Double-stranded RNA directly acts on fibroblast and myeloid lineages to induce necroptosis as in TNFα. Here, we investigated whether this type of cell death occurred in
cancer cells in response to
polyinosinic-polycytidylic acid (polyI:C) and the pan-
caspase inhibitor z-Val-
Ala-Asp fluromethyl
ketone (zVAD). We found that the
colon cancer cell line CT26 is highly susceptible to necroptosis, as revealed by staining with
annexin V/
propidium iodide. CT26 cells possess
RNA sensors, TLR3 and MDA5, which are upregulated by
interferon (IFN)-inducing pathways and linked to receptor-interacting
protein kinase (RIP) 1/3 activation via TICAM-1 or MAVS adaptor, respectively. Although exogenously added polyI:C alone marginally induced necroptosis in CT26 cells, a combined regimen of polyI:C and zVAD induced approximately 50% CT26 necroptosis in vitro without secondary effects of TNFα or type I IFNs. CT26 necroptosis depended on the TLR3-TICAM-1-RIP3 axis in the
tumor cells to produce
reactive oxygen species, but not on MDA5, MAVS, or the
caspases/
inflammasome activation. However, the
RNA-derived necroptosis was barely reproduced in vivo in a CT26
tumor-implanted Balb/c mouse model with administration of polyI:C + zVAD. Significant shrinkage of CT26
tumors was revealed only when polyI:C (100 μg) was injected intraperitoneally and zVAD (1 mg) subcutaneously into
tumor-bearing mice that were depleted of cytotoxic T lymphocytes and natural killer cells. The results were confirmed with immune-compromised mice with no lymphocytes. Although necroptosis-induced
tumor growth retardation appears mechanistically complicated and dependent on the injection routes of polyI:C and zVAD, anti-
caspase reagent directed to
tumor cells will make
RNA adjuvant
immunotherapy more effective by modulating the formation of the tumoricidal microenvironment and dendritic cell-inducing antitumor immune system.