Preclinical Evaluation of CAR T Cell Function: In Vitro and In Vivo Models

Abstract

Immunotherapy using chimeric antigen receptor (CAR) T cells is a rapidly emerging modality that engineers T cells to redirect tumor-specific cytotoxicity. CAR T cells have been well characterized for their efficacy against B cell malignancies, and rigorously studied in other types of tumors. Preclinical evaluation of CAR T cell function, including direct tumor killing, cytokine production, and memory responses, is crucial to the development and optimization of CAR T cell therapies. Such comprehensive examinations are usually performed in different types of models. Model establishment should focus on key challenges in the clinical setting and the capability to generate reliable data to indicate CAR T cell therapeutic potency in the clinic. Further, modeling the interaction between CAR T cells and tumor microenvironment provides additional insight for the future endeavors to enhance efficacy, especially against solid tumors. This review will summarize both in vitro and in vivo models for CAR T cell functional evaluation, including how they have evolved with the needs of CAR T cell research, the information they can provide for preclinical assessment of CAR T cell products, and recent technology advances to test CAR T cells in more clinically relevant models.

Design and preclinical testing of an anti‐CD41 CAR T cell for the treatment of acute megakaryoblastic leukaemia

“…and European Medicines Agency (EMA), targeting B-cell lymphomas, multiple myeloma and acute B-cell lymphoblastic leukaemia, while CAR T cells against solid tumours showed inconsistent results. [36][37][38][39][40] As the in vitro testing is essential before translating the study to animal models, the co-culture between CAR T cells and the target may indicate the potential efficacy of these CAR T cells. By lowering the ratio between effector and target, we can recreate the 'real life' situation when CAR T cells represent a minority in the organism after infusion, 41 thus by analysing the interaction between CAR T cells and the target at different ratios, many situations can be evaluated.…”

A bioengineered immunocompetent human leukemia chip for preclinical screening of CAR T cell immunotherapy

“…Other types of resistance or relapse scenarios such as acquired resistance via surface antigen loss, down-regulation, and mutation following CAR T cell challenge have not been tested on-chip, which can be directions for improvement. Nevertheless, our platform can be assembled rapidly (half day) and allows at least a 14-day long evaluation of CAR T cell responses, which is of great convenience compared to immunocompetent animal models that can take up to half year or longer to establish, let alone month-long experiments subsequently [21][22][23][24]. More importantly, based on multi-dimensional and spatiotemporal measurements, we develop a matrix-based analytical and functional index to comprehensively delineate therapeutic functionality of CAR T cell of different designs or generations and confirm that a limited Th2 function may compromise CAR T cell performance as shown in CAR stress test scenario and patientderived CAR T cell products.…”

“…Current strategies for preclinical assessment of CAR T cell function rely on in vitro and in vivo models [21][22][23]. Conventional in vitro assays such as two-dimensional (2D) cell co-cultures and 3D tumor spheroids/organoids demonstrate limited predictive value due to their lack of tumor associated stroma and/or immune components, thus are just useful for CAR T cell development at early-stage.…”

“…Conventional in vitro assays such as two-dimensional (2D) cell co-cultures and 3D tumor spheroids/organoids demonstrate limited predictive value due to their lack of tumor associated stroma and/or immune components, thus are just useful for CAR T cell development at early-stage. In vivo animal models have been widely established for preclinical cancer research, however, most of them are either immunodeficient or differing from host immunity, dictating the pursuit of humanized models [21][22][23][24].…”

Relapse after CAR-T cell therapy in B-cell malignancies: challenges and future approaches

“…Future approaches to address or prevent relapse after CAR-T cell therapy in B-cell malignancies are destined to involve multiple aspects in accordance with the nature of tumor cells themselves, the properties of CAR-T cells, and the landscape within the specific TME. Developing suitable preclinical evaluation models will provide deep insights into the mechanisms behind tumor relapse and indicate CAR-T cell therapeutic potency in the clinic (Si et al, 2022).…”