title: Harnessing Syk family tyrosine kinases as signaling domains for chimeric single chain of the variable domain receptors: optimal design for T cell activation.
abstract: T cells of tumor bearers often show defective TCR-mediated signaling events and, therefore, exhibit impaired immune responses. As such, patients with heavy tumor burden are often not amenable to adoptive T cell therapy. To overcome this limitation, we have developed a chimeric receptor that joins an extracellular single chain Fv (scFv) of a specific Ab for Ag recognition to an intracellular protein tyrosine kinase (PTK) for signal propagation. Stimulation through the scFv-PTK receptor should bypass defective TCR-proximal events and directly access the T cell's effector mechanisms. In this study we describe the optimization of a scFv-PTK configuration, leading to complete T cell activation. The cytosolic PTK Syk is superior to its family member, Zap-70, for intracellular signaling. As a transmembrane (TM) domain, CD4 performs better than CD8 when plastic-immobilized Ag serves as a stimulator. However, when APC are used to trigger chimeric receptors, the need for a flexible spacer between the scFv and TM domains becomes apparent. The CD8alpha-derived hinge successfully performs this task in chimeric scFv-Syk receptors regardless of its cysteine content. A cytotoxic T cell hybridoma expressing chimeric receptor genes composed of scFv-CD8(hinge)-CD8(TM)-Syk or scFv-CD8(hinge)-CD4(TM)-Syk is efficiently stimulated to produce IL-2 upon interaction with APC and specifically lyses appropriate target cells in a non-MHC-restricted manner.
authors.affiliation: Department of Immunology, The Weizmann Institute of Science, Rehovot, Israel.
authors.forename: C J
authors.initials: CJ
authors.lastname: Fitzer-Attas
authors.forename: D G
authors.initials: DG
authors.lastname: Schindler
authors.forename: T
authors.initials: T
authors.lastname: Waks
authors.forename: Z
authors.initials: Z
authors.lastname: Eshhar
mesh_terms: D000818:Animals; D015704:CD4 Antigens / Q000737:chemistry; D016827:CD8 Antigens / Q000737:chemistry; D002478:Cells, Cultured; D003545:Cysteine / Q000737:chemistry; D003602:Cytotoxicity, Immunologic; D019281:Dimerization; D004792:Enzyme Precursors / Q000502:physiology*; D047908:Intracellular Signaling Peptides and Proteins; D008213:Lymphocyte Activation; D051379:Mice; D011505:Protein-Tyrosine Kinases / Q000502:physiology*; D020794:Receptor Protein-Tyrosine Kinases / Q000737:chemistry*; D011948:Receptors, Antigen, T-Cell / Q000737:chemistry*; D011993:Recombinant Fusion Proteins; D013329:Structure-Activity Relationship; D000072377:Syk Kinase; D051746:ZAP-70 Protein-Tyrosine Kinase
chemical_list: D015704:CD4 Antigens; D016827:CD8 Antigens; D004792:Enzyme Precursors; D047908:Intracellular Signaling Peptides and Proteins; D011948:Receptors, Antigen, T-Cell; D011993:Recombinant Fusion Proteins; D011505:Protein-Tyrosine Kinases; D020794:Receptor Protein-Tyrosine Kinases; D000072377:Syk Kinase; C000606897:Syk protein, mouse; D051746:ZAP-70 Protein-Tyrosine Kinase; C497262:Zap70 protein, mouse; D003545:Cysteine