BRAIN CANCER STEM CELLS AND BEYOND
SCIENTIFIC REVIEW

Introduction:
In order for one to understand brain cancer stem cells, one must first understand stem cells in general as well as cancer stem cells.  Stem cells are the most primitive form of all cell types from all of the tissues and organs of a biological system.  The word “stem” lends to the straightforward interpretation of the kinds of cells they really are; cells that are the origin or beginning of all other cell types, and cells in which all these other cell types eventually can arise from.  Stem cells are often broadly classified as such, but many different kinds of stem cells exist providing for a broad spectrum of proliferated progenitor cells that ultimately lead to the creation of every adult cell type necessary for sustaining life.  Whether it be a hematopoietic stem cell giving rise to all of the cell types typically found in whole blood (Red blood cells, platelets, leukocytes such as neutrophils, eosinophils and basophils as well as monocytes (macrophages), and T and B cell lymphocytes), a cardiac stem cell giving rise to cardiac myocytes in the heart, a liver stem cell giving rise to hepatocytes in the liver, or a neural stem cell which can give rise to neurons, astrocytes, and oligodendrocytes (Kondo 2006), all stem cells are common with respect to their primitive nature and they ideally exist without any signs or markers of differentiation and lineage commitment.  The following diagram (figure 1) generalizes how some stem cells (neural stem cells for example) can proliferate and eventually become committed, specialized adult cells. 

Figure 1: Neural Stem Cell Hierarchy: An example of a stem cell proliferation pathway (Neural stem cell pathway) showing the proliferation pathways that are often seen in the development of mature, specialized cells of the nervous system.  This example was used because these specialized cells are often the subject of many brain cancers.

Stem Cell Biology:

Stem cells have characteristic properties that make them inherently distinct from other mature differentiated cells.  Their purpose and capabilities are quite different than those of a specialized mature cell which have been already maximally differentiated and reside in the organs or tissues in which they were educated for.  I know this because a stem cell possesses the biological property of self-renewal which will be explored and is essential in understanding cancer stem cells and ultimately brain cancer stem cells.  Another important property of stem cells is that of asymmetric division.  This is when each stem cell produces two progeny cells that are initially the same as the parent, but one has the genes that give it the potential to continuously proliferate and become a more differentiated mature cell.  The other progeny cell does not proliferate and remains identical to the mother stem cell from the original niche (Guo et al. 2006), thus stem cells are able to maintain themselves in numbers.  Both self-renewal and asymmetric division are hallmarks of stem cells. 

Self-renewal is the ability of a stem cell to produce a progeny cell that is identical to the mother stem cell.  When stem cells produce these clonogenic cells that will remain primitive and refrain from proliferation, the cells are said to have maintained stemness. The self-renewed stem cell is identical with regards to the expressions of cell-surface molecules, relative stemness and the nature of the chromosomes amongst other things.  Through the process of self-renewal, the mother stem cell is able to maintain the same quantity and arrangements of its DNA and chromosomes over time, giving the DNA of the stem cell an immortalized property.  This has been termed “Immortal DNA strand Cosegregation” by Rambhatla L et al.  Unlike the other daughter cell which subsequently may have increased gene expression and regulation due to its goal of becoming a specialized adult cell.  Figure 2 demonstrates self-renewal and asymmetric division.  Once a daughter cell has started the process of proliferation and commitment, it cannot dedifferentiate to its parent cell (Fig. 2: C cannot revert to A). In Figure 2, section C, the daughter cell is subjected to symmetrical division to generate identical copies of itself. 

