oncogenesis in neurosurgery

1. z
Oncogenesis in
Neurosurgery
Cell cycle, Telomeres and Antioxidants
Lee Wei Lun

2. z
Role of proto-oncogenes and suppresors
 Tumor suppressor: genes code for proteins that normally
operate to restrict cellular growth and division or even promote
programmed cell death (apoptosis)
 Proto-oncogenes: normal cellular gene that code for proteins
that function to drive the cell cycle forward
 Oncogenes: mutated gene or overexpressed gene contributing
to converting a normal cell into a cancer cell
altered

3. z
Loss of cell cycle control
 Mutation of proto-oncogenes
into oncogenes
 Inactivation of Tumor
suppressor genes
 Dysregulated cell growth
 Allows cells with damaged DNA
to progress to replication
 Tumor/Cancer cell formation

4. z
1) Point mutations will permanently activate proteins that normally interchange between active or
inactive states
2) chromosomal translocation. This occurs when the pieces of broken chromosomes reattach
haphazardly, leading either to the formation of a fusion protein and altered regulation of protein
expression
3) the proto-oncogene may exist in multiple copies in the cell, resulting in amplified expression

5. z
Cell Division and Cell Cycle

6. z
Cell Division
 Cell division is key to life
 To grow and develop: produce new cells—and allow for the death of
old cells
 Injury repair
 Reproduction
 Without sufficient cellular oversight, repeated rounds of unregulated
cell division can lead to life-threatening disease like cancer

7. z
 When a cell divides, it is essential that the new cell (also known
as the daughter cell) contains the same genetic information as
the old cell (also known as the parent cell)
 Mistakes during copying, or unequal division of the genetic
material between cells, can lead to cells that are unhealthy or
non-functional

8. z
Cell Cycle
 Cell cycle is an ordered series of events involving cell growth
and cell division that produces two new daughter cells
 The process are precisely timed and carefully regulated stages
of growth, DNA replication, and division
 Two major phases: interphase and the mitotic phase

9. z
 Interphase
 the cell grows and DNA is replicated
 the longest phase of the cell cycle
(90%)
 The three stages of interphase are
called G1, S, and G2.
 Mitotic (M) phase
 the replicated DNA and cytoplasmic
contents are separated, and the cell
divides.

10. z
M Phase
 First portion of the mitotic phase is called karyokinesis, or
nuclear division. Karyokinesis is divided into:
 prophase, metaphase, anaphase, and telophase
 The second portion of the mitotic phase, called cytokinesis, is
the physical separation of the cytoplasmic components

11. z
 Prophase: Nuclear envelope starts to dissociate into small
vesicles. Nucleolus disappears. The centrosomes begin to
move to opposite poles of the cell. microtubule fibers
lengthen.
 Prometaphase: Nuclear envelope is fully broken down.
Chromosomes are attached to microtubules from both
poles.
 Metaphase: all the chromosomes are aligned in the
equatorial plane.
 Anaphase: Sister chromatids separate at the centromere
and is pulled rapidly toward the centrosome to which its
microtubule is attached. Cell becomes visibly elongated.
 Telophase: Chromosomes reach the opposite poles and
begin to decondense, relaxing into a chromatin. Nuclear
envelopes form around the chromosomes.

12. z

13. z
Cell-cycle control
 Cell division is the result of a series of events involving DNA
replication (S phase), and the subsequent production of two
daughter cells (M phase)
 Fundamental task of the cell-cycle is to ensure that DNA is faithfully
replicated once during the S phase and that identical chromosomal
copies are distributed equally to two daughter cells during the M
phase
 Cell-cycle progression depends on discrete control points to achieve
that end

14. z
Checkpoints
 Control mechanisms that restrain cell-cycle transition or induce
apoptotic signalling pathways after cell stress are known as
checkpoints
 Key control points in the cell-cycle are at:
 G1 checkpoint - DNA quality
 end of the G2 checkpoint - chromosomal quality
 End Mitosis checkpoint

