Abrogating Cbl-b in effector CD8+ T cells improves the efficacy of adoptive therapy of leukemia in mice
Ingunn M. Stromnes, … , Hua Gu, Philip D. Greenberg
Ingunn M. Stromnes, … , Hua Gu, Philip D. Greenberg
Published September 20, 2010
Citation Information: J Clin Invest. 2010;120(10):3722-3734. https://doi.org/10.1172/JCI41991.
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Research Article Hematology

Abrogating Cbl-b in effector CD8+ T cells improves the efficacy of adoptive therapy of leukemia in mice

Abstract

The clinical use of adoptive immunotherapy with tumor-reactive T cells to treat established cancers is limited in part by the poor in vivo survival and function of the transferred T cells. Although administration of exogenous cytokines such as IL-2 can promote T cell survival, such strategies have many nonspecific activities and are often associated with toxicity. We show here that abrogating expression of Casitas B-lineage lymphoma b (Cbl-b), a negative regulator of lymphocyte activation, in tumor-reactive CD8+ T cells expanded ex vivo increased the efficacy of adoptive immunotherapy of disseminated leukemia in mice. Mechanistically, Cbl-b abrogation bypassed the requirement for exogenous IL-2 administration for tumor eradication in vivo. In addition, CD8+ T cells lacking Cbl-b demonstrated a lower threshold for activation, better survival following target recognition and stimulation, and enhanced proliferative responses as a result of both IL-2–dependent and –independent pathways. Importantly, siRNA knockdown of Cbl-b in human CD8+CD28– effector T cell clones similarly restored IL-2 production and proliferation following target recognition independent of exogenous IL-2, enhanced IFN-γ production, and increased target avidity. Thus, abrogating Cbl-b expression in effector T cells may improve the efficacy of adoptive therapy of some human malignancies.

Authors

Ingunn M. Stromnes, Joseph N. Blattman, Xiaoxia Tan, Sara Jeevanjee, Hua Gu, Philip D. Greenberg

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Figure 4

Ablating Cbl-b expression in both CD28– and CD28+ cells promotes Bcl-xL expression and cell survival.

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Ablating Cbl-b expression in both CD28– and CD28+ cells promotes Bcl-xL ...
(A and C) In vitro–derived effector cells were activated with antigen, with or without r-h-IL-2 (25 U/ml). After 4 days, the percentage of CD8+ transgenic cells stained for 7AAD and/or Annexin-V was determined by FACS analysis. The numbers in the top right quadrant indicate the percentage of transgenic T cells that are both Annexin-V+ and 7AAD+. The numbers in the bottom right quadrant indicate the percentage of transgenic cells that are Annexin-V+ and 7AAD–. The outlined numbers indicate the total percentage of transgenic cells that are Annexin-V+. (B and D) In vitro–derived Thy1.2+TCRgag effector cells of the indicated genotype were stimulated with peptide-pulsed Thy1.1+ splenocytes (5 μg/ml). After 4 days, CD8+Thy1.2+ were analyzed for intracellular expression of Bcl-2 and Bcl-xL using flow cytometry. The numbers in the plot indicate the percentage of transgenic cells that are positive for BCl-xL. Gray numbers correspond to (B) TCRgagCblb+/+ and (D) TCRgagCblb+/+CD28–/– cells, and black numbers correspond to (B) TCRgagCblb–/– and (D) TCRgagCblb–/–CD28–/– cells. The numbers in the parentheses indicate the MFI of the Bcl-xL+ population. Data represent 3 independent experiments.