“Glioma Stem Cells Promote Radioresistance by Preferential Activation of the DNA Damage Response (2006),” by Shideng Bao, Qiulian Wu, Roger McLendon, Yueling Hao, Qing Shi, Anita Hjelmeland, Mark W. Dewhirst, Darell Bigner, Jeremy Rich

In 2006, Shideng Bao and colleagues published “Glioma Stem Cells Promote Radioresistance by Preferential Activation of the DNA Damage Response,” hereafter “Glioma Stem Cells,” in Nature. The study describes how cells within a glioblastoma, a type of fast-growing brain tumor, have a high expression of a protein called CD133, which is associated with neural stem cells. Among those cells with high expression of CD133, there exist many stem cells called glioma stem cells. In the paper, Bao and colleagues demonstrate that glioma stem cells are more resistant to radiation compared to other cells within a glioblastoma tumor, and that their resistance has to do with their ability to repair DNA. “Glioma Stem Cells” was one of the first studies to identify the role of glioma stem cells in resistance to radiation and laid the framework for future studies that investigated their role in tumor progression and recurrence as well as novel treatments targeting those cells.

1. Background and Context
2. Article Summary
3. Impact

Background and Context

At the time of publication, the authors of “Glioma Stem Cells” were all affiliated with Duke University Medical Center in Durham, North Carolina. At that time, the primary author of the study, Shideng Bao, was an assistant professor at the Preston Robert Tisch Brain Tumor Center at the Duke University School of Medicine, where he studied glioma stem cells. Jeremy Rich, the senior author of the paper, studied cancers of the nervous system at Duke University Medical Center. His work involved understanding the importance of cancer stem cells to better treat cancer patients. Bao and Rich collaborated on their research at Duke. As of 2025, Bao is the director of the Center for Cancer Stem Cell Research at Cleveland Clinic Main Campus in Cleveland, Ohio. His laboratory focuses on understanding the interactions between glioma stem cells and the tumor microenvironment with the goal of developing new treatments for glioblastoma patients. As of 2025, Rich is the deputy director for research for the University of North Carolina Lineberger Comprehensive Cancer Center in Chapel Hill, North Carolina, where his research laboratory focuses on identifying novel treatment options for advanced cancers.

Throughout the study, the researchers discuss terms within the field of neuro-oncology, such as glioblastoma multiforme, brain cancer stem cells, and ionizing radiation. Glioblastoma multiforme, or GBM, is one of the most aggressive forms of brain cancer that originates directly from glial cells, which are non-neuronal cells that support the brain, neurons, and the neural system. The median survival for patients after diagnosis with GBM is roughly around fourteen to sixteen months. One factor that researchers have identified that contributes to the aggressiveness and recurrence of GBM tumors is their resistance to treatments, such as ionizing radiation. Ionizing radiation is a type of radiotherapy that uses beams of radiation to damage the DNA of tumor cells inside the body, which can destroy their ability to replicate into more tumor cells. If a cell cannot repair that damage, a damaged cell can even undergo apoptosis, which is the process of programmed cell death that occurs if a cell incurs too much damage. Ionizing radiation is a common treatment for patients with GBM.

The researchers in “Glioma Stem Cells” specifically focus on the relationship between how particular cells within the GBM brain tumor called brain cancer stem cells can contribute to a tumor overall being resistant to radiation, or radioresistant. Stem cells are cells in the body that can develop into many different types of cells through a process called differentiation. They can self-renew, a process in which stem cells can produce more of the same type of stem cell. Stem cells typically function to repair damaged tissue. In the field of cancer research, researchers hypothesize that there are a small number of stem cells within a tumor that are likely responsible for initiating a tumor and maintaining its growth. Even if radiation damages or destroys a majority of the tumor cells, a few stem cells in the tumor that survive can initiate the growth of a recurrent tumor. Those cancer stem cells are present in brain cancers, and, specifically within GBM, those cells are glioma stem cells.

Article Summary

“Glioma Stem Cells” consists of two major sections, an untitled main section and a titled “Methods.” In the first section, Bao and colleagues describe their experiments performed throughout the study and the main findings of those experiments, including their finding that populations of cells with high CD133 expression contain a large subset of glioma stem cells that are more resistant to radiation due to their ability to repair DNA damage. In “Methods,” Bao and colleagues describe the specific cells and laboratory techniques used throughout their experiments, including an alkaline comet assay to separate broken pieces of DNA as well as immunoblotting and immunofluorescence staining to detect which cells were expressing their proteins of interest.

Bao and colleagues first introduce the aggressive nature of glioblastoma tumors and describe how other researchers have argued that cancer stem cells drive tumor recurrence. They also state that one marker to identify neural stem cells and brain cancer stem cells is a protein called CD133, or Prominin-1, which exists on a cell’s surface. They cite previous studies in which researchers demonstrate that cells within a glioblastoma tumor expressing CD133, which they hypothesize to be glioma stem cells, have a greater ability to grow and form tumors compared to cells within that tumor that do not express the CD133 protein. Given the importance of CD133 in tumor growth and formation, Bao and colleagues investigated whether those glioma stem cells contribute to a tumor’s resistance to ionizing radiation.

