[2085] Detection of Her2/Neu Overexpression in Gastric Adenocarcinmas Using the Novel RNAScope Method

Aatur D Singhi, Yuling Luo, Hongwei Wang, Xiao-Jun Ma, Peter B Illei. University of Pittsburgh, Pittsburgh, PA; Advanced Cell Diagnostics, Inc, Hayward, CA; Johns Hopkins University, Baltimore, MD

Background: Approximately 20% of gastric adenocarcinomas are Her2 positive and may benefit from trastuzumab therapy. In gastric carcinoma Her2 protein expression and gene amplification dos not correlate as well as in breast carcinomas and therefore the use of more than one method is recommended. Her2 protein expression is strongly influenced by pre-analytical factors leading to false negative results. A robust RNA in-situ hybridization (ISH) assay using FFPE could represent an alternative method for accurately evaluating Her2/neu expression.
Design: Tissue microarrays of 55 cases of gastric adenocarcinomas were subjected to the RNAScope Her2 in situ hybridization method (Advanced Cell Diagnostics, Inc., Hayward, CA) and a previously established scoring method. Matching slides were also subjected to Her2 immunohistochemistry using Herceptest (Dako, Carpintera, CA) and the 4B5 monoclonal antibody (Ventana Medical Systems, Tucson, AZ) using standardized FDA approved protocols and scoring guidelines.
Results: Tumors were from 22 female and 33 male patients (28-81 years; mean 62.2 years). HER2 mRNA molecules were detected as punctate dots. Samples with >15 HER2 mRNA dots/cell were classified as positive. Seven tumors were positive by RNA ISH and 6 of them were positive by both IHC assay. RNA positive tumor was positive (3+) using the 4B5 antibody and equivocal (2+) using Herceptest. This case showed an average of 10 signal dots in 100% of the tumor cells. One more tumor showed similar RNA signaling (10 signals in 100% of tumor cells) and was negative by both immunohistochemistry method (score 0). One RNA ISH positive tumor was negative by Herceptest and one 4B5 positive tumor was negative by RNA ISH. All other cases were negative by all 3 assays.
Conclusions: Her2/neu protein expression evaluated by immunohistochemistry correlated well with increased Her2/neu RNA levels as assessed by the RNAScope RNA ISH method. The signals are crisp and are easy to read. Since the signal detection utilizes a chromogenic detection the signals can be localized to tumor cells (including high grade dysplasia). In contrast, this can be difficult to perform when using fluorescent in situ hybridization (FISH) especially in small biopsy samples where a large number of non-neoplastic cells can be present. Furthermore, the recent release of an automated method for performing the RNAScope assay (Ventana Discovery) has made the hybridization easier to perform and more reproducible.
Category: Techniques

Tuesday, March 5, 2013 2:30 PM

Proffered Papers: Section E, Tuesday Afternoon


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