Suppression of cervical carcinoma cell growth by intracytoplasmic codelivery of anti-oncoprotein E6 antibody and small interfering RNA

Courtête, Jérôme; Sibler, Annie-Paule; Zeder‐Lutz, Gabrielle; Dalkara, Deniz; Oulad‐Abdelghani, Mustapha; Zuber, Guy; Weiss, Étienne

doi:10.1158/1535-7163.mct-06-0808

Cited by 58 publications

(38 citation statements)

References 49 publications

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“…All antibodies used in this study were obtained from the following commercial sources, except for the 4C6 mouse monoclonal antibody to HPV16 E6 (5,28), generated by Etienne Weiss (University of Strasbourg) and provided through Arbor Vita Corporation (Sunnyvale, CA): actin (1501; Chemicon), E6AP (sc-25509; Santa Cruz), HA-horseradish peroxidase (Roche), IB␣ (9242; Cell Signaling), p53 (OP43; Calbiochem), ubiquitin (sc-9133; Santa Cruz), USP15 (H00009958-M01; Abnova), and Xpress (R910-25; Invitrogen).…”

Section: Methodsmentioning

confidence: 99%

The Ubiquitin-Specific Peptidase USP15 Regulates Human Papillomavirus Type 16 E6 Protein Stability

Vos

Altreuter

White

et al. 2009

J Virol

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Proteomic identification of human papillomavirus type 16 (HPV16) E6-interacting proteins revealed several proteins involved in ubiquitin-mediated proteolysis. In addition to the well-characterized E6AP ubiquitinprotein ligase, a second HECT domain protein (HERC2) and a deubiquitylating enzyme (USP15) were identified by tandem affinity purification of HPV16 E6-associated proteins. This study focuses on the functional consequences of the interaction of E6 with USP15. Overexpression of USP15 resulted in increased levels of the E6 protein, and the small interfering RNA-mediated knockdown of USP15 decreased E6 protein levels. These results implicate USP15 directly in the regulation of E6 protein stability and suggest that ubiquitylated E6 could be a substrate for USP15 ubiquitin peptidase activity. It remains possible that E6 could affect the activity of USP15 on specific cellular substrates, a hypothesis that can be tested as more is learned about the substrates and pathways controlled by USP15.Human papillomaviruses (HPVs) are associated with several human cancers, most notably human cervical cancer, the second most common cancer among women worldwide (43). Papillomaviruses cause proliferative squamous epithelial lesions, and more than 100 HPV types have been described (14). The HPV types associated with mucosal squamous epithelial lesions have been further classified into high-or low-risk types based on the propensity for the lesions with which they are associated to progress to cancer. Among the high-risk HPV types, HPV type 16 (HPV16) and HPV18 account for approximately 70% of cervical cancers (43). The high-risk HPV types carry two genes, the E6 and E7 genes, which have oncogenic properties and are always expressed in HPV-positive cancers. E6 and E7 interfere with the p53 and retinoblastoma (pRB) tumor suppressor pathways, respectively, and contribute directly to cell cycle alterations, protection from apoptosis, and transformation (14). The dysregulated expression of the E6 and E7 oncoproteins is an important step in the progression from a preneoplastic stage to cancer in HPV-infected cells and is often a consequence of the integration of the viral genome into the host chromosome.The interaction between E6 and p53 is mediated by the E3 ubiquitin ligase E6AP (15). E6, p53, and E6AP form a complex in which E6 directs the ligase activity of E6AP to p53, thereby targeting p53 for ubiquitin-mediated degradation (36). E6, however, has a number of other cellular partners and other functions. For instance, the C terminus of the high-risk E6 protein contains a PDZ binding motif (20,25) that mediates the interaction with several PDZ domain-containing proteins, including discs large (Dlg), Scribble (Scrib), the MAGI family of proteins, MUPP1, and PATJ (9, 10, 29). Some of these proteins are also targeted for degradation in an E6AP-dependent manner (22, 29). While the major mechanism of oncogenesis revolves around E6's ability to inhibit the proapoptotic effects of p53, recent work involving the PDZ domain proteins indica...

