Abstract PS18-25: Concordance of immunohistochemical assays between peri-operative and post-operative breast tumor specimens: A prospective observational study of 18 cases
Authors: Chang Martin C. , Ennis Marguerite , Escallon Jaime , Bukhanov Karina , Dowling Ryan JO , Stambolic Vuk , Goodwin Pamela J 
Published: February 2021

Abstract
Background and Rationale: Biomarker evaluation on breast tumor tissue is an important component of clinical research. There is a concern that tissues collected at different times (pre- vs intra-op) and with different techniques (core biopsy vs. surgical excision) produce different results that may confound comparisons of patient samples. Our objective is to address the difference between tumor tissue from the same patient collected pre- vs intra-op and by core biopsy (Core Bx) vs surgical excision, with an emphasis on ER/PgR/HER2/Ki-67 and biomarkers related to insulin metabolism.
Design: Following a protocol approved by the IRB, patients with Core Bx proven invasive breast cancer (BC) &gt;1.5 cm in size by imaging underwent a peri-operative Core Bx followed by surgical excision. No neoadjuvant therapy was administered. Formalin-fixed paraffin-embedded tumor sections of the diagnostic and peri-operative Core Bx and surgical excision were immunohistochemically (IHC) stained for ER, PgR, HER2, Ki-67, insulin receptor (IR), phospho-AKT (pAKT), and phospho-AMPK (pAMPK) using standardized protocols on the same platforms. A pathologist (MCC) scored all sections in blinded order to yield H-Scores (which combine percentage of stain-positive cells and staining intensity). The level of agreement for each assay between specimens was assessed by using pairwise models based on normal theory.
Results: 18 women (mean age = 66.8 years, 16/18 postmenopausal) provided all specimens. Mean (± SD) invasive tumor size was 2.7 (± 1.2) cm. 1 BC was Nottingham Grade 1 (6%), 9 Grade 2 (50%), and 8 Grade 3 (44%). An in situ component was present in 4/18 (22%) cases (all non-extensive). Lymphovascular invasion was present in 6/18 (33%) cases. 10/18 were node-negative (N0, 56%), 2/18 had isolated tumor cells (pN0[i+], 22%), 5/18 had 1 to 3 nodes (pN1, 28%), 1/18 had &gt;9 nodes involved (pN3, 6%).There was high concordance between diagnostic Core Bx, peri-op Core Bx, and excisions for the standard prognostic markers ER, PgR, and HER2. ER was concordant in all samples including 16/18 (89%) ER-positive and 2/19 (11%) ER-negative BCs. PgR was concordant in 16/18 (89%) of cases. In the 2 discordant cases, PgR was negative in 2 of 3 tissue samples with low expression in the 3rd sample (1 diagnostic Core Bx and 1 excision). HER2 was concordant in all samples in 16/18 (89%) HER2-negative cases and in 2/18 (11%) HER2-positive cases including 1/18 cases (6%) positive by IHC, and 1/18 cases (6%) positive by ISH.
Table 1 summarizes the agreement between samples for Ki-67, IR, pAKT, and pAMPK. Ki-67 scores were statistically similar between diagnostic and peri-op Core Bx and excision samples. The IR, pAKT, and pAMPK H-Scores were statistically similar between diagnostic and peri-op Core Bx, but significantly different between Core Bxs versus excisional specimens. There was a systematic tendency towards lower IHC H-Scores in the excisional specimen for IR, pAKT, and pAMPK.
Conclusion: Tissue from surgical excisions are susceptible to reduced IHC staining for metabolic markers such as IR, and phosphorylated kinases, when compared to core biopsies. When evaluating non-standard biomarkers for research, core biopsies should be used when possible.
Funding: This study was funded by the Hold'Em For Life Charity and the Breast Cancer Research Foundation
Table 1: Concordance Between IHC Scores for Ki-67, Insulin Receptor, pAKT, and pAMPKScore: Mean ± Standard DeviationPeri-Operative Core Bx versus Diagnostic Core BxExcision versus Diagnostic Core BxDiagnostic Core BxPeri-Operative Core BxExcisionDifferenceP-valueDifferenceP-valueKi-67 (% Positive)33.5 ± 28.235.9 ± 31.034.2 ± 29.1Mean ± SD: 2.4 ± 5.60.09Mean ± SD: 0.6 ± 6.30.68Range: -7.7 to 12.3Range: -11.5 to 12.2Insulin Receptor (H-Score**)136 ± 102124 ± 9986 ± 78Mean ± SD: -13 ± 530.32Mean ± SD: -51 ± 540.001*Range: -215 to 40Range: -150 to 5Phospho-AKT (H-Score**)102 ± 70100 ± 8641 ± 41Mean ± SD: -1 ± 750.95Mean ± SD: -61 ± 520.0001*Range: -170 to 170Range: -144 to 0Phospho-AMPK (H-Score**)201 ± 79185 ± 78157 ± 86Mean ± SD: -15 ± 760.41Mean ± SD: -44 ± 690.016*Range: -225 to 120Range: -215 to 70* Statistically significant difference in pairwise values based on t-test** H-Score is the percentage of stain-positive cells multiplied by the average intensity score (0 = absent, 1 = faint, 2 = moderate, 3 = strong)
Citation Format: Martin C. Chang, Marguerite Ennis, Jaime Escallon, Karina Bukhanov, Ryan JO Dowling, Vuk Stambolic, Pamela J Goodwin. Concordance of immunohistochemical assays between peri-operative and post-operative breast tumor specimens: A prospective observational study of 18 cases [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS18-25.