TROP2-Directed CAR-NK Cells for the Immunotherapy of Pancreatic Cancer

SUMMARY/ ABSTRACT Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer with a 5-year survival rate of only ~12%. Therefore, there is a critical unmet need for new treatment options. Chimeric antigen receptor (CAR)-T cells have led to a paradigm shift in the treatment of some hematologic cancers, but efficacy in solid tumors remains limited, partly due to the lack of highly specific targets and the immunosuppressive tumor microenvironment (TME). Moreover, the time required and the high cost of manufacturing an autologous cell product, and the toxicity of CAR-T cells call for novel products that are universal, safe, and potent. CAR-NK cells have emerged as a promising cell therapy for cancer due to the NK cells’ innate ability to kill tumor cells and their safety in the allogeneic setting. In a first-in-human study, our group showed the safety and efficacy of cord blood (CB)-derived CAR-NK cells targeting CD19 in B-lymphoid malignancies. This proposal aims to build on this platform to develop the next-generation NK cell therapies for PDAC by enhancing NK cell potency and persistence through optimal co-stimulatory signaling, cytokine armoring and checkpoint inhibition. We have identified TROP2 as a promising therapeutic target in PDAC and developed a novel strategy to target TROP2 by genetically modifying CB-NK cells with a retroviral vector encoding: (i) the humanized RS7 scFv targeting TROP2; (ii) DAP10 as an NK- specific co-stimulatory domain; (iii) IL-15 to support their survival and proliferation; and (iv) inducible caspase-9 (iC9) as a safety switch (iC9/TROP2CAR/IL-15). Our data show the efficacy and safety of this approach in vitro and in vivo and support its translation to the clinic. In addition, we have developed a robust strategy for the cryopreservation of CAR-NK cells, enabling the generation of a biobank of off-the-shelf engineered NK cells that could be thawed and infused at bedside, thus reducing cost and increasing accessibility. We have manufactured and cryopreserved 125 patient doses of GMP-grade iC9/TROP2CAR/IL-15 NK cells and a clinical study to test the safety and efficacy of this off-the-shelf product in PDAC was recently approved by the FDA (Protocol 2022- 0687; IND 29348). Finally, we have devised a novel strategy to target the immune metabolic checkpoint CREM to modulate the metabolic fitness and potency of CAR-NK cells in the PDAC TME. We hypothesize that targeting TROP2 with iC9/TROP2CAR/IL-15 NK cells will greatly improve outcomes in PDAC and that by deleting the metabolic immune checkpoint CREM we can further enhance the fitness and potency of NK cells. We will test our hypothesis in three specific aims: In Aim 1 we will conduct a Phase I/II clinical trial to test the safety and efficacy of intraperitoneally delivered iC9/TROP2CAR/IL-15 NK cells in patients with TROP2+ PDAC. In Aim 2 we will apply innovative single-cell transcriptomics and spatial proteomics to comprehensively characterize the fate of the adoptively transferred CAR-NK cells and their interaction with other cells within the TME, to uncover key mechanisms of efficacy and resistance. In Aim 3 we will investigate the mechanism by which targeting CREM enhances the antitumor activity of iC9/TROP2CAR/IL-15 NK cells in an orthotopic PDX mouse model of PDAC.