CSF FLUID-

BIOMARKERS FOR
CNS
MALIGNANCIES
Dr Rituraj
Dr Gireesha
Dr Charanjeet Ahluwalia
TABLE OF CONTENTS
 INTRODUCTION

 CURRENT METHODS AND WHY THE NEED FOR CSF BIOMARKERS?

 CSF CYTOLOGY

 NEW BIOMARKERS TARGETS

 CSF BIOMARKERS FOR MAJOR CNS MALIGNANCIES.

INTRODUCTION
 Central nervous system (CNS) tumors are a major health
problem and surpass leukemia as the leading cause of
cancer death among children and adolescents.
 CNS tumors are associated with high rates of mortality and
morbidity.
 The new 2016 WHO classification of CNS tumors is based
on molecular parameters in addition to histology to define
many tumor entities, thus formulating a concept for how CNS
tumor diagnoses should be structured in the molecular era.
WHY THE BIOMARKERS?
 Primary malignant intracranial tumors of the CNS can roughly be
divided into gliomas, primary CNS lymphomas (PCNSL),
medulloblastomas, and primary neuroectodermal tumors that are
common in children.
 CNS is also a frequent site of metastasis, with an estimated incidence
of 9–17% for all types of cancer combined.
 Lung cancer, breast cancer, and melanoma are the most common
cancers to metastasize to the brain.
 Current methods for diagnosing and monitoring CNS tumors, such as
cerebrospinal fluid (CSF) cytology and MRI, have low sensitivity
and specificity, respectively.
 Serial MRI and CT are suboptimal methods to detect
changes associated with treatment response or disease
progression.
 Recent therapeutic advances have the potential to shift
treatment approaches from disease-type-based therapy to
individual-genotype-based therapy.
 For genomic characterization of CNS tumors we require
tissue biopsy.
 The delicate nature of the nervous systems makes tumors
located in the CNS notoriously difficult to reach, which poses
several problems.
 Surgery may not be an option at all due to the location, size,
or nature of the anomaly.
 Pseudo- progression: The appearance of increased lesion
size on radiologic images that are caused by post surgery or
chemo-radiation effects and not to tumor progression.
 Biomarkers may help address these CNS tumor-specific
problems by providing information about the tumor from a
more accessible sampling location than the tumor site itself.
WHY CSF?

Tumor
markers
already
established
for various
tumors
 Plasma analysis helps in detection, mutation profiling,
evaluation of response to therapy and recurrence, and detection
of emergent resistance to ongoing therapies; in various cancers.
 Due to blood–brain barrier (BBB), blood is not an ideal fluid to
accurately evaluate biomarkers derived from CNS tumors.

A recent study on blood samples collected from 640 patients with various cancer
types suggested that ctDNA is a promising biomarker for solid tumors outside the
CNS, including bladder, breast, colorectal, gastro-oesophageal, hepatocellular,
ovarian, pancreatic, and head and neck cancer and melanoma.
However, ctDNA was detected in the blood of fewer than 10% of patients with
gliomas.

