By: Rachel Phillips
Chemotherapy and radiation are staples of cancer treatment, but the search to find a solution that kills tumor cells without any harmful side-effects is essential for increasing patient quality of life. Dr. Gershon describes an all too common occurrence in his practice: “Imagine a six-year old child is used to being normal, playing, [and] all of a sudden starts having headaches. Turns out he has [a] medulloblastoma, and has to undergo treatment. Instead of their life being like that of every other six year old, they’re going to the hospital every single day for six weeks to be sedated and treated with radiation. [This] makes them feel nauseated when they wake up, and when they’re done, they’re getting these drugs put in to their bodies that continue to make them feel nauseated… All of this while the doctors are insisting they’re trying to help them. It really creates kind of a sense of PTSD for the kids.”
Medulloblastomas are the most common malignant tumor in children, and the Gershon Lab in the Neurology Department at UNC Chapel-Hill is currently studying this tumor and cerebellar development, in mice. The lab is led by pediatric neuro-oncologist, Dr. Timothy R. Gershon, who grew up with a fascination of brain tumors and a desire to face the challenge of treating them. The research team aims to illuminate new treatment therapies for pediatric brain cancer that do not induce the debilitating neuro-cognitive impairments seen following traditional treatment.
Apoptosis Pathway- Cytotoxic stimuli leads to activation of Bcl-2 proteins which interact with Bcl-xL. Bcl-xL can no longer inhibit active Bax, and Bax leads to release of cytochrome C, ultimately causing cell death. Flowchart adapted from Katie Veleta.
One of the recent developments in the Gershon lab has been the discovery of roles of specific proteins involved in programmed cell death, or apoptosis, during cerebellar development. The human brain is essentially under construction for the first year of life when the cerebellum, a large portion in the brain located in the back of the skull responsible for regulating motor activity and coordination, is developing. Cerebellar granule neuron progenitors (CGNPs) are the cells most important during this formative time of cell division, also referred to as proliferation.
Signaling in developing brains regulates CGNP proliferation and is essential for preventing medulloblastoma development. CGNPs readily undergo apoptosis when they are subjected to DNA damage, which can come in the form of chemo, radiation, or drug therapy.
Regulation of apoptosis is performed in the cerebellum by proteins of the Bcl-2 family. Bax is one of the pro-apoptotic members of this group of proteins. Specifically, Bax undergoes a change in shape at the start of apoptosis, which is recognized by the antibody 6A7. Researchers wanted to know what other Bcl-2 proteins were also present during this time and if any of these were forming interactions with Bax.
To examine this interaction, whole cerebella were dissected and subjected to an antibody (6A7) and analyzed by Western blot, a way to identify the presence of various proteins. Two ages of mice were used; young (7 days old) and adult age.
Mouse cerebellum stained with PCNA to show proliferating cells (purple) and p27 for differentiating cells (green) at height of cerebellar proliferation stage, 7 days old, in a normally developing mouse. Image courtesy of Gershon Lab.
Main findings indicated that another Bcl-2 protein, called Bcl-xL, was interacting with Bax. Interestingly, this interaction only occurred at early stages of cerebellar development (in the 7 day old mice), and not once the brain was fully developed (in the adult mice). This led Gershon and his team to conclude that Bcl-xL and Bax work together during this crucial time of neurogenesis when CGNPs are proliferating. This is significant because establishing how cerebellar neurons are formed during development gives researchers insight for targeting medulloblastoma cells. For example, a protein important in regulating developmental cell death could be targeted in a potential treatment, causing apoptosis specifically in medulloblastoma cells.
Following this result, a small molecule inhibitor of Bcl-xL, called ABT737, was applied to CGNPs in order to disrupt this physical connection between Bcl-xL and Bax. ABT737 induced rapid apoptosis in cerebellar cells of young mice. This led to the discovery that induced apoptosis in CGNPs is developmentally regulated, and may be regulated similarly in medulloblastomas.
Brain sections of mice at developmental age, 20 days old. Cerebellum with deleted Bcl-xL gene sequence (right) shows increase apoptosis; control mouse with Bcl-xL intact (left). Image courtesy of Gershon Lab.
After finding the interaction between Bax and Bcl-xL leads to increase apoptosis in CGNPs, Ph.D. Candidate Katie Veleta of the Gershon lab deleted the gene sequence that codes for Bcl-xL in developing mouse cerebella. She predicted that through the apoptosis pathway (Figure 2), mice would display increased cell death. Recall that proliferating cells are those that are developing and moving to where they will remain as adult cells in the cerebellum. Veleta found that the size of the cerebellum is strikingly smaller and that the inner layer, normally housing the mature cells that previously proliferated, appears less populated in mice without Bcl-xL protein (Figure 3). This showed that while Bcl-xL deletion in medulloblastomas leads to increase in tumor apoptosis, it does not block tumor progression.
These findings are significant because they allow us to understand that other proteins must be involved in the fight to keep medulloblastoma cancer cells alive. Future directions utilizing apoptosis will explore how the activation or deactivation of related proteins prolongs the life of brain tumors. Advancements can lead to medulloblastoma treatment with higher specificity than chemo or radiation.
Currently, the graduate students in Dr. Gershon’s lab are exploring various routes of cerebellar development related to medulloblastomas, such as WNT signaling, DNA repair, and aerobic glycolysis. Genetic combinations have been discovered in recent years that ultimately “stop progenitors [aka CGNPs] from growing.” We know more about the brain now than ever, and our understanding of brain cancer is a direct result of neuroscience and molecular biology research. The era of expanding cancer care is upon researchers and physicians, allowing for more treatment routes than were readily available in the past century. Yet, as more and more children with medulloblastomas survive, their cancer treatment can still leave lasting effects on both the mind and body. The Gershon lab aims to expand these therapies even further, with the ultimate goal to give patients their ‘pre-diagnosis life’ back.