1,721,006 research outputs found
ELTD1 as a multi-focal target for malignant gliomas: Preclinical studies
No full text© 2021 The Author(s). Published by Oxford University Press, the Society for Neuro-Oncology and the European Association of Neuro-Oncology.Background: Glioblastoma (GBM) is the most aggressive malignant primary brain tumor in adults. These high-grade gliomas undergo unregulated vascular angiogenesis, migration and cell proliferation allowing the tumor cells to evade cell-cycle checkpoints and apoptotic pathways. The Epidermal growth factor, latrophilin, and seven transmembrane domain-containing 1 on chromosome 1 (ELTD1) is an angiogenic biomarker that is highly expressed in malignant gliomas. Novel treatments targeting ELTD1 with monovalent monoclonal (mmAb) and single chain variable fragment (scFv) antibodies were effective in increasing animal survival, decreasing tumor volume and normalizing the vasculature. Due to the success of our antibody treatments on angiogenesis, this study sought to determine if our anti-ELTD1 treatments affected other aspects of tumorigenesis (cell proliferation, migration, and apoptosis) in a G55 glioma xenograft preclinical mouse model. Methods: Tumor tissue from untreated, mmAb and scFv anti-ELTD1 treated animals was used to quantify the positivity levels of human mitochondrial antibody, c-MET and Ki-67 for cellular proliferation, migratory markers CD44v6, TRPM8, and BMP2, and cleaved caspase 3 to assess apoptotic activity. Results: This approach demonstrated that our anti-ELTD1 treatments directly affected and decreased the human tumor cells within the tumor region. Additionally, there was a significant decrease in both cellular proliferation and migration due to anti-ETLD1 therapy. Lastly, anti-ELTD1 treatments successfully increased apoptotic activity within the tumor region. Conclusion: Our data suggest that anti-ELTD1 therapies would be effective against malignant gliomas by having a multi-focal effect and targeting all four aspects of tumorigenesis.N
Optimization of peripheral blood volume for in silico reconstitution of the human B-cell receptor repertoire
No full textB cells recognize antigens via membrane-expressed B-cell receptors (BCR) and antibodies. Similar human BCR sequences are frequently found at a significantly higher frequency than that theoretically calculated. Patients infected with SARS-CoV2 and HIV or with autoimmune diseases share very similar BCRs. Therefore, in silico reconstitution of BCR repertoires and identification of stereotypical BCR sequences related to human pathology have diagnostic potential. Furthermore, monitoring changes of clinically significant BCR sequences and isotype conversion has prognostic potential. For BCR repertoire analysis, peripheral blood (PB) is the most convenient source. However, the optimal human PB volume for in silico reconstitution of the BCR repertoire has not been studied in detail. Here, we sampled 5, 10, and 20 mL PB from the left arm and 40 mL PB from the right arm of two volunteers, reconstituted in silico PB BCR repertoires, and compared their composition. In both volunteers, PB sampling over 20 mL resulted in slight increases in functional unique sequences (FUSs) or almost no increase in repertoire diversity. All FUSs with a frequency above 0.08% or 0.03% in the 40 mL PB BCR repertoire were detected even in the 5 mL PB BCR repertoire from each volunteer. FUSs with a higher frequency were more likely to be found in BCR repertoires from reduced PB volume, and those coexisting in two repertoires showed a statistically significant correlation in frequency irrespective of sampled anatomical site. The correlation was more significant in higher-frequency FUSs. These observations support the potential of BCR repertoire analysis for diagnosis.N
Assessment of an scFv Antibody Fragment Against ELTD1 in a G55 Glioblastoma Xenograft Model
