Vascular Endothelial Growth Factor Receptor Research Articles

Identification of a Novel Vascular Endothelial Growth Factor Receptor-3-Targeting Peptide for Molecular Imaging of Metastatic Lymph Nodes.

Because of the insidious nature of lymphatic metastatic cancer, accurate imaging tracing is very difficult to achieve in the clinic. Previous studies have developed the LARGR peptide (named TMVP1) as a radiotracer for vascular endothelial growth factor receptor-3 (VEGFR-3) imaging in cancer. However, its affinity for the target remains insufficient, resulting in low imaging sensitivity. In this study, we identified a high-affinity VEGFR-3 targeting peptide, named TMVP1446, using a multiplex screening platform. TMVP1446 demonstrated a dissociation constant of 8.97 × 10-8 M. Both in vitro and in vivo assays confirmed that fluorescently labeled TMVP1446 specifically bound to VEGFR-3. In a 4T1-luciferase tumor mouse model, cyanine 7-labeled TMVP1446 effectively discriminated between contralateral normal lymph nodes (c-LN) and cancer-metastatic sentinel lymph nodes (m-SLN). To evaluate the potential of TMVP1446, we developed a novel VEGFR-3 positron emission tomography radiotracer ([68Ga]Ga-DOTA-TMVP1446) for cancer-m-SLN imaging. [68Ga]Ga-DOTA-TMVP1446 accurately detected and assessed the status of lymph node metastasis, even in micrometastatic tumors, in the B16-F10 mouse tumor model. These findings suggest that TMVP1446 has great potential for advancing VEGFR-3 molecular imaging and metastatic sentinel lymph node imaging.

Effect of electroacupuncture at "Baihui" (GV20) and "Zusanli" (ST36) on angiogenesis in the brain of middle cerebral artery occlusion rats based on HIF/VEGF/Notch signaling pathway

To observe the effect of electroacupuncture (EA) on hypoxia-inducible factor (HIF)/vascular endothelial growth factor (VEGF)/Notch signaling pathway and related factors in rats with cerebral ischemia (CI), so as to explore its regulatory mechanism underlying improvement of CI by promoting cerebral microangiogenesis. Fifteen male SD rats were randomly selected as the sham surgery (sham) group, while other 85 rats were used to prepare CI model according to the modified Zea-Longa method. The successful CI model rats (n=60) were randomly allocated to model, EA, EA+inhibitor, and inhibitor groups, with 15 rats in each group. EA stimulation (1 Hz/20 Hz, 1-2 mA) was applied to "Baihui"(GV20) and right "Zusanli"(ST36) for 30 min, once a day for 7 days. Rats of the 2 inhibitor groups received intraperitoneal injection of YC-1 (HIF-1α inhibitor, 2.5 mg/kg), once a day for 7 days. The modified neurological severity scale (mNSS, including the motor ［muscular state, abnormal movement］, sensory ［visual, tactile and proprioceptive］, balance and reflex functions, 0-18 points) was used to assess the rats' neurological deficit state. The percentage of cerebral infarct volume was measured after 2, 3, 5-triphenyl tetrazolium chloride (TTC) staining, and pathological changes of the ischemic penumbra cortex were observed after H.E. staining. The immunoactivity of CD34 was determined using immunohistochemistry, followed by calculating the microvascular density (MVD) in the ischemic penumbra cortex, and the expression levels of HIF-1α, VEGF, vascular endothelial growth factor receptor 2 (VEGFR2), Notch1, and Delta like 4 ligand (DLL4) mRNAs and proteins in the ischemic penumbra of brain tissue were detected using real-time PCR and Western blot, respectively. In contrast to the sham group, the mNSS score, percentage of cerebral infarct volume, MVD, and expression levels of HIF-1α, VEGF, VEGFR2, Notch1 and DLL4 proteins and mRNAs were all significantly increased in the model group (P<0.01). In comparison with the model group, the mNSS score and percentage of cerebral infarct volume were significantly decreased in the EA group (P<0.01), and increased in the inhibitor group (P<0.01), and the MVD, and expression levels of HIF-1α, VEGF, VEGFR2, Notch1 and DLL4 proteins and mRNAs were further strikingly increased in the EA group (P<0.01), but obviously decreased in the inhibitor group (P<0.05, P<0.01). Comparison between EA and EA+inhibitor groups showed that the effects of EA were basically eliminated in lowering mNSS score and percentage of cerebral infarct volume (P<0.01), and in up-regulating MVD, and expression levels of HIF-1α, VEGF, VEGFR2, Notch1 and DLL4 mRNAs and proteins (P<0.01). The levels of mNSS score and percentage of cerebral infarct volume were markedly higher (P<0.01) in the inhibitor group than those in the EA+inhibitor group, while the MVD, expression levels of HIF-1α, VEGF, VEGFR2, Notch1 and DLL4 mRNAs and proteins were significantly lower in the inhibitor group than in the EA+inhibitor group (P<0.01), suggesting an elimination of EA after administration of HIF-1α inhibitor. H.E. staining showed loosened structure and disordered arrangement of neurons, swollen and vacuolized cells with ruptured nuclei, swollen and deformed microvessels in the ischemic penumbra of the brain tissue in the model group, which was improved in the EA group, including reduction of cellular swelling degree and vacuol-like changes, relatively intact of nuclei, and increase of new capillaries. EA of GV20 and ST36 can improve neurological deficit and reduce cerebral infarct volume in CI rats, which may be associated with its functions in promoting angiogenesis in ischemic penumbra area and in up-regulating the activities of HIF/VEGF/Notch signaling pathway and related factors.

