LB Flashcards

1
Q

What is a liquid biopsy?

A

→ Sampling and analysis of non-solid biological tissue, primarily blood

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2
Q

What other liquids are there for liquid biopsies?

A

→ urine,
→ plasma,
→ serum,
→ saliva

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3
Q

What are saliva biopsies useful for?

A

→ head and neck cancers

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4
Q

What is CSF biopsies used for?

A

→ circulating tumour DNA in brain tumours

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5
Q

What is an example of an established liquid biopsy?

A

→ amniotic fluid

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6
Q

What has amniotic fluid analysis been replaced with and why?

A

→ substituted with circulating foetal DNA in mothers blood

→ amniotic analysis is invasive

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7
Q

What are some markers for liquid biopsies?

A
→ cell free nucleotides
→ tumour educated platelets
→ circulating tumour cells
→ disseminated tumour cells
→ metabolites
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8
Q

Which cells are extracted for liquid biopsies?

A

Normally interested in somatic information because we can find germline information from any part of the body
→ germ line may not have tumour cells

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9
Q

Why are EDTAs used for venepuncture?

A

→ Preventing: blood clots
→ genomic DNA release (from white blood cells)
→ haemolysis

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10
Q

What does bursting of white blood cells release?

A

→ genomic DNA

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11
Q

What are the tubes used for liquid biopsises?

A

→ EDTA
→ citrate
→ cell free DNA

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12
Q

What are the properties that make EDTAs and citrates useful for LB?

A

→ contain anticoagulant to prevent clotting

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13
Q

What are the logistical and storage issues for EDTAs?

A

→ On-site centrifugation within 6hrs of collection to isolate plasma and avoid white cells apoptosis.
→ If not possible, sample can be stored at 4ºC for a up to a week

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14
Q

What are the properties of cell-free DNA tubes that makes them suitable for LB?

A

→ Contain a stabiliser to prevent release of gDNA from white blood and haemolysis of red blood cells

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15
Q

How can cell-free DNA tubes be stored?

A

→ Samples can be stored for 6-14 days at 6ºC-37ºC

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16
Q

What makers can be found in the plasma?

A

→ cfDNA
→ exosomes
→ hormones

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17
Q

What markers are found in the buffy coat layer?

A

→ WBC and CTC

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18
Q

What is found in the haemotocrit?

A

→ RBCs

19
Q

What are the actual LB biomarkers?

A

→ Circulating Tumour Cells (CTC)

→ Circulating Tumour DNA (ctDNA)

20
Q

What are CTCs?

A

→ Cells that have detached from a tumour and travel through the bloodstream to other parts of the body- single cells or clusters

21
Q

What is CTC a maker for?

A

→ tumour growth

→ negative cancer prognosis

22
Q

What may it mean if CTCs are found when on treatment?

A

→ treatment may not be working

23
Q

Why are specific methods needed to study CTCs?

A

→ Found in a high background of normal cells

24
Q

How are CTCs isolated and characterised?

A

→ based on transcripts- PCR done on total RNA extracted from the cells.
→ use cell curface markers eg FACS or magnetic beads

25
Q

What markers are found on CTC surfaces?

A

→ CD45 negative

→ EpCAM positive

26
Q

How are CTCs differentiated?

A

→ cell surface markers

→ OR use their physical properties, size, charge, density

27
Q

What is involved in CTC analysis?

A

→ study the omes with NGS, RTqPCR, flow cytometrry, FISH

28
Q

What is ctDNA?

A

→ DNA that comes from cancerous cells and tumours

29
Q

Where is ctDNA found?

A

→ plasma, serum, urine

→ Low concentration

30
Q

What does the amount of ctDNA depend on?

A

→ health status in the same person (increase in cancer, trauma)

31
Q

What is the size range of ctDNA?

A

→ Highly fragmented but with specific size range (<500bp)

32
Q

Why is the first stage of NGS not needed for ctDNA?

A

→ highly fragmented

33
Q

Where is the ctDNA supernatant transferred to?

A

→ clean polypropylene

34
Q

What are the three ways ctDNA is isolated?

A

→ magnetic beads
→ cellulose-based
→ sillica based

35
Q

What does use of NGS, and array CGH for ctDNA show?

A

→ Amplifications and deletions,
→ Translocations,
→ Point mutations,
→ Chromosomes abnormalities, epigenetic status (methylation)

36
Q

Why is ctDNA sequencing limited?

A

→ not working with cells so can only interrogate genome and epigenome

37
Q

What is ctDNA qPCR used for?

A

→ presence quantification

38
Q

What are the advantages of liquid biopsies over solid biopsies?

A

→ Lower invasiveness even for tissues of limited access
→ Higher patient compliance
→ Higher cost/effectiveness
→ Allow repeated access and multiple sampling
→ No special training required for extraction

39
Q

What are the advantages of disadvantages of liquid biopsies?

A

→ Low amount of material- need sensitive systems
→ Early diagnosis
→ Data interpretation

40
Q

Why use liquid biopsies?

A

→ cancer is is a heterogenous disease
→ properties within a tumour differ and also between metastatic sites.
→ Primary tumour information may not reflect the current disease condition
→ No need to identify the tumour site before taking a biopsy and allow repeating sampling
→ Allow analysis tissues difficult to access

41
Q

What is one way lung cancer be detected?

A

→ Detection of EGFR mutations

42
Q

What is LIQUID CDx?

A

→ Pan-tumour liquid biopsy test for patients with advanced solid cancer

43
Q

Why are exosomes cancer biomakers?

A

→ participate in cancer progression and metastasis by transferring bioactive molecules between cancer and various cells

44
Q

Why are miRNA markers for cancer?

A

→ plays a role is proliferative signalling

→ resisting cell death