Human Vascular Endothelial Growth Factor Research Articles

Base amount-dependent fluorescence enhancement for the assay of vascular endothelial growth factor 165 in human serum using hairpin DNA-silver nanoclusters and oxidized carbon nanoparticles.

A base amount-dependent fluorescence enhancement-based strategy is put forward to determine vascular endothelial growth factor 165 (VEGF165) in human serum by the use of hairpin DNA-silver nanoclusters (hDNA-AgNCs) and oxidized carbon nanoparticles (CNPs). The hDNA-AgNCs aptasensing probe consists of AgNCs-contained hairpin loop (that generates afluorescence signal), hairpin stem (that makes the structure stable), and the terminal aptamer 1 (that recognizesthe target together with aptamer 2). It has been demonstrated that the fluorescence intensity of hDNA-AgNCs is ~ 3-fold stronger than that of single-stranded DNA-AgNCs (ssDNA-AgNCs), and hDNA-AgNCs have a strong dependence of fluorescence enhancement on the base amount in hairpin stem and loop. Upon the addition of oxidized CNPs, the terminal aptamer 1 of hDNA-AgNCs can adsorb onto the surface of oxidized CNPs via π-π stacking, and the fluorescence of hDNA-AgNCs (with excitation/emission maxima at 490/567nm) is quenched via fluorescence resonance energy transfer (FRET). When aptamer 2 and VEGF165 are subsequently added, aptamer 1, VEGF165, and aptamer 2 reassemble into anintact tertiary structure, and the fluorescence is recovered because hDNA-AgNCs are far away from the surface of oxidized CNPs and the FRET efficiency decreases. Under the optimized conditions, the aptasensing probe can selectively assay VEGF165 with a detectionlimit of 14pM. The results provide a label-free and sensitive method to monitor VEGF165 in human serum. Schematic representation of the strong dependence of fluorescence enhancement on base amount in stem and loop of hairpin DNA-silver nanoclusters. The probe can be used to assay vascular endothelial growth factor 165 (VEGF165) and give a judgment whether human serum VEGF165 is at a normal or abnormal level for clinical diagnosis.

Targeted Molecular Therapies in the Treatment of Esophageal Adenocarcinoma, Are We There Yet?

Simple SummaryTraditional therapeutic approaches to esophageal adenocarcinoma involve a combination of surgery, chemotherapy, and radiation. Despite innovations in treatment, outcomes remain poor. Targeted molecular therapies and immunotherapies have been used to great effect in various other solid tumors. Several targeted agents show promise in treating esophageal adenocarcinoma. In this review, we aim to highlight recent developments in the arena of targeted therapeutics and suggest topics of future investigations.Esophageal adenocarcinoma is one of the leading causes of cancer-related deaths worldwide. The incidence of esophageal adenocarcinoma has increased at an alarming rate in the Western world and long-term survival remains poor. Current treatment approaches involve a combination of surgery, chemotherapy, and radiotherapy. Unfortunately, standard first-line approaches are met with high rates of recurrence and metastasis. More recent investigations into the distinct molecular composition of these tumors have uncovered key genetic and epigenetic alterations involved in tumorigenesis and progression. These discoveries have driven the development of targeted therapeutic agents in esophageal adenocarcinoma. While many agents have been studied, therapeutics targeting the human epidermal growth factor receptor (HER2) and vascular endothelial growth factor (VEGF) pathways have demonstrated improved survival. More recent advances in immunotherapies have also demonstrated survival advantages with monoclonal antibodies targeting the programmed death ligand 1 (PD-L1). In this review we highlight recent advances of targeted therapies, specifically agents targeting receptor tyrosine kinases, small molecule kinase inhibitors, and immune checkpoint inhibitors. While targeted therapeutics and immunotherapies have significantly improved survival, the benefits are limited to patients whose tumors express biomarkers such as PD-L1 and HER2. Survival remains poor for the remainder of patients with esophageal adenocarcinoma, underscoring the critical need for development of novel treatment strategies.