  Figure 2: Stem Cell Self-Renewal and Asymmetric Division

Cancer Stem Cells – Experimental Evidence of Existence
There was once a time when scientists did not know that cancer stem cells actually existed.  In recent years however, the true identity and origin of cancer stem cells have been brought to light and many research scientists have shown that they do indeed exist.  Many scientists accept the possibility that cancer stem cells may have arisen from two possible sources: Either they arose from normal stem cells that have acquired mutations somewhere along the line in the cell cycle, or they arose from progenitor cells that have themselves acquired mutations and picked up the property of self-renewal that progenitors generally have no or very little ability to possess (Guo et al. 2006).  This would essentially be “dedifferentiation” in that the progenitor cell in question originally differentiated to become the progenitor cell it is, from a stem cell, but now has regained some of its stemness and reverted to the stem cell level which is above the progenitors in the hierarchy.  Anyone would be able to say that genetically this could probably lead to some abnormalities and perhaps loss of some control of the once normal cells’ destiny, leading to cancer and the formation of these cancer stem cells.  Evidence of the stem cell origin of cancer stem cells is laid out by Guo et al.  In it they explain using the hematopoietic stem cell model, that indeed oncogenic events take place.  In chronic myelogenous leukemia (CML), “the t(9;22) chromosomal translocation which leads to the formation of the p210 BCR-ABL1 oncoprotein, is present in the hematopoietic stem cells of patients (with CML) (Guo et al. 2006).”  They add to this evidence in a case involving acute lymphoblastic leukemia (ALL).  “In a subset of ALL, this t(9;22) breakpoint was detected in the CD34+ CD38+ CD19- hematopoietic stem cells.  The transcripts of another leukemic fusion oncogene, AML1-ETO, were also detected in the CD34+ Thy+ CD38- Lin- hematopoietic stem cells of patients with acute myelogenous leukemia (AML) in long-term remission (Guo et al. 2006).”  The evidence above tends to lead one to believe cancer is linked to stem cells in some way, however as a scientist one must still be aware of many other factors and occurrences at play when dealing with oncogenic activation. 

In another experiment where the goal was to show cancer stem cells also have a progenitor origin, the B lymphocytes and their CD19 cell-surface marker was utilized.  Similar in ways to the previous experiment described above, Castor et al. set out to detect the presence of p190 BCR-ABL, an oncogenic fusion gene, in acute lymphoblastic leukemia (ALL) patients.  With the knowledge that B cells possess cluster designation (CD) marker 19 on its surface amongst many other markers, they proceeded with this experiment.  The fusion gene “could only be detected in CD34+ CD38- CD19+                 B progenitor population of some ALL patients.  From these patients, the purified CD19+ B cells, but not the CD34+ CD38- CD19- hematopoietic stem cells, exclusively reconstituted CD19+ leukemia,”(Castor et al.) in NOD/SCID mouse models.  Therefore this enabled scientists to have solid proof that they had a cancer stem cell whose origin was a B lymphocyte progenitor.  J.E. Dick and colleagues, true pioneers in this field were able to show through the use of murine SCID models, that there was a very small subset or population of cells that were able to essentially initiate cancer or leukemia in this instance.  The cells became known as LIC’s (leukemia-initiating cells), and these cells were determined to have a special tumorigenic capability that was unique from all the other clonogenic cancer (leukemia) cells in the mouse (Bonnet D and Dick JE 1997, Hope KJ et al. 2004).  This was showing that it wasn’t the mass numbers of cancer cells that actually propagated the disease over time, but rather a distinct set of cells (LIC’s) capable of directing and controlling other cells in ways scientists still aren’t completely familiar with today.  These LIC’s are often loosely termed cancer stem cells now, and this model is now known as the “stem cell model” for cancer.  With all of these advances in cancer biology, the discrepancy in this field concerning whether these “cancer stem cells” actually exist or not should be alleviated over time.  Keeping an open mind on the reality of these stem cells will inevitably lead to a more accurate and easier understanding of brain cancer stem cells.

Brain Cancer and Brain Cancer Stem Cells
Brain cancer is an extremely horrifying disease that thousands of people must come to terms with each and every year.  Brain tumors are usually given a name depending on where the tumor originated and in what tissue(s) it originated from.  The more generalized tumors and common ones will be explored.  The most common type of brain tumor is known as a glioma.  Within the glioma class of brain tumors, there are other specific names of brain tumors that reflect the origin and cells/tissues affected; there are astrocytomas which are derived from glial cells known as astrocytes, and they often take up residence in the cerebrum, brain stem, and the cerebellum (Galderisi U et al. 2006).  Another important brain tumor in the astrocytoma family is called glioblastoma multiforme, which has been classified by the World Health Organization as a grade IV glioma, making it the most malignant of all gliomas (Vescovi A et al. 2006).  “Prognosis is poor and the median survival when radiotherapy and chemotherapy are combined is 14.6 months” (Vescovi A et al. 2006).  Some other important brain tumors are medulloblastomas, oligodendrogliomas and ependymoma.  There are many more that involve mixes between the various types and ought to be studied as well.