Efficacy, Safety, and Influence on Tumor Microenvironment of Neoadjuvant Pembrolizumab plus Ramucirumab for PD-L1 Positive NSCLC: A Phase 2 Trial (EAST ENERGY).

Angiogenesis inhibitors are known to modify tumor immunity. Combination of angiogenesis inhibitors with immune checkpoint inhibitors (ICIs) has shown efficacy against many types of cancers, including non-small cell lung cancer (NSCLC). We investigated the feasibility of neoadjuvant therapy with pembrolizumab and ramucirumab, a vascular endothelial growth factor (VEGF) receptor-2 antagonist for patients with PD-L1-positive NSCLC and its influence on the tumor microenvironment (TME). Patients with pathologically proven NSCLC with PD-L1-positive, clinical stage IB-IIIA were eligible. Patients received two cycles of pembrolizumab (200 mg/body) and ramucirumab (10 mg/kg) every three weeks. Surgery was scheduled 4 to 8 weeks after the last dose. The primary endpoint was the major pathologic response (MPR) rate by a blinded independent pathology review. The sample size was 24 patients. Exploratory endpoints were evaluated to elucidate the effects of neoadjuvant therapy on TME. The 24 eligible patients were enrolled between July 2019 and April 2022. The MPR rate was 50.0% (90% confidence interval, 31.9-68.1%). Six patients showed pathological complete response. Grade 3 adverse events (AEs) occurred in 9 patients (37.5%), including 3 immune-related AEs (acute tubulointerstitial nephritis in 2 cases and polymyalgia rheumatica in one). There were no grade 4 or 5 AEs. The transcriptome and multiplexed immunohistochemistry results suggested that tumors with greater CD8+ T-cell infiltration and higher expression of effector molecules at the baseline could show better sensitivity to treatment. This new neoadjuvant combination of pembrolizumab plus ramucirumab was feasible and anti-VEGF agents may enhance the effects of ICIs.

Structural and dynamical insights revealed the anti-glioblastoma potential of withanolides from Withania coagulans against vascular endothelial growth factor receptor (VEGFR).

