Neuropilin-1 Research Articles

Intravitreal CendR peptides target laser-induced choroidal neovascularization sites in mice

Choroidal neovascularization (CNV) is a common ocular pathology that may be associated in a variety of eye diseases. Although intravitreal injection treatment of anti-vascular endothelial growth factor (anti-VEGF) drugs shows significant clinical benefits in CNV treatment, the limitations of the current therapy need to be addressed. The aim of our study was to investigate the potential utility of three C-end Rule (CendR) peptides (RPARPAR, PL3, iRGD) for CNV targeting and to evaluate the efficacy of peptides for treating experimental CNV in mice. We observed that the CendR peptides localize to the CNV lesion sites after intravitreal injection and were mainly found in the outer nuclear cell layer (ONL) of the mouse retina. Interestingly, experimental therapy with tenascin-C (TNC-C) and neuropilin-1 (NRP-1)-targeting PL3 peptide, reduced angiogenesis and decreased vascular leakage. The results suggest that PL3 and potentially other CendR peptides could serve as affinity targeting ligands and therapeutics for ocular diseases that involve pathological CNV.

New Crystal Form of Human Neuropilin-1 b1 Fragment with Six Electrostatic Mutations Complexed with KDKPPR Peptide Ligand.

Neuropilin 1 (NRP1), a cell-surface co-receptor of a number of growth factors and other signaling molecules, has long been the focus of attention due to its association with the development and the progression of several types of cancer. For example, the KDKPPR peptide has recently been combined with a photosensitizer and a contrast agent to bind NRP1 for the detection and treatment by photodynamic therapy of glioblastoma, an aggressive brain cancer. The main therapeutic target is a pocket of the fragment b1 of NRP1 (NRP1-b1), in which vascular endothelial growth factors (VEGFs) bind. In the crystal packing of native human NRP1-b1, the VEGF-binding site is obstructed by a crystallographic symmetry neighbor protein, which prevents the binding of ligands. Six charged amino acids located at the protein surface were mutated to allow the protein to form a new crystal packing. The structure of the mutated fragment b1 complexed with the KDKPPR peptide was determined by X-ray crystallography. The variant crystallized in a new crystal form with the VEGF-binding cleft exposed to the solvent and, as expected, filled by the C-terminal moiety of the peptide. The atomic interactions were analyzed using new approaches based on a multipolar electron density model. Among other things, these methods indicated the role played by Asp320 and Glu348 in the electrostatic steering of the ligand in its binding site. Molecular dynamics simulations were carried out to further analyze the peptide binding and motion of the wild-type and mutant proteins. The simulations revealed that specific loops interacting with the peptide exhibited mobility in both the unbound and bound forms.

Low Expression of the NRP1 Gene Is Associated with Shorter Overall Survival in Patients with Sonic Hedgehog and Group 3 Medulloblastoma.

Medulloblastoma (MB) is the most common type of malignant pediatric brain tumor. Neuropilin-1 (NRP1), encoded by the NRP1 gene, is a transmembrane glycoprotein overexpressed in several types of cancer. Previous studies indicate that NRP1 inhibition displays antitumor effects in MB models and higher NRP1 levels are associated with poorer prognosis in MB patients. Here, we used a large MB tumor dataset to examine NRP1 gene expression in different molecular subgroups and subtypes of MB. We found overall widespread NRP1 expression across MB samples. Tumors in the sonic hedgehog (SHH) subgroup showed significantly higher NRP1 transcript levels in comparison with Group 3 and Group 4 tumors, with SHH samples belonging to the α, β, Δ, and γ subtypes. When all MB subgroups were combined, lower NRP1 expression was associated with significantly shorter patient overall survival (OS). Further analysis showed that low NRP1 was related to poorer OS, specifically in MB subgroups SHH and Group 3 MB. Our findings indicate that patients with SHH and Group 3 tumors that show lower expression of NRP1 in MB have a worse prognosis, which highlights the need for subgroup-specific investigation of the NRP1 role in MB.

Dual Targeted Nanoparticles for the Codelivery of Doxorubicin and siRNA Cocktails to Overcome Ovarian Cancer Stem Cells.

