Combined inhibition of MET and VEGF enhances therapeutic efficacy of EGFR TKIs in EGFR-mutant non-small cell lung cancer with concomitant aberrant MET activation.

BACKGROUND: Aberrant activation of mesenchymal epithelial transition (MET) has been considered to mediate primary and acquired resistance to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) in EGFR-mutant non-small cell lung cancer (NSCLC). However, mechanisms underlying this process are not wholly clear and the effective therapeutic strategy remains to be determined. METHODS: The gefitinib-resistant NSCLC cell lines were induced by concentration increase method in vitro. Western blot and qPCR were used to investigate the relationship between MET and vascular endothelial growth factor (VEGF)/VEGF receptor 2 (VEGFR2) signaling pathway. Double luciferase reporter gene and co-immunoprecipitation were used to further reveal the regulation mechanism between MET and VEGF/VEGFR2. The effect of combined inhibition of MET and VEGF/VEGFR2 signaling pathway on the therapeutic sensitivity of EGFR-TKI in gefitinib resistant cell lines with MET aberration was verified ex vivo and in vivo. RESULTS: We successfully obtained two gefitinib-resistant NSCLC cell lines with EGFR mutation and abnormal activation of MET. We observed that MET formed a positive feedback loop with the VEGF/VEGFR2 signaling, leading to persistent downstream signaling activation. Specifically, MET up-regulated VEGFR2 expression in a MAPK/ERK/ETS1-dependent manner, while VEGF promoted physical interaction between VEGFR2 and MET, thereby facilitating MET phosphorylation. A MET inhibitor, crizotinib, combined with an anti-VEGF antibody, bevacizumab, enhanced the sensitivity of NSCLC cells to gefitinib and synergistically inhibited the activation of downstream signaling in vitro. Dual inhibition of MET and VEGF combined with EGFR TKIs markedly restrained tumor growth in both human NSCLC xenograft models and in an EGFR/MET co-altered case. CONCLUSIONS: Our work reveals a positive feedback loop between MET and VEGF/VEGFR2, resulting in continuous downstream signal activation. Combined inhibition of MET and VEGF/VEGFR2 signaling pathway may be beneficial for reversing EGFR TKIs resistance.

ß-arrestin2 promotes angiogenesis of liver sinusoidal endothelial cells through the VEGF/VEGFR2 pathway to aggravate cirrhosis.

Excessive extracellular matrix deposition and increased intrahepatic angiogenesis are prominent features of cirrhosis. ß-arrestin2 is thought to be involved in the pathological processes of various fibrotic diseases. This study aimed to investigate the role and possible mechanism of ß-arrestin2 in the angiogenesis of cirrhosis. Firstly, ß-arrestin2 expression in liver tissues of cirrhotic patients was detected, and the correlation between ß-arrestin2 and α-SMA, CD-31, PDGF, and VEGF indexes was analyzed. Then, after liver cirrhosis induced by CCL4 in Arrb2-KO mice (ß-arrestin2 coding gene), liver histopathological changes were observed, and the expressions of α-SMA, CD-31, PDGF, VEGF, and VEGFR2 were detected. Finally, VEGF-A was used to treat human liver sinusoidal endothelial cells (LSECs) to simulate pathological conditions. After transfection with si-ARRB2, the cell activity, MDA and GSH-PX activities, cell invasion, angiogenesis, and the expressions of α-SMA, CD-31, and VEGF/VEGFR2 pathway were detected. Results showed that ß-arrestin2 expression in the liver increased significantly during cirrhosis and was positively correlated with angiogenesis. In vivo, Arrb2-KO significantly inhibited fibrosis and angiogenesis in cirrhotic mice, and decreased the expressions of α-SMA, CD31, PDGF, VEGF, and VEGFR2. Studies using LSECs in vitro showed that after intervention of ARRB2, the activity of LSECs and the number of invasions and tubule formations were significantly reduced. Similarly, after transfection with si-ARRB2, the expressions of α-SMA, CD31, PDGF, VEGF, and VEGFR2 in LSECs were significantly decreased. Collectively, ß-arrestin2 aggravated cirrhosis by promoting the angiogenesis of LSECs. Blocking ß-arrestin2 may be an important target against angiogenesis and fibrosis in cirrhosis.

Development of bioassay platforms for biopharmaceuticals using Jurkat-CAR cells by AICD.