15. z
Cell Cycle

16. z
G1 Checkpoint
 Governs the transition from G1 to the S phase
 Rb protein:
 Under non-replicating conditions, Rb protein binds to and sequesters key
transcriptional factors E2F.
 When it is time for the cell to replicate, cyclin D binds to CDK4 and CDK6, forming
active kinases that phosphorylate Rb
 Phosphorylated Rb then releases factors that transcriptionally activate S phase
genes, permitting transition from G1 to the S phase.
 p16 protein:
 One of the most important regulators of the CDK4-cyclin D complex.
 It blocks the binding of CDK4 to cyclin D, preventing phosphorylation of Rb, thus
arresting the cell in the G1 phase

17. z
 p53 protein
 transcription factor that activates expression of proliferation-inhibiting
and apoptosis-promoting proteins in response to DNA damage
 Activated p53 upregulates p21 (a CDK inhibitor) which in turns inhibits
phosphorylation of Rb  Rb continues to bind to E2F
 Inactivation of p16, p53 or Rb; overexpression of cyclin D or CDK4
activity results in progression to the S phase without regard to
genomic integrity.
 common events in human glioma cells

18. z
G2 checkpoint
 Control of this checkpoint is, however, not as well understood as
the factors that regulate the G1 checkpoint
 Ataxia-telangiectasia-mutated (ATM)
 primary role of this cellular response to DNA damage is to prevent the
accumulation and propagation of genetic errors during cell division.
ATM is activated by the DNA damage sensor complex MRN (MRE1,
RAD50 and NBS1) and phosphorylates a great number of substrates.
 Cdc2 kinase is a mitogenic stimuli, ATM activates CHK2 that degrades
Cdc25 and induce arrest of cells before mitosis
 p53 protein also plays a role in G2 checkpoint

19. z

20. z
M Checkpoint: Spindle assembly checkpoint (SAC)
 Ensure that replicated DNA is partitioned equally between the two daughter cells
 Errors in mitotic spindle formation result in incorrect chromosome segregation
and chromosomal gains or losses in daughter cells
 diffusible mitotic checkpoint complex consisting of MAD2, BUBR1 and CDC20,
which acts as a potent inhibitor of APC/CCDC20 and anaphase initiation
 SAC prolonging mitosis until bipolar spindle attachment is achieved by all
chromosomes.
 Since mitotic entry is irreversible, cells cannot exit mitosis until the SAC is
satisfied. In cases where chromosome biorientation is not resolved following
prolonged mitotic arrest, cells follow two paths: either:
 apoptosis via caspase activation
 ‘slippage’, whereby cells exit M phase without chromosome segregation and enter the
next cell cycle as a single tetraploid cell

21. z

22. z
Oncogenes and Gliomas
 Low-grade gliomas usually express wild-type Rb and p16
proteins
 Inactivation of Rb protein, p16 and amplification of cyclin D1 and
CDK4 which regulate the phosphorylation of Rb protein, allowing
E2F activity  among the most frequent abnormalities that
occur in anaplastic astrocytomas
 Phosphatase and tensin homolog (PTEN) deletion – tumour
suppressor gene in glioma; upregulation of the CDK-inhibitor
p27 and modification of the function of cyclin D1

23. z
Oncogenes and Gliomas
 Human malignant gliomas with an abnormal regulation of the
p16/Rb pathways exceeds 80% of the cases
 p16 is deleted in approx 50% of glioblastomas
 Immunohistochemical studies showed lack of expression of the Rb
gene in 30% of glioblastomas
 BCL-2 protein is overexpressed in malignant gliomas
 Fas pathway, CD95 expression increases during malignant
progression from low-grade to anaplastic astrocytoma.
 Other chromosomal abnormalities: gain of chromosome 7 and
loss of chromosome 10, or EGFR amplification

24. z
Telomeres

25. z
Telomeres
 Telomeres consist of long tandem arrays of TTAGGG repeats,
bound by proteins, placed at the end of linear chromosomes.
 These non-coding telomeric repeats represent a buffer zone
preventing the adjacent coding region of the genome from erosion.
 In normal human cells, the telomeres decreases by some 5-20
repeats with every cell division. Therefore, telomere shortening
limits the number of times a cell can divide.
 Considered the mitotic clock by which cells count the number of
times they divided and regulate the onset of replicative
senescence in somatic cells.