Most anticancer treatments only induce the death of ordinary cancer cells, while cancer stem cells (CSCs) in the quiescent phase of cell division are difficult to kill, which eventually leads to cancer drug resistance, metastasis, and relapse. Therefore, CSCs are also important in targeted cancer therapy. Herein, we developed dual-targeted and glutathione (GSH)-responsive novel nanoparticles (SSBPEI-DOX@siRNAs/iRGD-PEG-HA) to efficiently and specifically deliver both doxorubicin and small interfering RNA cocktails (siRNAs) (survivin siRNA, Bcl-2 siRNA and ABCG2 siRNA) to ovarian CSCs. They are fabricated via electrostatic assembly of anionic siRNAs and cationic disulfide bond crosslinking-branched polyethyleneimine-doxorubicin (SSBPEI-DOX) as a core. Interestingly, the SSBPEI-DOX could be degraded into low-cytotoxic polyethyleneimine (PEI). Because of the enrichment of glutathione reductase in the tumor microenvironment, the disulfide bond (-SS-) in SSBPEI-DOX can be specifically reduced to promote the controlled release of siRNA and doxorubicin (DOX) in the CSCs. siRNA cocktails could specifically silence three key genes in CSCs, which, in combination with the traditional chemotherapy drug DOX, induces apoptosis or necrosis of CSCs. iRGD peptides and "sheddable" hyaluronic acid (HA) wrapped around the core could mediate CSC targeting by binding with neuropilin-1 (NRP1) and CD44 to enhance delivery. In summary, the multifunctional delivery system SSBPEI-DOX@siRNAs/iRGD-PEG-HA nanoparticles displays excellent biocompatibility, accurate CSC-targeting ability, and powerful anti-CSC ability, which demonstrates its potential value in future treatments to overcome ovarian cancer metastasis and relapse. To support this work, as exhaustive search was conducted for the literature on nanoparticle drug delivery research conducted in the last 17 years (2007-2023) using PubMed, Web of Science, and Google Scholar.

Deciphering intercellular signaling complexes by interaction-guided chemical proteomics

Indirect cell–cell interactions mediated by secreted proteins and their plasma membrane receptors play essential roles for regulating intercellular signaling. However, systematic profiling of the interactions between living cell surface receptors and secretome from neighboring cells remains challenging. Here we develop a chemical proteomics approach, termed interaction-guided crosslinking (IGC), to identify ligand-receptor interactions in situ. By introducing glycan-based ligation and click chemistry, the IGC approach via glycan-to-glycan crosslinking successfully captures receptors from as few as 0.1 million living cells using only 10 ng of secreted ligand. The unparalleled sensitivity and selectivity allow systematic crosslinking and identification of ligand-receptor complexes formed between cell secretome and surfaceome in an unbiased and all-to-all manner, leading to the discovery of a ligand-receptor interaction between pancreatic cancer cell-secreted urokinase (PLAU) and neuropilin 1 (NRP1) on pancreatic cancer-associated fibroblasts. This approach is thus useful for systematic exploring new ligand-receptor pairs and discovering critical intercellular signaling events.

Tumor Necrosis Factor and Interleukin-1β Upregulate NRP2 Expression and Promote SARS-CoV-2 Proliferation.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which causes coronavirus disease 2019 (COVID-19), utilizes the host receptor angiotensin-converting enzyme 2 (ACE2) and the auxiliary receptor Neuropilin-1 (NRP1) to enter host cells. NRP1 has another isoform, NRP2, whose function in COVID-19 has seldom been reported. In addition, although patients with severe cases of COVID-19 often exhibit increased levels of proinflammatory cytokines, the relationship between these cytokines and SARS-CoV-2 proliferation remains unknown. The aim of this study is to clarify the roles of proinflammatory cytokines in Neuropilin expressions and in SARS-CoV-2 infection. To identify the expression patterns of NRP under inflamed and noninflamed conditions, next-generation sequencing (RNA-seq), immunohistochemistry, quantitative real-time PCR, and Western blotting were performed using primary cultured fibroblast-like synoviocytes, MH7A (immortalized cell line of human rheumatoid fibroblast-like synoviocytes), immortalized MRC5 (human embryonic lung fibroblast), and synovial tissues. To measure viral proliferative capacity, SARS-CoV-2 infection experiments were also performed. NRP2 was upregulated in inflamed tissues. Cytokine-stimulated human fibroblast cell lines, such as MH7A and immortalized MRC5, revealed that NRP2 expression increased with co-stimulation of tumor necrosis factor α (TNFα) and interleukin-1 beta (IL-1β) and was suppressed with anti-TNFα antibody alone. TNFα and IL-1β promoted SARS-CoV-2 proliferation and Spike protein binding. The viral proliferation coincided with the expression of NRP2, which was modulated through plasmid transfections. Our results revealed that proinflammatory cytokines, including TNFα, contribute to NRP2 upregulation and SARS-CoV-2 proliferation in host human cells.