THE EFFECT OF AN 8-WEEK ANAEROBIC GYMNASTICS TRAINING ON BDNF, VEGF, AND SOME PHYSIOLOGICAL CHARACTERISTICS IN CHILDREN

The purpose of the present study was to observe changes in levels of brain-derived neurotrophic factor (BDNF), vascular endothelial growth factor (VEGF), resting metabolic rate (RMR) and maximum oxygen consumption (VO2max) in the gymnast children after an anaerobic gymnastics training program. Thirty beginner gymnasts aged 8-12 years old were randomly assigned to control (n = 15) and experimental (n = 15) groups. The anaerobic gymnastics training was conducted for 8 weeks, 3 times per a week. Each session lasted 45 minutes: 10 min warm-up, 30 min core exercise, and 5 min cool down. The anthropometric and body composition of subjects were measured and growth factors were measured by using human BDNF and VEGF PicoKine™ ELISA Kit and analysis was performed using sandwich enzyme-linked immunosorbent assay (Morland et al.) before and after the intervention, and VO2max, maximum heart rate and RMR were measured using a gas analyzer. At the baseline there were not any significant differences between both groups (p>0.05). But in the post-test, a significant difference was observed for BDNF(p=0.02) and VEGF(p=0.018) values between the two groups. Within-group there was a decrease in the value of the maximum heart rate indicator (P<0.05) and VO2max and BDNF increased significantly after an intervention (P<0.05). In conclusion, the results of the present study suggest that anaerobic gymnastic training increases the level of salivary BDNF and VEGF in children. These types of exercises may also improve cardiorespiratory fitness in children.

Design, synthesis, and biological evaluation of new challenging thalidomide analogs as potential anticancer immunomodulatory agents

Thalidomide and its analogs are immunomodulatory drugs that inhibit the production of certain inflammatory mediators associated with cancer. In the present work, a new series of thalidomide analogs was designed and synthesized to obtain new effective antitumor immunomodulatory agents. The synthesized compounds were evaluated for their cytotoxic activities against a panel of four cancer cell lines (HepG-2, HCT-116, PC3 and MCF-7). Compounds 33h, 33i, 42f and 42h showed strong potencies against all tested cell lines with IC50 values ranging from 14.63 to 49.90 µM comparable to that of thalidomide (IC50 values ranging from 32.12 to 76.91 µM). The most active compounds were further evaluated for their in vitro immunomodulatory activities via estimation of human tumor necrosis factor alpha (TNF-α), human caspase-8 (CASP8), human vascular endothelial growth factor (VEGF), and nuclear factor kappa-B P65 (NF-κB P65) in HCT-116 cells. Thalidomide was used as a positive control. Compounds 33h and 42f showed a significant reduction in TNF-α. Furthermore, compounds 33i and 42f exhibited significant elevation in CASP8 levels. Compounds 33i and 42f inhibited VEGF. In addition, compound 42f showed significant decrease in levels of NF-κB p65. Moreover, apoptosis and cell cycle tests of the most active compound 42f, were performed. The results indicated that compound 42f significantly induce apoptosis in HCT-116 cells and arrest cell cycle at the G2/M phase.

Associations between VEGF isoforms and impending retinopathy of prematurity.

Vascular Endothelial Growth Factor (VEGF) is the main driver of angiogenesis during neurodevelopment (i.e., brain and retina). VEGF165 and VEGF121 are the two most prevalent human VEGF isoforms. Although retinopathy of prematurity (ROP), a neuroretinal disorder, is associated with VEGF dysregulation, little is known about the interaction of VEGF isoforms on neuroretinal angiogenesis. We hypothesized that: (a) A specific VEGF165/VEGF121 correlation, at a given time point, is associated with normal retinal development (no ROP) and (b) An altered correlation, of such, is associated with aberrant retinal development (ROP). Utilizing pre-collected dried blood spots (DBS) from<1-week-old preterm infants, we aimed to determine whether correlations between VEGF165 and VEGF121 precede the diagnosis of early stage, non-proliferative ROP (NP-ROP). We conducted a case/control study, utilizing DBS from 65 preterm infants. We measured DBS levels of VEGF165 on the Mesoscale Discovery Platform and VEGF121 via Cloud Clone Elisa Assay. In infants with NP-ROP, VEGF165 is significantly higher in males (than females). In infants without ROP, there is a significant correlation between VEGF165 and VEGF121 in females (but not males). In infants with NP-ROP, the opposite is so; there is a significant correlation between VEGF165 and VEGF121 in males (but not females). This pilot study, utilizing de-identified data, suggests the potential importance of examining interactions between VEGF isoforms, at<1week after birth, to better understand ROP development. Our study also suggests that retinal angiogenesis may not be a sex-neutral process. A prospective study is needed to confirm our novel findings.

