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Yadveer Singh Grewal, Muhammad J A Shiddiky, Stephen M Mahler, Gerard A Cangelosi, Matt Trau
Rapid progress in disease biomarker discovery has increased the need for robust detection technologies. In the past several years, the designs of many immunoaffinity reagents have focused on lowering costs, improving specificity while also promoting stability. Antibody fragments (scFvs) have long been displayed on the surface of yeast and phage libraries for selection, however the stable production of such fragments presents challenges that hamper their widespread use in diagnostics. Membrane and cell wall proteins similarly suffer from stability problems when solubilized from their native environment...
October 20, 2016: ACS Applied Materials & Interfaces
Wei Li, Hongjia Yang, Dimiter S Dimitrov
CD16A (FcγRIIIA) is an activating receptor mostly expressed on natural killer (NK) cells and monocytes/macrophages. It can mediate antibody-dependent cell-mediated cytotoxicity (ADCC) through low-affinity interaction with human immunoglobulin G (IgG) Fc. It can also mediate cell lysis if NK cells are guided by bispecific killer cells engagers (BiKEs). BiKEs showed some success in clinical trials of cancer and are promising candidate therapeutics. However, currently reported BiKEs are based on antibody fragments (scFvs) of relatively large size...
October 3, 2016: Experimental and Molecular Pathology
Xiaochen Guo, Hongxue Cao, Yunxin Wang, Yang Liu, Yanmin Chen, Nan Wang, Shan Jiang, Shengqi Zhang, Qiang Wu, Tianhe Li, Yingjie Zhang, Bing Zhou, Jiechao Yin, Deshan Li, Guiping Ren
We have previously reported an antigen and antibody co-expression (AAC) technology to demonstrate the interaction between a known antigen and antibody. To validate the co-expression system for screening antibody libraries, a single chain fragment variable(scFv)antibody library was constructed from chickens immunized with the VP2 protein of infectious bursal disease virus (IBDV). The genes of both VP2 and scFv antibodies were inserted into the pBFD-Ab-Ag vector. The co-expression library was subjected to three rounds of screening by flow cytometry (FCM) using a polyclonal antibody against VP2 through a bacteria display system...
November 1, 2016: Veterinary Immunology and Immunopathology
Joerg U Schmohl, Martin Felices, Deborah Todhunter, Elizabeth Taras, Jeffrey S Miller, Daniel A Vallera
BACKGROUND: The design of a highly effective anti-cancer immune-engager would include targeting of highly drug refractory cancer stem cells (CSC). The design would promote effective antibody-dependent cell-mediated cytotoxicity (ADCC) and simultaneously promote costimulation to expand and self-sustain the effector NK cell population. Based on our bispecific NK cell engager platform we constructed a tetraspecific killer engager (TetraKE) comprising single-chain variable fragments (scFvs) binding FcγRIII (CD16) on NK cells, EpCAM on carcinoma cells and CD133 on cancer stem cells in order to promote ADCC...
September 16, 2016: Oncotarget
Morteza Motallebnezhad, Vahid Younesi, Leili Aghebati-Maleki, Hamid Nickho, Elham Safarzadeh, Majid Ahmadi, Ali Akbar Movassaghpour, Ahmad Hosseini, Mehdi Yousefi
Insulin-like growth factor I receptor (IGF-IR) is expressed on breast cancer cells and involves in metastasis, survival, and proliferation. Currently, application of IGF-IR-targeting monoclonal antibodies (mAbs), alone or in combination with other drugs, is a promising strategy for breast cancer therapy. Single-chain fragment variable (scFv) antibodies have been introduced as appropriate tools for tumor-targeting purposes because of their advantages over whole antibodies. In the present study, we employed a naïve phage library and isolated scFvs against a specific epitope from extracellular domain of IGF-IR by panning process...
September 17, 2016: Tumour Biology: the Journal of the International Society for Oncodevelopmental Biology and Medicine
Salma Teimoori, Watee Seesuay, Surasak Jittavisutthikul, Urai Chaisri, Nitat Sookrung, Jaslan Densumite, Nawannaporn Saelim, Monrat Chulanetra, Santi Maneewatch, Wanpen Chaicumpa
A direct acting anti-Ebola agent is needed. VP40, a conserved protein across Ebolavirus (EBOV) species has several pivotal roles in the virus life cycle. Inhibition of VP40 functions would lessen the virion integrity and interfere with the viral assembly, budding, and spread. In this study, cell penetrable human scFvs (HuscFvs) that bound to EBOV VP40 were produced by phage display technology. Gene sequences coding for VP40-bound-HuscFvs were subcloned from phagemids into protein expression plasmids downstream to a gene of cell penetrating peptide, i...
