Abstract PS5-33: Androgen receptor gene expression and prediction of clinical outcomes of breast cancers exhibiting triple negative or triple positive breast carcinoma cells procured by LCM
Authors: Wittliff James L , Daniels Michael W. 
Published: February 2021

Abstract
Background: Clinical utility of AR, assessed by IHC of protein or by RT-qPCR of gene expression of intact tissue biopsies, has been the focus of many studies without uniform agreement. Due to cellular heterogeneity of most breast cancer biopsies, it is often unclear if measurements of AR and certain clinically relevant biomarkers accurately reflect their content in isolated cells. Using Laser Capture Microdissection (LCM) to procure only populations of breast carcinoma cells from tissue biopsies provides the inimitable opportunity to examine relationships of AR to a variety of parameters. Methods: We performed retrospective studies using a unique, de-identified dataset of ER/ESR1, PR/PGR and HER2/ERBB2 results with AR gene expression determined from LCM-procured carcinoma cells of biopsies from 247 breast cancer patients with associated clinical follow-up. ER/PR protein levels, expressed as fmol/mg cytosol protein, were quantified from each carcinoma biopsy with FDA-approved kits using enzyme immunoassay (EIA, Abbott Labs) or radio-ligand binding assay (NEN/DuPont). HER2 protein content of biopsies was quantified by EIA (Oncogene Sciences). Microarray analyses of ~22,000 genes were performed on RNA isolated, purified and amplified from LCM-procured carcinoma cells to assess relative gene expression. Biomarker results and de-identified clinical outcomes were examined using REMARK criteria in a CLIA licensed laboratory. Gene expression, quantified biomarkers, features of primary breast cancers and clinical outcomes were analyzed by univariable and multivariable Cox regressions, Fisher’s Exact Test, Kaplan Meier plots and with R software v4.0.0. Clinical relevance of gene expression was externally validated with SurvExpress. (Aguirre-Gamboa et al. PLoS One e742502013). Results: Using the median value of relative AR expression in LCM-procured carcinoma cells as a cutoff, elevated AR mRNA, when considered independently, was associated with the following (given as the means): older patients (61.1 vs 55.8 yo), relative expression of either ESR1 (0.2 vs -3.1), PGR (0.4 vs 0.0) or ERBB2 (0.7 vs -0.1) as well as longer PFS (63.7 vs 52.6 mos) and OS (72.4 vs 61.8 mos). Also, carcinoma cells with increased AR mRNA exhibited higher levels of ER protein (242.4 vs 106.5 fmol/mcp) or PR protein (313.8 vs 90.2 fmol/mcp). Whereas, HER2 protein status was not significantly different between AR median bifurcated groups). Neither patient race nor tissue pathology was examined in these analyses. Significantly, Kaplan Meier analyses clearly indicated that addition of AR gene expression to TNBC status (so called QNBC, Quadruple Negative Breast Cancer) did not alter either PFS (5 yr = 0.78 vs 0.79 % probability) nor OS (5 yr = 0.57 vs 0.56 %) of patients. Similarly, Kaplan Meier analyses also indicated that addition of AR gene expression to TPBC status (so called QPBC, Quadruple Positive Breast Cancer) did not alter PFS (5 yr = 0.87 vs 0.87 % probability) nor OS (5 yr = 0.67 vs 0.66 %) of patients. Also, no differences were ascertained for PFS nor for OS of patients at 10 yrs of outcomes between TNBC and QNBC breast cancers clearly indicating that AR gene expression does not contribute to the assessment of a patient’s clinical course. Conclusions: Collectively, when cancers were compared as TNBC versus TPBC status, expression results of ESR1, PGR and ERBB2 genes using LCM-procured carcinoma cells indicated poorer prognosis and overall survival of breast cancer patients with TNBC. However, inclusion of AR gene expression in either of these bifurcated groups ascertained from LCM-procured cells, did not alter patient PFS nor OS suggesting AR utility in clinical management appears to be unwarranted. Supported in part by grants to JLW from Phi Beta Psi Sorority Charity Trust.
Citation Format: James L Wittliff, Michael W. Daniels. Androgen receptor gene expression and prediction of clinical outcomes of breast cancers exhibiting triple negative or triple positive breast carcinoma cells procured by LCM [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 PS5-33.