26. z

27. z
Telomeres in defining cellular lifespan
 Human telomeres are regions of 4-15 kilobases (kb) of repetitive
hexameric (TTAGGG)n guanine-rich DNA sequences at the ends of
each chromosome.
 Has a T-loop that shields the end of chromosome from DNA repair
and DNA damage-sensing mechanism
 a complex of telomere-specific proteins, named shelterin complex,
binds and caps telomeres, preventing chromosomal ends from
being recognized by the DNA damage response (DDR) machinery.
 Telomeres are incapable of being fully replicated during each round
of cell division and undergo progressive shortening during normal
cellular proliferation.
 Eventually, they become so short that they trigger the DDR, causing
cell crisis  results in replicative senescence and eventually
checkpoint-driven cell death and apoptosis, defining cellular
lifespan.

28. z
 As previously described, in order to prevent degradation by
exonucleases or processing as damaged DNA, the telomere 3’
single-strand overhang folds back into the D-loop of duplex
telomeric DNA to form a protective ‘T-loop’,

29. z  Shelterin complex proteins interact
selectively with telomeric DNA and
localize to telomeres.
 Composed of six core components:
TRF1, TRF2, POT1, TIN2, TPP1 and
RAP1
 through the interaction with the
shelterin complex proteins, telomeres
protect chromosomes from
recombination, end-to-end fusion, and
recognition as damaged DNA 
providing a means for complete
replication of chromosomes
TRF1 and TRF2 (Telomeric Repeat Factors 1 and 2)
recognize duplex telomeric DNA
POT1 (Protection of Telomere 1) associates with the
single-strand telomeric DNA present at the 3′-
overhang
scaffolding subunit TIN2
RAP1 (Repressor Activator Protein 1), is a TRF2-
associated factor

30. z
Shelterin complex
 TRF1 has a key role in the modulation of telomere length. Its ADP-
ribosylation, allow telomerase to bind telomeres and start their
elongation.
 TRF2 serves to block recognition of telomeres as double-strand
DNA breaks.  Loss of TRF2 function leads to the activation of the
ATM kinase, formation of telomere dysfunction-induced foci (TIFs),
 induction of either senescence or apoptosis.
 POT1 serves to hide the 3′-telomeric overhang from telomerase
and from DNA damage sensing mechanisms. loss of POT1 
activation of the ATR kinase (ATM and Rad3-related kinase) 
formation of TIFs  induction of either apoptosis or cell cycle arrest

31. z
Pathways regulating telomeres length
1. Telomerase
 highly specialized ribonuclear reverse transcriptase enzyme
 extension of 5’-ends of the lagging DNA strand by adding TTAGGG
repeats onto the telomeres using its intrinsic RNA as a template for
reverse transcription
 Two major subunit recognized:
 h-TERC – provide the template for telomeres elongation
 h-TERT - contains a reverse transcriptase domain that catalyzes
this reaction

32. z
 In most somatic cells telomerase is produced at very low levels.
In contrast, many malignant cells are able to upregulate this
enzyme and extend their survival through continuous telomeric
elongation
 Vast majority of tumor cells use telomerase as preferred
mechanism for telomere maintenance

33. z
2. Alternative Lengthening of Telomeres pathway (ALT)
 ALT is rarely found in carcinomas but frequently activated in tumors
of mesenchymal and neuroepithelial origin like osteosarcomas,
liposarcomas or astrocytomas
 ALT positive cells contain several classes of extrachromosomal
telomeric repeats (ECTRs) in the nucleus. ECTRs correlate with
ALT activity but specific function of ECTRs in the telomere
lengthening process is still unknown.