The assessment of bioactivity for therapeutic antibody release assay poses challenges, particularly when targeting immune checkpoints. An in vitro bioassay platform was developed using the chimeric antigen receptor on Jurkat cells (Jurkat-CAR) to analyze antibodies targeting immune checkpoints, such as CD47/SIRPα, VEGF/VEGFR1, PD-1/PD-L1, and CD70/CD27. For CD47/SIRPα, the platform involved a Jurkat-CAR cell line expressing the chimeric SIRPα receptor (CarSIRPα). CarSIRPα was created by sequentially fusing the SIRPα extracellular region with the CD8α hinge region, the transmembrane (TM) and intracellular (IC) domains of CD28, and the intracellular signaling domain of CD3ζ. The resulting Jurkat-CarSIRPα cells can undergo "activation-induced cell death (AICD)" upon incubation with purified or cellular CD47, as evidenced by the upregulation of CD69, IL-2, and IFN-γ. Similar results also appeared in Jurkat CarVEGFR1, Jurkat CarPD1 and Jurkat CARCD27 cells. These cells are perfectly utilized for the bioactivity analysis of therapeutic antibody. Our study indicates that the established in vitro assay platform based on Jurkat-CAR has been confirmed repeatedly and has shown robust reproducibility; thus, this platform can be used for screening or for release assays of given antibody drugs targeting immune checkpoints.

The Impact of the VEGF/VEGFR2/PI3K/AKT Signaling Axis on the Proliferation and Migration Abilities of Human Dental Pulp Stem Cells.

The potential therapeutic benefits of human dental pulp stem cells (HDPSCs) in dental regenerative medicine have been demonstrated. However, little is known about the molecular mechanisms regulating the biological characteristics of HDPSCs. The experiment aims to explore whether VEGF activates signaling pathways such as FAK, PI3K, Akt, and p38 in HDPSCs, and to investigate the molecular mechanisms by which VEGF influences proliferation and migration of HDPSCs. Normal and inflamed human dental pulp (HDP) samples were collected, and the levels of VEGF in HDP were assessed. HDPSCs were cultured and purified. HDPSCs were stimulated with lipopolysaccharide (LPS) at gradient concentrations, and real-time quantitative polymerase chain reaction (qPCR) was used to assess changes in VEGF mRNA. Gradient concentrations of VEGF were used to stimulate HDPSCs, and cell migration ability was evaluated through scratch assays and Transwell chamber experiments. Phosphorylation levels of FAK, AKT, and P38 were assessed using Western blotting. Inhibitors of VEGFR2, FAK, AKT, P38, and VEGF were separately applied to HDPSCs, and cell migration ability and phosphorylation levels of FAK, AKT, and P38 were determined. The results indicated significant differences in VEGF levels between normal and inflamed HDP tissues, with levels in the inflamed state reaching 435% of normal levels (normal: 87.91 ng/mL, inflamed: 382.76 ng/mL, P < 0.05). LPS stimulation of HDPSCs showed a significant increase in VEGF mRNA expression with increasing LPS concentrations (LPS concentrations of 0.01, 0.1, 1, and 10 µg/mL resulted in VEGF mRNA expressions of 181.2%, 274.2%, 345.8%, and 460.9%, respectively, P < 0.05). VEGF treatment significantly enhanced the migration ability of HDPSCs in Transwell chamber experiments, with migration rates increasing with VEGF concentrations (VEGF concentrations of 0, 1, 10, 20, 50, and 100 ng/mL resulted in migration rates of 8.41%, 9.34%, 21.33%, 28.41%, 42.87%, and 63.15%, respectively, P < 0.05). Inhibitors of VEGFR2, FAK, AKT, P38, and combined VEGF stimulation demonstrated significant migration inhibition, with migration rates decreasing to 8.31%, 12.64%, 13.43%, 18.32%, and 74.17%, respectively. The migration rate with combined VEGF stimulation showed a significant difference (P < 0.05). The analysis of phosphorylation levels revealed that VEGF stimulation significantly activated phosphorylation of FAK, AKT, and P38, with phosphorylation levels increasing with VEGF concentrations (P < 0.05). The VEGF/VEGFR2 signaling axis regulated the migration ability of HDPSCs through the FAK/PI3K/AKT and P38MAPK pathways. This finding highlighted not only the crucial role of VEGF in injury repair of HDPSCs but also provided important clues for a comprehensive understanding of the potential applications of this signaling axis in dental regenerative medicine.

Polyphyllin I ameliorates gefitinib resistance and inhibits the VEGF/VEGFR2/p38 pathway by targeting HIF-1a in lung adenocarcinoma.