No full textGlioblastoma (GBM), themost common primary brain tumor found in adults, is extremely aggressive. These high-grade gliomas, which are very diffuse, highly vascular, and invasive, undergo unregulated vascular angiogenesis. Despite available treatments, the median survival for patients is dismal. ELTD1 (EGF, latrophilin, and 7 transmembrane domain containing protein 1) is an angiogenic biomarker highly expressed in human high-grade gliomas. Recent studies have demonstrated that the blood-brain barrier, as well as the blood-tumor barrier, is not equally disrupted in GBM patients. This study therefore aimed to optimize an antibody treatment against ELTD1 using a smaller scFv fragment of a monoclonal antibody that binds against the external region of ELTD1 in a G55 glioma xenograft glioma preclinical model. Morphological magnetic resonance imaging (MRI) was used to determine tumor volumes and quantify perfusion rates. We also assessed percent survival following tumor postdetection. Tumor tissue was also assessed to confirm and quantify the presence of the ELTD1 scFvmolecular targetedMRI probe, aswell asmicrovessel density andNotch1 levels. In addition, we used molecular-targeted MRI to localize our antibodies in vivo. This approach showed that our scFv antibody attached-molecular MRI probe was effective in targeting and localizing diffuse tumor regions. Through this analysis, we determined that our anti-ELTD1 scFv antibody treatments were successful in increasing survival, decreasing tumor volumes, and normalizing vascular perfusion and Notch1 levels within tumor regions. This study demonstrates that our scFv fragment antibody against ELTD1 may be useful and potential antiangiogenic treatments against GBM. (C) 2019 The Authors. Published by Elsevier Inc. on behalf of Neoplasia Press, Inc.Y
Bispecific anti-mPDGFR beta x cotinine scFv-C-kappa-scFv fusion protein and cotinine-duocarmycin can form antibody-drug conjugate-like complexes that exert cytotoxicity against mPDGFR beta expressing cells
No full textAntibody selection for antibody-drug conjugates (ADCs) has traditionally depended on its internalization into the target cell, although ADC efficacy also relies on recycling of the receptor-ADC complex, endo-lysosomal trafficking, and subsequent linker/antibody proteolysis. In this study, we observed that a bispecific anti-murine platelet-derived growth factor receptor beta (mPDGFR beta) x cotinine single-chain variable fragment (scFv)-kappa constant region (C-kappa)-scFv fusion protein and cotinine-duocarmycin can form an ADC-like complex to induce cytotoxicity against rnPDGFR beta expressing cells. Multiple anti-mPDGFR beta antibody candidates can be produced in this bispecific scFv-C-kappa-scFv fusion protein format and tested for their ability to deliver cotinine-conjugated cytotoxic drugs, thus providing an improved approach for antibody selection in ADC development.N
Effective Combination Immunotherapy through Vessel Normalization Using a Cancer-Targeting Antiangiogenic Peptide–Antibody Hybrid
No full text© 2022 Wiley-VCH GmbH.Although cancer immunotherapy using immune checkpoint blockade (ICB) has changed the paradigm for treating patients with certain cancers, its therapeutic benefits are limited to approximately one-fourth of patients, highlighting the potential for combining immunotherapy with another therapeutic modality. Here, a treatment regimen that combines a cancer-targeting antiangiogenic agent that inhibits angiogenesis within the tumor and ICB to improve therapeutic outcome is reported. The cancer-targeting antiangiogenic modality is constructed as a hybrid complex, designated HyPEPEDB-VEGF, comprising a cotinine-labeled bispecific peptide targeting both extra domain B of fibronectin (EDB) and vascular endothelial growth factor (VEGF) and an anticotinine antibody (Abcot). The resulting HyPEPEDB-VEGF specifically bound to EDB-overexpressing CT26 murine colorectal cancer cells and inhibited VEGF-induced proliferation of human umbilical vascular endothelial cells. Upon intraperitoneal injection, HyPEPEDB-VEGF preferentially accumulates in CT26 syngeneic tumors and inhibits tumor growth in a dose-dependent manner. Furthermore, the combination of HyPEPEDB-VEGF with an anti-PD-1 antibody (αPD-1) in conjunction with dose optimization of the two modalities leads to substantial inhibition of tumor growth without loss of body weight due to vascular normalization within the tumor. These findings suggest that the combination of cancer-specific antiangiogenic therapy using HyPEPEDB-VEGF together with ICB may be a feasible approach for effective cancer therapy.N
Protein‐Encoding Free‐Standing RNA Hydrogel for Sub‐Compartmentalized Translation
