Retrovirally engineered T‐cell‐based immunotherapy targeting type III variant epidermal growth factor receptor, a glioma‐associated antigen

Abstract

The isotype of epidermal growth factor receptor variant III (EGFRvIII) is often identified in glioblastomas. Previously, we created a mouse monoclonal antibody, 3C10 (IgG2b), that specifically recognized EGFRvIII, and a recombinant single-chain variable fragment of 3C10. The aim of the current study was to develop genetically engineered T cells, termed T-bodies, that express a chimeric receptor consisting of the 3C10 single-chain variable fragment coupled to signaling modules such as the CD3zeta (f) chain, for the treatment of tumors expressing mutant EGFR. After successful construction of the chimeric 3C10 ⁄ CD3f T-cell receptor, its expression on the T-body was observed using western blotting and flow cytometry. The specificity of the T-body for EGFRvIII was evaluated using an interferon-gamma Elispot assay and a standard 51 Cr-release cytotoxicity assay. Furthermore, we demonstrated that the systemically delivered T-body infiltrated the intrabrain tumor and significantly delayed tumor growth. These results indicate that the T-body expressing the chimeric 3C10 ⁄ CD3f T-cell receptor specifically recognized glioma cells expressing EGFRvIII. In conclusion, T-body-based immunotherapy appears to be a promising approach for the treatment of glioma. (Cancer Sci 2010; 101: 2518-2524 T he expression of epidermal growth factor receptor (EGFR) is amplified in approximately 50% of glioblastomas (GBM).(1) The binding of a ligand to EGFR leads to receptor dimerization, autophosphorylation and activation of several downstream signaling pathways such as the Ras ⁄ Raf ⁄ MEK ⁄ ERK pathway, the PI3K ⁄ Akt pathway and the PLC-gamma (c) ⁄ PKC pathway, resulting in cell proliferation, motility and survival.(2) Approximately 40-70% of brain tumors with EGFR amplification express mutant EGFR variant III (EGFRvIII); EGFRvIII has a deletion of exons 2-7 that causes a defect in the extracellular ligand-binding domain and induces constitutive activation in a ligand-independent manner.(3-6) Notably, EGFRvIII is characterized by an 801-base pair (bp) in-frame deletion, which results in a unique sequence with a glycine residue at the fusion junction between amino acid residues 5 and 274. Epidermal growth factor receptor variant III is an attractive target antigen for cancer immunotherapy because it is not expressed in normal tissue and is associated with survival, invasion and angiogenesis in cancers.(4,7) Previously, we generated the monoclonal antibody (mAb) 3C10 and a recombinant singlechain variable fragment (scFv) antibody (Ab) that specifically recognizes EGFRvIII.

Complement‐mediated tumor‐specific cell lysis by antibody combinations targeting epidermal growth factor receptor (EGFR) and its variant III (EGFRvIII)

“…Due to its tumor specificity, EGFRvIII is currently also being investigated as a target antigen in vaccination trials, (13) and for other therapeutic approaches. (28,29) The EGFR mAb act via Fab-and Fc-mediated effector mechanisms. (5) For example, zalutumumab blocks EGFR signaling by inhibition of ligand binding and receptor downmodulation and induces efficient ADCC, a Fc-mediated mechanism.…”

CAR T Cells Targeting Podoplanin Reduce Orthotopic Glioblastomas in Mouse Brains

“…17). In addition, CAR-transduced T cells can migrate through the microvascular walls and penetrate tumors (14,16). Clinical trials are ongoing using CAR T-cell therapy against GBM targeting EGFRvIII and HER2 (18,19).…”

“…We and other groups have generated several CARs against the antigens expressed in GBM, including epidermal growth factor receptor variant III (EGFRvIII; ref. 14), human epidermal growth factor receptor 2 (HER2; ref. 15), IL13 receptor alpha 2 (IL13Ra2; ref.…”

Chimeric Antigen Receptor Therapeutic Strategies: The Future of Glioblastoma Management

“…These second and third generation CAR avoided a major pitfall of first generation CAR, namely the limited functional lifespan of CAR-T cells and the occurrence of quiescence in the absence of costimulatory signals. Clinical trials have demonstrated significant long-term disease free survival in patients with hematologic malignancies using second generation CAR-T cells [2,16], and both second and third generation CAR-T cells have been shown to be efficacious against murine glioma models [17][18][19][20][21][22].…”