Rationale MnTE-2-PyP (T2E), a Mn-based SOD mimic, is able to inhibit PC3 cell (advanced prostate cancer cell line) growth, but not normal fibroblasts growth in vitro. We hypothesize that T2E increases H2O2 levels in PC3 cells, but not normal fibroblasts, and that the oxidation of key proteins that regulate cell cycle contributes to the cancer growth arrest. Methods PC3 and mouse prostate fibroblasts (MPF) were used. A ROS-Glo assay was performed to measure H2O2 levels. Oxidized proteins were isolated by biotinylated iodoacetamide probes and identified by mass spectrometry. PP1 activity was measured using a Ser/Thr phosphatase assay kit. Cell cycle distribution was measured by FACS with PI and BrdU staining. Cyclin D and pRB levels and phosphorylation levels were examined by western blots. Results T2E increased H2O2 levels in PC3 cells by 63.3% (P = 0.0080) as compared to PBS-treated cells but not in normal fibroblasts. Accordingly, serine/threonine-protein phosphatase PP1 was oxidized by T2E, which resulted in a 22.5% (P=0.022) decrease in PP1 activity in T2E -treated PC3 cells as compared to controls. Cyclin D1 levels decreased 25.8% (P=0.042) while phospho-cyclin D1 to total cyclin D1 ratio increased to 165.3% (P=0.0018) in T2E-treated PC3 cells as compared to PBS-treated ones. Cyclin D1 regulates pRB phosphorylation and this led to a 46.2% decrease (P=0.017) of phospho-pRB to total pRB in PC3 cells, which might account for the reduced proliferation of PC3 at Day 4 (48.9% decrease as compared to PBS treatment, P=0.030). However, no significant change in cyclin D1 levels were observed in MPF. Conclusion T2E increases H2O2 levels in PC3 cells but not MPF, and this causes the oxidation and loss of PP1 activity, which may result in the hyper-phosphorylation of cyclin D1. The increased phospho-cyclin D1 level leads to hypo-phosphorylaiton of pRB and induces cell growth arrest in T2E-treated PC3 cells, but PMF cyclin D1 levels are not affected by T2E.