New insights into the interactions of HPV-16 E6*I and E6*II with p53 isoforms and induction of apoptosis in cancer-derived cell lines.

An important characteristic of cancers associated with high-risk human papillomaviruses (HR-HPV) is the inability of p53 to activate apoptosis due to the effect of the oncoprotein E6. However, the effect of HPV-16 E6 splice variant isoforms (namely E6*I and E6*II), their interaction with the existing p53 isoforms, and their influence on apoptosis is unclear. Here, we report the outcome of ectopic expression of HPV-16 E6, E6*I, and E6*II on the relative levels of p53 and p53 isoforms Δ40p53 and Δ133p53 and their interactions with these proteins. Additionally, we evaluated the effect of ectopic expression of p53, Δ40p53, and Δ133p53 on apoptosis in a p53 null pulmonary cell line (H1299) co-transfected with E6 isoforms and p53+/+ cell lines with HR-HPV (SiHa and HeLa), transfected with p53 isoforms and treated with cisplatin, a conventional drug used to treat cervical cancer. Our results show that E6 and E6*II induced a significant decrease in p53, but only E6 triggered a Δ40p53 decrease and that E6*II interacts with p53 but not with Δ40p53 and Δ133p53. On the other hand, E6*I did not show any effect or interaction with the p53 isoforms. We found that apoptosis was elevated in H1299 cells transfected with p53 (p = 0.0001) and Δ40p53 (p = 0.0001). A weak apoptotic effect was observed when Δ133p53 was ectopically expressed (p = 0.0195). We observed that both p53 (p = 0.0006) and Δ40p53 (p = 0.0014) induced apoptosis in cisplatin-treated SiHa cells; however in cisplatin-treated HeLa cells, only p53 induced apoptosis (p = 0.0029). No significant differences in apoptosis were observed upon ectopic expression of p53, Δ40p53, and Δ133p53 in SiHa and HeLa cells. Our findings suggest a possible therapeutic application for the combining of p53 or Δ40p53 with cisplatin to induce an increased apoptosis of cancer cells expressing E6 isoforms from HPV-16.

Nanog, in Cooperation with AP1, Increases the Expression of E6/E7 Oncogenes from HPV Types 16/18.

Persistent infections with some types of human papillomavirus (HPV) constitute the major etiological factor for cervical cancer development. Nanog, a stem cell transcription factor has been shown to increase during cancer progression. We wanted to determine whether Nanog could modulate transcription of E6 and E7 oncogenes. We used luciferase reporters under the regulation of the long control region (LCR) of HPV types 16 and 18 (HPV16/18) and performed RT-qPCR. We found that Nanog increases activity of both viral regulatory regions and elevates endogenous E6/E7 mRNA levels in cervical cancer-derived cells. We demonstrated by in vitro mutagenesis that changes at Nanog-binding sites found in the HPV18 LCR significantly inhibit transcriptional activation. Chromatin immunoprecipitation (ChIP) assays showed that Nanog binds in vivo to the HPV18 LCR, and its overexpression increases its binding as well as that of c-Jun. Surprisingly, we observed that mutation of AP1-binding sites also affect Nanog's ability to activate transcription, suggesting cooperation between the two factors. We searched for putative Nanog-binding sites in the LCR of several HPVs and surprisingly found them only in those types associated with cancer development. Our study shows, for the first time, a role for Nanog in the regulation of E6/E7 transcription of HPV16/18.

Human papillomavirus-16 DNA methylation patterns support a causal association of the virus with oral squamous cell carcinomas.

