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BackgroundThe tumor microenvironment (TME) often contains high levels of suppressive myeloid cells that contribute to innate checkpoint inhibitor (CPI) resistance. Pionyr’s Myeloid Tuning approach involves altering the composition and/or the function of myeloid cells in the TME. Myeloid reprogramming alters the function of immunosuppressive myeloid cells to acquire an immunostimulatory phenotype. Triggering receptor expressed on myeloid cells-1 (TREM1) is an immunoglobulin superfamily cell surface receptor enriched on tumor-associated myeloid cells. To investigate the potential of TREM1 modulation as an anti-cancer therapeutic strategy, Pionyr developed an afucosylated humanized anti-TREM1 monoclonal antibody termed PY159 and characterized it in pre-clinical and translational biomarker assays described below.MethodsPY159 responses in human whole blood and dissociated primary tumor cells in vitro were evaluated by flow cytometry and measurement of secreted cytokines and chemokines by MSD. TREM1 expression in human tumors was validated by scRNAseq, flow cytometry, and immunohistochemistry (IHC). In vivo efficacy and pharmacodynamic studies of a surrogate anti-mouse TREM1 antibody, termed PY159m, were evaluated using syngeneic mouse tumor models, either as a single-agent or in combination with anti-PD-1. To select tumor types and patients most likely to benefit from PY159 therapy, Pionyr developed qualitative and quantitative monoplex and multiplex IHC assays that detect TREM1 expression levels in human tumor tissues.ResultsPY159 treatment in vitro induced signaling, upregulated monocyte activation markers, and induced proinflammatory cytokines. In human tumors, TREM1 was detected on tumor-associated neutrophils, tumor-associated macrophages, and monocytic myeloid-derived suppressive cells. The surrogate PY159m anti-mouse TREM1 antibody exhibited anti-tumor efficacy in several syngeneic mouse tumor models, both as single-agent and in combination with anti-PD-1. Screening for TREM1 expression in tumor tissues demonstrated that TREM1+ tumor associated myeloid cells were highly enriched in the TME of multiple solid tumor indications. The monoplex and multiplex IHC assays offered insights into the localization of TREM1+ myeloid cells and their spatial relationship with other immune cells present in the TME to determine what immune composition will be more favorable for response to PY159 therapy.ConclusionsCollectively, the available nonclinical data support PY159 as a TREM1 agonist that reprograms myeloid cells and unleashes anti-tumor immunity. PY159 safety and efficacy are currently being evaluated in first-in-human clinical trial (NCT04682431) involving select advanced solid tumors patients resistant and refractory to standard of care therapies alone and in combination with a CPI. The TREM1 IHC assay is successfully being used on FFPE archival tumor tissues from enrolled patients to determine TREM1 expression levels.

