Quantitation of truncated HER2 in formalin-fixed paraffin-embedded breast carcinoma tissues.

Abstract

e11130 Background: Trastuzumab treatment of Her2+ breast cancer is the benchmark success story for targeted cancer therapy, however, a significant number of patients are either non-responsive or experience disease progression. A novel mechanism of resistance in some patients is the expression of truncated Her2 (p95Her2), formed by proteolytic cleavage of the HER2 molecule. Truncated p95Her2 loses its extracellular trastuzumab binding epitope, but retains a constitutively active kinase domain predicted to be sensitive to kinase inhibitors, like lapatinib and neratinib. At present there are no diagnostic assays for this p95Her2. Methods: Using the Liquid Tissue-SRM (selective reaction monitoring) platform we have developed multiple mass spectrometric assays to quantify specific cytoplasmic or extracellular domain (ECD) peptides from Her2 directly from formalin fixed paraffin embedded (FFPE) clinical tissue. Results: We validated these assays using recombinant human Her2, where we saw equivalent intracellular and extracellular Her2 levels. In formalin fixed SkBr3 breast carcinoma cells, we also detected equivalent levels of the Her2 peptides, as expected. We initiated clinical validation of this assay by analyzing a dozen FFPE human breast carcinoma tumors where Her2 expression and gene amplification levels had been determined. In 9 of 12 tumors (IHC: 1+ to 3+) we measured equivalent levels of intracellular and ECD peptides. Most interesting, 3 tumors showed dramatic differences in the levels of Her2 peptides detected. In two 3+ tumors, the intracellular peptide was 50% higher than the ECD peptide and in the last tumor more than twice the expression of the ECD Her2 peptide. These data suggest that in these 3 human tumors Her2 is missing a significant percent of the ECD, and is truncated. Continuing our clinical validation work we are analyzing larger numbers of Her2 positive breast tumors from clinical cohorts capable of confirming the clinical usefulness of the p95Her2 assay. Conclusions: Measuring Her2 truncation in Her2 positive breast tumors will benefit patients by enabling clinicians to select patient more likely to benefit to TKI (lapatinib, neratinib) therapy and will help physicians to make earlier informed therapeutic decisions.