ARTICLE ABSTRACTRetinoid X receptor alpha (RXRα) and its N-terminally truncated version, tRXRα, are widely implicated in cancer development and represent intriguing targets for cancer prevention and treatment. Successful manipulation of RXRα and tRXRα requires the identification of their modulators that could produce therapeutic effects. Here, we report that a class of nitrostyrene derivatives bind to RXRα by a unique mechanism, of which the nitro group of nitrostyrene derivatives and Cys432 of RXRα are required for binding. The binding results in the potent activation of Gal4-DBD-RXRα-LBD transactivation. However, the binding inhibits the transactivation of RXRα homodimer, which might be due to the distinct conformation of RXRα homodimer induced by these nitrostyrene derivatives. Two RXRα point mutants with Cys432 substituted with Tyr and Trp, respectively, could mimic the bindings of two nitrostyrene derivatives and have the ability of autotransactivation. In studying the functional consequences of the binding, we show that these nitrostyrene derivatives could potently inhibit the TNFα/NFκB signaling pathway in a tRXRα-dependent manner. tRXRα promotes TNFα-induced NF-κB activation through its interaction with TRAF2 and enhances TNFα-induced ubiquitination of RIP1, which is strongly inhibited by nitrostyrene derivatives. The inhibition of TNFα-induced NF-κB activation results in the synergistic effect of the combination of nitrostyrene derivatives and TNFα on the induction of cancer cell apoptosis. Together, our results show a new class of RXRα modulators that induce apoptosis of cancer cells through their unique binding mode and new mechanism of action. Cancer Res; 75(10); 2049–60. ©2015 AACR.