Several studies also reported the sensitivity for circulating tumor cells in the blood of
patients with glioblastoma as low as 21% to 39%.
CSF CYTOLOGY
 Sample can be obtained at the time of tumor surgery or by
lumbar or intracerebroventricular (ICV) reservoir puncture.
 To avoid false positive results due to sloughing of tumor cells
at the time of surgery, a recovering interval of one to two
weeks is suggested
 7.5-10.5 mL of CSF to be withdrawn
 Immediately processed (cell counts can diminish by up to
50% within 2 h of collection)
Processed by centrifugation
(CytospinÒ) at 800 g for 3-5
min, air-dried for 10-15 min
and stained with May-
Grunwald Giemsa (MGG)
stain solution for 10-15 min.
Liquid-based cytology (LBC)
method has been suggested to
better detect malignant cells in
CSF from solid tumors.
The collected samples need to be
added to 10 mL preservation
solution, mixed and stood for 15
min.
Slides are fixed in 95% ethanol for
15 min and stained by standard
Papanicolaou method.
LIMITATIONS OF CSF
CYTOLOGY
 Qualitative assay based on microscopic examination with high
inter-observer variability.
 Requires the presence of morphologically intact tumor cells to be
examined under the microscope (low sensitivity)
 False-negative results are common.
 Due to the shedding of malignant cells into the CSF
intermittently and in low numbers, inconsistent presence of cancer
cells in the CSF is expected.
 So it is recommended that CSF analysis should be repeated if
initially negative.
POTENTIAL BENEFITS OF CSF
BIOMARKERS FOR PATIENTS WITH
CNS TUMORS
(1) Early diagnosis before surgery
(2) Diagnosis of tumors that are not amenable or typically
treated by surgery
(3) Early identification of tumor recurrence
(4) Monitoring of response to treatment
(5) Facilitating use of precision-medicine therapies
WHAT ALL CAN BE USED AS
BIOMARKER?
METABOLOMICS AND
PROTEOMICS
 Metabolites are intermediates or products of biochemical reactions such
as glycolysis or synthesis of cellular macromolecules.
 Human Metabolome Database (HMDB) lists 450 metabolites that
have been identified in CSF.
 Cellular metabolites are altered in infective, inflammatory and
neoplastic brain tissues and are secreted in CSF.
Glycolysis and TCA cycle metabolites

 Tumor cells use glucose for energy and transform it into lactate by the
enzyme lactate dehydrogenase (LDH) (WARBURG EFFECT)
 But normal cells, transform glucose into pyruvate and then metabolize
pyruvate in the mitochondria by oxidative phosphorylation.
 So, tumor cells rely more in aerobic glycolysis and lactic acid
fermentation, than the TCA cycle, for ATP production.
 Lactate can be measured in the CSF of patients with brain tumors.
Chow et al retrospectively reviewed the
lactate concentration in CSF of 2268
patients from 1992 to 2002. Only 159
patients had lactate levels of 2 mmol/L or
greater, and 4% of them were diagnosed
with brain or meningeal tumors.

In a case report published in 2008, Blüher

et al reported that in a 3-year-old boy with
malignant meningeal melanoma,
measurement of pH in CSF showed central
lactic acidosis despite alkalosis in
peripheral blood.
 Glucose and glutamine are the main substrates that tumor cells use to
produce energy.
 Glucose and glutamine provide acetyl-CoA and NADPH, which are
needed to synthesize lipids.
 Hypoglycorrhachia, a decrease in glucose levels in the CSF, is
associated with bacterial and fungal infections, but it also has
been described in patients with brain tumors.
 This can be used as screening tool.
 Studies have shown elevated
D-2-HG levels in the CSF of
patients with IDH-mutant
gliomas.
 Also, citric acid and isocitric
acid have been shown to be
elevated in the CSF of
patients with IDH-mutant
gliomas.
CIRCULATING TUMOR CELLS
(CTCs)
 CSF fluid flow cytometry is a useful addition to CSF cytology to identify
CTCs.
 Cytology examines morphologic patterns, and flow cytometry has the
potential to provide information about cell surface protein expression.
 It has to be said that both false negative and false positive results
(especially at low cell counts, < 25 cells/uL) can occur with flow cytometry
too
 Poor differential ability between mitoses and neoplastic cells is also reported
 The cell count and the percentage of neoplastic cells reported in the CSF by
both cytology and flow cytometry is significantly higher compared with those
found to be positive by flow cytometry alone.
 So a combination of CSF cytology and FCM is currently considered
best
 CellSearch® is a recent technique using molecular markers to detect and
enumerate circulating tumor cells in the CSF.
 Only system approved by the FDA for the enumeration of CTCs in the
blood of patients suffering from metastatic breast cancer, prostate
cancer, or colorectal cancer
CIRCULATING TUMOR DNA (ctDNA)

