HER-2/Neu Status Can Be Reliably Determined in Cytologic Breast Cancer Specimens Using Bright-Field Microscopy and Silver In-Situ Hybridization (SISH).
Zeina Ghorab, Reda S Saad, Shawna Noy, Wedad M Hanna, Sharon Nofech-Mozes. Sunnybrook Health Sciences Centre, Toronto, ON, Canada
Background: Accurate HER-2 testing of all breast cancer patients at primary diagnosis is essential for optimal disease management. ASCO/CAP guideline recommendations for HER-2 testing emphasize the need for standardization of preanalytical variables, especially formalin fixation. Accordingly, any change in testing protocol should be validated. Alcohol fixation may alter immunoreactivity; however there is only limited data on reliability of in situ hybridization in alcohol fixed cytologic specimen (CS). This study was designed to validate SISH in alcohol fixed cell blocks.
Design: We identified 50 pairs of CS positive for malignant cells and a corresponding breast cancer surgical specimen. This set of cases was specifically enriched in HER-2 positive cases. The gold standard for validation was HER-2 status that was determined on the FFPE surgical specimens by IHC. Equivocal IHC cases were further tested by FISH or SISH. CS were fixed in alcohol based preparation: 48 Saccomanno Collection Fluid and 2 in Cytolyt. HER-2 gene amplification status was determined on cell blocks using the Ventana Inform HER-2 SISH kit (Tucson, Arizona). Amplification was determined when the SISH HER-2 /CEP17 ratio was greater than 2.2. Two pathologists reviewed the CS and determined the SISH ratio and quantify malignant cells in a 3 tiered system (cellularity:1= 1-20 cells, 2=21-60 cells, 3=>60 cells). The concordance between HER-2 testing by SISH on CS was compared to that of FFPE surgical specimens.
Results: There were 42 cases with 3+ cellularity, 5 cases with 2+ and only 2 cases with 1+ cellularity (both were amplified). SISH reaction was successful in 49/50 (98%) cases. In one case, the test was uninterpretable due to poor HER-2 signal. All 22 Her2-positive surgical cases were CS-amplified and all 27 Her2-negative surgical cases were CS-not amplified. Overall there was a 100% concordance between Her2 status determined on alcohol fixed CS by SISH when compared with FFPE surgical specimens.
Conclusions: SISH reaction was successful in 98% of the cases. Our study validated SISH as a reliable accurate test for Her2 on alcohol fixed cytologic specimens. SISH allows for bright field microscopy which is advantageous over dark field traditional florescence technique, in particular in CS even with low cellularity.
Tuesday, March 1, 2011 1:00 PM
Platform Session: Section D, Tuesday Afternoon