Purpose: Programmed death ligand 1 (PD-L1) is expressed on antigen-presenting cells and inhibits activation of T cells through its receptor PD-1. PD-L1 is aberrantly expressed on some epithelial malignancies and Hodgkin lymphomas and may prevent effective host antitumor immunity. The role of PD-L1 in non-Hodgkin lymphomas (NHL) is not well characterized.Experimental Design: PD-L1 expression was analyzed in cell lines and lymphoma specimens by using flow cytometry and immunohistochemistry. Functional activity of PD-L1 was studied by incubating irradiated lymphoma cells with allogeneic T cells with or without anti-PD-L1 blocking antibody; T-cell proliferation and IFN-g secretion served as measures of T-cell activation. Similar experiments were conducted using cultures of primary lymphoma specimens containing host T cells.Results: PD-L1 was expressed uniformly by anaplastic large cell lymphoma (ALCL) cell lines, but rarely in B-cell NHL, confined to a subset of diffuse large B-cell lymphomas (DLBCL) with activated B-cell features (3 of 28 cell lines and 24% of primary DLBCL). Anti-PD-L1 blocking antibody boosted proliferation and IFN-g secretion by allogeneic T cells responding to ALCL and DLBCL cells. In autologous cultures of primary ALCL and DLBCL, PD-L1 blockade enhanced secretion of inflammatory cytokines IFN-g, granulocyte macrophage colony-stimulating factor, interleukin (IL)-1, IL-6, IL-8, IL-13, TNF-a, and macrophage inflammatory protein-1a. In establishing cell lines from an aggressive PD-L1þ mature Bcell lymphoma, we also noted that PD-L1 expression could be lost under certain in vitro culture conditions. Conclusions: PD-L1 may thwart effective antitumor immune responses and represents an attractive target for lymphoma immunotherapy. Clin Cancer Res; 17(13); 4232-44. Ó2011 AACR.

Mitogen-activated protein kinase (MAPK) cascades have pivotal roles in plant innate immunity. However, downstream signaling of plant defense-related MAPKs is not well understood. Here, we provide evidence that the Nicotiana benthamiana WRKY8 transcription factor is a physiological substrate of SIPK, NTF4, and WIPK. Clustered Pro-directed Ser residues (SP cluster), which are conserved in group I WRKY proteins, in the N-terminal region of WRKY8 were phosphorylated by these MAPKs in vitro. Antiphosphopeptide antibodies indicated that Ser residues in the SP cluster of WRKY8 are phosphorylated by SIPK, NTF4, and WIPK in vivo. The interaction of WRKY8 with MAPKs depended on its D domain, which is a MAPKinteracting motif, and this interaction was required for effective phosphorylation of WRKY8 in plants. Phosphorylation of WRKY8 increased its DNA binding activity to the cognate W-box sequence. The phospho-mimicking mutant of WRKY8 showed higher transactivation activity, and its ectopic expression induced defense-related genes, such as 3-hydroxy-3-methylglutaryl CoA reductase 2 and NADP-malic enzyme. By contrast, silencing of WRKY8 decreased the expression of defense-related genes and increased disease susceptibility to the pathogens Phytophthora infestans and Colletotrichum orbiculare. Thus, MAPK-mediated phosphorylation of WRKY8 has an important role in the defense response through activation of downstream genes.

Purpose: The growth of non–Hodgkin lymphomas can be influenced by tumor–immune system interactions. Cytotoxic T-lymphocyte antigen 4 (CTLA-4) is a negative regulator of T-cell activation that serves to dampen antitumor immune responses. Blocking anti–CTLA-4 monoclonal antibodies improves host resistance to immunogenic tumors, and the anti–CTLA-4 antibody ipilimumab (MDX-010) has clinical activity against melanoma, prostate, and ovarian cancers. Experimental Design: We did a phase I trial of ipilimumab in patients with relapsed/refractory B-cell lymphoma to evaluate safety, immunologic activity, and potential clinical efficacy. Treatment consisted of ipilimumab at 3 mg/kg and then monthly at 1 mg/kg × 3 months (dose level 1), with subsequent escalation to 3 mg/kg monthly × 4 months (dose level 2). Results: Eighteen patients were treated, 12 at the lower dose level and 6 at the higher dose level. Ipilimumab was generally well tolerated, with common adverse events attributed to it, including diarrhea, headache, abdominal pain, anorexia, fatigue, neutropenia, and thrombocytopenia. Two patients had clinical responses; one patient with diffuse large B-cell lymphoma had an ongoing complete response (>31 months), and one with follicular lymphoma had a partial response lasting 19 months. In 5 of 16 cases tested (31%), T-cell proliferation to recall antigens was significantly increased (>2-fold) after ipilimumab therapy. Conclusions: Blockade of CTLA-4 signaling with the use of ipilimumab is well tolerated at the doses used and has antitumor activity in patients with B-cell lymphoma. Further evaluation of ipilimumab alone or in combination with other agents in B-cell lymphoma patients is therefore warranted. (Clin Cancer Res 2009;15(20):6446–53)