Anti-Tumor Activity of Human T Cells Expressing the CC49-zeta Chimeric Immune Receptor

Abstract

A chimeric immune receptor consisting of an extracellular antigen-binding domain derived from the CC49 humanized single-chain antibody, linked to the CD3zeta signaling domain of the T cell receptor, was generated (CC49-zeta). This receptor binds to TAG-72, a mucin antigen expressed by most human adenocarcinomas. CC49-zeta was expressed in CD4+ and CD8+ T cells and induced cytokine production on stimulation. Human T cells expressing CC49-zeta recognized and killed tumor cell lines and primary tumor cells expressing TAG-72. CC49-zeta T cells did not mediate bystander killing of TAG-72-negative cells. In addition, CC49-zeta T cells not only killed FasL-positive tumor cells in vitro and in vivo, but also survived in their presence, and were immunoprotective in intraperitoneal and subcutaneous murine tumor xenograft models with TAG-72-positive human tumor cells. Finally, receptor-positive T cells were still effective in killing TAG-72-positive targets in the presence of physiological levels of soluble TAG-72, and did not induce killing of TAG-72-negative cells under the same conditions. This approach is being currently being utilized in a phase I clinical trial for the treatment of colon cancer.

Impact of chimeric immune receptor extracellular protein domains on T cell function

“…To overcome these difficulties, several groups have described the genetic modification of T cells to express recombinant CIRs which mediate MHC-independent killing of target cells. These receptors comprise an extracellular antigen binding domain, derived from an antigen-specific antibody or ligand, coupled to an intracellular signaling domain derived either from the CD3 chain of the T cell receptor complex, [1][2][3][4][5] the Fc receptor (FcR) ␥ chain tumor [5][6][7] or more distal components of the T cell signal transduction pathway such as the syk molecule. 8 In this fashion, genemodified T cells can be redirected to recognize a broad assortment of tumor and viral antigens including HER-2/Neu (erb-B2), 3,5 HIV env , 4 folate binding protein, 6 IgE 9 and TAG-72.…”

“…8 In this fashion, genemodified T cells can be redirected to recognize a broad assortment of tumor and viral antigens including HER-2/Neu (erb-B2), 3,5 HIV env , 4 folate binding protein, 6 IgE 9 and TAG-72. 2 The scFv or ligand antigen binding domains in these CIRs must be coupled to the T cell receptor signaling moiety via a heterologous protein spacer domain to retain receptor function 10 or in certain receptors, even for receptor expression (D Smith, unpublished results). Most of the CIRs generated have employed the immunoglobulin hinge and constant regions as a heterologous protein spacer domain 1,2,4,5,7 although in the one comparative study of the impact of the length of the heterologous protein spacer domain on receptor expression and function, fragments of the CD4 or CD8 protein were employed.…”

“…2 The scFv or ligand antigen binding domains in these CIRs must be coupled to the T cell receptor signaling moiety via a heterologous protein spacer domain to retain receptor function 10 or in certain receptors, even for receptor expression (D Smith, unpublished results). Most of the CIRs generated have employed the immunoglobulin hinge and constant regions as a heterologous protein spacer domain 1,2,4,5,7 although in the one comparative study of the impact of the length of the heterologous protein spacer domain on receptor expression and function, fragments of the CD4 or CD8 protein were employed. 3 The different heterologous protein spacer domains used in the various CIRs that we have generated have shown a tremendous impact on receptor expression and function (J McArthur, unpublished results).…”

T-cell killing of heterogenous tumor or viral targets with bispecific chimeric immune receptors

“…11,12 The construction of the anti-gp120 ligand CD4-and anti-gp41 single-chain Ab 98.6-monospecific CIRs are described by Roberts et al, 17 the anti-HIV env gp120 447D-scFv CIR is described by Patel et al, 22 and the anti-TAG-72 scFv CIR is described by McGuinness et al 18 The BCIR-1 receptor (CC49-L205c-CEA(a 3 b 3 )-) directs the expression of a hybrid protein that consists of the V L secretion leader and scFv domain of CC49 joined at its C terminus (residue 245 of the mature scFv) through a 25-aa linker (SSADDAKKDAAKKDDAKKDDAKKDG (L205c)) to the a 3 b 3 region of CEA (residues 498 -675 of the mature polypeptide). The extracellular domains are joined through the C terminus of the CEA(a 3 b 3 ) domain to the CD4 transmembrane region (residues 372-395 of the mature polypeptide) and the CD3-cytoplasmic domains (residues 31-142 of the mature polypeptide).…”

“…15 The Ag recognition domain of all of these CIRs is linked to the intracellular-signaling domain of the CD3-subunit of the T-cell receptor (TCR), which, upon engagement of the receptor, induces T-cell activation, cytolytic function, and cytokine release. T cells expressing the CD4-and CC49-CIRs have been shown to kill HIV-infected cells 16,17 and TAG-72-expressing tumor cells, 18 respectively. A second anticancer CIR comprising the humanized scFv of the Col-1 Ab, 19 which binds CEA and mediates the lysis of CEA-expressing human adenocarcinoma cells, was also generated.…”