Abstract
Background: Assessment of HER-2/neu status in invasive breast cancer is crucial to establish eligibility for trastuzumab and taxane based chemotherapy. Next to immunohistochemistry (IHC) to evaluate protein overexpression, a second line gene amplification test is required for cases with equivocal protein expression. This study aimed to validate a new PCR based test, called Multiplex Ligation-dependent Probe Amplification (MLPA), as a simple and quick method to assess HER-2/neu gene amplification status in invasive breast cancer.Methods: MPLA results were compared with gene amplification status assessed by fluorescence in situ hybridization (FISH) and chromogenic in situ hybridization (CISH) as gold standard, and with protein overexpression by IHC in 518 breast carcinoma patients.Results: About 10% of cases overexpressed HER-2/neu at the protein level (IHC), and 11% of cases showed gene-amplification by MLPA. A high concordance was found between FISH and CISH, MLPA and IHC, and MLPA and CISH. MLPA showed amplification in 7/36 (19%) of the equivocal IHC 2+ cases. However, of the IHC 0/1+ cases, 6/434 (1.4%) were also amplified by MLPA, and amplification was confirmed in all of these cases by FISH/CISH. On the other hand, one of the 48 (2%) IHC 3+ cases was normal by MLPA and lack of amplification was confirmed by FISH/CISH.Conclusion: MLPA is a fast, accurate and cheap method to detect breast cancer HER-2/neu amplification in small quantities of DNA extracted from paraffin blocks, and thereby a reliable alternative to FISH and CISH.
Highlights
HER-2/neu is a proto-oncogene located on chromosome 17q21 that belongs to the human epidermal growth factor receptor (EGFR) family
HER-2 amplification status by multiplex ligation-dependent probe amplification (MLPA) was normal in 86% of cases, low level amplified in 3% and amplified in 11% of cases
1/48 (2%) IHC 3+ cases was normal by MLPA and lack of amplification was confirmed by fluorescence in situ hybridization (FISH)/Chromogenic in situ hybridization (CISH)
Summary
HER-2/neu is a proto-oncogene located on chromosome 17q21 that belongs to the human epidermal growth factor receptor (EGFR) family It encodes a 185 kD transmembrane protein that is involved in signal transduction [1,31]. Cinomas HER-2 is amplified and the expression of its receptor protein is increased [27,35,39] Such patients respond well to treatment with trastuzumab, a recombinant humanized monoclonal anti-HER-2 antibody [15, 40]. Assessment of HER-2/neu status in invasive breast cancer is crucial to establish eligibility for trastuzumab and taxane based chemotherapy. This study aimed to validate a new PCR based test, called Multiplex Ligation-dependent Probe Amplification (MLPA), as a simple and quick method to assess HER-2/neu gene amplification status in invasive breast cancer
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