Abstract 3354: Capturing tumor heterogeneity to personalize therapy in young patients with hormone receptor positive breast cancer
Authors: Segatto Ilenia , Mattevi Maria Chiara , Dall'Acqua Alessandra , Musco Lorena , Ruggiero Federica , Giustino Gabriele Di , Favero Andrea , Vinciguerra Gian Luca Rampioni , Crestan Nicole , Pellizzari Luca , Spizzo Riccardo , Bomben Riccardo , Vit Filippo , Vecchione Andrea , Gerratana Lorenzo , Puglisi Fabio , Massarut Samuele , Perin Tiziana , Baldassarre Gustavo , Belletti Barbara 
Published: March 2024

Abstract
Background: Breast cancer (BC) in young women (Y) is a distinct entity in terms of prognosis. Although triple negative and HER2 positive subtypes are enriched in Y patients, the hormone receptor (HR) positive subtype (luminal, LBC) remains the most common. A thorough understanding of LBC biology in Y women has not been reached yet, also hampered by lack of appropriate models that faithfully recapitulate these tumors in molecular and preclinical studies.
Methods: To investigate molecular determinants of progression and therapeutic resistance, we collected a large panel of fresh specimens (N=226) from Y (&lt; 40 years) and nY (&gt;55 years) LBC patients, characterized by targeted sequencing and used to generate Patient-Derived Organoids (PDO) and Xenografts (PDX). A selected panel of Y and nY LBC patients (N=96), sharing similar clinical-pathological features, was used for whole exome sequencing (WES) and transcriptomic analyses.
Results: We generated stable LBC PDO (N=32, 13 from Y patients), with a success rate of 50% and stable LBC PDX (N=15, 11 from Y patients), with a success rate 17%. Our targeted panel of NGS, covering most mutated genes in LBC at high coverage (1000x), revealed a significantly higher number of Y patients displaying mutations in PIK3CA and in PIK3CA+TP53. However, these data were not observable in WES analyses (200x), likely because many of those mutations displayed a VAF &lt; 5% (20% of PIK3CA and 30% of TP53). The transcriptomic analysis showed an enrichment in signatures of aggressiveness in the Y LBC subgroup. The EGF family member AREG (Amphiregulin) was the most enriched gene in tumors from pre- versus post-menopausal patients. Following validation in a larger panel of LBC, both at RNA and protein level, we tested the impact of AREG in LBC cell lines. AREG expression was timely induced by growth factor and hormonal stimulation. Furthermore, stimulation using amphiregulin induced the activation of EGFR and its downstream pathway, creating a positive feedback loop. Relevant to the Y LBC phenotype, a consistent and time dependent induction of AREG transcription was induced following estrogen and progesterone stimulation, indicating that, in LBC, AREG might act as a transcriptional target of ESR1. Finally, high levels of amphiregulin reduced response to tamoxifen and CDK4/6 inhibitors in LBC cells and PDOs.
Conclusions and Perspectives: Our study provides valuable molecular and biological insights into LBC in Y women. We identified distinct genetic and transcriptomic profiles, including PIK3CA and TP53 mutations and AREG overexpression, which may be exploited to plan more targeted and efficient treatments. The establishment of a living biobank of PDO and PDX from LBC patients of Y age will enable preclinical trials, facilitating discovery of biomarkers and therapeutic strategies tailored to the unique biology of Y LBC.
Citation Format: Ilenia Segatto, Maria Chiara Mattevi, Alessandra Dall'Acqua, Lorena Musco, Federica Ruggiero, Gabriele Di Giustino, Andrea Favero, Gian Luca Rampioni Vinciguerra, Nicole Crestan, Luca Pellizzari, Riccardo Spizzo, Riccardo Bomben, Filippo Vit, Andrea Vecchione, Lorenzo Gerratana, Fabio Puglisi, Samuele Massarut, Tiziana Perin, Gustavo Baldassarre, Barbara Belletti. Capturing tumor heterogeneity to personalize therapy in young patients with hormone receptor positive breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3354.