Chimeric antigen receptor containing ICOS signaling domain mediates specific and efficient antitumor effect of T cells against EGFRvIII expressing glioma

Abstract

BackgroundAdoptive transfer of chimeric antigen receptor (CAR)-modified T cells appears to be a promising immunotherapeutic strategy. CAR combines the specificity of antibody and cytotoxicity of cytotoxic T lymphocytes, enhancing T cells’ ability to specifically target antigens and to effectively kill cancer cells. Recent efforts have been made to integrate the costimulatory signals in the CAR to improve the antitumor efficacy. Epidermal growth factor receptor variant III (EGFRvIII) is an attractive therapeutic target as it frequently expresses in glioma and many other types of cancers. Our current study aimed to investigate the specific and efficient antitumor effect of T cells modified with CAR containing inducible costimulator (ICOS) signaling domain.MethodsA second generation of EGFRvIII/CAR was generated and it contained the EGFRvIII single chain variable fragment, ICOS signaling domain and CD3ζ chain. Lentiviral EGFRvIII/CAR was prepared and human CD3+ T cells were infected by lentivirus encoding EGFRvIII/CAR. The expression of EGFRvIII/CAR on CD3+ T cells was confirmed by flow cytometry and Western blot. The functions of EGFRvIII/CAR+ T cells were evaluated using in vitro and in vivo methods including cytotoxicity assay, cytokine release assay and xenograft tumor mouse model.ResultsChimeric EGFRvIIIscFv-ICOS-CD3ζ (EGFRvIII/CAR) was constructed and lentiviral EGFRvIII/CAR were made to titer of 106 TU/ml. The transduction efficiency of lentiviral EGFRvIII/CAR on T cells reached around 70% and expression of EGFRvIII/CAR protein was verified by immunoblotting as a band of about 57 kDa. Four hour 51Cr release assays demonstrated specific and efficient cytotoxicity of EGFRvIII/CAR+ T cells against EGFRvIII expressing U87 cells. A robust increase in the IFN-γ secretion was detected in the co-culture supernatant of the EGFRvIII/CAR+ T cells and the EGFRvIII expressing U87 cells. Intravenous and intratumor injection of EGFRvIII/CAR+ T cells inhibited the in vivo growth of the EGFRvIII expressing glioma cells.ConclusionsOur study demonstrates that the EGFRvIII/CAR-modified T cells can destroy glioma cells efficiently in an EGFRvIII specific manner and release IFN-γ in an antigen dependent manner. The specific recognition and effective killing activity of the EGFRvIII-directed T cells with ICOS signaling domain lays a foundation for us to employ such approach in future cancer treatment.

ICOS-based chimeric antigen receptors program bipolar TH17/TH1 cells

“…CAR T cells with ICOS domains retain a "bipolar" bias toward a T H 1 and T H 17 signature, which is a plasticity that appears to be a prerequisite for potent antitumor activity in several preclinical models. 17 Shen et al constructed an ICOS-based CAR that recognizes EGFRvIII, 33 and demonstrated that T H 1 cells redirected with this CAR could release IFN-g and destroy glioma cells in an antigen-dependent manner. Our results confirm and extend this work by performing a comprehensive analysis of CARs expressing ICOS signaling domains.…”

β-catenin and PI3Kδ inhibition expands precursor Th17 cells with heightened stemness and antitumor activity

“…In the context of adoptive immunotherapy, ICOS signaling enhances the antitumor activity of human and murine Th17 cells and Tc17 cells (9,51). Moreover, introducing the ICOS domain into CARs enhanced cellular engraftment in vivo (52)(53)(54). In light of our work, we suspect that combining drugs or genetic strategies to regulate p110δ and β-catenin induced in T cells could augment various forms of cancer immunotherapy, including cytokine therapy, checkpoint modulators, and cellular therapy.…”

CARTs for Solid Tumors: Feasible or Infeasible?

“…First-generation CARs include only immunoreceptor tyrosine-based activation motifs (ITAMs) from the CD3ζ chain or the FcR-γ chain as an intracellular signaling domain [8], whereas second-generation CARs include a single costimulatory domain derived from CD28 [42,43], CD134 (OX40) [44], CD137 (4-1BB) [45,46], CD27 [47], or inducible costimulator (ICOS) [48,49], and third-generation CARs include 2 or more costimulatory domains in addition to the CD3ζ chain or FcR-γ chain [50,51].…”