Abstract 2188: Generation and characterization of human placental-derived CD19 CAR-T cells using viral vectors
Authors: Karasiewicz Kathy , He Shuyang , Tess Kristina , Ling Weifang , Jhun Kevin , Kaufmann Gunnar , Zeldis Jerome B. , Ji Henry , Hariri Robert , Zhang Xiaokui 
Published: September 2020

Introduction: Celularity, Inc. is developing a CD19 CAR-T cell therapy using an allogeneic platform derived from postpartum human placental cells. To augment the CD19 CAR expression and activity of Placental-derived T (P-T) cells, we have evaluated the use of various lentiviral (LV) vectors, as well as a retroviral (RV) CD19 CAR construct and assessed the effects of each virus on the phenotypic characteristics and activity of our cells. Fully humanized scFv CD19 constructs have been shown to be less immunogenic with improved persistence and efficacy in patients (Sommermeyer, et. al. Leukemia, 2017), so human (Hu) vs. mouse (Ms) scFv CD19 CAR constructs were also evaluated.

Experimental Procedures: Isolated P-T cells were activated and transduced with various Ms and Hu scFv CD19 CAR viral vectors, including two Ms scFv LV sequences (containing 4-1BB costimulatory domains), three different Hu scFv LV sequences (two containing 4-1BB and one containing CD28 costimulatory domains), and one Ms scFv RV sequence (containing 4-1BB costimulatory domain). In vitro functional activity of these P-T CD19 CAR cells was assessed against CD19+ Burkitt's Lymphoma (Daudi) and Acute lymphoblastic Leukemia (NALM6) cell lines in cytotoxicity assays. In vivo anti-tumor activity of P-T Ms scFv CD19 CAR LV vs. RV cells was assessed using a disseminated lymphoma xenograft model in NSG mice.

Results: All CD19 CAR constructs containing the 4-1BB costimulatory domain resulted in high transduction efficiency in P-T cells: Ms LV: 32-70% (n=4); Hu LV: 26-42% (n=2); Ms RV: 22-52% (n=6). P-T cells transduced with LV resulted in lower frequency of CD8+ T cells (9-42%), as compared to cells transduced with RV (46-59%), and consequently had lower CD8+ CD19 CAR+ (of CD3+) expression (LV: 2.8-6.4%; RV: 14-30%). In vitro, all CD19 CAR constructs specifically lysed CD19+ Daudi/ Nalm6 targets in cytotoxicity assays. In vivo, both P-T Ms scFv CD19 CAR LV and RV cells significantly reduced tumor burden and improved survival over the vehicle control. P-T CD19 CAR RV cells demonstrated superior anti-tumor activity over P-T CD19 CAR LV with 100% survival out to day 120, while the CD19 CAR LV treated group succumbed to 100% mortality by day 70.

Summary: We have demonstrated the ability to generate P-T CD19 CAR cells using a variety of viral vectors containing the 4-1BB costimulatory domain. Transduction of P-T cells using Hu vs. Ms scFv CD19 CAR sequences resulted in comparable CD19 CAR expression and in vitro cytotoxic activity, whereas use of LV vs. RV had an impact on the phenotypic characteristic of the cells and resulted in differences in the in vivo tumor activity of the cells.

Citation Format: Kathy Karasiewicz, Shuyang He, Kristina Tess, Weifang Ling, Kevin Jhun, Gunnar Kaufmann, Jerome B. Zeldis, Henry Ji, Robert Hariri, Xiaokui Zhang. Generation and characterization of human placental-derived CD19 CAR-T cells using viral vectors [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2188.