Nature. 2022 Sep;609(7926):369-374. doi: 10.1038/s41586-022-05140-y. | PubMed

Jiqin Zhang¹, Yongxian Hu^2,3,4,5, Jiaxuan Yang⁶, Wei Li⁷, Mingming Zhang^2,3,4,5, Qingcan Wang⁷, Linjie Zhang⁶, Guoqing Wei^2,3,4,5, Yue Tian⁶, Kui Zhao⁸, Ang Chen^6,7, Binghe Tan^6,7, Jiazhen Cui^2,3,4,5, Deqi Li⁷, Yi Li^2,3,4,5, Yalei Qi⁶, Dongrui Wang^2,3,4,5, Yuxuan Wu^6,7, Dali Li^9,10, Bing Du^11,12, Mingyao Liu^13,14, He Huang^15,16,17,18

1. Shanghai Frontiers Science Center of Genome Editing and Cell Therapy, Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China. zjqjeremy@163.com.
2. Bone Marrow Transplantation Center, the First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China.
3. Liangzhu Laboratory, Zhejiang University Medical Center, Hangzhou, China.
4. Institute of Hematology, Zhejiang University, Hangzhou, China.
5. Zhejiang Province Engineering Laboratory for Stem Cell and Immunity Therapy, Hangzhou, China.
6. Shanghai Frontiers Science Center of Genome Editing and Cell Therapy, Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China.
7. BRL Medicine, Inc., Shanghai, China.
8. PETCT Center, The First Affiliated Hospital, School of Medicine, Zhejiang University, Hangzhou, China.
9. Shanghai Frontiers Science Center of Genome Editing and Cell Therapy, Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China. dlli@bio.ecnu.edu.cn.
10. BRL Medicine, Inc., Shanghai, China. dlli@bio.ecnu.edu.cn.
11. Shanghai Frontiers Science Center of Genome Editing and Cell Therapy, Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China. bdu@bio.ecnu.edu.cn.
12. BRL Medicine, Inc., Shanghai, China. bdu@bio.ecnu.edu.cn.
13. Shanghai Frontiers Science Center of Genome Editing and Cell Therapy, Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China. myliu@bio.ecnu.edu.cn.
14. BRL Medicine, Inc., Shanghai, China. myliu@bio.ecnu.edu.cn.
15. Bone Marrow Transplantation Center, the First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China. huanghe@zju.edu.cn.
16. Liangzhu Laboratory, Zhejiang University Medical Center, Hangzhou, China. huanghe@zju.edu.cn.
17. Institute of Hematology, Zhejiang University, Hangzhou, China. huanghe@zju.edu.cn.
18. Zhejiang Province Engineering Laboratory for Stem Cell and Immunity Therapy, Hangzhou, China. huanghe@zju.edu.cn.

Abstract

Recently, chimeric antigen receptor (CAR)-T cell therapy has shown great promise in treating haematological malignancies1-7. However, CAR-T cell therapy currently has several limitations8-12. Here we successfully developed a two-in-one approach to generate non-viral, gene-specific targeted CAR-T cells through CRISPR-Cas9. Using the optimized protocol, we demonstrated feasibility in a preclinical study by inserting an anti-CD19 CAR cassette into the AAVS1 safe-harbour locus. Furthermore, an innovative type of anti-CD19 CAR-T cell with PD1 integration was developed and showed superior ability to eradicate tumour cells in xenograft models. In adoptive therapy for relapsed/refractory aggressive B cell non-Hodgkin lymphoma (ClinicalTrials.gov, NCT04213469 ), we observed a high rate (87.5%) of complete remission and durable responses without serious adverse events in eight patients. Notably, these enhanced CAR-T cells were effective even at a low infusion dose and with a low percentage of CAR+ cells. Single-cell analysis showed that the electroporation method resulted in a high percentage of memory T cells in infusion products, and PD1 interference enhanced anti-tumour immune functions, further validating the advantages of non-viral, PD1-integrated CAR-T cells. Collectively, our results demonstrate the high safety and efficacy of non-viral, gene-specific integrated CAR-T cells, thus providing an innovative technology for CAR-T cell therapy.

Presented By Jiqin Zhang