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510 Background: Patients with NMIBC CIS unresponsive to BCG have limited treatment options. N-803 (Anktiva) is a mutant IL-15-based immunostimulatory fusion protein complex (IL-15RαFc) that promotes proliferation and activation of natural killer (NK) cells and CD8+ T cells, but not regulatory T cells. Phase Ib data in BCG-naive patients with NMIBC demonstrate that intravesical administration of N-803 with BCG induced complete response in all patients, without recurrences for the study duration of 24 months. An open-label, 3 cohort multicenter phase II/III study (QUILT 3.032) of intravesical BCG plus N-803 in patients with BCG-unresponsive high-grade NMIBC (NCT03022825) was opened. We report here the interim analysis of Cohort A, BCG-unresponsive (CIS) [with or without Ta or T1 disease], as of December 2020 data cutoff. Methods: All treated patients received intravesical N-803 plus BCG, consistent with the standard induction/maintenance treatment schedule. The primary endpoint for Cohort A of this phase II/III study is incidence of complete response (CR) of CIS at any time. Results: To date, 80 patients have enrolled in cohort A of this phase II/III trial. Evaluable analysis at this time shows CR rate at any time of 72% (N=51/71); for patients achieving CR, the probability of maintaining a CR for 12 months is 59%, with a median duration of complete response of 19.2 (7.6, 26.4) months. Low-grade treatment related AEs include dysuria, hematuria, and pollakiurua (all 16%), urgency (14%), and bladder spasm (8%), all other AEs were seen at 6% or less. A total of 9 subjects experienced at least 1 treatment emergent SAE (Severe Adverse event), the SAE rate is 1% for any given AE. No treatment emergent SAE’s were considered treatment related. No immune related SAE’s have been seen. To date, 10/80 (12.5%) patients proceeded to cystectomy in this BCG unresponsive population. Conclusions:With a CR rate of 72%, N-803 has met its primary endpoint with 59% probability of CR patients maintaining CR for at least 12 months. With the observed strong efficacy and an SAE rate of 1%, N-803 represents a novel treatment option for BCG unresponsive CIS with a favorable benefit:risk ratio, in a therapeutically challenging disease. Clinical trial information: NCT03022825.

INTRODUCTION AND OBJECTIVES:Patients with non-muscle-invasive bladder cancer (NMIBC) who do not respond to Bacillus Calmette-Guerin (BCG) therapy have limited treatment options: the standard of car...

Human immunodeficiency virus (HIV) persists indefinitely in individuals with HIV who receive antiretroviral therapy (ART) owing to a reservoir of latently infected cells that contain replication-competent virus

e15008 Background: The oncotherapeutic promise of IL-15, a potent immune stimulant, is limited by short serum half-life. The fusion protein N-803 is an IL-15 superagonist complex that has > 20-fold longer half-life in vivo vs IL-15. This study characterized the pharmacokinetic (PK) profile, biological activity, and safety of N-803 after subcutaneous administration to healthy human volunteers. Methods: Volunteers were randomized 1:1 to receive 1 mg/mL or 2 mg/mL N-803. Each subject received 2 doses of N-803: 10 µg/kg followed 24 days later by 20 µg/kg. After each dose, PK and safety measures were assayed for 9 successive days. Primary endpoint was the PK profile of N-803; secondary was safety; and exploratory endpoints were cytokine levels, immune cell characterization, and immunogenicity. Results: N-803 resulted in no grade ≥3 or serious adverse events (AEs). Mild injection site reactions, chills, and pyrexia were the most common AEs. Serum N-803 concentrations peaked 10.3-15.4 hours after administration and declined with a half-life of 20.0-20.7 hours. Peak N-803 serum concentrations were dose-dependent, with a 1.5-fold increase in Cmax after administration of 20 µg/kg vs 10 µg/kg. In the peripheral blood, N-803 induced a transient decline, followed by a significant increase (3-fold) in NK cell number that persisted for ≥24 days. N-803 also caused a significant proliferation of NK (22-fold increase in Ki-67+ cells), CD8+ (27-fold) and CD4+ T (11-fold) cells; however, increased cell number occurred only in NK cells. N-803 administration also increased serum levels of interferon gamma, IL-10, and IL-6. One of 14 evaluable subjects had measureable anti-N-803 antibodies at the end-of-study visit. Conclusions: N-803 results in prolonged elevation of drug serum concentrations, contrasting with rapid clearance of recombinant human IL-15 (ie, half-life of ~20 vs < 1 hour). N-803 administration was well-tolerated in healthy volunteers, without evidence of adverse systemic inflammatory responses, and resulted in proliferation of NK cells and CD8+ T cells, as well as sustained increases in NK cell number. Findings in this study are consistent with published results from N-803 administration in treating liquid tumors and lung cancer. Our results demonstrate N-803 administration potentiates the proliferation and activity of lymphocytes with antitumor and antivirus properties, and suggest this investigational product holds promise in treatment of cancer as well as infectious disease such as HIV. Clinical trial information: NCT03381586 .