Bao and colleagues discuss their first finding that when they treated glioblastoma cells with radiation, there was an increase in the number of cells expressing the CD133 protein after the radiation treatment. They irradiated those glioblastoma cells on their own. They also implanted human glioblastoma cells into mice and directly irradiated those mice. After irradiating the GBM cells on their own and in the mice, they reported an increase in the number of cells that express the CD133 protein. Bao and colleagues note the fact that the radiation did not cause cells to start expressing the CD133 protein. They state that the radiation likely killed the cells that did not express CD133, and the cells that had survived were the cells that expressed CD133. That means that there were more surviving cells with that CD133 protein that were then able to replicate to produce more cells in the tumor with CD133. Since CD133 is a marker to identify brain cancer stem cells, the authors argue that that result provided some evidence that glioblastoma stem cells might be more resistant to radiation than other cells. Cells with high levels of CD133 are CD133+, and those with low levels of CD133 are CD133-.

Bao and colleagues also investigated how implanting groups of cells with each group containing various amounts of the CD133 protein could affect GBM tumor growth. They found that groups where cells had higher CD133 protein levels showed a greater increase in tumor growth compared to groups where cells had lower CD133 levels.

Then, they attempted to confirm that those cells that expressed CD133 were truly cancer stem cells. The researchers report that they looked for several features of cancer stem cells. One feature of cancer stem cells in the brain is that the cells can grow into spherical cluster formations known as neurospheres. They found that CD133+ cells formed neurospheres, whereas CD133- cells rarely formed those neurospheres. A second feature of brain cancer stem cells is that those cells typically express certain proteins other than CD133, including SOX2, Musashi, and Nestin, and Bao and colleagues found that their glioblastoma cells that expressed CD133 also expressed other proteins that are markers for brain cancer stem cells. A third feature of neural stem cells is the ability to differentiate into more specialized cells in the nervous system such as astrocytes or oligodendrocytes, which are both specialized central nervous system cells. Bao and colleagues found that their glioblastoma cells that expressed CD133 were able to differentiate into those specialized cells of the central nervous system. As a result of all of those findings, the researchers reported that the glioblastoma cells in the experiments that expressed CD133 were likely glioma stem cells due to the fact that those cells with CD133 exhibited multiple characteristics of brain cancer stem cells.

Bao and colleagues further investigated why CD133+ glioblastoma cells were more resistant to radiation than CD133^- glioblastoma cells. To do that, they looked at the levels of particular proteins, such as Caspace-3 and annexin V, that are associated with apoptosis. Since radiation induces cell damage, cells often undergo apoptosis after radiation exposure. In Bao and colleagues’ experiment, they found that glioblastoma cells with high expression of CD133 expressed less of those proteins associated with apoptosis than cells with lower expression of CD133. Hence, cells with higher CD133 levels were likely experiencing less apoptosis than cells with lower CD133 levels. Since Bao and colleagues previously confirmed that the glioblastoma cells with higher CD133 levels were likely glioma stem cells, their results on apoptosis provided more evidence for the idea that glioma stem cells are more resistant to radiation than other glioma cell types.

The researchers also compared the growth rate of CD133+ cells both exposed to radiation and not exposed to radiation. They found that when CD133+ cells underwent radiation exposure, their growth increased over fourfold. That result confirmed again that the glioblastoma cells with high levels of CD133, which were glioma stem cells, were more resistant to radiation and were able to grow more rapidly after radiation than other CD133- glioblastoma cells that had low expression of CD133 and, thus, were not glioma stem cells.

Bao and colleagues also confirmed the ability of the CD133+ glioblastoma cells to form tumors after exposure to radiation compared to CD133- glioblastoma cells. They irradiated CD133+ and CD133- glioblastoma cells and then injected the cells into mice to assess the ability of that cell type to form a tumor after radiation exposure. The scientists found that when exposed to a low dose of radiation, treated irradiated CD133+ cells reformed tumors at a similar rate to untreated CD133+ cells. They found that cells with high expression of CD133 that took high doses of radiation had a decreased ability to form tumors. In comparison, CD133- glioblastoma cells exposed to that same low dose of radiation were completely unable to form tumors after the researchers injected them into mice. Based on that result and the results from previous experiments, Bao and colleagues state that populations of CD133+ glioblastoma cells have a higher proportion of cancer stem cells within the population when compared to populations of CD133- glioblastoma cells. They state that those glioma stem cells are more resistant to radiation compared to CD133- glioblastoma cells.

