Abstract A021: Enhancing NK cell immunotherapy in non-small cell lung cancer using dual-antigen targeting Tri-Specific Killer Engagers (TriKEs/TriKE PACCs)
Lung cancer is the leading cause of cancer-related death worldwide, with ∼230,000 new U.S. cases projected in 2026. Non-small cell lung cancer (NSCLC), the most common subtype, is treated with surgery, chemotherapy, radiation, targeted therapy, and immunotherapy. While checkpoint blockade therapies have improved outcomes, only ∼15% of advanced NSCLC patients achieve long-term survival, leaving most patients with limited options after relapse. This underscores the need for more effective treatments. Natural Killer (NK) cells play a key role in tumor surveillance, and higher NK cell levels correlate with better patient outcomes. To harness them, we developed Tri-specific Killer Engager (TriKE) molecules to enhance NK cell function through CD16 engagement, IL-15 signaling, and tumor antigen targeting. However, current TriKEs face limitations including rapid clearance (through the glomerulus) and restriction to single-antigen targeting. To overcome these challenges, we engineered TriKE-PACCs (Poly-Antigen Cytokine Complexes), a next-generation platform designed to extend NK cell persistence and enable dual-antigen targeting. TriKE-PACCs combine an anti-CD16 single domain antibody (sdAb), an IL-15 moiety, and an anti-mesothelin single chain variable fragment (scFv), along with a PACC module consisting of IL-15 receptor alpha linked to an anti-B7H3 sdAb. This bifunctional design enables dual targeting of mesothelin and B7H3, addressing both tumor heterogeneity and antigen escape, as seen in NSCLC. Using H322 NSCLC cells with B7-H3 and/or mesothelin CRISPR knockouts to model tumor heterogeneity and antigen loss, we evaluated NK cell degranulation (CD107a), cytokine production (IFNγ, TNFα), and cytotoxicity (Xcelligence and Incucyte live imaging assays). Treatment with TriKE-PACCs significantly enhanced NK cell degranulation, cytokine production, and tumor killing compared to single-antigen TriKEs and IL-15 alone, particularly in heterogeneous antigen settings. Next steps include validating our findings in NSCLC 3D organoid and spheroid models, as well as confirming the efficacy of TriKE-PACCs in vivo. To summarize, our findings show that TriKE-PACCs are a promising immunotherapy approach for NSCLC, with potential to overcome antigen escape, improve NK cell persistence, and enhance clinical outcomes for lung cancer patients. Rachel N. Steinmetz, Shee Kwan Phung, Joshua T. Walker, Tumpa Dasgupta, Yvette Soignier, Philippa R. Kennedy, Martin Felices. Enhancing NK cell immunotherapy in non-small cell lung cancer using dual-antigen targeting Tri-Specific Killer Engagers (TriKEs/TriKE PACCs) [abstract]. In: Proceedings of AACR Drug Discovery and Development (AACR D3) Conference; 2026 Jul 21-24; Boston, MA. Philadelphia (PA): AACR; Clin Cancer Res 2026;32(14_Suppl):Abstract nr A021.