IL-15 is a cytokine that is crucial for the development, survival, and activation of NK cells, and pre-clinical studies demonstrated that IL-15 augments antibody-dependent cellular cytotoxicity. We hypothesized that IL-15 would augment anti-CD20 mAb (rituximab)-directed responses in patients with indolent non-Hodgkin’s lymphoma. To address this, we performed a first-in-human combination clinical trial of the IL-15 super agonist N-803 with rituximab, and investigated the impact of this novel immune combination on NK, CD8 T cells, and monocytes. The combination was safe and demonstrated clinical activity, including in patients refractory to rituximab. To understand the in vivo impact on the patients’ immune systems, we performed mass cytometry and cellular indexing of transcriptomes and epitopes by sequencing (CITE-seq), which measures single-cell RNA-seq and cell surface proteins simultaneously. N-803 induced expansion of NK (12.5-fold) and CD8 T cells (2-fold) in serial blood samples and upregulated CD38, chemokines, and MHC class II family member expression while maintaining CD16 expression on NK. On CD56dim NK, N-803 increased activating receptors, decreased CD57, and modulated transcription factor expression (AP-1, Eomes, CEBPB/D). CD56bright NK cells were primed, with increases in granzyme A, B, and K. Therapy also markedly altered the monocyte compartment, significantly upregulating type 1 interferon and interferon gamma pathways in CD14+ monocytes, and decreased the percent of CD16+ monocytes, suggestive of potential trafficking to the tumor. Collectively, our data demonstrates that N-803 plus rituximab induces complex immune activation, and supports N-803 combination with other mAbs and immunotherapies.

Background Neuroblastoma (NB) is the most common malignant extracranial solid tumor in children. Despite significant survival improvements in multiple pediatric malignancies, children with relapsed neuroblastoma (NB) have dismal outcomes. ROR1 is highly expressed on the majority of NB (Rebagay/Yan Front Oncol, 2012). Our group has successfully expanded functional and active peripheral blood NK cells (exPBNK) with irradiated feeder cells and electroporated CAR mRNA to exPBNK (Chu/Cairo, et al, Can Imm Res 2015). N-803 (ALT-803) is a novel IL15 superagonist complex where mutant IL-15N72D is bound to an IL-15RαSu-Fc fusion protein with higher potency and longer half-life compared to IL-15 (Han/Wong, Cytokine, 2011). Objectives To determine if anti-ROR1 CAR engineered NK cells can efficiently target ROR1+ NB and if the cytotoxicity can be further enhanced in combination with N-803. Methods ExPBNK cells were expanded with lethally irradiated K562-mbIL21cells (Denman/Lee Plos One, 2012) and isolated as previously described (Chu/Cairo, Can Imm Res, 2015). ExPBNK cells were electroporated with anti-ROR1-CAR mRNA (ROR1-CAR was generously provided by Riddell S) using maxcyte electroporator. ALT-803 was generously provided by Altor BioScience. CHLA-255 and CHLA-136 were generously provided by Seeger R. In vitro cytotoxicity of anti-ROR1 CAR NK against NB cell lines were examined at different E:T ratios. Granzyme B, Perforin and IFN-γ levels were evaluated by ELISA assays. High dimentionsal CyTOF analysis was performed to character the immunophenotying of anti-ROR1-CAR-NK. NB xenografted NSG mice were generated and tumor burden was monitored as we previously described (Chu/Cairo, Oncoimmunology 2017). Results Anti-ROR1-CAR was efficiently expressed on exPBNK after CAR mRNA-electroporation at 24 hrs. Anti-ROR1-CAR-NK were effective in killing NB cells even at very low E:T ratio compared to mock NK with enhanced granzyme B, perforin and interferon- release (p Conclusion anti-ROR1-CAR NK had significantly enhanced cytotoxicity against ROR1+ NB cells in vitro and in vivo using human NB xenografted NSG mice compared to controls. ALT-803 further enhanced the anti-tumor effect of anti-ROR1 CAR NK (supported by W81XWH-17-1-0420).

