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Poster Display session

333 - Pooling signaling and costimulatory domains in a flexible CARpool design


14 Dec 2018


Poster Display session


Lorraine Springuel


Annals of Oncology (2018) 29 (suppl_10): x11-x16. 10.1093/annonc/mdy485


L. Springuel, J. Bolsée, A. Velghe, S. Agaugué, D. Gilham

Author affiliations

  • Research & Development, Celyad SA, 1435 - Mon-Saint-Guibert/BE


Abstract 333


Chimeric Antigen Receptors (CARs) consist of a target binding moiety fused to an extracellular spacer, a transmembrane region and an intracellular signaling tail comprising a tandem alignment of co-stimulatory and activatory domains. This linear configuration displays rigid spatial orientation and ratio of co-stimulation to activation domains. We have developed a novel mix and match approach (CARpool) where the costimulatory signal is provided in trans on accessory proteins that associate with the antigen binding chain via transmembrane interactions.


Several CD3ζ-containing CAR chains were designed using the transmembrane and cytoplasmic domains of NKG2D or NKp44, associating with DAP10 and DAP12 respectively. Each CAR contained a B7H6-targeting scFv and was co-expressed with corresponding accessory protein using a 2A site. Primary human T cells engineered with the diverse constructs were screened for CAR expression, phenotype and in vitro function.


NKG2D-based CAR complexes were moderately expressed at the cell surface but bound B7H6 and led to potent cells. Modification of the position of the charged residue within the transmembrane domain of the CAR is being used to modulate the surface expression and thus their potency. NKp44-based CAR complexes were more frequently expressed on primary T cells and bound B7H6, though functionality appears to be dependent on the nature of the extracellular spacer.


These studies provide proof-of-concept for a novel CAR design where it is possible to incorporate or interchange costimulatory domain(s) in a stoichiometrically controlled way. Recapitulating physiological TCR activation by providing co-stimulation in trans within the CARpool may result in optimal downstream signaling, thereby enhancing anti-tumoral activity.

Editorial acknowledgement

Clinical trial identification

Legal entity responsible for the study

Celyad SA.


Celyad SA.


L. Springuel, J. Bolsée, A. Velghe, S. Agaugué, D. Gilham: Employee of Celyad SA.

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