Recent accomplishments include the following: [ ] MECHANISMS INVOLVED IN IL-15 SUPERAGONIST ENHANCEMENT OF ANTI-PD-L1 THERAPY: Immunotherapy targeting PD-1/PD-L1 fails to induce clinical responses in most patients with solid cancers. N-803, formerly ALT-803, is an IL-15 superagonist mutant and dimeric IL-15RalphaSushi-Fc fusion protein complex that enhances CD8+ T and NK cell expansion and function and exhibits anti-tumor efficacy in preclinical models. Previous in vitro studies have shown that IL-15 increases PD-L1 expression, a negative regulator of CD8+ T and NK cell function. Most reported preclinical studies administered N-803 intraperitoneally not subcutaneously, the current clinical route of administration. N-803 is now being evaluated clinically in combination with PD-1/PD-L1 inhibitors. However, the mechanism of action has not been fully elucidated. We have examined the anti-tumor efficacy and immunomodulatory effects of combining N-803 with an anti-PD-L1 antibody in preclinical models of solid carcinomas refractory to anti-PD-L1 or N-803. Subcutaneous N-803 and an anti-PD-L1 monoclonal antibody were administered as monotherapy or in combination to 4T1 triple negative breast and MC38-CEA colon tumor-bearing mice. Anti-tumor efficacy was evaluated, and a comprehensive analysis of the immune-mediated effects of each therapy was performed on the primary tumor, lung as a site of metastasis, and spleen. We demonstrated that N-803 treatment increased PD-L1 expression on immune cells in vivo, supporting the combination of N-803 and anti-PD-L1. N-803 plus anti-PD-L1 was well-tolerated, reduced 4T1 lung metastasis and MC38-CEA tumor burden, and increased survival as compared to N-803 and anti-PD-L1 monotherapies. Efficacy of the combination therapy was dependent on both CD8+ T and NK cells and was associated with increased numbers of these activated immune cells in the lung and spleen. Most alterations to NK and CD8+ T cell phenotype and number were driven by N-803. However, the addition of anti-PD-L1 to N-803 significantly enhanced CD8+ T cell effector function versus N-803 and anti-PD-L1 monotherapies, as indicated by increased Granzyme B and IFNgamma production, at the site of metastasis and in the periphery. Increased CD8+ T cell effector function correlated with higher serum IFNgamma levels, without related toxicities, and enhanced anti-tumor efficacy of the N-803 plus anti-PD-L1 combination versus either monotherapy. We have provided a novel insight into the mechanism of action of N-803 plus anti-PD-L1 combination and offer preclinical proof of concept supporting clinical use of N-803 in combination with checkpoint inhibitors, including for patients non- and/or minimally responsive to either monotherapy. [ ] THE MULTI-FUNCTIONALITY OF N-809, A NOVEL FUSION PROTEIN ENCOMPASSING ANTI-PD-L1 AND THE IL-15 SUPERAGONIST FUSION COMPLEX: We have described a novel bifunctional fusion protein, designated N-809. This molecule comprises the IL-15/IL15Ralpha superagonist complex containing the Fc-domain of IgG1 (N-803, formerly designated as ALT-803) fused to two single chain anti-PD-L1 domains. The fully human IgG1 portion of the N-809 molecule was designed to potentially mediate antibody dependent cellular cytotoxicity (ADCC). These studies show that N-809 has the same ability to bind PD-L1 as an anti-PD-L1 monoclonal antibody. RNAseq studies show the ability of N-809 to alter the expression of an array of genes of both CD4+ and CD8+ human T cells, and to enhance their proliferation; CD8+ T cells exposed to N-809 also have enhanced ability to lyse human tumor cells. An array of genes was differentially expressed in human natural killer (NK) cells following N-809 treatment, and there was increased expression of several surface activating receptors; there was, however, no increase in the expression of inhibitory receptors known to be upregulated in exhausted NK cells. N-809 also increased the cytotoxic potential of NK cells, as shown by increased expression of granzyme B and perforin. The lysis of several tumor cell types was increased when either NK cells or tumor cells were exposed to N-809. Similarly, the highest level of ADCC was seen when both NK cells (from donors or cancer patients) and tumor cells were exposed to N-809. These studies thus demonstrate the multi-functionality of this novel agent. [ ] TEMPORAL CHANGES WITHIN THE (BLADDER) TUMOR MICROENVIRONMENT THAT ACCOMPANY THE THERAPEUTIC EFFECTS OF THE IMMUNOCYTOKINE NHS-IL12: While significant strides in the treatment of metastatic bladder cancer have been made with immune checkpoint inhibitors, the treatment of carcinoma in situ and non-muscle invasive, non-metastatic (superficial) human urothelial carcinoma, also termed