title: Expression profiling of Epstein-Barr virus-encoded microRNAs from paraffin-embedded formalin-fixed primary Epstein-Barr virus-positive B-cell lymphoma samples.
abstract: Epstein-Barr virus (EBV) is implicated in a range of B-cell malignancies and expresses unique microRNAs (EBV-miRNAs). Due to the requirements for high-quality RNA, studies profiling EBV-miRNA in EBV-positive lymphomas have been restricted to cell-lines or frozen samples. However, the most commonly available archived patient material is paraffin-embedded formalin-fixed (FFPE) tissue. This has impeded the widespread profiling of EBV-miRNA expression in clinical samples. The requirements for accurate EBV-miRNA real-time RT-PCR quantitation in FFPE tissues representing a broad-spectrum of EBV-positive lymphomas were determined systematically, including where the neoplastic cells are sparse relative to the non-malignant infiltrate. The level of cellular EBV-load correlated strongly with the sum of EBV-miRNA expression and the number of EBV-miRNAs detectable. As calibrators for cellular EBV-load, the sum EBV-miRNA was optimal to EBV-genome copy number and EBER2 expression level, with the added advantage of not requiring additional assays. EBV-miRNA was profiled reliably within archival FFPE tissue in 14/23 patients, but not in tissues with low abundance EBV. This method enabled specific and simultaneous detection of numerous EBV-miRNAs in FFPE lymphoma samples that contain EBV at high to medium levels, making it as a useful tool for studies of EBV-miRNA in the majority of diagnostic biopsies.
authors.affiliation: Clinical Immunohaematology Laboratory, Queensland Institute of Medical Research, Brisbane, Australia.
authors.forename: Jamie P
authors.initials: JP
authors.lastname: Nourse
authors.forename: Pauline
authors.initials: P
authors.lastname: Crooks
authors.forename: Colm
authors.initials: C
authors.lastname: Keane
authors.forename: Do
authors.initials: D
authors.lastname: Nguyen-Van
authors.forename: Sally
authors.initials: S
authors.lastname: Mujaj
authors.forename: Nathan
authors.initials: N
authors.lastname: Ross
authors.forename: Kimberley
authors.initials: K
authors.lastname: Jones
authors.forename: Frank
authors.initials: F
authors.lastname: Vari
authors.forename: Erica
authors.initials: E
authors.lastname: Han
authors.forename: Ralf
authors.initials: R
authors.lastname: Trappe
authors.forename: Susanne
authors.initials: S
authors.lastname: Fink
authors.forename: Maher K
authors.initials: MK
authors.lastname: Gandhi
mesh_terms: D000328:Adult; D000368:Aged; D005260:Female; D005557:Formaldehyde / Q000494:pharmacology; D020869:Gene Expression Profiling; D004854:Herpesvirus 4, Human / Q000235:genetics*; D006801:Humans; D016393:Lymphoma, B-Cell / Q000821:virology*; D008297:Male; D035683:MicroRNAs / Q000096:biosynthesis / Q000235:genetics*; D008875:Middle Aged; D016612:Paraffin Embedding; D057089:Pathology, Molecular / Q000379:methods*; D012367:RNA, Viral / Q000096:biosynthesis / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D016707:Tissue Fixation
chemical_list: D035683:MicroRNAs; D012367:RNA, Viral; D005557:Formaldehyde