Identification of human papillomavirus (HPV) 16 DNA integration and the ensuing patterns of methylation in HPV‐associated head and neck squamous cell carcinoma cell lines

Hatano, Takashi; Sano, Daisuke; Takahashi, Hideaki; Hyakusoku, Hiroshi; Isono, Yasuhiro; Shimada, Seiichiro; Sawakuma, Kae; Takada, Kazuya; Oikawa, Ritsuko; Watanabe, Yoshiyuki; Yamamoto, Hiroyuki; Itoh, Fumio; Myers, Jeffrey N.

doi:10.1002/ijc.30589

Cited by 22 publications

(21 citation statements)

References 38 publications

(80 reference statements)

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“…A similar observation was made by Park et al (2011) [41]. This can be explained by the results of Hatano et al (2017), who found that the methylation of the HPV genome and human region near the integration site display the same methylation pattern [42]. So, in comparison to the extrachromosomal form of the viral genome, the methylation level of the integrated form is likely to be more influenced by the site of integration.…”

Section: Discussionsupporting

confidence: 67%

Detailed Characteristics of Tonsillar Tumors with Extrachromosomal or Integrated Form of Human Papillomavirus

Pokrývková

Saláková

Smahelova

et al. 2019

Viruses

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The human papillomavirus (HPV) integration, the critical step in viral carcinogenesis, most frequently occurs in the E2 gene, which results in its inactivation and in an increase of E6/E7 transcription. However, in a substantial number of tumors, the virus is present in an extrachromosomal form. For those tumors, the transformation mechanisms are not fully elucidated. Here we evaluated the possible mechanism of inactivating the E2 without interruption of the gene, methylation or mutation of the E2 binding sites (E2BSs) in HPV16-positive tonsillar tumors by next-generation and Sanger sequencing. Viral genome status was analyzed by the amplification of papillomavirus oncogene transcripts assay (APOT) and mRNA mapping, and expression of viral oncogenes was performed by qPCR. The methylation of E2BSs was significantly higher in tumors with an integrated, in comparison to extrachromosomal, form of the viral genome. No mutations were detected in the E2BSs. The viral oncogenes were equally expressed in samples with an integrated and extrachromosomal form of the virus. Only the nucleotide variants were identified in the E2 gene. No proposed mechanism of E2 inactivation was confirmed in tonsillar tumors with an extrachromosomal form of the HPV genome. The expression of E6/E7 genes seems to be sufficient to initiate and maintain the carcinogenic process

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Section: Discussionsupporting

confidence: 67%

Detailed Characteristics of Tonsillar Tumors with Extrachromosomal or Integrated Form of Human Papillomavirus

Pokrývková

Saláková

Smahelova

et al. 2019

Viruses

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“…Other viruses such as HBV (hepatitis B virus) show that methylation of the HBx gene is in accordance with the methylation status of the flanking human gene [ 45 ]. And also in their HPV research, they showed the DNA methylation status of the integrated HPV16 genome was affected by the methylation status of the human genome flanking integration breakpoints in three neck and head carcinoma cell lines [ 46 ].Thus, we suppose that some HPV integrants might be activated or inactivated through self-methylation induced by flanking methylation status of host genome when integrated into human genome, leading to tumorigenesis. Whether this is the case for HPV16-induced cervical cancer remains to be confirmed.…”

Section: Discussionmentioning

confidence: 99%

Identification of reliable biomarkers of human papillomavirus 16 methylation in cervical lesions based on integration status using high-resolution melting analysis