Infection with human papillomavirus-16 (HPV-16) is the cause of most anogenital carcinomas. This virus is also detected in about 20% of all head and neck squamous cell carcinomas. While there is strong evidence for a causal etiological role in the case of tonsillar carcinomas, causal association with malignant lesions of the oral cavity is not yet conclusive. Our previous investigations of HPV-16 DNA methylation in anogenital sites have identified hypermethylation of the L1 gene and part of the long control region in many malignant lesions, but rarely in asymptomatic infections and low-grade precancerous lesions. Here, we report hypermethylation of this diagnostically important segment of the viral DNA in 10 out of 12 HPV-16 positive oral carcinomas from Mexican patients. These data indicate epigenetic changes of HPV-16 in oral carcinomas similar to those in anogenital carcinomas, suggesting carcinogenic processes under the influence of HPV-16 in most if not all of these oral malignant lesions.

[High-risk human papilloma virus and cervical intraepithelial neoplasia in women at 2 hospitals in Mexico City]. / Virus de papiloma humano de alto riesgo (VPH-AR) y neoplasia intraepitelial cervical (NIC) en mujeres de dos hospitales de la Ciudad de México.

OBJECTIVE: To determine the high risk HPV (HR-HPV) association with Cervical Intraepithelial Neoplasia (CIN) in women of two Dysplasia Clinics in Mexico City. MATERIAL AND METHODS: Prolective case-control study was done. Women with and without security affiliation attended in Instituto Mexicano del Seguro Social (Hospital 1) and Hospital General de México (Hospital 2) were included in the study. Cases were women with histopathologic diagnosis of CIN and controls were women with negative dysplasia in cytologic study (Pap). Information was obtained by direct interview. HR-HPV was determined by Hybrid Capture II assay, in cervical samples. Bivariate and logistic regression analysis was done. RESULTS: One hundred and two cases and 192 controls from Hospital 1 and 89 cases and 66 controls from Hospital 2 were included. 83.3% and 77.3% of women from Hospital 1 and 2 respectively were positive to HR-HPV. The association HR-HPV and CIN in Hospital 1 was ORa = 40.6, C.I. 95% = 17-96.8; while in Hospital 2 there was not association. Age was an effect modifier in the HR-HVP and CIN association, in Hospital 1. It was observed a correlation between viral load and CIN degree. CONCLUSIONS: The HR-HPV infection frequency in controls and CIN I was higher than the reported in other studies. Age was a modifier in the HR-HPV association and CIN. In dysplasia clinics without medical referral system of patients is possible to observe similar risk factors to cervical cancer.

Virus de papiloma humano de alto riesgo (VPH-AR) y neoplasia intraepitelial cervical (NIC) en mujeres de dos hospitales de la Ciudad de México / High-risk human papilloma virus and cervical intraepithelial neoplasia in women at 2 hospitals in Mexico City

OBJECTIVE: To determine the high risk HPV (HR-HPV) association with Cervical Intraepithelial Neoplasia (CIN) in women of two Dysplasia Clinics in Mexico City. MATERIAL AND METHODS: Prolective case-control study was done. Women with and without security affiliation attended in Instituto Mexicano del Seguro Social (Hospital 1) and Hospital General de MÚxico (Hospital 2) were included in the study. Cases were women with histopathologic diagnosis of CIN and controls were women with negative dysplasia in cytologic study (Pap). Information was obtained by direct interview. HR-HPV was determined by Hybrid Capture II assay, in cervical samples. Bivariate and logistic regression analysis was done. RESULTS: One hundred and two cases and 192 controls from Hospital 1 and 89 cases and 66 controls from Hospital 2 were included. 83.3 and 77.3 of women from Hospital 1 and 2 respectively were positive to HR-HPV. The association HR-HPV and CIN in Hospital 1 was ORa = 40.6, C.I. 95 = 17-96.8; while in Hospital 2 there was not association. Age was an effect modifier in the HR-HVP and CIN association, in Hospital 1. It was observed a correlation between viral load and CIN degree. CONCLUSIONS: The HR-HPV infection frequency in controls and CIN I was higher than the reported in other studies. Age was a modifier in the HR-HPV association and CIN. In dysplasia clinics without medical referral system of patients is possible to observe similar risk factors to cervical cancer.