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Engineering a single chain Fv antibody to ?v?6 integrin using the specificity-determining loop of a foot-and-mouth disease virus

Engineering a single chain Fv antibody to ?v?6 integrin using the specificity-determining loop of a foot-and-mouth disease virus
Engineering a single chain Fv antibody to ?v?6 integrin using the specificity-determining loop of a foot-and-mouth disease virus
The ?v?6 integrin is a promising target for cancer therapy. Its expression is up-regulated de novo on many types of carcinoma where it may activate transforming growth factor-?1 and transforming growth factor-?3, interact with the specific extracellular matrix proteins and promote migration and invasion of tumor cells. The viral protein 1 (VP1) coat protein of the O1 British field strain serotype of foot-and-mouth disease virus is a high-affinity ligand for ?v?6, and we recently reported that a peptide derived from VP1 exhibited ?v?6-specific binding in vitro and in vivo. We hypothesized that this peptide could confer binding specificity of an antibody to ?v?6. A 17-mer peptide of VP1 was inserted into the complementarity-determining region H3 loop of MFE-23, a murine single-chain Fv (scFv) antibody reactive with carcinoembryonic antigen (CEA). The resultant scFv (B6-1) bound to ?v?6 but retained residual reactivity with CEA. This was eliminated by point mutation (Y100bP) in the variable heavy-chain domain to create an scFv (B6-2) that was as structurally stable as MFE-23 and reacted specifically with ?v?6 but not with ?5?1, ?v?3, ?v?5, ?v?8 or CEA. B6-2 was internalized into ?v?6-expressing cells and inhibited ?v?6-dependent migration of carcinoma cells. B6-2 was subsequently humanized. The humanized form (B6-3) was obtained as a non-covalent dimer from secretion in Pichia pastoris (115 mg/l) and was a potent inhibitor of ?v?6-mediated cell adhesion. Thus, we have used a rational stepwise approach to create a humanized scFv with therapeutic potential to block ?v?6-mediated cancer cell invasion or to deliver and internalize toxins specifically to ?v?6-expressing tumors.
antibody engineering, scFv, integrin ?v?6, VP1 peptide, foot-and-mouse disease virus
0022-2836
385-401
Kogelberg, Heide
f1a1fcd6-c4bf-4355-8678-13c9256cd475
Tolner, Berend
beea1fdc-6576-4000-b810-2d179e78cfcb
Thomas, Gareth J.
2ff54aa9-a766-416b-91ee-cf1c5be74106
Di Cara, Danielle
b5735d75-fe5a-488b-8925-703c3c317d67
Minogue, Shane
8b9455b6-1171-496c-a53b-66ab32308ced
Ramesh, Bala
e827116a-a274-4bd2-ba10-75ded7c683d7
Sodha, Serena
0943e72f-5a3b-4f9f-a54d-35e33d018dea
Marsh, Dan
df5ab9c4-9889-4093-a635-92c5c8508104
Lowdell, Mark W.
399d3d5b-dcd3-4b23-a6f9-7141c0717f56
Meyer, Tim
28c8fc8e-873f-4278-9443-cb7c9f9f487f
Begent, Richard H.J.
8d732fc3-678d-439a-ba03-9cc57a69a7ca
Hart, Ian
303b970c-96ed-49ec-9112-c5b9dd6d4165
Marshall, John F.
1134b21f-7f23-4ee6-aaf5-c7580b73c10b
Chester, Kerry
c7014f1a-71d9-4ca0-959d-2164f2b9b3e2
Kogelberg, Heide
f1a1fcd6-c4bf-4355-8678-13c9256cd475
Tolner, Berend
beea1fdc-6576-4000-b810-2d179e78cfcb
Thomas, Gareth J.
2ff54aa9-a766-416b-91ee-cf1c5be74106
Di Cara, Danielle
b5735d75-fe5a-488b-8925-703c3c317d67
Minogue, Shane
8b9455b6-1171-496c-a53b-66ab32308ced
Ramesh, Bala
e827116a-a274-4bd2-ba10-75ded7c683d7
Sodha, Serena
0943e72f-5a3b-4f9f-a54d-35e33d018dea
Marsh, Dan
df5ab9c4-9889-4093-a635-92c5c8508104
Lowdell, Mark W.
399d3d5b-dcd3-4b23-a6f9-7141c0717f56
Meyer, Tim
28c8fc8e-873f-4278-9443-cb7c9f9f487f
Begent, Richard H.J.
8d732fc3-678d-439a-ba03-9cc57a69a7ca
Hart, Ian
303b970c-96ed-49ec-9112-c5b9dd6d4165
Marshall, John F.
1134b21f-7f23-4ee6-aaf5-c7580b73c10b
Chester, Kerry
c7014f1a-71d9-4ca0-959d-2164f2b9b3e2

