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Viral load, integration and methylation of E2BS3 and 4 in human papilloma virus (HPV) 16-positive vaginal and vulvar carcinomas.

Lillsunde Larsson G, Helenius G, Sorbe B, Karlsson MG - PLoS ONE (2014)

Bottom Line: Fifty-seven samples, positive for HPV16, were selected for the study.Vaginal tumors were found to have a higher viral load (p = 0.024) compared to vulvar tumors but a high copy number (> median value, 15,000) as well as high methylation (>50%) was significantly (p = 0.010 and p = 0.045) associated with a worse cancer-specific survival rate in vulvar carcinoma, but not in vaginal carcinoma.HPV16- related parameters were found to be of clinical importance in the vulvar series only.

View Article: PubMed Central - PubMed

Affiliation: Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden; School of Health and Medical Sciences, Örebro University, Örebro, Sweden.

ABSTRACT

Objective: To investigate if viral load, integration and methylation of E2BS3 and 4 represent different ways of tumor transformation in vaginal and vulvar carcinoma and to elucidate its clinical impact.

Methods: Fifty-seven samples, positive for HPV16, were selected for the study. Detection of viral load was made with realtime-PCR using copy numbers of E6 and integration was calculated from comparing E2 to E6-copies. Methylation of E2BS3 and 4 was analysed using bisulphite treatment of tumor DNA, followed by PCR and pyrosequencing.

Results: Vaginal tumors were found to have a higher viral load (p = 0.024) compared to vulvar tumors but a high copy number (> median value, 15,000) as well as high methylation (>50%) was significantly (p = 0.010 and p = 0.045) associated with a worse cancer-specific survival rate in vulvar carcinoma, but not in vaginal carcinoma. Four groups could be defined for the complete series using a Cluster Two step analysis; (1) tumors holding episomal viral DNA, viral load below 150,000 copies not highly methylated (n = 25, 46.3%); (2) tumors harboring episomal viral DNA and being highly methylated (>50%; n = 6, 11.1%); (3) tumors with viral DNA fully integrated (n = 11, 20.4%), and (4) tumors harboring episomal viral DNA and being medium- or unmethylated (<50%) and having a high viral load (> total mean value 150,000; n = 12, 22.2%). The completely integrated tumors were found to be distinct group, whilst some overlap between the groups with high methylation and high viral load was observed.

Conclusion: HPV16- related integration, methylation in E2BS3 and 4 and viral load may represent different viral characteristics driving vaginal and vulvar carcinogenesis. HPV16- related parameters were found to be of clinical importance in the vulvar series only.

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Related in: MedlinePlus

Cluster analysis data plotted in a 3-D box.Cluster identification: 1 = Mixed group (n = 25), 2 = Methylation >50% (n = 6), 3 = Integrated viral DNA (n = 11) and 4 = Viral load >150 000 (n = 12). Integrated tumors were found to be exclusive from the other groups. Overlap was seen between 4 highly methylated tumors that also had a high viral load (group 2).
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pone-0112839-g003: Cluster analysis data plotted in a 3-D box.Cluster identification: 1 = Mixed group (n = 25), 2 = Methylation >50% (n = 6), 3 = Integrated viral DNA (n = 11) and 4 = Viral load >150 000 (n = 12). Integrated tumors were found to be exclusive from the other groups. Overlap was seen between 4 highly methylated tumors that also had a high viral load (group 2).

Mentions: The classification seemed to be distinct for tumors that were integrated since none of them were highly methylated or having a high viral load. There were however some overlap between the tumor groups with high methylation in E2BS3 and 4 and a high viral load, table 3 and Figure 3. Tumors belonging to the first group all presented a low viral load. They could further be grouped as either mixed episomal/integrated and medium methylated (n = 2) or mixed episomal/integrated and unmethylated (n = 5). Also the fully episomal tumors were either medium methylated (n = 1) or unmethylated (n = 17).


