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Thursday, February 5, 2009

Christopher Maher, University of Michigan - “Integrative Transcriptome Sequencing to Discover Gene Fusion in Cancer”

80% of all known gene fusions are associated with 10% of human cancers. Epithelial cancers account for 80% of cancer deaths, but have only 10% of known fusions.

Mined publicly available datasets and looked for genes with outlier expression.

Will use next-gen sequencing to get direct sequence evidence of chimeric events. Decided to use both 454 and Illumina. Categorized: mapping reads, partially aligned reads, non-mapping reads. Used same samples [whoa.. classification just got extensive.... moving on.]

Chimera discovery using long read technology. Sequenced: VcaP, LNCaP, RWPE. Found 428, 247, 83 chimeras respectively.

Then added illumina. First checked that they could find the fusion that they know. 21 reads mapped there.

Found both intra- and inter-chromosomal candidates, and then validated 73% of them.

So, to recap: candidates found by both 454 and illumina were MUCH more selective and found they were throwing out false positives, but keeping all the known targets.

Confirmed results with FISH.

Next expt: Identification of novel chimeras in prostate tumour samples. Found candidate sequences from non-mapping read, then worked to validate. How does it work, and what's it's frequency? Found it in 7 metastatic prostate tissues and is androgen inducible. In a meta study, found the fusion of interest in about 50% of prostate cancers.

Came up with a chimeric classification system: 5: inter chromosomal translocation, inter chromsomeal complex rearrangements, intra chromsomal complex deletions, intra chromosomal complex rearrangements.

Summary: validated 14 novel chimeras
Demonstrated cell line can harbor non-specific fusions...
[too slow to catch last point]

Answer to question: 100bp reads would have been long enough to nominate fusions.



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