Kogelberg, Heide, Tolner, Berend, Thomas, Gareth J., Di Cara, Danielle, Minogue, Shane, Ramesh, Bala, Sodha, Serena, Marsh, Dan, Lowdell, Mark W., Meyer, Tim, Begent, Richard H.J., Hart, Ian, Marshall, John F. and Chester, Kerry (2008) Engineering a single chain Fv antibody to ?v?6 integrin using the specificity-determining loop of a foot-and-mouth disease virus. Journal of Molecular Biology, 382 (2), 385-401. (doi:10.1016/j.jmb.2008.07.013).

Record type: Article

Abstract

The ?v?6 integrin is a promising target for cancer therapy. Its expression is up-regulated de novo on many types of carcinoma where it may activate transforming growth factor-?1 and transforming growth factor-?3, interact with the specific extracellular matrix proteins and promote migration and invasion of tumor cells. The viral protein 1 (VP1) coat protein of the O1 British field strain serotype of foot-and-mouth disease virus is a high-affinity ligand for ?v?6, and we recently reported that a peptide derived from VP1 exhibited ?v?6-specific binding in vitro and in vivo. We hypothesized that this peptide could confer binding specificity of an antibody to ?v?6. A 17-mer peptide of VP1 was inserted into the complementarity-determining region H3 loop of MFE-23, a murine single-chain Fv (scFv) antibody reactive with carcinoembryonic antigen (CEA). The resultant scFv (B6-1) bound to ?v?6 but retained residual reactivity with CEA. This was eliminated by point mutation (Y100bP) in the variable heavy-chain domain to create an scFv (B6-2) that was as structurally stable as MFE-23 and reacted specifically with ?v?6 but not with ?5?1, ?v?3, ?v?5, ?v?8 or CEA. B6-2 was internalized into ?v?6-expressing cells and inhibited ?v?6-dependent migration of carcinoma cells. B6-2 was subsequently humanized. The humanized form (B6-3) was obtained as a non-covalent dimer from secretion in Pichia pastoris (115 mg/l) and was a potent inhibitor of ?v?6-mediated cell adhesion. Thus, we have used a rational stepwise approach to create a humanized scFv with therapeutic potential to block ?v?6-mediated cancer cell invasion or to deliver and internalize toxins specifically to ?v?6-expressing tumors.

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More information

Published date: 3 October 2008
Keywords: antibody engineering, scFv, integrin ?v?6, VP1 peptide, foot-and-mouse disease virus

Identifiers

Local EPrints ID: 66684
URI: http://eprints.soton.ac.uk/id/eprint/66684
ISSN: 0022-2836
PURE UUID: d7c8a2bb-61d3-4e61-8fd9-4c95a0f2d2dd

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Date deposited: 10 Jul 2009
Last modified: 13 Mar 2024 18:28

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Contributors

Author: Heide Kogelberg
Author: Berend Tolner
Author: Danielle Di Cara
Author: Shane Minogue
Author: Bala Ramesh
Author: Serena Sodha
Author: Dan Marsh
Author: Mark W. Lowdell
Author: Tim Meyer
Author: Richard H.J. Begent
Author: Ian Hart
Author: John F. Marshall
Author: Kerry Chester

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