Combinatorial cellular therapy in pediatric solid tumors with natural killer (NK) and genetically engineered myeloid cells (GEMys).

Abstract

2561 Background: Early human studies with Natural Killer (NK) cells have shown promise, harnessing their ability to infiltrate into tumors and be tumoricidal. However, these effects can be limited by adenosine, reactive oxygen species, and TGF-β in the highly immune suppressive tumor microenvironment (TME). TGF-β and downstream SMAD3 signaling in Natural Killer (NK) cells have been shown to cause decreased cytokine production, downregulation of activating cell surface receptors, and attenuated cytotoxic function against solid tumors. Exposure to TGF-β during cell culture generates expanded NK cells with increased resistance to TGF-β signaling, dubbed TGF-β imprinted NK cells. In vitrostudies have shown that these cells have increased cytokine production and cytotoxic function in the presence of TGF-β. However, their in vivo trafficking, modulation of the TME, and therapeutic efficacy remain understudied. Our lab has previously shown that when given to tumor-bearing mice, myeloid cells modified to produce IL-12 (IL-12-Genetically Engineered Myeloid cells, aka IL-12 GEMys) are effective at homing to tumor and metastatic sites and, via IL-12 secretion and TME modulation, increase NK cell presence and activation in these sites. They hold great promise as combination therapy, enhancing the effectiveness of other cellular therapies such as NK cell therapy. A current clinical trial exploring TGF-β Imprinted, Ex Vivo Expanded, Universal Donor NK Cell Infusions in relapsed sarcomas is underway (NCT05634369). Methods: In vitro experiments were conducted with the xCELLigence real-time cell analysis system. The human osteosarcoma cell line 143B and human rhabdomyosarcoma cell line RH-30 were evaluated for cell death after co-culture with healthy donor NK cells or TGF-β imprinted NK cells (E:T ratio of 5:1) in isolation and in combination with IL-12 GEMys (E:T ratio of 2:1). Results: TGF-β imprinted NK cells are significantly more effective at tumor cell killing of both 143B and RH30 cell lines in-vitro compared to regular expanded NK cells (WT NK cells), with >90% decrease in cell index compared to 50-60%, respectively, after 48 hours of co-culture. IL-12 GEMys also significantly enhance the cytotoxic effects of WT NK cells, increasing cell killing to 70-80% compared to co-culture with untransduced GEMy controls. Conclusions: These initial in-vitro experiments provide insight into the promising effectiveness of NK-myeloid cell combination therapies in the treatment of pediatric sarcomas. Ongoing in vitroexperiments include cytokine quantification by ELISA and flow cytometric analysis of NK cell activation and proliferation markers. Our preliminary results provide rationale to continue studying the in vivo efficacy of TGF-β imprinted NK cell monotherapy and in combination with IL-12 GEMys in the treatment of tumor-bearing NSG mice.