Simultaneous Engagement of Tumor and Stroma Targeting Antibodies by Engineered NK-92 Cells Expressing CD64 Controls Prostate Cancer Growth

Abstract

Metastatic castration-resistant prostate cancer (mCRPC) has been largely resistant to immunotherapy. Natural killer (NK) cells are cytotoxic lymphocytes that detect and kill transformed cells without prior sensitization, and their infiltration into prostate tumors corresponds with an increased overall survival among patients with mCRPC. We sought to harness this knowledge to develop an approach to NK-cell based immunotherapy for mCRPC. We engineered an NK cell line (NK-92MI) to express CD64, the sole human high-affinity IgG Fcγ receptor (FcγR1), and bound these cells with antibodies to provide interchangeable tumor-targeting elements. NK-92MICD64 cells were evaluated for cell-activation mechanisms and antibody-dependent cell-mediated cytotoxicity (ADCC). A combination of mAbs was used to target the prostate tumor antigen tumor-associated calcium signal transducer 2 (TROP2) and the cancer-associated fibroblast marker fibroblast activation protein alpha (FAP). We found that CD64, which is normally expressed by myeloid cells and associates with the adaptor molecule FcRγ, can be expressed by NK-92MI cells and mediate ADCC through an association with CD3ζ. Cytotoxicity from the combination approach was two-fold higher compared to treatment with NK-92MICD64 cells and either mAb alone, and seven-fold higher than NK-92MICD64 cells alone at an effector–target cell ratio of 20:1. The cytotoxic effect was lost when using isotype control antibodies, indicating a selective targeting mechanism. The combination approach demonstrated efficacy in vivo as well and significantly reduced tumor growth compared with the saline control. This combination therapy presents a potential approach for treating mCRPC and could improve immunotherapy response.

A Hot Topic: Cancer Immunotherapy and Natural Killer Cells

“…Knowing that NK-92MI cells are CD16-, they observed a high level of ADCC against both prostate cell lines in the presence of appropriated antibodies. The data was confirmed in an in vivo xenograft mouse model [ 82 ], so that with this approach, immunotherapy of prostate cancer and other solid tumors could become possible. The results also emphasize the benefits of including the TME into the therapeutic strategy.…”

“…The NK-92 cell line was transduced with the CAR construct and efficiently killed estrogen-resistant MCF-7 breast cancer cell line variants in vitro [ 81 ]. Hintz et al used the NK-92MI derivative to test its efficiency against a prostate cancer cell line and in parallel a prostate stromal cell line (to mimic in vitro the targeting of the tumor and the TME in parallel) [ 82 ]. To do so, they transduced the NK cell line with full-length CD64, a high affinity Fc γ receptor physiologically expressed only by myeloid cells.…”

Expression of Fibroblast Activation Protein Is Enriched in Neuroendocrine Prostate Cancer and Predicts Worse Survival

“…Interestingly, when 68 Ga-FAPI was compared head-to-head with 18 F-FDG PET/CT in a multicenter study of 71 patients with various primaries, the former had similar quantitative tumor uptake, but lower background uptake, yielding improved diagnostic information particularly in tumor areas with high physiological 18 F-FDG uptake [ 22 ]. From a therapeutic perspective, targeting of stromal FAP with monoclocal antibodies can be effective, particularly when combined with tumor targeting approaches against the prostate tumor antigen tumor-associated calcium signal transducer 2 (TROP2) using engineered natural killer NK-92 cells expressing CD64 [ 23 ].…”

Examination of IgG Fc Receptor CD16A and CD64 Expression by Canine Leukocytes and Their ADCC Activity in Engineered NK Cells

“…FreeStyle 293-F cells were obtained from Thermo Fisher Scientific (Waltham, MA). All cells were cultured per the manufacturer’s directions, as we have previously described ( 17 , 18 ). For cell activation, leukocytes were stimulated with phorbol-12-myristate-13-acetate (PMA) (MilliporeSigma, Burlington, MA), as previously described ( 33 ).…”

“…CD64, however, is expressed by myeloid leukocyte populations but not lymphocytes, including NK cells ( 10 ). Therefore, we have engineered human NK cells to express recombinant versions of CD64 to increase their attachment efficiency to antibody-coated tumor cells to kill these cells ( 17 , 18 ). Moreover, due to its high affinity state, NK cells expressing recombinant CD64 can be “armed” with anti-tumor mAbs, which can be switched for universal tumor antigen targeting ( 14 , 17 , 18 ).…”