34. z
Telomeres and Astrocytic tumor
 Functional telomeres protect chromosome ends from recombination and
fusion, and are therefore essential for maintenance of chromosomal stability
 Alteration of telomere length leads to genomic instability
 Almost all malignant tumors, world health organization (WHO) high grade
brain tumors are associated to higher telomerase activity than benign
tumors
 Increased telomerase expression has been also associated with higher
proliferative index, tumor grading, age, vascular endothelial proliferation,
poor outcome
 presence of elongated telomeres and of an ALT pathway was demonstrated
to be associated to a better prognosis in patients affected by glioblastomas.
 median survival of 542 days compared with 247 days in those without the ALT
phenotype.
 associated with elongated telomeres, benign biology and better prognosis

35. z
Telomeres and Astrocytic tumor
 In summary, high telomerase activity and reduced telomere
length seem to be features of high grade astrocytic tumors,
whereas ALT phenotype are specific of low grade ones and,
therefore, correlates with a better prognosis.

36. z
Telomeres and Oligodendrogliomas
 According to Nurnberg and Hiraga, oligodendrogliomas seem to
have compatible or longer telomeres than normal brain tissues
and detectable telomerase activity.
 telomerase activity associated to reduced telomeres as markers
of aggressive behavior in these neoplasms  anaplastic variant

37. z
Telomeres and Meningiomas
 Benign mengiomas (WHO grade I) show a variable telomere
length, sometimes slightly shorter than normal meningeal tissue
 Whereas atypical and anaplastic meningiomas show telomerase
activity associated with a significant telomere length shortening
 Summary: Telomere shortening together with telomerase activity
may be a critical step in pathogenesis of atypical and malignant
meningiomas and may correlate with their malignant behavior

38. z
Telomeres and Schwannomas
 Schwannomas usually show reduced telomeres, benign
pathological features, low proliferative indices and a lack of
telomerase activity.
 Rarely they can assume an aggressive behaviour that is
characterized by malignant pathological features and high
replicative potential that are usually associated to elongated
telomeres.
 Long telomeres allow tumor cells to maintain an high
proliferative capacity without the need of telomerase activity

39. z
Antioxidants

40. z
Oxidative metabolism
 Oxidation is a natural process of cellular metabolism. Reactive
oxygen species (ROS) is constantly produced during oxidative
metabolism.
 The formation of ROS is a consequence of aerobic metabolism
 low and moderate levels of ROS may act as modulator of cellular
proliferation and differentiation and also may be involved in the
expression of antioxidant genes, but high levels induce severe
cellular damages and also cellular death
 In normal conditions, the countenance in the intracellular ROS
levels is maintained with the contribution of antioxidant scavenging
systems and defense components.

41. z
 Brain tissue displays high oxygen consumption
around 20% of the total metabolic activity, which
are required to support normal function thus the
brain is particularly susceptible to oxidative
damage since does not have much antioxidant
storage
 Additionally, the brain has a high amount of fatty
acids  particularly sensitive to free radical attacks
 Compared to other tissue, brain tissue generate an
excessive amount of ROS

42. z

43. z
Oxidative stress
 Oxidative stress can be defined as the disturbance in the
oxidant-antioxidant balance.
 Enhanced oxidative stress can lead to modification of
cellular targets and induce cell damage and death.
 Cell damage and the following deficiency in cellular repair
processes due to the constant oxidative damage are
correlated with carcinogenesis.
 ROS interaction with DNA results in fragmentation with the
loss of bases and causes DNA strand breaks  can lead
oncogenic transformation

44. z

45. z

46. z
 Cancer cells have high levels of oxidative stress.
 Elevated levels of intrinsic oxidative stress due to the clustering of
factors such as enhanced metabolism, mitochondrial mutation,
inflammation and cytokines.
 Mitochondria in malignant cells present an overproduction of ROS
and structural and functional differences from mitochondria of
normal cells
 These malignant cells maintain excessive oxidative stress
compared to normal cells
 Malignant cells also tend to adapt aggressively and may deplete
cellular antioxidant capacity  lower levels of antioxidant enzymes

