A Study of T Cell Tolerance to the Tumor-Associated Antigen MDM2: Cytokines Can Restore Antigen Responsiveness, but Not High Avidity T Cell Function

BACKGROUND: Most tumor-associated antigens (TAA) currently used for immunotherapy of cancer are also expressed in normal tissues, which may induce tolerance and impair T cell-mediated immunity. However, there is limited information about how physiological expression in normal tissues alters the function of TAA-specific T cells. METHODOLOGY/PRINCIPAL FINDINGS: We used a T cell receptor transgenic model to study how MDM2 expression in normal tissues affects the function of T cells specific for this TAA that is found at high levels in many different types of tumors. We found that some MDM2-specific T cells escaped thymic deletion and persisted in the peripheral T cell pool. When stimulated with antigen, these T cells readily initiated cell division but failed to proliferate and expand, which was associated with a high rate of apoptosis. Both IL-2 and IL-15 efficiently rescued T cell survival and antigen-specific T cell proliferation, while IL-7 and IL-21 were ineffective. Antigen-stimulated T cells showed impaired expression of the effector molecules CD43, granzyme-B and IFN-γ, a defect that was completely restored when T cells were stimulated in the presence of IL-2. In contrast, IL-15 and IL-21 only restored the expression of CD43 and granzyme-B, but not IFN-γ production. Finally, peptide titration experiments with IL-2 rescued T cells indicated that they were of lower avidity than non-tolerant control T cells expressing the same TCR. CONCLUSIONS/SIGNIFICANCE: These data indicate that cytokines can rescue the antigen-specific proliferation and effector function of MDM2-specific T cells, although this does not lead to the recovery of high avidity T cell function. This study sheds light on possible limitations of immunotherapy approaches that target widely expressed TAA, such as MDM2

Nanomolar-potency small molecule inhibitor of STAT5 protein

© 2014 American Chemical Society. We herein report the design and synthesis of the first nanomolar binding inhibitor of STAT5 protein. Lead compound 13a, possessing a phosphotyrosyl-mimicking salicylic acid group, potently and selectively binds to STAT5 over STAT3, inhibits STAT5-SH2 domain complexation events in vitro, silences activated STAT5 in leukemic cells, as well as STAT5's downstream transcriptional targets, including MYC and MCL1, and, as a result, leads to apoptosis. We believe 13a represents a useful probe for interrogating STAT5 function in cells as well as being a potential candidate for advanced preclinical trials

Synthesis and structure-activity relationships of 3,5-disubstituted-pyrrolo[2,3-b]pyridines as inhibitors of adaptor-sssociated kinase 1 with antiviral activity

There are currently no approved drugs for the treatment of emerging viral infections, such as dengue and Ebola. Adaptor-associated kinase 1 (AAK1) is a cellular serine-threonine protein kinase that functions as a key regulator of the clathrin-associated host adaptor proteins and regulates the intracellular trafficking of multiple unrelated RNA viruses. Moreover, AAK1 is overexpressed specifically in dengue virus-infected but not bystander cells. Because AAK1 is a promising antiviral drug target, we have embarked on an optimization campaign of a previously identified 7-azaindole analogue, yielding novel pyrrolo[2,3- b]pyridines with high AAK1 affinity. The optimized compounds demonstrate improved activity against dengue virus both in vitro and in human primary dendritic cells and the unrelated Ebola virus. These findings demonstrate that targeting cellular AAK1 may represent a promising broad-spectrum antiviral strategy

Anticancer kinase inhibitors impair intracellular viral trafficking and exert broad-spectrum antiviral effects

Global health is threatened by emerging viral infections, which largely lack effective vaccines or therapies. Targeting host pathways that are exploited by multiple viruses could offer broad-spectrum solutions. We previously reported that AAK1 and GAK, kinase regulators of the host adaptor proteins AP1 and AP2, are essential for hepatitis C virus (HCV) infection, but the underlying mechanism and relevance to other viruses or in vivo infections remained unknown. Here, we have discovered that AP1 and AP2 cotraffic with HCV particles in live cells. Moreover, we found that multiple viruses, including dengue and Ebola, exploit AAK1 and GAK during entry and infectious virus production. In cultured cells, treatment with sunitinib and erlotinib, approved anticancer drugs that inhibit AAK1 or GAK activity, or with more selective compounds inhibited intracellular trafficking of HCV and multiple unrelated RNA viruses with a high barrier to resistance. In murine models of dengue and Ebola infection, sunitinib/erlotinib combination protected against morbidity and mortality. We validated sunitinib- and erlotinib-mediated inhibition of AAK1 and GAK activity as an important mechanism of antiviral action. Additionally, we revealed potential roles for additional kinase targets. These findings advance our understanding of virus-host interactions and establish a proof of principle for a repurposed, host-targeted approach to combat emerging viruses.status: publishe

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

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