Glioblastoma (GBM), well known as grade 4 tumors due to its progressive malignant features such as vascular proliferation and necrosis, is the most aggressive form of primary brain tumor found in adults. Mutations and amplifications in the vascular endothelial growth factor receptor (VEGFR) contribute to almost 25% of GBM tumors. And thus, VEGFR has been declared the primary target in glioblastoma therapeutic strategies. However, many studies have been previously reported that include GBM as global therapeutics challenge, but they lack the molecular level insights that could help in understanding the biological function of a therapeutically important protein playing a major role in the disease and design the best strategies to develop the potential drugs. Therefore, to the best of our knowledge, the present study is the first time of kind, which involves multi-in silico approaches to predict the inhibition potential of withanolides from Withania coagulan against VEGFR. The study is actually based on determining the mode of action of five isolates: withanolide J, withaperuvin, 27-hydroxywithanolide I, coagule E, and coagule E, along with their respective binding energies. Molecular docking simulations revealed primarily four ligands, withanolide J (- 7.33kJ/mol), 27-withanolide (- 7.01kJ/mol), ajugine, withaperuvin (- 6.89kJ/mol), and ajugine E (- 6.39kJ/mol), to have significant binding potencies against the protein. Ligand binding was found to enhance the confirmational stability of the protein revealed through RMSD analysis, and RMSF assessment revealed the protein residues especially from 900-1000 surrounding the binding of the protein. Structural and dynamics of the protein via dynamics cross-correlation movement (DCCM) and principal component analysis (PCA) in both the unbound form and complexed with most potent ligand, withanolide J, reveal the ligand binding affecting the entire conformational integrity of the protein stabilized by hydrogen bonds and electrostatic attractions. Free energy of binding estimations by means of molecular mechanics Poisson-Boltzmann surface area (MMPBSA) method further revealed the withanolide J to have maximum binding potency of the all ligands. Withanolide J in final was also found to have suitable molecular characterizations to cross the blood-brain barrier (BBB +) and reasonable human intestinal absorption ability determined by ADMET profiling via admetSAR tools.

Chinese multidisciplinary expert consensus on the rational use of surufatinib in clinical practice（2024 edition）

Neuroendocrine neoplasms are a group of heterogeneous tumors originating from the neuroendocrine system, which can occur in any part of the body. Pulmonary and gastroenteropancreatic neuroendocrine tumors are the most common. In recent years, the global incidence of neuroendocrine tumors has increased more significantly than other types of tumors, especially in the past 40 years. The drug treatment of neuroendocrine tumors includes somatostatin analogues, anti-angiogenesis targeting drugs, nuclide therapy and chemotherapy. Surufatinib is an oral tyrosine kinase receptor inhibitors that targets for vascular endothelial growth factor receptor (VEGFR1-3), fibroblast growth factor receptor 1 (FGFR1), and colony-stimulating factor-1 receptor (CSF-1R), has received approval in 2020 and 2021 for the treatment of locally advanced or metastatic, progressive nonfunctional, well-differentiated (G1, G2) neuroendocrine tumors (NETs) of both pancreatic and extrapancreatic origin that are unresectable. Ongoing exploratory studies are investigating its potential application in other tumor types. Common adverse reactions observed during surufatinib treatment include hypertension, proteinuria, bleeding events, and hepatic lab test abnormal and diarrhea. Chinese multidisciplinary expert consensus on the rational use of surufatinib in clinical practice (2024 edition) aims to provide standardized guidance for its rational use and enhance patient compliance to maximize therapeutic benefits.

4-Anilinoquinazoline Derivatives as the First Potent NOD1-RIPK2 Signaling Pathway Inhibitors at the Nanomolar Range.

Inflammation is a defense mechanism that restores tissue damage and eliminates pathogens. Among the pattern recognition receptors that recognize danger or pathogenic signals, nucleotide oligomerization domains 1 and 2 (NOD1/2) have been identified to play an important role in innate immunity responses, and inhibition of NOD1 could be interesting to treat severe infections and inflammatory diseases. In this work, we identified the first selective NOD1 versus NOD2 pathway inhibitors at the nanomolar range based on a 4-anilinoquinazoline scaffold. We demonstrated that NOD1 inhibition occurs through the inhibition of receptor interacting protein kinase 2 (RIPK2), which is involved in its downstream signaling pathways. Compound 37 demonstrates no cytotoxicity, a selectivity for RIPK2 over epithelial and vascular endothelial growth factor receptors (EGFR/VEGFR), and a capacity to reduce pro-inflammatory cytokine IL-8 secretion. The structure of the RIPK2-compound 37 complex was resolved by crystallography. The 4-anilinoquinazoline scaffold offers novel perspectives to design NOD1-RIPK2 signaling inhibitors.

