Pre-emptive immunotherapy after HSC transplantation based on minimal residual disease (MRD) status with donor lymphocyte infusions (DLI) using cytokine-induced killer (CIK) cells may be beneficial to prevent relapse with a reduced risk of causing graft-versus-host-disease (GvHD) compared to conventional T cell infusion. CIK cells are a heterogeneous effector cell population including T cells (CD3+CD56-), natural killer (NK) cells (CD3-CD56+) and natural killer T (T-NK) cells (CD3+CD56+), that exhibit non-MHC-restricted cytotoxic activity and are generated by ex vivo expansion of peripheral blood mononuclear cells (PBMC) through the addition of interferon (IFN)-γ, anti-CD3 antibody, IL-2 and IL-15.
While CIK cells have shown potent in vivo activity against various cancer types such as lymphomas or colorectal cancer, their cytotoxicity against B-ALL, characterized by the expression of CD19, has been limited. Hence, retargeting of CIK cells using chimeric antigen receptors (CARs) to facilitate selective target cell recognition and enhance specific cytotoxicity represents a promising approach. CAR comprise an extracellular scFv antibody fragment as an antigen-binding domain, linked via a flexible hinge region and a transmembrane domain to an intracellular signaling moiety such as CD3 zeta chain (first generation CAR), or zeta chain fused to a co-stimulatory protein domain such as CD28 (second generation CAR).
We established an optimized protocol for transduction of CIK cells with CD19-specific lentiviral CAR constructs, and characterized cells for expression of an EGFP marker gene and CAR surface expression. Effects of exposure to lentiviral vector particles on the development of CIK cell subpopulations were monitored. In in vitro cytotoxicity assays we could demonstrate potent and selective cytotoxicity of retargeted CIK cells towards established cancer cell lines expressing CD19 and primary pre-B-ALL blasts. In addition, we observed significantly enhanced degranulation of CAR-CIK cells upon target cell contact and showed increased secretion of pro-inflammatory cytokines, while no secretion of immunosuppressive IL-10 could be detected. Cytotoxity towards non-malignant allo-PBMCs remained low.
In addition, we investigated anti-leukemic activity of retargeted CIK cells in vivo in NOD/SCID common gamma chain knockout (NSG) mouse models using bioluminescence and fluorescence imaging. We observed potent reduction of tumor load after only two CIK cell applications in mice with an engrafted primary pre-B-ALL yielding long-term survivors with a negative MRD status (4/6). In a separate model, we demonstrated migration of CD19-specific CIK cells into local, subcutaneous tumor sites.
In conclusion, CAR-CIK cells represent a promising alternative to CAR-T cell therapy, as relatively high numbers of conventional CIK cells have already been infused in clinical trials, which demonstrated a low risk of causing severe side effects.
Citation Format: Sarah Oelsner, Juliane Wagner, Miriam E. Friede, Verena Pfirrmann, Eva Rettinger, Ralf Schubert, Heike Pfeifer, Evelyn Ullrich, Peter Bader, Winfried S. Wels. Genetically modified cytokine-induced killer (CIK) cells for targeted cancer therapy. [abstract]. In: Proceedings of the CRI-CIMT-EATI-AACR Inaugural International Cancer Immunotherapy Conference: Translating Science into Survival; September 16-19, 2015; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(1 Suppl):Abstract nr A164.