Abstract 4539: Tumor evolution and immune microenvironment dynamics define response to neoadjuvant treatment of esophageal adenocarcinoma
Authors: Barroux Melissa , Househam Jacob , Lakatos Eszter , Ronel Tahel , Baker Ann-Marie , Salié Henrike , Mossner Max , Smith Kane , Kimberley Chris , Nowinski Salpie , Berner Alison , Gunasri Vinaya , Jansen Marnix , Caravagna Giulio , Slotta-Huspenina Julia , Weichert Wilko , Alberstmeier Markus , Chain Benny , Friess Helmut , Bengsch Bertram , Schmid Roland M. , Siveke Jens T. , Quante Michael , Graham Trevor A. 
Published: April 2023

Abstract
Introduction: Locally advanced esophageal adenocarcinoma (EAC) remains difficult to treat, and resistance is common. The ecological and evolutionary dynamics responsible for treatment failure are incompletely understood.
Aim & Methods: We performed a multi-omic study with a multi-timepoint strategy to examine neoadjuvant treatment response at clonal resolution and in the surrounding tumor microenvironment. EAC samples from chemotherapy responding (REs) and chemotherapy non-responding (NRs) patients with locally advanced EAC were collected at three time points (prior, during, and after neoadjuvant therapy) within the multicenter MEMORI trial. Whole exome sequencing (mean depth 300x) was performed on 47 samples from 17 REs and on 24 samples from 10 NRs. Matched RNA sequencing was performed on 53 samples from 17 REs and on 26 samples from 10 NRs. To characterize immune response we performed imaging mass cytometry (IMC) with a 18-marker panel on 26 samples from 9 REs and 16 samples from 6 NRs and T-cell receptor sequencing in 18 RE-samples and 9 NR-samples.
Results: We observed profound changes in mutation signatures over time, characterized by C&gt;A transitions after exposure to neoadjuvant chemotherapy (FOLFOX), which is consistent with earlier observed oxaliplatin induced mutational signatures. Phylogenetic analysis showed no major changes in the clonal make-up during treatment, suggesting phenotypic plasticity rather than clonal evolution caused treatment resistance. EAC samples displayed widespread copy number alterations (CNAs), as previously described. CNAs arising during treatment were significantly more likely to be small, focal alterations rather than large alterations. At the transcriptome level neoadjuvant treatment led to significant changes with significant upregulation of immune and stromal pathways and oncogenic pathways such as KRAS, Hedgehog and WNT. The presence of immune escape mechanisms (defined as LOH or mutations in HLA, B2M mutations or PDL-1 overexpression) and a lack of clonal T-cell expansions were linked to poor clinical treatment response. Moreover, IMC analyses showed a less activated T-cell phenotype in NRs than in REs throughout treatment.
Conclusion: Using a multi-timepoint approach, we integrated genetic analyses with transcriptomic analyses and analyses of the tumor immune microenvironment for a holistic understanding of treatment resistance. This study identifies profound transcriptional changes during treatment with limited evidence that clonal replacement is the cause, suggesting phenotypic plasticity as a mechanism for therapy resistance. The presence of immune escape, a less activated T-cell phenotype and a lack of clonal T-cell expansions in patients with poor clinical treatment response has high pharmacological relevance and could be exploited via combined immune-chemotherapy treatments.
Citation Format: Melissa Barroux, Jacob Househam, Eszter Lakatos, Tahel Ronel, Ann-Marie Baker, Henrike Salié, Max Mossner, Kane Smith, Chris Kimberley, Salpie Nowinski, Alison Berner, Vinaya Gunasri, Marnix Jansen, Giulio Caravagna, Julia Slotta-Huspenina, Wilko Weichert, Markus Alberstmeier, Benny Chain, Helmut Friess, Bertram Bengsch, Roland M. Schmid, Jens T. Siveke, Michael Quante, Trevor A. Graham. Tumor evolution and immune microenvironment dynamics define response to neoadjuvant treatment of esophageal adenocarcinoma. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4539.