Dickkopf-1: An indirect target for oncogenic HPVs and its role in Cisplatin-mediated apoptosis in cervical cancer cells

Oncogenic human papillomavirus (HPV) types cause cervical cancer. The continuous expression of the viral E6/E7 oncogenes controls the malignant phenotype of HPV-positive cancer cells, as they interfere with major tumor-suppressive pathways and thereby induce uncontrolled cell proliferation and cellular resistance towards apoptosis. The low vaccination rates with the prophylactic vaccines protecting against oncogenic HPV infections as well as the overall poor therapeutic response of advanced or recurrent cervical carcinomas emphasize the importance to improve our concepts of HPV-associated carcinogenesis as a basis to develop more efficient therapeutic options. The overall aim of this thesis was to characterize the regulation and functional role of the putative tumor suppressor protein Dickkopf-1 (Dkk1) in cervical cancer cells. My studies revealed that Dkk1 levels are strongly restricted by HPV oncogene expression. Mechanistically, E6 was found to be a key driver in this process by interfering with the ability of p53 to activate Dkk1 expression. Moreover, epigenetic mechanisms as well as metabolic changes at hypoxic conditions, such as the inhibition of oxidative phosphorylation (OXPHOS), were shown to be additional driving forces in silencing Dkk1 expression levels in cervical cancer cells. Further, I could demonstrate that Dkk1 levels are a critical determinant for the Cisplatin response of cervical cancer cells: While treatment with Dkk1 conditioned medium sensitized HeLa cells towards Cisplatin treatment, RNAi- or CRISPR/Cas9-induced Dkk1 repression or depletion efficiently protected cervical cancer cells from Cisplatin-induced apoptosis. Despite the well-characterized role of Dkk1 in antagonizing canonical Wnt signaling, the Dkk1-dependent Cisplatin response was not associated with alterations of this pathway. Instead, transcriptome analyses using Affymetrix GeneChip arrays uncovered that multiple members of the AP-1 protein family were less efficiently induced upon Cisplatin treatment in Dkk1 knockout HeLa cells compared to parental HeLa cells. Functional studies revealed that Dkk1 expression was required to induce pro-apoptotic JNK signaling, which acts upstream of AP-1 and is necessary to trigger an efficient apoptotic Cisplatin response in cervical cancer cells. Collectively, this work discloses that Dkk1 expression in cervical cancer cells is negatively controlled by oncogenic HPVs through E6-mediated p53 degradation. Moreover, it shows that Dkk1 repression provides Cisplatin resistance in cervical cancer cells, which is mechanistically caused by the impairment of Cisplatin-induced pro-apoptotic JNK signaling

Effects of the Antifungal Agent Ciclopirox in HPV-Positive Cancer Cells : Repression of Viral E6/E7 Oncogene Expression and Induction of Senescence and Apoptosis

The malignant growth of human papillomavirus (HPV)-positive cancer cells is dependent on the continuous expression of the viral E6/E7 oncogenes. Here, we examined the effects of iron deprivation on the phenotype of HPV-positive cervical cancer cells. We found that iron chelators, such as the topical antifungal agent ciclopirox (CPX), strongly repress HPV E6/E7 oncogene expression, both at the transcript and protein level. CPX efficiently blocks the proliferation of HPV-positive cancer cells by inducing cellular senescence. Although active mTOR signaling is considered to be critical for the cellular senescence response towards a variety of pro-senescent agents, CPX-induced senescence occurs under conditions of severely impaired mTOR signaling. Prolonged CPX treatment leads to p53-independent Caspase-3/7 activation and induction of apoptosis. CPX also eliminates HPV-positive cancer cells under hypoxic conditions through induction of apoptosis. Taken together, these results show that iron deprivation exerts profound anti-viral and anti-proliferative effects in HPV-positive cancer cells and suggest that iron chelators, such as CPX, possess therapeutic potential as HPV-inhibitory, pro-senescent and pro-apoptotic agents in both normoxic and hypoxic environments. This article is protected by copyright. All rights reserved.publishe