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J Proteomics. 2008 Apr 30;71(1):61-70. doi: 10.1016/j.jprot.2007.11.009. Epub 2007 Dec 5.

Mass spectrometric quantification of asparagine synthetase in circulating leukemia cells from acute lymphoblastic leukemia patients.

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1
Department of Pharmacology, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA.

Abstract

The appearance of asparaginase-resistant acute lymphoblastic leukemia (ALL) in transformed cell lines has been correlated with increased expression of asparagine synthetase (ASNS). Recent measurements using mRNA-based assays have raised doubts, however, as to the importance of ASNS protein in the cellular mechanisms that confer drug resistance upon the leukemic cells. Studies aimed at determining the concentration of ASNS protein in human leukemias are therefore needed to resolve this issue. A mass spectrometry (MS)-based procedure is presented for the direct quantification of ASNS protein concentration in complex sample mixtures. This assay is able to distinguish samples from transformed cell lines that express ASNS over a wide dynamic range of concentration. Importantly, this method directly detects ASNS protein, the functional entity that may be synthesizing sufficient asparagine to render leukemia cells resistant to asparaginase-treatment. We also report the successful use of this MS method, which has lower limits of detection and quantification of 30 and 100 attomoles, respectively, for the first direct measurements of ASNS protein concentrations in four patient blast samples.

PMID:
18541474
DOI:
10.1016/j.jprot.2007.11.009
[Indexed for MEDLINE]
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