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Circulating tumor cells(CTCs)
Isolation methods and
applications
Sanaz Raeisi M.S of Biotecnology
Supervisor:Dr.Zarei
11 June 2019
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content
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Cancer
the uncontrolled growth and division
of cells without paying attention to
cell's ون
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Cancer in general
population
* The second cause of death after
* More than 70% will di
2020
er until
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Cancer diagnose and
survival!
mEarly(stage1)m Late(stage4)
100%
92% 92%
breast cancer colorectal cancer ovarian cancer
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We are vulnerable without early
warning system
* No good screening method for some
cancers such as ovarian , liver or
pancreatic .
* Our weapon against cancer should
ARE YOU
PREPARED
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“Seed in the Soil” Hypothesis
Steven Paget - 1889
Tumor
تت ۱ 1
لامع مع ايلا 00060
Cancer cells are the seeds and
the microenvironment is the soil.
“When a plant goes to seed, its seeds are carried in all
(oligo را( فا فا
".ازه؟ اهوأمععمم
— Stephen Paget, 1889
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CTC : a circulating tumor cell that
has enter into the bloodstream or
lymp* ~*~
Cancer foci
Circulating Tumor Cell
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10
Primary site of tumor formation
Blood vessel and Lymphatic
Cell survives and
becomes a ctc
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EMT in cancer cells
۰ The epithelial mesenchymal
transition (EMT) is a necessary step
for the detachment and migration of
cancer cells.
* In this way migratory ability ,
invasiveness an nce of
Nema Primary phat موه
Memtrane Piha cal caer cts
Seen ptr
3 گر
6۳-۰ 0 ae
oma Epitheism Pat من corm اث wer
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Advantages forward traditional
method
1.tissue biopsy:
It can be painful and invasive , not
easily obtain
2.Lequid biopsy:
مهللا أطال35أ١
painful , ea:
12
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Disadvantages of ctcs
1. Short presence time in circulation(1-
2.5h)
2. Heterogeneity nature of these cells.
3. Low amoun hem.
.
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Isolation methods
1. Biological Properties
a.
6 2 © Erythrocytes
© Leukocytes
Immunoafinity 6 وعم
2. Physical Properties
GR sizetdotormavitty | ts
electrical properties,
Whole blood
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Density-based isolation
technologies based-density :
OncoQuick and
pot
Centrifugation
Diluted blood 4- — ~~ Blood plasma
—» Ficoll solution
Ficoll solution ع4 wv Red blood cells
PBMC + CTCs
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Size-Based isolation
technologies that use size-based:
Parsortix™, ScreenCell® and ISET®.
Current
Techniques
poresize
‘Sie es
هعلوم
slitsize
(6
0 3 6 9 12 15 18 21 4
Cell Diameter (um)
16
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Isolation based on biological
properties
* CTCs can be positively or negatively
enriched based on the expression of
surface proteins.
* Separation of CTCs based on surface
antigens of two types of technology:
Immunomagnetic beads and
Microfluidic device.
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Immunomagnetic
separation
* This approach utilizes capture agent-
labeled magnetic beads for positive
selection of CTCs using cell-surface
markers(EpCAM).
* Epithelial cell adhesion molecule
over € x
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derive
دعا مك
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microfluidic
* Positive selection of ctc using
antibodies attached to microfluidic
عم
Non-cancerous
Blood inlet
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* Technologies based on positive
selection:
AdnatTest , MACS®, IsoFlux™ , and
the CellSearch® and
MagSwepeer system.
* Antibody based negative selection:
Deple © d cells using
1 1 سم للح جح حم جلا
anti_c 5 5 etic
@ crc
af * ox + + که White blood cell
Magnetic depletion *G immunomagnetic beac
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Cell search
MagNest™
fcelsaye 4eelltracks
3
|
Cell Search
Epithelial Cell Kit
۳ ن Enrichment of CTC with
anti-EpCAM ferro fluids
Captures tumour cells with very
low EpCAM expression
CeliTracks® Analyzer It
\w/ Linux operating system
21
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Images of tumor cell
CK-PE ۱ DAPI اْ Tumour cell
neg
pos pos :
0 ۱ ۳13 ا ْ i
Leukocyte
Leukocyte nucleus 0045 6 \
Tumour cell
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maintrac
* Quantitative determination of vital
cells.
٠ work processes:
RBC lysis , sentrifugation , labled
anti-EpCAM ,
adding propidium iodide, analysis
a1 eee
i ۳۱ 1 ۱ ۱ ۳
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Society of ctcs
Circulating tumor cell can exist as a
single cellular as hetero cellular
cluster of cells that travel through
Primary tumour
Plakoglobin-expressing
ell
tumour c
Metastasis
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Immune cells collaborate with cancer cel
(a key to identifying new cancer therapie
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Ctc associated with platelets
* Ctc interact with platelet during
intravasation and trigger a factor
called TCIPA on platelet.
* TCIPA activates platelets allowing
them to attach to ctc and shield them
from immune attack.
* Platelets can also transfer
MHC ,further which allows evasion of
immune destruction.
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Ctcs and neutrophils
the neutrophils enhance the
metastasis-seeding ability of CTCs by
releasing specific messenger
substances, such as cytokines.
the presence of CTC-neutrophil
clusters in the bloodstream also
correlates with a poor prognosis of
breast cancer patients.
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Clinical application of CTCs
* CTCs as prognostic markers in
cancer(CTCs enumeration)
25
<5ctcs/7ml blood >5ctcs/7ml blood 28
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* Screening and early detection
Cancer screening aims to detect
cancer before symptoms appear.
blood tests, urine tests, DNA tests or
medical imaging can be done.
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۰ A tool to monitor treatment
effectiveness
Do the metastase relapse?
Does the tumor answer to our
therapy or become resistant?
Do our drugs work?
Answer to all these questions come
back to their numbers
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, ۵
hel
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references
https://www.maintrac.de/En/maintrac
4
https://www.ncbi.nim.nih.gov/pmc/art
icles/PMC6023422/
https://www.labce.com/spg1629022 t
he_advantages_and_limitations_of_lig
uid_biops
* tumj.tums.ac.ir/article-1-51-fa.pdf
aums.abzums.ac. it/article-1-410-fa.p
https://www.sciencedaily.com/release,
¢/2019/07/190706131933 htm