Background
Cancer cells of solid tumors are not isolated entities, but are sustained in an environment of non-malignant cell populations, extracellular matrix and secreted factors. This tumor microenvironment (TME) is a heterogeneous and dynamic system that supports tumor growth and cancer metastasis, shapes treatment response and ultimately impacts the prognosis of the patient (1).
Colorectal cancer (CRC) is the second most common type of cancer in Norway. The tumors have a highly heterogeneous TME, showing large variation in immune cell infiltration and stromal components both within and among tumors. Immune infiltration is typically associated with a favorable patient prognosis, while cancer-associated fibroblasts are markers of an invasive phenotype and confer poor response to conventional chemotherapies. However, there is strong context dependency. For example, regulatory T cells can suppress immune function, while macrophages can be either tumor promoting or tumor suppressing (2,3). Improved interpretation of the TME in relation to patient outcomes is dependent on spatial contexts and the interaction between cancer cells and their microenvironment.
The project has been approved by the Regional Committee for Medical and Health Research Ethics South East Norway (project numbers 2005.1629, 2010/1805 and 2016/414), the Norwegian Data Protection Authority (05/01527-3) and the Norwegian Directorate of Health (05/5447).
Aims of the project
Multiplex immunohistochemistry is used for precise in situ analysis of protein marker expression, their potential co-localisation and for defining cell populations in tumors. This master project will be included in the framework of the Biomarker Norway (BIOMAN) study (https://www.ous-research.no/home/lothe/Research+interests/24818). This is a nationwide multi-center collaborative translational research study for biomarker analyses of CRC treated according to standard guidelines. The patient material is population-representative over a 5 year period (2008-13) and a series of up to 4 000 patients will be included. At each hospital, including all health regions, a pathologist and a clinician are responsible for selecting representative diagnostic blocks and for quality control of detailed clinical data.
Tissue microarrays (TMAs) from cases included in the BIOMAN study will be subjected to immunostaining with a combination of validated antibody panels directed against cancer cell-intrinsic (i.e. expressed by malignant epithelial cells) and TME-associated biomarkers (e.g. immune cell biomarkers) and analyzed through our imaging and digital image analysis platform. This approach will allow us to score each patient sample based, for instance, on their level of biomarker expression or on the spatial distribution of cell populations within the tumor and their pattern of interaction. The generated data will then be used to interrogate databases containing the patients' clinical data, including data on the therapy, to draw clinicopathological association using established statistical methodologies. The ultimate aim of the study will be to assess the clinical utility of our analyses, by evaluating the biomarkers associated to the patients prognosis and/or their ability to predict chemotherapy benefit.
We hypothesize that co-expression and the spatial context of protein markers of malignant epithelial cells and diverse cell populations of the immune TME and tumor stroma will improve the prediction of clinical outcome.
Methods
The student will perform staining of selected marker cocktails on TMAs of the BIOMAN cohort, digital image analyses and statistical analyses for tumor phenotyping and clinicopathological associations (REK 2016/414). The clinical data base has been designed by Professor, senior consultant Marianne G Guren, and responsible clinicians at each participating site has added data for their patients into a joint database. For patients from Oslo University Hospital a cohort of samples in TMAs and a number of molecular data exist at Dept. of Molecular Oncology. These TMAs will be used for initial analyses and potentially validated in the BIOMAN cohort and existing genomic data will guide the selection of protein marker cocktails (REK 2005.1629, 2010/1805).
Study and research environment
The project will be conducted in the Lothe group at the Department of Molecular Oncology, Institute for Cancer Research, Oslo University Hospital (https://www.ous-research.no/molecularoncology/). This research environment has established a molecular pathology platform for fluorescence-based immunohistochemistry and digital image analysis, using the PhenoImager HT system from Akoya Biosciences. This gives the opportunity to perform multiplex fluorescence staining of up to 9 markers on the same tissue section, with semi-automated extraction of quantitative data using dedicated image analysis software. The analysis will be performed on TMAs containing 1 mm tissue cores of formalin-fixed paraffin-embedded tumor samples included in the BIOMAN study series. The samples are collected from the diagnostic biobanks at each hospital, and tissue cores are taken from representative tumor areas marked on a hematoxylin-eosin stained slide as well as on the corresponding block by a pathologist. TMA construction has been completed at the Dept. of Molecular Oncology, and sections of them are available for the MSc project. Several multiplex marker cocktails have been optimized. The candidate will participate and learn this optimization protocol.
Supervisors
Dr. Christian Kranjec (scientist, main supervisor; will follow the student on daily basis, ensure midterm-evaluation and supervise in theoretical and practical matters of the MSc degree); Associate Professor Anita Sveen (project group leader, co-supervisor; will contribute with expertise in CRC biology and medicine); Dr. Ina A. Eilertsen (scientist, co-supervisor; will contribute will teaching of lab protocols) and Professor Ragnhild A. Lothe (Department head, Group leader; will ensure that all necessary resources are available, internal supervisor).
References
1. de Visser KE, Joyce JA. The evolving tumor microenvironment: From cancer initiation to metastatic outgrowth. Cancer Cell. 2023 Mar 13;41(3):374–403.
2. Bergsland CH, Jeanmougin M, Moosavi SH, Svindland A, Bruun J, Nesbakken A, et al. Spatial analysis and CD25-expression identify regulatory T cells as predictors of a poor prognosis in colorectal cancer. Mod Pathol Off J U S Can Acad Pathol Inc. 2022 Sep;35(9):1236–46.
3. Wang H, Tian T, Zhang J. Tumor-Associated Macrophages (TAMs) in Colorectal Cancer (CRC): From Mechanism to Therapy and Prognosis. Int J Mol Sci. 2021 Aug 6;22(16):8470.
4. Tan WCC, Nerurkar SN, Cai HY, Ng HHM, Wu D, Wee YTF, et al. Overview of multiplex immunohistochemistry/immunofluorescence techniques in the era of cancer immunotherapy. Cancer Commun Lond Engl. 2020 Apr;40(4):135–53.