Degradation of AHR by PROTACS : A novel approach for cancer chemoprevention

321 Activation of the aryl hydrocarbon receptor (AHR) leads to many adverse biological effects including tumor promotion, epithelial hyperplasia, endocrine disruption and immunodeficiency. Emerging evidence also links activation of the AHR to the development of type II diabetes. The AHR is thought to mediate its biological effects via its upregulation of a number of target genes. Agonists of the AHR includes dioxin (2, 3, 7, 8 tetrachlorodibenzo-p-dioxin) and polyhalogenated aromatic hydrocarbons found in cigarette smoke and other air pollutants. Our hypothesis is that blocking the actions of the AHR via appropriate AHR antagonists is a valid approach to be used for treatment of a number of human disease states. In the current project we have focused on the development of an AHR antagonist using the PROTACS (PROteolysis TArgeting Chimeric moleculeS) approach. PROTACS is a novel approach of tagging small recognition sequences of a specific E3 ubiquitin ligase complex on to a known ligand for the receptor of interest (AHR) for targeting its degradation via the 26S proteosomal pathway. PROTACS contain a ligand for the AHR, a recognition sequence for the pVHL E3 ubiquitin ligase complex and a linker moiety. The initial, parent compound that serves as the AHR ligand in these studies is apigenin. Here, we demonstrate the design, efficacy and specificity of apigenin bound PROTAC molecules. The design of the apigenin-Protacs was initiated by synthesizing apigenin derivatives that retained binding to the AHR and antagonistic activity. The Apigenin-PROTACS proved to be capable of degrading the AHR in both HepG2 (human hepatoma) and NHK (Primary human keratinocytes) cells in a time-dependent and concentration-dependent manner. These initial experiments demonstrate that the PROTAC approach can be effective in inhibiting the actions of the AHR blocking AHR-mediated effects.