Clinical impact of TP53 functional mutations in patients with metastatic colorectal cancer treated with bevacizumab and chemotherapy.

Clinical and experimental studies indicate that the tumor protein p53 (TP53) gene loss of function due to missense mutations (MMs) may confer sensitivity to anti-angiogenics. This effect seems to be linked to cross-talk mechanisms among TP53, vascular endothelial growth factor (VEGF), and VEGF receptors. We investigated whether specific TP53 MMs are associated with clinical outcomes of patients with metastatic colorectal cancer (mCRC) treated with first-line chemotherapy plus Bevacizumab. The study focused on KRAS-mutated, liver-only mCRC cases as a homogeneous subgroup that may represent a relevant setting for exploring this association. MMs were identified on primary tumors. MMs were classified by mutant-specific residual transcriptional activity scores (TP53RTAS) as transcriptionally inactive (TP53inactive = TP53RTAS 0%) or active (TP53active = TP53RTAS ≥ 1%) and used for stratifying patients in progression-free survival (PFS), response rate, and overall survival (OS) analyses. The study population consisted of 62 patients. MMs were found in 39 cases (62%) with 16 having TP53inactive and 23 TP53active MMs. Patients with TP53inactive MMs showed better PFS in comparison with the remaining groups (wild-type and TP53active). This effect was retained in the multivariate model. A similar clinical impact was observed in the OS analysis. There was a significant difference in the overall response rate and rate of post-treatment resection of liver metastases between the TP53inactive and the wild-type or TP53active MMs cases. Specific TP53 MMs may identify sub-groups of patients who benefit from Bevacizumab-based systemic therapy and these findings could lead to novel tailored treatment strategies in this setting.

Near-Infrared II Fluorescence Imaging Highlights Tumor Angiogenesis in Hepatocellular Carcinoma with a VEGFR-Targeted Probe.

Hepatocellular carcinoma (HCC) is typically characterized by rich vascularity, with angiogenesis playing a crucial role in its growth and invasion. Molecular imaging of specific receptors in blood vessels is crucial in HCC diagnosis. In particular, in vivo imaging utilizing the second near-infrared (NIR-II) window offers improved tissue penetration, reduced light scattering, and lower autofluorescence. Despite the great potential of the NIR-II window, developing safe and effective probes to provide better imaging performance for HCC is urgently needed. In this study, NIR-II imaging integrated with a vascular endothelial growth factor receptor (VEGFR)-targeted probe generated by combining a VEGFR-targeted peptide with indocyanine green (ICG) is used to characterize HCC-related angiogenesis at a resolution of 56.0µm. For the first time, liver metabolic curves and parameters of liver function reserve (LFR) are obtained by fitting NIR-II fluorescence signals with high spatiotemporal resolution, showing significant differences between HCC mice and controls. Moreover, unlike ICG, the targeting probe has a targeted effect on blood vessels in vivo. The tumor-to-normal (T/N) ratio in NIR-II imaging reaches up to 3.30 after post-injection of the targeting probe. The results indicate that the VEGFR-targeted probe is a powerful tool for NIR-II fluorescence imaging to enhance early diagnosis of HCC.

Empagliflozin attenuating renal interstitial fibrosis in diabetic kidney disease by inhibiting lymphangiogenesis and lymphatic endothelial-to-mesenchymal transition via the VEGF-C/VEGFR3 pathway

