Novel Affibody Molecules Targeting the HPV16 E6 Oncoprotein Inhibited the Proliferation of Cervical Cancer Cells

Abstract

Despite prophylactic vaccination campaigns, high-risk human papillomavirus (HPV)-induced cervical cancer remains a significant health threat among women, especially in developing countries. The initial occurrence and consequent progression of this cancer type primarily rely on, E6 and E7, two key viral oncogenes expressed constitutively, inducing carcinogenesis. Thus, E6/E7 have been proposed as ideal targets for HPV-related cancer diagnosis and treatment. In this study, three novel HPV16 E6-binding affibody molecules (ZHPV16E61115, ZHPV16E61171, and ZHPV16E61235) were isolated from a randomized phage display library and cloned for bacterial production. These affibody molecules showed high binding affinity and specificity for recombinant and native HPV16 E6 as determined by surface plasmon resonance, indirect immunofluorescence, immunohistochemistry, and near-infrared small animal optical imaging in vitro and in vivo. Moreover, by binding to HPV16 E6 protein, ZHPV16E61235 blocked E6-mediated p53 degradation, which increased the expression of some key p53 target genes, including BAX, PUMA and p21, and thereby selectively reduced the viability and proliferation of HPV16-positive cells. Importantly, ZHPV16E61235 was applied in combination with HPV16 E7-binding affibody ZHPV16E7384 to simultaneously target the HPV16 E6/E7 oncoproteins, and this combination inhibited cell proliferation more potently than either modality alone. Mechanistic studies revealed that the synergistic antiproliferative activity depends primarily on the induction of cell apoptosis and senescence but not cell cycle arrest. Our findings provide strong evidence that three novel HPV16 E6-binding affibody molecules could form a novel basis for the development of rational strategies for molecular imaging and targeted therapy in HPV16-positive preneoplastic and neoplastic lesions.

The HPV16E7 Affibody as a Novel Potential Therapeutic Agent for Treating Cervical Cancer Is Likely Internalized through Dynamin and Caveolin-1 Dependent Endocytosis

“…Some therapeutic affibodies have now entered preclinical and clinical trials for cancer treatment [ 5 ]. Recently, several affibodies using intracellular proteins as targets show great potential in cancer and other disease treatment [ 6 , 7 , 8 , 9 , 10 , 11 ]. We have previously developed the affibody Z HPV16E7 384 as a cervical cancer therapeutic agent [ 6 , 7 ].…”

“…Several therapeutic affibodies have now entered preclinical and clinical trials for cancer treatment [ 5 ]. In more recent studies, several affibodies that target intracellular proteins show great potential in cancer and other disease therapy, which include the affibodies targeting HPV16 E6 or E7 for cervical cancer, those targeting the EBV LMP1 C-terminal domain or LMP2A N-terminal domain for nasopharyngeal carcinoma, and those targeting Chlamydia trachomatis MOMP for Chlamydia trachomatis [ 6 , 7 , 8 , 9 , 10 , 11 ]. However, a key question that remains to be answered is whether and how the affibodies enter the target cells to interact with intracellular target proteins, leading to the inhibition of the target cell proliferation and finally to destroy the target cells.…”

The Effect of Oncogene Proteins of Human Papillomaviruses on Apoptosis Pathways in Prostate Cancer

“…Destabilizes p63 and p73 with direct impact [74,75] It inhibits p53 activity and reduces apoptosis in the cancer cell [76,77] It can also inhibit TRAIL-induced apoptosis by targeting FADD and caspase-8 [78] Activates AKT by binding to p53, p21, and p27 proteins [79] Inactivates PTEN via PDZ proteins [80,81] E5 Prevents Fas ligand (FASL or CD95L) from inhibiting FASL, TRAILdependent apoptosis [82,83] Prevent TRAIL and prevent the break of pro-caspases-3 and 8 also of PARP [84] Increase the sensitivity of EGFR to EGF stimulation [85][86][87] Activates AKT, then Bax binding to mitochondria inhibited [12,66,74] E2…”

Bacteria-derived chimeric toxins as potential anticancer agents

“…Affibody is a small cell-penetrating molecule derived from the Z domain of Staphylococcus aureus protein A (SP-A) ( 87 ). One of the major challenges of immunotoxin-based cancer therapy is the relatively large sizes of the antibodies used.…”