Brain Cancer Stem Cells – Evidence of Existence
There are mixed feelings and thoughts in the scientific world as to where brain cancer stem cells come from, if they even do exist.  Historically, it has been thought that tumors and cancers of the brain derived from the same things that cause the majority of cancers: oncogenic mutations in adult neuronal cells and/or their precursors (Kondo 2006).  The discovery of neural stem cells however has changed the way brain cancer has been studied.  It is now widely thought these neural stem cells take the brunt of these mutations and subsequently cause abnormal behavior of the specialized cells they give rise to.  Some evidence of this is that most deadly brain tumors such as medulloblastoma and glioblastoma multiforme (GBM) show markers of neural stem cell (NSC), such as Nestin, Bmi1, and Sox2, and differentiation markers, including the neuronal marker microtubule associated protein (MAP) 2, the astrocyte marker glial fibrillary acidic protein (GFAP), and the oligodendrocyte marker galactocerebroside” (Kondo 2006).  This suggests that these brain tumors share properties with NSCs, and this is consistent with the argument that brain tumors might be stem cell derived.  A final exemplification of evidence for brain cancer stem cells and with close ties with NSCs came from research done by Singh SK et al (2004).  This group of scientists, and others were able to isolate what they believed to be “cancer stem cells,” from two of the more common brain tumors.  They expand the tumor cells in “serum-free media containing basic fibroblast growth factor (bFGF) and EGF. They were able to culture floating aggregates (neurospheres) similar to what is expected of NSCs if similar culture condition was used in the expansion.  These aggregates self-renewed in culture and expressed markers of NSCs, such as Nestin, CD133, and Notch, as well as differentiation markers such as MAP2, GFAP and myelin proteins” (Singh SK et al 2004). Together, the evidence is moving towards a bigger picture. Most importantly, the aggregates from the glioblastomas and medulloblastomas were able to generate characteristic malignancies in animal models.  The conclusion is that these tumors must possess “cancer-initiating neural stem cell-like cells” (Kondo 2006).  From this evidence, immunopurification techniques were applied to identify the stem cell population with anti-CD133 (Kondo 2006) and later Singh and other scientists on the same team were able to purify brain cancer stem cells from human medulloblastomas and glioblastomas.  “They demonstrated that as few as 100 CD133-positive glioblastoma cells can form tumors in NOD/SCID brain, suggesting that CD133 is an excellent marker of brain cancer stem cells, as well as of normal stem cells” (Kondo 2006).  A key point in this experiment that I will stress because it should not be overlooked, was that the CD133-positive cells they had could actually possess the phenotype of the tumor from the patient in which the sample had originated from.  After differentiation and culturing in a dish, they could reproduce the exact same tumor that was identified beforehand.  Scientists are getting very close to solving this puzzle.

Implications and Potential Future Therapies
The knowledge that both cancer stem cells and brain cancer stem cells exist now challenge the method by which cancer is treated. The model of cancer as originating from a small subset of stem cells ensures survival of the brain tumor, while the non-tumorigenic cells are there for other purposes and make up the bulk mass of the tumor. Scientists need to develop specific ways to target the brain cancer stem cells and destroy the tumors’ ability to thrive.  Traditional treatment regimens have had the goal of shrinking the brain tumor and killing off as many “homogenous” tumor cells as possible.  The current “status quo” of cancer therapy basically involves using chemotherapeutic agents that target the characteristic rapidly cycling tumor cells (usually non-tumorigenic).
This is good, but if it doesn’t address the root of the problem being the (brain) cancer stem cells themselves, then it is highly unlikely success in treating brain tumors will be achieved.  This traditional way is the old model, and we now know that the brain tumor population is heterogeneous containing both non-tumorigenic and tumorigenic cells (cancer stem cells).  Others have suggested indepth examination of normal stem cells for areas of potential therapeutic targets for cancer treatment (Galderisi et al. 2006).  In the future it may be wise to take an in depth look into many of the genes associated with normal stem cell self renewal like Wnt, Sonic Hedgehog, Notch etc.  These genes may give informative insights and direction on how to manipulate and control self renewal of brain cancer stem cells.  Also, if we could somehow perfect our in vitro methods with brain cancer stem cells, we might be able to learn more and more about them and ultimately prevail over the existence of brain tumors.  The blueprint is laid out and any scientist familiar with cancer stem cells probably has a good idea of what the broad goal of this new age of cancer therapy should be.  The next step is implementing successful therapy policy, and start saving lives today.

References:

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Castor A, Nilsson L, Astrand-Grundstrom I, Buitenhuis M, Ramirez C, Anderson K,
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Hope KJ, Jin L, Dick JE (2004) Acute myeloid leukemia originates from a hierarchy of
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Kondo, Toru (2006) Brain cancer stem-like cells.  Eur J Cancer 42:1237-42.

Muhammad AH, Clarke MF (2004) Self-renewal and solid tumor stem cells. 
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Acknowledgements
This review was prepared by Michael J. Poynton III, a graduate student in the Stem Cell Biology Class, Graduate School of Biomedical Sciences, University of Medicine and Dentistry of New Jersey:

Teaching Assistant: Kathy Trzaska