Small molecule inhibitors of necrosis

#This invention was made in part with Government funding, and the Government therefore has certain rights in the invention.

#In general, the invention relates to methods and compounds used to decrease necrosis.

#In many diseases, cell death is mediated through apoptotic and/or necrotic pathways. While much is known about the mechanisms of action that control apoptosis, control of necrosis is not as well understood. Understanding the mechanisms regulating both necrosis and apoptosis in cells is essential to being able to treat conditions, such as neurodegenerative diseases, stroke, coronary heart disease, kidney disease, and liver disease. A thorough understanding of necrotic and apoptotic cell death pathways is also crucial to treating AIDS and the conditions associated with AIDS, such as retinal necrosis.

#FIG. 1 is a schematic representation of the chemical structure of a molecule which may be used to decrease necrosis. In this chemical structure each R1 is independently selected from the group consisting of hydrogen, methyl, carboxy, hydroxyl, methoxyl, amino, and nitro; R2 is selected from the group consisting of hydrogen, alkyl, and acyl; R3 is selected from the group consisting of alkyl, acyl, halogen, hydrogen, or hydroxyl; R4 is selected from the group consisting of methyl, hydroxyl, carboxyl, and linear and branching alkyl groups; X is selected from the group consisting of ═O, —OH and —H; Y is selected from the group consisting of ═S and —SR5, where R5 is either hydrogen or an alkyl group; and each of the bonds (a), (b), and (c) independently is either a double or single bond, provided, however, that bond (a) and bond (b) are not both double bonds.

#FIG. 2 is a schematic representation of the chemical structure of chemical compound ID number 115807 from the ChemBridge chemical compound library.

#FIG. 3 is a schematic representation of the chemical structure of a molecule which may be used to decrease necrosis. In this chemical structure, each of X1 and X2 is independently selected from the group consisting of ═O, —OH and —H; R1 is selected from the group consisting of hydrogen and a hydroxyl; R2 is selected from the group consisting of hydrogen, sulfate, nitro, and halide; and the bond (a) is either a single or double bond.

#Described herein are methods for decreasing necrosis, as well as for treating a condition in a subject. Techniques for carrying out the methods of the invention are now described in detail.

#Identification of Chemical Compounds that Decrease Cell Necrosis

#Assays that measure cell necrosis may be used to facilitate the identification of molecules that decrease necrosis induced by stimuli, such as zVAD-fmk/TNFα. In one approach, zVAD-fmk is added to the culture media of cells at high density (for example, 5×105 or 7.5×105 cells/ml), which are capable of undergoing necrosis in response to zVAD-fmk/TNFα. Candidate molecules, for example, chemical compounds from the ChemBridge chemical library are added, in varying concentrations to the cells, and the cells are then exposed to TNFα.