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Detection of a rare BCR-ABL tyrosine kinase fusion protein in H929 multiple myeloma cells using immunoprecipitation (IP)-tandem mass spectrometry (MS/MS).

Abstract：

:Hypothesis directed proteomics offers higher throughput over global analyses. We show that immunoprecipitation (IP)-tandem mass spectrometry (LC-MS/MS) in H929 multiple myeloma (MM) cancer cells led to the discovery of a rare and unexpected BCR-ABL fusion, informing a therapeutic intervention using imatinib (Gleevec). BCR-ABL is the driving mutation in chronic myeloid leukemia (CML) and is uncommon to other cancers. Three different IP-MS experiments central to cell signaling pathways were sufficient to discover a BCR-ABL fusion in H929 cells: phosphotyrosine (pY) peptide IP, p85 regulatory subunit of phosphoinositide-3-kinase (PI3K) IP, and the GRB2 adaptor IP. The pY peptides inform tyrosine kinase activity, p85 IP informs the activating adaptors and receptor tyrosine kinases (RTKs) involved in AKT activation and GRB2 IP identifies RTKs and adaptors leading to ERK activation. Integration of the bait-prey data from the three separate experiments identified the BCR-ABL protein complex, which was confirmed by biochemistry, cytogenetic methods, and DNA sequencing revealed the e14a2 fusion transcript. The tyrosine phosphatase SHP2 and the GAB2 adaptor protein, important for MAPK signaling, were common to all three IP-MS experiments. The comparative treatment of tyrosine kinase inhibitor (TKI) drugs revealed only imatinib, the standard of care in CML, was inhibitory to BCR-ABL leading to down-regulation of pERK and pS6K and inhibiting cell proliferation. These data suggest a model for directed proteomics from patient tumor samples for selecting the appropriate TKI drug(s) based on IP and LC-MS/MS. The data also suggest that MM patients, in addition to CML patients, may benefit from BCR-ABL diagnostic screening.

journal_name

Proc Natl Acad Sci U S A

journal_title

Proceedings of the National Academy of Sciences of the United States of America

authors

Breitkopf SB,Yuan M,Pihan GA,Asara JM

doi

10.1073/pnas.1212759109

subject

Has Abstract

pub_date

2012-10-02 00:00:00

pages

16190-5

issue

eissn

0027-8424

issn

1091-6490

pii

1212759109

journal_volume

109

pub_type

杂志文章

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Detection of a rare BCR-ABL tyrosine kinase fusion protein in H929 multiple myeloma cells using immunoprecipitation (IP)-tandem mass spectrometry (MS/MS).