BACKGROUND: Lung adenocarcinoma (LUAD) is the most common pathological type of lung cancer. Epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) have been administered as the first-line therapy for patients with EGFR mutations in LUAD, but it is almost inevitable that resistance to EGFR-TKIs therapy eventually arises. Polyphyllin I (PPI), derived from Paris polyphylla rhizomes, has been shown to have potent anti-cancer properties in a range of human cancer types including LUAD. However, the role of PPI in gefitinib resistance and the underlying mechanism remain elusive. PURPOSE: To evaluate the antitumor impacts of PPI on gefitinib resistance cells and investigate its molecular mechanism. METHODS: CCK-8, wound healing, transwell assay, and xenograft model were performed to determine the anti-cancer effects of PPI as well as its ability to overcome gefitinib resistance. Immunoblotting, co-immunoprecipitation, phospho-RTK antibody array, qRT-PCR, and immunofluorescence were utilized to explore the mechanism by which PPI overrides gefitinib resistance. RESULTS: PPI inhibited cell survival, growth, and migration/invasion in both gefitinib-sensitive (PC9) and -resistant (PC9/GR) LUAD cells (IC50 at 2.0 µM). Significantly, treatment with PPI at 1.0 µM resensitized the resistant cells to gefitinib. Moreover, cell-derived xenograft experiments revealed that the combination of PPI and gefitinib overcame gefitinib resistance. The phospho-RTK array and immunoblotting analyses showed PPI significant inhibition of the VEGFR2/p38 pathway. In addition, molecular docking suggested the interaction between PPI and HIF-1α. Mechanistically, PPI reduced the protein expression of HIF-1α in both normoxia and hypoxia conditions by triggering HIF-1α degradation. Moreover, HIF-1α protein but not mRNA level was elevated in gefitinib-resistant LUAD. We further demonstrated that PPI considerably facilitated the binding of HIF-1α to VHL. CONCLUSIONS: We present a novel discovery demonstrating that PPI effectively counteracts gefitinib resistance in LUAD by modulating the VEGF/VEGFR2/p38 pathway. Mechanistic investigations unveil that PPI facilitates the formation of the HIF-1α /VHL complex, leading to the degradation of HIF-1α and subsequent inhibition of angiogenesis. These findings uncover a previously unidentified mechanism governing HIF-1α expression in reaction to PPI, providing a promising method for therapeutic interventions targeting EGFR-TKI resistance in LUAD.

Suppression of inflammation in ulcerative colitis rats by avocado and pomegranate.

BACKGROUND: Pomegranate seed oil (PSO) and avocado seed oil (ASO) are natural polyphenols with established anti-inflammatory activity. PURPOSE: This study aimed to investigate the molecular mechanisms underlying the therapeutic efficacy of PSO and ASO in experimental ulcerative colitis (UC) with reference to sulfasalazine (SLZ). METHODS: Eighty male albino rats were divided equally into 8 groups; Normal, PSO, ASO, SLZ, UC-control, (UC + PSO), (UC + ASO) and (UC + SLZ) groups. Colitis was induced by intra-rectal injection of acetic acid. PSO (0.5ml/200g), ASO (1ml/250g) and SLZ (100 mg/kg) were administered orally once/day for 14 days, 24h after colitis induction. Colitis was evaluated by measuring disease activity index (DAI), colon weight/length ratio and histologic inflammatory score. Vascular endothelial growth factor receptor-2 (VEGFR-2), colonic macrophage migration inhibitory factor (MIF), and malondialdehyde (MDA) were determined. Colonic gene expression of TNF-α, VEGF and heme oxygenase-1 (HO-1) were also estimated. RESULTS: PSO and ASO treatments to UC rats significantly reduced DAI, weight/length ratio, VEGFR-2, and colon histologic inflammatory score versus UC-controls. ASO significantly suppressed MIF levels and TNF-α expression greater than PSO. However, PSO was more significant than ASO in reducing MDA levels and up-regulating HO-1 expression. Both oils significantly down-regulated VEGF expression. The obtained biochemical and histological changes induced by UC were nearly corrected by SLZ. CONCLUSION: The proved beneficial effect of PSO and ASO as anti-inflammatory, anti-angiogenic, and antioxidant in UC rats could be mediated by suppression of TNF-α, VEGF, and MIF and up-regulation of HO-1.

Metabolic Syndrome and Cardiac Vessel Remodeling Associated with Vessel Rarefaction: A Possible Underlying Mechanism May Result from a Poor Angiogenic Response to Altered VEGF Signaling Pathways.

