Related Experiment Video
Updated: Aug 4, 2026
A Syngeneic Mouse B-Cell Lymphoma Model for Pre-Clinical Evaluation of CD19 CAR T Cells
Published on: October 16, 2018
Engineering CAR-NK cells to secrete IL-15 sustains their anti-AML functionality but is associated with systemic
Ilias Christodoulou1,2, Won Jin Ho1, Andrew Marple1
1Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
Background:
The prognosis of patients with recurrent/refractory acute myelogenous leukemia (AML) remains poor and cell-based immunotherapies hold promise to improve outcomes. Natural Killer (NK) cells can elicit an antileukemic response via a repertoire of activating receptors that bind AML surface ligands. NK-cell adoptive transfer is safe but thus far has shown limited anti-AML efficacy. Here, we aimed to overcome this limitation by engineering NK cells to express chimeric antigen receptors (CARs) to boost their anti-AML activity and interleukin (IL)-15 to enhance their persistence.
Methods:
We characterized in detail NK-cell populations expressing a panel of AML (CD123)-specific CARs and/or IL-15 in vitro and in AML xenograft models.
Results:
CARs with 2B4.ζ or 4-1BB.ζ signaling domains demonstrated greater cell surface expression and endowed NK cells with improved anti-AML activity in vitro. Initial in vivo testing revealed that only 2B4.ζ Chimeric Antigen Receptor (CAR)-NK cells had improved anti-AML activity in comparison to untransduced (UTD) and 4-1BB.ζ CAR-NK cells. However, the benefit was transient due to limited CAR-NK-cell persistence. Transgenic expression of secretory interleukin (sIL)-15 in 2B4.ζ CAR and UTD NK cells improved their effector function in the setting of chronic antigen simulation in vitro. Multiparameter flow analysis after chronic antigen exposure identified the expansion of unique NK-cell subsets. 2B4.ζ/sIL-15 CAR and sIL-15 NK cells maintained an overall activated NK-cell phenotype. This was confirmed by transcriptomic analysis, which revealed a highly proliferative and activated signature in these NK-cell groups. In vivo, 2B4.ζ/sIL-15 CAR-NK cells had potent anti-AML activity in one model, while 2B4.ζ/sIL-15 CAR and sIL-15 NK cells induced lethal toxicity in a second model.
Conclusion:
Transgenic expression of CD123-CARs and sIL-15 enabled NK cells to function in the setting of chronic antigen exposure but was associated with systemic toxicities. Thus, our study provides the impetus to explore inducible and controllable expression systems to provide cytokine signals to AML-specific CAR-NK cells before embarking on early-phase clinical testing.
Insights
Engineering Natural Killer (NK) cells with chimeric antigen receptors (CARs) and interleukin-15 (IL-15) showed promise against acute myelogenous leukemia (AML). However, this approach led to toxicities, suggesting a need for controllable expression systems for future clinical trials.
Area of Science:
- Immunotherapy
- Hematologic Malignancies
- Cellular Engineering
Background:
- Recurrent/refractory acute myelogenous leukemia (AML) has a poor prognosis, necessitating novel therapeutic strategies.
- Natural Killer (NK) cells offer potential for antileukemic responses, but adoptive transfer has shown limited efficacy.
- Engineering NK cells with chimeric antigen receptors (CARs) and interleukin-15 (IL-15) aims to enhance anti-AML activity and persistence.
Purpose of the Study:
- To engineer NK cells expressing CD123-specific CARs and/or IL-15 to improve anti-AML efficacy.
- To evaluate the in vitro and in vivo performance of these engineered NK cells in AML models.
Main Methods:
- Characterization of NK cells engineered with CD123-specific CARs (2B4.ζ or 4-1BB.ζ signaling domains) and/or IL-15.
- In vitro assessment of anti-AML activity and effector function under chronic antigen stimulation.
- In vivo evaluation in AML xenograft models.
- Multiparameter flow cytometry and transcriptomic analysis to assess NK cell phenotype and function.
Main Results:
- CARs with 2B4.ζ signaling domains enhanced NK cell anti-AML activity in vitro.
- Transgenic IL-15 improved NK cell effector function and persistence, leading to potent anti-AML activity in one model.
- However, combined CAR and IL-15 expression in NK cells induced lethal toxicities in a second model, linked to systemic effects.
Conclusions:
- Engineered NK cells with CD123-CARs and IL-15 can overcome chronic antigen exposure challenges but pose systemic toxicity risks.
- Controllable and inducible expression systems are crucial for safe clinical translation of CAR-NK cell therapies for AML.
