TNFalpha is a prominent proinflammatory cytokine and a critical mediator for the development of many types of cancer such as breast, colon, prostate, cervical, skin, liver, and chronic lymphocytic leukemia. Binding of TNFalpha to TNFR1 can lead to divergent signaling pathways promoting predominantly NF-kappaB activation but also cell death. We report here that the nitric oxide (NO) donor glyceryl trinitrate (GTN) converts TNFalpha, generated from immune cells or cancer cells stimulated by chemotherapy, into a prodeath mediator in colon and mammary cancer cells. GTN-mediated S-nitrosylation of cIAP1 on cysteines 571 and 574 inhibited its E3 ubiquitin ligase activity, which in turn reduced Lys63-linked ubiquitination of RIP1 and initiated assembly of a death complex. These findings provide insights into how NO can harness advantageous aspects of inflammation in cancer and provide new therapeutic strategies.Significance: Combination of an NO donor with chemotherapeutic drug-induced TNFalpha represents a potentially valuable anticancer strategy. Cancer Res; 78(8); 1948-57. (c)2018 AACR.