Apo2 ligand or tumour necrosis factor-related apoptosisinducing ligand (Apo2L/TRAIL) is one of the several members of the tumour necrosis factor (TNF) gene superfamily that induce apoptosis through engagement of death receptors (DRs). Apo2L/TRAIL interacts with an unusually complex receptor system of two DRs and three decoys. This protein has garnered intense interest as a potential candidate for cancer therapy because as a trimer it selectively induces apoptosis in many transformed cells but not in normal cells. While much of the early characterisation of Apo2L/TRAIL and its receptors relied on overexpression studies, recent work using untransfected cells has clarified how endogenous proteins transmit apoptotic signals from this ligand. In this review, we focus on the apoptotic signalling pathways stimulated by Apo2L/TRAIL and summarise what is known about its physiological role.

The programmed death-1 (PD-1) receptor serves as an immunologic checkpoint, limiting bystander tissue damage and preventing the development of autoimmunity during inflammatory responses. PD-1 is expressed by activated T cells and downmodulates T-cell effector functions upon binding to its ligands, PD-L1 and PD-L2, on antigen-presenting cells. In patients with cancer, the expression of PD-1 on tumor-infiltrating lymphocytes and its interaction with the ligands on tumor and immune cells in the tumor microenvironment undermine antitumor immunity and support its rationale for PD-1 blockade in cancer immunotherapy. This report details the development and characterization of nivolumab, a fully human IgG4 (S228P) anti-PD-1 receptor-blocking monoclonal antibody. Nivolumab binds to PD-1 with high affinity and specificity, and effectively inhibits the interaction between PD-1 and its ligands. In vitro assays demonstrated the ability of nivolumab to potently enhance T-cell responses and cytokine production in the mixed lymphocyte reaction and superantigen or cytomegalovirus stimulation assays. No in vitro antibody-dependent cell-mediated or complement-dependent cytotoxicity was observed with the use of nivolumab and activated T cells as targets. Nivolumab treatment did not induce adverse immune-related events when given to cynomolgus macaques at high concentrations, independent of circulating anti-nivolumab antibodies where observed. These data provide a comprehensive preclinical characterization of nivolumab, for which antitumor activity and safety have been demonstrated in human clinical trials in various solid tumors. Cancer Immunol Res; 2(9); 846-56. Ó2014 AACR.

The importance of Bax for induction of tumor apoptosis through death receptors remains unclear. Here we show that Bax can be essential for death receptor--mediated apoptosis in cancer cells. Bax-deficient human colon carcinoma cells were resistant to death-receptor ligands, whereas Bax-expressing sister clones were sensitive. Bax was dispensable for apical death-receptor signaling events including caspase-8 activation, but crucial for mitochondrial changes and downstream caspase activation. Treatment of colon tumor cells deficient in DNA mismatch repair with the death-receptor ligand apo2 ligand (Apo2L)/tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) selected in vitro or in vivo for refractory subclones with Bax frameshift mutations including deletions at a novel site. Chemotherapeutic agents upregulated expression of the Apo2L/TRAIL receptor DR5 and the Bax homolog Bak in Baxminus sign/minus sign cells, and restored Apo2L/TRAIL sensitivity in vitro and in vivo. Thus, Bax mutation in mismatch repair--deficient tumors can cause resistance to death receptor--targeted therapy, but pre-exposure to chemotherapy rescues tumor sensitivity.

Caspase-8 is believed to play an obligatory role in apoptosis initiation by death receptors, but the role of its structural relative, caspase-10, remains controversial. Although earlier evidence implicated caspase-10 in apoptosis signaling by CD95L and Apo2L/TRAIL, recent studies indicated that these death receptor ligands recruit caspase-8 but not caspase-10 to their deathinducing signaling complex (DISC) even in presence of abundant caspase-10. We characterized a series of caspase-10-specific antibodies and found that certain commercially available antibodies cross-react with HSP60, shedding new light on previous results. The majority of 55 lung and breast carcinoma cell lines expressed mRNA for both caspase-8 and -10; however, immunoblot analysis revealed that caspase-10 protein expression was more frequently absent than that of caspase-8, suggesting a possible selective pressure against caspase-10 production in cancer cells. In nontransfected cells expressing both caspases, CD95L and Apo2L/TRAIL recruited endogenous caspase-10 as well as caspase-8 to their DISC, where both enzymes were proteolytically processed with similar kinetics. Caspase-10 recruitment required the adaptor FADD/ Mort1, and caspase-10 cleavage in vitro required DISC assembly, consistent with the processing of an apoptosis initiator. Cells expressing only one of the caspases underwent ligand-induced apoptosis, indicating that each caspase can initiate apoptosis independently of the other. Thus, apoptosis signaling by death receptors involves not only caspase-8 but also caspase-10, and both caspases may have equally important roles in apoptosis initiation.

Innate immunity is the first line of defense against infection, protecting the host during the development of adaptive immunity and critically affecting the nature of the adaptive response. We show that, in contrast to tumor necrosis factor alpha (TNF-alpha), the related protein TWEAK attenuates the transition from innate to adaptive mechanisms. TWEAK-/- mice had overabundant natural killer (NK) cells and displayed hypersensitivity to bacterial endotoxin, with their innate immune cells producing excess interferon (IFN)-gamma and interleukin (IL)-12. TWEAK inhibited stimulation of the transcriptional activator STAT-1 and induced p65 nuclear factor (NF)-kappaB association with histone deacetylase 1, repressing cytokine production. TWEAK-/- mice developed oversized spleens with expanded memory and T helper 1 (TH1) subtype cells upon aging and mounted stronger innate and adaptive TH1-based responses against tumor challenge. Thus, TWEAK suppresses production of IFN-gamma and IL-12, curtailing the innate response and its transition to adaptive TH1 immunity.