 Molecular techniques such as digital droplet PCR (ddPCR) and

BEAMing were implemented to detect specific mutations in bio fluids in
past.
 However, next-generation sequencing (NGS) provides an opportunity to
screen for novel mutations and even gene fusions and amplifications
with a limit of detection below 1%.
 The rate of ctDNA detection in CSF is not 100%, but relatively similar to
that of other biofluids.
 The rate of ctDNA detection in CSF is higher than the rate of detection
of intact tumor cells by microscopic examination.
RNA
 MicroRNAs (miRNAs): discovered in the 1990s.
 They are short, non translated fragments of RNA that bind to 3’
untranslated regions of messenger RNA and repress protein
translation in several molecular pathways.
 4 major methods for detection of miRNAs in CSF: qRT-PCR,
DNA microarrays, in situ hybridization and RNA sequencing.
 qRT-PCR is quantitative, extremely sensitive, highly specific and
reproducible-PREFERRED
 It offers the advantage of amplifying small amounts of RNA into a
much larger sample of cDNA; thus only a few neoplastic cells are
needed in the CSF.
ROADMAP
GLIOMAS
 Gliomas are the most common malignant primary brain tumors.
 They account for approximately 27% of all primary brain tumors and
80% of malignant primary brain tumors.
 Gliomas recur in almost all cases despite maximal treatment.
However, in low-grade gliomas this may take years.
 With time, gliomas acquire new mutations that can drive progression
to a higher tumor grade.
 Glioblastoma (GBM) is most lethal and most common form of
primary CNS tumor.
GLIOMA-DERIVED PROTEINS
 Formation of new vasculature is important for tumor growth.
 Proteins that stimulate neovascularization are important, both as
therapeutic targets and as possible biomarkers. Eg. vascular
endothelial growth factor (VEGF).
Sampath et al. (2004) reported very low VEGF levels in CSF samples from
Hypoxia
 control -- glioma
patients, cells
while the increase the
concentration wasproduction
significantlyof VEGF--secreted
higher in
into thewith
patients tumor microenvironment
high-grade astrocytic gliomas but not in nonastrocytic
tumors, suggesting that VEGF levels in CSF may potentially have
diagnostic value for the identification of astrocytic tumors.

When correlated to overall survival, however, they found no significant

correlation, indicating that CSF VEGF levels might not be of prognostic
value
Ribom and colleagues (2003) looked solely at grade II
gliomas and did not find any detectable VEGF in CSF.
 Fibroblast growth factors (FGFs) are a family of growth
factors that are involved in embryonic development,
angiogenesis, cell differentiation, proliferation, and migration.
 Acidic and basic types.

Peles et al. (2004) reported significantly higher median

bFGF levels for high-grade gliomas.
A CSF concentration of bFGF above the median was
significantly associated with shorter overall survival.

However, another research group detected only very low

levels of bFGF in the CSF of 2 of 7 patients with low-grade
astrocytoma (Ribom et al., 2003).
 Extracellular matrix (ECM) is a significant component of tumor local
micro-environment.
 Matricellular proteins are nonstructural secreted proteins that
interact with the ECM.
 Several matricellular proteins are under investigation as cancer
biomarkers.
• Tenascin-C is a glycoprotein that is Tenascin C levels in CSF were reported
highly expressed during development to be increased in astrocytic tumors
in migratory cells. Expression in adult when compared to nonastrocytic
tissue is limited unless the tissue primary CNS tumors, metastases, and
undergoes remodeling. noncancerous controls.
• This can be due to physiologic Patients with high-grade astrocytomas
processes in, for example, wound had higher tenascin-C levels than
healing, but is also seen in patients with low-grade astrocytomas
malignancies. (Yoshida et al., 1994).
 Osteopontin (OPN) is a secreted phosphoprotein that binds to
integrins and can activate intracellular signaling.
 OPN plays an active role in cell adhesion, angiogenesis, and immune
responses.
 It has been implicated in several types of cancer, including glioma,
carcinomas of the breast and prostate, colon carcinoma,
medulloblastoma, and atypical teratoid/rhabdoid tumors.