Rapid transient expression of functional human vascular endothelial growth factor in Nicotiana benthamiana and characterization of its biological activity

Human vascular endothelial growth factor (VEGF) is a potent pro-angiogenic growth factor essential for wound healing. Due to its potential applications, many expression strategies have been developed to produce high levels of VEGF. Here, we have optimized the expression conditions for the production of recombinant VEGF in Nicotiana benthamiana by using a geminiviral vector. Four different expression constructs that differ by the location of a C- or N-terminal histidine tag and SEKDEL sequence were developed and utilized for plant transient expression. The recombinant VEGF was further purified by using affinity chromatography and confirmed by SDS-PAGE and Western blotting probed with anti-VEGF antibody. Furthermore, our results showed that the recombinant VEGF in all tested concentrations did not exhibit any cytotoxic effect on HaCaT cells and induced cell migration in vitro. These findings show that the plant-produced VEGF has the potential to be used in regenerative medicine and cosmetic industry.

Analysis of plant-derived phytochemicals as anti-cancer agents targeting cyclin dependent kinase-2, human topoisomerase IIa and vascular endothelial growth factor receptor-2

Cancer is caused by a variety of pathways, involving numerous types of enzymes. Among them three enzymes i.e. Cyclin-dependent kinase-2 (CDK-2), Human topoisomerase IIα, and Vascular Endothelial Growth Factor Receptor-2 (VEGFR-2) are three of the most common enzymes that are involved in the cancer development. Although many chemical drugs are already available in the market for cancer treatment, plant sources are known to contain a wide variety of agents that are proved to possess potential anticancer activity. In this experiment, total thirty phytochemicals were analyzed against the mentioned three enzymes using different tools of bioinformatics and in silico biology like molecular docking study, drug likeness property experiment, ADME/T test, PASS prediction, and P450 site of metabolism prediction as well as DFT calculation to determine the three best ligands among them that have the capability to inhibit the mentioned enzymes. From the experiment, Epigallocatechin gallate was found to be the best ligand to inhibit CDK-2, Daidzein showed the best inhibitory activities towards the Human topoisomerase IIα, and Quercetin was predicted to be the best agent against VEGFR-2. They were also predicted to be quite safe and effective agents to treat cancer. However, more in vivo and in vitro analyses are required to finally confirm their safety and efficacy in this regard.

Exosomes derived from human mesenchymal stem cells preserve mouse islet survival and insulin secretion function.

Islet cell death and loss of function after isolation and before transplantation is considered a key barrier to successful islet transplantation outcomes. Mesenchymal stem cells (MSCs) have been used to protect isolated islets owing to their paracrine potential partially through the secretion of vascular endothelial growth factor (VEGF). The paracrine functions of MSCs are also mediated, at least in part, by the release of extracellular vesicles including exosomes. In the present study, we examined (i) the effect of exosomes from human MSCs on the survival and function of isolated mouse islets and (ii) whether exosomes contain VEGF and the potential impact of exosomal VEGF on the survival of mouse islets. Isolated mouse islets were cultured for three days with MSC-derived exosomes (MSC-Exo), MSCs, or MSC-conditioned media without exosomes (MSC-CM-without-Exo). We investigated the effects of the exosomes, MSCs, and conditioned media on islet viability, apoptosis and function. Besides the expression of apoptotic and pro-survival genes, the production of human and mouse VEGF proteins was evaluated. The MSCs and MSC-Exo, but not the MSC-CM-without-Exo, significantly decreased the percentage of apoptotic cells and increased islet viability following the downregulation of pro-apoptotic genes and the upregulation of pro-survival factors, as well as the promotion of insulin secretion. Human VEGF was observed in the isolated exosomes, and the gene expression and protein production of mouse VEGF significantly increased in islets cultured with MSC-Exo. MSC-derived exosomes are as efficient as parent MSCs for mitigating cell death and improving islet survival and function. This cytoprotective effect was probably mediated by VEGF transfer, suggesting a pivotal strategy for ameliorating islet transplantation outcomes.

Persistent Pulmonary Hypertension of the Newborn: Pathophysiological Mechanisms and Novel Therapeutic Approaches.