47. z
Glutathione - the main Non-enzymatic
antioxidant defence system
 Glutathione (GSH) is the most abundant intracellular non-
enzymatic antioxidant involved in the protection of cells against
oxidative damage.
 Oxidized into glutathione disulfide, GSSG
 elevated GSH levels increase the antioxidant capacity of many
cancer cells enhancing their resistance to oxidative stress. high
levels of GSH in cells are related to apoptosis resistance.
 This demonstrates the importance of the antioxidants in
favoring tumor progression
 GSH status in blood and in cancer cells were associated with
tumor growth in vivo

48. z
Enzymatic antioxidant defence systems
 Body’s endogenous defense mechanisms consist of several
enzyme systems that catalyze reactions to neutralize free radicals.
 They protect against free radical-induced cell damage. Between
them, the most studied includes:
 Superoxide dismutase (SOD),
 Glutathione peroxidase (GPx)
 Catalase (CAT)
 SOD eliminates superoxide radical (O2-).
 CAT and GPx are responsible for the disintegration of H2O2 and in
this way protect the cell against the formation of the most reactive
hydroxyl radical.

49. z
Superoxide Dismutases
 In mammals, there are three distinctive SODs:
 copper/zinc SOD or SOD1
 manganese SOD or SOD2 and
 extracellular SOD or SOD3.
 SOD1 is closely linked to cancer. Loss of SOD1 increases ROS levels
which cause oxidative DNA damage and promotes carcinogenesis
 On the other hand, cancer cells become increasingly dependent on
activated antioxidants such as SOD1 to prevent excessive cellular damage
and apoptosis during tumor progression.
 SOD2 also considered as a tumor suppressor - however its activity
appears to be tumor type/stage dependant.
 SOD3 is less well understood

50. z
SOD in brain tumors
 Lower SOD activity in astrocytomas, meningiomas, metastatic
tumors and other types of tumors was observed when compared
with their peritumoral tissues.

51. z
Catalase
 A enzyme that converts hydrogen peroxide into water and
oxygen.
 Gliomas appear to overexpress catalase when compared with
normal astrocytes.
 Although the enzyme production is increased, its effectiveness is
diminished as a consequence of tumor process, but it could be
also considered as a compensatory mechanism.

52. z
Glutathione Peroxidases
 GPx are another group of enzymes capable of reducing
hydroperoxides, using GSH as a substrate and generating
GSSG which is, once again reduced by the enzyme glutathione
reductase (GR) to GSH

53. z

54. z
 There is a well documented association between increased
consumption of antioxidants and decreased incidence of cancer.
And antioxidant supplements are recommended as part of a cancer
prevention diet.
 Summary: Antioxidant enzymes can antagonize initiation and
promotion steps of carcinogenesis
  yet antioxidant supplementation by cancer patients during
treatment is quite contentious
 Main strategies as a potential treatment are the inhibition of the
antioxidant enzymes and molecules in cancer cells and the
production of ROS leading to induced-apoptosis.

55. z
References
 Fueyo, J., Gomez-Manzano, C., McDonnell, T.J. (2005). Regulation of Cell-Cycle and
Apoptosis in Human Brain Tumors. In: Ali-Osman, F. (eds) Brain Tumors. Contemporary
Cancer Research. Humana Press.
 Chow, A. Y. (2010) Cell Cycle Control by Oncogenes and Tumor Suppressors: Driving the
Transformation of Normal Cells into Cancerous Cells. Nature Education 3(9):7
 Matthews, H.K., Bertoli, C. & de Bruin, R.A.M. Cell cycle control in cancer. Nat Rev Mol Cell
Biol 23, 74–88 (2022).
 Lichtor, Terry (2013). Clinical Management and Evolving Novel Therapeutic Strategies for
Patients with Brain Tumors || Telomeres and Brain Tumors. , 10.5772/45956(Chapter 16)
 Lichtor, Terry (2013). Clinical Management and Evolving Novel Therapeutic Strategies for
Patients with Brain Tumors || The Stance of Antioxidants in Brain Tumors. ,
10.5772/45956(Chapter 23)
 Ramírez-Expósito MJ, Martínez-Martos JM. The Delicate Equilibrium between Oxidants and
Antioxidants in Brain Glioma. Curr Neuropharmacol.