NKTR-255, a Polymer-Conjugated IL-15 Enhances Antibody-Dependent Cellular Cytotoxicity Mediated By NK Cells in a B Cell Lymphoma Model
Authors: Miyazaki Takahiro , Kivimäe Saul , Hennessy Marlene , Pena Rhoneil , Quach Phi , Moffet Andrew , Nieves Wildaliz , Zhang Ping , Marcondes Mario Q. , Madakamutil Loui , Zalevsky Jonathan 
Published: November 2019

Background
IL-15 is a cytokine that activates and provides survival benefit to T and NK cells and has great potential as an immunotherapeutic agent for the treatment of cancer. Exploiting the therapeutic value of native IL-15 has been challenging due to its unfavorable pharmacokinetic properties and poor tolerability. NKTR-255 is a polymer-conjugated IL-15 designed to retain binding affinity to the IL-15 receptor alpha chain and have decreased clearance to provide a sustained pharmacodynamics response. NKTR-255 has an enhanced immunotherapeutic effect when combined with certain targeted monoclonal antibodies that mediate tumor killing by antibody dependent cellular cytotoxicity (ADCC). Here we further characterized the effect of NKTR-255 on NK cell-mediated ADCC in a B cell lymphoma model.
Methods
For in vitro ADCC, NKTR-255 pre-treated human NK cells (effector cells) were co-cultured with RPMI-8226 (target cells) and with daratumumab for 3 hours. The ability of NK cells to lyse target cells was evaluated by detecting 7-AAD stained RPMI-8226. In the Daudi lymphoma model, SCID or SCID beige mice were inoculated IV with 1x107 Daudi cells on Day 0. NKTR-255 (0.3 mg/kg IV) was administered on Days 14, 21 and 28 and a single dose of antibody treatment was given on Day 14 (daratumumab 0.5 mg/kg IP). Survival rate was determined by onset of hind limb paralysis or moribundity as surrogate parameters. Concurrent vs staggered regimen assessment was conducted with NKTR-255 dosed on day 14 and daratumumab on Day 14, 17, or 21. For dose examination, high (0.5 mg/kg) and low (0.05 mg/kg) doses of daratumumab were combined with low (0.03 mg/kg) and high (0.3 mg/kg) doses of NKTR-255 in a concurrent manner. Daudi cells in the bone marrow were assessed 10 days after the NKTR-255 dose by flow cytometry.
Results
In vitro, NKTR-255 pre-treatment of human purified NK cells enhanced daratumumab-mediated ADCC against RPMI-8226. The ability of NKTR-255 to enhance in vitro ADCC was translated into an enhanced therapeutic efficacy of daratumumab in the Daudi lymphoma model with SCID mice. NKTR-255 combined with daratumumab synergistically provided long-term survival benefit in a NKTR-255 dose-dependent manner. To confirm the contribution of NK cells as effector cells for ADCC in vivo, the therapeutic effect of the combination treatment was assessed in the tumor model in SCID beige mice, which have a selective impairment of NK cell cytotoxic function. The long-term survival benefit by the combination treatment, which was observed in SCID mice (median survival of 27 days in control and 59.5 days in treatment), was attenuated in SCID beige mice (median survival of 23 days in control and 30 days in treatment). Concurrent vs staggered dose regimens and dose ranging for NKTR-255 and daratumumab were evaluated in the Daudi model with SCID mice to understand optimal therapeutic use of NKTR-255 in future clinical studies. Tumor depletion in bone marrow was most effective in concurrent treatment of NKTR-255 and daratumumab and three days-staggered treatment (NKTR-255 first and the antibody 3 days later). In contrast, a 7-day stagger showed no benefit of the combination in tumor depletion. Combination of low dose daratumumab and high dose NKTR-255 was equally effective in tumor depletion as a combination of high dose daratumumab and low dose NKTR-255.
Conclusions
NKTR-255 was shown to enhance in vivo therapeutic efficacy of daratumumab, a monoclonal antibody that mediates tumor killing by ADCC. The effect was mainly mediated by NK cells. The treatment scheduling experiments demonstrated that antibody should be present during NKTR-255-driven NK cell activation/expansion phase to efficiently deplete Daudi tumor cells in bone marrow. In addition, even low dose of daratumumab treatment showed effective tumor depletion when combined with high dose of NKTR-255. This suggests that NKTR-255 has a potential to expand the therapeutic window of existing tumor-targeted antibodies to patients with, for example, low antibody concentration in target tumor site or low expression of tumor antigen.

Disclosures
Miyazaki: Nektar Therapeutics: Employment, Equity Ownership. Kivimäe:Nektar Therapeutics: Employment, Equity Ownership. Hennessy:Nektar Therapeutics: Employment, Equity Ownership. Pena:Nektar Therapeutics: Employment, Equity Ownership. Quach:Nektar Therapeutics: Employment, Equity Ownership. Moffet:Nektar Therapeutics: Employment, Equity Ownership. Nieves:Nektar Therapeutics: Employment, Equity Ownership. Zhang:Nektar Therapeutics: Employment, Equity Ownership. Marcondes:Nektar Therapeutics: Employment, Equity Ownership. Madakamutil:Nektar Therapeutics: Employment, Equity Ownership. Zalevsky:Nektar Therapeutics: Employment, Equity Ownership.