Persistent pulmonary hypertension of the newborn (PPHN) is one of the main causes of neonatal morbidity and mortality. It is characterized by sustained elevation of pulmonary vascular resistance (PVR), preventing an increase in pulmonary blood flow after birth. The affected neonates fail to establish blood oxygenation, precipitating severe respiratory distress, hypoxemia, and eventually death. Inhaled nitric oxide (iNO), the only approved pulmonary vasodilator for PPHN, constitutes, alongside supportive therapy, the basis of its treatment. However, nearly 40% of infants are iNO resistant. The cornerstones of increased PVR in PPHN are pulmonary vasoconstriction and vascular remodeling. A better understanding of PPHN pathophysiology may enlighten targeted and more effective therapies. Sildenafil, prostaglandins, milrinone, and bosentan, acting as vasodilators, besides glucocorticoids, playing a role on reducing inflammation, have all shown potential beneficial effects on newborns with PPHN. Furthermore, experimental evidence in PPHN animal models supports prospective use of emergent therapies, such as soluble guanylyl cyclase (sGC) activators/stimulators, l-citrulline, Rho-kinase inhibitors, peroxisome proliferator-activated receptor-γ (PPAR-γ) agonists, recombinant superoxide dismutase (rhSOD), tetrahydrobiopterin (BH4) analogs, ω-3 long-chain polyunsaturated fatty acids (LC-PUFAs), 5-HT2A receptor antagonists, and recombinant human vascular endothelial growth factor (rhVEGF). This review focuses on current knowledge on alternative and novel pathways involved in PPHN pathogenesis, as well as recent progress regarding experimental and clinical evidence on potential therapeutic approaches for PPHN.

Alginate/laponite hydrogel microspheres co-encapsulating dental pulp stem cells and VEGF for endodontic regeneration

Considering the complicated and irregular anatomical structure of root canal systems, injectable microspheres have received considerable attention as cell carriers in endodontic regeneration. Herein, we developed injectable hybrid RGD-alginate/laponite (RGD-Alg/Lap) hydrogel microspheres, co-encapsulating human dental pulp stem cells (hDPSCs) and vascular endothelial growth factor (VEGF). These microspheres were prepared by the electrostatic microdroplet method with an average size of 350~450 μm. By adjusting the content of laponite, the rheological properties and the degradation rate of the microspheres in vitro could be conditioned. The release of VEGF from the RGD-Alg/0.5%Lap microspheres was in a sustained manner for 28 days while the bioactivity of VEGF was preserved. In addition, the encapsulated hDPSCs were evenly distributed in microspheres with a cell viability exceeding 85%. The deposition of abundant extracellular matrix such as fibronectin (FN) and collagen type I (Col-I) was shown in microspheres after 7 days. The laponite in the system significantly up-regulated the expression of odontogenic-related genes of hDPSCs at day 7. Furthermore, after subcutaneous implantation with tooth slices in a nude mouse model for 1 month, the hDPSCs-laden RGD-Alg/0.5%Lap+VEGF microspheres significantly promoted the regeneration of pulp-like tissues as well as the formation of new micro-vessels. These results demonstrated the great potential of laponite-enhanced hydrogel microspheres in vascularized dental pulp regeneration. Statement of SignificanceInjectable cell-laden microspheres have recently gained great attention in endodontic regeneration. Here we first developed hybrid alginate/laponite hydrogel microspheres (size about 350~450 μm) by electrostatic microdroplet method, which exhibited tunability in mechanical property and sustained release ability. The incorporation of laponite and the sustained release of VEGF supported not only dental pulp stem cells differentiation in vitro but neotissue regeneration in vivo. These features combined with the simplicity in preparation, made the microspheres ideally suited to simultaneous cells and growth factors delivery in dental pulp regeneration and even other tissue regeneration application.

Intracerebroventricular delivery of vascular endothelial growth factor in patients with amyotrophic lateral sclerosis, a phase I study.

