Cancer and Molecular Methods

Question 1

List the five most common cancers in Ireland, in descending order.

Answer 1

Non-melanoma
Prostate
Breast
Bowel
Lung

Question 2

Most common cancers in males are non-melanoma, prostate, bowel, _______, melanoma, and testicular cancer.

Answer 2

Lung

Question 3

Most common female cancers are non-melanoma, breast, ______, lung, melanoma, and gynae cancers.

Answer 3

Bowel

Question 4

____________ cells divide repeatedly, out of control, and therefore crowd out normal cells, and they function abnormally.

Answer 4

Cancerous

Question 5

List four known carcinogens.

Answer 5

Ionising radiation, such as x-rays and UV light
Chemicals, such as cigarette tar
Viral infections, such as HPV
Hereditary dispositions (cancer cannot be inherited, but families may have predispositions)

Question 6

What two classes of genes lead to cancer when mutations occur in them?

Answer 6

Oncogenes and tumour-suppressor genes

Question 7

True or false: cells that are old or malfunctioning normally self-destruct, and are replaced by new cells.

Answer 7

True

Question 8

DNA damage, in the form of mutations, is caused by __________.

Answer 8

Mutagens

Question 9

Describe carcinomas.

Answer 9

The begin in the skin or tissue, and usually form solid tumours. They are the most common form of cancer. Examples include prostate, lung, colon, etc.

Question 10

Where do sarcoma cancers begin?

Answer 10

Tissues that support and connect the body

Question 11

True or false: sarcomas cannot develop in fat, muscle, nerve, tendons, etc..

Answer 11

False

Question 12

_______________ occurs when healthy blood cells change and grow uncontrollably.

Answer 12

Leukaemia

Question 13

_______ tumours do not spread from their site of origin, but can crowd out (squash) surrounding cells, e.g., brain tumour, warts, etc..

Answer 13

Benign

Question 14

Malignant tumours can spread from the original site and cause secondary tumours, in a phenomenon known as ___________.

Answer 14

Metastasis

Question 15

A cancer diagnosis pathway in pathology involves macroscopy, microscopy (routine H&E staining, and ______________) as well as molecular testing.

Answer 15

Immunohistochemistry

Question 16

________ ______ compares the size, shape, and other characteristics to those of healthy cells.

Answer 16

Histologic grade

Question 17

A tumour with cells that look more like healthy cells is called _____ _________ or well differentiated, and is often associated with a better prognosis, or chance of recovery.

Answer 17

Low grade

Question 18

_______ rate details how often the cancer cells are dividing.

Answer 18

Mitotic

Question 19

If the tumour has invaded the blood vessels or lymph vessels that flow into the ________ ______, there is a greater chance that the cancer has metastasized (spread) to other parts of the body.

Answer 19

Lymph nodes

Question 20

‘Targeted’ therapies (precision medicine) are used where _______ __________ ________ identify subgroups of tumours, which may or may not respond to novel drug treatments.

Answer 20

Specific molecular markers

Question 21

List two methods used in molecular techniques.

Answer 21

DNA/RNA and protein methods

Question 22

All molecular techniques must fulfil stringent accreditation requirements, and must be ___________.

Answer 22

Affordable

Question 23

_____________ of the pre-analytical workflow is important, to support downstream analysis.

Answer 23

Optimisation

Question 24

List three other considerations of molecular techniques.

Answer 24

Minimisation of wastage, to maximise utility of tissues and specimens
Training
Recording and reporting of results

Question 25

Outline the workflow steps of molecular techniques.

Answer 25

Preparation and assessment
Extraction/isolation of target molecules (in PCR, this involves the extraction of nucleic acids (DNA) to provide a template)
Amplification and measurement
Real-time sequencing can be used
Analysis and interpretation
Reporting of findings
Should be done within the assay context

Question 26

Describe in-situ hybridisation.

Answer 26

Application of a labelled probe (oligonucleotide) to detect target sequences of interest

Probe is specific to a chromosome, or to a gene mutation

Label is usually fluorescent, but may be biotin, or alkaline phosphatase

ISH methods are ideal for the detection of a low copy number

Indirect method employs a secondary layer in the form of a chromagen

Question 27

Describe fluorescent ISH (FISH)

Answer 27

Basic elements include a DNA probe and a target sequence

Before hybridisation, the DNA probe is labelled with a fluorophore by various means, such as nick translation, random primed labelling, and PCR

The labelling probe and target DNA (on a slide) are denatures

The denatured probe and the target are combined, and this allows the annealing of complementary DNA sequences

This is known as hybridisation

A fluorescent signal is emitted

Question 28

PCR is based on the amplification of specific sequences, as defined by __________.

Answer 28

Primers

Question 29

True or false: in PCR, paraffin wax-embedded tissues’ DNA/RNA can be harder to extract, or fragmented.

Answer 29

True

Question 30

What type of sample is best for PCR?

Answer 30

Fresh, frozen tissue

Question 31

List three points regarding PCR extraction.

Answer 31

Usually involves the proteolytic digestion of tissue to release DNA/RNA
Spin columns, such as AllPrep columns, can be used
Resulting DNA is of high quality

Question 32

Write a note on Sanger sequencing.

Answer 32

Mainstay in many laboratories for 30+ years. Amplified PCR products are subjected to a second round of PCR, using a set of fluorescently-tagged ddNTP, which terminate chain extension when incorporated.
Labelled product is run through a capillary, and subjected to electrophoresis, based on size

Question 33

What are two advantages of real-time PCR (qPCR)?

Answer 33

Allows for quantification of PCR products, using fluorescent probes

High degree of sensitivity

Question 34

List three uses of PCR?

Answer 34

Detection of EGFR1 gene in non-small cell lung cancer

Detects mutations in the BRAF gene in melanoma

KRAS and NRAS gene mutations in colorectal cancer can be detected via PCR; testing for these mutations is mandatory

Question 35

Describe array analysis.

Answer 35

Allows for multiple types of DNA or RNA to be analysed simultaneously

DNA or mRNA is extracted, and applied to an array, where it hybridises

Computerised reading allows for results to be generated