Antibody Phage Display Libraries: Contributions to Oncology

Dantas‐Barbosa, Carmela; Brigido, Marcelo M.; Maranhão, Andréa Queiróz

doi:10.3390/ijms13055420

Cited by 40 publications

(25 citation statements)

References 108 publications

(112 reference statements)

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“…2 In the past few decades, technologies such as transgenic mice encompassing human antibody gene repertoires, and phage display of antibody libraries, have become available, facilitating the rapid flourishing of therapeutic antibodies in the drug discovery field. 3 …”

Section: Introductionmentioning

confidence: 99%

Next-generation sequencing enables the discovery of more diverse positive clones from a phage-displayed antibody library

et al. 2017

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Phage display technology provides a powerful tool to screen a library for a binding molecule via an enrichment process. It has been adopted as a critical technology in the development of therapeutic antibodies. However, a major drawback of phage display technology is that because the degree of the enrichment cannot be controlled during the bio-panning process, it frequently results in a limited number of clones. In this study, we applied next-generation sequencing (NGS) to screen clones from a library and determine whether a greater number of clones can be identified using NGS than using conventional methods. Three chicken immune single-chain variable fragment (scFv) libraries were subjected to bio-panning on prostate-specific antigen (PSA). Phagemid DNA prepared from the original libraries as well as from the Escherichia coli pool after each round of bio-panning was analyzed using NGS, and the heavy chain complementarity-determining region 3 (HCDR3) sequences of the scFv clones were determined. Subsequently, through two-step linker PCR and cloning, the entire scFv gene was retrieved and analyzed for its reactivity to PSA in a phage enzyme immunoassay. After four rounds of bio-panning, the conventional colony screening method was performed for comparison. The scFv clones retrieved from NGS analysis included all clones identified by the conventional colony screening method as well as many additional clones. The enrichment of the HCDR3 sequence throughout the bio-panning process was a positive predictive factor for the selection of PSA-reactive scFv clones.

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Section: Introductionmentioning

confidence: 99%

Next-generation sequencing enables the discovery of more diverse positive clones from a phage-displayed antibody library

et al. 2017

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“…4-6 In peptide phage display, bacteriophage are engineered to display random peptide sequences off of coat proteins to create a peptide library. One popular commercial platform, the Phage Display (Ph.D.) system, uses M13 bacteriophage modified for pentavalent peptide display, where the randomized peptide is fused to each of the five pIII bacteriophage coat proteins.…”

Section: Introductionmentioning

confidence: 99%

Efficient Identification of Murine M2 Macrophage Peptide Targeting Ligands by Phage Display and Next-Generation Sequencing

et al. 2015

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Peptide ligands are used to increase the specificity of drug carriers to their target cells and to facilitate intracellular delivery. One method to identify such peptide ligands, phage display, enables high-throughput screening of peptide libraries for ligands binding to therapeutic targets of interest. However, conventional methods for identifying target binders in a library by Sanger sequencing are low-throughput, labor-intensive, and provide a limited perspective (< 0.01%) of the complete sequence space. Moreover, the small sample space can be dominated by non-specific, preferentially amplifying “parasitic sequences” and plastic-binding sequences, which may lead to the identification of false positives or exclude the identification of target-binding sequences. To overcome these challenges, we employed next-generation Illumina sequencing to couple high-throughput screening and high-throughput sequencing, enabling more comprehensive access to the phage display library sequence space. In this work, we define the hallmarks of binding sequences in next-generation sequencing data, and develop a method that identifies several target-binding phage clones for murine, alternatively-activated (M2) macrophages with a high (100%) success rate: sequences and binding motifs were reproducibly present across biological replicates; binding motifs were identified across multiple unique sequences; and an unselected, amplified library accurately filtered out parasitic sequences. In addition, we validate the Multiple Em for Motif Elicitation tool as an efficient and principled means of discovering binding sequences.

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“…With the advent of recombinant DNA technology, antibody genes can be selected and amplified using phage display, yeast display, bacterial display, ribosome display, mRNA display, DNA display or mammalian cell surface display [69][70][71][72][73] and see chapter in this book: "Display technologies for the selection of monoclonal antibodies for clinical use" by Tsuruta et al…”

Section: Engineered Sdab Fragments From Vertebratesmentioning

confidence: 99%

The Development of Single Domain Antibodies for Diagnostic and Therapeutic Applications

Leow¹,

Cheng²,

Fischer³

et al. 2018

Antibody Engineering

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Monoclonal antibodies have become increasingly accepted as diagnostics and therapeutics for various human diseases due to their high affinity and specificity. However, several practical drawbacks are apparent for the reagents based on conventional IgG antibodies. With the emergence of antibody engineering, many problems were overcome when the recombinant antibody fragments such as Fabs, scFvs, diabodies and single domain antibodies (sdAbs), are developed. These fragments not only retain the specificity of the whole monoclonal antibodies, but are also easy to express and produce in prokaryotic expression systems. Rather unexpectedly, the natural sdAbs namely V HH s, V NAR s and variable lymphocyte receptors (VLRs) that comprise excellent biological activities were recently discovered in camelids, cartilaginous fish and lampreys, respectively. Due to their unique characteristics, including small size, high thermostability, stable folding in the nucleus and cytosol and long CDR3 regions which have access to cavities or clefts on the surface of proteins, these new binders are now investigated extensively as a substitute for conventional antibodies. This review describes the potential of sdAbs selected using in vitro display systems and their use in multiple applications.(polyclonal Abs) are heterogeneous antibody mixtures that are derived from multiple plasma cell lines. Because polyclonal antibodies comprise a mixture of different antibodies carrying numerous paratopes, they have excellent properties for recognizing antigens [2]. A monoclonal antibody (mAb) is a homogeneous antibody generated from a single B lymphocyte clone. Antibodies produced in mAb format have an extremely high specificity against a single epitope on antigens [3]. Recombinant antibodies (rAbs) are antibodies generated using molecular biology techniques. They are aimed to improve the sensitivity, selectivity, stability and immobilization properties in diagnostic applications, for example, in biosensors [4]. In making decision to use or generate polyclonal, monoclonal or recombinant antibodies, several factors should be considered, including commercial availability, possibility to raise animals, types of applications, time length of a project and costs [1]. Although a vast number of rAbs has been proposed [5][6][7][8], the natural sdAb fragments that were recently discovered from camelids (V HH s), sharks (V NAR s) and lampreys (VLRs) have shown to possess extraordinary features that are not found in conventional antibodies, such as a small dimension, an elevated stability and the capability of recognizing cavities and clefts on the surface of proteins that cannot be reached by conventional recombinant antibodies [9][10][11]. This chapter will discuss the availability of new binders derived from vertebrates and give an overview of their applications in a biomedical platform by recognizing specified targets from various diseases. Monoclonal antibodies and their limitationsThe first description of monoclonal antibody (mAbs) production was published ...

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Antibody Phage Display Libraries: Contributions to Oncology

Cited by 40 publications

References 108 publications

Next-generation sequencing enables the discovery of more diverse positive clones from a phage-displayed antibody library

Next-generation sequencing enables the discovery of more diverse positive clones from a phage-displayed antibody library

Efficient Identification of Murine M2 Macrophage Peptide Targeting Ligands by Phage Display and Next-Generation Sequencing

The Development of Single Domain Antibodies for Diagnostic and Therapeutic Applications

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