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Abstract

CD4⁺ T cells with pre-defined MHC-unrestricted specificity to type II collagen (CII) were engineered for cell-based anti-inflammatory gene therapy of autoimmune arthritis. To this end, recombinant chimeric immunoreceptors, C2γ or C2ζ, were expressed in primary mouse keyhole limpet hemocyanin (KLH)-specific T_h1 and T_h2 cells using retrovirus vector-based somatic cell gene transfer. The ectodomain of these tyrosine-based activation motif (ITAM)-containing immunoreceptors is a single-chain IgG variable domain of an anti-CII mAb. The engineered cells might arrest migration when they encounter CII in articular cartilage. Up to 19 and 55% of transduced CD4⁺ T cells expressed respectively C2γ and C2ζ. The expression of C2γ or C2ζ on the surface of CD4⁺ T cells was down-regulated upon binding CII, and cells activated in such a way proliferated, up-regulated CD25 expression and produced cytokines. Comparison of cytokine levels normalized by the number of producer cells revealed that C2γ and C2ζ were as potent as TCR in the induction of IFN-γ, but induced lower levels of IL-4. It appears that the reason why CD4⁺ T cells stimulated through C2γ and C2ζ produce low levels of IL-4 is a lack of integration between co-stimulatory signals required for the optimal production of this cytokine and the ITAM-dependent signals generated by the immunoreceptors. The significance of these data for the development of anti-inflammatory gene therapy based on CD4⁺ T cells targeted to a tissue-specific protein is discussed.