Covalent Ligands that Target the Human Papillomavirus E6 Oncoprotein

The high‐risk human papillomavirus (HPV) early protein 6 (E6) viral oncoprotein is a key player in HPV‐mediated cancers and is the major driver of cervical, anal, and a substantial portion of head and neck cancers. HPV E6 recruits the cellular ubiquitin E3‐ligase E6‐associated protein (E6AP), also termed UBE3A, and this E6/E6AP complex in turn recruits the tumor suppressor p53, resulting in the ubiquitin‐mediated degradation of p53. The cysteine‐rich E6 protein presents an attractive target for chemical intervention through covalent, cysteine‐reactive chemistries. While previous studies have uncovered peptidic covalent inhibitors of E6, we sought to determine whether we could identify nonpeptidic functional binders. We screened a library of cysteine‐reactive covalent ligands and identified a pyrazoline hit, EN218, bearing a chloroacetamide cysteine‐reactive warhead. Evaluation of pyrazoline analogs led to the development of an improved molecule, ( S )‐WX48A, that stereoselectively reacted with C58 on E6 to inhibit E6‐mediated p53 ubiquitination. While this molecule is still in an early stage and requires further medicinal chemistry optimization to exert on‐target cellular activity, our study demonstrates that E6 can be directly and stereoselectively targeted with a covalent nonpeptidic small‐molecule and provides a foundation for developing more drug‐like E6 inhibitors for HPV therapy.