BACKGROUND: Elevated mortality rates in patients with metabolic syndrome (MetS) are partly due to adverse remodeling of multiple organs, which may lead to cardiovascular disease, nonalcoholic fatty liver disease, kidney failure, or other conditions. MetS symptoms, such as obesity, hypertension, hyperglycemia, dyslipidemia, associated with insulin and leptin resistance, are recognized as major cardiovascular risk factors that adversely affect the heart. SUMMARY: Pathological cardiac remodeling is accompanied by endothelial cell dysfunction which may result in diminished coronary flow, dysregulated oxygen demand/supply balance, as well as vessel rarefaction. The reduced number of vessels and delayed or inhibited formation of collaterals after myocardial infarction in MetS heart may be due to unfavorable changes in endothelial cell metabolism but also to altered expression of vascular endothelial growth factor molecules, their receptors, and changes in signal transduction from the cell membrane, which severely affect angiogenesis. KEY MESSAGES: Given the established role of cardiac vessel endothelial cells in maintaining tissue homeostasis, defining the molecular background underlying vessel dysfunction associated with impaired angiogenesis is of great importance for future therapeutic purposes. Therefore, the aim of this paper was to present current information regarding vascular endothelial growth factor signaling in the myocardium of MetS individuals.

A case of primary pulmonary angiosarcoma arising from tuberculous scar with fatal capillary bleeding.

The etiology of primary pulmonary angiosarcoma is still unknown. Here we report a case of primary pulmonary angiosarcoma originated from a tuberculous scar and presented as aggressive deterioration with uncontrolled bleeding from capillaries with angiodysplasia.

Efficacy and safety of VEGF/VEGFR inhibitors for platinum-resistant ovarian cancer: a systematic review and meta-analysis of randomized controlled trials.

BACKGROUND: Almost all patients with ovarian cancer will experience relapse and eventually develop platinum-resistant. The poor prognosis and limited treatment options have prompted the search for novel approaches in managing platinum-resistant ovarian cancer (PROC). Therefore, a meta-analysis was conducted to evaluate the efficacy and safety of combination therapy with vascular endothelial growth factor (VEGF) /VEGF receptor (VEGFR) inhibitors for PROC. METHODS: A comprehensive search of online databases was conducted to identify randomized clinical trials published until December 31, 2022. Pooled hazard ratios (HR) was calculated for overall survival (OS) and progression-free survival (PFS), while pooled odds ratio (OR) was calculated for objective response rate (ORR) and treatment-related adverse events (TRAEs). Subgroup analysis was further performed to investigate the source of heterogeneity. RESULTS: In total, 1097 patients from eight randomized clinical trials were included in this meta-analysis. The pooled HRs of OS (HR = 0.72; 95% CI: 0.62-0.84, p < 0.0001) and PFS (HR = 0.52; 95% CI: 0.45-0.59, p < 0.0001) demonstrated a significant prolongation in the combination group compared to chemotherapy alone for PROC. In addition, combination therapy demonstrated a superior ORR compared to monotherapy (OR = 2.34; 95%CI: 1.27-4.32, p < 0.0001). Subgroup analysis indicated that the combination treatment of VEGF/VEGFR inhibitors and chemotherapy was significantly more effective than monochemotherapy in terms of OS (HR = 0.71; 95% CI: 0.61-0.84, p < 0.0001), PFS (HR = 0.49; 95% CI: 0.42-0.57, p < 0.0001), and ORR (OR = 2.97; 95% CI: 1.89-4.67, p < 0.0001). Although the combination therapy was associated with higher incidences of hypertension, mucositis, proteinuria, diarrhea, and hand-foot syndrome compared to monochemotherapy, these toxicities were manageable and well-tolerated. CONCLUSIONS: The meta-analysis demonstrated that combination therapy with VEGF/VEGFR inhibitors yielded better clinical outcomes for patients with PROC compared to monochemotherapy, especially when combined with chemotherapy. This analysis provides more treatment options for patients with PROC. SYSTEMATIC REVIEW REGISTRATION: [ https://www.crd.york.ac.uk/PROSPERO ], Prospective Register of Systematic Reviews (PROSPERO), identifier: CRD42023402050.

VEGF signaling: Role in angiogenesis and beyond.

Angiogenesis is a crucial process for tissue development, repair, and tumor survival. Vascular endothelial growth factor (VEGF) is a key driver secreted by cancer cells, promoting neovascularization. While VEGF's role in angiogenesis is well-documented, its influence on the other aspects in tumor microenvironemt is less discussed. This review elaborates on VEGF's impact on intercellular interactions within the tumor microenvironment, including how VEGF affects pericyte proliferation and migration and mediates interactions between tumor-associated macrophages and cancer cells, resulting in PDL-1-mediated immunosuppression and Nrf2-mediated epithelial-mesenchymal transition. The review discusses VEGF's involvement in intra-organelle crosstalk, tumor metabolism, stemness, and epithelial-mesenchymal transition. It also provides insights into current anti-VEGF therapies and their limitations in cancer treatment. Overall, this review aims to provide a thorough overview of the current state of knowledge concerning VEGF signaling and its impact, not only on angiogenesis but also on various other oncogenic processes.

