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Holger Thie

Bio: Holger Thie is an academic researcher from Miltenyi Biotec. The author has contributed to research in topics: Phage display & Affinity maturation. The author has an hindex of 10, co-authored 19 publications receiving 553 citations. Previous affiliations of Holger Thie include Braunschweig University of Technology.

Papers
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Journal ArticleDOI
TL;DR: This work presents the first comprehensive comparison of V gene subfamily use for all steps of an antibody phage display pipeline, and describes a compatible cassette vector set, allowing in vivo biotinylation, enzyme fusion and Fc fusion.

145 citations

Journal ArticleDOI
TL;DR: This article gives an overview about the development of human therapeutic antibodies generated by phage display and those currently in clinical trials, 14 of which are described in detail.
Abstract: This article gives an overview about the development of human therapeutic antibodies generated by phage display After an introduction to the method, the focus is on approved antibodies and those currently in clinical trials, 14 of which are described in detail

86 citations

Journal ArticleDOI
TL;DR: This work has analysed the influence of various leader peptides on antibody phage display efficiency and production yields of soluble antibody fragments and the applicability of the SRP pathway to antibody phages display and the production of functional scFvs is shown.

75 citations

Journal ArticleDOI
14 Jan 2011-PLOS ONE
TL;DR: Antibody phage display starting with immune libraries and followed by affinity maturation is a powerful strategy to generate high affinity human antibodies to difficult targets, in this case shown by the creation of a highly specific antibody with subnanomolar affinity to a very small epitope consisting of four amino acids.
Abstract: Background So far, human antibodies with good affinity and specificity for MUC1, a transmembrane protein overexpressed on breast cancers and ovarian carcinomas, and thus a promising target for therapy, were very difficult to generate. Results A human scFv antibody was isolated from an immune library derived from breast cancer patients immunised with MUC1. The anti-MUC1 scFv reacted with tumour cells in more than 80% of 228 tissue sections of mamma carcinoma samples, while showing very low reactivity with a large panel of non-tumour tissues. By mutagenesis and phage display, affinity of scFvs was increased up to 500fold to 5,7×10−10 M. Half-life in serum was improved from below 1 day to more than 4 weeks and was correlated with the dimerisation tendency of the individual scFvs. The scFv bound to T47D and MCF-7 mammalian cancer cell lines were recloned into the scFv-Fc and IgG format resulting in decrease of affinity of one binder. The IgG variants with the highest affinity were tested in mouse xenograft models using MCF-7 and OVCAR tumour cells. However, the experiments showed no significant decrease in tumour growth or increase in the survival rates. To study the reasons for the failure of the xenograft experiments, ADCC was analysed in vitro using MCF-7 and OVCAR3 target cells, revealing a low ADCC, possibly due to internalisation, as detected for MCF-7 cells. Conclusions Antibody phage display starting with immune libraries and followed by affinity maturation is a powerful strategy to generate high affinity human antibodies to difficult targets, in this case shown by the creation of a highly specific antibody with subnanomolar affinity to a very small epitope consisting of four amino acids. Despite these “best in class” binding parameters, the therapeutic success of this antibody was prevented by the target biology.

75 citations

Journal ArticleDOI
29 Mar 2018-mAbs
TL;DR: The most abundant mRNA transcripts found in a hybridoma cell line did not necessarily encode the antibody chains providing the correct specificity, reiterating the importance of using sequence-defined recombinant antibodies for research or diagnostic use.
Abstract: Monoclonal antibodies are commonly assumed to be monospecific, but anecdotal studies have reported genetic diversity in antibody heavy chain and light chain genes found within individual hybridomas. As the prevalence of such diversity has never been explored, we analyzed 185 random hybridomas, in a large multicenter dataset. The hybridomas analyzed were not biased towards those with cloning difficulties or known to have additional chains. Of the hybridomas we evaluated, 126 (68.1%) contained no additional productive chains, while the remaining 59 (31.9%) contained one or more additional productive heavy or light chains. The expression of additional chains degraded properties of the antibodies, including specificity, binding signal and/or signal-to-noise ratio, as determined by enzyme-linked immunosorbent assay and immunohistochemistry. The most abundant mRNA transcripts found in a hybridoma cell line did not necessarily encode the antibody chains providing the correct specificity. Consequently, when cloning antibody genes, functional validation of all possible VH and VL combinations is required to identify those with the highest affinity and lowest cross-reactivity. These findings, reflecting the current state of hybridomas used in research, reiterate the importance of using sequence-defined recombinant antibodies for research or diagnostic use.

