnancy concentrations of soluble fms-like tyrosine kinase-1, placental growth factor, and PAI-2 are associated with uteroplacental vascular resistance, placental weight, and birth weight. Moreover, first-trimester placental growth factor and PAI-2 are associated with an increased risk of adverse pregnancy outcomes. LEVEL OF EVIDENCE: II.

Serum hepcidin: reference ranges and biochemical correlates in the general population.

To date, concentrations of the promising biomarker hepcidin have only been assessed in serum of relatively small series of healthy volunteers and patients. We assessed age- and sex-stratified reference ranges of serum hepcidin concentration in a selected reference set and performed regression analyses to study associations between hepcidin and (biochemical) variables in a large, well-phenotyped sample of the general population (n = 2998). All participants filled out a questionnaire on lifestyle, health status, and medical history. Serum measurements of iron parameters, liver enzyme alanine aminotransferase, creatinine and C-reactive protein were available. Serum hepcidin concentrations were lower for premenopausal than for postmenopausal women (median, 4.1 nM vs 8.5 nM, respectively). Hepcidin concentrations in men were constant over age (median, 7.8 nM). Serum hepcidin was strongly associated with serum ferritin in men and women: ß-coefficient of log-transformed variables (95% confidence interval): 0.78 (0.74-0.82) and 0.83 (0.78-0.88), respectively. Additional significant, though less strong, associations were observed for C-reactive protein and total iron binding capacity in men and for total iron binding capacity, alanine aminotransferase, and glomerular filtration rate in women. Our study provides age- and sex-specific reference ranges of serum hepcidin concentration and indicates ferritin as the primary correlate of serum hepcidin concentration.