Abstract 3268: Costimulatory T-cell engagement by PRS-342, a GPC3/4-1BB bispecific molecule, leads to activation of T-cells and tumor growth inhibition in a HCC humanized mouse model

e14604 Background: There is great interest in investigating immune checkpoint inhibition in combination with novel agents, however, preclinical studies are limited by the lack of immune-competent human cancer cell line xenograft models. We developed a “hematopoietic” humanized mouse model to address this unmet need. HDAC inhibitors (HDACi) induce T-cell chemokine expression and enhance response to PD1 inhibitors in lung cancer models. We evaluated the combination of OKI-179, a novel Class I HDACi and nivolumab in our humanized mouse model. Methods: BRGS newborn pups were transplanted with CD34+ cells purified from umbilical cord blood. At 16 weeks, MDA-MB-231 cells were implanted in the flanks of the mice. When the average tumor size reached ~150-300 mm3, the mice were randomized into vehicle, nivolumab, OKI-179, or the combination treatment according to % chimerism. At the end of the treatment, mice were euthanized and tissues were collected for further analysis. Results: We observed a statistically significant improvement in tumor growth inhibition with the combination of OKI-179 and nivolumab (%TGI = 77%) as compared to single agent nivolumab (45%TGI). Nivolumab treatment led to a significant decrease in PD-1 expression on T cells. We also observed an increased number of TILs (CD8+ T cells) and T cells in the lymph nodes of treated mice suggesting T cell expansion. All mice were highly chimeric and responding tumors exhibited an increase in CD44 high IFN+ T cells. We observed a higher CD8% and a higher effector memory % (HLADR+, CD45RO+) in these mice. Conclusions: We successfully established a humanized TNBC human cancer cell line xenograft model, with tumor engraftment occurred in all humanized mice. Mice treated with nivolumab demonstrating the development of lymph nodes that were populated by activated T cells. The combo resulted in superior tumor growth inhibition. These preliminary results demonstrate that human immunity and PD-1 expressing T cells exist in these models and provide the basis for planned immunotherapy combination studies.

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Lactate Increases Stemness of CD8+ T Cells to Augment Anti-Tumor Immunity

10.1101/2021.10.18.464724 ◽

2021 ◽

Author(s):

Qiang Feng ◽

Zhida Liu ◽

Xuexin Yu ◽

Tongyi Huang ◽

Jiahui Chen ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Tumor Growth ◽

Growth Inhibition ◽

Tumor Immunity ◽

Cd8 T Cells ◽

Subcutaneous Administration ◽

Tumor Growth Inhibition ◽

Mouse Splenocytes

Nutrients and metabolites play important roles in immune functions. Recent studies show lactate instead of glucose can serve as a primary carbon fuel source for most tissues. The role of lactate in tumor immunity is not well understood with immune suppressive functions reported for lactic acid, the conjugate acid form of lactate. In this study, we report lactate increases the stemness of CD8+ T cells and augments anti-tumor immunity. Subcutaneous administration of lactate but not glucose shows CD8+ T cell-dependent tumor growth inhibition. Single cell transcriptomics analysis revealed lactate treatment increased a subpopulation of stem-like TCF-1-expressing CD8+ T cells, which is further validated by ex vivo culture of CD8+ T cells from mouse splenocytes and human peripheral blood mononuclear cells. The inhibition of histone deacetylase activity by lactate increased acetylation in the histone H3K27 site at the Tcf7 super enhancer locus and increased the gene expression of Tcf7. Adoptive transfer of CD8+ T cells pretreated with lactate in vitro showed potent tumor growth inhibition in vivo. Our results elucidate the immune protective role of lactate in anti-tumor immunity without the masking effect of acid. These results may have broad implications for T cell therapy and the understanding of lactate in immune metabolism.

