Several quassinoids were identified in a high-throughput screening assay as inhibitors of the transcription factor AP-1. Further biological characterization revealed that while their effect was not specific to AP-1, protein synthesis inhibition and cell growth assays were inconsistent with a mechanism of simple protein synthesis inhibition. Numerous plant extracts from the plant family Simaroubaceae were also identified in the same screen; bioassay-guided fractionation of one extract (Ailanthus triphylla) yielded two known quassinoids, ailanthinone (3) and glaucarubinone (4), which were also identified in the pure compound screening procedure.The quassinoids are a group of several hundred complex nortriterpenoids originally isolated from plants in the family Simaroubaceae on the basis of their bitter taste.1 A number of quassinoids have been found to have antileukemic and other anticancer activity,2 -5 most notably bruceantin, which advanced to phase II clinical trials in breast cancer and melanoma before failing due to toxicity (hypotension, nausea, vomiting and fever) and poor efficacy. 6, 7 Although the first pharmacologic studies of quassinoids identified inhibition of protein synthesis as a probable mechanism of action, recent workers have proposed a number of other mechanisms to explain the cell growth inhibition caused by these compounds.8 Among these are inhibition of plasma membrane NADH oxidase activity, induced differentiation of leukemia and lymphoma cells via c-myc down-regulation and G 1 arrest, 8,9 and mitochondrial membrane depolarization with caspase-3 activation. 9,10 Quassinoids have also shown activity in antimalarial assays, and this activity has been correlated with inhibition of protein synthesis in the parasite. 11 Quassinoids comprise a relatively homogeneous group of structures with a common tetracyclic skeleton, which can be categorized by the presence or absence of a bridging ether oxygen from carbon 17, and whether that bridge is connected to carbon 11 or 13. † Dedicated to Dr. David G.I. Kingston of Virginia Polytechnic Institute and State University for his pioneering work on bioactive natural products.*Author to whom correspondence should be addressed. Tel.: 301-8546-1942. Fax: 301-846-6177. beutlerj@mail We have conducted a high-throughput screening assay for inhibitors of the transcription factor activating protein-1 (AP-1). 12 Among the confirmed hits in the screen were two purified quassinoids, as well as numerous extracts and their fractions from the Simaroubaceae. This contribution reports the evaluation of a set of quassinoids for specificity against the AP-1 target, the structure-activity requirements for such activity, as well as their patterns of cell growth inhibition in the NCI 60-cell screen and their effect on protein synthesis.The two pure compounds initially identified in the AP-1 screen were 6α-tigloyloxyglaucarubol (1, NSC#374763) 13 and 15-methylcarbamoyl-bruceolide (2, NSC#377097), 14 however, samples of these compounds were not available for further te...