Engineering enhanced CAR T-cells for improved cancer therapy

Abstract

Chimeric antigen receptor (CAR) T-cell therapies have evolved from a research tool to a paradigm-shifting therapy with impressive responses in B cell malignancies. This review summarizes the current state of the CAR T-cell field, focusing on CD19- and B cell maturation antigen-directed CAR T-cells, the most developed of the CAR T-cell therapies. We discuss the many challenges to CAR-T therapeutic success and innovations in CAR design and T-cell engineering aimed at extending this therapeutic platform beyond hematologic malignancies.

Keywords: CAR; T cell; antigen; cancer; chimeric.

Figure 1.

B cell malignancies at the different stages of B cell development. The normal B-cell developmental lymphocytes shown at the top, often share the same immunophenotypic characteristics with the malignant counterparts depicted at the bottom, reflecting the expansion of a dominant clone leading to development of leukemia or lymphoma.

Figure 2.

Operational pipeline for integrating correlative studies in translational science laboratories. Novel therapies developed and pre-clinically validated in research laboratories are handed off to the process development (PD) team for scale-up and the development of a current Good Manufacturing Practice (GMP) process. In collaboration with the GMP teams Standard Operating Procedures (SOP) and documentation forms are developed and GMP staff trained in the new procedures. The Correlative Studies Laboratory will, in parallel, ensure that all supportive assays, protocols, and forms are in place, that staff is trained, and that routine, qualified assays are developed and biobanking ensured. This same team is also involved in protocol development, which is lead by the Clinical Operations team with feedback from the study clinicians and the research laboratory that developed the new process. When a new clinical trial begins the Correlative Studies laboratory starts receiving biospecimens from the clinic, manufacturing facility, or collaborating laboratories, and logs these samples into the Laboratory Information Management System (LIMS), to be processed as specified by standard operating procedures and examined using validated assays by qualified personnel. Aliquots are retained from each specimen for future translational studies. The data are reviewed by subject matter experts (SME) before being reviewed by the quality control (QC) manager and entered into a database. A staff statistician cleans and analyzes the data for reporting purposes, e.g. to FDA or for scientific meetings and manuscript preparation.

Figure 3.

Current strategies to overcome the hurdles of poor response to autologous CAR-T-cell therapy. Several factors, such as low frequencies of early memory CAR-T-cells in the infusion product, over-expression of checkpoint inhibitory molecules on the apheresis T-cells and loss of target antigen on the tumor, have been shown to contribute to the lack of efficacy of CAR-T-cells in many patients. Optimizing the manufacturing process by laboratory-based engineering approaches, such as memory T-cells enrichment, dural CAR development and specific gene editing, is essential to improve the quality of CAR-T-cell product, thereby enhancing its capacity of tumor clearance and in vivo persistence.

Figure 4.

CAR design limitations that affect clinical responses following CAR T-cell treatment and potential solutions. Most CARs are made up of an tumor-associated antigen-binding scFv fragment (e.g. CD19, fused in-frame with a T-cell signaling domain), enhanced with a co-stimulatory domain (e.g. CD28 or 4–1BB) that is separated from the scFv by a spacer sequence. The design of this synthetic receptor affects various aspects of its in vivo performance and ultimately clinical responses. Additionally, small molecules such as Dasatinib may tone dysfunction-inducing CAR signaling .