72 citations


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Journal ArticleDOI
TL;DR: The preeminent antibody engineering technologies used in the development of therapeutic antibody drugs, such as humanization of monoclonal antibodies, phage display, the human antibody mouse, single B cell antibody technology, and affinity maturation are outlined.
Abstract: It has been more than three decades since the first monoclonal antibody was approved by the United States Food and Drug Administration (US FDA) in 1986, and during this time, antibody engineering has dramatically evolved. Current antibody drugs have increasingly fewer adverse effects due to their high specificity. As a result, therapeutic antibodies have become the predominant class of new drugs developed in recent years. Over the past five years, antibodies have become the best-selling drugs in the pharmaceutical market, and in 2018, eight of the top ten bestselling drugs worldwide were biologics. The global therapeutic monoclonal antibody market was valued at approximately US$115.2 billion in 2018 and is expected to generate revenue of $150 billion by the end of 2019 and $300 billion by 2025. Thus, the market for therapeutic antibody drugs has experienced explosive growth as new drugs have been approved for treating various human diseases, including many cancers, autoimmune, metabolic and infectious diseases. As of December 2019, 79 therapeutic mAbs have been approved by the US FDA, but there is still significant growth potential. This review summarizes the latest market trends and outlines the preeminent antibody engineering technologies used in the development of therapeutic antibody drugs, such as humanization of monoclonal antibodies, phage display, the human antibody mouse, single B cell antibody technology, and affinity maturation. Finally, future applications and perspectives are also discussed.

1,025 citations

Journal ArticleDOI
TL;DR: This review efforts to describe the various proteomics approaches, the recent developments and their application in research and analysis.
Abstract: Proteomics involves the applications of technologies for the identification and quantification of overall proteins present content of a cell, tissue or an organism. It supplements the other "omics" technologies such as genomic and transcriptomics to expound the identity of proteins of an organism, and to cognize the structure and functions of a particular protein. Proteomics-based technologies are utilized in various capacities for different research settings such as detection of various diagnostic markers, candidates for vaccine production, understanding pathogenicity mechanisms, alteration of expression patterns in response to different signals and interpretation of functional protein pathways in different diseases. Proteomics is practically intricate because it includes the analysis and categorization of overall protein signatures of a genome. Mass spectrometry with LC-MS-MS and MALDI-TOF/TOF being widely used equipment is the central among current proteomics. However, utilization of proteomics facilities including the software for equipment, databases and the requirement of skilled personnel substantially increase the costs, therefore limit their wider use especially in the developing world. Furthermore, the proteome is highly dynamic because of complex regulatory systems that control the expression levels of proteins. This review efforts to describe the various proteomics approaches, the recent developments and their application in research and analysis.

528 citations

Journal ArticleDOI
TL;DR: The amenability of in vitro display to high-throughput applications broadens the prospects for their wider use in basic and applied research.
Abstract: In vitro display technologies, best exemplified by phage and yeast display, were first described for the selection of antibodies some 20 years ago. Since then, many antibodies have been selected and improved upon using these methods. Although it is not widely recognized, many of the antibodies derived using in vitro display methods have properties that would be extremely difficult, if not impossible, to obtain by immunizing animals. The first antibodies derived using in vitro display methods are now in the clinic, with many more waiting in the wings. Unlike immunization, in vitro display permits the use of defined selection conditions and provides immediate availability of the sequence encoding the antibody. The amenability of in vitro display to high-throughput applications broadens the prospects for their wider use in basic and applied research.

499 citations

Journal ArticleDOI
TL;DR: The first crystal structure of the C-terminal adhesion domain of InvD revealed a distinct Ig-related fold that, apart from the canonical β-sheets, comprises various modifications of and insertions into the Ig-core structure, suggesting that InvD modulates Ig functions in the intestine and affects direct interactions with a subset of cell surface-exposed B-cell receptors.

497 citations

Journal ArticleDOI
TL;DR: The technology shows a trend toward consolidation with a smaller set of systems that are being applied against multiple targets and in different settings, with emphasis on the development of drug candidates for therapy or in vivo diagnostics: Adnectins, Affibodies, Anticalins, DARPins, and engineered Kunitz-type inhibitors, among others.

458 citations