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254 Development of Clinical Quantitative Multiplex High-Throughput Mass Spectrometric Immunoassay for Detecting Low Abundance Cancer Related Proteins/Peptides in Bodily Fluids

Number of anticipated awards: 4
(Fast-Track proposals will be accepted.)
Budget (total costs): Phase I: $150,000;
Phase II: $1,000,000

The deadline for receipt of all contract proposals submitted in response to this solicitation is: 5:00 p.m. Eastern Standard Time Monday, November 5, 2007

The application of proteomics tools in the clinical setting lags far behind their use in basic science and drug discovery. In the past, protein/peptide biomarkers were tested individually to determine their value using common techniques such as ELISA, 2-D gels, and mass spectrometry. Each of these technologies has its advantages, but they still suffer from an inability to quantitatively evaluate multiple markers in a single reaction. However, recent applications of affinity mass spectrometry into clinical laboratories brought a renewed interest in mass spectrometric immunoassays as a more specific affinity method capable of selectively targeting and studying protein biomarkers. In mass spectrometry-based immunoassays, proteins are affinity retrieved from biological samples via surface-immobilized antibodies, and are then detected via mass spectrometric analysis. The assays benefit from dual specificity, which is brought about by the affinity of the antibody and the protein mass readout. The mass spectrometric aspect of the assays enables single-step detection of protein isoforms and their individual quantification.

Therefore, the NCI is interested in proposals that focus on developing a multiplexed mass spectrometric immunoassay for the detection of low abundance cancer related proteins/peptides from bodily fluids (examples of “bodily fluids” include plasma or serum, serous fluids collected from ductal lavage, but not cell lysates or tissue culture media). Proposals should describe how the proposed technology will be highly specific, highly selective and have ultra-sensitive detection capabilities (at least within the ng/mL range) with limited sample preparation. Proposals should also distinguish any new methods of multiplex fabrication, novel immunoaffinity capture systems, and/or new detection/quantification systems. All responses must deliver a reasonable method for working with complex bodily fluids. In addition, maximum level of multiplexing, volume of sample requirement, with sample processing/analysis time must be addressed. Surface enhanced laser desorption ionization (SELDI) MS will not be considered for this SBIR due to its limited ability to comprehensively measure and identify low abundance proteins in serum or plasma considered to be within the dynamic range of proteins released from cancer cells.

Project goals:

Of the hundreds of thousands of proteins believed to be found in different body fluids, it is likely that cancer-related proteins will be in relatively low abundance. The development of effective technologies to accurately measure these proteins and improve our diagnostic capabilities by discerning diseased from non-diseased states requires the development of next-generation proteomic technologies. The purpose of this project is to stimulate the development of multiplexed mass spectrometric immunoassays for the detection of low abundance cancer related proteins/peptides from bodily fluids in support of the Clinical Proteomic Technologies Initiative (http://proteomics.cancer.gov). In addition, this tool as conceived is to be applicable in Cancer Centers and other settings where NCI Investigators conduct clinical care.

Phase I activities and expected deliverables:

During the Phase I proof-of-concept stage, offerors are encouraged (but not required) to pursue any of the biomarkers listed in the following reference: Malu Polanski and N. Leigh Anderson. (2006) “A List of Candidate Cancer Biomarkers for Targeted Proteomics.” Biomarker Insights 2:1-48

  • Demonstration of feasibility of the innovative approach.
  • Produce an initial product prototype in working with the Clinical Proteomic Technologies Initiative community.
  • Conduct usability testing with product prototype with representative users (e.g., Clinical Proteomic Technologies Initiative community).
  • Make modifications to the prototype based on results obtained from usability testing.
  • Compare findings to ELISA-based technologies. Detection limits should aim to be measured and reported as absolute quantitations that equal or surpass current ELISA measurements.
  • Prototype requirements include sample volumes less than 50 microliters, multiplex a minimum of 5 markers, high sensitivity (detection limit lower than 5 picogram/microliter), high reproducibility (CV’s less than 10%), and broad dynamic range (gram/Liter to nanogram/Liter).
  • Establish prototype revisions/additions to be implemented and tested in Phase II.
  • Present findings to an NCI Evaluation Panel.
  • Research should be proposed with quantitative feasibility milestones.

Phase II activities and expected deliverables:

  • Implement strategy and project plan for a fully functional quantitative, automated high-throughput multiplex affinity/protein capture technology for detecting low abundance cancer related proteins/peptides from bodily fluids.
  • Specificity greater than 95%.
  • Development of an affinity/protein capture technology with multiplexing capability up to 50 analytes (proteins/peptides) that implements the features, functions, and requirements developed in Phase I.
  • Project to be done in coordination with the Clinical Proteomic Technologies Initiative community to integrate platform into the technology assessment programs and greater scientific community.
  • Validate findings to ELISA.
  • Research should be proposed with quantitative feasibility milestones.

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