The Development of a Canine Single-chain Phage Antibody Library to Isolate Recombinant Antibodies for use in Translational Cancer Research

Małgorzata  Lisowska; Erin G. Worralll; Filip Zavadil Kokáš; Keith Charlton; Euan Murray; M Aiman Mohtar; Radovan Krejcir; Václav  Hrabal; Jack Brydon; Ainhoa Gonzalez Urionabarrenetxea; David Saliba; Mariana Grima; Umesh Kalathiya; Petr Muller; Adam Krejci; Bořivoj Vojtěšek; Kathryn L Ball; Robin Fahraeus; David Argyle; Maciej Parys; Ted R Hupp

doi:10.1016/j.crmeth.2025.101008

The Development of a Canine Single-chain Phage Antibody Library to Isolate Recombinant Antibodies for use in Translational Cancer Research

Małgorzata Lisowska , Erin G. Worralll, Filip Zavadil Kokáš, Keith Charlton, Euan Murray, M Aiman Mohtar, Radovan Krejcir, Václav Hrabal, Jack Brydon, Ainhoa Gonzalez Urionabarrenetxea, David Saliba, Mariana Grima, Umesh Kalathiya, Petr Muller, Adam Krejci, Bořivoj Vojtěšek, Kathryn L Ball, Robin Fahraeus, David Argyle, Maciej ParysTed R Hupp

Research output: Contribution to journal › Article › peer-review

Abstract

The development of canine immunotolerant monoclonal antibodies can accelerate the invention of new medicines for both canine and human diseases. A methodology was developed to clone the naïve, somatically mutated variable domain repertoire from canine B-cell mRNA using 5’RACE PCR. A set of degenerate primers were then designed and used to clone variable domain genes into archival “holding” plasmid libraries. These archived variable domain genes were then combinatorially ligated to produce a scFv M13 phage library. Next-generation long-read and short-read DNA sequencing methodologies were developed to annotate features of the cloned library including CDR diversity and IGHV/IGKV/IGLV subfamily distribution. A synthetic IgG was developed from this scFv library to the canine immune checkpoint receptor PD-1. This synthetic platform can be used to clone and annotate archived antibody variable domain genes for use in perpetuity in order to develop improved preclinical models for the treatment of complex human diseases

Original language	English
Journal	Cell Reports
Early online date	24 Mar 2025
DOIs	https://doi.org/10.1016/j.crmeth.2025.101008
Publication status	E-pub ahead of print - 24 Mar 2025

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Lisowska , M., Worralll, E. G., Zavadil Kokáš, F., Charlton, K., Murray, E., Mohtar, M. A., Krejcir, R., Hrabal, V., Brydon, J., Gonzalez Urionabarrenetxea, A., Saliba, D., Grima, M., Kalathiya, U., Muller, P., Krejci, A., Vojtěšek, B., Ball, K. L., Fahraeus, R., Argyle, D., ... Hupp, T. R. (2025). The Development of a Canine Single-chain Phage Antibody Library to Isolate Recombinant Antibodies for use in Translational Cancer Research. Cell Reports. Advance online publication. https://doi.org/10.1016/j.crmeth.2025.101008

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title = "The Development of a Canine Single-chain Phage Antibody Library to Isolate Recombinant Antibodies for use in Translational Cancer Research",

abstract = "The development of canine immunotolerant monoclonal antibodies can accelerate the invention of new medicines for both canine and human diseases. A methodology was developed to clone the na{\"i}ve, somatically mutated variable domain repertoire from canine B-cell mRNA using 5{\textquoteright}RACE PCR. A set of degenerate primers were then designed and used to clone variable domain genes into archival “holding” plasmid libraries. These archived variable domain genes were then combinatorially ligated to produce a scFv M13 phage library. Next-generation long-read and short-read DNA sequencing methodologies were developed to annotate features of the cloned library including CDR diversity and IGHV/IGKV/IGLV subfamily distribution. A synthetic IgG was developed from this scFv library to the canine immune checkpoint receptor PD-1. This synthetic platform can be used to clone and annotate archived antibody variable domain genes for use in perpetuity in order to develop improved preclinical models for the treatment of complex human diseases",

