In situ quantitative measurement of mRNA to predict response to trastuzumab in a cohort of metastatic breast cancer patients.

Abstract

573 Background: Trastuzumab therapy is currently selected based on assessment of HER2 by either immunohistochemistry (IHC) for protein over-expression or fluorescence in situ hybridization (FISH) to detect gene amplification. We sought to determine the predictive value of in situ quantitative measurement of mRNA on a cohort of trastuzumab-treated metastatic breast cancer patients. Methods: A tissue microarray composed of ninety metastatic breast cancers treated with various chemotherapy regimens combined with trastuzumab was constructed. HER2 intracellular domain (ICD), HER2 extracellular domain (ECD) and HER2 mRNA were assessed using the AQUA method for quantitative immunofluorescence. For HER2 protein evaluation CB11was used to measure ICD and SP3 to measure ECD of the HER2 receptor. In addition, HER2 mRNA status was assessed using the RNAscope assay ERRB2 probe according to manufacturer’s protocol modified for detection with Cy-5 Tyramide. Cytokeratin was used in order to create the tumor mask and signal was quantified within the mask. Primary endpoints included time to progression (TTP) from trastuzumab initiation and overall survival (OS) times. Statistical analysis was performed using Kaplan-Meier analysis and the Cox proportional hazard model. Results: HER mRNA was tightly correlated with ICD HER2, as measured by CB11 (r2=0.35), but not with ECD HER2 as measured by SP3 (r2=0.14). Both ICD HER2 and HER2 mRNA were predictive of response to trastuzumab, but ECD was not. Multivariate analysis including age, histological grade and hormone receptor status shows the ICD to be predictive of TTP (p=0.0377). HER2 mRNA levels were significant for prediction of TTP (p=0.0344) in multivariate analysis including age and histological grade. Conclusions: The expression of ICD, as detected by CB11 and HER2 mRNA levels were significantly associated with TTP in this trastuzumab-treated metastatic cohort. The ECD, as detected by SP3 is neither predictive of response, nor tightly correlated with HER2 mRNA. In situ assessment of HER2 mRNA has the potential to identify breast cancer patients who derive benefit from trastuzumab treatment.