Schuhmann et al. (2010) identified Another study showed that cleaved

increased levels of OPN in CSF of GBM OPN fragments in CSF were markedly
patients when compared to controls. increased in glioma patients compared
to controls with systemic cancer, and
noncancer patients.
 Matrix metalloproteinases (MMPs) are peptidases that can cleave
almost all components of the ECM.
 Different subtypes of MMPs include collagenases and gelatinases,
 Exist in both membrane-bound and secreted forms
 Detected by ZYMOGRAPHY.
 MMP-2 (gelatinase A) and MMP-9 (gelatinase B) are associated with
tumor invasion and are the most upregulated MMPs in gliomas.

The increasing activity of MMP-9 in
the CSF long before progression
was seen on magnetic resonance
imaging (MRI) suggested it could
possibly be a marker for disease
progression (Wong et al., 2008).
(Forsyth et al., 1999; Raithatha et al., 2000; Liu et
al., 2010b). Friedberg et al. (1998) noted that the
uncleaved form of MMP-9 (proMMP-9) was present
in the CSF of all tested patients with cancer located
in the CNS, whether primary or metastatic, while it
could not be detected in any of the control samples.
GLIOMA-DERIVED ctDNA
 Wang et al. (2015) found that ctDNA could be detected in CSF of
almost all high-grade tumors, but not in low grade gliomas.

 Proximity of the tumor to a CSF reservoir (e.g., the ventricles) greatly

increased the likeliness of finding detectible ctDNA levels.

 The overall detection level of ctDNA was 70%.

 This indicates that ctDNA detection is a strong candidate for diagnostic
clinical application
 Alu elements are the most common retrotransposable elements in
human DNA. They comprise 11% of the human genome.
 Cancer cells release ctDNA with more variable lengths than
apoptotic cells, and release longer fragments into their
environment compared to normal cells.

Shi et al. investigated whether the integrity of DNA

fragments in serum and CSF from glioma patients
differed from controls.
The ratio of short Alu repeats (Alu115) versus long
Alu repeats (Alu247) was used as a readout.
The long:short DNA ratio was significantly different
when in CSF but not in serum.
GLIOMA-DERIVED RNAs
 The most notable miRNA in glioma is miR-21.
 Anti apoptotic effects and promotion of invasion
 Expressed in normal brain tissue, levels are upregulated in GBM
 miR-15b is a regulator of cell cycle progression in glioma cells
 The combined presence of mir-15b and miR-21 differentiates
glioma from other intracranial pathologies with a sensitivity of
90% and a specificity of 100%
When miR-21 levels were compared in longitudinal
samples of GBM patients (intraoperative vs.
posttreatment), disease activity and treatment
response were reflected in the concentration of miR-
21 (Teplyuk et al., 2012).
GLIOMA-DERIVED EXTRACELLULAR VESICLES(EV)

 Akers et al. (2013) isolated EVs from the CSF of 13 GBM patients and 14
noncancerous controls.
 The miR-21 content of EVs from GBM patients was on average 10-fold
higher than that of the control group.
 None of the samples from the control group contained more than 0.25 miR-21
copies per EV
 85% of samples from GBM patients carried >0.25 copies/EV (p < 0.001).
 This result suggets the possibility of EV miR-21 content >0.25 copies/EV as a
biomarker for GBM diagnosis.
 Sample taken prior to surgery contained 50-fold more miR-21 per EV
compared to the sample taken 3 months after surgery.
 This suggests a correlation between tumor burden and EV miR-21
content in CSF

Shi et al. (2015) showed that higher exosomal miR-21 levels in

CSF significantly correlate with glioma recurrence, which could
provide a useful tool for tracking disease activity.