Bao and colleagues also investigated the mechanisms by which CD133+ cells have an increased resistance to radiation. Radiation typically induces damage to the DNA of a cell. If a cell can repair the DNA damage, then the cell may survive and continue to replicate. As a result, Bao and colleagues investigated whether there is an upregulation in pathways involved in DNA repair in glioblastoma cells after radiation. To do that, they induced DNA damage through either radiation or a drug that mimics the effects of radiation. They examined whether there were increases in the activation of proteins that are involved in DNA repair, such as RAD17, CHK1, and CHK2. They found that overall, CD133+ cells had higher levels of those activated proteins involved in DNA repair after radiation compared to CD133- cells. Bao and colleagues state that the result suggests cells with higher CD133 expression can activate more proteins to help repair damaged DNA, making them more resistant and better able to survive after damage from radiation. To confirm that result, Bao and colleagues used an inhibitor of proteins in the DNA damage repair pathway to see if that would impact the ability of CD133+ cells to repair their DNA. They found that disrupting the DNA repair pathway with that inhibitor disrupted the ability of CD133+ cells to repair their DNA and in turn, resist radiation. Since Bao and colleagues state that most of their CD133+ glioblastoma cells are likely glioma stem cells, that result suggests that glioma stem cells are more resistant to radiation than other glioblastoma cells due to their ability to repair DNA damage induced by radiation treatments.

Lastly, Bao and colleagues state that CD133+ cells are more resistant than other glioblastoma cells to radiation through the ability to repair DNA damage caused by that radiation. Since they state that most of their glioblastoma cells with high CD133 levels are likely glioma stem cells, their results suggest that glioma stem cells are the cells that are more resistant to radiation than CD133- cells. The glioma stem cells within a glioblastoma tumor contribute to the tumor’s overall resistance to radiation and potential for recurrence after radiation treatment. They state that therapies that target the ability of those glioma stem cells to repair DNA may decrease a glioblastoma tumor’s resistance to radiation and increase the efficacy of radiation as a treatment for patients with glioblastoma tumors.

The second main section of the “Glioma Stem Cells” is “Methods,” which walks through specific details on the cells as well as the laboratory techniques that Bao and colleagues used throughout the experiment. They state that they received samples from GBM tumors in pediatric patients and implanted those cells into immunocompromised mice. After the tumors in the mice grew, Bao and colleagues used the cells from those mice tumors for their experiments. They also used cells directly received from human glioblastoma patients who underwent surgical removal of their tumor. To assess the amount of DNA damage and repair in the cells, Bao and colleagues used the alkaline comet assay. The alkaline comet assay involves placing cells in a gel and applying an electric field that separates broken pieces of DNA. To investigate the expression of proteins, for example, proteins that are markers for stem cells or proteins involved in the DNA repair pathway, they used techniques such as immunoblotting and immunofluorescence staining, which both use antibodies that bind to proteins of interest and allow for analysis of that protein. To identify which cells had high CD133 expression and which cells had low CD133 expression, they labeled each cell type with fluorescent dyes.

Impact

As of 2025, researchers have cited “Glioma Stem Cells” over 7,600 times. Bao and colleagues’ study was one of the first to demonstrate that cancer stem cells are more resistant to radiation and paved the way for future research on the role of cancer stem cells in other cancers. For example, researchers at the David Geffen School of Medicine at the University of California at Los Angeles in Los Angeles, California, demonstrated that breast cancer-initiating stem cells are resistant to radiation. Other studies that cited Bao and colleagues’ study involve further understanding of how glioma stem cells behave within a tumor. In 2014, researchers at the Broad Institute of Harvard University, the Massachusetts Institute of Technology, and Massachusetts General Hospital in Cambridge, Massachusetts, investigated the cellular heterogeneity of GBM tumors. They found that cells in glioblastoma tumors can exist in different stem-cell-like states. That finding built onto Bao and colleagues’ study because both provided more knowledge on the behavior of glioma cells, which were then integral for therapeutic strategies.

Bao and colleagues’ work in “Glioma Stem Cells” also set the stage for future work on how to target those cells for therapeutic benefit. A 2018 Cleveland Clinic News Release discusses Bao and colleagues’ research investigating a drug called ibrutinib, which inhibits glioma stem cells. That study demonstrated the effectiveness of the drug in mice and led to a clinical trial of the drug with radiation to treat patients with GBM. The researchers found that the combination of radiation and the drug increased lifespan and repressed resistance. In 2023, researchers at the Baptist Health Miami Cancer Institute in Miami, Florida, along with colleagues at the Cleveland Clinic in Cleveland, Ohio, demonstrated that ibrutinib is safe in combination with chemotherapy and radiation in a clinical trial with GBM patients. The survival outcomes of patients treated with ibrutinib were promising compared to historical survival data. Because of their role in the initiation, progression, and recurrence of GBM, glioma stem cells have become a target for novel cancer therapies that target those glioma stem cells to improve treatment for people with GBM.

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