Functional characterization of secreted proteins mediating glioma cell invasion
Glioma belong to the most common primary tumors of the brain. Glioblastoma is the most commonly diagnosed and the most aggressive malignant brain tumor. Despite multiple forms of treatment options (surgery, radiation treatment and chemotherapy), the median survival rate of patients with this form of brain cancer is about 14 months. One important reason for the poor prognosis is the aggressive growth of these tumor cells as they infiltrate the surrounding brain tissues. As a result, performing a complete surgical resection of the area is not possible. The remaining tumor cells have proven themselves to be extremely resistent against further therapeutic methods, such as radiation and chemotherapy. Relapse often occurs as a result. The invasion of the surrounding healthy brain tissues by glioma tumors does not happen randomly. It has been found to be associated with distinct anatomic structures such as the basement membranes of blood vessels as well as the subependyma. With regards to migration and invasion, the inhibitory myelin pathways also serve as essential structures for glioma in non-glioma cells (neurons, fibroblasts, other tumor cells as well as CNS metastases). The currently used standard therapies, such as radio and alkylating chemotherapy (including Temodal), target dividing cells. However, the invading cells do not divide, since the responsible components of the cytoskeleton direct mobility and cell division mechanisms, but not both at the same time (“go or grow” hypothesis). These cells are therefore therapy-resistant, which creates a major problem for treatment. New therapeutic agents can help stop the invasion and make the cells more susceptible to established therapeutic methods. In this connection, the Unfolded Protein Response shows great potential as a goal for therapeutic interventions.
Towards a transcriptome-wide and integrated vision of the translation branch of the unfolded protein response in glioma
The elevated proliferation of cancer cells necessitates a high metabolic rate and elevated protein synthesis. The tumor microenvironment often poses an additional challenge to the tumor cells with limited nutrient availability and hypoxic conditions. In sum, this results in persistent and on-going Endoplasmic Reticulum (ER) stress which triggers the Unfolded Protein Response (UPR), a signaling cascade that aims at reinstating and maintaining cellular homeostasis, controlling cell fate and allowing the cells to persist.
The UPR leads to a transcriptional and translational reprogramming in order to alleviate ER stress. Upon continuous ER stress and when cellular homeostasis cannot be reinstated, the UPR can also trigger apoptosis. Tumor cells make use of the prosurvival response of the UPR while evading apoptosis even in the presence of continuous ER stress. Moreover, it has been demonstrated that the active UPR in tumors conveys chemoresistance. Hence the UPR has emerged as a target for therapeutic intervention with the aim to tilt the balance of the protective effects of the UPR towards the induction of apoptosis in tumors.
In general the UPR utilizes three main branches to sense perturbations in ER homeostasis and to trigger the appropriate cellular responses: inositol-requiring enzyme-1 (IRE1), activating transcription factor 6 (ATF6) and PKR-like endoplasmic reticulum kinase (PERK). While the first two pathways mainly act via transcriptional reprogramming, PERK controls cellular translation via the phosphorylation of eukaryotic initiation factor 2 alpha (eIF2a). Strikingly though, while continuous ER stress (as encountered in tumors) normally results in attenuation of both the IRE1 and ATF6 branches of the UPR, PERK is thought to remain continuously active, suggesting that the PERK branch might be the predominant UPR signal cascade in tumors.
Despite its broad clinical importance, qualitative and quantitative models that describe the UPR in cancer cells are missing so far. We aim for a better understanding of the UPR in glioma in particular in the context of clinically important cell fate decisions such as invasion (‘go versus grow’) or resistance to pharmacological treatment (survival vs. apoptosis). To this end, we employ state of the art methodology to gain novel, deep and systems-wide insights into translation and its regulation during the UPR in glioma.
A systems biology approach to determine the equilibrium of the unfolded protein response
Rapid expansion of glioma cells coupled with inadequate vascularization increases levels of cellular stress and triggers the unfolded protein response (UPR), which is typical for many aggressive cancers. The UPR can be considered a survival response mechanism promoting an adaptation to a changing microenvironment. All three branches of the UPR can promote survival in response to stress, persisting or prolonged UPR however can result in apoptotic cell death.
The mechanisms that determine the cell fate decision during ER stress are not well understood but increasing evidence suggests that the multiple signals that determine specific outcomes are complex and highly integrated. Until now it is not clear at which point the decision of survival versus apoptosis is made. Quantitative measurements of the equilibrium of the three UPR pathways are not available yet, but it appears that the UPR in glioma, in dependency of its strength, has a strong impact on proliferation, invasion and apoptosis.
Systems biology of the Unfolded Protein Response in Glioma (SUPR-G)
The unfolded protein response (UPR) is a protective mechanism of the cell, coupled to the endoplasmic reticulum (ER), that is deregulated in many diseases. Especially in glial tumours, which make up 82% of the largest group of malignant tumours of the central nervous system, the UPR can have a negative impact on the successful outcome of treatment. Treatment resistance may develop leading to an undesirable survival of abnormal cells. A basic understanding of the signalling pathways involved, and their cell-stress induced reprogramming, is needed to determine the underlying pathogenetic mechanisms of this effect, and to enable an improved treatment of patients.
Although genome-wide genomic, epigenomic and transcriptional profiles improved the understanding of metabolic changes and molecular aberrations in glioma, the resulting impact on ER Stress and the mechanism driving cell fate decisions via the UPR remain unresolved. Therefore, our objectives are to:
- a) construct an UPR and ER Stress interaction network
- b) compare the therapeutic response pathways, and
- c) predict the UPR system behavior to identify targets which modulate migration and invasion characteristics of glioma cells
Towards understanding the UPR in infiltrating glioma cells
Diffuse human gliomas are graded according to the WHO classification into WHO grade II, III or IV depending on their histological characteristics, with the grade IV tumor being the most malignant diffuse glioma, the glioblastoma multiforme with a median survival of around one year. One of the major therapeutic challenges in the therapy of diffuse gliomas is their inherently diffuse growth within the brain. While the tumor bulk is often surgically accessible and the resection area can adjuvantly be treated using radiation therapy, the tumor cells, which have already diffusely infiltrated into the adjacent brain are not accessible to these therapeutic approaches.
Genetically, the vast majority of diffuse gliomas of WHO grades II and III carry mutations in IDH1 and about half of these tumors harbor mutations in the TP53 gene. In contrast, primary glioblastomas show a different mutation profile with either mutations in the PTEN gene and/or amplification with or without activating mutation of the EGFR gene besides mutations in other genes encoding proteins involved in the activation of the Pi3K/PKB signaling pathway.
The identification of novel therapies specifically targeting the infiltrating tumor cells in the context of their genetic alterations will be invaluable. One molecular mechanism which we are particularly interested in in this context is the Unfolded Protein Response (UPR) which occurs upon the induction of endoplasmatic reticulum (ER) stress. The UPR is a cellular stress response pathway that can for example be elicited upon increased loads of protein synthesis due to changes in cellular behaviour such as migration. Our project aims at a better understanding on how migrating glial precursor and infiltrating glioma cells are distinct from non-migrating glial precursor and infiltrating glioma cells especially also with respect to the UPR and how these properties can be exploited to therapeutically target infiltrating glioma cells outside of the bulk tumor.