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Second Half of San Francisco Mass Spec Conference Focuses on Assays & Biomarkers

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This is the second installment of a two-part conference report on the Sixth International Symposium on Mass Spectrometry in the Life Sciences (see PM, 9-5-03 for Part I)

The last two days of the symposium in San Francisco focused on new protein separation and analysis methods, as well as the buzz about biomarkers.

Wednesday, Aug. 27

Michel Desjardins, associate professor of pathology and cell biology at the University of Mont-real, discussed his use of 1D gel organelle-based separation and nano ESI MS/MS to look at 600 proteins found in the phagosome membrane at different stages of phagosome formation and function. Desjardins encouraged others to also run organelles rather than whole cell lysates through mass specs, calling cell fractionation “the PCR of proteins.” Desjardins’ results from earlier studies revealed that membranes of the ER fused with the plasma membrane to create phagosomes. Updated studies with nano-ESI provided greater resolution. “Now we’re looking at phagosomes with a modern digital camera, not with five pixels,” he said.

Leonard Foster, a postdoc in the department of biochemistry and molecular biology at the University of Southern Denmark, spoke about an isotope-labeling technique that he used as an alternative to the ICAT technique commonly used in differential protein expression studies. The new technique — stable-isotope labeling by amino acids in cell culture, or SILAC — differs from ICAT, which was developed by Ruedi Aebersold, in that it labels metabolically rather than chemically, according to Foster.

Rebecca Heald, assistant professor of molecular and cell biology at UC Berkeley, also outsourced her samples for mass spec analysis by sending them to John Yates’ lab at the Scripps Research Institute (see Pioneer, p. 9). Yates used high-throughput shotgun LC/LC MS/MS analysis to look at Heald’s purified samples of midbodies — cellular structures involved in mitosis. After receiving a “huge list of proteins” from Yates, Heald’s lab determined the functions of many of the proteins by knocking out gene expression with RNAi and assaying the resulting cells for mitotic activity.

Thursday, Aug. 28

Biomarkers were the hot topic on the final day of the conference, with four talks devoted in large part to biomarker discovery. Richard Caprioli, director of the mass spectrometry research center at Vanderbilt University Medical Center, started off by discussing his research — recently published in the Lancet — describing the discovery of serum biomarker patterns for non-small cell lung cancer using bioinformatic clustering (see PM 9-5-03). Caprioli said that he was able to find patterns that predicted poor versus good prognosis with 95 percent accuracy, and was also able to look at the effects of drugs on tumor tissue over time. “These are not just fingerprints — they are molecular weight annotated patterns,” he said.

Paul Tempst, director of the Protein Center at Memorial Sloan-Kettering Cancer Center, collabor-ated with Bruker Daltonics — using their Ultraflex MALDI MS/MS platform with reverse phase magnetic separation beads (see PM 9-5-03) — to look at serum protein patterns for solid tumor cancers. Referring to the similar work of Lance Liotta and Emanuel Petricoin (see PM 7-18-03), who looked at serum patterns for ovarian cancer using Ciphergen’s SELDI techno-logy, Tempst asked, “If they can get this data with their methods, what about if you use a high-end platform?” Tempst said he got better reproducibility and more peaks using the Bruker instrument and the magnetic beads, rather than SELDI. Like Caprioli, he used clustering analysis to identify patterns.

Stanley Hefta, executive director for proteomics at Bristol-Myers Squibb, showed how the company used multi-dimensional micro-LC and LC-ESI-MS to identify the differential expression of proteins in hepatic cell lines that are receiving toxic versus non-toxic drugs. To emphasize to his audience the potential significance of biomarker discovery, he noted, “We are making business decisions based on this.”

Steven Carr, senior director of discovery technologies at Millen-nium Pharmaceuticals, discussed how he worked with Roche Diagnostics to look for prognostic biomarkers for rheumatoid arthritis. Carr used LC MS/MS to look for the biomarkers, then did preliminary validation using antibody screening. Carr used Agilent’s abundant protein removal column (see PM 8-18-03) before serum analysis. Roche is now trying to develop immunological tests against the markers that Carr found.

Stephen Naylor, chief technology officer of Beyond Genomics, also spoke Thursday about systems biology techniques that analyze a disease by combining data from genomics, proteomics, and metabolomics into one large data set for integrated cluster analysis.

— KAM

 

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