title: Molecular characterization of p16-immunopositive but HPV DNA-negative oropharyngeal carcinomas.
abstract: Recent studies have reported that p16 protein overexpression qualifies as a surrogate marker identifying an oncogenic human papillomavirus (HPV) infection in oropharyngeal squamous cell carcinoma (OPSCC). However, there is still a percentage of OPSCCs that are positive for p16 immunohistochemistry (p16 IHC) but lack HPV DNA. The objective of this study was to characterize this group at the molecular level by performing sensitive HPV DNA- and RNA-based PCR methods and genetic profiling. All patients diagnosed with an OPSCC in the period 2000-2006 in two Dutch university medical centers were included (n = 841). The presence of HPV in a tumor sample was tested by p16 IHC followed by an HPV DNA GP5+/6+ PCR. p16 IHC scored positive in 195 samples, of which 161 were HPV DNA-positive and 34 (17%) HPV DNA-negative. In the latter group, a SPF10-LiPA25 assay, an HPV16 type-specific E7 PCR and an E6 mRNA RT-PCR were performed. Next, ten of these cases were further analyzed for loss of heterozygosity (LOH) of 15 microsatellite markers at chromosome arms 3p, 9p and 17p. Of the 34 p16-positive but PCR-negative OPSCCs, two samples tested positive by SPF10 assay, HPV16 E7 PCR and HPV16 E6 mRNA RT-PCR. Three samples tested positive by SPF10 assay but negative by the HPV16-specific assays. Nine of ten cases that were tested for LOH showed a genetic pattern comparable to that of HPV-negative tumors. This study categorizes p16-positive but HPV DNA-negative OPSCCs as HPV-negative tumors based on genetic profiling. This study highlights the importance of performing HPV testing in addition to p16 IHC for proper identification of HPV-associated OPSCCs.
authors.affiliation: Department of Otolaryngology/Head and Neck Surgery, VU University Medical Center, Amsterdam, The Netherlands.
authors.forename: Michelle M
authors.initials: MM
authors.lastname: Rietbergen
authors.forename: Peter J F
authors.initials: PJ
authors.lastname: Snijders
authors.forename: Derakshan
authors.initials: D
authors.lastname: Beekzada
authors.forename: Boudewijn J M
authors.initials: BJ
authors.lastname: Braakhuis
authors.forename: Arjen
authors.initials: A
authors.lastname: Brink
authors.forename: Daniëlle A M
authors.initials: DA
authors.lastname: Heideman
authors.forename: Albertus T
authors.initials: AT
authors.lastname: Hesselink
authors.forename: Birgit I
authors.initials: BI
authors.lastname: Witte
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Bloemena
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Baatenburg-De Jong
authors.forename: C René
authors.initials: CR
authors.lastname: Leemans
authors.forename: Ruud H
authors.initials: RH
authors.lastname: Brakenhoff
mesh_terms: D002294:Carcinoma, Squamous Cell / Q000235:genetics / Q000378:metabolism* / Q000821:virology; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000032:analysis*; D004252:DNA Mutational Analysis; D004279:DNA, Viral / Q000235:genetics*; D054884:Host-Pathogen Interactions; D052162:Human papillomavirus 16 / Q000235:genetics* / Q000502:physiology; D006801:Humans; D007150:Immunohistochemistry; D019656:Loss of Heterozygosity; D018895:Microsatellite Repeats / Q000235:genetics; D009154:Mutation; D009856:Oncogene Proteins, Viral / Q000235:genetics; D009959:Oropharyngeal Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000821:virology; D050725:Papillomavirus E7 Proteins / Q000235:genetics; D030361:Papillomavirus Infections / Q000235:genetics / Q000378:metabolism* / Q000821:virology; D016133:Polymerase Chain Reaction; D012367:RNA, Viral / Q000235:genetics; D012097:Repressor Proteins / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016159:Tumor Suppressor Protein p53 / Q000235:genetics
chemical_list: D019941:Cyclin-Dependent Kinase Inhibitor p16; D004279:DNA, Viral; C068651:E6 protein, Human papillomavirus type 16; D009856:Oncogene Proteins, Viral; D050725:Papillomavirus E7 Proteins; D012367:RNA, Viral; D012097:Repressor Proteins; D016159:Tumor Suppressor Protein p53; C059731:oncogene protein E7, Human papillomavirus type 16
keywords: HPV; LOH; p16 immunohistochemistry