Background Tumor-associated myeloid cells can impede productive anti-tumor immunity. One strategy for targeting immunosuppression is myeloid reprogramming, which drives immunosuppressive myeloid cells to acquire an immunostimulatory phenotype. Triggering receptor expressed on myeloid cells-1 (TREM1) is an immunoglobulin superfamily cell surface receptor expressed on neutrophils and subsets of monocytes and tissue macrophages. TREM1 associates with DAP12 adaptor and induces proinflammatory signaling, amplifies innate immune responses, and is implicated in the development of acute and chronic inflammatory diseases. TREM1 is also enriched in tumors, specifically on tumor-associated myeloid cells. To investigate the potential of TREM1 modulation as an anti-cancer therapeutic strategy, we developed PY159, an afucosylated humanized anti-TREM1 monoclonal antibody, and characterized it in the pre-clinical assays described below. Materials and Methods An FcγR binding ELISA and a Jurkat TREM1/DAP12 NFAT-luciferase reporter cell line were used to assess PY159 binding to human FcγRs and TREM1 signaling, respectively. PY159 responses in human whole blood in vitro were evaluated by flow cytometry, transcriptional analysis of sorted leukocyte subsets, and measurement of secreted cytokines/chemokines by MSD. A Transwell system was used to evaluate PY159 effects on neutrophil chemotaxis. TREM1 expression in human tumors was validated by scRNAseq, immunohistochemistry, and flow cytometry. Anti-tumor efficacy of a surrogate anti-mouse TREM1 antibody, PY159m, was evaluated using syngeneic mouse tumor models, either as a single agent or in combination with anti-PD-1. Results PY159 afucosylation increased its binding affinity for FcγR and its ability to activate TREM1/DAP12 signaling. In human blood assays, PY159 treatment did not induce depletion of TREM1-expressing cells. Rather, it upregulated monocyte activation markers, promoted neutrophil chemotaxis, and induced proinflammatory cytokines and chemokines, which was dependent on PY159 afucosylation. In human tumors, TREM1 was detected on tumor-associated neutrophils, tumor-associated macrophages, and monocytic myeloid-derived suppressive cells. PY159 induced proinflammatory cytokines and chemokines in dissociated human tumors in vitro, demonstrating that PY159 can reprogram tumor-associated myeloid cells. A surrogate anti-mouse TREM1 antibody, PY159m, exhibited anti-tumor efficacy in several syngeneic mouse tumor models, both as single-agent and in combination with anti-PD-1. Conclusions These results show that PY159 is a TREM1 agonist that reprograms myeloid cells and unleashes anti-tumor immunity. PY159 safety and efficacy are currently being evaluated in first-in-human clinical trial (NCT04682431) involving patients resistant and refractory to standard of care therapies. Disclosure Information V. Juric: A. Employment (full or part-time); Significant; Pionyr Immunotherapeutics Inc. E. Mayes: A. Employment (full or part-time); Significant; Pionyr Immunotherapeutics Inc. M. Binnewies: A. Employment (full or part-time); Significant; Pionyr Immunotherapeutics Inc. P. Canaday: A. Employment (full or part-time); Significant; Pionyr Immunotherapeutics Inc. T. Lee: A. Employment (full or part-time); Significant; Pionyr Immunotherapeutics Inc. S. Dash: A. Employment (full or part-time); Significant; Pionyr Immunotherapeutics Inc. J.L. Pollack: A. Employment (full or part-time); Significant; Pionyr Immunotherapeutics Inc. J. Rudolph: A. Employment (full or part-time); Significant; Pionyr Immunotherapeutics Inc. V. Huang: A. Employment (full or part-time); Significant; Pionyr Immunotherapeutics Inc. X. Du: A. Employment (full or part-time); Significant; Pionyr Immunotherapeutics Inc. N. Jahchan: A. Employment (full or part-time); Significant; Pionyr Immunotherapeutics Inc. A.J. Ramoth: A. Employment (full or part-time); Significant; Pionyr Immunotherapeutics Inc. S. Mankikar: A. Employment (full or part-time); Significant; Pionyr Immunotherapeutics Inc. M. Norng: A. Employment (full or part-time); Significant; Pionyr Immunotherapeutics Inc. C. Santamaria: A. Employment (full or part-time); Significant; Pionyr Immunotherapeutics Inc. K.P. Baker: A. Employment (full or part-time); Significant; Pionyr Immunotherapeutics Inc. L. Liang: A. Employment (full or part-time); Significant; Pionyr Immunotherapeutics Inc.

Summary: Converting checkpoint inhibitor (CPI)-resistant individuals to being responsive requires identifying suppressive mechanisms. We identify TREM2+ tumor-associated macrophages (TAMs) as being correlated with exhausted CD8+ tumor-infiltrating lymphocytes (TILs) in mouse syngeneic tumor models and human solid tumors of multiple histological types. Fc domain-enhanced anti-TREM2 monoclonal antibody (mAb) therapy promotes anti-tumor immunity by elimination and modulation of TAM populations, which leads to enhanced CD8+ TIL infiltration and effector function. TREM2+ TAMs are most enriched in individuals with ovarian cancer, where TREM2 expression corresponds to disease grade accompanied by worse recurrence-free survival. In an aggressive orthotopic ovarian cancer model, anti-TREM2 mAb therapy drives potent anti-tumor immunity. These results highlight TREM2 as a highly attractive target for immunotherapy modulation in individuals who are refractory to CPI therapy and likely have a TAM-rich tumor microenvironment.

Converting checkpoint inhibitor (CPI) resistant patients to responsive requires identifying novel suppressive mechanisms. We identified TREM2 + tumor associated macrophages (TAMs) as being correlated with exhausted CD8 + tumor infiltrating lymphocytes (TILs) in both mouse syngeneic tumor models and human solid tumors of multiple histological types. Fc-domain enhanced anti-TREM2 monoclonal antibody (mAb) therapy promoted anti-tumor immunity by elimination and modulation of TAM populations which led to enhanced CD8 + TIL infiltration and effector function. TREM2 + TAMs were most enriched in human ovarian cancer patients, where TREM2 expression corresponded to disease grade accompanied by worse recurrence-free survival. In an aggressive orthotopic ovarian cancer model, anti-TREM2 mAb therapy drove potent anti-tumor immunity. Together, these results highlight TREM2 as a highly attractive target for immunotherapeutic modulation in patients who are refractory to CPI therapy and likely have a TAM-rich tumor microenvironment.