HPV E7 oncogene expression impairs Rb function and confers CDK4/6 inhibitor resistance in cervical cancer.

Abstract

e17504 Background: Malignant tumor proliferation is one of the important factors for poor prognosis. CDK4/6 inhibitors can delay tumor progression by inhibiting cell cycle and inducing apoptosis, so they are potentially effective broad-spectrum anti-tumor targeted drugs, but drug resistance limits their clinical applications. Our previous research found that the CDK4/6 inhibitor LEE011 has anti-tumor effects on cervical cancer C33A cells, but no anti-tumor activity against HeLa. The mechanism is not very clear, we speculate that it is related to HPV infection. In this research, our aim is to clarify the correlation between HPV and tumor drug sensitivity and explore the possible mechanisms. Methods: Two HPV positive human cervical cancer cell lines, SiHa and Caski were chosen to verify the sensitivity to LEE011 by cell morphology, cell cycle and apoptosis. The antitumor activity of CDK4/6 inhibitor LEE011 by E7 knockin C33A and E7 knock-down HeLa cell lines was assessed by in vitro clonogenic assay, flow cytometry, and target inhibition verified by immunoblotting. Effects of LEE011 and E7 knock-down combination in vivo were studied by xenograft tumor regrowth delay, and tissue section immunohistochemistry. Results: SiHa and Caski were insensitive to LEE011 antitumor activity. Enhancement of drugsensitivity was lost in cell lines with HPV positive. After down-regulation of E7, LEE011 treatment increased the inhibition cell proliferation and pro-apoptosis of HeLa. Mechanistically, the loss of E7-induced Rb inactivation and LEE011 inhibited Rb phosphorylation, leading to cell-cycle arrest. In vivo, LEE011 inhibited tumor regrowth, with sustained inhibition of cyclin D-CDK4/6-Rb-E2F1 activity in E7 knock-down HeLa. However, up-regulation of E7 can reduce the ability of LEE011 to inhibit cell proliferation and pro-apoptosis of C33A. In summary, our study signifies inhibiting the CDK4/6 pathway by LEE011 in combination with down-regulation of E7 as a promising therapeutic strategy to treat cervical cancer. Conclusions: These findings suggested that HPV can affect Rb function by expressing oncoprotein E7 leading to resistance to targeted drug therapy, which means that HPV may be a biomarkers of drug therapy efficacy.