Exosomal miRNA content can be used for prognostic purposes

in the future.
CNS METASTASES
 Incidence reports of intracranial metastasis vary from 7% to 26%
 The most common primary tumors that metastasize to the CNS are
lung carcinoma, followed by carcinoma of the breast and
melanoma.
 CNS metastases are a major negative prognostic factor in cancer
and a significant cause of morbidity and mortality.
 The brain parenchyma is the most frequent site of intracranial
metastasis. Other localizations within the CNS are the meninges
(leptomeningeal metastases, LM) or and the cranial nerves.
 LM is a serious complication of metastatic tumor associated
with significant mortality.
 The gold standard for diagnosis is CSF cytology (poor
sensitivity)
 The poor prognosis of patients with CNS metastases and the
fast rate of disease progression demands early diagnosis.
 Therefore, sensitive tests are of great importance.
PROTIEN DERIVED FROM CNS
METS
 The most significant example of how analysis of CSF proteins has
impacted the clinical management of CNS cancer is in the case of
intracranial malignant germ cell tumors.
 bHCG and AFP -- markedly elevated in the CSF of intracranial
malignant germ cell tumor patients
 Both markers are currently utilized clinically as diagnostic and accurate
indicators of response to therapy.
 Assessment of AFP and total bHCG in both serum and CSF is
mandatory to distinguish between germinoma and NGGCT (non-
germinoma germ cell tumors).
 CSF AFP > 1000 ng/mL at diagnosis, or age < 6 years,
intracranial malignant germ cell tumor patients are stratified
as high risk and are treated more intensively.
 Also the verification of bHCG and AFP levels prior to surgical
resection provides a reference point that can be used to
assess recurrence during follow-up
 But, their absence does not rule out a germ cell tumor.
 CEA is known serum biomarker for colorectal carcinoma
 It can be seen in CSF of pts with mets from breast,
carcinoma, lung carcinoma and seminoma.
 Increased CSF levels of CEA with CYFRA 21-1 and NSE
are linked to diagnosis of LM by lung carcinoma.
ctDNA DERIVED FROM CNS METS
 EGFR mutations are found in patients with adenocarcinoma of the
lung.
 These mutations can also be found in the ctDNA in the CSF of these
patients, even in patients with negative cytologic results.
Promoter methylation of the human telomerase reverse transcriptase (hTERT) is
found in most cancer cells and can therefore serve as a general marker of cancer.
• Using real-time PCR and real-time methylation- sensitive high-resolution melting (MS-
HRM), CSF samples of 67 patients with leptomeningeal metastasis were analyzed for
hTERT methylation.
• This led to detection levels of hTERT-methylated DNA as low as 1% in a background of
unmethylated DNA.
• The sensitivity of this test was 92% and the specificity 100%, which makes this a
promising tool for the diagnosis of LM (Bougel et al., 2013).
 RNA DERIVED FROM CNS

METS
Melanoma has a strong propensity to metastasize to the brain.
 CSF cytology is used to search for melanoma-derived CNS metastasis. (not
sensitive)
 Hoon et al. (2001) investigated the presence of mRNA of three melanoma-
associated markers in CSF to diagnose CNS metastasis: MAGE-3, MART-
1, and tyrosinase.
 Using traditional methods, only 1/37 patients showed melanoma- positive
cytology and IHC.
 But 30% of patients had positive PCR result for at least one of the RNA
markers.
 The correlation between the detection of melanoma-associated RNA in
the CSF and the development of CNS metastases after 3 months was
also significant.
 Patel et al. (2011) used CellSearch® for detection of breast tumor cells in
CSF of a patient
 CTCs can be isolated using FCM separation based on epithelial cell adhesion
molecule (EpCAM).
 EpCAM is expressed in various cancers of epithelial origin.
 CSF CTCs levels correlate with treatment response and disease burden.

