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

Affibodies targeting intracellular proteins have a great potential to function as ideal therapeutic agents. However, little is known about how the affibodies enter target cells to interact with intracellular target proteins. We have previously developed the HPV16E7 affibody (Z HPV16E7 384) for HPV16 positive cervical cancer treatment. Here, we explored the underlying mechanisms of Z HPV16E7 384 and found that Z HPV16E7 384 significantly inhibited the proliferation of target cells and induced a G1/S phase cell cycle arrest. Furthermore, Z HPV16E7 384 treatment resulted in the upregulation of retinoblastoma protein (Rb) and downregulation of phosphorylated Rb (pRb), E2F1, cyclin D1, and CDK4 in the target cells. Moreover, treatment with dynamin or the caveolin-1 inhibitor not only significantly suppressed the internalization of Z HPV16E7 384 into target cells but also reversed the regulation of cell cycle factors by Z HPV16E7 384. Overall, these results indicate that Z HPV16E7 384 was likely internalized specifically into target cells through dynamin- and caveolin-1 mediated endocytosis. Z HPV16E7 384 induced the cell cycle arrest in the G1/S phase at least partially by interrupting HPV16E7 binding to and degrading Rb, subsequently leading to the downregulation of E2F1, cyclin D1, CDK4, and pRb, which ultimately inhibited target cell proliferation. These findings provide a rationale of using Z HPV16E7 384 to conduct a clinical trial for target therapy in cervical cancer.