Single-cell analysis of the survival mechanisms of fratricidal CAR-T targeting of T cell malignancies

Hui Hu; Ling Tang; Yuyan Zhao; Jiali Cheng; Mei Huang; Yong You; Ping Zou; Qian Lei; Xiaojian Zhu; An-Yuan Guo

doi:10.1016/j.omtn.2024.102225

Single-cell analysis of the survival mechanisms of fratricidal CAR-T targeting of T cell malignancies

Mol Ther Nucleic Acids. 2024 May 20;35(2):102225. doi: 10.1016/j.omtn.2024.102225. eCollection 2024 Jun 11.

Authors

Hui Hu^{1

2}, Ling Tang³, Yuyan Zhao⁴, Jiali Cheng⁵, Mei Huang⁵, Yong You⁶, Ping Zou⁶, Qian Lei¹, Xiaojian Zhu⁵, An-Yuan Guo^{1

4}

Affiliations

¹ Department of Hematology, West China Biomedical Big Data Center, West China Hospital, Med-X Center for Informatics, Sichuan University, Chengdu 610041, China.
² Department of Laboratory Medicine, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430030, China.
³ Department of Hematology, Renmin Hospital of Wuhan University, Wuhan 430060, China.
⁴ Hubei Bioinformatics & Molecular Imaging Key Laboratory, College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan, Hubei 430074, China.
⁵ Department of Hematology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
⁶ Institute of Hematology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China.

Abstract

Chimeric antigen receptor T (CAR-T) cell therapy targeting T cell tumors still faces many challenges, one of which is its fratricide due to the target gene expressed on CAR-T cells. Despite this, these CAR-T cells can be expanded in vitro by extending the culture time and effectively eliminating malignant T cells. However, the mechanisms underlying CAR-T cell survival in cell subpopulations, the molecules involved, and their regulation are still unknown. We performed single-cell transcriptome profiling to investigate the fratricidal CAR-T products (CD26 CAR-Ts and CD44v6 CAR-Ts) targeting T cells, taking CD19 CAR-Ts targeting B cells from the same donor as a control. Compared with CD19 CAR-Ts, fratricidal CAR-T cells exhibit no unique cell subpopulation, but have more exhausted T cells, fewer cytotoxic T cells, and more T cell receptor (TCR) clonal amplification. Furthermore, we observed that fratricidal CAR-T cell survival was accompanied by target gene expression. Gene expression results suggest that fratricidal CAR-T cells may downregulate their human leukocyte antigen (HLA) molecules to evade T cell recognition. Single-cell regulatory network analysis and suppression experiments revealed that exhaustion mediated by critical regulatory factors may contribute to fratricidal CAR-T cell survival. Together, these data provide valuable and first-time insights into the survival of fratricidal CAR-T cells.

Keywords: MT: Bioinformatics; cell subpopulations; chimeric antigen receptor T cell; exhausted T cells; regulatory network; single-cell sequencing.