Reduced glutathione rather than oxygen concentration determines the reduction rate of nitroimizadol probes used as hypoxia markers

Introduction: Tumor hypoxia results from the negative balance between the oxygen demands of the tissue and the capacity of the vasculature to deliver satisfactory oxygen provisions. The resulting oxygen deficit has important consequences in the aggressiveness and malignancy of the tumors as well as on their resistance to therapy, endowing the imaging of hypoxia with vital repercussions in tumor prognosis and therapy design (1). Nitroimidazolyl derivatives (EF5, Pimonidzole, 18F-MISO) have been extensively used as molecular markers of hypoxia in combination with different techniques as immunohistochemistry (2), PET (3) or MRI (4). Their use is based on the in vivo reduction of the nitro-group and the subsequent trapping of reactive imidazolyl derivatives in regions of low oxygen tension (5). A key issue to be solved in order to fully understand the activity of these markers is the mechanism of reduction. However the details of the reduction mechanism and its rate determining steps have remained elusive through the years. Here we investigate the kinetics of in vitro reduction of commercially available, as well as newly synthesized nitroimidazole based hypoxia probes.