CAR-T cells dual-target CD123 and NKG2DLs to eradicate AML cells and selectively target immunosuppressive cells

ABSTRACT

Chimeric antigen receptor (CAR)-T cells have not made significant progress in the treatment of acute myeloid leukemia (AML) in earlyclinical studies. This lack of progress could be attributed in part to the immunosuppressive microenvironment of AML, such as monocyte-like myeloid-derived suppressor cells (M-MDSCs) and alternatively activated macrophages (M2 cells), which can inhibit the antitumor activity of CAR-T cells. Furthermore, AML cells are usually heterogeneous, and single-target CAR-T cells may not be able to eliminate all AML cells, leading to disease relapse. CD123 and NKG2D ligands (NKG2DLs) are commonly used targets for CAR-T therapy of AML, and M-MDSCs and M2 cells express both antigens. We developed dual-targeted CAR-T (123NL CAR-T) cells targeting CD123 and NKG2DL by various structural optimization screens. Our study reveals that 123NL CAR-T cells eradicate AML cells and selectively target immunosuppressive cells. A highly compact marker/suicide gene, RQR8, which binds targeting epitopes of CD34 and CD20 antigens, was also incorporated in front of the CAR structure. The binding of Rituximab to RQR8 leads to the elimination of 123NL CAR-T cells and cessation of their cytotoxicity. In conclusion, we successfully developed dual effects of 123NL CAR-T cells against tumor cells and immunosuppressive cells, which can avoid target escape and resist the effects of immunosuppressive microenvironment.

KEYWORDS:

• AML
• CAR-T
• CD123
• immunotherapy
• NKG2DL

Acknowledgments

We thank Zongjin Li (Nankai University School of Medicine) and Shang Chen (Nankai University School of Medicine) for their guidance and assistance in animal experiments.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Authors’ contributions

XJ and MFZ designed the research. XJ, DNX, RS, XX, MZ, JXW, YBY, XYH, and JXM performed the research. XJ, WYL, and MFZ analyzed the data. XJ wrote the manuscript. XJ, DNX, RS and MFZ revised the manuscript.

Availability of data and materials

All data obtained and/or analyzed during the current study are available from the corresponding authors upon reasonable request.

Abbreviations

CAR-T	=	Chimeric antigen receptor-engineered T cells
AML	=	Acute myeloid leukemia
M-MDSCs	=	Monocyte-like myeloid-derived suppressor cells
M2 cells	=	Alternately activated macrophages
NKG2DLs	=	NKG2D ligands
123NL CAR-T	=	Dual-target CAR-T cells targeting CD123 and NKG2DLs
B-ALL	=	B cell acute lymphoblastic leukemia
LFA-1	=	Lymphocyte function-associated antigen 1
BMMCs	=	Bone marrow mononuclear cells

Ethics approval and consent to participate

This study was approved by the Ethics Committee of Tianjin First Central Hospital. All patients signed an informed consent form. The animal experiment protocol was approved by the Animal Care and Use Committee of Tianjin Medical University.

Supplementary material

Supplemental data for this article can be accessed online at https://doi.org/10.1080/2162402X.2023.2248826

Additional information

Funding

This work was supported by grants from the General Project of the National Natural Science Foundation of China (81970180 to MZ), the Science and Technology Project of Tianjin Municipal Health Committee (TJWJ2022QN030 to MZ), Key projects of Tianjin Applied Basic Research and Multi-Investment Fund (21JCZDJC01240), Science and Technology Project of Tianjin Municipal Health Committee (TJWJ2022XK018 to MZ), the Key Science and Technology Support Project of Tianjin Science and Technology Bureau (20YFZCSY00800 to MZ), Tianjin Health Bureau Project (ZC20074 to XX), and Tianjin Key Medical Discipline (Specialty) Construction Project (TJYXZDXK-056B), as well as China Postdoctoral Science Foundation (2022M722748).