Abstract
BackgroundImatinib mesylate is an effective treatment for metastatic gastrointestinal stromal tumor (GIST). However, most patients eventually develop resistance and there are few other treatment options. Immunotherapy using genetically modified or designer T cells (dTc) has gained increased attention for several malignancies in recent years. The aims of this study were to develop and test novel anti-KIT dTc engineered to target GIST cells.MethodsHuman anti-KIT dTc were created by retroviral transduction with novel chimeric immune receptors (CIR). The gene for stem cell factor (SCF), the natural ligand for KIT, was cloned into 1st generation (SCF-CD3ζ, 1st gen) and 2nd generation (SCF-CD28-CD3ζ, 2nd gen) CIR constructs. In vitro dTc proliferation and tumoricidal capacity in the presence of KIT+ tumor cells were measured. In vivo assessment of dTc anti-tumor efficacy was performed by treating immunodeficient mice harboring subcutaneous GIST xenografts with dTc tail vein infusions.ResultsWe successfully produced the 1st and 2nd gen anti-KIT CIR and transduced murine and human T cells. Average transduction efficiencies for human 1st and 2nd gen dTc were 50% and 42%. When co-cultured with KIT+ tumor cells, both 1st and 2nd gen dTc proliferated and produced IFNγ. Human anti-KIT dTc were efficient at lysing GIST in vitro compared to untransduced T cells. In mice with established GIST xenografts, treatment with either 1st or 2nd gen human anti-KIT dTc led to significant reductions in tumor growth rates.ConclusionsWe have constructed a novel anti-KIT CIR for production of dTc that possess specific activity against KIT+ GIST in vitro and in vivo. Further studies are warranted to evaluate the therapeutic potential and safety of anti-KIT dTc.
Highlights
Imatinib mesylate is an effective treatment for metastatic gastrointestinal stromal tumor (GIST)
peripheral blood mononuclear cells (PBMC) were seeded at a density of 2 × 106 cells/ml, and activated on anti-CD3 coated (OKT3, eBioscience, San Diego, CA) 750 ml flasks with 2 ug/mL anti-CD28 (CD28.2, eBioscience) and 300 U/mL of human IL-2 in AIM V medium (Invitrogen, Grand Island, NY) supplemented with 5% heat inactivated sterile human serum (Valley Biomedical, Winchester, VA). 293T-HEK phoenix amphotropic cells (Orbigen, Allele Biotechnology, San Diego, CA) were transfected with 50 μg 1st or 2nd gen c-KIT ligand chimeric immune receptor (CIR) retroviral plasmid using LipoD283 (SignaGen Laboratories, Rockville, MD)
The KL component is expressed on the extracellular aspect of the CIR to enable interaction with KIT on the surface of target tumor cells (Figure 1B)
Summary
Imatinib mesylate is an effective treatment for metastatic gastrointestinal stromal tumor (GIST). Immunotherapy using genetically modified or designer T cells (dTc) has gained increased attention for several malignancies in recent years. The aims of this study were to develop and test novel anti-KIT dTc engineered to target GIST cells. The majority of patients with metastatic GIST who are treated with imatinib develop resistance and subsequently progressive disease [4]. Cell-based immunotherapy using tumor infiltrating lymphocytes (TIL) has shown success for specific diseases [5]. The genetically modified or designer T cell (dTc) strategy allows for the production of tumor-specific lymphocytes for any patient with a suitable target tumor antigen. Expression of CIR on the surface of modified T cells allows for highly specific recognition of tumor cells expressing the cognate antigenic moiety
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