author = "Ma{\l}gorzata Lisowska and Worralll, \{Erin G.\} and \{Zavadil Kok{\'a}{\v s}\}, Filip and Keith Charlton and Euan Murray and Mohtar, \{M Aiman\} and Radovan Krejcir and V{\'a}clav Hrabal and Jack Brydon and \{Gonzalez Urionabarrenetxea\}, Ainhoa and David Saliba and Mariana Grima and Umesh Kalathiya and Petr Muller and Adam Krejci and Bo{\v r}ivoj Vojt{\v e}{\v s}ek and Ball, \{Kathryn L\} and Robin Fahraeus and David Argyle and Maciej Parys and Hupp, \{Ted R\}",

year = "2025",

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doi = "10.1016/j.crmeth.2025.101008",

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Lisowska , M, Worralll, EG, Zavadil Kokáš, F, Charlton, K, Murray, E, Mohtar, MA, Krejcir, R, Hrabal, V, Brydon, J, Gonzalez Urionabarrenetxea, A, Saliba, D, Grima, M, Kalathiya, U, Muller, P, Krejci, A, Vojtěšek, B, Ball, KL, Fahraeus, R, Argyle, D , Parys, M & Hupp, TR 2025, 'The Development of a Canine Single-chain Phage Antibody Library to Isolate Recombinant Antibodies for use in Translational Cancer Research', Cell Reports. https://doi.org/10.1016/j.crmeth.2025.101008

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T1 - The Development of a Canine Single-chain Phage Antibody Library to Isolate Recombinant Antibodies for use in Translational Cancer Research

AU - Lisowska , Małgorzata

AU - Worralll, Erin G.

AU - Zavadil Kokáš, Filip

AU - Charlton, Keith

AU - Murray, Euan

AU - Mohtar, M Aiman

AU - Krejcir, Radovan

AU - Hrabal, Václav

AU - Brydon, Jack

AU - Gonzalez Urionabarrenetxea, Ainhoa

AU - Saliba, David

AU - Grima, Mariana

AU - Kalathiya, Umesh

AU - Muller, Petr

AU - Krejci, Adam

AU - Vojtěšek, Bořivoj

AU - Ball, Kathryn L

AU - Fahraeus, Robin

AU - Argyle, David

AU - Parys, Maciej

AU - Hupp, Ted R

PY - 2025/3/24

Y1 - 2025/3/24

N2 - The development of canine immunotolerant monoclonal antibodies can accelerate the invention of new medicines for both canine and human diseases. A methodology was developed to clone the naïve, somatically mutated variable domain repertoire from canine B-cell mRNA using 5’RACE PCR. A set of degenerate primers were then designed and used to clone variable domain genes into archival “holding” plasmid libraries. These archived variable domain genes were then combinatorially ligated to produce a scFv M13 phage library. Next-generation long-read and short-read DNA sequencing methodologies were developed to annotate features of the cloned library including CDR diversity and IGHV/IGKV/IGLV subfamily distribution. A synthetic IgG was developed from this scFv library to the canine immune checkpoint receptor PD-1. This synthetic platform can be used to clone and annotate archived antibody variable domain genes for use in perpetuity in order to develop improved preclinical models for the treatment of complex human diseases

AB - The development of canine immunotolerant monoclonal antibodies can accelerate the invention of new medicines for both canine and human diseases. A methodology was developed to clone the naïve, somatically mutated variable domain repertoire from canine B-cell mRNA using 5’RACE PCR. A set of degenerate primers were then designed and used to clone variable domain genes into archival “holding” plasmid libraries. These archived variable domain genes were then combinatorially ligated to produce a scFv M13 phage library. Next-generation long-read and short-read DNA sequencing methodologies were developed to annotate features of the cloned library including CDR diversity and IGHV/IGKV/IGLV subfamily distribution. A synthetic IgG was developed from this scFv library to the canine immune checkpoint receptor PD-1. This synthetic platform can be used to clone and annotate archived antibody variable domain genes for use in perpetuity in order to develop improved preclinical models for the treatment of complex human diseases

U2 - 10.1016/j.crmeth.2025.101008

DO - 10.1016/j.crmeth.2025.101008

M3 - Article

SN - 2211-1247

JO - Cell Reports

JF - Cell Reports

ER -

The Development of a Canine Single-chain Phage Antibody Library to Isolate Recombinant Antibodies for use in Translational Cancer Research

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