DEVELOPMENT AND OPTIMISATION OF A FULLY HUMAN FVIII MIMETIC BISPECIFIC ANTIBODY FOR PATIENTS WITH HAEMOPHILIA A
Author(s): ,
Wei Wang
Affiliations:
Kymab Ltd,Cambridge,United Kingdom
,
John Blackwood
Affiliations:
Kymab Ltd,Cambridge,United Kingdom
,
Roberto Magliozzi
Affiliations:
Kymab Ltd,Cambridge,United Kingdom
,
Leonardo Moraes
Affiliations:
Kymab Ltd,Cambridge,United Kingdom
,
Alla Fane-Dremucheva
Affiliations:
Kymab Ltd,Cambridge,United Kingdom
,
Ana Camacho
Affiliations:
University of Bath,Bath,United Kingdom;Kymab Ltd,Cambridge,United Kingdom
,
Andrew Wood
Affiliations:
Kymab Ltd,Cambridge,United Kingdom
,
Benjamin Grimshaw
Affiliations:
Kymab Ltd,Cambridge,United Kingdom
,
Benjamin Jenkins
Affiliations:
University of Bath,Bath,United Kingdom;Kymab Ltd,Cambridge,United Kingdom
,
Hannah Craig
Affiliations:
Kymab Ltd,Cambridge,United Kingdom
,
Jacob Galson
Affiliations:
Kymab Ltd,Cambridge,United Kingdom
,
Hui Liu
Affiliations:
Kymab Ltd,Cambridge,United Kingdom
,
Lauriane Gamand
Affiliations:
Kymab Ltd,Cambridge,United Kingdom
,
Luca Badiali
Affiliations:
Kymab Ltd,Cambridge,United Kingdom
,
Margot Billaud
Affiliations:
Kymab Ltd,Cambridge,United Kingdom
,
Nick England
Affiliations:
Kymab Ltd,Cambridge,United Kingdom
,
Peter Thomas
Affiliations:
Kymab Ltd,Cambridge,United Kingdom
,
Vivian Wong
Affiliations:
Kymab Ltd,Cambridge,United Kingdom
,
Volker Germaschewski
Affiliations:
Kymab Ltd,Cambridge,United Kingdom
,
Allan Bradley
Affiliations:
Kymab Ltd,Cambridge,United Kingdom
E-Chiang Lee
Affiliations:
Kymab Ltd,Cambridge,United Kingdom
EHA Library. Wang W. Jun 14, 2019; 266138; PF338
Wei Wang
Wei Wang
Contributions
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Abstract

Abstract: PF338

Type: Poster Presentation

Presentation during EHA24: On Friday, June 14, 2019 from 17:30 - 19:00

Location: Poster area

Background

Recently a range of alternative novel therapies have been developed to improve treatment options for patients with Hemophilia A. One approach is to generate a Factor VIII (F.VIII) mimetic molecule using a humanised bispecific antibody, as was done for Hemlibra. Taking advantage of Kymab’s fully human antibody discovery platform, we describe the selection and optimisation of an FVIII mimetic common light chain (CLC) bispecific antibody which can similarly catalyse the generation of Factor Xa (FXase) and normalise the activated partial thromboplastin time (aPTT).

Aims

To generate a functionally active F.VIII mimetic bispecific antibody for Haemophilia A treatment 

Methods

F.IX and F.X. binding antibodies were generated by immunizing Kymouse™, which contain the full human immunoglobin repertoire, with F.IX or F.X, respectively. Isolated F.IX and F.X specific arms were co-expressed as 2-heavy-2-light-chain (2H2L) bispecific antibodies. Purified 2H2L bispecifics were screened using a high-throughput chromogenic FXase assay. The light chain of a promiscuous F.IX arm was chosen to generate transgenic mice expressing this bespoke common light chain (CLC) in the Kymouse™ background. By immunizing these transgenic mice with F.X, F.X binding antibodies containing the CLC were recovered. The heavy chains of these F.X binding antibodies were co-expressed with the heavy/light chains of the chosen F.IX arm as CLC bispecific antibodies. One biologically active CLC bispecific antibody was identified by functional assays, and chosen for further optimization. The optimization of the lead bispecific antibody, KY1049, was achieved by data mining of next generation sequencing information using Kymab’s IntelliSelect™ bioinformatic platform, coupled with site-specific mutagenesis.

Results

More than 8,000 2H2L bispecifics were screened by a chromogenic FXase assay to select the most active and versatile F.IX arm (Figure 1A). More than 400 F.X heavy chains subsequently isolated from the bespoke CLC Kymouse™ were screened to identify a highly active CLC bispecific (Figure 1B). Further optimisation of the molecule was carried out to iteratively increase FVIII mimetic activity by deep data-mining of heavy chain NGS sequence data, or site-specific mutagenesis. The combinatorial optimization process resulted in a highly functional CLC bispecific, KY1049 with comparable F.VIII mimetic activities to a sequence-identical analogue of Hemlibra (Figure 1C).

Conclusion

Our bispecific antibody discovery platform consisting of four-chain matrix screening, common light chain transgenic mouse technology, B cell network analysis and site-specific mutagenesis was applied to develop KY1049, a potent FV.III mimetic bispecific antibody, which shows equivalent activity to Hemlibra in vitro and holds promise as a future therapeutic in Haemophilia A.

Session topic: 33. Bleeding disorders (congenital and acquired)

Keyword(s): Hemophilia A, Hemostasis

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