21st Century Science Initiative Grant: Researching Brain Cancer

Neuro-oncology has witnessed some important therapeutic advances, particularly in the treatment of pediatric brain tumors. Unfortunately the outcome for adult patients with the most common intrinsic primary brain tumor, glioblastoma (WHO grade IV astrocytoma), continues to be extremely poor: even the most exciting advances provide only minimal improvement in median survival in clinical trials. Fundamental changes in our paradigm in the development of prognostic markers, imaging, and therapy must occur for real changes in patient outcome. To develop a new therapeutic model, incorporating lessons from another area of medicine, infectious disease, may be useful. Mycobacterium tuberculosis (Mtb) is a major health burden in the developing world and in immunocompromised hosts, but few new effective anti-tuberculosis agents have been developed. Recent studies suggest that traditional high-throughput Mtb drug development assays that non-specifically target proliferation may not be useful in improving patient outcome because this model does not recapitulate in vivo conditions (Nathan et al. 2008). Rather, Mtb displays a cellular heterogeneity with a small fraction of the total population that is resistant to conventional therapies and is relatively quiescent. Non-replicating bacteria may be critical to the problem of persistent Mtb infection. The striking parallels to cancer biology cannot be ignored and are not surprising as nature tends to repeat patterns. While not all cancers may display a clear cellular hierarchy of tumor growth, the heterogeneity of cancers is essential to incorporate in models.

Cancer stem cells, which have been also described as tumor initiating cells or tumor propagating cells, are tumor cells that make copies of themselves (self renew) and propagate tumors similar to the parental tumor. Cancer stem cells from glioblastomas share some characteristics with normal brain stem cells including the expression of neural stem cell markers, the capacity for self renewal and long term proliferation, and the ability to differentiate into multiple nervous system lineages (neurons, astrocytes, and oligodendrocytes). However, brain tumor stem cells exhibit significant distinctions from normal stem cells in frequency, proliferation, aberrant expression of differentiation markers, chromosomal abnormalities, and tumor formation. The potent tumorigenic capacity of cancer stem cells coupled with increasing evidence of radioresistance and chemoresistance suggests that cancer stem cells contribute to tumor maintenance and recurrence. Thus, targeting cancer stem cells may offer new avenues of therapeutic intervention. This hypothesis has been recently validated in clinical trial of breast cancer in which patients undergoing treatment with cytotoxic chemotherapy experienced an increase in breast cancer stem cells in the surviving tumor while the use of a therapy targeting the stem cell population stabilized the cancer stem cell population.

Cancer cells grow in harsh environments with low levels of oxygen and nutrients and high levels of acid and metabolic byproducts. Rather than causing detrimental effects to cancer cells, the toxic tumor environment is associated with resistance to therapy and increased invasion into normal tissue. Cells that can survive in tumor regions with low oxygen and nutrients activate survival mechanisms with direct consequences for patients afflicted with cancer. These findings have led to the development of new therapies or techniques that may reverse the survival effects of the tumor regions of low oxygen/nutrition with limited success to date in clinical trials. We hypothesized that cancer stem cells may respond to low oxygen levels in a manner different from the rest of the tumor cells. In work currently in publication, we have discovered that brain tumor stem cells have specific molecular responses to low oxygen and inhibiting these responses blocks the ability of cancer stem cells to survive and grow. We are currently extending these studies to examine other potential therapeutic targets that may be downstream from the effects of low oxygen responses. Another extension of these studies has been to understand how low oxygen may contribute to the resistance of cancer stem cells to radiation and chemotherapy.

Many models of cancer stem cells imply a one-way transition from a stem-like cancer cell towards a more differentiated cell that is less likely to be able to grow long term and resist radiation and chemotherapy. Some investigators have suggested that treatments that kill cancer stem cells are the essential weapon required to improve cancer treatment. Indeed, a recent study from Germany has suggested that the chemotherapy most commonly used to treat glioblastoma patients, temozolomide (Temodar), kills cancer stem cells. We and others have demonstrated that another therapy in clinical trial for glioblastomas, the anti-angiogenic antibody bevacizumab (Avastin), can also kill cancer stem cells by disrupting a location (niche) that supports the growth of cancer stem cells. These studies are exciting because they suggest that known effective treatments may achieve results due, in part, to effects on cancer stem cells. However, there is a significant caveat: non-stem cancer cells (i.e. the bulk of the tumor) may be able to move back toward a more stem-like state.

The field of stem cell biology has been remarkably energized by the 2006 discovery that normal adult cells could be “reprogrammed” to become undifferentiated multi-potent stem cells capable of becoming any cell in the human body. The recipe for reprogramming cells included only four genes, three of which have been linked to brain tumor stem cells. More recent studies have shown that brain (neural) stem cells can be reprogrammed to the multi-potent state with only a single gene. The advances in nuclear reprogramming have caused excitement as we envision the creation of patient-specific resources for tissue regeneration and repair. In our excitement, we cannot fail to acknowledge the potential for disease that could arise from the acquisition of stem-like characteristics with the ability to overcome the body’s natural defenses. Normal stem cells must be able to move to locations of injury and survive in dangerous environments, so it is of little surprise that cancer stem cells acquire these traits. While we must guess the constituent requirements needed for reprogramming, cancer cells face evolutionary pressures that select for altering the expression or structure of genes that give advantage. Cancer biologists have typically considered proliferation the goal of cancer cells but acquisition of other characteristics – high motility, resistance to toxins, and possessing increased capacity for recruiting blood vessels – may provide a greater advantage to the neoplastic population than amplified proliferation alone. We therefore hypothesize that cancer cells face strong incentives to acquire these characteristics by adopting a stem cell program.

The search for new treatments has become intertwined with the identification of genes and their products that directly contribute to the malignancy of cancer. However, the function of a gene is dependent on the state and environment for the cell. We again find a parallel with Mtb in which “essentiality is conditional.” (Nathan et al. 2008). For example, an inhibitor of the Mtb proteosome is ineffectual with the bacteria in normal growth conditions but the same drug becomes potent when the Mtb is grown in conditions similar to the human body (N.B. the proteosome has been recently suggested to be a brain tumor stem cell target). If we apply these ideas to cancer, we may need to consider the utility of new drugs within the environmental conditions native for the tumor cells. Based on preliminary studies, we suggest that the local tumor environment (the microenvironment) may direct brain tumor cells to acquire stem cell-like characteristics and become more malignant. We believe that understanding the interaction between pathways that govern cancer stem cells and the microenvironment will be critical in designing better therapies for brain tumor therapy.

Based on this background, we propose to study both cancer stem cells and non-stem cells in specific environments to understand the potential utility of anti-cancer stem cell therapies, such as STAT3 or PI3K/Akt inhibitors, both alone in combination with conventional therapies. In addition, the microenvironment profoundly impacts the metabolism of a cell. Very recent studies have detected mutations in enzymes that regulate the ability of a cell to extract energy from nutrition. We will investigate the role of nutritional restriction on cancer stem cells and their response to therapies. We hope that the insights gained from these studies will inform the development of new treatments that can be translated directly into clinical trial.