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Inhibition of LDH-A by Oxamate Enhances the Efficacy of Anti-PD-1 Treatment in an NSCLC Humanized Mouse Model

Frontiers in Oncology ◽

10.3389/fonc.2021.632364 ◽

2021 ◽

Vol 11 ◽

Author(s):

Tianyun Qiao ◽

Yanlu Xiong ◽

Yangbo Feng ◽

Wenwen Guo ◽

Yongsheng Zhou ◽

...

Keyword(s):

T Cells ◽

Mouse Model ◽

Tumor Growth ◽

Cd8 T Cells ◽

Immune Cells ◽

Preclinical Model ◽

Small Subset ◽

Therapeutic Effects ◽

Humanized Mouse ◽

Humanized Mouse Model

Immunotherapy is a curable treatment for certain cancers, but it is still only effective in a small subset of patients, partly because of the lack of sufficient immune cells in the tumor. It is reported that targeted lactate dehydrogenase (LDH) to reduce lactic acid production can promote the infiltration and activity of immune cells and turn tumors into hot tumors. Therefore, we constructed a humanized mouse model to evaluate the efficacy of using classical LDH inhibitor oxamate and pembrolizumab alone or in combination in non-small cell lung cancer (NSCLC). We found that both oxamate and pembrolizumab monotherapy significantly delayed tumor growth; moreover, combination therapy showed better results. Immunofluorescence analysis showed that oxamate treatment increased the infiltration of activated CD8+ T cells in the tumor, which might have enhanced the therapeutic effects of pembrolizumab. Treatment of the humanized mice with anti-CD8 abrogated the therapeutic effects of oxamate, indicating CD8+ T cells as the main force mediating the effect of oxamate. In conclusion, Our preclinical findings position that oxamate not only inhibits tumor growth at a high safe dose but also enhances the efficacy of pembrolizumab in Hu-PBMC-CDX mice. Our study also provides a preclinical model for exploring the efficacy of other immune-based combination therapies for NSCLC.

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Successful Regulatory T Cell-Based Therapy Relies on Inhibition of T Cell Effector Function and Enrichment of FOXP3+ Cells in a Humanized Mouse Model of Skin Inflammation

Journal of Immunology Research ◽

10.1155/2020/7680131 ◽

2020 ◽

Vol 2020 ◽

pp. 1-11

Author(s):

S. Landman ◽

V. L. de Oliveira ◽

M. Peppelman ◽

E. Fasse ◽

E. van Rijssen ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Mouse Model ◽

Human Skin ◽

Skin Inflammation ◽

Effector T Cells ◽

Effector Function ◽

Humanized Mouse ◽

Humanized Mouse Model

Background. Recent clinical trials using regulatory T cells (Treg) support the therapeutic potential of Treg-based therapy in transplantation and autoinflammatory diseases. Despite these clinical successes, the effect of Treg on inflamed tissues, as well as their impact on immune effector function in vivo, is poorly understood. Therefore, we here evaluated the effect of human Treg injection on cutaneous inflammatory processes in vivo using a humanized mouse model of human skin inflammation (huPBL-SCID-huSkin). Methods. SCID beige mice were transplanted with human skin followed by intraperitoneal (IP) injection of 20‐40×106 allogeneic human PBMCs. This typically results in human skin inflammation as indicated by epidermal thickening (hyperkeratosis) and changes in dermal inflammatory markers such as the antimicrobial peptide hBD2 and epidermal barrier cytokeratins K10 and K16, as well as T cell infiltration in the dermis. Ex vivo-expanded human Treg were infused intraperitoneally. Human cutaneous inflammation and systemic immune responses were analysed by immunohistochemistry and flow cytometry. Results. We confirmed that human Treg injection inhibits skin inflammation and the influx of effector T cells. As a novel finding, we demonstrate that human Treg injection led to a reduction of IL-17-secreting cells while promoting a relative increase in immunosuppressive FOXP3+ Treg in the human skin, indicating active immune regulation in controlling the local proinflammatory response. Consistent with the local control (skin), systemically (splenocytes), we observed that Treg injection led to lower frequencies of IFNγ and IL-17A-expressing human T cells, while a trend towards enrichment of FOXP3+ Treg was observed. Conclusion. Taken together, we demonstrate that inhibition of skin inflammation by Treg infusion, next to a reduction of infiltrating effector T cells, is mediated by restoring both the local and systemic balance between cytokine-producing effector T cells and immunoregulatory T cells. This work furthers our understanding of Treg-based immunotherapy.