Overexpression of NRP1 is Associated with Poor Prognosis via Accelerating Immunosuppression in Head and Neck Squamous Cell Carcinoma.

Neuropilin-1 (NRP1) is a significant molecular structure that participates in many diseases progress including malignant tumors. However, its role in head and neck squamous cell carcinoma (HNSCC) remains to be uncovered. In this study, we determined the function of NRP1 as a crucial biomarker in proliferation, metastasis and immunosuppression in HNSCC. We collected samples of normal tissues (n = 18) and HNSCC tissues (n = 202) for immunohistochemical staining of NRP1 and evaluated its correlation to clinical prognostic characteristics. Furthermore, we enrolled 37 HNSCC patients received immune checkpoint blockade (ICB) treatment with defined therapeutic effects records. The biological process, signal pathways, and immune infiltration relevance to NRP1 were analyzed utilized transcriptome data from The Cancer Genome Atlas (TCGA). The NRP1 protein expression was significantly upregulated in HNSCC tissue and was associated with T stage, N stage, histological differentiation, recurrence and NRP1 expression. The high expression of NRP1 indicated poor survival rate and was found to be an independent prognosis factor. Enrichment analysis showed that NRP1 was associated with cell adhesion, extracellular matrix organization, homophilic cell adhesion via plasma membrane in biological process and neuroactive ligand-receptor interaction, protein digestion and absorption, calcium signal pathways. Moreover, NRP1 mRNA level was found positively correlated to cancer associated fibroblast cells, Treg cells and macrophage/monocyte cells. NRP1 might be likely to develop into a potential immunoregulation target as well as a predictive biomarker in HNSCC immune treatment.

Commensal bacteria weaken the intestinal barrier by suppressing epithelial neuropilin-1 and Hedgehog signaling

The gut microbiota influences intestinal barrier integrity through mechanisms that are incompletely understood. Here we show that the commensal microbiota weakens the intestinal barrier by suppressing epithelial neuropilin-1 (NRP1) and Hedgehog (Hh) signaling. Microbial colonization of germ-free mice dampens signaling of the intestinal Hh pathway through epithelial Toll-like receptor (TLR)-2, resulting in decreased epithelial NRP1 protein levels. Following activation via TLR2/TLR6, epithelial NRP1, a positive-feedback regulator of Hh signaling, is lysosomally degraded. Conversely, elevated epithelial NRP1 levels in germ-free mice are associated with a strengthened gut barrier. Functionally, intestinal epithelial cell-specific Nrp1 deficiency (Nrp1ΔIEC) results in decreased Hh pathway activity and a weakened gut barrier. In addition, Nrp1ΔIEC mice have a reduced density of capillary networks in their small intestinal villus structures. Collectively, our results reveal a role for the commensal microbiota and epithelial NRP1 signaling in the regulation of intestinal barrier function through postnatal control of Hh signaling.

Chlorotoxin binds to both matrix metalloproteinase 2 and neuropilin 1

Chlorotoxin (CTX), a scorpion venom-derived 36-residue miniprotein, binds to and is taken up selectively by glioblastoma cells. Previous studies provided controversial results concerning target protein(s) of CTX. These included CLC3 chloride channel, matrix metalloproteinase 2 (MMP-2), regulators of MMP-2, annexin A2, and neuropilin 1 (NRP1). The present study aimed at clarifying which of the proposed binding partners can really interact with CTX using biochemical methods and recombinant proteins. For this purpose, we established two new binding assays based on anchoring the tested proteins to microbeads and quantifying the binding of CTX by flow cytometry. Screening of His-tagged proteins anchored to cobalt-coated beads indicated strong interaction of CTX with MMP-2 and NRP1, whereas binding to annexin A2 was not confirmed. Similar results were obtained with fluorophore-labeled CTX and CTX-displaying phages. Affinity of CTX to MMP-2 and NRP1 was assessed by the "immunoglobulin-coated bead" test, in which the proteins were anchored to beads by specific antibodies. This assay yielded highly reproducible data using both direct titration and displacement approach. The affinities of labeled and unlabeled CTX appeared to be similar for both MMP-2 and NRP1 with estimated KD values of 0.5 to 0.7μM. Contrary to previous reports, we found that CTX does not inhibit the activity of MMP-2 and that CTX not only with free carboxyl end but also with carboxamide terminal end binds to NRP1. We conclude that the presented robust assays could also be applied for affinity-improving studies of CTX to its genuine targets using phage display libraries.