We studied the feasibility, safety, tolerability and pharmacokinetics of intracerebroventricular delivery of recombinant human vascular endothelial growth factor in patients with amyotrophic lateral sclerosis. In this phase I study in patients with amyotrophic lateral sclerosis, the study drug was delivered using an implantable programmable pump connected to a catheter inserted in the frontal horn of the lateral cerebral ventricle. A first cohort received open label vascular endothelial growth factor (0.2, 0.8 and 2 µg/day), a second cohort received placebo, 0.8 or 2 µg/day of study dug. After the 3-month study period, all patients could participate in an open label extension study. In total, 18 patients with amyotrophic lateral sclerosis, seen at the University Hospitals in Leuven were included. The surgical procedure was well tolerated in most patients. One patient had transient postoperative seizures, due to an ischemic lesion along the catheter tract. The first 3-month study period was completed by 15/18 patients. Administration of 2 µg/day vascular endothelial growth factor resulted in sustained detectable levels in cerebrospinal fluid. A pulmonary embolus occurred in 3 patients, in 1 patient in the first 3-month study, and in 2 patients during the open label extension study. The study drug was well tolerated in the other patients, for up to 6 years in the open label extension study. Our study shows that intracerebroventricular administration of 2 µg/day of vascular endothelial growth factor to patients with amyotrophic lateral sclerosis is feasible, results in detectable cerebrospinal fluid levels and is well tolerated in most patients. The most common serious adverse event was a pulmonary embolus.

Transplantation of human dental pulp stem cells ameliorates diabetic polyneuropathy in streptozotocin-induced diabetic nude mice: the role of angiogenic and neurotrophic factors

BackgroundDental pulp stem cells (DPSCs) have high proliferation and multi-differentiation capabilities that maintain their functionality after cryopreservation. In our previous study, we demonstrated that cryopreserved rat DPSCs improved diabetic polyneuropathy and that the efficacy of cryopreserved rat DPSCs was equivalent to that of freshly isolated rat DPSCs. The present study was conducted to evaluate whether transplantation of cryopreserved human DPSCs (hDPSCs) is also effective for the treatment of diabetic polyneuropathy.MethodshDPSCs were isolated from human impacted third molars being extracted for orthodontic reasons. Eight weeks after the induction of diabetes in nude mice, hDPSCs (1 × 105/limb) were unilaterally transplanted into the hindlimb skeletal muscle, and vehicle (saline) was injected into the opposite side as a control. The effects of hDPSCs were analyzed at 4 weeks after transplantation.ResultshDPSC transplantation significantly ameliorated reduced sensory perception thresholds, delayed nerve conduction velocity, and decreased the blood flow to the sciatic nerve in diabetic mice 4 weeks post-transplantation. Cultured hDPSCs secreted the vascular endothelial growth factor (VEGF) and nerve growth factor (NGF) proteins. A subset of the transplanted hDPSCs was localized around the muscle bundles and expressed the human VEGF and NGF genes at the transplanted site. The capillary/muscle bundle ratio was significantly increased on the hDPSC-transplanted side of the gastrocnemius muscles in diabetic mice. Neutralizing antibodies against VEGF and NGF negated the effects of hDPSC transplantation on the nerve conduction velocity in diabetic mice, suggesting that VEGF and NGF may play roles in the effects of hDPSC transplantation on diabetic polyneuropathy.ConclusionsThese results suggest that stem cell transplantation with hDPSCs may be efficacious in treating diabetic polyneuropathy via the angiogenic and neurotrophic mechanisms of hDPSC-secreted factors.

Modular protein-oligonucleotide signal exchange.

While many methods are available to measure the concentrations of proteins in solution, the development of a method to quantitatively report both increases and decreases in different protein concentrations in real-time using changes in the concentrations of other molecules, such as DNA outputs, has remained a challenge. Here, we present a biomolecular reaction process that reports the concentration of an input protein in situ as the concentration of an output DNA oligonucleotide strand. This method uses DNA oligonucleotide aptamers that bind either to a specific protein selectively or to a complementary DNA oligonucleotide reversibly using toehold-mediated DNA strand-displacement. It is possible to choose the sequence of output strand almost independent of the sensing protein. Using this strategy, we implemented four different exchange processes to report the concentrations of clinically relevant human α-thrombin and vascular endothelial growth factor using changes in concentrations of DNA oligonucleotide outputs. These exchange processes can operate in tandem such that the same or different output signals can indicate changes in concentration of distinct or identical input proteins. The simplicity of our approach suggests a pathway to build devices that can direct diverse output responses in response to changes in concentrations of specific proteins.

