Liquid biopsy - Recent advance in pathology

2. LIQUID BIOPSY
DR. SEENA SUSAN ITTY
JR 2,PATHOLOGY

3. CONTENTS
• Introduction
• Definition
• Indication
• What comes under liquid biopsy?
• Advantages
• Limitations
• Summary
• References

4. INTRODUCTION
• Cancer is a heterogenous disease.
Molecular properties differ within a tumour
Primary tumour biopsy may not reflect current
disease condition.
Therapy causes changes in tumour cells.
• Tumour releases a multitude of biomarkers into
blood.
• Liquid biopsy is a minimally invasive alternative
to surgical biopsy for tumour molecular profiling.

5. DEFINITION
• Liquid biopsy (blood based biomarkers) are
non invasive blood tests that detect:-
Circulating tumour cells(CTCs)
Fragments of tumour DNA(ctCNA)/cell free
DNA(cfDNA)
Exosomes and microvesicles
that are shed into blood from primary tumour
and from metastatic sites.
• Tumour educated platelets(TEPs)

7. CLINICAL APPLICATIONS

9. NEW ADDITION:-TUMOUR EDUCATED PLATELETS (TEP)

11. • 1869 – T.R.Ashworth first described the
presence of epithelial cells in the blood of a
woman with metastatic breast cancer that were
similar in appearance to her primary tumour
cells.
• 2002 and 2003 – Thiery and Steeg : Found the
formation mechanism of CTC.
• 2007 – CTC has been recommended as new
tumour biomarker by ASCO

12. CTCs
• Represent the clones within the tumour that
has the metastatic potential.
• CTCs represent a rare cell population - less
than 10 cells/mL.
• Number of cells reflect the stage of the tumour
• Sample collection – Cell save preservative
tube

16. • Immunost
ain
• FISH
• Sequencin
g
• qRT-PCR
• Expression
analysis
• Cell
culture

18. CTCs
ADVANTAGES
• Minimally invasive
prognostic marker
• Both phenotypic and
genotypic analysis
• Potential relation with
tumour progression and
metastasis
• High specificity
• Availability of FDA approved
method for isolation
DISADVANTAGES
• Low sensitivity in low
volume diseases
• Low concentration
• Fragility of cells
• Presence of benign
circulating epithelial cells
• Cannot decrease the
diagnosis bias from tumour
heterogeneity

19. Complex heterogeneity
• Morphology of CTCs derived from different tumour
tissues through EMT is sigificantly different.
• Even within only one cancer,the morphology and
amount of CTCs derived from different molecular
subtypes of solid tumour or distant sites are distinct.
• The percentage of patients whose CTCs can be detected
is also different between cancers.
Eg: Colorectal,ovarian and breast cancer : 50-70%
Non small cell lung cancer is only 30%

24. ctDNA
ADVANTAGES
• High sensitivity
• Allow analysis of DNA
sequence and
methylation,including PCR
and NGS
• Can decrease the diagnosis
bias from tumour
heterogeneity
• Timely and dynamically
monitortumour progression
LIMITATIONS
• ctDNA is unstable , so
requires fast processing
• Low specificity because of
cfDNA from normal tissue
• False positive and false
negative results

26. • Role in tumour initiation,progression,metastasis
and drug resistance
• Based on these results,researchers have
developed some exosomes targeted drugs and
tried to use exosomes for drug delivery.
• Enrichment methods: Similar to those of CTC
• Analysis – Electron microscope,RT-PCR,Western
blot,FISH,Flow cytometry and NGS.

27. EXOSOMES
ADVANTAGES
• Allow analysis of
DNA,RNA and proteins
from solid tumour
• Potential relation with the
tumour drug-resistance and
metastasis
LIMITATIONS
• Lacking effective
enrichment method

28. TUMOUR EDUCATED PLATELETS
• Besides tumour cells and their products,normal
cells present in the tumour microenvironment
are also released into the blood stream.
• They harbour important information about the
tumour.
• Platelets have been studied extensively and
gave promising results.

29. • Interaction between blood platelets and tumour
cells:
Not only affects the expression of relevant
genes in the tumour cells,but also alters the
RNA profile of blood platelets.
• mRNA sequencing of TEPs can be done and
can be used to distinguish from platelets of
healthy individuals.

31. ADVANTAGES
• Non invasively take repeated tumour samples.
• Fewer side effects
• Rapid testing speed
• Decreasing the diagnosis bias from tumour
heterogeneity.
• Dynamically reflect the tumour progression.

32. LIMITATIONS
• Lack of standardisation of techniques
• Sensitivity and specificity and current detection
technolgies need to be improved
• Low concentration of CTC,ctDNA and exosomes
• However cannot be used in general:
High cost
Requirement for high quality DNA(not
characteristic of cfDNA)
Extensive data analysis requiring a dedicated
bioinformation

34. IS THIS THE END OF TISSUE BIOPSIES?
• Tissue biopsy will remain as the gold standard.
• Liquid biopsies are recommended when tissue
biopsy is difficult,or when primary site of
disease unknown
• Help patients get the right treatment.

35. REFERENCES
• Liquid biospsy in cancer patients – The hand lens
for tumour evaluation Antonio Russo,Antonio
Giordano,Christian Rolfo
• Current and future applications of liquid biopsy in
non small cell lung cancer from early to advanced
stages Nicolas Guibert,Anne Pradines et al
European respiratory Review 2020 29
• Liquid biopsy of cancer:a multiodal diagnostic
toolin clinical oncology Raffaele palmirotta et al
Ther Adv Med Oncol.2018;10