Project

EGFR-targeted therapy has been approved for the treatment of advanced CRC, but response rates are low, not durable and treatment is limited to those patients with Ras-wildtype disease. Moreover, TNBC and CRC patients are prone to develop metastases and have an especially poor prognosis underpinning the need for new targeted and broadly applicable therapeutic strategies.

The high proliferation rate and the invasive properties common to cancer cells rely on active secretory machineries. Tumour cell secretion contributes to the following hallmarks of cancer: 1) hyperproliferation; 2) evasion of growth suppression; 4) loss of cell polarity; 5) activation of cell motility, invasion and metastasis; 6) shaping of the tumour microenvironment through altered presentation of proteins and the cancer secretome; 5) resistance to cell death.

Dysregulated secretion can thus be considered as a major driver of cancer progression and therefore holds promise as a general therapeutic target for the treatment of solid cancers including TNBC and CRC, irrespective of the mutational status.

However, while it is intuitively clear that tumour growth and metastasis are linked to secretion, strategies for therapeutic exploitation of the secretory pathway are still in their infancy. This can be explained by the incomplete understanding of how the secretory pathway is dysregulated by aberrant signalling in cancer cells9. This lack of knowledge hinders efforts to exploit the secretory pathway for therapeutic and diagnostic purposes.

SECRET Early Stage Researchers (ESRs) will interrogate the impact of secretion to TNBC and CRC progression in a training network combining researchers with different expertise from academia and industry providing a unique combination of tools, instrumentation and knowledge, which will generate an interdisciplinary and intersectoral platform integrating cell biology, biochemistry, proteomics, molecular biology, organoid and animal models, systems biology, and computational modelling.

Drawing on this unique consortium expertise each ESR project will further be supported by innovative systems cancer cell biology and/or integrative proteomic analysis approaches. SECRETs research objectives are i) to obtain a mechanistic and quantitative understanding of the mutual regulation of the secretory pathway through signalling and delineate its contribution to cancer progression and ii) to define secretory pathway- linked candidate biomarker genes suitable for cancer diagnosis and prognosis.

SECRET will thus address current unmet needs in the cancer field and pave the road for the development of novel therapeutic strategies.

Individual Research Projects (IRPs) at a glance:

Work Package 1 – Mechanisms

IRP 1.1 – Oncogenic kinase signaling driving secretion and TNBC progression

IRP 1.2 – Impact of biomechanical ECM properties on Golgi-controlled cancer cell secretion and communication

IRP 1.3 – Aberrant Rho regulation in cancer cell secretion

IRP 1.4 – Systematic analysis of the role of Golgi fragmentation in TNBC progression

IRP 1.5 – Regulation of STING trafficking

IRP 1.6 – How do MAPK inhibitors affect the interplay of proliferation, secretion and invasion of cancer cells?

IRP 1.7 – Secretome mediated tumor-macrophage interactions and treatment resistance

IRP 1.8 – Links between the autoregulation of cargo export from the ER and the control of TNBC cell growth

Work Package 2 – Biomarkers

IRP 2.1 – Proteomics investigation of the malignant functions and diagnostic potential of the cancer secretome: Focus on classically secreted proteins

IRP 2.2 – Proteomics investigation of the malignant functions and diagnostic potential of the cancer secretome: Focus on exosomes

IRP 2.3 – Data science and systems biology approaches to derive biological insights from secretome data sets

IRP 2.4 – Changes in the secretome during tumor stroma interaction

IRP 2.5 – Influence of the secretome on invasion and metastasis in vivo

IRP 2.6 – Pathway construction via multiplex phosphoproteomic data

IRP 2.7 – Development of a validated mathematical model to predict druggable nodes within the secretory pathway

Work Package 1 – SECRET Mechanisms

1.1 – Oncogenic kinase signaling driving secretion and TNBC progression

Project description:

TNBC is an especially aggressive disease for which efficient molecular therapies are still lacking. The serine-threonine protein kinase PKD3 is elevated in TNBC cell lines and tissues and pharmacological PKD inhibition potently suppressed the proliferation of TNBC cells in vitro and in vivo. Additionally, PKD3 regulates the fission of vesicles at the Golgi complex thereby controlling secretion. Using different techniques such as multiplexing and in vivo xenograft models the candidate will establish PKD3 and downstream cargo as novel druggable nodes for TNBC treatment.