non-muscle invasive bladder cancer (NMIBC), remains intractable with bacillus Calmette-Guerin (BCG) employed as the standard of care. In this study, an immunocytokine, NHS-muIL12, which consists of two molecules of murine IL-12 fused to NHS76, a tumor necrosis-targeting human IgG1, was examined as an immunotherapeutic in an orthotopic MB49luc bladder tumor model. The antitumor activity of systemic administration of NHS-muIL12 was investigated on MB49luc tumors, an aggressive, bioluminescent orthotopic bladder cancer model. Temporal studies were carried out on MB49luc bladder tumors harvested during various time points during NHS-muIL12 treatment and cellular changes associated with the reduction in tumor burden following NHS-muIL12 were determined by flow cytometry. Effects of those changes on the proliferation/activation of lymphoid cells were also determined. Studies revealed a significant reduction in MB49luc bladder tumor burden occurring between days 3 and 6 after the third and final systemic administration of NHS-muIL12. Temporal analyses of the MB49luc bladder tumor microenvironment (TME) initially revealed a large accumulation of myeloid-derived suppressor cells (MDSCs) and macrophages that elicited potent immunosuppression. Immunosuppression was characterized by the inability of CD4+ and CD8+ T cells to respond to broad-based immune stimulants. NHS-muIL12 administration resulted in temporal-dependent reductions in the number of MDSCs, macrophages and tumor-associated TGF-beta, which culminated in a re-ignition of CD4+ and CD8+ T cells to elicit potent antitumor responses against MB49luc bladder tumors. These findings provide strong evidence that the systemic administration of an immunocytokine consisting of a tumor-targeting Ig through recognition of DNA and DNA-histone complexes coupled to muIL-12 can effectively target the bladder TME; this significantly reduces the myeloid cellular compartment and reverts an immunosuppressive to an immunopermissive TME, ultimately resulting in antitumor effects. These studies provide further rationale for the employment of NHS-IL12 as an immunomodulator and clinical immunotherapeutic for NMIBC.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Investigator-Initiated Intramural Research Projects (ZIA)
Project #
Application #
Study Section
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
National Cancer Institute Division of Basic Sciences
Zip Code
Knudson, Karin M; Hicks, Kristin C; Luo, Xiaoling et al. (2018) M7824, a novel bifunctional anti-PD-L1/TGF? Trap fusion protein, promotes anti-tumor efficacy as monotherapy and in combination with vaccine. Oncoimmunology 7:e1426519
Fujii, Rika; Schlom, Jeffrey; Hodge, James W (2018) A potential therapy for chordoma via antibody-dependent cell-mediated cytotoxicity employing NK or high-affinity NK cells in combination with cetuximab. J Neurosurg 128:1419-1427
McGee, Heather M; Daly, Megan E; Azghadi, Sohelia et al. (2018) Stereotactic Ablative Radiation Therapy Induces Systemic Differences in Peripheral Blood Immunophenotype Dependent on Irradiated Site. Int J Radiat Oncol Biol Phys 101:1259-1270
Friedman, Jay; Morisada, Megan; Sun, Lillian et al. (2018) Inhibition of WEE1 kinase and cell cycle checkpoint activation sensitizes head and neck cancers to natural killer cell therapies. J Immunother Cancer 6:59
Grenga, Italia; Donahue, Renee N; Gargulak, Morgan L et al. (2018) Anti-PD-L1/TGF?R2 (M7824) fusion protein induces immunogenic modulation of human urothelial carcinoma cell lines, rendering them more susceptible to immune-mediated recognition and lysis. Urol Oncol 36:93.e1-93.e11
Fujii, Rika; Jochems, Caroline; Tritsch, Sarah R et al. (2018) An IL-15 superagonist/IL-15R? fusion complex protects and rescues NK cell-cytotoxic function from TGF-?1-mediated immunosuppression. Cancer Immunol Immunother 67:675-689
Malamas, Anthony S; Hammond, Scott A; Schlom, Jeffrey et al. (2017) Combination therapy with an OX40L fusion protein and a vaccine targeting the transcription factor twist inhibits metastasis in a murine model of breast cancer. Oncotarget 8:90825-90841
Jochems, Caroline; Tritsch, Sarah R; Pellom, Samuel Troy et al. (2017) Analyses of functions of an anti-PD-L1/TGF?R2 bispecific fusion protein (M7824). Oncotarget 8:75217-75231
Morillon 2nd, Y Maurice; Hammond, Scott A; Durham, Nicholas M et al. (2017) Enhanced immunotherapy by combining a vaccine with a novel murine GITR ligand fusion protein. Oncotarget 8:73469-73482
Fallon, Jonathan K; Vandeveer, Amanda J; Schlom, Jeffrey et al. (2017) Enhanced antitumor effects by combining an IL-12/anti-DNA fusion protein with avelumab, an anti-PD-L1 antibody. Oncotarget 8:20558-20571

Showing the most recent 10 out of 55 publications