October 14, 2016: Biochemical and Biophysical Research Communications
Yanling Wu, Shibo Jiang, Tianlei Ying
INTRODUCTION: A variety of approaches are being pursued to improve the safety and antitumor potency of chimeric antigen receptor (CAR) T-cell therapy. However, most engineering efforts have thus far been focused on its intracellular signaling domain, while its extracellular antigen-binding domain has received less attention. AREAS COVERED: Herein, the authors summarize the current knowledge of CAR T-cell therapy. Accordingly, they focus on its antigen-binding domain, discuss key considerations for selecting an optimal single-chain variable fragment (scFv) when designing a CAR, and suggest potential directions aimed at developing the next-generation CARs...
September 19, 2016: Expert Opinion on Biological Therapy
Surasak Jittavisutthikul, Watee Seesuay, Jeeraphong Thanongsaksrikul, Kanyarat Thueng-In, Potjanee Srimanote, Rolf G Werner, Wanpen Chaicumpa
A safe and effective direct acting anti-hepatitis C virus (HCV) agent is still needed. In this study, human single chain variable fragments of antibody (scFvs) that bound to HCV NS3/4A protein were produced by phage display technology. The engineered scFvs were linked to nonaarginines (R9) for making them cell penetrable. HCV-RNA-transfected Huh7 cells treated with the transbodies produced from four different transformed E. coli clones had reduced HCV-RNA inside the cells and in the cell spent media, as well as fewer HCV foci in the cell monolayer compared to the transfected cells in culture medium alone...
2016: Frontiers in Immunology
William F Goins, Bonnie Hall, Justus B Cohen, Joseph C Glorioso
Gene therapy applications depend on vector delivery and gene expression in the appropriate target cell. Vector infection relies on the distribution of natural virus receptors that may either not be present on the desired target cell or distributed in a manner to give off-target gene expression. Some viruses display a very limited host range, while others, including herpes simplex virus (HSV), can infect almost every cell within the human body. It is often an advantage to retarget virus infectivity to achieve selective target cell infection...
September 7, 2016: Current Opinion in Virology
Brian Spencer, Stephanie Williams, Edward Rockenstein, Elvira Valera, Wei Xin, Michael Mante, Jazmin Florio, Anthony Adame, Eliezer Masliah, Michael R Sierks
OBJECTIVE: Progressive accumulation of α-synuclein (α-syn) has been associated with Parkinson's disease (PD) and Dementia with Lewy body (DLB). The mechanisms through which α-syn leads to neurodegeneration are not completely clear; however, the formation of various oligomeric species have been proposed to play a role. Antibody therapy has shown effectiveness at reducing α-syn accumulation in the central nervous system (CNS); however, most of these studies have been conducted utilizing antibodies that recognize both monomeric and higher molecular weight α-syn...
August 2016: Annals of Clinical and Translational Neurology
Yue Sun, Bhupal Ban, Andrew Bradbury, G A Shakeel Ansari, Diane A Blake
The development of antibodies to low molecular weight haptens remains challenging due to both the low immunogenicity of many haptens and the cross-reactivity of the protein carriers used to generate the immune response. Recombinant antibodies and novel display technologies have greatly advanced antibody development; however, new techniques are still required to select rare hapten-specific antibodies from large recombinant libraries. In the present study, we used a combination of phage and yeast display to screen an immune antibody library (size, 4...
September 20, 2016: Analytical Chemistry
Shuai Dong, Hongxi Shi, Donghui Cao, Yicun Wang, Xintong Zhang, Yan Li, Xiang Gao, Li Wang
Candida albicans (C. albicans) is an important human commensal and opportunistic fungal pathogen. Secreted aspartyl proteinases (Saps) are a major virulence trait of C. albicans, and among these proteases Sap2 has the highest expression levels. It is possible that antibodies against Sap2 could provide an antifungal effect. In this study, two phages displaying anti-rSap2 single chain variable fragments (scFvs) were screened from human single fold scFv libraries, and their potential therapeutic roles were evaluated using a murine model infected by C...
2016: Scientific Reports
Leila Rahbarnia, Safar Farajnia, Hossein Babaei, Jafar Majidi, Kamal Veisi, Asghar Tanomand, Bahman Akbari
Phage display is a prominent screening technique for development of novel high affinity antibodies against almost any antigen. However, removing false positive clones in screening process remains a challenge. The aim of this study was to develop an efficient and rapid method for isolation of high affinity scFvs by removing NSBs without losing rare specific clones. Therefore, a novel two rounds strategy called invert biopanning was developed for isolating high affinity scFvs against EGFRvIII antigen from human scFv library...
August 9, 2016: Biologicals: Journal of the International Association of Biological Standardization
Carol A Harley, Greg Starek, David K Jones, Andreia S Fernandes, Gail A Robertson, João H Morais-Cabral
The human human ether-à-go-go-related gene (hERG) potassium channel plays a critical role in the repolarization of the cardiac action potential. Changes in hERG channel function underlie long QT syndrome (LQTS) and are associated with cardiac arrhythmias and sudden death. A striking feature of this channel and KCNH channels in general is the presence of an N-terminal Per-Arnt-Sim (PAS) domain. In other proteins, PAS domains bind ligands and modulate effector domains. However, the PAS domains of KCNH channels are orphan receptors...