Renal interstitial fibrosis (RIF) is a significant pathological change in diabetic kidney disease (DKD) that can be induced by endothelial-to-mesenchymal transition (EndMT). Lymphangiogenesis, mediated by the vascular endothelial growth factor-C (VEGF-C)/vascular endothelial growth factor receptor-3 (VEGFR-3) pathway, plays a crucial role in the development of RIF in DKD. Although numerous studies have demonstrated the efficacy of empagliflozin in treating renal injury, its effects on lymphangiogenesis in DKD-related RIF and the underlying mechanisms remain unclear. In the present study, significant lymphangiogenesis was assessed in the renal interstitium of patients with DKD. We subsequently explored the relationship between DKD-related RIF and lymphangiogenesis in mouse models, high-glucose (HG)-stimulated renal HK-2 cell lines, and human lymphatic endothelial cells (hLECs). Additionally, we evaluated the effects of empagliflozin on these processes. The results revealed that HG induces lymphangiogenesis, which exacerbates RIF by promoting inflammatory responses. Furthermore, hLECs directly contributed to the progression of DKD-related RIF through EndMT. Further analysis revealed that tubular epithelial cells (TECs) act as effector cells for VEGF-C, with the epithelial-to-mesenchymal transition (EMT) of TECs occurring concurrently with the EndMT of lymphatic vessels. Empagliflozin inhibited RIF in DKD by suppressing the VEGF-C/VEGFR3 pathway and reducing lymphangiogenesis. In conclusion, this study elucidates the interplay between lymphangiogenesis, EndMT, and RIF in DKD and provides new insights into the mechanism by which empagliflozin treats DKD.

Epidermal growth factor receptor (EGFR) and vascular endothelial growth factor A (VEGF-A) expressions in Ethiopian female breast cancer and their association with histopathologic features.

Epidermal growth factor receptor (EGFR) and vascular endothelial growth factor receptor (VEGF) play important role in breast tumor growth, invasion, metastasis, patient survival and drug resistance. The aim of this study was to evaluate the protein expression status of EGFR and VEGF-A, as well as their association with hormone receptor status and histopathological characteristics in the invasive type of female breast cancer among Ethiopians. The primary breast tumor tissues were obtained from 85 Ethiopian invasive breast cancer cases that underwent modified radical mastectomy (MRM) from June 2014 to June 2015. Their FFPE blocks were analyzed for EGFR and VEGF protein expressions using immunohistochemical techniques. The expressions were also correlated with histopathologic features. Epidermal growth factor receptor over-expression was observed in 22% of the tumor samples. VEGF-A expression was negative in 13.41%, low in 63.41%, moderate in 20.73%, and high in 2.44%. EGFR expression, but not VEGF-A, showed a significant inverse correlation with both estrogen receptor (ER) (P = 0.01) and progesterone receptor (PR) statuses (P = 0.04). EGFR and VEGF expressions did not show significant association with tumor size, grade, lymph node status or age at diagnosis. Epidermal growth factor receptor expression was most likely associated with ER and PR negative tumors. Assessments of multiple molecular markers aid to understand the biological behavior of the disease in Ethiopian population. It might also help to predict which group of patients might get more benefit from the selected treatment strategies and which are not.

Zinc Finger Protein 536 is a Potential Prognostic Biomarker that Promotes Neuroblastoma Progression via VEGFR2-PI3K-AKT Pathway.

Neuroblastoma (NB) is a well-known pediatric malignancy intertwined with neurodevelopment. Previously implicated in neuronal differentiation, Zinc Finger Protein 536 (ZNF536) has emerged as a promising prognostic and immune-related biomarker in our pan-cancer analysis. Single-cell RNA transcriptome sequencing, bulk transcriptome analysis, and immuno-histochemistry were used to assess ZNF536 expression and its association with prognosis. Cell proliferation, migration, invasion, and differentiation in ZNF536-knockdown NB cell lines were detected to evaluate the effect of ZNF536 on tumor cells. Vascular Endothelial Growth Factor Receptor 2 (VEGFR2), a potential target of ZNF536, and its downstream PI3K/AKT signaling cascade were investigated using transcriptome sequencing, CUT&Tag, quantitative real-time PCR (qRT-PCR), and Western blotting. The role of ZNF536 in tumorigenesis and the potential regulation axis was evaluated in vivo using a BALB/c nude mouse xenograft tumor model. ZNF536 mRNA and protein expression were significantly higher in NB patients with poor prognosis. In vitro, ZNF536 knockdown curtailed proliferation, migration, and invasion of NB cells while fostering differentiation. ZNF536 regulated VEGFR2 expression, thus activating the PI3K-AKT pathway. In vivo, ZNF536 knockdown reduced tumor growth and proliferation via the VEGFR2-PI3K-AKT pathway. ZNF536 resulted as a novel prognostic biomarker in NB, promoting oncogenesis through VEGFR2-PI3K-AKT signaling axis modulation, suggesting its therapeutic potential in managing NB progression.