Location: University of Stuttgart, Germany 

Principal Investigator: Dr Angelika Hausser (USTUTT)

Collaborators: Prof Leonidas Alexopoulos, ProtATonce, Greece (PAO) and Prof Tilman Brummer, University Medical Center Freiburg, Germany (UKLFR)

1.2 – Impact of biomechanical ECM properties on Golgi-controlled cancer cell secretion and communication

Project description:

Breast tumorigenesis is accompanied by collagen crosslinking, extracellular matrix (ECM) stiffening and increased contractility. The Golgi complex has been recently established as a mechanosensor organelle, but how matrix stiffness impinges on the secretory pathway on the level of the Golgi complex remains unknown. The candidate will thus investigate how the cytoskeleton senses the ECM density to coordinate Golgi secretory function with the increased demand for factors such as proteases or cytokines required for ECM remodeling during invasive 3D migration.

Location: University of Stuttgart, Germany 

Principal Investigator: Dr Angelika Hausser (USTUTT)

Collaborators: Prof Connie Jimenez, VU University Medical Center, Netherlands (VUmc); Dr Judith Farres, Anaxomics, Spain (AX); Dr Helmut Dolznig, Medical University Vienna, Austria (MUW).

1.3 – Aberrant Rho regulation in cancer cell secretion

Project description:

Metastatic dissemination of cancer cells is driven by aberrant Rho GTPase signalling, which is often caused by the deregulated expression of Rho regulators, the GAP and GEF proteins. By controlling cytoskeletal remodelling, Rho GTPases are not only critical regulators of cell morphology and motility, they also drive membrane trafficking and polarized secretion. In cells with deregulated Rho signalling, the candidate will therefore identify by quantitative mass spectrometry the changes in plasma membrane protein composition and the cellular secretome. The candidate will then explore in 3D co-culture models how these changes affect the interaction with stromal cells, e.g. primary human macrophages and fibroblasts, both of which play important supportive roles in tumor cell invasion. 

Location: University of Stuttgart, Germany 

Principal Investigator: Prof Monilola Olayioye (USTUTT)

Collaborators: Prof Connie Jimenez, VU University Medical Center, Netherlands (VUmc); Prof Matthias Schwab, Robert-Bosch-Krankenhaus, Stuttgart, Germany (RBK)

1.4 – Systematic analysis of the role of Golgi fragmentation in TNBC progression

Project description:

The candidate will address the question of why breast cancer cells exhibit a fragmentation of the Golgi apparatus and what advantage this condition confers to the cancer cells. This will be addressed using a combination of live cell imaging and siRNA screening. 

Location: University of Oslo, Norway

Principal Investigator: Prof Hesso Farhan (UiO)

Collaborators: Prof Roded Sharan, Tel Aviv University, Israel (TAU); and Prof Gunhild Maelandsmo, Oslo University Hospital, Norway (OUH)

1.5 – Regulation of STING trafficking

Previously: Cleaved proteins in exosomes and their contribution to the establishment of the metastatic niche

Project description:

Stimulator of interferon genes (STING) is a key component of the innate immune response by detecting DNA-containing pathogens and tumour-derived DNA. Upon activation, STING translocates to the endoplasmic reticulum-Golgi intermediate compartment (ERGIC) and the Golgi in a process that is dependent on COPII vesicles, initiating production of type I interferon. The aim of this study is to identify the COPII components required for STING cellular trafficking and to elucidate the coupling of trafficking and activation of STING. Our work will contribute to a deeper understanding of STING trafficking and the spatial regulation of its activation, ultimately elucidating its role in anti-cancer immunity.

Location: University of Oslo, Norway 

Principal Investigator: Prof Hesso Farhan (UiO)

Collaborators: Prof Christian Behrends, Ludwig Maximilian University of Munich, Germany (LMU)

1.6 – How do MAPK inhibitors affect the interplay of proliferation, secretion and invasion of cancer cells?

Project description:

The RAS/ERK MAPK pathway is frequently dysregulated in colorectal and triple negative breast cancer (TNBC) and is pursued as a target in precision medicine with a growing arsenal of kinase inhibitors and therapeutic antibodies. In this project, we will investigate how these compounds interfere with the secretory pathway and thereby affect the hallmarks of cancer such as proliferation, survival, motility and invasiveness.

Location: University Medical Center Freiburg, Germany

Principal Investigator: Prof Tilman Brummer (UKLFR)

Collaborators: Prof Hesso Farhan, University of Oslo, Norway (UiO) and Dr Klaus Mauch, Insilico Biotechnology AG, Germany (INS)

1.7 – Secretome mediated tumor-macrophage interactions and treatment resistance

Project description:

Acquisition of chemoresistance in TNBC is one of the major clinical problems. The resistance can be facilitated via cancer cell interactions with Tumour Associated Macrophages (TAM). Cancer-secreted cytokines can modify macrophages so that they reduce treatment effect. By revealing mechanisms by which TNBC-TAM interactions mediate the resistance, novel therapeutic strategies will be designed. 

