Abstract 2906: Memory-like differentiation, tumor targeting monoclonal antibodies, and chimeric antigen receptors enhance natural killer cell responses to head and neck cancer

Abstract Background: Head and neck cancer (HNC) is an aggressive tumor with low response rates to frontline PD-1 blockade. Natural killer (NK) cells are a promising cellular therapy for T cell-refractory cancers, but are frequently dysfunctional in patients with HNC. New strategies are needed to enhance NK cell responses against HNC. Activation of NK cells with IL-12, 15 and 18 results in memory-like (ML) differentiation improving NK cell responses in hematological malignancies. We hypothesized that ML NK cell differentiation, tumor targeting with cetuximab, and engineering with α-ephA2 chimeric antigen receptor (CAR) would enhance NK cell responses against HNC. Methods: ML NK and conventional (c) NK cells were generated from healthy donors (HD) and evaluated for their ability to produce IFNγ, TNF, degranulate (CD107a) and kill HNC cell lines and primary HNC cells alone or in combination with cetuximab in vitro and in xenograft models. To identify activating receptors involved in NK cell recognition of HNC, blocking experiments were used. ML and cNK cells were engineered to express α-EphA2 CAR-CD8A-41BB-CD3z and functional responses were assessed in vitro against HNC cell lines and primary HNC targets. Results: ML NK cells generated from HD displayed enhanced IFNγ response compared to cNK cells against HNC cell lines SCC1(11±2% vs. 24±1%, mean ± SEM p<0.001), SCC47 (13±2% vs. 36±2%, p<0.01) and primary HNC targets (17±2% vs. 32±3%, p<0.01) which were further enhanced by cetuximab, SCC1 (cNK + cetuximab vs. ML + cetuximab 16±2% vs. 28±3%, p<0.001), SCC47 (39±3% vs. 56±2%, p<0.001), primary HNC targets (23±2% vs. 40±2%, p<0.0001). The combination of ML NK cells and cetuximab was superior to cNK and cetuximab in 51 Cr release assay and IncuCyte® killing assays for HNC cell lines (p<0.01) and primary cells (p<0.01). Findings were consistent in SCC1 HNC xenograft model, where tumor bearing NSG mice treated with cetuximab and ML NK cells had significantly lower tumor burden at 21 days compared to untreated mice, mice that received cNK cells + cetuximab or cetuximab only (p<0.001). Mechanistically, NK cell IFNγ, CD107a and cytotoxicity against HNC targets is partially dependent on activating receptors NKG2D, CD2 and DNAM-1. ML NK cells expressing α-ephA2 CAR were specific to ephA2+ cell lines and demonstrated increased IFNγ (37±4% vs. 16±3%, p<0.001) and CD107a (36±4% vs. 20±1%, p<0.001) against NK resistant ephA2+ UD-SCC2, compared to CAR negative cells. EphA2 CAR ML NK cells had increased CD107a (26±4% vs. 39±4%, p<0.01) and IFNγ (45±5% vs. 66±3%, p<0.001) against primary ephA2+ HNC targets. Conclusion: These pre-clinical findings show that ML differentiation alone or in concert with cetuximab directed targeting or ephA2 CAR engineering, was effective against HNC and provide the rationale for investigation in early phase clinical trials for HNC patients. Citation Format: Miriam T. Jacobs, Pamela Wong, Alice Zhou, Michelle Becker-Hapak, Nancy Marin, Lynne Marsala, Mark Foster, Jennifer Foltz, Celia Cubitt, Jennifer Tran, Carly Neal, David Russler-Germain, Lily Chang, Timothy Schappe, Ethan McClain, Samantha Kersting-Schadek, Carl DeSelm, Melissa Berrien-Elliott, Sidarth V. Puram, Todd A. Fehniger. Memory-like differentiation, tumor targeting monoclonal antibodies, and chimeric antigen receptors enhance natural killer cell responses to head and neck cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2906.