Effect and mechanism of Panax notoginseng saponins on wound healing after anal fistula surgery in rats / 中国药房

OBJECTIVE To investigate the effect and mechanism of Panax notoginseng saponins （PNS） on wound healing after anal fistula surgery in rats by regulating the hypoxia-inducible factor-1α （HIF-1α）/vascular endothelial growth factor （VEGF）/ vascular endothelial growth factor receptor-2 （VEGFR2） signaling pathway. METHODS SD rats were selected to establish a postoperative rat model of anal fistula by infecting wound with Escherichia coli. The model rats were randomly grouped into model group， PNS low-dose and high-dose groups （15， 30 mg/cm2）， high-dose of PNS+2-methoxyestradiol （2ME2） group （PNS 30 mg/cm2+HIF-1α inhibitor 2ME2 4 mg/kg）， with 10 rats in each group. Another 10 normal rats were selected for back hair removal treatment as the control group. Each drug group was injected with the corresponding drug solution intramuscularly or （and） intraperitoneally， once a day， for 3 weeks. After the last administration， the wound healing rate （excluding the control group）， microvascular density （MVD）， the expression of collagen Ⅰ and fibronectin （FN） in the wound tissue were detected in each group； the levels of angiogenic factors [VEGF， E-mail：842710813@qq.com angiopoietin-Ⅰ （Ang-Ⅰ）， Ang-Ⅱ] in serum， the levels of inflammatory factors [interleukin-6 （IL-6） and IL-2] in serum binggui7183@163.com and wound tissue as well as the expressions of the related proteins of HIF-1α/VEGF/VEGFR2 signaling pathway in the wound tissue of rats were also detected in each group. RESULTS The MVD， the expression of collagen Ⅰ and FN in the wound tissue， and the levels of IL-6 and IL-2 in serum and wound tissue of rats increased significantly in the model group， compared to the control group （P＜0.05）， while the serum levels of VEGF， Ang- Ⅰ and Ang-Ⅱ decreased significantly （P＜0.05）. The wound healing rate， the MVD in wound tissue， the serum levels of VEGF， Ang-Ⅰ and Ang-Ⅱ， the expressions of collagen Ⅰ and FN in the wound tissue， and protein expressions of HIF-1α， VEGF and VEGFR2 in the PNS low-dose and high-dose groups increased significantly， compared to the model group （P＜0.05）， while the levels of IL-6 and IL-2 in serum and wound tissue decreased significantly （P＜0.05）； the high-dose PNS had a stronger effect （P＜ 0.05）. 2ME2 could weaken the effect of PNS on above indicators of rats after anal fistula surgery （P＜0.05）. CONCLUSIONS PNS can promote the production of angiogenic factors and inhibit the production of pro-inflammatory factors， thereby promoting wound healing in rats after anal fistula surgery. The above effects are related to the activation of HIF-1α/VEGF/VEGFR2 signaling pathway.

Saponin of Aralia taibaiensis promotes angiogenesis through VEGF/VEGFR2 signaling pathway in cerebral ischemic mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Aralia taibaiensis is known for its ability to promote blood circulation and dispel blood stasis, activate meridians and remove arthralgia. The saponins of Aralia taibaiensis (sAT) are the main active components that are often used to treat cardiovascular and cerebrovascular diseases. However, it has not been reported whether sAT can improve ischemic stroke (IS) by promoting angiogenesis. AIM OF THE STUDY: In this study, we investigated the potential of sAT to promote post-ischemic angiogenesis in mice and determined the underlying mechanism through in vitro experiments. METHODS: To establish the middle cerebral artery occlusion (MCAO) mice model in vivo. First of all, we examined the neurological function, brain infarct volume, and degree of brain swelling in MCAO mice. We also observed pathological changes in brain tissue, ultrastructural changes in blood vessels and neurons, and the degree of vascular neovascularization. Additionally, we established the oxygen-glucose deprivation/reoxygenation (OGD/R) -human umbilical vein endothelial cells (HUVECs) model in vitro to detect the survival, proliferation, migration and tube formation of OGD/R HUVECs. Finally, we verified the regulatory mechanism of Src and PLCγ1 siRNA on sAT promoting angiogenesis by cell transfection technique. RESULTS: In the cerebral ischemia-reperfusion mice, sAT distinctly improved the cerebral infarct volume, brain swelling degree, neurological dysfunction, and brain histopathological morphology due to cerebral ischemia/reperfusion injury. It also increased the double positive expression of BrdU and CD31 in brain tissue, promoted the release of VEGF and NO and decreased the release of NSE and LDH. In the OGD/R HUVECs, sAT significantly improved cell survival, proliferation, migration and tube formation, promoted the release of VEGF and NO, and increased the expression of VEGF, VEGFR2, PLCγ1, ERK1/2, Src and eNOS. Surprisingly, the effect of sAT on angiogenesis was inhibited by Src siRNA and PLCγ1 siRNA in OGD/R HUVECs. CONCLUSION: The results proved that sAT promotes angiogenesis in cerebral ischemia-reperfusion mice and its mechanism is to regulate VEGF/VEGFR2 and then regulate Src/eNOS and PLCγ1/ERK1/2.

