P4-09-04: Do Decreased CEP17 Signals Indicate the Presence of “Monosomy” in Breast Cancer? A Study of HER2 Gene Status and HER2 Immunohistochemistry.
Authors: Tse CH , Hwang HC , Goldstein LC , Kandalaft PL , Duesterhoeft SM , Kussick SL , Gown AM 
Published: June 2012

Abstract
Background:
Detection of HER2 gene amplification by fluorescence in-situ hybridization (FISH) is one of the key tests predicting clinical response of breast cancer to trastuzumab. According to the recent ASCO/CAP guidelines for HER2 testing, the tumor is considered positive for HER2 gene amplification when the HER2/CEP17 ratio is greater than 2.2, negative if less than 1.8, and equivocal if the ratio is between 1.8 and 2.2. The need to use CEP17 as reference for HER2 assumes that the copy number of CEP17 is equivalent to the copy number of chromosome 17. Recent studies, however, have shown that this may not be true in cases of increased CEP17 signals interpreted as “polysomy”. This study aims to examine the effect of decreased CEP17 signals interpreted as “monosomy” on HER2 protein expression and gene amplification, and to test the hypothesis that a subset of these cases may be inappropriately classified as amplified or equivocal.
Material and Methods:
59 cases tested in parallel by IHC and FISH were selected based on HER2 signals &gt; 2 and CEP17 signals &lt; 1.5. To further examine chromosome 17 copy number, FISH mapping studies were performed on 5 cases, using probes for SMS, RARA, and TP53 loci which are telomeric to the CEP17 and HER2 loci. Chromosome 17 status was determined by correlating the mean copy numbers of HER2, CEP17, SMS, RARA, and TP53. A gene locus with signal number/cell between 1.4 and 2.4 was considered consistent with eusomy.
Results:
The 59 cases were divided into three groups based on HER2 signals/cell. Group 1, comprising 11 cases, had HER2 signals &gt; 4; all were amplified with HER2/CEP17 ratios between 3 and 5; 10 of these 11 cases showed equivocal IHC (2+). Group 2, comprising 17 cases, had HER2 signals &gt; 3 and &lt; 4; all were HER2 amplified with HER2/CEP17 ratios between 2.3 and 3.6, and 15 showed equivocal IHC (2+). Group 3, comprising 32 cases, had HER2 signals &gt; 2 and &lt; 3; 19 of these showed HER2/CEP17 ratios in the equivocal range (1.8−2.2) and 13 were amplified with ratios between 2.3 and 2.8; 25 showed equivocal IHC (2+), and 7 were negative by IHC (0,1+). Five cases were further tested using FISH probes to alternative chromosome 17 genes (SMS, RARA, and TP53). The number of CEP17 signals/cell of these 5 cases ranged from 1.01 to 1.15 and the HER2 signals/cell ranged from 2.05 to 2.84. In these cases when additional probes to chromosome 17 were used as reference genes, 4/5 cases remained amplified or equivocal. One case with HER2 signals of 2.05 changed from equivocal to negative. 4/5 of these cases showed equivocal IHC (2+).
Discussion:
Application of strict ASCO-CAP guidelines on 59 “monosomy” cases showed that all were either amplified or equivocal by FISH, with the vast majority showing equivocal (2+) IHC. In cases with decreased CEP17 signals, most cases remained as “monosomy” using the alternative chromosome 17 reference genes. This may be due to deletion of the short arm of chromosome 17 or true monosomy of chromosome 17. Further studies are warranted on the cases with 2–4 HER2 signals per cell and “monosomy” in order to permit accurate determination of HER2 gene status and appropriate treatment with anti-HER2 therapy.
Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P4-09-04.