Combining Dinutuximab with ALT-803 (IL-15 Superagonist) Significantly Improves the Activity of Ex Vivo Expanded Natural Killer Cells Against GD2 + Pediatric Solid Tumors (ST)

Background: Despite significant survival improvements in multiple pediatric malignancies, children with recurrent and/or metastatic osteosarcoma (OS) (Allison/Menendez, Sarcoma, 2012), relapsed neuroblastoma (NB) (London/Matthay, Journal of Clinical Oncology, 2011) and glioblastoma multiforme (GBM) (Knisely/Baerhing, The Lancet, 2009) continue to have dismal outcomes. GD2 is highly expressed in NB, OS and GBM (Woo/Joo, Anat Cell Biol, 2015). Dinutuximab is an anti-GD2 monoclonal antibody that has significantly increased EFS in children with GD2+ neuroblastoma (Yu/Sondel, NEJM, 2010). ALT-803 is a novel IL15 superagonist complex where mutant IL-15N72D is bound to an IL-15RαSu-Fc fusion protein (Xu/Wong, Cancer Res, 2013). ALT-803 has superior immunostimulatory activity, more favorable pharmacokinetics and pharmacodynamics compared to IL-15 (Rhode/Wong 2016), and is currently being tested in several clinical trials. Objective: We hypothesized that the combination of Dinutuximab with ALT-803 will enhance the in vitro cytotoxicity of exPBNK cells against GD2+ OS, NB and GBM. Method: PBMCs were isolated, and expanded them ex vivo with lethally irradiated K562-mbIL21cells (Denman/Lee Plos One, 2012). ExPBNK cells were isolated using Miltenyi NK cell isolation kits as previously described (Chu/Cairo, Can Imm Res, 2015). NK proliferation, NK receptors expression and cytotoxicity were assessed as previously described (Chu/Cairo, Can Imm Res, 2015). ALT-803 was generously provided by Altor BioScience. Dinutuximab (generously provided by United Therapeutics) was used for antibody-dependent cellular cytotoxicity (ADCC) assays. IFN-g and perforin levels were evaluated by ELISA assays. GD2+ OS (U2OS), NB (SKNF2) and GBM (M059K) cell lines were used as target cells. Results: ALT-803 significantly promoted in vitro proliferation of exPBNK by increasing the phosphorylation of Akt, Stat3/5 and p38 MAPK. ALT-803 also increased the expression of NK activating receptors: NKG2D, NKp30, NKp44, and NKp46. ALT-803 significantly enhanced exPBNK in vivo persistence (P < .05) and significantly reduced tumor burden in OS xenografted mice when combined with exPBNK cells (P < .05). Furthermore, ALT-803 significantly enhanced exPBNK mediated ADCC with dinutuximab against OS, NB and GBM cells in a E:T dependent manner (P < .001) (Figure 1). ALT-803 significantly enhanced IFN-g (P < .001) and perforin (P < .001) release from exPBNK when it was combined with dinutuximab against OS, NB and GBM cells compared to exPBNK, ALT-803 + exPBNK, or dinutuximab + exPBNK. Conclusions: These results show that combining Dinutuximab with ALT-803 significantly enhanced exPBNK ADCC, and IFN-g and perforin release against GD2+ OS, NB and GBM cells. Future studies using NOD/SCID human solid tumor xenograft mouse model are being planned to investigate the in vivo efficacy of this combination.