LAUSR

Constitutive EGFR activation plays an important role in the pathophysiology of various solid cancers, such as colorectal cancer, non-small cell lung cancer or squamous cell carcinomas of the head and neck. Tyrosine kinase inhibitors (TKI) and monoclonal antibodies (mAbs), which interfere with signal transduction and activation of EGFR, are approved for treatment of such cancers. However, intrinsic or acquired resistance to these treatments has been described for many patients. Natural killer cells (NK-cells) are important effectors of innate immunity and NK-cell engagers have shown evidence of improved safety in patients compared to T-cell engagers. To specifically utilize the cytotoxic potential of NK-cells to eliminate EGFR-expressing tumors, we developed tetravalent bispecific EGFR/CD16A TandAbs comprising fully human Fv domains recognizing human and cynomolgus EGFR and CD16A. TandAbs recognizing epitopes in the extracellular domain of EGFR differing from epitopes targeted by other mAbs were characterized. Lead candidate AFM24 shows superior cytotoxicity in terms of ADCC (main mode of action) and reduced inhibition of EGFR-mediated phosphorylation compared to cetuximab. Importantly, inhibition of EGFR-signaling is believed to contribute to skin toxicity caused by therapeutic mAbs and TKI’s. AFM24’s cytotoxic activity was tested against EGFR+ tumor cell lines including some carrying a Ras mutation, which is a negative prognostic biomarker and renders cells less susceptible to cetuximab or panitumumab. The cetuximab-resistant CRC cell line HCT-116 or the NSCLC cell line A549 (both with Ras mutations) were efficiently killed with EGFR/CD16A TandAbs in vitro. In vivo data in the HCT-116 model indicate anti-tumor efficacy of AFM24, while no efficacy of cetuximab was seen. Importantly, AFM24 does not activate NK-cells without target cell binding and does not bind to any other members of the EGFR family. While binding and cytotoxic efficacy of many therapeutic mAbs are impaired by serum IgG, no substantial change in AFM24’s binding affinity to NK-cells was observed in the presence of high concentrations of human IgG. In calcein-release cytotoxicity assays with NK-cells as effectors, we showed that the presence of IgG had only little inhibitory effect on AFM24 efficacy compared to cetuximab. In addition, competition of an anti-CD16 mAb with AFM24 in cytotoxicity assays was substantially lower than with cetuximab. Taken together our data demonstrate that AFM24 is a highly potent human antibody displaying favorable biological properties over existing mAbs. This human/cynomolgus cross-reactive agent is currently in preclinical development to treat EGFR+ malignancies and has the potential to exhibit a favorable side effect profile and reduced toxicity and to overcome resistance to other targeted anti-EGFR therapeutic agents. Citation Format: Michael Kluge, Kristina Ellwanger, Uwe Reusch, Ivica Fucek, Michael Weichel, Torsten Haneke, Stefan Knackmus, Joachim Koch, Martin Treder. EGFR/CD16A TandAbs are efficacious NK-cell engagers with favorable biological properties which potently kill EGFR+ tumors with and without Ras mutation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 3641. doi:10.1158/1538-7445.AM2017-3641