Neuropilin-1 is essential for vascular endothelial growth factor A-mediated increase of sensory neuron activity and development of pain-like behaviors.

Neuropilin-1 (NRP-1) is a transmembrane glycoprotein that binds numerous ligands including vascular endothelial growth factor A (VEGFA). Binding of this ligand to NRP-1 and the co-receptor, the tyrosine kinase receptor VEGFR2, elicits nociceptor sensitization resulting in pain through the enhancement of the activity of voltage-gated sodium and calcium channels. We previously reported that blocking the interaction between VEGFA and NRP-1 with the Spike protein of SARS-CoV-2 attenuates VEGFA-induced dorsal root ganglion (DRG) neuronal excitability and alleviates neuropathic pain, pointing to the VEGFA/NRP-1 signaling as a novel therapeutic target of pain. Here, we investigated whether peripheral sensory neurons and spinal cord hyperexcitability and pain behaviors were affected by the loss of NRP-1. Nrp-1 is expressed in both peptidergic and nonpeptidergic sensory neurons. A CRIPSR/Cas9 strategy targeting the second exon of nrp-1 gene was used to knockdown NRP-1. Neuropilin-1 editing in DRG neurons reduced VEGFA-mediated increases in CaV2.2 currents and sodium currents through NaV1.7. Neuropilin-1 editing had no impact on voltage-gated potassium channels. Following in vivo editing of NRP-1, lumbar dorsal horn slices showed a decrease in the frequency of VEGFA-mediated increases in spontaneous excitatory postsynaptic currents. Finally, intrathecal injection of a lentivirus packaged with an NRP-1 guide RNA and Cas9 enzyme prevented spinal nerve injury-induced mechanical allodynia and thermal hyperalgesia in both male and female rats. Collectively, our findings highlight a key role of NRP-1 in modulating pain pathways in the sensory nervous system.

P-793 Human trophoblastic spheroid derived from human expanded potential stem cells (hEPSC) for the identification of adhesion molecules for embryo implantation.

Abstract Study question Are cell surface proteins on trophoblastic spheroid BAP-EB derived from hEPSC involved in the endometrial epithelial cell attachment? Summary answer E-cadherin (CDH1) and Neuropilin-1 (NRP1) are two potential embryonic surface molecules involved in the human implantation process. What is known already Study of the mechanisms of human implantation in vivo is challenging. The availability of high-quality human embryos for in vitro research is limited. We have recently derived morula-like hEPSC lines (hEPSC-em) from donated human embryos. Trophoblastic spheroids (em-BAP-EB) derived from hEPSC-em resemble human trophectoderm during the early implantation process. The em-BAP-EBs attach specifically onto receptive endometrial epithelial cells, suggesting its usefulness as human embryo surrogate for studying embryonic surface molecules involving in early implantation process. Study design, size, duration hEPSC-em was subjected to embryoid body formation and induced to differentiate into trophoblastic spheroids (em-BAP-EB) for 48h and 72h. Endometrial aspirates were collected from infertile women in their natural cycles 2 or 7 days after LH surge (LH + 2/+7). Endometrial epithelial cells (EEC) were isolated and cultured for the attachment assays. Biotin-labelled apical surface proteins on em-BAP-EB were identified by mass spectrometry. Antibody blocking assays were performed to confirm the biological functions. Participants/materials, setting, methods The attachment rates of em-BAP-EB onto EEC were compared between pre-receptive (LH + 2 day) and receptive (LH + 7 day) phases. The differentially expressed apical surface protein on em-BAP-EB-48h and -72h were subjected to plasma membrane proteins (PMP) identification and pathway analysis. The gene and protein expression patterns of the target proteins were analyzed by single cell RNA sequencing (scRNA-seq) and western blotting respectively. The functional roles of the target proteins were studied by antibody blocking during attachment Main results and the role of chance The attachment rates of em-BAP-EB-72h onto receptive stage (LH + 7, n = 17) EEC were significantly higher than those onto pre-receptive stage (LH + 2, n = 5) EEC. Gene Ontology analysis indicated that the PMP with induced expression levels on em-BAP-EB-72h when compared to -48h (72h>48h) were enriched for “protein binding” and “cadherin binding” in molecular functions, and “cell-cell adhesion” in biological processes. Among them, 28 PMP were trophectoderm-specific. CDH1 and NRP1 were selected for further analysis. Analysis based on a published dataset showed that significantly higher expressions of CDH1 and NRP1 were detected in polar trophectoderm as compared to mural trophectoderm of E7 human blastocyst. CDH1 and NRP1 were significantly up-regulated in trophectoderm-like em-BAP-EB by scRNA-seq analysis. Live-cell immunocytochemistry demonstrated more intense apical surface expression of CDH1 and NRP1 on em-BAP-EB-72h as compared to -48h. Antibody blocking experiments indicated that pre-treatment of the em-BAP-EB-72h with specific antibody individually against the ectodomains of CDH1 and NRP1 significantly reduced the attachment rates onto receptive endometrial epithelial cells. The data suggested that CDH1 and NRP1 are two potential apical surface molecules essential for blastocyst attachment. Limitations, reasons for caution The lack of embryonic compartment in the em-BAP-EB and the absence of other key players like decidualized stromal cells and immune cells in the coculture model might not fully represent the early attachment process. The limited sample size in this study may also limit the interpretation of the data. Wider implications of the findings Despite advancement in in vitro fertilization, the success rate remains unsatisfactory. It is not known if embryological factor(s) other than embryo aneuploidy are involved. The current data may suggest the potential roles of CDH1 and NRP1 as non-chromosomal embryological markers of its implantation potential. Trial registration number Not applicable

