Forman, Stephen J.,Brown, Christine E.,Jonnalagadda, Umamaheswararao,Mardiros, Armen
申请号：
ES14877876
公开号：
ES2767423T3
申请日:
2014.03.14
申请国别(地区):
ES
年份:
2020
代理人:
摘要：
Adoptive immunotherapy using T cells genetically redirected via expression of chimeric antigen receptors (CARs) is a promising approach for cancer treatment. However, this immunotherapy is dependent in part on the optimal molecular design of the CAR, which involves an extracellular ligand-binding domain connected to an intracellular signaling domain by spacer and/or transmembrane sequences. CAR designs frequently incorporate extracellular linker regions based on the immunoglobulin constant regions of either IgG1 or IgG4. In this study, the ability of the IgG4-Fc linker to result in off-target interactions between the CAR and Fc gamma receptors (FcγRs) was examined. As proof of principle, a CD19-specific scFv-IgG4-CD28-zeta CAR was studied, and it was determined that, in contrast to CAR-negative T cells that only expressed an EGFRt tracking marker, CAR+ T cells did not engraft in NSG mice and, indeed, were bound to soluble FcγRs. As such, mutations to avoid FcγR binding would improve CAR+ T cell persistence and anti-tumor efficacy were designed. Consequently, a CD19-specific CAR that has been mutated at two sites within the CH2 region (L235E； N297Q) of the IgG4 Fc spacer (CD19R(EQ)), as well as a CD19-specific CAR that has a CH2 deletion in its IgG4 Fc spacer (CD19Rch2Δ) were generated. These mutations/the deletion do not alter the functional ability of the CAR, when expressed by T cells, to mediate antigen-specific lysis of tumor cells. However, compared to T cells that express a non-mutated CAR, T cells expressing the CD19R(EQ) and CD19Rch2Δ exhibit impaired binding to recombinant soluble FcγRs. These CD19R(EQ) and CD19Rch2Δ T cells also exhibit improved engraftment in NSG mice, to levels similar to that seen with CAR-negative/EGFRt+ T cells. Importantly, the CD19R(EQ) and CD19Rch2Δ T cells also exhibited significantly improved CD19-specific anti-lymphoma efficacy in NSG mice. Together these studies provide evidence that optimal CAR function necessitates the elimin
