Abstract A22: Stromal cell signature in luminal breast cancer associated with response to neoadjuvant chemotherapy
Authors: Maria Lucia H. Katayama, Rene A da Costa Vieira, Rosimeire A. Roela, Victor P. Andrade, Luiz Guilherme C. A. de Lima, Giselly Encinas, Ligia M. Kerr, Simone Maistro, M. Mitzi Brentani, Maria A. A. Koike Folgueira
Journal: Clinical Cancer Research (2018)

Abstract Background: In locally advanced breast cancer, neoadjuvant chemotherapy may reduce tumor dimension and allow breast-conserving surgeries. However, it is not yet possible to predict which patients will benefit from neoadjuvant chemotherapy, as some patients may not experience tumor reduction, especially in luminal tumor samples. Tumor behavior reflects an intense crosstalk between malignant and stromal cells; however, little importance has been placed on the influence of the stromal cell in tumor response to chemotherapy. Objective: Our aim was to evaluate whether a stromal cell transcriptional signature might be associated with response to neoadjuvant anthracycline and taxane, in locally advanced luminal breast cancer, using a direct approach of stromal cell selection. Methodology: Twenty-nine patients diagnosed with locally advanced luminal breast cancer received neoadjuvant chemotherapy (doxorubicin and cyclophosphamide followed by paclitaxel). Breast cancer slices were submitted to laser-capture microdissection (LCM) to select stromal cells. Total RNA from stromal enriched cells were isolated using Arcturus PicoPure RNA isolation (Life technologies) according to the manufacturer's protocol. Linearly amplified two-round RiboAmp HSPlus2-round and Low Input Quick Amp (Agilent Technologies) protocols were adapted to optimize the amplification for the small numbers of cells produced by LCM. The labelled cRNAs were hybridized onto the Sure Print G3 (8×60K, Agilent Technologies). After having washed the slides, the arrays were scanned with the Agilent Bundle Model B Microarray Scanner System (Agilent Technologies). Scanned image files were visually inspected for artifacts, and the fluorescence intensities were extracted and preprocessed with Agilent Feature Extraction software (v10.7.1) and to normalize the results geneSpring GX12.1 software (Agilent Technologies) was used. Expression levels were compared using MultiExperiment Viewer (MeV) software, applying significance analysis of microarrays (SAM). Gene set enrichment analysis (GSEA) was used to identify whether predefined gene sets might associate with gene expression differences between phenotypes (http://software.broadinstitute.org/gsea/index.jsp). This methodology makes it possible to detect situations where all genes, in a predefined set, change in a small but coordinated way. Results: In estrogen receptor (ER) positive tumors, GSEA (FDR &amp;lt; 0.1) indicated that tumor down-staging was correlated with one KEGG gene set: antigen processing antigen presentation (comprehending CD74, CD8A, CD8B, and 9 molecules of HLA complex, among others) and with four Gene Ontology (GO) gene sets (biologic process), including one related with immune response: regulation of T cell activation (comprehending genes such as ZAP70, LCK, CD24, CD47, CD3E, among others). Gene sets associated with tumor non-down-staging (GSEA FDR &amp;lt; 0.25) were KEGG gene set: extracellular cell matrix (ECM) receptor interaction (including 5 types of integrins, 6 collagens, 4 laminins, fibronectin 1, and trombospondin 1), as well as seven GO gene sets (biologic process): cell adhesion; cell recognition; embryonic development; (cellular component): cortical cytoskeleton; cell projection part; cell surface; cytosolic part. Conclusion: A stromal signature associated with immune response in locally advanced luminal breast cancer may be associated with tumor down-staging following neoadjuvant chemotherapy. Financial Support: FAPESP 09/100088-7. Citation Format: Maria Lucia H. Katayama, Rene A da Costa Vieira, Rosimeire A. Roela, Victor P. Andrade, Luiz Guilherme C. A. de Lima, Giselly Encinas, Ligia M. Kerr, Simone Maistro, M. Mitzi Brentani, Maria A. A. Koike Folgueira. Stromal cell signature in luminal breast cancer associated with response to neoadjuvant chemotherapy [abstract]. In: Proceedings of the AACR International Conference held in cooperation with the Latin American Cooperative Oncology Group (LACOG) on Translational Cancer Medicine; May 4-6, 2017; São Paulo, Brazil. Philadelphia (PA): AACR; Clin Cancer Res 2018;24(1_Suppl):Abstract nr A22.