Neoplasia molecular pathology: Chromosomal change
14 important questions on Neoplasia molecular pathology: Chromosomal change
What do we use for numerical alterations and copy number ....?
Copy number -> MPLA
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Sequence is done on RNA not DNA, Why?
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Oncogenic mutations can be induced by:
translocation
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To which family of membrane receptors does the ErbB2 receptor belong?
ERBB2 = HER2
On which biomarker does Transtuzumab work, and which pathway is inhibited?
Where does FISH stand for, and what is it?
- Fluorescent In Situ Hybridization
- use of: tissue slides, well defined probes, a hybridisation protocol and a visualisation tool such as fluorescence microscopy
- focus on a specific gene
- mainly used for detection of amplifications, numerical alterations
What is HER2 amplification?
What is in the pathology FFPE tissue?
What does MLPA stand for, and what does it mean?
- Multiplex Ligation Dependent Probe Amplification
- Amplification of MLPA probes, not of sample DNA
- Probes are composed of synthetic oligonucleotides complementary to the target sequence, primers sequence X and Y that are needed for PCR after probe hybridization and a stuffer sequence which differs in length between the different probes in the assay.
- up to 50 probes are present in one DNA sample
- Detection of chromosomal deletions (losses)
- Multiplex ligation dependent probe amplification
- focus on multiple genes
How do MLPA result look like?
Translocation, gene fusion > Biological effect
- By the fusion; the coding region of gene X fuses to the coding region of gene Y
- Generation of a chimeric fusion gene
- chimaera transcript
- chimaera protein
- Oncogenic properties, by merge of protein domains that originate from both fusion partners
- examples: gene fusions/translocations that are associated with sarcoma or with lung cancer.
Are break points in fusion genes mostly in intronic or exotic?
- Breakpoints in fusion genes are mostly intronic
- different breakpoints in an intron and / or breakpoints in different introns
- RT-PCR:
- chromosomal translocations (in sarcoma) result in a fusion transcript
- analysis of mRNA > cDNA (and need for 1 forward primer to detect multiple breakpoints.
What should be kept in mind by using DNA/RNA from tissue?
- Sufficient tissue, DNA/RNA
- DNA/RNA quality (from FFPE) sufficient for the analysis?
- from tissue that is representative
To what can chromosomal translocations lead ?
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