Abstract P4-09-05: Characterization and validation of biologically-driven HER2-positive breast cancer subgroups in the ALTTO and NeoALTTO clinical trials
Authors: Rediti Mattia , Venet David , Garcia Andrea Joaquin , Agbor-Tarh Dominique , Maetens Marion , Vincent Delphine , Majjaj Samira , El-Abed Sarra , Ueno Takayuki , Cosimo Serena Di , Piccart Martine , Pusztai Lajos , Loi Sherene , Salgado Roberto , Viale Giuseppe , Rothé Françoise , Sotiriou Christos 
Published: March 2023

Abstract
Background: Tumor and microenvironment features, including luminal phenotype as well as metabolic, immune and stroma activation, impact prognosis and treatment response in HER2-positive breast cancer. Here, we aimed to identify subgroups depicting biological processes associated with prognosis in patients receiving adjuvant trastuzumab in the phase III ALTTO trial, and to validate their biological and prognostic characteristics in the phase III neoadjuvant NeoALTTO trial. Methods: By applying a case-control approach (1:2), we selected from the ALTTO trastuzumab arm 134 and 268 patients with and without a distant relapse respectively, matched for clinicopathological characteristics. In ALTTO, RNA was obtained from FFPE tumor cores from surgical samples. RNA sequencing (RNAseq) data available from 254 frozen pre-treatment samples were used for validation in patients receiving trastuzumab and/or lapatinib in NeoALTTO. The Absolute Intrinsic Molecular Subtyping (AIMS) was used to compute PAM50 subtypes. Prognostic genes were identified using a multivariable Cox proportional hazard model (controlling for clinicopathological characteristics and PAM50 HER2-enriched [HER2-E] vs others) for distant relapse-free survival (DRFS). Clusters were identified using non-negative matrix factorization (NMF) and k-means clustering, and characterized with gene expression profiling and Gene Set Variation Analysis (GSVA, hallmark gene sets from MSigDB). Aiming at identifying the subgroups in external cohorts, we identified relevant genes to develop group-specific signatures with LASSO regression, and used the derived scores to build a multinomial classifier. Results: The case-control cohort includes a high-risk population, with higher proportions of &gt;2cm, node positive and G3 tumors compared to the whole ALTTO trastuzumab arm. RNAseq data were generated for 386/402 patients. NMF and k-means clustering performed on genes associated with DRFS (false discovery rate &lt; 0.05) identified 4 groups with distinct biological characteristics and prognosis: immune-enriched (IM, N = 69), proliferative/metabolic (P/M, N = 87, characterized by glycolysis, cholesterol homeostasis and proliferation pathways), mesenchymal/stroma-enriched (M/S, N = 76, characterized by epithelial-mesenchymal transition, angiogenesis and TGF-beta signaling), and hormone receptor positive-enriched (N = 154), further divided into PAM50 HER2-E (N = 91) and non-HER2-E (N = 63, LUM, mainly luminal A/B tumors). The IM and LUM groups presented better DRFS (91% and 87% 5-year DRFS, respectively) compared to the others (5-year DRFS of 72%, 58% and 51% for HER2-E, M/S and P/M, respectively). The gene expression-based classifier identified the same 5 groups in NeoALTTO. GSVA analysis and comparisons among the 5 groups showed similar results in the two studies. Of interest, we observed significant differences in event-free survival in NeoALTTO (P = 0.035 and P = 0.041 in the whole population and in the subgroup with residual disease, respectively), with IM and LUM presenting better outcomes compared to the other groups. Sensitivity to neoadjuvant treatment as described by pathological complete response (pCR, ypT0/is ypN0) differed across the groups, with LUM presenting lower pCR rates (8%) compared to HER2-E (40%), IM (35%), P/M (32%), and M/S (32%). Conclusions: Five biologically-driven HER2-positive breast cancer subgroups were identified in ALTTO, highlighting the heterogeneity of this disease. Of note, their biological features and clinical behavior were validated in the NeoALTTO population, suggesting the robustness of our findings. IM and LUM tumors could be considered for treatment de-escalation approaches. Additional validation in cohorts receiving standard (neo)adjuvant therapies is warranted.
Citation Format: Mattia Rediti, David Venet, Andrea Joaquin Garcia, Dominique Agbor-Tarh, Marion Maetens, Delphine Vincent, Samira Majjaj, Sarra El-Abed, Takayuki Ueno, Serena Di Cosimo, Martine Piccart, Lajos Pusztai, Sherene Loi, Roberto Salgado, Giuseppe Viale, Françoise Rothé, Christos Sotiriou. Characterization and validation of biologically-driven HER2-positive breast cancer subgroups in the ALTTO and NeoALTTO clinical trials [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P4-09-05.