Session: 703. Cellular Immunotherapies other than CAR-T Cells: Basic and Translational: Poster I
Hematology Disease Topics & Pathways:
Research, Fundamental Science, Translational Research
Methods: In the current study, a novel NKG2D based CAR was inserted into a target gene locus by the non-viral site-specific integration technology. Specific killing activities of Super-γδ T cells to AML cell lines (HL60, THP1) and several tumor cell lines (including NSCLCs, HCCs, RCCs and OCs) were measured by a luciferase-based method. To investigate the killing activity for CLL1 negative tumor cells, a CLL1 knocked out tumor cell line—THP1-CLL1-KO was generated, and the specific killing of unmodified γδ T, CLL1 CAR-αβ T, and Super-γδ T to THP1- CLL1- KO were studied in vitro. To evaluate the toxicity of Super-γδ T cells to PBMCs, unmodified γδ T, CLL1 CAR-αβ T, and Super-γδ T were co-cultured with human PBMC, respectively, and PBMC composition changes were measured by a flow-based assay.
Results: To minimize off-target editing, we used site-specific integration technology PrecisionGENE for the manufacturing of Super-γδ T cells and have achieved over 70% integration efficiency. Cytotoxicity of the newly developed Super -γδ T cells to AML cell lines was tested in vitro. Notably, Super-γδ T cells showed almost 100% killing to AML cell lines (HL60 cells) in vitro at effector-to-target ratio of 3:1, which is comparable to CLL1 CAR-γδ T, but over 5-fold more potent than unmodified γδ T with 20%-30% killing. In addition, there was significantly increased cytokine secretion from Super -γδ T cells when co-cultured with AML tumor cells compared to γδ T. To evaluate whether Super -γδ T cells could overcome cancer cell heterogenicity, a CLL1 negative target cell THP1-CLL1-KO cell line was used for comparison. Importantly, Super-γδ T cells showed almost 80% killing efficiency against the THP1-CLL1-KO at a E:T ratio of 1:3, while CLL1 CAR- αβ T’s killing was minimal. Moreover, normal PBMCs and fibroblast cells were used to study for the off-target cytotoxicity of Super-γδ T cells. It is worth mentioning that while there was no obvious off-target toxicity by Super-γδ T for monocyte when co-culturing with PBMC, CLL1 CAR-αβ T showed killing toward monocytes. To further investigate the killing potential toward different tumor cell lines, cytotoxicity of Super-γδ T cells to various blood and solid tumor cell lines was studied in vitro. Super-γδ T showed enhanced cytotoxicity to a broad-spectrum of tumor cell lines including NSCLC, HCC, RCC and OC cell lines, independent of their mutation background. In vivo efficacy for Super-γδ T is under-going and the result will be presented.
Conclusions:
Taken together, Super-γδT showed much enhanced anti-tumor functionality and superior safety profile in vitro. Super-γδT demonstrated anti-tumor activity against both blood and solid tumor cell lines independent of their mutation background. Therefore, Super-γδ T has the potential to be used as a novel cellular therapeutic agent for treating R/R AML and multiple solid tumors, and it could be efficacious toward cancers of various types, stages and forms.
Disclosures: No relevant conflicts of interest to declare.