Viral load, integration and methylation of E2BS3 and 4 in human papilloma virus (HPV) 16-positive vaginal and vulvar carcinomas.

Lillsunde Larsson G, Helenius G, Sorbe B, Karlsson MG - PLoS ONE (2014)

Cluster analysis data plotted in a 3-D box.Cluster identification: 1 = Mixed group (n = 25), 2 = Methylation >50% (n = 6), 3 = Integrated viral DNA (n = 11) and 4 = Viral load >150 000 (n = 12). Integrated tumors were found to be exclusive from the other groups. Overlap was seen between 4 highly methylated tumors that also had a high viral load (group 2).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4231157&req=5

pone-0112839-g003: Cluster analysis data plotted in a 3-D box.Cluster identification: 1 = Mixed group (n = 25), 2 = Methylation >50% (n = 6), 3 = Integrated viral DNA (n = 11) and 4 = Viral load >150 000 (n = 12). Integrated tumors were found to be exclusive from the other groups. Overlap was seen between 4 highly methylated tumors that also had a high viral load (group 2).
Mentions: The classification seemed to be distinct for tumors that were integrated since none of them were highly methylated or having a high viral load. There were however some overlap between the tumor groups with high methylation in E2BS3 and 4 and a high viral load, table 3 and Figure 3. Tumors belonging to the first group all presented a low viral load. They could further be grouped as either mixed episomal/integrated and medium methylated (n = 2) or mixed episomal/integrated and unmethylated (n = 5). Also the fully episomal tumors were either medium methylated (n = 1) or unmethylated (n = 17).

Bottom Line: Fifty-seven samples, positive for HPV16, were selected for the study.Vaginal tumors were found to have a higher viral load (p = 0.024) compared to vulvar tumors but a high copy number (> median value, 15,000) as well as high methylation (>50%) was significantly (p = 0.010 and p = 0.045) associated with a worse cancer-specific survival rate in vulvar carcinoma, but not in vaginal carcinoma.HPV16- related parameters were found to be of clinical importance in the vulvar series only.

View Article: PubMed Central - PubMed

Affiliation: Department of Laboratory Medicine, Örebro University Hospital, Örebro, Sweden; School of Health and Medical Sciences, Örebro University, Örebro, Sweden.

ABSTRACT

Objective: To investigate if viral load, integration and methylation of E2BS3 and 4 represent different ways of tumor transformation in vaginal and vulvar carcinoma and to elucidate its clinical impact.

Methods: Fifty-seven samples, positive for HPV16, were selected for the study. Detection of viral load was made with realtime-PCR using copy numbers of E6 and integration was calculated from comparing E2 to E6-copies. Methylation of E2BS3 and 4 was analysed using bisulphite treatment of tumor DNA, followed by PCR and pyrosequencing.

Results: Vaginal tumors were found to have a higher viral load (p = 0.024) compared to vulvar tumors but a high copy number (> median value, 15,000) as well as high methylation (>50%) was significantly (p = 0.010 and p = 0.045) associated with a worse cancer-specific survival rate in vulvar carcinoma, but not in vaginal carcinoma. Four groups could be defined for the complete series using a Cluster Two step analysis; (1) tumors holding episomal viral DNA, viral load below 150,000 copies not highly methylated (n = 25, 46.3%); (2) tumors harboring episomal viral DNA and being highly methylated (>50%; n = 6, 11.1%); (3) tumors with viral DNA fully integrated (n = 11, 20.4%), and (4) tumors harboring episomal viral DNA and being medium- or unmethylated (<50%) and having a high viral load (> total mean value 150,000; n = 12, 22.2%). The completely integrated tumors were found to be distinct group, whilst some overlap between the groups with high methylation and high viral load was observed.

Conclusion: HPV16- related integration, methylation in E2BS3 and 4 and viral load may represent different viral characteristics driving vaginal and vulvar carcinogenesis. HPV16- related parameters were found to be of clinical importance in the vulvar series only.

Show MeSH
Related in: MedlinePlus