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Path - Intro to molecular Pathology (ii) (Tumour suppressor genes (APC…
Path - Intro to molecular Pathology (ii)
Nucleic acid-based techniques
mutation detection
certain clinically significant mutations won't lead to any IHC-detectable changes
extract DNA/RNA from tissue, identify areas of interest
PCR
traditional (DNA) or real time (mRNA)
DNAP exponentially amplifies DNA with heat, DNA sticks + forms new strands when cooled
sensitive
cheap
full mutation info not always available
single base variant detection
Sequencing
exact mutation identified
traditional
gold-standard
cheap but inefficient for a large no. of targets
lots of experience
has a max read length (500-600, absolute max = 800 bps)
next generation
massive parallel sequencing
multiple overlapping short reads
bioinformatics
a computer makes sense of the raw sequence + assembles DNA into readable genes
a tumour is never fully comprised of tumour cells - some parallel sequences won't have a mutation, but most will
Oncogenes
Ras
Myc
Burkitt's lymphoma
Abl
translocation: BCR-Abl = philadelphia chromo
CML
1 of 1st targets for targeted tx
Ret
thyroid carcinoma
sometimes targeted tx doesn't work
if 1 pathway is blocked some tumour cells can get another pathway to take over
Tumour suppressor genes
most common tumour mutations
accumulate over time
Rb
1st one identified
APC
FAP
AD
1000s of adenomatous neoplastic polyps, some undertow malignant change
prophylactic colectomy indicated
1% of all CR cancers
somatically mutated in most CR cancer cases
P53
NB in preventing cancer
mice studies: both P53 genes knocked out, 100% developed cancer (different types)
molecular policeman (safeguards integrity of genome)
Li-Fraumeni Syndrome
rare
AD
1 mutant P53 in germline + acquired abnormality of 2nd allele
predisposes to sarcoma, leukaemia, breast carcinoma + brain tumours
DNA repair genes
MMR
mutations lead to HNPCC (hereditary non-polyposis CR cancer), new name = Lynch syndrome
microsatellite instability (hypermutability) in 15% of CR cancers
mucinous/poorly differentiated morphology
tumour infiltrating lymphocytes (TILs) + proximal location (right colon)
also endometrial carcinoma + rarely other types
clinical criteria (Hx + exam) insensitive
can be detected with electrophoresis (compare to normal)
screen with IHC, confirm with sequencing
MLH1 + PMS2
epigenetic mutation: promotor methylation
stops expression upstream, hence no protein downstream
hypermethylator phenotype
usually sporadic
MSH2 + MSH6
repair damage from chemicals/UV light/radiation
BRCA