Nitric oxide short-circuits interleukin-12-mediated tumor regression

Abstract

Interleukin-12 (IL-12) can promote tumor regression via activation of multiple lymphocytic and myelocytic eVectors. Whereas the cytotoxic mechanisms employed by T/NK/NKT cells in IL-12-mediated tumor kill are well deWned, the antitumor role of macrophage-produced cytotoxic metabolites has been more controversial. To this end, we investigated the speciWc role of nitric oxide (NO), a major macrophage eVector molecule, in post-IL-12 tumor regression. Analysis of tumors following a single intratumoral injection of slow-release IL-12 microspheres showed an IFN�-dependent sevenfold increase in inducible nitric oxide synthase (iNOS) expression within 48 h. Flow cytometric analysis of tumor-resident leukocytes and in vivo depletion studies identiWed CD11b+ F4/80+ Gr1lo macrophages as the primary source of iNOS. Blocking of post-therapy iNOS activity with N-nitro-L-arginine methyl ester (L-NAME)dramatically enhanced tumor suppression revealing the inhibitory eVect of NO on IL-12-driven antitumor immunity. Superior tumor regression in mice receiving combination treatment was associated with enhanced survival and proliferation of activated tumor-resident CD8+ T-eVector/memory cells (Tem). These Wndings demonstrate that macrophageproduced NO negatively regulates the antitumor activity of IL-12 via its detrimental eVects on CD8+ T cells and identify L-NAME as a potent adjuvant in IL-12 therapy of cancer. Abstract Interleukin-12 (IL-12) can promote tumor regression via activation of multiple lymphocytic and myelocytic eVectors. Whereas the cytotoxic mechanisms employed by T/NK/NKT cells in IL-12-mediated tumor kill are well deWned, the antitumor role of macrophage-produced cytotoxic metabolites has been more controversial. To this end, we investigated the speciWc role of nitric oxide (NO), a major macrophage eVector molecule, in post-IL-12 tumor regression. Analysis of tumors following a single intratumoral injection of slow-release IL-12 microspheres showed an IFN -dependent sevenfold increase in inducible nitric oxide synthase (iNOS) expression within 48 h. Flow cytometric analysis of tumor-resident leukocytes and in vivo depletion studies identiWed CD11b + F4/80 + Gr1 lo macrophages as the primary source of iNOS. Blocking of post-therapy iNOS activity with N-nitro-L-arginine methyl ester (L-NAME) dramatically enhanced tumor suppression revealing the inhibitory eVect of NO on IL-12-driven antitumor immunity. Superior tumor regression in mice receiving combination treatment was associated with enhanced survival and proliferation of activated tumor-resident CD8+ T-eVector/memory cells (Tem). These Wndings demonstrate that macrophageproduced NO negatively regulates the antitumor activity of IL-12 via its detrimental eVects on CD8+ T cells and identify L-NAME as a potent adjuvant in IL-12 therapy of cancer.

Tolerogenic Phenotype of IFN-γ–Induced IDO+ Dendritic Cells Is Maintained via an Autocrine IDO–Kynurenine/AhR–IDO Loop

“…Induction of IL-10 (3), inducible NO synthase (4), and, more recently, regulatory T cell (Treg) expansion (5) have been identified as potential mechanisms that limit the duration of IL-12-driven cytotoxic T cell activity. Whereas production of IL-10 and inducible NO synthase are rapid but transient events (6,7), post-IL-12 Treg rebound is progressive and long-lasting (5). Recent work revealed that IFN-g, which is essential to the antitumor activity of IL-12, also induced IDO + dendritic cells (DC) in the tumor and the tumor-draining lymph node (TDLN), and that IDO + DC orchestrated the post-IL-12 Treg expansion (8).…”

Effect of complement C5a in tumor progression and the local tumor microenvironment.

“…Also, NO is a major macrophage effector molecule stimulated to be released under inflammatory conditions. Macrophage NO can be cytotoxic to tumor cells; however, NO produced by the infiltrating macrophage can also be cytotoxic to anti-tumor lymphocytes [72]. Immune inhibition by iNOS was found to be stronger than tumor cell cytotoxic effector functions; therefore macrophage NO had dramatic negative effects on the survival of anti-tumor CD8+ T cells in a murine tumor model.…”

“…TNF-a release may be restricted for extended amounts of time [72]. NO is a lipophilic molecule and easily migrates across cell membranes to act intracellularly, and signaling and post-translational modifications lead to anti-apoptosis and increased cell growth of recipient cells [72]. NO is a shortlived molecule; however, iNOS can induce sustained levels of NO, expanding the duration and levels of NO in the environment [57].…”

“…TNF-a release may be restricted for extended amounts of time [72]. NO is a lipophilic molecule and easily migrates across cell membranes to act intracellularly, and signaling and post-translational modifications lead to anti-apoptosis and increased cell growth of recipient cells [72].…”

“…Immune inhibition by iNOS was found to be stronger than tumor cell cytotoxic effector functions; therefore macrophage NO had dramatic negative effects on the survival of anti-tumor CD8+ T cells in a murine tumor model. SKOV -3 CV and WT potentially generated NO, through elevated levels of iNOS, may enhance tumor growth by promoting angiogenesis and invasion or the preferential destruction of anti-tumor immune cells [72].…”