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1146 NKG2D co-stimulation via anti-sMIC/MIC targeting antibody enhances CD8 T cell functional heterogeneity and memory development in mouse prostate tumors
  1. Tyler Smith1,
  2. Sizhe Liu1,
  3. Lisha Zhu2 and
  4. Jennifer Wu1
  1. 1Northwestern University Feinberg, Chicago, IL, USA
  2. 2University of Chicago, Chicago, IL, USA


Background Effective co-stimulation of CD8 T cells is critical for anti-tumor effector responses and maintenance of antigen-specific TCF-1+ memory CD8 T cells necessary for long-term tumor control. NKG2D co-stimulation of CD8 T cells via binding of stress ligands such as membrane-bound MIC enhances CD8 T cell effector functions and is important for promoting memory development. Cancer cells often shed MIC into a soluble form (sMIC) that inhibits CD8 T cell activation. Humanized TRAMP/MICB prostate tumor mice treated with anti-sMIC/MIC targeting antibody develop potent anti-tumor responses by CD8 T cells by 1) sequestering sMIC and 2) inducing NKG2D pathway signaling, thereby re-invigorating CD8 T cell immunity. However, the mechanistic underpinnings of CD8 T cell reprogramming by NKG2D co-stimulation in the tumor microenvironment are poorly understood. In this preliminary study, we used scRNA-seq of CD8 TILs from TRAMP/MICB mice treated or untreated with the anti-sMIC/MIC targeting antibody B10G5 to investigate the hypothesis that B10G5 differentially reprograms CD8 T cells at the transcriptional and epigenetic level via sustained NKG2D pathway co-stimulation, thereby optimizing TCR-dependent effector and memory responses.

Methods Adult TRAMP/MICB mice were treated with 200µg B10G5 intraperitoneally twice per week for 4 weeks. Prostate tumors from B10G5-treated mice, untreated mice with well-differentiated tumors, and untreated mice with poorly differentiated tumors were collected, and their CD45+ cells isolated via FACS for scRNA-seq. Subclusters specific to CD8 T cells were the focus of subsequent transcriptomic analyses.

Results Prostate tumors from mice treated with anti-sMIC/MIC antibody B10G5 revealed enhanced functional heterogeneity of CD8 T cell subtypes compared to tumors from untreated mice. In contrast to tumors from untreated mice that were populated with primarily effector CD8 T cells expressing CD226hi, CXCR3hi, GZMBhi and/or NKG7, tumors from B10G5-treated mice were enriched in TCF7hi IL7R+ EOMEShi stem-like memory CD8 T cells and EOMES+ CD27hi PD-1+ effector memory CD8 T cells. These memory populations upregulated epigenetic modifiers (KMT2A/E) and transcription factors (NR4A1/2/3) important in metabolic reprogramming and memory CD8 T cell differentiation.

Conclusions These results establish a groundwork for identifying targets for epigenetic and metabolic alteration of CD8 T cells via NKG2D co-stimulation within the tumor microenvironment.

Ethics Approval All animal studies were approved by the Institutional Animal Care and Use Committee (IACUC) committee of Northwestern University.

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