A novel series of benzoylsulfonamide derivatives were synthesized and biologically evaluated. Among them, 4-(biphenyl-4-ylmethylsulfanylmethyl)- N -(hexane-1-sulfonyl)benzamide (compound 18K ) was identified as a protein tyrosine phosphatase 1B (PTP1B) inhibitor with potent and selective inhibitory activity against PTP1B (IC₅₀=0.25 µM). Compound 18K functioned as a non-competitive inhibitor and bound to the allosteric site of PTP1B. It also showed high oral absorption in mice (the maximum drug concentration ( C _max)=45.5 µM at 30 mg/kg), rats ( C _max=53.6 µM at 30 mg/kg), and beagles ( C _max=37.8 µM at 10 mg/kg), and significantly reduced plasma glucose levels at 30 mg/kg/d ( per os ( p.o. )) for one week with no side effects in db / db mice. In conclusion, the substituted benzoylsulfonamide was shown to be a novel scaffold of a non-competitive and allosteric PTP1B inhibitor, and compound 18K has potential as an efficacious and safe anti-diabetic drug as well as a useful tool for investigations of the physiological and pathophysiological effects of allosteric PTP1B inhibition.