Phase 1 LymfactinⓇ Study: Short-term Safety of Combined Adenoviral VEGF-C and Lymph Node Transfer Treatment for Upper Extremity Lymphedema

To study the safety and tolerability of LymfactinⓇ treatment combined with microvascular lymph node transfer surgery in patients with upper limb lymphedema. Upper limb lymphedema is a common clinical challenge after breast cancer surgery and/or radiotherapy. LymfactinⓇ is an adenovirus type 5-based gene therapy involving expression of human vascular endothelial growth factor C (VEGF-C) in the damaged tissue. It aims to correct deficient lymphatic flow by promoting the growth and repair of lymphatic vessels. In Phase I, LymfactinⓇ was combined with microvascular lymph node transfer surgery to study the safety and tolerability of LymfactinⓇ and the biodistribution of the viral vector in patients with upper limb lymphedema. Fifteen patients with breast cancer-associated secondary lymphedema of the upper arm were recruited between December 2016 and February 2018. Three patients received a lower dose (1 × 1010) and 12 a higher dose (1 × 1011) of viral particles, respectively. No dose-limiting toxicities were observed, and the study was completed with the pre-determined maximum dose. Commonly reported adverse events during the 12-month follow-up were common cold, fever, gastroenteritis, pain in the operation area, headache, muscle ache and elevated liver enzymes. Serious adverse events consisted of two erysipelas infections in the lymphedema arm (requiring hospitalization) and one hematoma of the flap donor site. After 12 months' follow-up, results indicate that LymfactinⓇ is well tolerated. The study continues with a 36-months efficacy and 5 years safety follow-up of the patients. The oncological safety aspects of LymfactinⓇ will require a longer follow-up period.

Experimental abdominal aortic aneurysm growth is inhibited by blocking the JAK2/STAT3 pathway

BackgroundThe JAK/STAT pathway is a vital transcription signaling pathway that regulates gene expression and cellular activity. Our recently published study highlighted the role of IL-17A in abdominal aortic aneurysm (AAA) formation and rupture. IL-17A has been proven to upregulate vascular endothelial growth factor (VEGF) expression in some diseases. However, no study has demonstrated the relationships among JAK2/STAT3, IL-17A and VEGF. Therefore, we hypothesized that IL-17A may up-regulate VEGF expression via the JAK2/STAT3 signaling pathway to amplify the inflammatory response, exacerbate neovascularization, and accelerate AAA progression. MethodsTo fully verify our hypothesis, two separate studies were performed: i) a study investigating the influence of JAK2/STAT3 on AAA formation and progression. ii) a study evaluating the relationship among IL-17A, JAK2/STAT3 and VEGF. Human tissues were collected from 7 AAA patients who underwent open surgery and 7 liver transplantation donors. All human aortic tissues were examined by histological and immunohistochemical staining, and Western blotting. Furthermore, mouse aortic tissues were also examined by histological and immunohistochemical staining and Western blotting, and the mouse aortic diameters were assessed by high-resolution Vevo 2100 microimaging system. ResultsAmong human aortic tissues, JAK2/STAT3, IL-17A and VEGF expression levels were higher in AAA tissues than in control tissues. Group treated with WP1066 (a selective JAK2/STAT3 pathway inhibitor), IL-17A, and VEGF groups had AAA incidences of 25%, 40%, and 65%, respectively, while the control group had an incidence of 75%. Histopathological analysis revealed that the IL-17A- and VEGF-related inflammatory responses were attenuated by WP1066. Thus, blocking the JAK2/STAT3 pathway with WP1066 attenuated experimental AAA progression. In addition, in study ii, we found that IL-17A siRNA seemed to attenuate the expression of IL-17A and VEGF in vivo study; treatment with VEGF siRNA decreased the expression of VEGF, while IL-17A expression remained high. In an in vitro study, rhIL-17A treatment increased JAK2/STAT3 and VEGF expression in macrophages in a dose-dependent manner. ConclusionBlocking the JAK2/STAT3 pathway with WP1066 (a JAK2/STAT3 specific inhibitor) attenuates experimental AAA progression. During AAA progression, IL-17A may influence the expression of VEGF via the JAK2/STAT3 signaling pathway. This potential mechanism may suggest a novel strategy for nonsurgical AAA treatment.

