Oncolytic viruses can exert their antitumor activity via direct oncolysis or activation of antitumor immunity. Although reovirus is currently under clinical investigation for the treatment of localized or disseminated cancer, any potential immune contribution to its efficacy has not been addressed. This is the first study to investigate the ability of reovirus to activate human dendritic cells (DC), key regulators of both innate and adaptive immune responses. Reovirus induced DC maturation and stimulated the production of the proinflammatory cytokines IFN-α, TNF-α, IL-12p70, and IL-6. Activation of DC by reovirus was not dependent on viral replication, while cytokine production (but not phenotypic maturation) was inhibited by blockade of PKR and NF-κB signaling. Upon coculture with autologous NK cells, reovirus-activated DC up-regulated IFN-γ production and increased NK cytolytic activity. Moreover, short-term coculture of reovirus-activated DC with autologous T cells also enhanced T cell cytokine secretion (IL-2 and IFN-γ) and induced non-Ag restricted tumor cell killing. These data demonstrate for the first time that reovirus directly activates human DC and that reovirus-activated DC stimulate innate killing by not only NK cells, but also T cells, suggesting a novel potential role for T cells in oncolytic virus-induced local tumor cell death. Hence reovirus recognition by DC may trigger innate effector mechanisms to complement the virus’s direct cytotoxicity, potentially enhancing the efficacy of reovirus as a therapeutic agent.

Purpose Reovirus is a naturally occurring oncolytic virus in clinical trials. Although tumor infection by reovirus can generate adaptive antitumor immunity, its therapeutic importance versus direct viral oncolysis is undefined. This study addresses the requirement for viral oncolysis and replication, and the relative importance of antitumor immunity and direct oncolysis in therapy. Experimental Design Nonantigen specific T cells loaded with reovirus were delivered i.v. to C57BL/6 and severe combined immunodeficient mice bearing lymph node and splenic metastases from the murine melanoma, B16ova, with assessment of viral replication, metastatic clearance by tumor colony outgrowth, and immune priming. Human cytotoxic lymphocyte priming assays were done with dendritic cells loaded with Mel888 cells before the addition of reovirus. Results B16ova was resistant to direct oncolysis in vitro, and failed to support reovirus replication in vitro or in vivo. Nevertheless, reovirus purged lymph node and splenic metastases in C57BL/6 mice and generated antitumor immunity. In contrast, reovirus failed to reduce tumor burden in severe combined immunodeficient mice bearing either B16ova or reovirus-sensitive B16tk metastases. In the human system, reovirus acted solely as an adjuvant when added to dendritic cells already loaded with Mel888, supporting priming of specific antitumor cytotoxic lymphocyte, in the absence of significant direct tumor oncolysis; UV-treated nonreplicating reovirus was similarly immunogenic. Conclusion The immune response is critical in mediating the efficacy of reovirus, and does not depend upon direct viral oncolysis or replication. The findings are of direct relevance to fulfilling the potential of this novel anticancer agent.

Oncolytic viruses (OV) are promising treatments for cancer, with several currently undergoing testing in randomised clinical trials. Measles virus (MV) has not yet been tested in models of human melanoma. This study demonstrates the efficacy of MV against human melanoma. It is increasingly recognised that an essential component of therapy with OV is the recruitment of host anti-tumour immune responses, both innate and adaptive. MV-mediated melanoma cell death is an inflammatory process, causing the release of inflammatory cytokines including type-1 interferons and the potent danger signal HMGB1. Here, using human in vitro models, we demonstrate that MV enhances innate antitumour activity, and that MV-mediated melanoma cell death is capable of stimulating a melanoma-specific adaptive immune response.

Membrane-presented CD40 agonists can induce apoptosis in carcinoma, but not normal homologous epithelial cells, whereas soluble agonists are growth inhibitory but not proapoptotic unless protein synthesis is blocked. Here we demonstrate that membrane-presented CD40 ligand (CD154) (mCD40L), but not soluble agonists, triggers cell death in malignant human urothelial cells via a direct mechanism involving rapid upregulation of TNFR-associated factor (TRAF)3 protein, without concomitant upregulation of TRAF3 mRNA, followed by activation of the c-Jun N-terminal kinase (JNK)/activator protein-1 (AP-1) pathway and induction of the caspase-9/caspase-3-associated intrinsic apoptotic machinery. TRAF3 knockdown abrogated JNK/AP-1 activation and prevented CD40-mediated apoptosis, whereas restoration of CD40 expression in CD40-negative carcinoma cells restored apoptotic susceptibility via the TRAF3/AP-1-dependent mechanism. In normal human urothelial cells, mCD40L did not trigger apoptosis, but induced rapid downregulation of TRAF2 and 3, thereby paralleling the situation in Blymphocytes. Thus, TRAF3 stabilization, JNK activation and caspase-9 induction define a novel pathway of CD40-mediated apoptosis in carcinoma cells.