American Association for Cancer Research
23266066cir180637-sup-208596_2_supp_5261052_pkygym.pdf (4.44 MB)

Supplementary Data from CD96 Is an Immune Checkpoint That Regulates CD8+ T-cell Antitumor Function

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journal contribution
posted on 2023-04-04, 00:46 authored by Deepak Mittal, Ailin Lepletier, Jason Madore, Amelia Roman Aguilera, Kimberley Stannard, Stephen J. Blake, Vicki L.J. Whitehall, Cheng Liu, Mark L. Bettington, Kazuyoshi Takeda, Georgina V. Long, Richard A. Scolyer, Ruth Lan, Nathan Siemers, Alan Korman, Michele W.L. Teng, Robert J. Johnston, William C. Dougall, Mark J. Smyth

Supplementary Figures 1-10


National Health and Medical Research Council

Cancer Council of Queensland

Senior Principal Research




CD96 is a novel target for cancer immunotherapy shown to regulate NK cell effector function and metastasis. Here, we demonstrated that blocking CD96 suppressed primary tumor growth in a number of experimental mouse tumor models in a CD8+ T cell–dependent manner. DNAM-1/CD226, Batf3, IL12p35, and IFNγ were also critical, and CD96-deficient CD8+ T cells promoted greater tumor control than CD96-sufficient CD8+ T cells. The antitumor activity of anti-CD96 therapy was independent of Fc-mediated effector function and was more effective in dual combination with blockade of a number of immune checkpoints, including PD-1, PD-L1, TIGIT, and CTLA-4. We consistently observed coexpression of PD-1 with CD96 on CD8+ T lymphocytes in tumor-infiltrating leukocytes both in mouse and human cancers using mRNA analysis, flow cytometry, and multiplex IHF. The combination of anti-CD96 with anti–PD-1 increased the percentage of IFNγ-expressing CD8+ T lymphocytes. Addition of anti-CD96 to anti–PD-1 and anti-TIGIT resulted in superior antitumor responses, regardless of the ability of the anti-TIGIT isotype to engage FcR. The optimal triple combination was also dependent upon CD8+ T cells and IFNγ. Overall, these data demonstrate that CD96 is an immune checkpoint on CD8+ T cells and that blocking CD96 in combination with other immune-checkpoint inhibitors is a strategy to enhance T-cell activity and suppress tumor growth.

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