Inducing high levels of reactive oxygen species (ROS) inside tumor cells is a cancer therapy method termed chemodynamic therapy (CDT). Relying on delivery of Fenton reaction promoters such as Fe2+, CDT takes advantage of overproduced ROS in the tumor microenvironment. We developed a peptide‐H2S donor conjugate, complexed with Fe2+, termed AAN‐PTC–Fe2+. The AAN tripeptide was specifically cleaved by legumain, an enzyme overexpressed in glioma cells, to release carbonyl sulfide (COS). Hydrolysis of COS by carbonic anhydrase formed H2S, an inhibitor of catalase, an enzyme that detoxifies H2O2. Fe2+ and H2S together increased intracellular ROS levels and decreased viability in C6 glioma cells compared with controls lacking either Fe2+, the AAN sequence, or the ability to generate H2S. AAN‐PTC–Fe2+ performed better than temezolimide while exhibiting no cytotoxicity toward H9C2 cardiomyocytes. This study provides an H2S‐amplified, enzyme‐responsive platform for synergistic cancer treatment.

Inducing high levels of reactive oxygen species (ROS) inside tumor cells is a cancer therapy method termed chemodynamic therapy (CDT). Relying on delivery of Fenton reaction promoters such as Fe 2 + , CDT takes advantage of overproduced ROS in the tumor microenvironment. We developed a peptide-H 2 S donor conjugate, complexed with Fe 2 + , termed AAN-PTC-Fe 2 + . The AAN tripeptide was specifically cleaved by legumain, an enzyme overexpressed in glioma cells, to release carbonyl sulfide (COS). Hydrolysis of COS by carbonic anhydrase formed H 2 S, an inhibitor of catalase, an enzyme that detoxifies H 2 O 2 . Fe 2 + and H 2 S together increased intracellular ROS levels and decreased viability in C6 glioma cells compared with controls lacking either Fe 2 + , the AAN sequence, or the ability to generate H 2 S. AAN-PTC-Fe 2 + performed better than temezolimide while exhibiting no cytotoxicity toward H9C2 cardiomyocytes. This study provides an H 2 Samplified, enzyme-responsive platform for synergistic cancer treatment.