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4308 DEEP-PRIMED IL-15 SUPERAGONIST IMPROVES ANTIVIRAL EFFICACY OF HIV-SPECIFIC CD8+ T-CELLS IN HUMANIZED MOUSE MODEL

Journal of Clinical and Translational Science ◽

10.1017/cts.2020.59 ◽

2020 ◽

Vol 4 (s1) ◽

pp. 4-5

Author(s):

Chase Daniel McCann ◽

Elizabeth Zale ◽

Adam Ward ◽

Thomas Dilling ◽

Ali Danesh ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Mouse Model ◽

Critical Role ◽

Humanized Mouse ◽

Humanized Mouse Model ◽

Viral Loads ◽

Specific Ctls ◽

The Impact

OBJECTIVES/GOALS: HIV-specific CD8+ T-cells play a critical role in partially controlling viral replication in infected-individuals, but ultimately fail to eliminate infection. Enhancing these T-cell responses through lymphocyte engineering approaches has the potential as a novel therapy capable of achieving durable control or eradication of infection. METHODS/STUDY POPULATION: IL-15 Superagonist (IL-15SA) potently supports the in vivo persistence and antiviral activity of adoptively transferred CD8+ T-cells. The Deep-PrimingTM technology platform, developed by Torque, allows for loading of immunomodulators onto the surface of T-cells via electrostatic ‘nanogels’, which slowly release to deliver sustained autocrine immune stimulation without the harmful effects of systemic exposure. Here, we investigate the impact of IL-15SA Deep-Priming on HIV-specific CD8+ T-cells in a humanized mouse model of HIV infection. Humanized mice were generated by engrafting NOD-scid-IL2Rgnull mice with memory CD4+ T-cells isolated from an ARV-suppressed HIV+ donor. An autologous HIV-specific Cytotoxic T-Lymphocyte (CTL) clone was isolated, and killing potential confirmed. Four weeks post humanization, mice were infected with HIV and received an infusion of unmodified HIV-Specific CTLs, or IL-15SA Deep-Primed HIV-specific CTLs (CTL-DP). T-cell numbers and plasma viral loads were quantified weekly by flow cytometry and qRT-PCR. RESULTS/ANTICIPATED RESULTS: Mice receiving unmodified CTLs trended toward reduced viral loads compared to the No Treatment condition, while mice receiving CTL-DP saw significant, 2-Log10 reductions in VL (p < 0.01). At 41 days post-infection 100% (5/5) of the No Treatment, 66.7% (4/6) of the CTL treatment, and 16.7% (1/6) of CTL-DP treatment mice had detectable viremia. IL-15SA Deep-Priming increased CTL expansion and persistence in peripheral blood which correlated with improved CD4+T-cell preservation. DISCUSSION/SIGNIFICANCE OF IMPACT: Here we demonstrate the first in vivo analysis of IL-15SA Deep-Priming of HIV-Specific CTLs. These data suggest that Deep-Priming of patient T-cells can enhance in vivo function and persistence, leading to improved viral suppression; a significant advancement in the field of HIV cure research. CONFLICT OF INTEREST DESCRIPTION: Austin Boesch, Thomas Andresen, and Douglas Jones are employees of Torque. Darrell Irvine is a co-founder of Torque and Chairman of Torque’s Scientific Advisory Board.

