A Fusion protein complex that combines il-12, il-15, and il-18 signaling to induce memory-like nk cells for cancer immunotherapy

Natural killer (NK) cells are a promising cellular therapy for cancer, with challenges in the field including persistence, functional activity, and tumor recognition.Briefly, priming blood NK cells with recombinant human (rh)IL-12, rhIL-15, and rhIL-18 (12/15/18) results in memory-like NK cell differentiation and enhanced responses against cancer.However, the lack of available, scalable Good Manufacturing Process (GMP)-grade reagents required to advance this approach beyond early-phase clinical trials is limiting.To address this challenge, we developed a novel platform centered upon an inert tissue factor scaffold for production of heteromeric fusion protein complexes (HFPC).The first use of this platform combined IL-12, IL-15, and IL-18 receptor engagement (HCW9201), and the second adds CD16 engagement (HCW9207).This unique HFPC expression platform was scalable with equivalent protein quality characteristics in small- A nd GMP-scale production HCW9201 and HCW9207 stimulated activation and proliferation signals in NK cells, but HCW9207 had decreased IL-18 receptor signaling.RNA sequencing and multidimensional mass cytometry revealed parallels between HCW9201 and 12/15/18.HCW9201 stimulation improved NK cell metabolic fitness and resulted in the DNA methylation remodeling characteristic of memory-like differentiation.HCW9201 and 12/15/18 primed similar increases in short-term and memory-like NK cell cytotoxicity and IFNg production against leukemia targets, as well as equivalent control of leukemia in NSG mice.Thus, HFPCs represent a protein engineering approach that solves many problems associated with multisignal receptor engagement on immune cells, and HCW9201-primed NK cells can be advanced as an ideal approach for clinical GMP-grade memory-like NK cell production for cancer therapy.