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Background  Until now, FISH has been the gold standard technique to identify  HER2  amplification status in ambiguous cases of breast cancer. Alternative techniques have been developed to increase the capacities of investigating  HER2  amplification status. The aims of this multicenter study in a large series of breast cancer patients were to prospectively compare the level of performance of CISH, SISH, and qPCR alternative techniques on paraffin-embedded core biopsies with “gold standard FISH” for evaluation of  HER2  amplification status.    Methods  This study was performed on 840 cases scored by immunohistochemistry (IHC): 0=317 (38%), 1+=183 (22%), 2+=109 (13%), 3+=231 (27%). Each of the 15 French centers participating in the study analyzed 56 breast carcinoma cases diagnosed on fixed paraffin-embedded core biopsies.  HER2  amplification status was determined by commercially available FISH used as the reference technique with determination of the HER2/CEN17 ratio or  HER2  copy number status. The alternative techniques performed on the same cases were commercially available SISH or CISH and a common qPCR method especially designed for the study including a set of 10 primer pairs: 2 for  HER2  (exons 8 and 26), 5 to evaluate chromosome 17 polysomy  TAOK1 ,  UTP6 ,  MRM1 ,  MKS1 ,  SSTR2  and 3 for diploidy control  TSN ,  LAP3  and  ADAMTS16 .    Results  The concordance between IHC and FISH was 96% to 95% based on the HER2/CEN17 ratio (n=766) or  HER2  copy number (n=840), respectively. The concordance of the alternative techniques with FISH was excellent: 97% and 98% for SISH (498 and 587 cases), 98% and 75% for CISH (108 and 204 cases) and 95% and 93% (699 and 773 cases) for qPCR based on the HER2/CEN17 ratio or  HER2  copy number, respectively. Similarly, sensitivity ranged from 99% to 95% for SISH, 100% to 99% for CISH and 89% to 80% for qPCR. The concordance with FISH (ratio) in the 2+ cases was 89% for SISH, 100% for CISH and 93% for qPCR.    Conclusion  These alternative techniques showed an excellent concordance with FISH in core biopsies allowing their use in routine clinical practice. This newly designed qPCR on paraffin-embedded core biopsies deserves special attention, as it is reliable, easy to perform and less expensive than ISH tests.

bmc cancer

SISH/CISH or qPCR as alternative techniques to FISH for determination of HER2 amplification status on breast tumors core needle biopsies: a multicenter experience based on 840 cases