Paradoxical expression of NRP1 in decidual stromal and immune cells reveals a novel inflammation balancing mechanism during early pregnancy.

To investigate the balancing mechanisms between decidualization-associated inflammation and pregnancy-related immunotolerance. Decidual samples from women with normal pregnancy (n = 58) or unexplained spontaneous miscarriage (n = 13), peripheral blood from normal pregnancy and endometria from non-pregnancy (n = 10) were collected. Primary endometrial stromal cells (ESCs), decidual stromal cells (DSCs), decidual immune cells (DICs) and peripheral blood mononuclear cells (PBMCs) were isolated. The plasmid carrying neuropilin-1 (NRP1) gene was transfected into ESC for overexpression. To induce decidualization in vitro, ESCs were treated with a combination of 10nM estradiol, 100nM progesterone and 0.5mM cAMP. Anti-Sema3a and anti-NRP1 neutralizing antibodies were applied to block the ligand-receptor interactions. RNA-seq analysis was performed to identify differentially expressed genes in DSCs and DICs, and NRP1 expression was verified by Western blotting and flow cytometry. The secretion of inflammatory mediators was measured using a multifactor cytometric bead array. The effects of Sema3a-NRP1 pathway on DICs were determined by flow cytometry. Statistical differences between groups were compared using the T test and one way or two-way ANOVA. Combined with five RNA-seq datasets, NRP1 was the only immune checkpoint changing oppositely between DSCs and DICs. The decreased expression of NRP1 in DSCs allowed intrinsic inflammatory responses required for decidualization, while its increased expression in DICs enhanced tolerant phenotypes beneficial to pregnancy maintenance. DSC-secreted Sema3a promoted immunosuppression in DICs via NRP1 binding. In women with miscarriage, NRP1 was abnormally elevated in DSCs but diminished in decidual macrophages and NK cells. NRP1 is a multifunctional controller that balances the inflammatory states of DSCs and DICs in gravid uterus. Abnormal expression of NRP1 is implicated in miscarriage.

Systematic pan-cancer analysis identified neuropilin 1 as an immunological and prognostic biomarker.

Neuropilin 1 (NRP1) is a transmembrane glycoprotein, nontyrosine kinase receptor that plays an important role in axonal growth and angiogenesis in the nervous system. Although currently more and more studies have shown that NRP1 plays an important role in some cancers, no systematic pan-cancer analysis of NRP-1 has been performed to date. Therefore, we aimed to investigate the associated immune function and prognostic value of NRP1 in 33 tumors of various cancer types. In this study, based on The Cancer Genome Atlas, Cancer Cell Line Encyclopedia, Genotype Tissue Expression, cBioportal for cancer genomics, and Human Protein Atlas (HPA databases), various bioinformatics analysis methods were used to investigate the potential carcinogenic effects of NRP1 activation, pan-cancer analysis of NRP1 expression, and the relationship between NRP1 expression and prognosis indicators including overall survival, disease-specific survival, disease-free interval, and progression-free interval, tumor mutational burden (TMB), and microsatellite instability (MSI). The results showed that NRP1 was highly expressed in most tumors. In addition, NRP1 was found to be positively or negatively correlated with the prognosis of different tumors. Also, the expression of NRP1 was associated with TMB and MSI in in 27 and 21 different types of tumors, respectively, and with DNA methylation in almost all the various types of tumors. The expression of the NRP1 gene was negatively correlated with the infiltration levels of most immune cells. In addition, the correlation between the level of immune cell infiltration and NRP1 expression varied according to immune cell subtype. Our study suggests that NRP1 plays an important role in tumor development and tumor immunity and could potentially be used as a prognostic indicator in a variety of malignancies.

