Synergistic activation of innate immunity by double-stranded RNA and CpG DNA promotes enhanced antitumor activity.
Double-stranded RNA (dsRNA) and unmethylated CpG sequences in DNA are pathogen-associated molecular patterns of viruses and bacteria that activate innate immunity. To examine whether dsRNA and CpG DNA could combine to provide enhanced stimulation of innate immune cells, murine macrophages were stimulated with poly-rI:rC (pIC), a dsRNA analog, and CpG-containing oligodeoxynucleotides (CpG-ODN). Combined treatments demonstrated synergy in nitric oxide, interleukin (IL)-12, tumor necrosis factor alpha, and IL-6 production. Studies using neutralizing antibodies for type I interferons (IFNs), IFN-alpha and IFN-beta, indicated that nitric oxide synthase synergism is mediated by paracrine/autocrine effects of IFN-beta. In contrast, enhanced cytokine production occurred independent of type I IFN and was maintained in macrophages from IFN-alpha/beta receptor knockout mice. Cotransfection of human Toll-like receptors 3 and 9 (receptors for dsRNA and CpG DNA, respectively) into 293T cells supported synergistic activation of an IL-8 promoter reporter construct by pIC, indicating interaction of the signaling pathways in driving the synergy response. In vivo stimulation of mice with pIC and CpG-ODN demonstrated synergy for serum IL-6 and IL-12p40 levels that correlated with an enhanced antitumor effect against established B16-F10 experimental pulmonary metastases. Treatment of tumor-bearing mice with pIC and CpG-ODN in combination resulted in enhanced nitric oxide synthase expression in lung tissue and enhanced up-regulation of class I major histocompatibility complex on splenic dendritic cells relative to treatments with either agent alone. In conclusion, the combined detection of viral pathogen-associated molecular patterns, i.e., dsRNA and CpG DNA, may mimic definitive viral recognition, resulting in an enhanced innate immune response that could be used for tumor vaccination or immunotherapy.
Department of Cancer Biology, Taussig Cancer Center, The Cleveland Clinic Foundation, Cleveland, Ohio 44195, USA.
Mark M
MM
Whitmore
Michael J
MJ
DeVeer
Andrea
A
Edling
Rhonda K
RK
Oates
Brenna
B
Simons
Daniel
D
Lindner
Bryan R G
BR
Williams
D000818:Animals; D002460:Cell Line; D018899:CpG Islands / Q000276:immunology; D004247:DNA / Q000276:immunology*; D004268:DNA-Binding Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D003713:Dendritic Cells / Q000276:immunology / Q000378:metabolism; D004789:Enzyme Activation; D006801:Humans; D007370:Interferon Type I / Q000276:immunology; D018664:Interleukin-12 / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D015850:Interleukin-6 / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D016209:Interleukin-8 / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D008168:Lung / Q000378:metabolism; D008262:Macrophage Activation / Q000276:immunology; D008264:Macrophages / Q000276:immunology* / Q000378:metabolism / Q000502:physiology; D008546:Melanoma, Experimental / Q000276:immunology; D008562:Membrane Glycoproteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D019001:Nitric Oxide Synthase / Q000096:biosynthesis / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II; D011070:Poly I-C / Q000494:pharmacology; D011401:Promoter Regions, Genetic; D012330:RNA, Double-Stranded / Q000276:immunology*; D011956:Receptors, Cell Surface / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D051217:Toll-Like Receptor 9; D051193:Toll-Like Receptors; D014162:Transfection; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology
D004268:DNA-Binding Proteins; D007370:Interferon Type I; D015850:Interleukin-6; D016209:Interleukin-8; D008562:Membrane Glycoproteins; D012330:RNA, Double-Stranded; D011956:Receptors, Cell Surface; C495371:TLR9 protein, human; C495370:Tlr9 protein, mouse; D051217:Toll-Like Receptor 9; D051193:Toll-Like Receptors; D014409:Tumor Necrosis Factor-alpha; D018664:Interleukin-12; D004247:DNA; C496319:NOS2 protein, human; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse; D011070:Poly I-C