title: Performance of a Branch Chain RNA In Situ Hybridization Assay for the Detection of High-risk Human Papillomavirus in Head and Neck Squamous Cell Carcinoma.
abstract: High-risk human papillomavirus (HR-HPV) is a major etiologic agent in a subset of head and neck squamous cell carcinomas (HNSCCs), and its recognition has prognostic and predictive implications. The availability of a sensitive and specific test to assess HR-HPV status is limited. We evaluate an RNA in situ hybridization (ISH) method using branch chain technology to detect HR-HPV and compare its results with DNA ISH, p16 immunohistochemistry, and polymerase chain reaction (PCR). Tissue sections from 54 patients were stained with a manual RNA ISH assay (ViewRNA), which detects 14 HR-HPV types, an automated DNA ISH assay, and p16 immunohistochemistry. Most cases (83%, n=45) were also tested on an automated platform for 14 HR-HPV types and 1 limited to HPV 16/18. PCR was performed in all cases and was successful in 93% (n=50). The RNA ISH assay produced results in 96% of the cases with strong signals and was easily interpreted. HR-HPV was detected in more cases (63%, n=34) by RNA ISH than by DNA ISH (39%, n=21). Compared with PCR, both ISH platforms were 94% specific. RNA ISH was more sensitive (91%) than DNA ISH (65%), and RNA ISH correlated more strongly with p16 immunostaining. HPV 16 represented 89% of HR-HPV detected. The cocktail HPV 16/18 platform was concordant with the pooled HR-HPV assay in all expected cases. The automated assay demonstrated high concordance (96%) with the manual version, showed decreased background, and should allow for easy implementation into the workflow of the diagnostic pathology laboratory.
authors.affiliation: Departments of *Pathology ‡Surgery ∥Tucker Gosnell Center for Gastrointestinal Cancers, Massachusetts General Hospital §Department of Pathology, Brigham and Women's Hospital †Harvard Medical School, Boston, MA.
authors.forename: Darcy A
authors.initials: DA
authors.lastname: Kerr
authors.forename: Kshitij S
authors.initials: KS
authors.lastname: Arora
authors.forename: Krishnan K
authors.initials: KK
authors.lastname: Mahadevan
authors.forename: Jason L
authors.initials: JL
authors.lastname: Hornick
authors.forename: Jeffrey F
authors.initials: JF
authors.lastname: Krane
authors.forename: Miguel N
authors.initials: MN
authors.lastname: Rivera
authors.forename: David T
authors.initials: DT
authors.lastname: Ting
authors.forename: Vikram
authors.initials: V
authors.lastname: Deshpande
authors.forename: William C
authors.initials: WC
authors.lastname: Faquin
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D057205:Automation, Laboratory; D014408:Biomarkers, Tumor / Q000032:analysis; D001706:Biopsy; D001900:Boston; D002294:Carcinoma, Squamous Cell / Q000737:chemistry / Q000473:pathology / Q000821:virology*; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000032:analysis; D004279:DNA, Viral / Q000235:genetics; D005260:Female; D006258:Head and Neck Neoplasms / Q000737:chemistry / Q000473:pathology / Q000821:virology*; D061809:Human Papillomavirus DNA Tests; D052162:Human papillomavirus 16 / Q000235:genetics*; D052161:Human papillomavirus 18 / Q000235:genetics*; D006801:Humans; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D008297:Male; D008875:Middle Aged; D025202:Molecular Diagnostic Techniques; D030361:Papillomavirus Infections / Q000378:metabolism / Q000473:pathology / Q000821:virology*; D011237:Predictive Value of Tests; D012367:RNA, Viral / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D015203:Reproducibility of Results; D000077195:Squamous Cell Carcinoma of Head and Neck; D020298:U937 Cells; D057188:Workflow
chemical_list: D014408:Biomarkers, Tumor; D019941:Cyclin-Dependent Kinase Inhibitor p16; D004279:DNA, Viral; D012367:RNA, Viral