CRISPR/Cas systems to overcome challenges in developing the next generation of T cells for cancer therapy

Abstract

Genetically engineered immune cells with chimeric antigen receptors (CAR) or modified T cell receptors (TCR) have demonstrated their potential as a potent class of new cancer therapeutic strategy. Despite the clinical success of autologous CD19 CAR T cells in hematological malignancies, allogeneic T cells exhibit many advantages over their autologous counterparts and have recently gathered widespread attention due to the emergence of multiplex genome editing techniques, particularly CRISPR/Cas systems. Furthermore, genetically engineered T cells face a host of major challenges in solid tumors that are not as significant for blood cancers such as T cell targeted delivery, target specificity, proliferation, persistence, and the immunosuppressive tumor microenvironment. We take this opportunity to analyze recent strategies to develop allogeneic T cells, specifically in consideration of CRISPR/Cas and its delivery systems for multiplex gene editing. Additionally, we discuss the current methods used to delivery CRISPR/Cas systems for immunotherapeutic applications, and the challenges to continued development of novel delivery systems. We also provide a comprehensive analysis of the major challenges that genetically engineered T cells face in solid tumors along with the most recent strategies to overcome these barriers, with an emphasis on CRISPR-based approaches. We illustrate the synergistic prospects for how the combination of synthetic biology and immune-oncology could pave the way for designing the next generation of precision cancer therapy.

Keywords: Adoptive cell therapy; Allogeneic T cells; CRISPR/Cas delivery systems; Cancer immunotherapy; Chimeric antigen receptor; Solid tumor.

Figure 1.

Illustration depicting the structure of T cell receptors and chimeric antigen receptors. Reprinted from [12]. Copyright 2018, with permission from Elsevier.

Figure 2.

A single guided RNA molecule (sgRNA), constructed by fusing the tracrRNA with the crRNA, can base pair with complementary regions of the DNA located next to a protospacer adjacent motif (PAM). With help of the Cas9 nuclease, the CRISPR/Cas9 system creates double-stranded breaks in the DNA, which can then be repaired using one of two pathways: non-homologous end joining or homology-directed repair.

Figure 3.

Schematic of the tumor microenvironment and the major barriers associated with T cell infiltration and signaling.

Figure 4.

Schematic of various strategies utilizing “AND”, “OR”, and “NOT” Boolean logic gates to enhance tumor specificity of adoptive cell therapy.

Figure 5.

Schematic of the titratable and reversible “ON-switch” mechanism through a heterodimerizing small molecule to allow for precise remote control. From [116]. Reprinted with permission from AAAS.

Figure 6.

Schematic of various costimulatory and coinhibitory signals for T cell activation and related pro-inflammatory and anti-inflammatory cytokines.