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Soluble GARP has potent antiinflammatory and immunomodulatory impact on human CD4+ T cells

Blood ◽

10.1182/blood-2012-12-474478 ◽

2013 ◽

Vol 122 (7) ◽

pp. 1182-1191 ◽

Cited By ~ 33

Author(s):

Susanne A. Hahn ◽

Heiko F. Stahl ◽

Christian Becker ◽

Anita Correll ◽

Franz-Joseph Schneider ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

Regulatory T Cells ◽

Mouse Model ◽

Cd4 T Cells ◽

Humanized Mouse ◽

Humanized Mouse Model ◽

Key Points ◽

Destructive Inflammation

Key Points GARP efficiently represses proliferation of naïve and resting CD4+ T cells and is involved in the induction of adaptive regulatory T cells. In vivo, GARP prevents T cell–mediated destructive inflammation in a preclinical humanized mouse model of GVHD.

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14 Novel CD3 epsilon humanized N-terminal epitope model for assessment of efficacy of T-cell engagers

Journal for ImmunoTherapy of Cancer ◽

10.1136/jitc-2020-sitc2020.0014 ◽

2020 ◽

Vol 8 (Suppl 3) ◽

pp. A14-A14

Author(s):

Gaëlle Martin ◽

Fabiane Sônego ◽

Audrey Beringer ◽

Chloé Beuraud ◽

Yacine Cherifi ◽

...

Keyword(s):

T Cells ◽

T Cell ◽

B Cells ◽

Mouse Model ◽

Humanized Mice ◽

Dependent Manner ◽

Salt Bridges ◽

Humanized Mouse ◽

Humanized Mouse Model ◽

Concentration Dependent Manner

BackgroundT-cell engagers have proved to be a promising therapeutic strategy in immunotherapy, for redirecting T cells activity against tumor cells. To facilitate the preclinical assessment of novel T-cell engagers and their translatability, we have developed an immunocompetent CD3 epsilon N-terminal epitope humanized mouse model.MethodsThis model was developed to express the human epitope of the CD3 epsilon chain, which is recognized by approximately 70% of the T-cell engagers (clone SP34). The rest of the extracellular domain was kept from mouse origin to preserve the amino acids involved in the interaction with CD3 gamma and delta. Similarly, the transmembrane domains and the intracellular domains where kept murine to enable salt bridges interaction, interaction with the CD3 zeta and the signaling into mouse cells.ResultsT cells from CD3 epsilon epitope humanized mice are found in comparable frequency in spleen, blood and bone marrow from WT mice. B cells, monocytes, dendritic cells and NK frequencies are also similar to the frequencies of these cell types in WT mice, suggesting that the humanization of the epitope of CD3 epsilon did not alter the immune cells distribution in these mice. Activation of T cells with antibodies targeting human CD3 (clone SP34) induced CD4 and CD8 T cell proliferation, as well as production of IL-2 and IFN-gamma. The CD3 functionality was demonstrated in vitro by the ability of B cells to produce IgM upon activation of T cells, suggesting a proper cooperation between T and B cells. Additionally, a first class of T-cell engagers targeting both human CD3 and a tumoral antigen, induced tumor cell lysis of MC38-Ag in a concentration-dependent manner. A second class of T cell engagers, also targeting CD3 and a tumoral antigen, showed an anti-tumor effect in vivo, and this effect was also shown to be dose-dependent.ConclusionsThese data suggest that the CD3 epsilon N-terminal epitope humanized mouse model enables the assessment of efficacy and mechanism of action of T-cell engagers.This model is currently being intercrossed with immunostimulatory humanized mouse models to provide new opportunities for assessment of bi-specific antibodies targeting the CD3 and immunostimulatory molecules. This model is the first generation of a broader program aiming at developping a Pan CD3 humanized model, where the gamma, delta and epsilon chains of the CD3 complex will be humanized. The Pan CD3 humanized mice are currently being investigated for immune responses and would provide a broader tool for assessment of T-cell engagers.

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