Does HPV RNA Chromogenic In Situ Hybridization (CISH) Discriminate between Low and High Grade Cervical Squamous Intraepithelial Lesions (SIL)?
Kelli M Clark, Mark F Evans, Xiao-Jun Ma, Xingyong Wu, Yuling Luo, Zhihua Peng, Kumarasen Cooper. Fletcher Allen Health Care, Burlington, VT; University of Vermont, Burlington, VT; Advanced Cell Diagnostics, Inc., Hayward, CA
Background: We previously demonstrated that HPV DNA in situ hybridization signal patterns distinguish CIN1 from CIN2/3 lesions. A diffuse intranuclear signal represents episomal virus, whilst a punctate signal represents integrated HPV DNA. This study explores a potential grading discriminatory role of a novel HPV RNA CISH assay in cervical SILs.
Design: 40 formalin-fixed, paraffin-embedded cervical biopsies were reviewed (KMC, KC) and the following diagnoses confirmed: 20 CIN1, 10 CIN2, and 10 CIN3. Samples were screened by RNAscope® CISH for high-risk HPV E6/E7 (18 types). CISH for ubiquitin C RNA served as a positive control. The RNA probe for Bacillus subtilis gene dapB served as a negative control.
Results: Tissue was exhausted in 4 lesions. HPV E6/E7 RNA was detected in 100% of the CIN1 and CIN2/3 lesions. Positive staining was defined as granular cytoplasmic and/or nuclear brown staining stronger than the background signal in non-lesional tissues. Two distinct staining patterns emerged: 15/18 CIN1, 8/9 CIN2 and 2/9 CIN3 lesions showed abundant finely granular nuclear and cytoplasmic signals in the lower half to two-thirds of the epithelium, with a distinctive segregation of diffuse strong nuclear staining in the superficial third to one-half of the epithelium (type I signal pattern). One CIN2 and 7/9 of the CIN3 lesions showed abundant finely granular nuclear and cytoplasmic signals throughout the entire epithelium, with focal randomly distributed diffuse nuclear staining (type II signal pattern). The remaining 3/18 CIN1 lesions showed granular nuclear and cytoplasmic signals in the lower half of the epithelium.
Conclusions: The CIN1/2 lesions (type I pattern) confirmed the productive (episomal) cycle with low level viral replication (granular signal) in the lower third to one-half of the SIL, with viral genome amplification (diffuse signal) in the upper half to two-thirds supporting viral assembly and packaging of episomes. In contrast, the majority of CIN3 cells (type II pattern) poorly supported viral gene amplification, were scarce and varied in their location and distribution, consistent with the proliferative/transformative (integrated) cycle. The unexpected finding of the majority of CIN2 lesions aggregating with the CIN1 RNA CISH pattern raises biological questions regarding potential progression (regression vs persistence), prognostic, and management implications for CIN2.
Category: Gynecologic & Obstetrics
Tuesday, March 20, 2012 2:00 PM
Platform Session: Section B, Tuesday Afternoon