Blocking oncogenic RAS enhances tumour cell surface MHC class I expression but does not alter susceptibility to cytotoxic lymphocytes.

Mutations in the RAS family of oncogenes are highly prevalent in human cancer and, amongst its manifold effects, oncogenic RAS impairs the expression of components of the antigen presentation pathway. This allows evasion of cytotoxic T lymphocytes (CTL). CTL and natural killer (NK) cells are reciprocally regulated by MHC class I molecules and any gain in CTL recognition obtained by therapeutic inactivation of oncogenic RAS may be offset by reduced NK cell activation. We have investigated the consequences of targeted inactivation of oncogenic RAS on the recognition by both CTL and NK cells. Inactivation of oncogenic RAS, either by genetic deletion or inactivation with an inducible intracellular domain antibody (iDAb), increased MHC class I expression in human colorectal cell lines. The common RAS mutations, at codons 12, 13 and 61, all inhibited antigen presentation. Although MHC class I modulates the activity of both CTL and NK cells, the enhanced MHC class I expression resulting from inactivation of mutant KRAS did not significantly affect the in vitro recognition of these cell lines by either class of cytotoxic lymphocyte. These results show that oncogenic RAS and its downstream signalling pathways modulate the antigen presentation pathway and that this inhibition is reversible. However, the magnitude of these effects was not sufficient to alter the in vitro recognition of tumour cell lines by either CTL or NK cells.

Reciprocal human dendritic cell-natural killer cell interactions induce antitumor activity following tumor cell infection by oncolytic reovirus.

Oncolytic virotherapy may mediate antitumor effects via direct oncolysis or immune-mediated tumor regression. Although the ability of oncolytic viruses to generate adaptive antitumor immunity has been characterized, their interactions with the innate immune system are relatively unclear. Using a human in vitro system, this study investigates the innate immunological consequences of reovirus therapy and its potential to activate NK cell-mediated antitumor activity. Dendritic cells (DC) loaded with reovirus-infected human melanoma Mel888 cells (DC-MelReo), but not reovirus-infected tumor cells alone, induced IFN-gamma production within the NK cell population upon coculture with PBMC, in a cell-to-cell contact-dependent manner. DC-MelReo secreted the chemokines CCL2, 3, 4, 5, 7, 8, 11, and CXCL10; these culture supernatants induced NK cell chemotaxis. Coculture of DC-MelReo with purified NK cells induced reciprocal contact-dependent phenotypic DC maturation, while DC-MelReo elicited up-regulation of the activation marker CD69 on NK cells, in a partially contact and partially IL-12 dependent manner. Significantly, DC-MelReo induced NK cell cytotoxicity toward tumor cells by a type I IFN dependent mechanism. These data demonstrate that tumor infection by reovirus can act via DC to induce NK cell recruitment, activation, and cytotoxicity, along with reciprocal DC maturation. These findings suggest that reciprocal DC-NK cell interactions, following reovirus therapy, may play an important role in altering the immune milieu of the tumor microenvironment and mediating tumor regression.

Tumor infection by oncolytic reovirus primes adaptive antitumor immunity.

PURPOSE: Early clinical trials are under way exploring the direct oncolytic potential of reovirus. This study addresses whether tumor infection by reovirus is also able to generate bystander, adaptive antitumor immunity. EXPERIMENTAL DESIGN: Reovirus was delivered intravenously to C57BL/6 mice bearing lymph node metastases from the murine melanoma, B16-tk, with assessment of nodal metastatic clearance, priming of antitumor immunity against the tumor-associated antigen tyrosinase-related protein-2, and cytokine responses. In an in vitro human system, the effect of reovirus infection on the ability of Mel888 melanoma cells to activate and load dendritic cells for cytotoxic lymphocyte (CTL) priming was investigated. RESULTS: In the murine model, a single intravenous dose of reovirus reduced metastatic lymph node burden and induced antitumor immunity (splenocyte response to tyrosinase-related protein-2 and interleukin-12 production in disaggregated lymph nodes). In vitro human assays revealed that uninfected Mel888 cells failed to induce dendritic cell maturation or support priming of an anti-Mel888 CTL response. In contrast, reovirus-infected Mel888 cells (reo-Mel) matured dendritic cells in a reovirus dose-dependent manner. When cultured with autologous peripheral blood lymphocytes, dendritic cells loaded with reo-Mel induced lymphocyte expansion, IFN-gamma production, specific anti-Mel888 cell cytotoxicity, and cross-primed CD8+ T cells specific against the human tumor-associated antigen MART-1. CONCLUSION: Reovirus infection of tumor cells reduces metastatic disease burden and primes antitumor immunity. Future clinical trials should be designed to explore both direct cytotoxic and immunotherapeutic effects of reovirus.