Tu et al., 2015 studied for the presence of

CTCs in the CSF of patients with
confirmed LM from lung cancer, and found
CTCs with a sensitivity of 78% compared
to 44% for conventional cytology.
 Kerklaan et al. (2015) collected samples from patients with a clinical
suspicion of LM but a negative or inconclusive MRI.
 The flow cytometry assay showed 100% sensitivity and 100%
specificity for early diagnosis of LM, while CSF cytology had a
sensitivity of only 62%.
 Subirá et al., 2012 confirmed higher sensitivity and same specificity as
CSF cytology in a similar study on 77 pts.
 The follow up study (2015) suggested a cutoff value of 8% EpCAM-
positive cells in the CSF to predict survival in patients.
CNS INVOLVEMENT IN
HEMATOLOGIC CANCERS
 Involvement of the CNS in hematologic cancers can occur in two ways:
o Metastasis of malignant hematopoietic or lymphoid cells to the CNS.
(LYMPHOMA/LEUKEMIA)
o Primary tumors that arise from lymphoid cells in CNS. (PCNSL-4% of
all intracranial malignancies)
 LM is the most common presentation of CNS metastasis form
hematological malignancies.
 FCM is currently recommended to be used together with MRI and
CSF cytology in order to diagnose LM.
 Acute leukemias account for 30% of pediatric malignancies.
 Improved survival: A 72% decrease in leukemia-related deaths in
children up to the age of 20 between 1970 and 2012.
 The increase in survival is partly due to therapeutic prophylaxis to
prevent CNS metastasis, which was implemented in the 1970s.
 10% of patients show signs of CNS metastasis at the time of diagnosis
 CNS recurrence after successful treatment of systemic disease is one
of the most serious complications with poor prognosis and a median
survival of less than 6 months.
 Hence it is important to diagnose these cases at early stages to
improve survival.
 PCNSL has a poor prognosis compared to its peripheral counterpart,
with a median survival of only 1–2 years.
 More than 90% of PCNSL are extranodal diffuse large B-cell non-
Hodgkin lymphomas.
 The only known risk factor for PCNSL is congenital or acquired
immunodeficiency leading to Epstein–Barr virus (EBV) infection-
associated disease
PROTEINS DERIVED FROM HEMATOLOGIC
CANCERS WITHIN CNS
 Interleukin-10 (IL-10) is a cytokine secreted by many cells of the
immune system including monocytes, T-helper type 2 cells, and B cells.
 IL-10 protects malignant B cells from apoptosis
 It is found to be elevated in the serum of patients with diffuse large B-
cell lymphoma.

In a cohort of patients with PCNSL and a control group of patients with other CNS
tumors, IL-10 levels were significantly higher in the CSF of PCNSL patients.

In follow-up 5/6 patients with high IL-10 levels received the pathologic diagnosis of
PCNSL, and no patients with undetectable IL-10 received the diagnosis of PCNSL.
Sasayama et al. (2012)
 CXCL13 is a cytokine also known as B-lymphocyte chemoattractant.
 It is expressed by malignant B cells and involved in compartmental
homing of B cells.
Levels of CXCL13 were While using CXCL13 alone, >70% of
significantly increased in the CNSL cases were positively identified
CSF of patients with either with a specificity of 94.9%.
PCNSL or secondary CNSL. The sensitivity increased to 84% if
When CXCL13 was there was elevation of both CXCL13
longitudinally measured, and IL-10 in the CSF, while a specificity
levels went down in all above 90% was maintained.
patients responsive to This is significantly higher than
treatment (Fischer et al., standard tests such as cytology and
2009). flow cytometry (Rubenstein et al.,
2013).
RNA DERIVED FROM HEMATOLOGIC
CANCERS WITHIN CNS

 The combination of miR-21 with miR-19 and miR-92a has 95.7%

sensitivity and 96.7% specificity for the diagnosis of PCNSL.
 A follow-up study with more patients supported the biomarker potential
of the miRNA signature.
CTC FROM HEMATOLOGIC
CANCERS WITHIN CNS
• Bromberg et al. (2007)-- study on newly diagnosed patients with
hematologic neoplasms
• FCM was more sensitive than cytology
• But no prognostic value alone.
• Patients with a positive FCM+cytology more often had CNS- related
symptoms and a shorter overall survival compared to patients with
only a positive FCM.
• Few patients had cytologic aberrations and CNS symptoms, but a
negative-flow cytometry.

