We have recently shown that covalent attachment of the NO moiety to the HIV protease inhibitor Saquinavir (Saq) produced a qualitatively new chemical entity, named Saquinavir-NO (Saq-NO), with enhanced anticancer properties and reduced toxicity. In this study we evaluated the impact of Saq-NO on the growth of A375 human melanoma cells, as a prototype of NO-dependent cancer model. The novel compound strongly affected the in vitro and in vivo progression of A375 melanoma cell growth. The mechanism of antimelanoma action comprised dual drug activity—induction of apoptotic cell death and acquisition of melanoma cell responsiveness to TRAIL. Saq-NO-triggered apoptosis was dependent on transient AKT up-regulation and reduced pERK and iNOS expression that were observed within the first 12 h of exposure to the drug. Thereafter, however, Saq-NO up-regulated both iNOS transcription and NO endogenous synthesis and sensitized A375 cells to TRAIL. Furthermore, reduced YY1 expression was observed after 24 h of Saq-NO exposure, which correlated with increased expression of DR5. The biological relevance of this complex and powerful action of Saq-NO was consistent with the marked drug-induced inhibition of the growth of A375 xenotransplants in nude mice. J. Cell. Physiol. 226: 1803–1812, 2011. © 2010 Wiley-Liss, Inc.