4561 Background: Patients with non-muscle-invasive bladder cancer (NMIBC) unresponsive to BCG therapy have limited treatment options. N-803 (also known as ALT-803) is an IL-15-based immunostimulatory protein complex (IL-15RαFc) that promotes proliferation and activation of natural killer (NK) cells and CD8+ T cells, but not regulatory T cells. Phase Ib data in BCG-naïve patients with NMIBC demonstrate that intravesical administration of N-803 with BCG induced complete response in all patients, without recurrences for the study duration of 24 months. Methods: An open-label, single-arm multicenter Phase 2 study of intravesical BCG plus N-803 in patients with BCG-unresponsive high-grade NMIBC (NCT03022825) was opened. The study has two cohorts: Cohort A, patients with BCG-unresponsive carcinoma in situ (CIS) [with or without Ta or T1 disease] and Cohort B, patients with BCG-unresponsive high-grade Ta/T1 disease. All treated patients receive intravesical N-803 plus BCG, similar to a standard induction and maintenance treatment schedule. The primary endpoint for Cohort A is incidence of complete response (CR) of CIS at any time, and the primary endpoint for Cohort B is disease-free rate at 12 months. Results: To date, forty-six patients have enrolled in this phase 2 trial (Cohort A (CIS), n = 23, Cohort B (Papillary), n = 23). Of eleven evaluable patients in Cohort A, nine patients (82%) have a reported CR. In addition, seven out of nine (78%) patients in Cohort A demonstrated CR at their 6-month response assessment. Of thirteen evaluable patients in Cohort B, ten patients (77%) showed no evidence of recurrence at their 3-month response assessment; of these, none (0/8) evaluated past 3 months have had disease recurrence. Three serious adverse events (AEs) have been reported (E coli infection, anemia, and bacteremia), with no immune-related AEs. Conclusions: Nine out of eleven (82%) patients with BCG-unresponsive CIS of the bladder demonstrated a complete response. Ten out of thirteen patients with BCG-unresponsive papillary NMIBC show no evidence of disease at first assessment. Intravesical N-803 plus BCG was well-tolerated and no patients experienced immune-related AEs. Clinical trial information: NCT03022825.

Background The overall survival rate for pediatric osteosarcoma (OS), neuroblastoma (NB) and glioblastoma (GBM) in children with relapsed/refractory and metastatic disease remains dismal. Our group has successfully expanded peripheral blood Natural Killer cells (exPBNK) with irradiated feeder cells (Chu/Cairo, et al, Can Imm Res 2015). ALT-803 (N-803) is a superagonist of an IL-15 variant bound to an IL-15RαSu-Fc fusion with enhanced IL-15 biological activity (Zhu et al. 2009 J Immunol). Tim-3 was found expressed on active NK cells and its expression in cancer patientsO NK cells correlated with disease stage and prognosis. Objective To determine if the combination of N-803 and TIM-3 blockage significantly enhances exPBNK cell cytotoxicity against OS, GBM and NB Method PBMCs were expanded with lethally irradiated K562-mbIL21-41BBL cells. CD56+CD3− exPBNK cells were isolated using Miltenyi NK cell isolation kit. N-803 was generously provided by Altor BioScience. ExPBNK were cultured with 3.5ng/ml N-803 and 10 or 20ug/ml anti-TIM-3 antibody. NK proliferation, NK receptors expression and cytotoxicity were examined (Chu/Cairo, oncoimmunology, 2017). Results N-803 increased NK activating receptorO expression: NKG2D, NKp30, NKp44, and NKp46. N-803 significantly enhanced exPBNK in vitro cytotoxicity against OS, NB and GBM (p Conclusions The combination of N-803 and anti-Tim-3 blockage significantly enhanced exPBNK in-vitro cytotoxicity against OS, NB and GBM cells. In-vivo effects of the combination are under investigation.