Immunoexpression of neuropilin-1 in the chorionic villi of HIV-infected preeclamptic South African women of African ancestry.

Neuropilin-1 (NRP-1) is an essential regulator of maternal immune tolerance, placentation, and angiogenesis. Its dysregulation in preeclampsia (PE) and human immunodeficiency virus (HIV) infection implicates NRP-1 in disease susceptibility and progression. Therefore, this study investigates placental NRP-1 immunoexpression in HIV-complicated preeclamptic pregnancies in South African women of African ancestry receiving antiretroviral therapy. Immunohistochemistry of recombinant anti-neuropilin-1 antibody was performed on placental tissue from 30 normotensive and 60 early onset (EOPE) and late-onset (LOPE) preeclamptic women stratified by HIV status. Qualitative analysis of NRP-1 immunostaining within the chorionic villi revealed a predominant localization in trophoblasts and syncytial knots as well as endothelial, fibroblast-like, and Hofbauer cells. Following morphometric evaluation, we report that PE and HIV infection and/or antiretroviral usage independently downregulate placental NRP-1 immunoexpression; however, as a comorbidity, this decline is further augmented within the conducting and exchange villi. Furthermore, reduced immunoexpression of NRP-1 in EOPE compared with LOPE villi may be due to maternal-fetal maladaptation. It is plausible that the decreased NRP-1 immunoexpression in PE placentae facilitates syncytiotrophoblast apoptosis and subsequent deportation of NRP-1 into the maternal circulation, contributing to the anti-angiogenic milieu of PE. We hypothesize that the intense NRP-1 immunoreactivity observed in Hofbauer cells at the maternal-fetal interface may contribute to the natural prevention mechanism of HIV vertical transmission.

In Silico Prediction and Molecular Docking of SNPs in NRP1 Gene Associated with SARS-COV-2.

Neuropilin-1 (NRP1) which is a main transmembrane cell surface receptor acts as a host cell mediator resulting in increasing the SARS-Cov-2 infectivity and also plays a role in neuronal development, angiogenesis and axonal outgrowth. The goal of this study is to estimate the impact of single nucleotide polymorphisms (SNPs) in the NRP1 gene on the function, structure and stabilization of protein as well as on the miRNA-mRNA binding regions using bioinformatical tools. It is also aimed to investigate the changes caused by SNPs in NRP1 on interactions with drug molecule and spike protein. The missense type of SNPs was analyzed using SIFT, PolyPhen-2, SNAP2, PROVEAN, Mutation Assessor, SNPs&GO, PhD-SNP, I-Mutant 3.0, MUpro, STRING, Project HOPE, ConSurf, and PolymiRTS. Docking analyses were conducted by AutoDock Vina program. As a result, a total of 733 missense SNPs were determined within the NRP1 gene and nine SNPs were specified as damaging to the protein. The modelling results showed that wild and mutant type amino acids had some different properties such as size, charge, and hydrophobicity. Additionally, their three-dimensional structures of protein were utilized for confirmation of these differences. After evaluating the results, nine polymorphisms rs141633354, rs142121081, rs145954532, rs200028992, rs200660300, rs369312020, rs370117610, rs370551432, rs370641686 were determined to be damaging on the structure and function of NRP1 protein and located in conserved regions. The results of molecular docking showed that the binding affinity values are nearly the same for wild-type and mutant structures support that the mutations carried out are not in the focus of the binding site, therefore the ligand does not affect the binding energy. It is expected that the results will be useful for future studies.

Direct anti-tumoral effects of sulfated fucans isolated from echinoderms: a possible role of neuropilin-1/β1 integrin endocytosis and focal adhesion kinase (FAK) degradation.

