Abstract 2174: The genomic and anti-tumor immune evolution of breast cancer
Authors: Blanco-Heredia Juan , Souza Carla Anjos , Gonçalves-Ribeiro Samuel , Gonzalez-Cao Maria , Callari Maurizio , Tresserra Francesc , Rossell Rafael , Caldas Carlos , De Mattos-Arruda Leticia 
Published: July 2021

Abstract
Background: Immunotherapy has changed the paradigm for a fraction of cancer patients. However, the evolution of tumor and immune microenvironment may uncover therapy resistance and immune evasion and its impact on clinical outcome. Here, in order to analyze the genomic and anti-tumor immune evolution, we performed extensive characterization of a dataset of a triple negative breast cancer patient including longitudinal and synchronous metastases (mets) and blood samples, collected over 2,039 days (d) of clinical follow-up.
Methods: Multi-platform profiling included whole exome seq, RNAseq, targeted multiplex gene expression, immunohistochemistry, and ELISpot T-cell assay of longitudinal chest wall biopsies (n=17 samples) and peripheral blood (n=14 samples), synchronous multi-regional mets at research autopsy (n=20 mets) and primary tumor analysis. State-of-the-art bioinformatics was applied to the data.
Results: Tumor mutation burden (TMB) estimates from the immunotherapy naïve chest wall lesion (M1) (d799) and 20 multi-regional mets (immunotherapy experienced, IE-mets) (d2,033) were 1.5 mut/mb and had median of 3.9 mut/mb, respectively (delta TMB 1.6 mut/mb). There were 73 neoantigens in M1 (subclonal/clonal ratio=0.4), and a median of 226 neoantigens per IE-mets (subclonal/clonal ratio=1.1). TP53 T256fs was validated as a T-cell specific clonal neoantigen and was targeted as liquid biopsy cell-free tumor DNA since cancer diagnosis. Clonal ordering and clonal evolution models for M1 and IE-mets evidenced that clustered mutations shared the same evolutionary history but with a mixed pattern of seeding: metastatic clones were shed by polyclonal and monoclonal seeding. Distinct seeding events occurred from early M1 to late IE-mets, and among late IE-mets (CI 0.95, p&lt; 0.0001). Longitudinal tumor inflammation (IFNy signaling) and antigen presenting machinery (APM) signatures defined immunophenotype statuses (i-inflamed, i-desert, i-excluded). They were modulated over clinical responses towards immune evasion and higher tumor cell proliferation. Immunophenotype statuses were also assigned to IE-mets that showed an overall heterogeneity and significant positive correlation to TMB (p=0.012), but not with the number of predicted neoantigens. M1 had HLA machinery status intact at the genomic level. IE-mets immunophenotyped as i-desert displayed statistically significant HLALOH (p&lt;0.05), indicating APM failure. However, an i-inflamed monoclonal IE-met evolving from M1 kept HLA status intact. This suggested APM machinery failure in IE-mets likely went through clonal evolution as a result of an immunoediting process.
Conclusion: By analysis of metastases genomes, we show that the immune tumor microenvironment shapes triple negative breast cancer progression by promoting and activating immune evasion mechanisms, culminating in heterogeneous lethal cancer.
Citation Format: Juan Blanco-Heredia, Carla Anjos Souza, Samuel Gonçalves-Ribeiro, Maria Gonzalez-Cao, Maurizio Callari, Francesc Tresserra, Rafael Rossell, Carlos Caldas, Leticia De Mattos-Arruda. The genomic and anti-tumor immune evolution of breast cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2174.