Abstract Mitochondrial respiratory complex II (CII), also known as succinate dehydrogenase, plays a critical role in mitochondrial metabolism. Known but low potency CII inhibitors are selectively cytotoxic to cancer cells including the benzothiadiazine‐based anti‐hypoglycemic diazoxide. Herein, we study the structure‐activity relationship of benzothiadiazine derivatives for CII inhibition and their effect on cancer cells for the first time. A 15‐fold increase in CII inhibition was achieved over diazoxide, albeit with micromolar IC 50 values. Cytotoxicity evaluation of the novel derivatives resulted in the identification of compounds with much greater antineoplastic effect than diazoxide, the most potent of which possesses an IC 50 of 2.93±0.07 μM in a cellular model of triple‐negative breast cancer, with high selectivity over nonmalignant cells and more than double the potency of the clinical agent 5‐fluorouracil. No correlation between cytotoxicity and CII inhibition was found, thus indicating an as‐yet‐undefined mechanism of action of this scaffold. The derivatives described herein represent valuable hit compounds for therapeutic discovery in triple‐negative breast cancer.