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Section: Methodsmentioning

confidence: 99%

The Ubiquitin-Specific Peptidase USP15 Regulates Human Papillomavirus Type 16 E6 Protein Stability

Vos

Altreuter

White

et al. 2009

J Virol

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“…The inhibition of E6 was obtained at the mRNA level by RNA interference (Butz et al, 2003) or at the protein level by transduced full-length anti-E6 antibody (Courtete et al, 2007) or recombinant antibody fragments (Griffin et al, 2006;Lagrange et al, 2007). In these studies, selective E6 inhibition was found to induce cell growth arrest accompanied in some cases by apoptosis.…”

Section: Introductionmentioning

confidence: 99%

A single-codon mutation converts HPV16 E6 oncoprotein into a potential tumor suppressor, which induces p53-dependent senescence of HPV-positive HeLa cervical cancer cells

et al. 2008

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High-risk mucosal human papillomaviruses (HPV), mainly HPV16 and HPV18, are implicated in cervical carcinogenesis. HPV16 E6 oncoprotein binds and often targets for degradation numerous cell proteins, including the tumor suppressor p53 and several PDZ domain proteins. Here, we show that a single-point mutation, F47R, is sufficient to convert the HPV16 E6 oncoprotein into a suppressor of HPV-positive HeLa cervical cancer cells proliferation. The E6 F47R mutant is defective for polyubiquitination and subsequent degradation of p53. When expressed in HPV-positive cervical cancer cells, E6 F47R acts as a dominant negative mutant by counteracting the p53 degradation activity of endogenous E6 and restoring high p53 protein levels. Moreover, the prolonged expression of E6 F47R leads to suppression of HeLa cells proliferation through the induction of premature senescence. This phenotype is independent on the PDZ-binding activity of E6. F47R-senescent HeLa cells exhibit a sustained expression of p53, hMDM2 and p21 CIP proteins and a reduced expression of endogenous HPV18 E6 protein. Finally, small interfering RNAs directed against p53 counteract the effect of E6 F47R expression, indicating that E6 F47R-induced cellular senescence is strongly dependent on p53 signaling pathway.

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“…In another study, delivery of monoclonal antibodies that bind to HPV-16 E6 and neutralize its biological activity in vitro could restore p53 function in SiHa cells but not in HeLa cells. Proliferation of SiHa cells was markedly diminished, but no apoptosis was detectable in HeLa cells 47. On that basis, involvement of GRP58 in the cell death process may vary from cell type to cell type (eg, glandular versus squamous), HPV copy number, expression of oncoproteins, or other variables that remain to be elucidated.…”

Section: Resultsmentioning

confidence: 99%

Potential implications of GRP58 expression and susceptibility of cervical cancer to cisplatin and thymoquinone-based therapy

Hafiza¹,

Latifah²

2014

OTT

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A new therapeutic approach of looking at the expression of glucose-regulated protein (GRP) 58 as an indication of cisplatin sensitivity may eradicate fruitless treatment and side effects in patients with cervical cancer. Thymoquinone, the bioactive compound in Nigella sativa, has been reported to have an antiproliferative effect on cervical cancer cells. This study compared the cytotoxic effects of cisplatin, a drug commonly used in the treatment of cervical cancer, and thymoquinone in cervical cancer (HeLa and SiHa) cell lines by 3-(4,5-Dimethyl thiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, and measured GRP58 expression in the cells by quantitative real-time polymerase chain reaction and Western blotting. Cisplatin had higher antiproliferative activity towards the cervical cancer cell lines than thymoquinone in a dose-dependent and time-dependent manner. However, cisplatin was more toxic to normal 3T3 and Vero cell lines than thymoquinone. The half maximal inhibitory concentration (IC50) of cisplatin in HeLa and SiHa cells at 72 hours was 13.3±2.52 μM and 19.5±2.12 μM, respectively. Meanwhile, the IC50 of thymoquinone in HeLa and SiHa cells was 29.57±5.81 μM and 23.41±1.51 μM, respectively (P<0.05). A significant correlation was found between the cytotoxicity of cisplatin and expression of GRP58, but this relationship was not significant for thymoquinone. Therefore, the response of cervical cancer cells to cisplatin can be predicted on the basis of GRP58 expression.

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Suppression of cervical carcinoma cell growth by intracytoplasmic codelivery of anti-oncoprotein E6 antibody and small interfering RNA

Cited by 58 publications

References 49 publications

The Ubiquitin-Specific Peptidase USP15 Regulates Human Papillomavirus Type 16 E6 Protein Stability

The Ubiquitin-Specific Peptidase USP15 Regulates Human Papillomavirus Type 16 E6 Protein Stability

A single-codon mutation converts HPV16 E6 oncoprotein into a potential tumor suppressor, which induces p53-dependent senescence of HPV-positive HeLa cervical cancer cells

Potential implications of GRP58 expression and susceptibility of cervical cancer to cisplatin and thymoquinone-based therapy

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