Targeting the Two Oncogenic Functional Sites of the HPV E6 Oncoprotein with a High‐Affinity Bivalent Ligand

Abstract

The E6 oncoproteins of high-risk mucosal (hrm) HPVs contain a pocket that captures LxxLL motifs and a C-terminal motif that recruits PDZ domains, both functions being crucial for HPV-induced oncogenesis. Here, we fused together a PDZ domain and a LxxLL motif both previously known to bind E6. Using NMR, calorimetry and mammalian protein complementation assay, we show that the resulting PDZ-LxxLL chimera is a bivalent nanomolar ligand of E6, while its separated PDZ and LxxLL components are only micromolar binders. The chimera binds to all hrm-HPV E6 proteins tested but not to low-risk mucosal or cutaneous HPV E6. Adenovirus-mediated expression of the chimera specifically induces the death of HPV-positive cells, concomitant with increased levels of tumour suppressor P53, of its transcriptional target p21, and of apoptosis marker cleaved caspase 3. The bifunctional PDZ-LxxLL chimera opens new perspectives both for diagnostics and therapy of HPV-induced cervical and head and neck cancers.

Mitotic control of human papillomavirus genome-containing cells is regulated by the function of the PDZ-binding motif of the E6 oncoprotein

“…The nature of the E6 PBM interaction with PDZ domains makes it an amenable peptide-protein interaction for inhibition by small molecules with the intention to develop novel therapeutics for the treatment of HPV–associated cancers [54, 55]. Since the E6 PBM is essential for viral replication [27], inhibition of this function may be considered a route of anti-viral intervention to treat active infections.…”

PDZ Domains as Drug Targets

“…[257] The human adenovirus type 9 (Ad9) E4 region ORF1 gene product, E4-ORF1, is another viral protein, which targets several PDZ-containing proteins through a C-terminal PDZ binding motif. d) Graphical model of the PDZbody [259,275] where the HPV18-E6 protein binds to the canonical binding pocket in the optimized SAP-97-PDZ2 through its C-terminal residues. [258] Adv.…”

“…[258] Adv. [275] Human Papillomavirus (HPV) E6 and E7 hijack the cell during infection by interacting with endogenous proteins such as the well-described transcription factor and tumor suppressor, p53, and several PDZ-containing proteins. 2019, 2, show very similar insertion of the two peptides into the binding pocket.…”

“…[272] This activation of PI3K deregulates cytosolic signalling pathway, which leads to cell survival and metabolism, and enhances virus replication. [275] In addition to the more well-studied interactions with viral proteins from HTLV-1, Ad9, and HPV, SAP-97 and Scribble have also been associated with Hepatitis C virus. [274] Because of the role HPV E6 plays in transforming infected cells into cancer cells, it is considered a potential drug target.…”

The Human Papillomavirus E6 PDZ Binding Motif: From Life Cycle to Malignancy

“…These results are particularly intriguing since the PBM deletion mutants of E6 target p53 as efficiently as wild type E6. This indicates that at least one aspect of E6 PDZ targeting links directly to pathways that are controlled by p53 [ 67 , 68 ]. It is also possible that the PDZ-PBM interaction of wild type E6 contributes towards the abrogation of a p53 function that is independent of the degradation of p53, and this functional ablation is necessary for viral genome maintenance.…”