Using chemo-sensitive and -resistant in vivo xenograft models and ex vivo cultures the ECR will identify respective secretomes, their influence on macrophages and association with treatment response. Profound analysis of resistance-associated characteristics will suggest novel candidate for targeting that will be explored in TNBC-macrophage co-cultures as well as in vivo models. 

Location: Oslo University Hospital, Norway

Principal Investigators: Drs Gunhild M. Mælandsmo and Lina Prasmickaite (OUH)

Collaborators: Dr. Helmut Dolznig, Medical University of Vienna, Austria  (MUW) and Dr. Leonidas Alexopoulos, ProtATonce, Greece (PAO)

1.8 – Links between the autoregulation of cargo export form the ER and the control of TNBC cell growth

Project description:

The secretory pathway responds to signals induced by the availability of environmental factors such as nutrients and growth factors. However, little is known about autochthonous regulation of the secretory pathway by signalling molecules. A recently uncovered signaling complex named AREX (Autoregulation of endoplasmic Reticulum EXport) exerts a positive regulatory action on the ER export machinery, effectively coupling protein folding with the ER export process. The candidate will characterize autoregulatory signalling at the ER-Golgi interface and explore its role in TNBC progression in 3D cell culture models and in vivo. 

Location: National Research Council of Italy, Naples 

Principal Investigator: Prof Alberto Luini (CNR)

Collaborators: Dr Julia Schüler, Charles River Discovery Services, Germany (CRDS) and Dr Helmut Dolznig, Medical University Vienna, Austria (MUW)

Work Package 2 – SECRET Biomarkers

2.1 – Proteomics investigation of the malignant functions and diagnostic potential of the cancer secretome: Focus on exosomes

2.2 – Proteomics investigation of the malignant functions and diagnostic potential of the cancer secretome: Focus on classically secreted proteins

Project description:

The cancer secretome harbors classically secreted (focus candidate Project 2.2) and vesicle (exosome)-based proteins (focus candidate Project 2.1) promoting invasion and proliferation. Therefore unbiased analysis using mass spectrometry-based proteomics combined with functional data mining has great potential to uncover drug targets and biomarkers for non-invasive diagnostic applications. 

Secretome and exosome proteomics of clinical material (focus colorectal cancer) will be further explored in pan-cancer cell models and clinical material. Drug target potential of selected regulatory proteins/pathways will be tested in functional experiments in vitro and the biomarker potential in clinical cohorts. 

Location: Cancer Center Amsterdam, Amsterdam University Medical Center, location VUmc, Amsterdam, The Netherlands

Principal Investigator: Prof Connie Jimenez (VUmc)

Collaborators IRP 2.1: Dr. Irene Bijnsdorp, Cancer Center Amsterdam, Amsterdam University Medical Center, Dr. Meike de Wit, Netherlands Cancer Institute, Amsterdam, The Netherlands (VUmc), Dr Angelika Hausser, University of Stuttgart, Germany (USTUTT) and Dr Julia Schüler, Charles River Discovery Services, Germany (CRDS)

Collaborators IRP 2.2: Dr. Irene Bijnsdorp, Cancer Center Amsterdam, Amsterdam University Medical Center, Dr. Meike de Wit, Netherlands Cancer Institute, Amsterdam, The Netherlands (VUmc); Prof Matthias Schwab, Robert-Bosch-Krankenhaus, Germany (RBK); Prof Leonidas Alexopoulos, ProtATonce, Greece (PAO)

2.3 – Data science and systems biology approaches to derive biological insights from secretome data sets

Project description:

The project will combine the results of statistical analyses in a manner that incorporates the known complementarity of the underlying biological functions. The ESR will develop semi-automated, iterative processes that can be optimized around a metric of biomarker quality to screen different data sets. This approaches will be applied with the aim of identifying relevant cross-talks between the measured secreted factors and key molecular players in cancer and identify new potential points of monitoring or therapeutic intervention.