Application of VEGF/VEGFR peptide vaccines in cancer: A systematic review of clinical trials.

Peptide vaccines that target vascular endothelial growth factor (VEGF) pathway have shown promising results in inducing strong anti-tumor immune responses with minimal toxicity in various clinical studies. This systematic review was conducted to provide a comprehensive evaluation of the therapeutic efficacy, immune response, survival rate, and side effects of VEGF/VEGF receptor-based peptide vaccines. VEGF/VEGFR2 peptide vaccines were found to be safe and effective in inducing anti-tumor immune responses, while induced moderate clinical benefit. In this regard, further clinical trials are necessary to fully evaluate their clinical effects and the exact correlation between induction of immune response and clinical outcomes.

The neurovasculature as a target in temporal lobe epilepsy.

The blood-brain barrier (BBB) is a physiological barrier maintaining a specialized brain micromilieu that is necessary for proper neuronal function. Endothelial tight junctions and specific transcellular/efflux transport systems provide a protective barrier against toxins, pathogens, and immune cells. The barrier function is critically supported by other cell types of the neurovascular unit, including pericytes, astrocytes, microglia, and interneurons. The dysfunctionality of the BBB is a hallmark of neurological diseases, such as ischemia, brain tumors, neurodegenerative diseases, infections, and autoimmune neuroinflammatory disorders. Moreover, BBB dysfunction is critically involved in epilepsy, a brain disorder characterized by spontaneously occurring seizures because of abnormally synchronized neuronal activity. While resistance to antiseizure drugs that aim to reduce neuronal hyperexcitability remains a clinical challenge, drugs targeting the neurovasculature in epilepsy patients have not been explored. The use of novel imaging techniques permits early detection of BBB leakage in epilepsy; however, the detailed mechanistic understanding of causes and consequences of BBB compromise remains unknown. Here, we discuss the current knowledge of BBB involvement in temporal lobe epilepsy with the emphasis on the neurovasculature as a therapeutic target.

Fruquintinib/HMPL-013 ameliorates cognitive impairments and pathology in a mouse model of cerebral amyloid angiopathy (CAA).

Cerebral amyloid angiopathy (CAA) is characterized by the cerebrovascular amyloid-ß (Aß) accumulation, and always accompanied by Alzheimer's disease (AD). The mechanisms revealing CAA pathogenesis are still unclear, and it is challenging to develop an efficient therapeutic strategy for its treatment. Vascular endothelial growth factor (VEGF) and its receptors including VEGFR-1,-2,-3 activation are involved in Aß processing, and modulate numerous cellular events associated with central nervous system (CNS) diseases. In the present study, we attempted to explore the regulatory function of fruquintinib (also named as HMPL-013), a highly selective inhibitor of VEGFR-1,-2,-3 tyrosine kinases, on CAA progression in Tg-SwDI mice. Here, we found that HMPL-013-rich diet consumption for 12 months significantly improved the behavioral performances and cerebral blood flow (CBF) of Tg-SwDI mice compared with the vehicle group. Importantly, HMPL-013 administration considerably reduced Aß1-40 and Aß1-42 burden in cortex and hippocampus of Tg-SwDI mice through regulating Aß metabolism process. Congo red staining confirmed Aß deposition in vessel walls, reflecting CAA formation, which was, however, strongly ameliorated after HMPL-013 treatment. Neuron death, aberrant glial activation and pro-inflammatory response in brain tissues of Tg-SwDI mice were dramatically alleviated after HMPL-013 consumption. More studies showed that the protective effects of HMPL-013 against CAA might be partially attributed to its regulation on the expression of genes associated with blood vasculature. Intriguingly, VEGF and phosphorylated VEGFR-1,-2 protein expression levels were remarkably decreased by HMPL-013 in cortex and hippocampus of Tg-SwDI mice, which were validated in HMPL-013-treated brain vascular endothelial cells (BVECs) under hypoxia. Finally, we found that VEGF-induced human umbilical vein endothelial cells (HUVEC) proliferation and tube formation were strongly abolished upon HMPL-013 exposure. Collectively, all these findings demonstrated that oral administration of HMPL-013 had therapeutic potential against CAA by reducing Aß deposition, inflammation and neuron death via suppressing VEGF/VEGFR-1,-2 signaling.

