Abstract P2-05-07: Comparison of the Xpert breast cancer stratifier mRNA assay with central ER, PR, HER2, and Ki67 immunohistochemistry (IHC) for rapid biomarker analysis in developing countries
Authors: Brock JE , Milner DA , Ho K , Natalie W , Victor C , Annaliza R , Teresa B , Kathryn G-F , Edwin LW , Jodi W , Wendy W , Michael B 
Published: May 2019

Abstract
Breast cancer care in the developing world is limited by access to quality ER and HER2 IHC diagnostic assays needed to justify hormone and HER2 therapeutics. Shipping pathology specimens to a central testing site often out of country delays therapy and is costly. The Xpert Breast Cancer Stratifier assay makes quantitative measurements of ESR1, PGR, ERBB2, and MKi67 mRNAs from FFPE specimens in &lt;2 hours on an easy-to-use automated diagnostic platform, the GeneXpert (GX). 10,000 GX machines are currently in use in 182 countries offering the possibility of a point-of-care solution. We compared concordance in tumor samples between IHC and mRNA intending to challenge the limits of the GX mRNA assay.
83 breast tumor samples were chosen including those with low cellularity, small volume disease, unusual subtypes, ER- tumors with surrounding benign epithelium, and low level HER2+ tumors. mRNA, IHC and FISH assays were performed. Slides were tested following macrodissection of invasive carcinoma and as non-macrodissected whole sections. GX measurements for Ki67 were compared with mitotic rate as an alternative to Ki67 IHC.
Overall percent agreement following macrodissection was 95% for ER, 89% for HER2, 76% for PR, and 80% for Ki67 (&gt;20% positive cut), and using whole section, 99% for ER, 80% for PR, 92% for HER2, and 73% for Ki67. Concordance was 92% for both macrodissection and whole section using mitotic rate to assess proliferation. Ignoring HER2 2+ calls which represented low level amplified tumors by FISH, the concordance rates were 95% for macrodissection and 99% for whole section. Discordance when testing long-term stored 4μm sections was resolved in a number of cases by using a fresh cut from the FFPE block. Half the ER discrepancies were in very small volume tumors ≤25mm2 and 75% were classified as ER-ve by IHC, and positive by Stratifier. 80% of ER IHC- cases were appropriately identified as ER- by the Stratifier in the presence of benign breast epithelium. HER2+ DCIS adjacent to HER2- invasive tumor resulted in a discrepant HER2 mRNA result even with macrodissection. No ER or HER2 discrepancies occurred in low cellularity tumors (≤30% cellularity) nor in lobular and mucinous subtypes.
In a study intended to challenge an mRNA breast biomarker assay, concordance between mRNA results and IHC was high for ER and HER2, the two most important prognostic markers needed for therapeutic decision making. Use of whole sections rather than tumor macrodissection did not decrease concordance. Discrepant ER cases were more prevalent when analyzing low volumes of tumor and in this setting were seen in ER IHC- tumors surrounded by ER+ normal epithelium, or with weak IHC expression, highlighting predictable limitations of the assay. Concordance was better between Ki67 mRNA and mitotic rate than with IHC. Re-test data suggested that a fresh cut of the FFPE block yields the best results by GX, perhaps due to mRNA degradation in stored 4μm sections. The Xpert Breast Cancer Stratifier may provide a rapid, cost-effective solution to the problem of obtaining accurate diagnostic results at the point-of-care in low resource settings, and deserves further evaluation in developing countries.
Citation Format: Brock JE, Milner DA, Ho K, Natalie W, Victor C, Annaliza R, Teresa B, Kathryn G-F, Edwin LW, Jodi W, Wendy W, Michael B. Comparison of the Xpert breast cancer stratifier mRNA assay with central ER, PR, HER2, and Ki67 immunohistochemistry (IHC) for rapid biomarker analysis in developing countries [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P2-05-07.