Location: Anaxomics Biotech S.L., Spain

Principal Investigators: The candidate will be directed within Anaxomics by Dr. Judith Farrés and Dr. José Manuel Mas and enrolled in the PhD program on Bioinformatics at Universitat Autònoma de Barcelona and be co-directed by Prof. Xavier Daura (AX)

Collaborators: Prof Connie Jimenez, VU University Medical Center, The Netherlands (VUmc) and Prof Xavier Daura, University Autonoma de Barcelona, Spain (UAB)

2.4 – Changes in the secretome during tumor stroma interaction

Project description:

Interactions between the tumour and its microenvironment strongly contribute to tumour progression, growth and metastasis. However, many details of these interactions are still unresolved. Therefore, in this project the candidate will address the secretome-dependent communication of colorectal carcinoma (CRC) cells with stromal fibroblasts and myeloid cells. The candidate will aim to delineate the changes in the secretome of the individual cell types when communicating with each other and to functionally test its impact on cancer progression. Specifically, this question will be addressed using 3D organotypic assays (OTAs) and primary cancer organoid cultures in a modular fashion in mono-cultures, co-cultures or triple-cultures of the respective cell types. All this data will be further bioinformatically evaluated in a biological network analysis. Identified molecules will be functionally validated in vitro and in orthotopic tumour xenograft models. 

Location: Medical University Vienna, Austria

Principal Investigator: Dr Helmut Dolznig (MUW)

Collaborators: Prof Olivier de Wever, Ghent University, Belgium (GU); and Dr Julia Schüler, Charles River Discovery Services, Germany (CRDS)

2.5 – Influence of the secretome on invasion and metastasis in vivo

Project description:

In recent years, there have been multiple efforts in the establishment and characterization of large collections of patient derived tumour xenograft (PDX) models for cancer research. PDX models mainly retain the histological and genetic characteristics of the donor tumour and remain stable across passages. They preserve cell-autonomous heterogeneity, thereby representing very well the molecular landscape of the corresponding disease. Currently, PDX panels display most consistently the complexity of tumour heterogeneity and molecular diversity of human cancers. The addition of human stromal as well as immune cells has an impact on the tumour biology of PDX models in vivo. The candidate will investigate the bi-directional cross-talk between these cell types and their murine counterparts by using an already established orthotopic colorectal carcinoma (CRC) as well as triple negative breast cancer (TNBC) PDX model. The candidate will determine the changes in the secretome of the different involved cell types and the influence of the bi-directional cross-talk on tumour invasion and metastatic behaviour of the cancer cells. After identification of key players in the secretome, their functional role will be evaluated via small molecule inhibitors or siRNA treatment experiments. 

Location: Charles River Discovery Services, Freiburg, Germany

Principal Investigator: Dr Julia Schüler (CRDS)

Collaborators: Dr Helmut Dolznig, Medical University Vienna, Austria (MUW) and Prof Christine Sers, Charite Universitätsmedizin Berlin, Germany (CUB)

2.6 – Pathway construction via multiplex phosphoproteomic data

Project description:

The candidate will aim to identify baseline and post-treatment phosphoproteomic and cytokine release signatures and associated logical models that can predict carcinoma progression. Using computational methodology to compare signaling networks and to identify drug effects by discrete logical models that build on integer linear programming formulation, the candidate will attempt to understand systems-level signaling effects of kinase inhibitors, and to develop systems biomarker candidates predictive of treatment responsiveness. The responsiveness of TNBC and CRC cell lines to drugs will be employed to develop computational models that link the phosphorylation activity to cell response. 

Location: ProtATonce, Athens, Greece

Principal Investigator: Prof Leonidas Alexopoulos (PAO)

Collaborators: Prof Connie Jimenez, VU University Medical Center, Netherlands (VUmc) and Dr Angelika Hausser, University of Stuttgart, Germany (USTUTT)

2.7 – Development of a validated mathematical model to predict druggable nodes within the secretory pathway

Project description:

Cell proliferation requires an increased rate of protein synthesis and need to sort and secrete those proteins through the secretory pathway. Understanding secretory pathway regulation is thus a basis for the development of therapeutic strategies targeting secretion and thus aberrant cancer proliferation. In particular, the induction of ER stress through inhibition of ER export is a promising strategy against cancer. The goal of this sub-project is to develop and validate a prototype systems-biology-based tool that can predict druggable secretory pathway nodes in TNBC and CRC by combining combinatorial optimization and machine learning techniques.

Location: Tel Aviv University, Israel

Principal Investigator: Prof. Roded Sharan (TAU)

Collaborators: Prof Hesso Farhan, University of Oslo, Norway (UiO) and Prof. Tilman Brummer, University Medical Center Freiburg, Germany (UKLFR)