 These findings conclude that the two methods are complementary and should be
used in conjunction with each other
PEDIATRIC CNS TUMORS
 Tumors of the CNS are the most common type of solid pediatric cancer and the leading cause
of mortality in children between 0 and 14 years of age
 Some tumors arise in the context of genetic syndromes. Eg. Neurofibromatosis 1 and 2 and
von Hippel–Lindau disease.
 However, majority are sporadic.
 The most common CNS tumor in children is the benign pilocytic astrocytoma (grade I), which
accounts for 15.4% of all tumors.
 Embryonic tumors occur relatively frequently in children as well.
 Malignant gliomas account for 11.7% of all primary CNS tumors in children
 The overall survival for low-grade tumors after 20 years is 87%.
 For the treatment of
children with a brain
tumor, placement of an
Ommaya reservoir is
common.
 It can be used to facilitate
the delivery of medication
into the CSF, to remove
CSF for analysis and
cytology, and to alleviate
increased intracranial
pressure.
PROTEINS DERIVED FROM
PEDIATRIC CNS MALIGNANCIES
 IGF- binding protein 3 (IGFBP-3) levels and fragmented IGFBP-3
levels are increased in pediatric medulloblastoma patients, but not in
ependymoma patients or non- cancerous controls.
A proteomics approach identified prostaglandin D2 synthase
(PGD2S) as a potential biomarker for medulloblastoma in pediatric
patients.
 PGD2S is physiologically PDG2S
expressed was atinleast
thetwofold
brainreduced
and isin the
relatively
CSF of
abundant in CSF. pediatric patients with medulloblastoma (Rajagopal
et al., 2011).
 Detection of PGD2S is used in corresponds
This a clinical setting to determine
to an earlier report of whether
a twofold
fluid leaking from ears or nose is CSF.
PDG2S reduction in patients with nonspecified brain
tumors when compared to controls (Saso et al.,
1998).
ctDNA DERIVED FROM
PEDIATRIC CNS MALIGNANCIES
 Wang et al. (2015) examined the CSF of both pediatric and adult
patients with several types of primary CNS tumors.
 CSF was sampled from the ventricles at the time of surgery.
 Whole-exome sequencing on tumor material after resection
revealed patient-specific somatic mutations.
 ctDNA could be detected in the CSF of all patients with
medulloblastoma, ependymoma, or high-grade astrocytoma
whose tumors lay directly adjacent to a CSF space such as the
ventricles.
 ctDNA and patient-specific somatic mutations were detected
in the CSF of 74% of all patients
RNA DERIVED FROM PEDIATRIC
CNS MALIGNANCIES
 Retinoblastoma is principally an ocular disease, but dissemination
to the CNS, mostly by leptomeningeal dissemination, is the most
common cause of mortality.
 mRNA of the ganglioside GD2 and its synthase has been investigated
as a possible bio- marker of CNS spreading of retinoblastoma.
 When CNS spreading is detected, an intensive The chemotherapy
ganglioside GD2 regimen
and its
can be started to best treat this complication.synthase are expressed in
When the CSF of children with disseminated retinoblastoma, and cells positive
retinoblastoma was examined for GD2 for GD2 have been detected in
synthase expression, it could be detected in all the CSF of retinoblastoma
cases. GD2 synthase mRNA was never patients with extraocular disease
detected in the CSF of low-risk patients or (Laurent et al., 2010; Shen et al.,
control subjects (Laurent et al., 2013). 2013).
CIRCULATING TUMOR CELLS DERIVED
FROM PEDIATRIC CNS MALIGNANCIES

 CSF cytology is the main stay in identification of pediatric CNS tumors.

 Disseminated disease is seen in 2-30% of ependymoma.
 Primary neuroectodermal tumor and medulloblastoma also show this
property too.
 Meta-analytic study 2010, showed that positive CSF cytology is an
independent prognostic factor for event free survival in pts with
ependymoma.
CONCLUSION AND SUMMARY
 CSF is an invaluable diagnostic window to the pathological state of
CNS.
 It is easily accessible by minimally-invasive standard clinical methods.
• The reported sample size in the literature is
 Biochemical molecules secreted by brain tumors to the CSF hold great
small.
promise as diagnostic/prognostic
• Mostmarkers.
data were generated by a limited number of
research
 However, despite lots of published groups the
literature, using different protocols
reproducibility betweenor
studies is variable, and none technologies.
of them have been validated for clinical
• No universally implemented guidelines are
use.
available yet for the CSF sample collection and
preparation or for protein profiling or miRNA
extraction from CSF and importantly for data
analysis.
THANK YOU!!