Increasing the safety and efficacy of chimeric antigen receptor T cell therapy

Abstract

Chimeric antigen receptor (CAR) T cell therapy is a promising cancer treatment that has recently been undergoing rapid development. However, there are still some major challenges, including precise tumor targeting to avoid off-target or "on-target/off-tumor" toxicity, adequate T cell infiltration and migration to solid tumors and T cell proliferation and persistence across the physical and biochemical barriers of solid tumors. In this review, we focus on the primary challenges and strategies to design safe and effective CAR T cells, including using novel cutting-edge technologies for CAR and vector designs to increase both the safety and efficacy, further T cell modification to overcome the tumor-associated immune suppression, and using gene editing technologies to generate universal CAR T cells. All these efforts promote the development and evolution of CAR T cell therapy and move toward our ultimate goal-curing cancer with high safety, high efficacy, and low cost.

Keywords: T lymphocytes; cancer adoptive immunotherapy; chimeric antigen receptors; gene editing; gene therapy.

Figure 1

CAR design. CARs are engineered membrane proteins that consist of three main components: an extracellular antigen-recognition domain, a hinge and transmembrane domain, and an intracellular T cell activation signaling domain. First-generation CARs have only a CD3ζ signaling domain in the intracellular part providing signal 1 for T cell activation. Second-generation CARs have an extra signal 2 domain in the intracellular domain. Third-generation CARs have three domains in the intracellular part, two signal 2 domains and one signal 1 domain. ScFv, single chain variable fragment; ICOS, inducible T cell costimulator; TM, transmembrane

Figure 2

Antigen A AND antigen B combinatorial antigen recognition to optimize the specificity of CAR T recognition and reduce the “on-target/off-tumor” toxicity. (A) Two-CAR design strategy: one CAR molecule for activation and another CAR molecule for costimulation in the same T cell. (B) SynNotch AND-gate circuit strategy: the synNotch receptor recognizes antigen B first, then induces the expression of CAR receptor, which recognizes antigen A, and finally activates the T cell. SynNotch, synthetic Notch receptor, which recognizes a target antigen B, then cleaves the receptor to release a transcriptional activator domain to enter the nucleus and induce the expression of CAR; ScFv, single chain variable fragment; ICOS, inducible T cell costimulator; TM, transmembrane

Figure 3

Armed CAR T cell with “OFF switch,” suicide gene—iCasp9 to kill itself by apoptosis. The iCasp9 gene and CAR gene are linked by the 2A sequence in the CAR vector, and therefore, iCasp9 is co-expressed with the CAR molecule in CAR T cells. Once the small molecule dimerizer chemical inducer of dimerization (CID) AP20187 is added, it binds to iCasp9, triggering apoptosis. iCasp9, inducible suicide gene caspase-9; Casp3, caspase-3; ScFv, single chain variable fragment; ICOS, inducible T cell costimulator; LTR, long terminal repeats; TM, transmembrane

Figure 4

CAR T cell with “OFF switch”—co-expression with a truncated protein or a small epitope peptide. In the presence of the mAb of the anti-epitope/truncated protein, the mAb binds the truncated protein or epitope peptide and mediates antibody-dependent cell-mediated cytotoxicity (ADCC) or complement-dependent cytotoxicity (CDC) to trigger cell lysis. The design strategy of an AND logic gate that requires “antigen + mAb”. NK cell, natural killer cells; FcR, Fc receptor; cetuximab, a chimeric monoclonal antibody against epidermal growth factor receptor; huEGFRt, a truncated human EGFR polypeptide; ScFv, single chain variable fragment; ICOS, inducible T cell costimulator; TM, transmembrane; EF1p, elongation factor 1 promoter sequences; GMCSFRss, the GM-CSF receptor-α chain signal sequences; RQR8, a 136-amino-acid marker with epitopes for CD34 and CD20 as a suicide molecule. SAR, scaffold attachment region

Figure 5

CAR T cells with “ON switch”—small molecules. Only when the small molecule is present can two parts of CAR be reassembled with the split construction to activate CAR T cells. The design strategy of an AND logic gate that requires “antigen + small molecule” together for T cell activation. ScFv, single chain variable fragment; ICOS, inducible T cell costimulator; TM, transmembrane

Figure 6

CAR T cells with “ON/OFF switch”. The “ON” switch utilizes the Tet-On inducible system. Only when tetracycline (Tet) is present, rtTA (r everse t etracycline-controlled t rans a ctivator) binds with Tet and induces the expression of the CAR molecule and marker protein, such as truncated EGFR (tEGFR). The “OFF” switch utilizes mAb-mediated ADCC or CDC. Only when the anti-marker protein Ab (such as cetuximab) is present does the mAb bind with the marker protein and mediate ADCC or CDC to lyse CAR T cells

Figure 7

Strategies for CAR T therapy to overcome challenges in treating cancers. CAR T therapy is not only a complex technology that requires the optimization of the CAR T cell design but also a clinical application that can be combined with other therapies to achieve high efficacy and safety