et al. 2018

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BackgroundThe dynamic methylation of human papillomavirus (HPV) 16 DNA is thought to be associated with the progression of cervical lesions. Previous studies that did not consider the physical status of HPV 16 may have incorrectly mapped HPV 16 methylomes. In order to identify reliable biomarkers for squamous cervical cancer (SCC), we comprehensively evaluated the methylation of HPV 16 depending on the integration incidence of each sample.MethodsBased on the integration status of 115 HPV 16-infected patients (50 SCC, 30 high-grade squamous intraepithelial lesion [HSIL], and 35 low-grade squamous intraepithelial lesion [LSIL]) and HPV 16-infected Caski cell lines by PCR detection of integrated papillomavirus sequences, we designed a series of primers that would not be influenced by breakpoints for a high-resolution melting (HRM) PCR method to detect the genome methylation.ResultsA few regions with recurrent interruptions were identified in E1, E2/E4, L1, and L2 despite scattering of breakpoints throughout all eight genes of HPV 16. Frequent integration sites often occurred concomitantly with methylated CpG sites. The HRM PCR method showed 100% agreement with pyrosequencing when 3% was set as the cutoff value. A panel of CpG sites such as nt5606, nt5609, nt5615, and nt5378 can be combined in reweighing calculations to distinguish SCC from HSIL and LSIL patients which have high sensitivity and specificity (88% and 92.31%, respectively).ConclusionsOur research shows that combination of CpG sites nt5606, nt5609, nt5615, and nt5378 can be used as potential diagnosis biomarkers for SCC, and the HRM PCR method is suitable for clinical methylation analysis.Electronic supplementary materialThe online version of this article (10.1186/s13148-018-0445-8) contains supplementary material, which is available to authorized users.

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“…Overall, the analysis of the methylation pattern of the integrate HPV16 and human host genome in cultured cells from HNSCC, including SCC from the base of the tongue, revealed that the methylation status of HPV16 is dramatically affected by the methylation status of the host DNA flanking the integration site with HPV16-DNA being highly methylated when integrated into intergenic highly methylated host genome sites, while remaining largely unmethylated when incorporated into poorly methylated intergenic regions [ 66 ]. This observation would argue for a bystander role of the virus methylome rather than an important phenomenon in HPV-driven carcinogenesis.…”

Section: Dna Methylationmentioning

confidence: 99%

Novel insights into epigenetic drivers of oropharyngeal squamous cell carcinoma: role of HPV and lifestyle factors

et al. 2017

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In the last years, the explosion of high throughput sequencing technologies has enabled epigenome-wide analyses, allowing a more comprehensive overview of the oropharyngeal squamous cell carcinoma (OPSCC) epigenetic landscape. In this setting, the cellular pathways contributing to the neoplastic phenotype, including cell cycle regulation, cell signaling, DNA repair, and apoptosis have been demonstrated to be potential targets of epigenetic alterations in OPSCC. Of note, it has becoming increasingly clear that HPV infection and OPSCC lifestyle risk factors differently drive the epigenetic machinery in cancer cells. Epigenetic changes, including DNA methylation, histone modifications, and non-coding RNA expression, can be used as powerful and reliable tools for early diagnosis of OPSCC patients and improve prognostication. Since epigenetic changes are dynamic and reversible, epigenetic enzymes may also represent suitable targets for the development of more effective OPSCC therapeutic strategies. Thus, this review will focus on the main known epigenetic modifications that can occur in OPSCC and their exploitation as potential biomarkers and therapeutic targets. Furthermore, we will address epigenetic alterations to OPSCC risk factors, with a particular focus on HPV infection, tobacco exposure, and heavy alcohol consumption.

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Identification of human papillomavirus (HPV) 16 DNA integration and the ensuing patterns of methylation in HPV‐associated head and neck squamous cell carcinoma cell lines

Cited by 22 publications

References 38 publications

Detailed Characteristics of Tonsillar Tumors with Extrachromosomal or Integrated Form of Human Papillomavirus

Detailed Characteristics of Tonsillar Tumors with Extrachromosomal or Integrated Form of Human Papillomavirus

Identification of reliable biomarkers of human papillomavirus 16 methylation in cervical lesions based on integration status using high-resolution melting analysis

Novel insights into epigenetic drivers of oropharyngeal squamous cell carcinoma: role of HPV and lifestyle factors

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