Hypercoagulability, a major complication of metastatic cancers, has usually been treated with heparins from natural sources, or with their synthetic derivatives, which are under intense investigation in clinical oncology. However, the use of heparin has been challenging for patients with risk of severe bleeding. While the systemic administration of heparins, in pre-clinical models, has shown primarily attenuating effects on metastasis, their direct effect on established solid tumors has generated contradictory outcomes. We investigated the direct antitumoral properties of two sulfated fucans isolated from marine echinoderms, FucSulf1 and FucSulf2, which exhibit anticoagulant activity with mild hemorrhagic potential. Unlike heparin, sulfated fucans significantly inhibited tumor cell proliferation (by ~30-50 percent), and inhibited tumor migration and invasion in vitro. We found that FucSulf1 and FucSulf2 interacted with fibronectin (FN) as efficiently as heparin, leading to loss of prostate cancer and melanoma cell spreading. The sulfated fucans increased the endocytosis of β1 integrin and neuropilin-1 (NRP-1) chains, two cell receptors implicated in FN-dependent adhesion. The treatment of cancer cells with both sulfated fucans, but not with heparin, also triggered intracellular focal adhesion kinase (FAK) degradation, with a consequent overall decrease in activated FAK levels. Finally, only sulfated fucans inhibited the growth of B16-F10 melanoma cells implanted in the dermis of syngeneic C57/BL6 mice. FucSulf1 and FucSulf2 arise from this study as candidates for the design of possible alternatives to long-term treatments of cancer patients with heparins, with the advantage of also controlling local growth and invasion of malignant cells.

A phase Ib/II, open-label, multicenter study to evaluate the safety, efficacy, and pharmacokinetics/pharmacodynamics of αvβ3/5 integrin and neuropilin-1 targeting peptide LSTA1 in patients with metastatic pancreatic ductal adenocarcinoma.

e16290 Background: LSTA1, formerly known as CEND-1, is a novel cyclic peptide which selectively binds to αv/β3 and β5 integrins, which are upregulated in solid tumors, where it undergoes protease cleavage producing a C-end rule peptide fragment (CendR) that selectively binds to and activates neuropilin-1 (NRP1). Once CendR binds NRP1, the CendR pathway is activated, leading to increased vascular permeability of tumors resulting in increased drug delivery and tumor accumulation of co-administered chemotherapeutic agents. Here we report the safety and efficacy results from a Phase Ib/II, multicenter study of LSTA1 plus (SoC) gemcitabine and locally produced nab-paclitaxel in patients with metastatic pancreatic ductal adenocarcinoma (mPDAC). Methods: The study included a 7-day safety run-in and dose expansion/extension parts. Previously untreated patients with histologically confirmed mPDAC and ≥1 target lesion were treated with systemic chemotherapy and received LSTA1 (1.6 mg/kg or 3.2 mg/kg), gemcitabine (1000 mg/m²) and nab-paclitaxel (125 mg/m2) on days 1, 8, 15, of each 28-day treatment cycle. LSTA1 alone was given during run-in to observe PK/PD. Treatment continued until disease progression or other discontinuation events. The primary endpoints were safety and preliminary efficacy. Results: As of 30 Nov 2022, 38 patients were enrolled at 10 sites in China (12 at 1.6 mg/kg; 26 at 3.2mg/kg). The median follow-up time was 6.28 months. 37 (97.4%) patients experienced TEAEs and 31 (81.6%) patients experienced grade≥3 TEAEs. The most common TEAEs were hematologic toxicity related to chemotherapy. The ORR was 42.9% (15/35) and DCR was 82.9% (29/35). The ORR in 1.6 mg/kg and 3.2 mg/kg groups was 45.5% and 41.7%, respectively. The mPFS in 1.6 mg/kg and 3.2 mg/kg groups was 7.43 months (95% CI: 0.59-NE) and 5.75 months (95% CI: 3.25-7.46) respectively. mOS was not reached. NRP1 expression levels were tested at baseline; six of whom had expression ≥40%. Of those, 4 had PRs and 2 had SD (ORR=66.7%; DCR=100%). The mPFS was 7.5 months. Conclusions: LSTA1 combined with SoC showed an acceptable safety profile in patients with mPDAC. Compared with historical SoC data, LSTA1 plus SoC numerically improved ORR and PFS. Higher NRP1 expression is associated with better ORR and PFS, suggesting NRP1 may be a potential biomarker of LSTA1 plus chemotherapy for mPDAC treatment. Further study is ongoing. Clinical trial information: NCT05052567 .

