Spermidine/spermine-N1-acetyltransferase (SSAT) is a mitochondrial-localized enzyme that is highly inducible and tightly controlled and is the rate-limiting enzyme in polyamine catabolism. It is known that SSAT is induced when polyamine level increases. Although multiple mechanisms have been implicated, translational control is thought to be paramount. Previous studies with transgenic and knockout mice suggested that for certain human conditions, the modulation of SSAT levels could offer therapeutic benefits. Besides polyamines and their analogs, certain stimuli can increase SSAT levels, suggesting that the development of reporters for high throughput screening can lead to the identification of novel pharmacophores that can modulate SSAT translation. Here we report the development and validation of a luciferase-based biosensor system for the identification of compounds that are able to either promote or prevent the translation of SSAT. The system uses HEK293T cells transfected with a construct composed of SSAT mRNA modified to lack upstream open reading frame (uORF) function, is mutated to reduce translational repression and is linked with luciferase. As a proof of principle of the utility of the SSAT translation sensor, we screened the Prestwick drug library (1,200 FDA Approved compounds). The library contained 15 compounds that activated SSAT translation by at least 40% more than the basal expression, but none exceeded the positive control N1, N11-diethylnorspermine. On the other hand, 38 compounds were found to strongly inhibit SSAT translation. We conclude that this biosensor can lead to the identification of novel pharmacophores that are able to modulate the translation of SSAT.