PKCθ is required for alloreactivity and GVHD but not for immune responses toward leukemia and infection in mice
Javier O. Valenzuela, Cristina Iclozan, Mohammad S. Hossain, Martin Prlic, Emily Hopewell, Crystina C. Bronk, Junmei Wang, Esteban Celis, Robert W. Engelman, Bruce R. Blazar, Michael J. Bevan, Edmund K. Waller, Xue-Zhong Yu, Amer A. Beg
Javier O. Valenzuela, Cristina Iclozan, Mohammad S. Hossain, Martin Prlic, Emily Hopewell, Crystina C. Bronk, Junmei Wang, Esteban Celis, Robert W. Engelman, Bruce R. Blazar, Michael J. Bevan, Edmund K. Waller, Xue-Zhong Yu, Amer A. Beg
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Research Article Immunology

PKCθ is required for alloreactivity and GVHD but not for immune responses toward leukemia and infection in mice

Abstract

When used as therapy for hematopoietic malignancies, allogeneic BM transplantation (BMT) relies on the graft-versus-leukemia (GVL) effect to eradicate residual tumor cells through immunologic mechanisms. However, graft-versus-host disease (GVHD), which is initiated by alloreactive donor T cells that recognize mismatched major and/or minor histocompatibility antigens and cause severe damage to hematopoietic and epithelial tissues, is a potentially lethal complication of allogeneic BMT. To enhance the therapeutic potential of BMT, we sought to find therapeutic targets that could inhibit GVHD while preserving GVL and immune responses to infectious agents. We show here that T cell responses triggered in mice by either Listeria monocytogenes or administration of antigen and adjuvant were relatively well preserved in the absence of PKC isoform θ (PKCθ), a key regulator of TCR signaling. In contrast, PKCθ was required for alloreactivity and GVHD induction. Furthermore, absence of PKCθ raised the threshold for T cell activation, which selectively affected alloresponses. Most importantly, PKCθ-deficient T cells retained the ability to respond to virus infection and to induce GVL effect after BMT. These findings suggest PKCθ is a potentially unique therapeutic target required for GVHD induction but not for GVL or protective responses to infectious agents.

Authors

Javier O. Valenzuela, Cristina Iclozan, Mohammad S. Hossain, Martin Prlic, Emily Hopewell, Crystina C. Bronk, Junmei Wang, Esteban Celis, Robert W. Engelman, Bruce R. Blazar, Michael J. Bevan, Edmund K. Waller, Xue-Zhong Yu, Amer A. Beg

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Figure 2

Crucial role of NF-κB in mediating PKCθ-dependent and -independent responses.

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Crucial role of NF-κB in mediating PKCθ-dependent and -independent respo...
(A) 1 × 105 MACS-purified WT and PKCθ–/– OT-1 CD8 T cells were stimulated with 1 × 104 BM-derived DCs in the presence of increasing amounts of OVAp ranging from 0 μM to 10 μM, as indicated. (B) 1 × 105 MACS-purified WT and PKCθ–/– OT-1 CD8 T cells were activated with microspheres bearing DimerX H-2Kb:Ig (pulsed with 0.1 μM OVAp) and B7.1:Fc or (C) 1 × 104 BM-derived DCs in the presence of 1 μM OVAp. Untreated (UT) samples received no stimulation. NF-κB nuclear localization was measured at the indicated times by EMSA. Prior to generating EMSA extracts, DCs were removed by positive selection of CD11c+ cells on a MACS column. (D) 1 × 105 MACS-purified WT, PKCθ–/–, and p50–/–cRel–/– CD8 T cells were activated with 0.1 μg anti-TCR ± anti-CD28 agonistic antibodies (coated on plates) in the presence or absence of 1 × 104 BM-derived DCs. T cell proliferation was measured after 3 days by 3H-thymidine incorporation. (E) 1 × 105 MACS-purified WT and PKCθ–/– OT-1 CD8 T cells were activated with microspheres coated with H-2Kb/OVA plus B7.1:Fc as described above. After 24 and 48 hours of activation, the cells were infected with the GFP-expressing retroviral vectors MIG or MIG-IKK. GFP+ cells were sorted by FACS and restimulated with Kb/OVA/B7 microspheres for an additional 48 hours. Proliferation was measured by 3H-thymidine incorporation during the last 10 hours of restimulation. Error bars indicate SD in triplicate samples.