Oncofetal MCB1 Is a Functional Biomarker for HCC Personalized Therapy.

Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related death worldwide and lacks biomarkers for personalized therapy. Herein, it is reported that MCB1 could be a novel oncofetal protein that is upregulated in the preneoplastic lesions and serum of early HCC patients. Functional studies reveal that MCB1 modulated p53 protein degradation to promote T-IC generation and drive HCC initiation. Furthermore, the MCB1/p53 axis is shown to determine the responses of hepatoma cells to conventional chemotherapeutics and predict transcatheter arterial chemoembolization (TACE) benefits in patients. Importantly, MCB1 can mediate sorafenib/lenvatinib resistance by downregulating two essential drug targets fibroblast growth factor receptor 1 (FGFR1) and vascular endothelial growth factor receptor 3 (VEGFR3) expression in a proteasome-dependent manner. Patient-derived tumor organoids (PDOs), patient-derived xenografts (PDXs), and patient cohorts analysis suggested that MCB1 levels in HCCs may determine the distinct responses to conventional therapeutics and targeted drugs. Furthermore, treatment of targeted drugs-resistant HCC with adeno-associated virus (AAV) targeting MCB1 or a proteasome inhibitor restores targeted drug response, suggesting their clinical significance in HCC combinational therapy. In conclusion, these findings demonstrate that MCB1 could act as a driver for HCC initiation, a contributor to drug resistance, and a biomarker for individualized HCC therapy.

Computer-Aided Design of VEGFR-2 Inhibitors as Anticancer Agents: A Review.

Due to its intricate molecular and structural characteristics, vascular endothelial growth factor receptor 2 (VEGFR-2) is essential for the development of new blood vessels in various pathological processes and conditions, especially in cancers. VEGFR-2 inhibitors have demonstrated significant anticancer effects by blocking many signaling pathways linked to tumor growth, metastasis, and angiogenesis. Several small compounds, including the well-tolerated sunitinib and sorafenib, have been approved as VEGFR-2 inhibitors. However, the widespread side effects linked to these VEGFR-2 inhibitors-hypertension, epistaxis, proteinuria, and upper respiratory infection-motivate researchers to search for new VEGFR-2 inhibitors with better pharmacokinetic profiles. The key molecular interactions required for the interaction of the small molecules with the protein target to produce the desired pharmacological effects are identified using computer-aided drug design (CADD) methods such as pharmacophore and QSAR modeling, structure-based virtual screening, molecular docking, molecular dynamics (MD) simulation coupled with MM/PB(GB)SA, and other computational strategies. This review discusses the applications of these methods for VEGFR-2 inhibitor design. Future VEGFR-2 inhibitor designs may be influenced by this review, which focuses on the current trends of using multiple screening layers to design better inhibitors.

Combination of anlotinib and sintilimab for the treatment of recurrent or metastatic head and neck squamous cell carcinoma: a single-arm prospective study

To investigate whether blocking both programmed cell death protein and vascular endothelial growth factor receptor could offer superior anticancer activity in these patients without compromising safety. In this study, patients were administered oral anlotinib (12 mg/day) on days 1–14 and intravenous sintilimab (200 mg) on day 1 of a 3-weekly cycle. The primary endpoints included the objective response rate and disease control rate. The secondary endpoints included overall survival (OS) and safety. Ten eligible patients were enrolled between June 2019 and May 2022, and eight patients underwent radiographic assessments. The results showed an objective response rate of 50% (partial and complete response in four and zero patients, respectively) and a disease control rate of 100%; four patients demonstrated stable disease for at least 8 weeks. The median OS was 4.37 (in our study, the score was 7), and the OS rate at 12 months was 37.5%. The Kaplan–Meier survival curve showed that the group with high blood glucose levels had a significantly shorter duration of survival than those with normal blood glucose levels. Adverse events of grade 3 and higher occurred in 50% of patients, and the most common severe adverse events included tumor pain (50%), hypertension (37.5%), tumor hemorrhage (25%), and decreased appetite (25%). The combination of anlotinib and sintilimab showed promising efficacy in controlling tumor size. However, the disappointing OS rate suggests that anti-vascular endothelial growth factor receptor agents should be used cautiously after radical radiation therapy. The combination used in this study demonstrated a toxicity profile comparable to that of other agents used in this setting. These findings warrant further investigation into the potential clinical utility of this combination.

