(word count: 239):Murine syngeneic tumor models are critical to novel immuno-based therapy development but the molecular and immunological features of these models are still not clearly defined. The translational relevance of differences between the models is not fully understood, impeding appropriate preclinical model selection for target validation, and ultimately hindering drug development. Across a panel of commonly-used murine syngeneic tumor models, we showed variable responsiveness to immunotherapies. We employed array comparative genomic hybridization, whole-exome sequencing, exon microarray analysis, and flow cytometry to extensively characterize these models, which revealed striking differences that may underlie these contrasting response profiles. We identified strong differential gene expression in immune-related pathways and changes in immune cell-specific genes that suggested differences in tumor immune infiltrates between models. Further investigation using flow cytometry showed differences in both the composition and magnitude of the tumor immune infiltrates, identifying models that harbor 'inflamed' and 'non-inflamed' tumor immune infiltrate phenotypes. We also found that immunosuppressive cell types predominated in syngeneic mouse tumor models that did not respond to immune-checkpoint blockade, whereas cytotoxic effector immune cells were enriched in responsive models. A cytotoxic cell-rich tumor immune infiltrate has been correlated with increased efficacy of immunotherapies in the clinic and these differences could underlie the varying response profiles to immunotherapy between the syngeneic models. This characterization highlighted the importance of extensive profiling and will enable investigators to select appropriate models to interrogate the activity of immunotherapies as well as combinations with targeted therapies in vivo.4

Programmed cell-death 1 ligand 1 (PD-L1) is a member of the B7/CD28 family of proteins that control T-cell activation. Many tumors can upregulate expression of PD-L1, inhibiting antitumor T-cell responses and avoiding immune surveillance and elimination. We have identified and characterized MEDI4736, a human IgG1 monoclonal antibody that binds with high affinity and specificity to PD-L1 and is uniquely engineered to prevent antibody-dependent cell-mediated cytotoxicity. In vitro assays demonstrate that MEDI4736 is a potent antagonist of PD-L1 function, blocking interaction with PD-1 and CD80 to overcome inhibition of primary human T-cell activation. In vivo MEDI4736 significantly inhibits the growth of human tumors in a novel xenograft model containing coimplanted human T cells. This activity is entirely dependent on the presence of transplanted T cells, supporting the immunological mechanism of action for MEDI4736. To further determine the utility of PD-L1 blockade, an anti-mouse PD-L1 antibody was investigated in immunocompetent mice. Here, anti-mouse PD-L1 significantly improved survival of mice implanted with CT26 colorectal cancer cells. The antitumor activity of anti-PD-L1 was enhanced by combination with oxaliplatin, which resulted in increased release of HMGB1 within CT26 tumors. Taken together, our results demonstrate that inhibition of PD-L1 function can have potent antitumor activity when used as monotherapy or in combination in preclinical models, and suggest it may be a promising therapeutic approach for the treatment of cancer. MEDI4736 is currently in several clinical trials both alone and in combination with other agents, including anti-CTLA-4, anti-PD-1, and inhibitors of IDO, MEK, BRAF, and EGFR.

In this study, we compared human immunodeficiency virus (HIV) type 1-specific proliferative responses with HIV-1-induced intracellular cytokine production in a cohort of clinically nonprogressing patients and individuals with progressive HIV-1 infection. We found strong HIV-1-specific proliferative responses in the clinical nonprogressor cohort that correlated with significant numbers of circulating HIV-1-specific CD4(+) T cells. In contrast, HIV-1-specific proliferative responses were absent in most individuals with progressive HIV-1 infection, even though interferon-gamma-producing HIV-1-specific CD4(+) T cells were detectable by flow cytometry. The implication of these data is that the important dysfunction seen in most HIV-positive patients from very early in disease may be an inability of HIV-1-specific CD4(+) memory T cells to proliferate in response to HIV antigens rather than an absolute loss of circulating virus-specific CD4(+) T cells.

Purpose: While immune checkpoint inhibitors such as anti–PD-L1 are rapidly becoming the standard of care in the treatment of many cancers, only a subset of treated patients have long-term responses. IL12 promotes antitumor immunity in mouse models; however, systemic recombinant IL12 had significant toxicity and limited efficacy in early clinical trials. Experimental Design: We therefore designed a novel intratumoral IL12 mRNA therapy to promote local IL12 tumor production while mitigating systemic effects. Results: A single intratumoral dose of mouse (m)IL12 mRNA induced IFNγ and CD8+ T-cell–dependent tumor regression in multiple syngeneic mouse models, and animals with a complete response demonstrated immunity to rechallenge. Antitumor activity of mIL12 mRNA did not require NK and NKT cells. mIL12 mRNA antitumor activity correlated with TH1 tumor microenvironment (TME) transformation. In a PD-L1 blockade monotherapy-resistant model, antitumor immunity induced by mIL12 mRNA was enhanced by anti–PD-L1. mIL12 mRNA also drove regression of uninjected distal lesions, and anti–PD-L1 potentiated this response. Importantly, intratumoral delivery of mRNA encoding membrane-tethered mIL12 also drove rejection of uninjected lesions with very limited circulating IL12p70, supporting the hypothesis that local IL12 could induce a systemic antitumor immune response against distal lesions. Furthermore, in ex vivo patient tumor slice cultures, human IL12 mRNA (MEDI1191) induced dose-dependent IL12 production, downstream IFNγ expression and TH1 gene expression. Conclusions: These data demonstrate the potential for intratumorally delivered IL12 mRNA to promote TH1 TME transformation and robust antitumor immunity. See related commentary by Cirella et al., p. 6080

BackgroundThe ability to modulate immune-inhibitory pathways using checkpoint blockade antibodies such as αPD-1, αPD-L1, and αCTLA-4 represents a significant breakthrough in cancer therapy in recent years. This has driven interest in identifying small-molecule-immunotherapy combinations to increase the proportion of responses. Murine syngeneic models, which have a functional immune system, represent an essential tool for pre-clinical evaluation of new immunotherapies. However, immune response varies widely between models and the translational relevance of each model is not fully understood, making selection of an appropriate pre-clinical model for drug target validation challenging.MethodsUsing flow cytometry, O-link protein analysis, RT-PCR, and RNAseq we have characterized kinetic changes in immune-cell populations over the course of tumor development in commonly used syngeneic models.ResultsThis longitudinal profiling of syngeneic models enables pharmacodynamic time point selection within each model, dependent on the immune population of interest. Additionally, we have characterized the changes in immune populations in each of these models after treatment with the combination of α-PD-L1 and α-CTLA-4 antibodies, enabling benchmarking to known immune modulating treatments within each model.ConclusionsTaken together, this dataset will provide a framework for characterization and enable the selection of the optimal models for immunotherapy combinations and generate potential biomarkers for clinical evaluation in identifying responders and non-responders to immunotherapy combinations.