August 30, 2016: Proceedings of the National Academy of Sciences of the United States of America
Mira Woitok, Diana Klose, Judith Niesen, Wolfgang Richter, Muhammad Abbas, Christoph Stein, Rolf Fendel, Magdalena Bialon, Christiane Püttmann, Rainer Fischer, Stefan Barth, Katharina Kolberg
Antibody-drug conjugates (ADCs) combine the potency of cytotoxic drugs with the specificity of monoclonal antibodies (mAbs). Most ADCs are currently generated by the nonspecific conjugation of drug-linker reagents to certain amino acid residues in mAbs, resulting in a heterogeneous product. To overcome this limitation and prepare ADCs with a defined stoichiometry, we use SNAP-tag technology as an alternative conjugation strategy. This allows the site-specific conjugation of O(6)-benzylguanine (BG)-modified small molecules to SNAP-tag fusion proteins...
October 28, 2016: Cancer Letters
Haiyan Zhao, Estefania Fernandez, Kimberly A Dowd, Scott D Speer, Derek J Platt, Matthew J Gorman, Jennifer Govero, Christopher A Nelson, Theodore C Pierson, Michael S Diamond, Daved H Fremont
Zika virus (ZIKV) infection during pregnancy has emerged as a global public health problem because of its ability to cause severe congenital disease. Here, we developed six mouse monoclonal antibodies (mAbs) against ZIKV including four (ZV-48, ZV-54, ZV-64, and ZV-67) that were ZIKV specific and neutralized infection of African, Asian, and American strains to varying degrees. X-ray crystallographic and competition binding analyses of Fab fragments and scFvs defined three spatially distinct epitopes in DIII of the envelope protein corresponding to the lateral ridge (ZV-54 and ZV-67), C-C' loop (ZV-48 and ZV-64), and ABDE sheet (ZV-2) regions...
August 11, 2016: Cell
Sanam Sadreddini, Mehrnosh Seifi-Najmi, Babollah Ghasemi, Hossein Samadi Kafil, Vahideh Alinejad, Sevil Sadreddini, Vahid Younesi, Farhad Jadidi-Niaragh, Mehdi Yousefi
BACKGROUND: Tetanus neurotoxin (TeNT) is composed of a light (LC) and heavy chain (HC) polypeptides, released by anaerobic bacterium Clostridium tetani and can cause fatal life-threatening infectious disease. Toxin HC and LC modules represents receptor binding and zinc metalloprotease activity, respectively. The passive administration of animal-derived antibodies against tetanus toxin has been considered as the mainstay therapy for years. However, this treatment is associated with several adverse effects due to the presence of anti-isotype antibodies...
December 23, 2015: Human Antibodies
Raheleh Toughiri, Xiufeng Wu, Diana Ruiz, Flora Huang, John W Crissman, Mark Dickey, Karen Froning, Elaine M Conner, Thomas P Cujec, Stephen J Demarest
IgG antibodies are multi-domain proteins with complex inter-domain interactions. Human IgG heavy chains (HCs) associate with light chains (LCs) of the κ or λ isotype to form mature antibodies capable of binding antigen. The HC/LC interaction involves four domains: VH and CH1 from the HC and VL and CL from the LC. Human Fabs with κ LCs have been well characterized for their unfolding behaviors and demonstrate a significant level of cooperativity and stabilization when all four domains are intact. Very little is known regarding the thermodynamic properties of human Fabs with λ LCs...
July 25, 2016: MAbs
Xiaocong Yu, Mark Duval, Melissa Gawron, Marshall R Posner, Lisa A Cavacini
Innovative strategies are necessary to maximize the clinical application of HIV neutralizing antibodies. To this end, bispecific constructs of human antibody F240, reactive with well-conserved gp41 epitope and antibody 14A8, reactive with the IgA receptor (CD89) on effector cells, were constructed. A F240 × 14A8 bispecific single chain variable region (scFv) molecule was constructed by linking two scFvs using a conventional GGGGS linker. Despite immunoreactivity with HIV gp41 and neutrophils, this bispecific scFv failed to inhibit HIV infection...
2016: Journal of Immunology Research
Ryutaro Asano, Noriaki Koyama, Yasuyo Hagiwara, Yosuke Masakari, Ryota Orimo, Kyoko Arai, Hiromi Ogata, Shozo Furumoto, Mitsuo Umetsu, Izumi Kumagai
The development of single-chain variable fragments (scFvs) as therapeutic agents has the potential to reduce the high cost of antibody production, but the development process often impairs scFv functions such as binding affinity and pharmacokinetics. Multimerization is one strategy for recovering or enhancing these lost functions. Previously, we constructed several antiepidermal growth factor receptor (EGFR) scFv multimers by modifying linker length and domain order. Antitumor effects comparable with those of the currently approved anti-EGFR therapeutic antibodies were observed for scFv trimers...
June 2016: FEBS Open Bio
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