In Silico Screening of 1,3,4-Thiadiazole Derivatives as Inhibitors of Vascular Endothelial Growth Factor Receptor-2 (VEGFR-2).

Angiogenesis plays a pivotal role in the growth, survival, and metastasis of solid tumors, with Vascular Endothelial Growth Factor Receptor-2 (VEGFR-2) being overexpressed in many human solid tumors, making it an appealing target for anti-cancer therapies. This study aimed to identify potential lead compounds with azole moiety exhibiting VEGFR-2 inhibitory effects. A ligand-based pharmacophore model was constructed using the X-ray crystallographic structure of VEGFR-2 complexed with tivozanib (PDB ID: 4ASE) to screen the ZINC15 database. Following virtual screening, six compounds demonstrated promising docking scores and drug-likeness comparable to tivozanib. These hits underwent detailed pharmacokinetic analysis to assess their absorption, distribution, metabolism, excretion, and toxicity (ADMET) properties. Furthermore, Density Functional Theory (DFT) analysis was employed to investigate the molecular orbital properties of the top hits from molecular docking. Molecular dynamics (MD) simulations were conducted to evaluate the conformational stability of the complexes over a 100 ns run. Results indicated that the compounds (ZINC8914312, ZINC8739578, ZINC8927502, and ZINC17138581) exhibited the most promising lead requirements for inhibiting VEGFR-2 and suppressing angiogenesis in cancer therapy. This integrated approach, combining pharmacophore modeling, molecular docking, ADMET studies, DFT analysis, and MD simulations, provides valuable insights into the identification of potential anti-cancer agents targeting VEGFR-2.

7410 Bevacizumab Induced Hypothyroidism

Abstract Disclosure: A.L. McKenna: None. S. Thota-Kammili: None. K. Grennan: None. S. Rao: None. It is known that thyroid dysfunction is an adverse effect of tyrosine kinase inhibitors (TKIs) via inhibition of the vascular endothelial growth factor receptor (VEGFR). The VEGF signaling pathway is important in regulating angiogenesis in tumors and normal tissue. As a highly vascularized gland, it is expected the thyroid may be compromised because of VEGFR inhibitors. Proposed mechanisms of thyroid dysfunction include capillary regression and constriction leading to reduced vascular perfusion to the gland. Bevacizumab is a monoclonal antibody that targets VEGF and inhibits activation of VEGFR, reducing angiogenesis; interestingly, this is not commonly reported in the literature. Here we present a case where bevacizumab altered thyroid function in a patient with pre-existing primary hypothyroidism. A 60-year-old woman presented with worsening hypothyroidism. She reported a history of nonautoimmune primary hypothyroidism diagnosed in her early 20s; she still had an intact thyroid with no history of head/neck radiation. In the months leading to the presentation, she had been receiving maintenance chemotherapy with 5 fluorouracil and bevacizumab every 2 weeks for metastatic duodenal carcinoma. Prior to starting chemotherapy, she had been on a steady dose of 112 mcg of levothyroxine (LT4) daily. After 6 months on chemotherapy, she reported symptoms of fatigue and hair loss. Workup revealed an elevated TSH of 41 mIU/L (reference 0.3-4.2 mIU/L). She denied any significant changes in weight or malabsorption issues. Aside from the chemotherapy, no other new medications were introduced. Her LT4 dose was increased to 137 mcg daily, and 3 months later, there was improvement in TSH to 6.5 mIU/L as well as her symptoms. At this point, bevacizumab was briefly on hold for 2 months due to an upcoming procedure, hence there was no change to LT4 to avoid over correction. Her TSH normalized and was trending lower while bevacizumab was on hold. Once bevacizumab resumed, LT4 was preemptively increased to 150 mcg daily. Four months after resuming bevacizumab, her TSH remains within the normal reference range on 150 mcg LT4. It is hypothesized that this VEGF blocker led to worsening of her underlying hypothyroidism.Given the shared down regulation in the VEGF signaling pathway, it would be expected that TKIs and monoclonal antibodies against VEGF would share similar adverse reactions to the thyroid. However, to our knowledge, there have been limited reports of bevacizumab induced thyroid dysfunction in the literature. Explanations to this may include the more targeted VEGF inhibition with bevacizumab versus the inhibition of multiple kinase receptors from the TKIs. Further research into this will be needed. This case demonstrates that bevacizumab can impact thyroid function and thyroid labs may need to be monitored at the onset and during VEGF inhibitor therapies. Presentation: 6/3/2024