No full textRNA can self-fold into complex structures that can serve as major biological regulators in protein synthesis and in catalysis. Due to the abundance of structural primitives and functional diversity, RNA has been utilized for designing nature-defined goals despite its intrinsic chemical instability and lack of technologies. Here, a robust, free-standing RNA hydrogel is developed through a sequential process involving both ligation and rolling circle transcription to form RNA G-quadruplexes, capable of both catalytic activity and enhancing expression of several proteins in sub-compartmentalized, phase-separated translation environments. The observations suggest that this hydrogel will expand RNA research and impact practical RNA principles and applications.N
Specific ablation of PDGFRβ-overexpressing pericytes with antibody-drug conjugate potently inhibits pathologic ocular neovascularization in mouse models
No full textBACKGROUND: Crosstalk between pericytes and endothelial cells is critical for ocular neovascularization. Endothelial cells secrete platelet-derived growth factor (PDGF)-BB and recruit PDGF receptor β (PDGFRβ)–overexpressing pericytes, which in turn cover and stabilize neovessels, independent of vascular endothelial growth factor (VEGF). Therapeutic agents inhibiting PDGF-BB/PDGFRβ signaling were tested in clinical trials but failed to provide additional benefits over anti-VEGF agents. We tested whether an antibody-drug conjugate (ADC) – an engineered monoclonal antibody linked to a cytotoxic agent - could selectively ablate pericytes and suppress retinal and choroidal neovascularization. METHODS: Immunoblotting, flow cytometry, cell viability test, and confocal microscopy were conducted to assess the internalization and cytotoxic effect of ADC targeting mPDGFRβ in an in vitro setting. Immunofluorescence staining of whole-mount retinas and retinal pigment epithelium-choroid-scleral complexes, electroretinography, and OptoMotry test were used to evaluate the effect and safety of ADC targeting mPDGFRβ in the mouse models of pathologic ocular neovascularization. RESULTS: ADC targeting mPDGFRβ is effectively internalized into mouse brain vascular pericytes and showed significant cytotoxicity compared with the control ADC. We also show that specific ablation of PDGFRβ-overexpressing pericytes using an ADC potently inhibits pathologic ocular neovascularization in mouse models of oxygen-induced retinopathy and laser-induced choroidal neovascularization, while not provoking generalized retinal toxicity. CONCLUSION: Our results suggest that removing PDGFRβ-expressing pericytes by an ADC targeting PDGFRβ could be a potential therapeutic strategy for pathologic ocular neovascularization
A High-Throughput Single-Clone Phage Fluorescence Microwell Immunoassay and Laser-Driven Clonal Retrieval System
No full textPhage display is one of the most frequently used platform technologies utilized to screen and select therapeutic antibodies, and has contributed to the development of more than 10 therapeutic antibodies used in the clinic. Despite advantages like efficiency and low cost, it has intrinsic technical limitations, such as the asymmetrical amplification of the library after each round of biopanning, which is regarded as a reason for it yielding a very limited number of antigen binders. In this study, we developed a high-throughput single-clonal screening system comprised of fluorescence immunoassays and a laser-driven clonal DNA retrieval system using microchip technology. Using this system, from a single-chain variable fragment (scFv) library displayed on phages with a complexity of 5.21 × 105 harboring random mutations at five amino acid residues, more than 70,000 clones—corresponding to ~14% of the library complexity—were screened, resulting in 78 antigen-reactive scFv sequences with mutations restricted to the randomized residues. Our results demonstrate that this system can significantly reduce the number of biopanning rounds, or even eliminate the need for this process for libraries with lower complexity, providing an opportunity to obtain more diverse clones from the library
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
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