Skip to main content
Premium Trial:

Request an Annual Quote

Moffitt Team Developing MRM-MS Assay Database Focused on Key Cancer Proteins


By Adam Bonislawski

Researchers at the Moffitt Cancer Center are developing a database of multiple-reaction monitoring mass spectrometry assays focused on key cancer proteins.

Called the Quantitative Assay Database, or QUAD, the resource provides complete workflows for MRM-MS-based quantitation and is intended as a narrowly focused but more in-depth complement to resources like the SRMAtlas developed by the Institute for Systems Biology and the Swiss Federal Institute of Technology Zurich, said John Koomen, a Moffitt researcher and leader of the QUAD project.

With more than 170,000 single-reaction monitoring assays, SRMAtlas offers "a significantly larger number of entries … but I think we're providing a little bit more detail," Koomen told ProteoMonitor. "We hope that over time [QUAD] will become a valuable resource for people who want to study cancer biology and help bridge the gap between biologists and resources like SRMAtlas that are really intended for people who are experts in mass spec or analytical chemistry."

The goal was to develop “something that would allow us, as we progressed, to integrate the [MRM] assays together," he said. "So one of the things that we've tried to do is to link everything up at a biological pathway level. We've taken a lot of the canonical signaling pathways that are involved in cancer and put together a list and developed assays against a number of the major players in each category."

These proteins are presented in QUAD as components within a cancer pathway diagram. Users can then click on individual proteins to get information on isolating and separating the protein as well as what peptides the Moffitt researchers think will work best for measuring that particular molecule. The layer below that contains data on these particular peptides – both the endogenous forms and the stable isotope-labeled varieties developed for the assays.

Koomen's team builds the assays using SDS-PAGE-fractionated lysates from cancer cell lines. SDS-PAGE's simplicity makes it a relatively easy technique for any lab to apply, he said, while using cancer cell lines as sample sources makes for easier assay development given that the target proteins are often significantly overexpressed in these lines compared to normal cells.

The database, which launched publicly in July, currently has around 100 MRM assays for proteins involved in colon and lung cancer, melanoma, leukemias, and myeloma. This includes assays for roughly 20 proteins involved in beta-catenin signaling – a key pathway in colon cancer – that Koomen's team developed as part of a study published last year in the Journal of Proteome Research, as well as assays for proteins linked to drug resistance in multiple myeloma that were generated for a study currently in press at Molecular & Cellular Proteomics.

QUAD's focus on particular cancer pathways mirrors the recent move by some proteomics databases — SRMAtlas among them — to include more data relevant to specific research questions. At the annual meeting of the Human Proteome Organization in September, ISB researcher Rob Moritz, one of the leaders of the SRMAtlas project, presented an addition to the database called Passel that's meant to allow researchers to upload data on SRM assays performed in particular biological systems (PM 9/9/2011).

Passel, Moritz said, will enable researchers to compile "multiple views of the same peptide by different groups," letting them "analyze how well a particular peptide perform[s] in the different biological experiments that people are doing."

Given the variables inherent in protein quantitation, such multiple views are important to maximizing the utility of these databases, Koomen noted. For instance, he said, his lab uses SDS-PAGE to fractionate the proteins. “But if you put, say, digest of whole cell lysate into the mass spectrometer, do these assays still hold up? We don't really know because what works in one assay is dependent on all the pre-analytical and analytical variables of that assay."

One issue that Koomen said he hopes to explore is how different sample collection techniques affect assay performance. Moffitt has an extensive collection of frozen tissue samples that its researchers use in their work, but for many other labs, formalin-fixed paraffin-embedded tissues are a primary sample source.

"Most of the time the two are comparable," he said, "but there have been occasions when the peptides that are appropriate for formalin-fixed tissue are different from those that work well for frozen tissue."

"I think it's very important for LC-MRM-MS that [researchers] be as straightforward and as focused on data sharing as possible," Koomen said, adding that he hoped the resource would help move MRM mass spec beyond its traditional role as a biomarker quantitation tool and into more general biological research.

"Most of the people who are doing LC-MRM are using it for biomarker evaluation, and it's perhaps a situation in which people are very protective of their data and their intellectual property," he said. "By sharing the information here, we want to make this a tool that's relevant to the broader study of biology, in addition to being a biomarker measurement device."

Moffitt hopes to include data from outside researchers as part of QUAD, Koomen said, and the database includes a system by which groups can submit assay information. His team also plans in the future to add MRM assay information from the literature to the resource.

Koomen's group is also working with UK-based proteomics company Proteome Sciences on MRM assay development. In July, the firm signed an agreement to support the development at Moffitt of mass spec assays measuring signaling and repair pathway proteins for clinical tumor biopsy analysis (PM 7/8/2011).

Have topics you'd like to see covered in ProteoMonitor? Contact the editor at abonislawski [at] genomeweb [.] com.
The Scan

New Study Highlights Role of Genetics in ADHD

Researchers report in Nature Genetics on differences in genetic architecture between ADHD affecting children versus ADHD that persists into adulthood or is diagnosed in adults.

Study Highlights Pitfall of Large Gene Panels in Clinical Genomic Analysis

An analysis in Genetics in Medicine finds that as gene panels get larger, there is an increased chance of uncovering benign candidate variants.

Single-Cell Atlas of Drosophila Embryogenesis

A new paper in Science presents a single-cell atlas of fruit fly embryonic development over time.

Phage Cocktail Holds Promise for IBD

Researchers uncovered a combination phage therapy that targets Klebsiella pneumonia strains among individuals experiencing inflammatory bowel disease flare ups, as they report in Cell.