Exploring the Anti‐Cancer Properties of Efavirenz, in Comparison to Thalidomide

IntroductionIt has been suggested that “highly active antiretroviral therapy” (HAART) caused resolution of secondary pulmonary cancers. The question then is: ‘Do antiretroviral drugs (ARD) possess anti‐cancer properties, and if so, then how? Efavirenz, a Non‐Nucleoside Reverse Transcriptase Inhibitor (NNRTI) and a first line drug used to manage HIV‐1 infection in both adults and children, is hereby investigated, in comparison with the known anti‐angiogenic and anti‐cancer drug, thalidomide.MethodologyEmbryonic CAMs of 30 fertile eggs of the domestic fowl (Gallus gallus variant Domesticus) were used as in vivo vascular test environment as follows: they were randomly allocated (n=6) to Human recombinant VEGF (R&amp;D Systems), Thalidomide (Sigma‐Aldrich), Control 1 group treated with 70% ethanol (diluent), Control 2 group of embryos that were not manipulated and Efavirenz, (Sequoia Research Products).ResultsEfavirenz suppressed angiogenesis in Chick CAM in 100% of the subjects, showing more anti‐angiogenic properties than thalidomide. Embryonic viability (as a product of CAM angiogenesis) in all the experimental groups (n=6) was as follows: Control 6 (100%); VEGF 5 (83.3%); Thalidomide 1 (17%) and Efavirenz 0 (0.0%). Thalidomide failed to suppress angiogenesis in 17% of the subjects. Moreover, unlike thalidomide, Efavirenz also suppressed erythropoiesis.Discussion and ConclusionThe role of vascular endothelial growth factor (VEGF) in both developmental and pathological angiogenesis has been studied (e.g. Ferrara, 2000; Hagedorn et al. 2004; Losordo et al., 2004; Maharaj, 2006; Makanya et al., 2007). VEGF is a specific and potent mitogen for endothelial cells that is expressed in a distinct temporo‐spatial pattern during mammalian lung development. Since cancer cell growth and viability depends on neo‐vascularization, this property of Efavirenz, which is antagonistic to VEGF activities, may cause spontaneous resolution of HIV‐associated cancers in patients placed on HAART. In a future work, the mechanism of action of this notable ARD will be further investigated.Support or Funding InformationSupported by Sefako Makgatho Health Sciences University (SMU), Ga‐Rankuwa, Gauteng province of South Africa CAM Images CAM IMAGESCAM ImagesFigure 1 Results Group Mean Angiogenic effect Standard Deviation F P‐value VEGF 0.83 0.41 Thalidomide 0.17 0.41 9.306 *0.0005 Efavirenz 0 0 Control 1 0.67 0.52 Control 2 1 0 *indicates the p value is significant, p &lt; 0.005. Possessing 17% angiogenicity implies the thalidomide failed as an anti‐angiogenic drug in the one embryo. Moreover, one of the embryos the thalidomide group reached the end of the experiment but died within seconds of exposure. CAM had one short thriving blood vessel on the CAM surface. In Efavirenz‐treated CAMS, no blood vessels were seen and neither was there any sign of erythropoiesis.

In Vitro Selection of New DNA Aptamers for Human Vascular Endothelial Growth Factor 165

Two DNA aptamers that bind the heparin-binding domain (HBD) of the human vascular endothelial growth factor 165 (VEGF-165) have been previously reported. Although VEGF-165 is a homodimeric protein and the two aptamers have different sequences and secondary structures, the aptamers appear to occupy the same binding site and cannot form a 2 : 1 aptamer/protein complex, thus making them unsuitable for creating a higher-affinity dimeric DNA aptamer. This has motivated us to conduct a new in vitro selection experiment to search for new VEGF-165-binding DNA aptamers with different properties. We undertook a multistream selection strategy in which the concentration of VEGF-165 was varied significantly. We carried out 11 rounds of selection, and next-generation sequencing was conducted for every round in each stream. From comprehensive sequence analysis, we identified four classes of DNA aptamers, of which two were reported before, but two are new DNA aptamers. One of the new aptamers exhibits a unique property that has never been observed before: it is capable of forming the 2 : 1 aptamer/protein complex with VEGF-165. This work has expanded the repertoire of VEGF-165-binding DNA aptamers and creates a possibility to engineer a higher affinity homodimeric aptamer for VEGF-165.

