PXD049393-2

PXD049393 is an original dataset announced via ProteomeXchange.

Title	SpyMask Enables Combinatorial Assembly of Bispecific Binders
Description	Bispecific antibodies are a successful and expanding therapeutic class, bridging two cell-types or engaging two different molecules on the same cell. Standard approaches to generate bispecifics are complicated by the need for disulfide reduction/oxidation or specialized formats. Here we present SpyMask, a modular approach to bispecifics using SpyTag/SpyCatcher spontaneous amidation. Two SpyTag-fused antigen-binding modules can be precisely conjugated onto DoubleCatcher, a tandem SpyCatcher where the second SpyCatcher is protease-activatable. We engineer a panel of structurally-distinct DoubleCatchers, from which binders project in different directions. We establish a generalized methodology for one-pot assembly and purification of bispecifics in 96-well plate format. A panel of binders recognizing different HER2 epitopes were coupled to DoubleCatcher, revealing unexpected combinations with anti-proliferative or pro-proliferative activity on HER2-addicted cancer cells. Bispecific activity depended sensitively on both binder orientation and the geometry of DoubleCatcher scaffolds. These findings support the need for straightforward assembly in different formats. SpyMask provides a scalable tool to discover synergy in bispecific activity, through modulating receptor organization and geometry. We use mass spectrometry to validate the protein building blocks of our bispecific assemblies, as well as to confirm precise heterodimerization of antigen-binding proteins.
HostingRepository	PRIDE
AnnounceDate	2024-10-22
AnnouncementXML	Submission_2024-10-22_06:33:59.434.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Claudia Driscoll
SpeciesList	scientific name: Escherichia coli; NCBI TaxID: 562; scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	No PTMs are included in the dataset
Instrument	6550 iFunnel Q-TOF LC/MS; Xevo G2-S QTof

Revision	Datetime	Status	ChangeLog Entry
0	2024-02-15 04:07:36	ID requested
1	2024-03-18 10:15:03	announced
⏵ 2	2024-10-22 06:33:59	announced	2024-10-22: Updated project metadata.

10.1038/S41467-024-46599-9;

submitter keyword: chemical biology, immunotherapy, uncaging, tyrosine kinase, bifunctional

Mark R. Howarth
contact affiliation	Department of Biochemistry, University of Oxford, South Parks Road, Oxford, OX1 3QU, UK; Department of Pharmacology, University of Cambridge, Tennis Court Road, Cambridge, CB2 1PD, UK.
contact email	mh2186@cam.ac.uk
lab head
Claudia Driscoll
contact affiliation	University of Oxford, University of Cambridge
contact email	claudia.driscoll@bioch.ox.ac.uk
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2024/03/PXD049393

PRIDE project URI

• PRIDE
  1. PXD049393
     1. Label: PRIDE project
     2. Name: SpyMask Enables Combinatorial Assembly of Bispecific Binders