Reactive oxygen and nitrogen species have emerged as important endogenous regulators of angiogenesis (blood vessel growth), which can contribute to the growth of cancers. Reactive oxygen species (ROS) include superoxide ions and hydrogen peroxide. NADPH oxidases, the mitochondrial respiratory chain, and endothelial NO-synthase (NOS) are all major sources of reactive oxygen species that lead to endothelial dysfunction in the cell. Nitric oxide regulates the cell’s response to metabolic stress and low oxygen tension. NO-synthase catalyzes the production of nitric oxide from L-arginine.
The family of NO-synthases perform a range of functions in the body, including maintenance of vascular tone, secretion of insulin, peristalsis, and angiogenesis. Nitric oxide and ROS create a balance between oxidation and reduction in healthy cells. However, in some cancerous cells, nitric oxide promotes tumor growth and metastasis. NADPH oxidases and endothelial NO-synthase are therefore targets of interest in pharmaceutical development for cancer and cardiovascular disease.
Each molecule of NO-synthase contains an N-terminal oxygenase domain and a multi-domain C-terminal reductase. Within the oxygen domain, there is a heme-a ferrous iron contained within a porphyrin ring. NOS inhibitors are usually designed to target the heme site within the oxygenase domain of the molecule. Those inhibitors don’t prevent ROS formation in the reductase domain. A drug targeting the reductase domain would inhibit ROS production as well as the formation of NO.
Researchers based in France and Belgium designed a novel probe they named a nanoshutter (NS1) that can bind to NOS at its NADPH binding site in the reductase domain of the molecule. (Rouaud et al. 2014) NS1 inhibits formation of NO by competing with NADPH at the binding site. Binding activates fluorescence in NS1, allowing imaging of endothelial NOS in live cells.