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DAP12-based activating chimeric antigen receptor for NK cell tumor immunotherapy
Katrin Töpfer1, Marc Cartellieri2, Susanne Michen1
1Section of Experimental Neurosurgery and Tumor Immunology, Department of Neurosurgery, University Hospital Carl Gustav Carus, TU Dresden, 01307 Dresden, Germany;
Abstract:
NK cells are emerging as new effectors for immunotherapy of cancer. In particular, the genetic engraftment of chimeric Ag receptors (CARs) in NK cells is a promising strategy to redirect NK cells to otherwise NK cell-resistant tumor cells. On the basis of DNAX-activation protein 12 (DAP12), a signaling adaptor molecule involved in signal transduction of activating NK cell receptors, we generated a new type of CAR targeting the prostate stem cell Ag (PSCA). We demonstrate in this article that this CAR, designated anti-PSCA-DAP12, consisting of DAP12 fused to the anti-PSCA single-chain Ab fragment scFv(AM1) confers improved cytotoxicity to the NK cell line YTS against PSCA-positive tumor cells when compared with a CAR containing the CD3ζ signaling chain. Further analyses revealed phosphorylation of the DAP12-associated ZAP-70 kinase and IFN-γ release of CAR-engineered cells after contact with PSCA-positive target cells. YTS cells modified with DAP12 alone or with a CAR bearing a phosphorylation-defective ITAM were not activated. Notably, infused YTS cells armed with anti-PSCA-DAP12 caused delayed tumor xenograft growth and resulted in complete tumor eradication in a significant fraction of treated mice. The feasibility of the DAP12-based CAR was further tested in human primary NK cells and confers specific cytotoxicity against KIR/HLA-matched PSCA-positive tumor cells, which was further enhanced by KIR-HLA mismatches. We conclude that NK cells engineered with DAP12-based CARs are a promising tool for adoptive tumor immunotherapy.
Insights
Engineered NK cells with a novel DNAX-activation protein 12 (DAP12)-based chimeric antigen receptor (CAR) targeting prostate stem cell antigen (PSCA) show enhanced cancer cell killing. This DAP12-CAR strategy holds promise for effective adoptive tumor immunotherapy.
Area of Science:
- Immunology
- Cancer Biology
- Cell Therapy
Background:
- Natural Killer (NK) cells are crucial for cancer immunotherapy.
- Chimeric antigen receptors (CARs) genetically engineered into NK cells can target resistant tumors.
- DNAX-activation protein 12 (DAP12) is a key signaling adaptor in NK cell activation.
Purpose of the Study:
- To develop and evaluate a novel DAP12-based CAR targeting prostate stem cell antigen (PSCA).
- To compare the efficacy of the DAP12-CAR with a traditional CD3ζ-based CAR in NK cells.
- To assess the in vivo therapeutic potential of DAP12-CAR engineered NK cells against PSCA-positive tumors.
Main Methods:
- Generation of a DAP12-CAR construct targeting PSCA (anti-PSCA-DAP12).
- Engineering of NK cell line YTS and primary human NK cells with the anti-PSCA-DAP12 CAR.
- In vitro cytotoxicity assays against PSCA-positive tumor cells.
- In vivo studies using tumor xenograft models in mice.
- Analysis of signaling pathways (ZAP-70 phosphorylation) and cytokine release (IFN-γ).
Main Results:
- The anti-PSCA-DAP12 CAR significantly enhanced YTS NK cell cytotoxicity against PSCA-positive tumor cells compared to a CD3ζ-CAR.
- CAR-engineered NK cells showed DAP12-dependent ZAP-70 phosphorylation and IFN-γ release upon target cell contact.
- Infusion of anti-PSCA-DAP12 CAR-engineered YTS cells delayed tumor growth and achieved complete eradication in a fraction of treated mice.
- DAP12-CARs were feasible in primary human NK cells, demonstrating specific cytotoxicity enhanced by KIR-HLA mismatches.
Conclusions:
- DAP12-based CARs represent a novel and effective strategy for engineering NK cells in cancer immunotherapy.
- This approach overcomes NK cell resistance in tumors and shows significant therapeutic potential in preclinical models.
- DAP12-CARs offer a promising alternative signaling module for NK cell-based adoptive immunotherapy.
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