DKTK-Young Investigator Group "Oncogenic Signaling in Colorectal Cancer" (Dr Peter Jung)
left: normal human colonic organoids grown from a mucosal tissue sample / right: Tumor organoid line established from liver metastasis of a colon adenocarcinoma (Courtesy: Dr Jung)
Colorectal cancer (CRC) represents the second most diagnosed form of cancer disease in Germany (Robert Koch Institute 2011/2012). In most cases, the disease follows the “classic” adenoma-carcinoma sequence, and underlying genetic alterations of disease driver pathways such as WNT, KRAS/MAPK, TGF-β/SMAD4, and p53 have been well known since decades (“Vogelgram”, named after Bert Vogelstein). Novel strategies that apply targeted therapies have improved the survival of individuals suffering from metastatic CRC disease. However, disease recurrence after initial curative surgery and chemoresistence strongly limit the success of nowadays cancer therapy. These obstacles can be only overcome by a better understanding and in‑depth characterization of the disease on an individual basis. Essential progress of molecular tumor pathology, which has adopted state-of-the-art next generation sequencing techniques, allows for a more precise patient stratification. However, the development of innovative patient-derived pre-clinical tumor models, which better reflect the characteristics of the primary tumor, are key to better understand disease progression and to develop personalized anti-cancer treatments.
We have set up an ex vivo culture system that allows us to propagate patient-derived samples of primary colorectal tumors and liver metastasis in the culture dish. Building this “living biobank” of human colorectal cancer at the DKTK partner side munich is done in collaboration with the Institute of Pathology of the Ludwig-Maximilian University (LMU) (Head: Prof. Dr. Thomas Kirchner) and with the support of PD Dr. Jens Neumann. In our pre-clinical model, primary tumor cells expand in three dimensions (3-D) as so-called “patient-derived tumor organoids” (PDO`s) which are embedded in an extracellular matrix mainly composed of laminin and collagen. Propagating cancer cells in three rather than two dimensions has been shown previously to create a reasonable heterogeneity at the cellular level with respect to self-renewal and proliferative capacity, hypoxic compartments, and signaling pathway activities which represent critical features of colorectal cancer stemness. The tumor organoid culture technique is currently employed by international consortia with the goal to implement this techniques in the development of custom-made, individualized anti-cancer therapy approaches. From what we now until now, it can be expected that the tumor organoid model can provide the missing link between cell culture and in vivo modelling of the disease. Intruigingly, as has been shown very recently, xeno-transplantation of primary tumors into immune-deficient mice leads to a host-specific tumor evolution that differs from the actual tumor progression in patients, presumably due to a foreign microenvironment in the host organism which imposes an artificial selective pressure on the xeno-transplanted human tumor cells. We see that patient-derived tumor organoids, dependent on their mutational status, indeed show dependencies on certain niche growth factors that can be provided ectopically in order to prevent this inappropriate bias. Importantly, primary tumor organoid lines have been shown also by others to maintain the genetic diversity of the original tumor for several weeks in the culture dish which makes them especially suitable to perform comprehensive analyses of the molecular events that confer chemotolerance and drug resistance in a patient-specific manner.
While tumors had been considered as a uniform cell cluster for many years, recent progress in the field has refined our view of cancer as hierarchical entities composed of a few self-renewing stem-like, metastasis-initiating cells (CSCs), and a bulk tumor mass of differentiated-like cells which have lost the ability to spread the disease to distant organs. As an additional layer of disease complexity, tumor-adjacent stromal cells, after their re-programming by epithelial cancer cells, are indeed capable to increase the frequency of cancer-initiating cells and the likelihood of disease progression and metastasis. We have optimized a 3-D co-culture of patient-derived tumor organoids with cancer-associated fibroblasts isolated from the same tumor samples. These co-cultures show increased tumor growth and cancer-stemness in form of LGR5 gene expression ex vivo. Hence, they represent a valuable tool to better understand the microenvironmental contribution to tumor aggressiveness and chemo-resistance. One of our interests is to study how advanced, metastatic CRC cells communicate to and polarize their adjacent microenvironment. In a collaboration with the Proteomics Laboratory of Prof. Dr. Bernhard Kuster in Freising, we are seeking to reveal how the secretome of patient-derived CRC organoids changes with increasing cancer stem cell features and metastatic disease progression