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Heinrich & Corless Laboratories
GIST Research Updates: May 2012
GIST Research Updates: May 2012
• Corless
– Will discuss new technologies for DNA sequencing
– The goal is to further subclassify GISTs and better
understand the genes that contribute to
malignancy
• Heinrich
– Will discuss studies aimed at understanding how
to kill persistent GIST cells that imatinib and
sunitinib cannot always control
‘Next Generation’ DNA Sequencing
• Traditional DNA sequencing is limited to
just one part of one gene at a time
• Second- and third-generation DNA
sequencers are now available at OHSU
• We have been using these new
technologies to study GISTs
Second-Generation DNA Sequencing
Sequencing GIST ‘Exomes’
• Human genome consists of 3.2 billion
letters (A, C, T, G)
• Genes comprise only ~2% of our DNA
(about 64 million letters)
• We ‘pre-select’ the genes, so that only
the 64 million letters of interest get
sequenced – this is the ‘exome’
• Cost: $2,000 per sample
GIST Exome Sequencing
Specimen Summary
Genotype
Sequencing completed
Sequencing in progress
Patients
Specimens
Patients
Specimens
KIT exon 9
5
8
1
1
KIT exon 11
9
13
PDGFRA
3
3
Wildtype
1
1
BRAF
1
1
18
25
5
Pts
Specimens
4
6
3
3
1
1
8
10
1
3
unknown
TOTAL
Available
11
Additional matched normal specimens: 19 completed; 5 in progress; 8 available
Sequencing results: Mutant genes by group
Apoptosis,
autophagy , 6
Cell adhesion, 5
Cell cycle, 1
DNA damage
response, 4
Epigenetic regulation, 8
•25 specimens
from 18 patients
•285 non-synonymous
mutations
•276 unique genes
GTPase regulation
(Ras family) , 6
Golgi, intracellular
transport , 7
Kinase, adaptor,
RTK regulation , 15
Ribosome
biogenesis, RNA
Polymerase, RNA
splicing, 13
Transcription
factor, 15
unknown, analysis
in progress, 173
Tumor suppressor,
TP53-related , 6
Ubiquitin , 9
Misc potentially
interesting , 8
GIST Analysis
Using Third-Generation DNA Sequencing
• The Knight Labs have
partnered with Ion Torrent/Life
Tech on the development of
genotyping panels using thirdgeneration DNA sequencing
technology
• Sequencing is performed on a
semiconductor chip
Sequencing Directly On A Chip
Traditional Sequence
3rd Generation Sequence
ACTGGTCCTGCTGGTTAG
ACTGGTCCTGCTGGTTAG
ACTGGTCCTGCTGGTTAG
ACTGGTCCTGCTGGTTAG
ACTGGTCCTGCTGGTTAG
ACTGGTCCAGCTGGTTAG
ACTGGTCCAGCTGGTTAG
ACTGGTCCAGCTGGTTAG
ACTGGTCCAGCTGGTTAG
ACTGGTCCAGCTGGTTAG
Knight Diagnostic Labs
GIST Panel
AKT1
AKT2
AKT3
ATM
BRAF
CDKN2A
HRAS
KIT
KRAS
MAP2K1
NF1
NRAS
PDGFRA
PIK3CA
PTEN
PTPN11
SDHA
SDHAF1
SDHAF2
SDHB
SDHC
SDHD
TP53
Common GIST mutations
Important in WT GISTs
Predicting GIST prognosis
Rare GIST mutations of research
interest