Repair Mechanism of Jianpi Huogu Prescription on Vascular Injury in Alcohol-induced Osteonecrosis of Femoral Head Based on VEGF/VEGFR2/PI3K/Akt Signaling Pathway / 中国实验方剂学杂志

MethodIn the experiment， 46% vol Red Star Erguotou （10 mL·kg·d-1） was used to establish the AONFH rat model， and the intervention effect of JPHGP at different doses （2.5， 5.0， 10.0 g·kg-1） was observed. Jiangusheng pill （JGS， 1.53 g·kg-1） was selected as the positive control. After 8 weeks of administration， the bone histomorphometry of the femoral head was analyzed by Micro-CT imaging， and the area of medullary microvessels in the femoral head was detected by ink perfusion. The pathological change was observed by hematoxylin and eosin （HE） staining. The protein expressions of Platelet endothelial cell adhesion molecule-1 （CD31）， VEGF， VEGFR2， PI3K， phosphor-Akt （p-Akt） and phosphatase and Tensin homologue deleted on chromosome 10 （PTEN） in the femoral head were determined by immunohistochemistry and Western blot. ResultCompared with normal group， the model group presented the fracture and thinning of trabeculae in the femoral head， increased empty bone lacunae， and elevated number and diameter of adipocytes （P<0.01）. Micro-CT imaging revealed a decrease in bone mineral density （BMD）， bone volume fraction （BV/TV）， trabecular thickness （Tb.Th） and trabecular number （Tb.N） （P<0.05， P<0.01） while an increase in bone surface-to-volume ratio （BS/BV） and trabecular separation （Tb.Sp） （P<0.01）. The results of ink perfusion showed that the area of medullary microvessels in the femoral head was reduced （P<0.01）. Compared with model group， JPHGP lowered the empty bone lacunae rate as well as the number and diameter of adipocytes in the femoral head of AONFH rats. Micro-CT imaging indicated that JPHGP low-dose group had elevated BV/TV， Tb.Th and Tb.N （P<0.05， P<0.01） while decreased BS/BV （P<0.01）， and there was an upward trend in BMD while a downward trend in Tb.Sp， but without statistical difference. In addition， JPHGP medium- and high-dose groups had a rise in BMD， BV/TV， Tb.Th and Tb.N （P<0.05， P<0.01）， a decrease in BS/BV and Tb.Sp （P<0.05， P<0.01） and enlarged area of medullary microvessels in the femoral head （P<0.05， P<0.01）. The expressions of CD31， VEGF， VEGFR2， PI3K， p-Akt in the model group were lower than those in the normal group （P<0.01）， and after medium and high doses of JPHGP treatment， the expressions of CD31， PI3K and p-Akt in the femoral head of rats were up-regulated （P<0.01） while the protein expression of PTEN was down-regulated （P<0.01）. Moreover， JPHGP up-regulated the expressions of VEGF and VEGFR2 （P<0.05， P<0.01）. ConclusionJPHGP can repair the vascular injury in AONFH， and its mechanism may be related to the activation of VEGF/VEGFR2/PI3K/Akt signaling pathway. This study provides certain scientific basis and reference for the clinical application of JPHGP. ObjecctiveTo observe the repair effect of Jianpi Huogu prescription （JPHGP） on vascular injury in experimental alcohol-induced osteonecrosis of femoral head （AONFH）， and to explore its mechanism based on vascular endothelial growth factor （VEGF）/VEGFR2/phosphoinositide 3-kinase （PI3K）/protein kinase B （Akt） signaling pathway.

Alkaloid Derivative (Z)-3ß-Ethylamino-Pregn-17(20)-en Inhibits Triple-Negative Breast Cancer Metastasis and Angiogenesis by Targeting HSP90α.

Metastasis is an important cause of cancer-related death. Previous studies in our laboratory found that pregnane alkaloids from Pachysandra terminalis had antimetastatic activity against breast cancer cells. In the current study, we demonstrated that treatment with one of the alkaloid derivatives, (Z)-3ß-ethylamino-pregn-17(20)-en (1), led to the downregulation of the HIF-1α/VEGF/VEGFR2 pathway, suppressed the phosphorylation of downstream molecules Akt, mTOR, FAK, and inhibited breast cancer metastasis and angiogenesis both in vitro and in vivo. Furthermore, the antimetastasis and antiangiogenesis effects of 1 treatment (40 mg/kg) were more effective than that of Sorafenib (50 mg/kg). Surface plasmon resonance (SPR) analysis was performed and the result suggested that HSP90α was a direct target of 1. Taken together, our results suggested that compound 1 might represent a candidate antitumor agent for metastatic breast cancer.

