Affinity Maturation of a High-affinity Human Monoclonal Antibody Against the Third Hypervariable Loop of Human Immunodeficiency Virus: Use of Phage Display to Improve Affinity and Broaden Strain Reactivity

Thompson, Julia; Pope, Tony; Tung, Jwu-Sheng; Chan, Christine P.; Hollis, Gregory; Mark, George E.; Johnson, Kevin S.

doi:10.1006/jmbi.1996.0069

Cited by 106 publications

(38 citation statements)

References 39 publications

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“…As this requires selection against various antigens simultaneously, we chose to work with a naive library, which is known to be robust and easy to use, and has been used to select antibodies towards several antigens [11]. The affinity of antibodies identified from this library is normally in the micromolar range, but can be improved by affinity maturation [27][28][29][30][31].…”

Section: Resultsmentioning

confidence: 99%

A model phage display subtraction method with potential for analysis of differential gene expression

et al. 1996

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In order to establish a subtractive procedure that makes it possible to enrich selectively phage displayed antibodies directed against proteins constituting a difference between two populations of cells, a competitive selection strategy utilising two solid phases was developed and tested. Antibodies recognising a defined difference between two otherwise identical protein mixtures were isolated and their specificity confirmed. To test further the efficacy of selection inhibition during the competitive selections, selections towards a total cell extract were performed with and without competition from the same extract. An analysis of the resulting phage antibodies confirmed the subtractive nature of the system described.

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Section: Resultsmentioning

confidence: 99%

A model phage display subtraction method with potential for analysis of differential gene expression

et al. 1996

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“…29) with recombinant human 5T4 using a 48-day rest/boost immunization protocol (30) and was identified from a biochemical assay of hybridoma supernatants for binding to 5T4. This antibody underwent affinity optimization by CDR-directed mutagenesis (31) and phage display selection (32,33). The lead antibody, 5T4_0108, was identified in a biochemical assay, where it demonstrated improved binding to 5T4 over the parental A07.…”

Section: Generation Of the Lead Anti-5t4 Antibody 5t4_0108mentioning

confidence: 99%

Preclinical Evaluation of MEDI0641, a Pyrrolobenzodiazepine-Conjugated Antibody–Drug Conjugate Targeting 5T4

Harper¹,

Lloyd²,

Dimasi³

et al. 2017

Molecular Cancer Therapeutics

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Antibody-drug conjugates (ADC) are used to selectively deliver cytotoxic agents to tumors and have the potential for increased clinical benefit to cancer patients. 5T4 is an oncofetal antigen overexpressed on the cell surface in many carcinomas on both bulk tumor cells as well as cancer stem cells (CSC), has very limited normal tissue expression, and can internalize when bound by an antibody. An anti-5T4 antibody was identified and optimized for efficient binding and internalization in a target-specific manner, and engineered cysteines were incorporated into the molecule for site-specific conjugation. ADCs targeting 5T4 were constructed by site-specifically conjugating the antibody with payloads that possess different mechanisms of action, either a DNA cross-linking pyrrolobenzodiazepine (PBD) dimer or a microtubule-destabilizing tubulysin, so that each ADC had a drug:antibody ratio of 2. The resulting ADCs demonstrated significant target-dependent activity in vitro and in vivo; however, the ADC conjugated with a PBD payload (5T4-PBD) elicited more durable antitumor responses in vivo than the tubulysin conjugate in xenograft models. Likewise, the 5T4-PBD more potently inhibited the growth of 5T4-positive CSCs in vivo, which likely contributed to its superior antitumor activity. Given that the 5T4-PBD possessed both potent antitumor activity as well as anti-CSC activity, and thus could potentially target bulk tumor cells and CSCs in target-positive indications, it was further evaluated in non-GLP rat toxicology studies that demonstrated excellent in vivo stability with an acceptable safety profile. Taken together, these preclinical data support further development of 5T4-PBD, also known as MEDI0641, against 5T4 þ cancer indications. Mol Cancer Ther;16(8); 1576-87. Ó2017 AACR.

