The EuroTARGET project .1 Objectives .1 Objectives

EuroTARGET denotes for “European collaborative project on Targeted therapy in Renal cell cancer: Genetic and Tumor-related biomarkers for response and toxicity”.

The general aim was to identify and characterize host- and tumor-related predictive biomarkers for response to targeted therapy in patients with metastasized renal cell carcinoma (mRCC). For this purpose several state-of-the-art approaches were combined to generate as much information as possible on host- and tumor-specific factors which could potentially be predictive for therapy outcome.

Hence, the main aim can be further divided into several objectives, where each work package contributed results with different approaches and methodology. A complete description of the work packages can be found in Section 3.2.2, Figure 3.1 and Table 3.6.

The following list comprises all project objectives ordered according to the study protocol without assigning them to specific work packages or project partners:

− Create a standardized European clinical databank and bio-repository (germline DNA of all patients and serum and frozen tumor tissue of a subgroup) of a large series of patients with mRCC treated with different targeted agents.

− Identify genetic markers for treatment response and toxicity by performing high-resolution germline whole-genome profiling in patients treated with sunitinib or sorafenib

− Identify exon and microRNA expression markers for treatment response and toxicity by gene expression profiling of tumors from patients with and without good response

− Identify kinase activity profiles related to TKI response

− Identify promoter hypermethylations markers in TKI response

− Identify resulting protein profiles corresponding to genomic, epigenetic and expression alterations related to TKI response

− Replicate all identified markers in independent patient series

− Study the functional relevance of replicated markers/networks in vitro by knock-out and knock-in transfection experiments

− Identify differentially expressed proteins before and after knock-down/

upregulation of genes of interest

− Identify plasma drug and metabolite levels as phenotype of results

− Explore the possibility of individualizing dosage regimens by integrating biomarker concentration – time profiles into PK/PD models for sunitinib (and pazopanib)

− Conduct integrated bioinformatical analyses of the results obtained by all different approaches in order to maximize the probability to find new markers and to understand the interrelatedness between them

− Construct new risk stratification criteria to be used for personalized mRCC patient management

− Disseminate the new knowledge to medical oncologists, urologists and the scientific community

3.2.2 Project design and duration

The EuroTARGET project was designed as a European-wide non-interventional prospective cohort study with an overall duration of 5 years [123].

Based on their expertise, project partners were assigned to 12 different work packages with one partner taking supervision on all tasks of the respective package. An overview on all work packages, their main subjects and the responsible project partner can be found in Table 3.6. Focus of the underlying work were the results of work packages 2 and 7 in a subcohort of the EuroTARGET study population which was additionally

Fig. 3.1: General concept of the EuroTARGET project.

monitored over a defined period during the first-line therapy with sunitinib or pazopanib (Section 3.3). Therefore, methods used in other work packages are not explained here in detail as it would be beyond the scope of this thesis.

Design and methods of work packages 2 and 7 are addressed in the following paragraphs beginning with 3.3.

WP = Work package

Tab. 3.6: Overview of EuroTARGET work packages

WP Description Lead participant

1 Retrospective and prospective collection of clinical data, genomic DNA and tumor tissue

University of Cambridge

2 Genetic profiling of mRCC patients Leiden University Medical Center 3 Transcription profiling of tumor

material from mRCC patients

Radboud University Medical Center, Nijmegen

4 Kinase activity profiling of mRCC PamGene, s-Hertogenbosch 5 DNA methylation biomarkers of

mRCC

Bellvitge Biomedical Research Institute, Barcelona

6 Functional studies University Hospital Homburg/Saar 7 PK/PD studies (EuT-PK/PD

substudy)

University of Bonn

8 Data management deCODE Genetics, Reykjavik

9 Integrated data analysis Netherlands Cancer Institute, Amsterdam

10 Dissemination and Training Central European Society for Anticancer Drug Research, Vienna

11 Ethics Institut national de santé et de la

recherché médicale, Paris 12 Project Management Radboud University medical

Center, Nijmegen

3.2.3 Patients

Patients were applicable for the EuroTARGET project if they were 18 and above years old, newly diagnosed with metastatic renal cell carcinoma and did not receive a treatment for their metastatic disease. Furthermore, patients had to be able to understand the patient information and to give informed consent. Recruitment was conducted in Germany, Austria, Switzerland, The Netherlands, United Kingdom, Iceland and Romania. Although the study was planned as prospective study, retrospective inclusion of patients was also possible under the requirement that at least one blood sample was available for germline analysis.

It was planned to include a total of 1100 patients. There was no intention to test a formal hypothesis. Hence, a sample size calculation was not conducted.

3.2.4 Medication

All possible targeted first-line therapies according to the recent guidelines for metastasized renal cell carcinoma were allowed. Since this study had a purely observational purpose, no dose or treatment recommendations were given.

3.2.5 Sampling procedure

Each patient was asked to donate 8-10 mL whole blood either collected in Lithium-Heparin or EDTA tubes for germline analysis. Blood samples were then stored at -20°C or, preferably, -80°C until analysis.

Tumor tissue was collected from each patient, if available. Fresh frozen tumor material stored at -80°C was preferable; however, paraffin blocks were also a valid option. All tumor samples were reviewed by a designated reference pathologist and then sent to the respective project partners for further analyses.

3.2.6 Endpoints

Progression-free survival (PFS) was chosen as primary endpoint with progression ideally defined with respect to the Response Evaluation Criteria In Solid Tumors (RECIST) [124]. Usually, the RECIST assessment is not part of the clinical routine;

hence, progression documented based on the treating physicians’ expert opinion was also viable when no RECIST information was available.

Treatment toxicity was determined as secondary endpoint. Documentation was handled using the Common Terminology Criteria for Adverse events (CTCAE,v 4.03) [105]. Only adverse events with grade 3 or higher had to be captured in the electronic case report form (eCRF). However, low-grade toxicity could be documented indirectly, when it was the reason for a dose reduction of the drug.

3.3 The EuroTARGET-PK/PD (EuT-PKPD) sub-study