Anlotinib exerts potent antileukemic activities in Ph chromosome negative and positive B-cell acute lymphoblastic leukemia via perturbation of PI3K/AKT/mTOR pathway.

OBJECTIVES: Despite advances in the development of novel targeted therapies, the need for B-ALL alternative treatments has not been met. Anlotinib could blunt the proangiogenic activity of VEGFR, PDGFR, and FGFR, and has shown strong antitumor activities across multiple tumors. However, anlotinib cytotoxicity against B-ALL has not ever been evaluated, thus prompting us to initiate this study. METHODS: Expression2Kinases program was used to identify potential treatment targets. Cell viability and apoptosis were determined by CCK-8 and Annexin V/PI staining kit, respectively. qRT-PCR and Western blotting were utilized to investigate the molecular mechanisms. In vivo antileukemia activity of Anlotinib was evaluated in a Ph+ B-ALL patient-Derived Xenograft (PDX) model. RESULTS: Compared with treatment-naive B-ALL cases, RR B-ALL patients had higher activities in the VEGF/VEGFR signaling and the PI3K/AKT/mTOR pathway. Exposure of Ph- and Ph+ B-ALL cells to anlotinib resulted in significant cell viability reduction, apoptosis enhancement, and cell cycle arrest at G2/M phase. Importantly, anlotinib treatment led to remarkably decreased leukemia burdens and extended the survival period in a Ph+ B-ALL PDX model. Blockade of the role of the proangiogenic mediators, comprising VEGFR2, PDGFR-beta, and FGFR3, played a critical role in the cytotoxicity of anlotinib against Ph- and Ph+ B-ALL. Moreover, anlotinib dampened the activity of PI3K/AKT/mTOR pathway that resides in the convergence of the three mentioned proangiogenic signals. CONCLUSION: This work provides impressive preclinical evidence of anlotinib against Ph- and Ph+ B-ALL and raises a rationale for future clinical evaluation of this drug in the management of Ph- and Ph+ B-ALL.

VEGF/VEGFR-Targeted Therapy and Immunotherapy in Non-small Cell Lung Cancer: Targeting the Tumor Microenvironment.

Non-small cell lung cancer (NSCLC) is the leading cause of death by cancer worldwide. Despite developments in therapeutic approaches for the past few decades, the 5-year survival rate of patients with NSCLC remains low. NSCLC tumor is a complex, heterogeneous microenvironment, comprising blood vessels, cancer cells, immune cells, and stroma cells. Vascular endothelial growth factors (VEGFs) are a major mediator to induce tumor microvasculature and are associated with the progression, recurrence, and metastasis of NSCLC. Current treatment medicines targeting VEGF/VEGF receptor (VEGFR) pathway, including neutralizing antibodies to VEGF or VEGFR and receptor tyrosine kinase inhibitors, have shown good treatment efficacy in patients with NSCLC. VEGF is not only an important angiogenic factor but also an immunomodulator of tumor microenvironment (TME). VEGFs can suppress antigen presentation, stimulate activity of regulatory T (Treg) cells, and tumor-associated macrophages, which in turn promote an immune suppressive microenvironment in NSCLC. The present review focuses on the angiogenic and non-angiogenic functions of VEGF in NSCLC, especially the interaction between VEGF and the cellular components of the TME. Additionally, we discuss recent preclinical and clinical studies to explore VEGF/VEGFR-targeted compounds and immunotherapy as novel approaches targeting the TME for the treatment of NSCLC.

Novel Drugs with High Efficacy against Tumor Angiogenesis.

Angiogenesis is involved in physiological and pathological processes in the body. Tumor angiogenesis is a key factor associated with tumor growth, progression, and metastasis. Therefore, there is great interest in developing antiangiogenic strategies. Hypoxia is the basic initiating factor of tumor angiogenesis, which leads to the increase of vascular endothelial growth factor (VEGF), angiopoietin (Ang), hypoxia-inducible factor (HIF-1), etc. in hypoxic cells. The pathways of VEGF and Ang are considered to be critical steps in tumor angiogenesis. A number of antiangiogenic drugs targeting VEGF/VEGFR (VEGF receptor) or ANG/Tie2, or both, are currently being used for cancer treatment, or are still in various stages of clinical development or preclinical evaluation. This article aims to review the mechanisms of angiogenesis and tumor angiogenesis and to focus on new drugs and strategies for the treatment of antiangiogenesis. However, antitumor angiogenic drugs alone may not be sufficient to eradicate tumors. The molecular chaperone heat shock protein 90 (HSP90) is considered a promising molecular target. The VEGFR system and its downstream signaling molecules depend on the function of HSP90. This article also briefly introduces the role of HSP90 in angiogenesis and some HSP90 inhibitors.