Targeting of CD122 enhances antitumor immunity by altering the tumor immune environment

Abstract

Mounting evidence demonstrates that CD8⁺CD122⁺ T cells have suppressive properties with the capacity to inhibit T cell responses. Therefore, these cells are rational targets for cancer immunotherapy. Here, we demonstrate that CD122 monoclonal antibody (mAb; aCD122) therapy significantly suppressed tumor growth and improved long-term survival in tumor-bearing mice. This therapeutic effect correlated with enhanced polyfunctional, cytolytic intratumoral CD8⁺ T cells and a decrease in granulocytic myeloid-derived suppressor cells (G-MDSCs). In addition, aCD122 treatment synergized with a vaccine to augment vaccine-induced antigen (Ag)-specific CD8⁺ T cell responses, reject established tumors and generate memory T cells. Furthermore, aCD122 mAb synergized with an anti-GITR (aGITR) mAb to confer significant control of tumor growth. These results suggest CD122 might be a promising target for cancer immunotherapy, either as a single agent or in combination with other forms of immunotherapy.

Keywords: CD122; CD8 T cells; GITR; immunotherapy; vaccines.

Figure 1. Treatment with 5H4 mAb reduced CD8⁺CD122⁺ T cell population

Naïve B6 non-tumor bearing mice were injected i.p. with 100 ug of 5H4 anti-CD122 mAb or with the isotype (rat IgG2a) control. Three days after injection, splenocytes were analyzed for CD8⁺CD122⁺ T cells by flow cytometry. (A-B) Flow plot analysis and representative data showing the percentages of CD8⁺CD122⁺ T cells and the MFI of CD122 on CD8⁺CD122⁺ T cells in total splenic lymphocytes. Isotype indicates CD8⁺ splenocytes stained with the isotype control antibody for CD122 staining. Results are representative of 2 independent experiments. ^****P<0.0001. Errors bars indicate SEM of n = 5/group.

Figure 2. CD122 mAb treatment suppressed CT26 and B16-OVA tumor growth and enhanced long-term survival

Treatment regimen, individual tumor growth, group tumor measurements, and survival of CT26 (A) and B16-OVA (B) implanted mice following treatment are indicated. Figures represent 2 (CT26) and 3 (B16-OVA) independent experiments. ^**P<0.01; ^***P<0.001; ^****P<0.0001.

Figure 3. CD122 mAb treatment altered the cellular composition of the tumor immune microenvironment in the B16-OVA melanoma model

TILs from tumors of B16-OVA mice were harvested 16 days after tumor implantation. (A) CD45⁺ leukocyte infiltrate and CD8⁺ and CD4⁺ TILs as percentage of total CD45⁺ cells are shown in treated versus untreated groups. TIL populations, including CD4⁺ Tregs (B) and M-MDSC (CD11b⁺Ly6C⁺Ly6G^-) and G-MDSC (CD11b⁺Ly6C^-Ly6G⁺) (C) were identified by flow cytometry. (D) CD122 expression by G-MDSC was shown by flow cytometry. M-MDSCs also expressed CD122 (data not shown). ^*P<0.05; ^**P<0.01; ns not significant. Error bars indicate SEM of n = 5/group.

Figure 4. CD122 mAb treatment of B16-OVA tumors increased the frequency and function of specific inflammatory cytokine production of tumor infiltrating T cells and efficacy is dependent on CD8⁺ T cells

(A) Summary data showing percentage of CD4⁺CD45⁺ TILs expressing IFNγ. (B-C) Representative and summary data showing single-, double- and triple positive CD8⁺ T cells releasing IFNγ, TNFα and/or co-expressing the degranulation marker, CD107a, following OVA_257-264 and OVA_323-339 peptide incubation with PMA/ION stimulation. (D) Tumor growth curves and survival over time for B16-OVA tumor-bearing mice treated aCD122 and with/without anti-CD4 or anti-CD8 mAbs. Experiments were repeated at least two times. ^*P<0.05; ^**P<0.01; ^***P<0.001. Errors bars indicate SEM of n = 5/group (A-C); n= 10/group in (D).

Figure 5. Anti-CD122 mAb treatment synergizes with an aGITR mAb therapy

On day +4 post-implantation of B16-OVA, mice were treated with aCD122 or aGITR or the combination as indicated. Mean tumor growth and survival are depicted. Experiments were repeated at least two times. ^*P<0.05; ^**P<0.01; ^***P<0.001; ^****P<0.0001. Errors bars indicate SEM; n = 10/group.

Figure 6. CD122 mAb synergized with a tumor vaccine to inhibit the growth of B16-OVA tumors

(A) On day +7 post-implantation, mice were treated with aCD122 mAb (administered 5 times at 2-3 day intervals) or with peptide vaccine (1 dose) or the combination as indicated. Mean tumor growth and survival are depicted (B). TILs were harvested at day +16, followed by analysis of percent of tumor infiltrating G-MDSCs, IFNγ/TNFα cytokine secreting CD8⁺ T cells upon ex-vivo stimulation with OVA_257-264 peptide, OVA tetramer-specific CD8⁺ TILs, and Tregs. (C) On day +7 post-implantation, mice were treated with aCD122 in combination with peptide vaccine (3 doses) as indicated. Survival curve on the left shows primary treatment results; curve on the right shows rechallenged mice that rejected tumors after aCD122/vaccine treatment. Data are representative of 2-3 independent experiments. ^*P<0.05; ^**P<0.01; ^***P<0.001; ^****P<0.0001. Error bars indicate SEM of n = 5/group for the bar graphs; n=10/group for the efficacy studies in B, and D.