title: Viral load, gene expression and mapping of viral integration sites in HPV16-associated HNSCC cell lines.
abstract: HPV-related HNSCC generally have a better prognosis than HPV-negative HNSCC. However, a subgroup of HPV-positive tumors with poor prognosis has been recognized, particularly related to smoking, EGFR overexpression and chromosomal instability. Viral integration into the host genome might contribute to carcinogenesis, as is shown for cervical carcinomas. Therefore, all HPV16-positive HNSCC cell lines currently available have been carefully analyzed for viral and host genome parameters. The viral integration status, viral load, viral gene expression and the presence of aneusomies was evaluated in the cell lines UD-SCC-2, UM-SCC-047, UM-SCC-104, UPCI:SCC090, UPCI:SCC152, UPCI:SCC154 and 93VU147T. HPV integration was examined using FISH, APOT-PCR and DIPS-PCR. Viral load and the expression of the viral genes E2, E6 and E7 were determined via quantitative PCR. All cell lines showed integration-specific staining patterns and signals indicating transcriptional activity using FISH. APOT- and DIPS-PCR identified integration-derived fusion products in six cell lines and only episomal products for UM-SCC-104. Despite the observed differences in viral load and the number of viral integration sites, this did not relate to the identified viral oncogene expression. Furthermore, cell lines exhibited EGFR expression and aneusomy (except UPCI:SCC154). In conclusion, all HPV16-positive HNSCC cell lines showed integrated and/or episomal viral DNA that is transcriptionally active, although viral oncogene expression was independent of viral copy number and the number of viral integration sites. Because these cell lines also contain EGFR expression and aneusomy, which are parameters of poor prognosis, they should be considered suitable model systems for the development of new antiviral therapies.
authors.affiliation: Department of Otorhinolaryngology and Head and Neck Surgery, GROW-School for Oncology and Developmental Biology, Maastricht University Medical Centre, The Netherlands; Department of Molecular Cell Biology, GROW-School for Oncology and Developmental Biology, Maastricht University Medical Centre, The Netherlands.
authors.forename: Nadine C
authors.initials: NC
authors.lastname: Olthof
authors.forename: Christian U
authors.initials: CU
authors.lastname: Huebbers
authors.forename: Jutta
authors.initials: J
authors.lastname: Kolligs
authors.forename: Mieke
authors.initials: M
authors.lastname: Henfling
authors.forename: Frans C S
authors.initials: FC
authors.lastname: Ramaekers
authors.forename: Iris
authors.initials: I
authors.lastname: Cornet
authors.forename: Josefa A
authors.initials: JA
authors.lastname: van Lent-Albrechts
authors.forename: Alexander P A
authors.initials: AP
authors.lastname: Stegmann
authors.forename: Steffi
authors.initials: S
authors.lastname: Silling
authors.forename: Ulrike
authors.initials: U
authors.lastname: Wieland
authors.forename: Thomas E
authors.initials: TE
authors.lastname: Carey
authors.forename: Heather M
authors.initials: HM
authors.lastname: Walline
authors.forename: Susanne M
authors.initials: SM
authors.lastname: Gollin
authors.forename: Thomas K
authors.initials: TK
authors.lastname: Hoffmann
authors.forename: Johan
authors.initials: J
authors.lastname: de Winter
authors.forename: Bernd
authors.initials: B
authors.lastname: Kremer
authors.forename: Jens P
authors.initials: JP
authors.lastname: Klussmann
authors.forename: Ernst-Jan M
authors.initials: EJ
authors.lastname: Speel
mesh_terms: D002294:Carcinoma, Squamous Cell / Q000235:genetics* / Q000473:pathology / Q000821:virology; D004279:DNA, Viral / Q000235:genetics; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism; D005260:Female; D020869:Gene Expression Profiling; D006258:Head and Neck Neoplasms / Q000235:genetics* / Q000473:pathology / Q000821:virology; D052162:Human papillomavirus 16 / Q000235:genetics* / Q000302:isolation & purification; D006801:Humans; D007124:Immunoenzyme Techniques; D017404:In Situ Hybridization, Fluorescence; D008297:Male; D008875:Middle Aged; D009856:Oncogene Proteins, Viral / Q000235:genetics*; D030361:Papillomavirus Infections / Q000235:genetics* / Q000473:pathology / Q000821:virology; D016133:Polymerase Chain Reaction; D014407:Tumor Cells, Cultured; D019562:Viral Load; D016662:Virus Integration / Q000235:genetics*
chemical_list: D004279:DNA, Viral; D009856:Oncogene Proteins, Viral; C512478:EGFR protein, human; D066246:ErbB Receptors
keywords: APOT-PCR; DIPS-PCR; genetic localization; integration; viral integration
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