The Molecular Pharmacology and Therapeutics of Angiogenesis

Angiogenesis is the formation of new vasculature on top of the pre-existing ones, which is critical during the species’ lifespan. It is responsible for delivering oxygen and nutrients to metabolically active tissues, therefore playing an essentialrole in physiological conditions, such as wound healing and pathological circumstances, like malignancies. Althoughsharing similar characteristics, the blood vessels in pathological angiogenesis develop irregularly and disorganizedly.Therefore, the leakage of materials from blood vessels is familiar in abnormal angiogenesis, which may result inmuch higher interstitial pressure than internal pressure. Whether the angiogenesis is physiological or pathological,the proliferation of normal dormant endothelial cells is an essential requirement. Additionally, angiogenesis is alsoinitiated by vascular endothelial growth factor A (VEGF-A) interacting with the vascular endothelial growth factorreceptor 2 (VEGFR2) or the co-receptor neuropilin-1 (NRP-1). The Vegfa gene can be spliced in many ways, creatingnumerous VEGF-A isoforms, each with unique interactions with VEGFR2. Several drugs have been approved as thefirst- or second-line of angiogenesis, thereby preventing cancer progression. The widely used is monoclonal antibodies(bevacizumab) and receptor tyrosine kinase inhibitor (sorafenib and axitinib). However, they lack selectivity and, asa result, are less effective and have considerable adverse effects. As a natural drug, Paclitaxel has an anti-angiogeniceffect in addition to its chemotherapeutic effect. However, different doses of paclitaxel have different anti-angiogenicmechanisms and different effects on different species.

The Molecular Pharmacology and Therapeutics of Angiogenesis

Angiogenesis is the formation of new vasculature on top of the pre-existing ones, which is critical during the species’ lifespan. It is responsible for delivering oxygen and nutrients to metabolically active tissues, therefore playing an essentialrole in physiological conditions, such as wound healing and pathological circumstances, like malignancies. Althoughsharing similar characteristics, the blood vessels in pathological angiogenesis develop irregularly and disorganizedly.Therefore, the leakage of materials from blood vessels is familiar in abnormal angiogenesis, which may result inmuch higher interstitial pressure than internal pressure. Whether the angiogenesis is physiological or pathological,the proliferation of normal dormant endothelial cells is an essential requirement. Additionally, angiogenesis is alsoinitiated by vascular endothelial growth factor A (VEGF-A) interacting with the vascular endothelial growth factorreceptor 2 (VEGFR2) or the co-receptor neuropilin-1 (NRP-1). The Vegfa gene can be spliced in many ways, creatingnumerous VEGF-A isoforms, each with unique interactions with VEGFR2. Several drugs have been approved as thefirst- or second-line of angiogenesis, thereby preventing cancer progression. The widely used is monoclonal antibodies(bevacizumab) and receptor tyrosine kinase inhibitor (sorafenib and axitinib). However, they lack selectivity and, asa result, are less effective and have considerable adverse effects. As a natural drug, Paclitaxel has an anti-angiogeniceffect in addition to its chemotherapeutic effect. However, different doses of paclitaxel have different anti-angiogenicmechanisms and different effects on different species.

The Molecular Pharmacology and Therapeutics of Angiogenesis

Angiogenesis is the formation of new vasculature on top of the pre-existing ones, which is critical during the species’ lifespan. It is responsible for delivering oxygen and nutrients to metabolically active tissues, therefore playing an essentialrole in physiological conditions, such as wound healing and pathological circumstances, like malignancies. Althoughsharing similar characteristics, the blood vessels in pathological angiogenesis develop irregularly and disorganizedly.Therefore, the leakage of materials from blood vessels is familiar in abnormal angiogenesis, which may result inmuch higher interstitial pressure than internal pressure. Whether the angiogenesis is physiological or pathological,the proliferation of normal dormant endothelial cells is an essential requirement. Additionally, angiogenesis is alsoinitiated by vascular endothelial growth factor A (VEGF-A) interacting with the vascular endothelial growth factorreceptor 2 (VEGFR2) or the co-receptor neuropilin-1 (NRP-1). The Vegfa gene can be spliced in many ways, creatingnumerous VEGF-A isoforms, each with unique interactions with VEGFR2. Several drugs have been approved as thefirst- or second-line of angiogenesis, thereby preventing cancer progression. The widely used is monoclonal antibodies(bevacizumab) and receptor tyrosine kinase inhibitor (sorafenib and axitinib). However, they lack selectivity and, asa result, are less effective and have considerable adverse effects. As a natural drug, Paclitaxel has an anti-angiogeniceffect in addition to its chemotherapeutic effect. However, different doses of paclitaxel have different anti-angiogenicmechanisms and different effects on different species.