Cutaneous Surgery Outcomes in Patients on Vascular Endothelial Growth Factor Receptor Inhibitors.

Cutaneous Surgery Outcomes in Patients on Vascular Endothelial Growth Factor Receptor Inhibitors.

The expression level of VEGFR2 regulates mechanotransduction, tumor growth and metastasis of high grade serous ovarian cancer cells

Recent data shows that alterations in the expression and/or activation of the vascular endothelial growth factor receptor 2 (VEGFR2) in high grade serous ovarian cancer (HGSOC) modulate tumor progression. However, controversial results have been obtained, showing that in some cases VEGFR2 inhibition can promote tumorigenesis and metastasis. Thus, it is urgent to better define the role of the VEGF/VEGFR2 system to understand/predict the effects of its inhibitors administered as anti-angiogenic in HGSOC. Here, we modulated the expression levels of VEGFR2 and analyzed the effects in two cellular models of HGSOC. VEGFR2 silencing (or its pharmacological inhibition) promote the growth and invasive potential of OVCAR3 cells in vitro and in vivo. Consistent with this, the low levels of VEGFR2 in OV7 cells are associated with more pronounced proliferative and motile phenotypes when compared to OVCAR3 cells, and VEGFR2 overexpression in OV7 cells inhibits cell growth. In vitro data confirmed that VEGFR2 silencing in OVCAR3 cells favors the acquisition of an invasive phenotype by loosening cell-ECM contacts, reducing the size and the signaling of focal adhesion contacts (FAs). This is translated into a reduced FAK activity at FAs, ECM-dependent alterations of mechanical forces through FAs and YAP nuclear translocation. Together, the data show that low expression, silencing or inhibition of VEGFR2 in HGSOC cells alter mechanotransduction and lead to the acquisition of a pro-proliferative/invasive phenotype which explains the need for a more cautious use of anti-VEGFR2 drugs in ovarian cancer.

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

Abstract 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.

Isolation-protocol, characterization, and in-vitro performance of equine umbilical vein endothelial cells.

Angiogenesis plays a crucial role in various physiological and pathological conditions. However, research in equine angiogenesis is relative limited, necessitating the development of suitable in-vitro models. To effectively analyze angiogenesis in-vitro, it is essential to target the specific cells responsible for this process, namely endothelial cells. Human umbilical vein endothelial cells (HUVECs) are one of the most used in vitro models for studying angiogenesis in humans. Serving as an equivalent to HUVECs, we present a comprehensive isolation protocol for equine umbilical vein endothelial cells (EqUVECs) with relatively minimal requirements, thereby enhancing accessibility for researchers. Umbilical cords obtained from five foals were used to isolate endothelial cells, followed by morphological and immunohistochemical identification. Performance of the cells in various assays commonly used in angiogenesis research was studied. Additionally, EqUVEC expression of vascular endothelial growth factor (VEGF) was assessed using ELISA. EqUVECs exhibited endothelial characteristics, forming a homogeneous monolayer with distinctive morphology. Immunohistochemical staining confirmed positive expression of key endothelial markers including von Willebrand factor (vWF), CD31, and vascular endothelial growth factor receptor-2 (VEGFR-2). Furthermore, performance assessments in in-vitro assays demonstrated the viability, proliferation, migration, tube formation and VEGF-expression capabilities of EqUVECs. The findings suggest that EqUVECs are a promising in-vitro model for studying equine angiogenesis, offering a foundation for further investigations into equine-specific vascular processes and therapeutic interventions.