Thiol reagents were shown to act as potent inhibitors of L5178-Y murine leukemia cell proliferation. A series of aryl maleimides (AMI) was synthesized and evaluated theoretically for global and local reactivity, showing their selectivity for thiol groups, due to a reaction of the vinyl moiety (a soft acid) with thiols (a soft base). Two AMI that are benzoic acid derivatives (1f and 1h) were tested with an in vitro and ex vivo model to evaluate their reactivity with thiols and their activity in L5178-Y cells. The in vitro reactions clearly showed a selective Michael type 1,4-addition reaction between thiols (glutathione and N-acetylcysteine, which are nucleophiles) and the AMI (1f and 1h, which are electrophiles). In cell cultures, the compounds induced a decreasing cellular viability and an apoptotic effect of up to 59.8% at 48 h. The ex vivo experimental model showed an important reduction of thiol levels in cells treated with 1h. Decreased cellular viability and increased apoptosis were confirmed by flow cytometry, DNA fragmentation and microscopy analysis (cytological studies). The increase in apoptosis on L5178-Y cells probably occurred, at least in part, by a decrease in glutathione levels and an increase in free radicals concentration. The decreased glutathione levels seem to make cancer cells more susceptible to death by apoptosis, and should certainly make them more vulnerable to a less aggressive treatment.