ARREST

Project Coordinator

Liam McDONNELL
- Fondazione Pisana per la Scienza ONLUS
- Pisa - Italy

Project Partners

Benjamin BALLUFF
- Maastricht Multimodal Molecular Imaging Institute (M4I), Maastricht University
- Maastricht - The Netherlands
Axel WALCH
- Helmholtz Zentrum München, Research Unit Analytical Pathology
- Oberschleißheim - Germany
Robert SCHNEIDER
- IGBMC, CNRS UMR 7104 - Inserm U 964
- Illkirch - France
Jens SIVEKE
- Klinikum rechts der Isar, Technical University of Munich
- Munich - Germany

a) Background and rationale:
Many tumors including esophageal and esophagogastric junction adenocarcinomas (EAC) suffer from typical tumor characteristics responsible for poor prognosis: inherent and acquired therapy resistance, early metastasis and recurrence. All of these features are closely linked to inter- and intratumor molecular heterogeneity.
b) Hypothesis:
We hypothesize to identify robust classifiers to direct therapy based on comprehensive analysis of sequential tumor material from the MEMORI trial (Metabolic and Molecular Response Evaluation for Therapy Individualization in esophageal and esophagogastric junction adenocarcinomas, EUDRA-CT Number 2014-000860-16). In this clinical trial, 14 days before and after induction of neoadjuvant chemotherapy tumor response is evaluated using PET and sequential tumor biopsies obtained. This setting gives the unique opportunity to study the evolution of intratumor heterogeneity of EAC in the context of therapy response through a comprehensive molecular examination of the tissue specimens before, under and after neoadjuvant therapy.
c) Aims:
We want to characterize and understand intratumor heterogeneity in EAC samples before, under and after therapy within the MEMORI trial on a proteomic, genomic, epigenomic and metabolomic level where mass spectrometry imaging (MSI) will be used to identify tumor-biologically and therapeutically relevant tumor subpopulations. In parallel, we want to compare the natural and therapy-induced molecular heterogeneity using a second, readily available EAC sample collective (n=120) with the aim to get insights into the molecular mechanisms of resistance.
d) Methods:
Formalin-fixed, paraffin-embedded (FFPE) and PAXgene-based samples of the 75 patients are collected before and at 14 days post chemotherapy and at resection. MSI of these will be used to obtain spatially-resolved molecular profiles (Norris et al. Chem Rev 2013). Histologically annotated tumor areas will then be in-silico segmented on the basis of the molecular profiles for uncovering spatial molecular intratumor heterogeneity and identifying those tumor subpopulations that are associated with the resistance of EAC patients (Balluff et al. J Pathol 2015). MALDI-FTICR MSI and LA-ICP MSI will be performed to visualize the distributions of metabolites and drugs in the same tissues for comparison with the segmented intratumor heterogeneity. In parallel or subsequent MSI-guided next generation sequencing on the full sample or dissected material, respectively, will be performed for genomic and epigenetic (histone profiling) analyses combined with quantitative label-free proteomics. e) Expected results and potential impact: Integrative bioinformatics will allow for a comprehensive understanding of therapy-induced intratumor heterogeneity and its molecular mechanisms of resistance. This will thus deliver potential alternative therapeutic targets for EAC and robust classifiers for therapy decision making.

Many tumors including esophagogastric junction adenocarcinomas (EAC) suffer from typical tumor characteristics responsible for poor prognosis: therapy resistance, early metastasis and recurrence. All of these features are closely linked to inter- and intratumor molecular heterogeneity (ITH). Understanding the molecular mechanisms unpinning ITH and treatment resistance requires the identification of the adverse tumor subpopulations, their comprehensive molecular characterization and, most importantly, exquisitely defined patient data and samples.
In the ARREST project we hypothesize to identify detrimental tumor subpopulations based on a spatially differentiated analysis of sequential tumor material from the MEMORI trial (n=75; EUDRA-CT Number 2014-000860-16). In this clinical trial, tumor specimens are obtained before, 14 days after induction of therapy start, and at surgery. This longitudinal setting gives the unique opportunity to study the evolution of ITH in the context of therapy response through a comprehensive molecular examination of the tissue specimens. In parallel, the natural molecular ITH using a second, readily available EAC sample cohort (n=120) will be determined.
First, biomolecular ITH in tissue specimens is visualized by mass spectrometry imaging (MSI), an unlabeled molecular imaging technique that we have previously demonstrated can be used to identify tumor subpopulations. After MSI-guided identification and isolation of the resistant tumor subpopulations, they will be characterized comprehensively using next generation sequencing for genomic and epigenetic profiling combined with quantitative label-free proteomics.
Ultimately, we expect that this comprehensive characterization of detrimental tumor clones will help understanding resistance mechanisms and hence deliver potential alternative therapeutic targets for EAC and robust classifiers for therapy decision making.