Cancer and Molecular Methods Flashcards
List the five most common cancers in Ireland, in descending order.
Non-melanoma
Prostate
Breast
Bowel
Lung
Most common cancers in males are non-melanoma, prostate, bowel, _______, melanoma, and testicular cancer.
Lung
Most common female cancers are non-melanoma, breast, ______, lung, melanoma, and gynae cancers.
Bowel
____________ cells divide repeatedly, out of control, and therefore crowd out normal cells, and they function abnormally.
Cancerous
List four known carcinogens.
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)
What two classes of genes lead to cancer when mutations occur in them?
Oncogenes and tumour-suppressor genes
True or false: cells that are old or malfunctioning normally self-destruct, and are replaced by new cells.
True
DNA damage, in the form of mutations, is caused by __________.
Mutagens
Describe carcinomas.
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.
Where do sarcoma cancers begin?
Tissues that support and connect the body
True or false: sarcomas cannot develop in fat, muscle, nerve, tendons, etc..
False
_______________ occurs when healthy blood cells change and grow uncontrollably.
Leukaemia
_______ tumours do not spread from their site of origin, but can crowd out (squash) surrounding cells, e.g., brain tumour, warts, etc..
Benign
Malignant tumours can spread from the original site and cause secondary tumours, in a phenomenon known as ___________.
Metastasis
A cancer diagnosis pathway in pathology involves macroscopy, microscopy (routine H&E staining, and ______________) as well as molecular testing.
Immunohistochemistry
________ ______ compares the size, shape, and other characteristics to those of healthy cells.
Histologic grade
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.
Low grade
_______ rate details how often the cancer cells are dividing.
Mitotic
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.
Lymph nodes
‘Targeted’ therapies (precision medicine) are used where _______ __________ ________ identify subgroups of tumours, which may or may not respond to novel drug treatments.
Specific molecular markers
List two methods used in molecular techniques.
DNA/RNA and protein methods
All molecular techniques must fulfil stringent accreditation requirements, and must be ___________.
Affordable
_____________ of the pre-analytical workflow is important, to support downstream analysis.
Optimisation
List three other considerations of molecular techniques.
Minimisation of wastage, to maximise utility of tissues and specimens
Training
Recording and reporting of results
Outline the workflow steps of molecular techniques.
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
Describe in-situ hybridisation.
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
Describe fluorescent ISH (FISH)
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
PCR is based on the amplification of specific sequences, as defined by __________.
Primers
True or false: in PCR, paraffin wax-embedded tissues’ DNA/RNA can be harder to extract, or fragmented.
True
What type of sample is best for PCR?
Fresh, frozen tissue
List three points regarding PCR extraction.
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
Write a note on Sanger sequencing.
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
What are two advantages of real-time PCR (qPCR)?
Allows for quantification of PCR products, using fluorescent probes
High degree of sensitivity
List three uses of PCR?
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
Describe array analysis.
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