Specific humoral response in cancer patients treated with a VEGF-specific active immunotherapy procedure within a compassionate use program

BackgroundCIGB-247 is a cancer therapeutic vaccine that uses as antigen a variant of human vascular endothelial growth factor (VEGF) mixed with the bacterially-derived adjuvant VSSP. CIGB-247 has been already evaluated in two phase I clinical trials (CENTAURO and CENTAURO-2), showing to be safe and immunogenic in advanced cancer patients selected under well-defined and controlled clinical conditions. Surviving patients were submitted to monthly re-immunizations and some of them showed objective clinical benefits. Based on these results, a compassionate use program (CUP) with CIGB-247 was initiated for patients that did not meet the strict entry criteria applied for the CENTAURO and CENTAURO-2 clinical trials, but could potentially benefit from the application of this cancer therapeutic vaccine.ResultsPolyclonal IgM, IgA and IgG antibodies specific for VEGF were detected by ELISA in serum samples from patients vaccinated with 400 μg of antigen combined with 200 μg of VSSP. Polyclonal antibody response showed no cross reactivity for other VEGF family member molecules like VEGF-C and VEGF-D. Serum from immunized individuals was able to block the binding of VEGF to its receptors VEGFR2 and VEGFR1. IgG fraction purified from immune sera shared the aforementioned characteristics and also inhibited the interaction between VEGF and the therapeutic recombinant antibody bevacizumab, an anti-angiogenic drug approved for the treatment of different tumors. No serious adverse events attributable to CIGB-247 have been documented yet in participants of the CIGB-247 CUP.The present paper is a first report of our findings concerning the humoral response and safety characteristics in treated CIGB-247 CUP cancer patients. The study has provided the unique opportunity of not only testing CIGB-247 in a broader clinical spectrum sample of Cuban cancer patients, but also within the context of the day-to-day clinical practice and treatment settings for these diseases in Cuban medical institutions.ConclusionsThe CIGB-247 CUP has demonstrated that immunization and follow-up of a variety of cancer patients, under day-to-day clinical practice conditions in several Cuban medical institutions, replicate our previous findings in clinical trials: CIGB-247 is safe and immunogenic.

Neutralization of Bombina variegata peptide 8 suppresses retinal neovascularization in two different murine models: The oxygen-induced retinopathy model and the rhodopsin promoter/VEGF transgenic mouse model

Bombina variegata 8 (Bv8), also known as prokineticin-2 (PK-2), is a potent pro-angiogenic factor. However, its role in retinal neovascularization (RNV) remains unknown. In this study, we explored the role of Bv8 in the pathogenesis of RNV. We found that the expression of Bv8 was significantly increased in two different models of retinal neovascularization: the oxygen-induced retinopathy (OIR) mouse model and the rhodopsin promoter (rho)/VEGF transgenic mouse model. Neutralization of Bv8 by intravitreal injections of its antibody, not only inhibited retinal and subretinal neovascularization but also decreased the mRNA and protein levels of several pro-angiogenic factors. Our in vitro assay showed that recombinant human Bv8 (RhBv8) protein promoted human retinal microvascular endothelial cells (HRECs) tube-formation, cell proliferation and vascular endothelial growth factor receptor 1 (VEGFR1) and receptor 2 (VEGFR2) expression. Our findings suggest that Bv8 could be used as a novel target for the treatment of RNV-related ocular diseases.

Carbon Dots Conjugated with Vascular Endothelial Growth Factor for Protein Tracking in Angiogenic Therapy.

One of the challenges of using growth factors for tissue regeneration is to monitor their biodistributions and delivery to injured tissues for minimally invasive detection. In the present study, tracking of human vascular endothelial growth factor (VEGF) was achieved by chemically linking it to photoluminescent carbon dots (CDs). Carbon dots were synthesized by the hydrothermal method and, subsequently, conjugated with VEGF using carbodiimide coupling. ELISA and western blot analysis revealed that VEGF-conjugated CDs preserve the binding affinity of VEGF to its antibodies. We also show that VEGF-conjugated CDs maintain the functionality of VEGF for tube formation and cell migration. The VEGF-conjugated CDs were also used for in vitro imaging of human umbilical vein endothelial cells. The results of this work suggest that cell-penetrating VEGF-conjugated CDs can be used for growth factor protein tracking in therapeutic and tissue engineering applications.