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“…Phage display of antibody libraries has provided a powerful tool for the isolation of human MAbs to important viral pathogens (4,7,12,16,24,51). Large repertoires of antibodies can be displayed on the surface of filamentous phage particles (e.g., M13), and antibodies with desired specificity can be isolated by panning on the antigens of interest (6,13,61).…”

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confidence: 99%

Identification by Phage Display and Characterization of Two Neutralizing Chimpanzee Monoclonal Antibodies to the Hepatitis E Virus Capsid Protein

Schofield

Glamann

Emerson³

et al. 2000

J Virol

155

109

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Two monoclonal antibodies (MAbs) against the ORF2 protein of the SAR-55 strain of hepatitis E virus (HEV) were isolated by phage display from a cDNA library of chimpanzee (Pan troglodytes) ␥1/ antibody genes. Both MAbs, HEV#4 and HEV#31, bound to reduced, denatured open reading frame 2 (ORF2) protein in a Western blot, suggesting that they recognize linear epitopes. The affinities (equilibrium dissociation constants, K d ) for the SAR-55 ORF2 protein were 1.7 nM for HEV#4 and 5.4 nM for HEV#31. The two MAbs also reacted in an enzyme-linked immunosorbent assay with recombinant ORF2 protein from a heterologous HEV, the Meng strain. Each MAb blocked the subsequent binding of the other MAb to homologous ORF2 protein in indirect competition assays, suggesting that they recognize the same or overlapping epitopes. Radioimmunoprecipitation assays suggested that at least part of the linear epitope(s) recognized by the two MAbs is located between amino acids 578 and 607. MAbs were mixed with homologous HEV in vitro and then inoculated into rhesus monkeys (Macaca mulatta) to determine their neutralizing ability. Whereas all control animals developed hepatitis (elevated liver enzyme levels in serum) and seroconverted to HEV, those receiving an inoculum incubated with either HEV#4 or HEV#31 were not infected. Therefore, each MAb neutralized the SAR-55 strain of HEV in vitro.Hepatitis E is an acute disease endemic in many countries throughout developing parts of the world, in particular on the continents of Africa and Asia, where epidemics have also occurred. The causative agent, hepatitis E virus (HEV), is transmitted via the fecal-oral route, predominantly through contaminated water (45). HEV is an RNA virus with a positive-sense genome approximately 7.5 kb in length. The genome contains three open reading frames (ORFs); ORF2 encodes the putative capsid protein (45). Hepatitis E has a low mortality rate in young adults (38). However, mortality rates can reach 20% in women in the third trimester of pregnancy (32, 58). Surprisingly, in industrialized countries such as the United States, where hepatitis E is not endemic, a significant proportion of healthy individuals within the general population are seropositive (over 20% in some areas; 39, 50). However, clinical hepatitis E is rare in these countries and usually is seen in individuals who acquired the infection during travel to a region in which HEV is endemic or epidemic.It has been suggested that animals serve as reservoirs for HEV and that human infections may, in part, be zoonoses. There have been several reports of HEV-specific (anti-HEV) antibody in animals (1, 9, 28, 29). Furthermore, an HEV-like virus was recently isolated from naturally infected swine in the United States (40). Although four genotypes of HEV have been identified, to date, only one serotype of HEV has been recognized. Therefore, it may be possible to produce a broadly protective vaccine. Passively transferred anti-HEV serum significantly reduced virus shedding in feces and abrogated disease in nonhu...

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Affinity Maturation of a High-affinity Human Monoclonal Antibody Against the Third Hypervariable Loop of Human Immunodeficiency Virus: Use of Phage Display to Improve Affinity and Broaden Strain Reactivity

Cited by 106 publications

References 39 publications

A model phage display subtraction method with potential for analysis of differential gene expression

A model phage display subtraction method with potential for analysis of differential gene expression

Preclinical Evaluation of MEDI0641, a Pyrrolobenzodiazepine-Conjugated Antibody–Drug Conjugate Targeting 5T4

Identification by Phage Display and Characterization of Two Neutralizing Chimpanzee Monoclonal Antibodies to the Hepatitis E Virus Capsid Protein

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