University of Pittsburgh Cancer Institute (UPCI)

Overview

The Mass Spectrometry Platform provides metabolomic and clinical proteomic services within the Cancer Biomarker Facility. MSP provides:

• Proteomics
  • specific scientific and technical resources and expertise focused on clinical proteomics
  • proteomic profiling
  • analyses of biological samples
  • broad scientific expertise, knowledge, and technical support for experimental design, protocol development, data analysis, and interpretation
  • state-of-the-art proteomics instrumentation and technologies, with a primary emphasis on mass spectrometry-based systems
• Metabolomics
  • Metabolomic profiling and discovery
  • High-throughput quantitative metabolite assay development and deployment

MSP develops and validates approaches for focused clinical applications, including:

• early detection and screening for cancer
• molecular classification for diagnosis, prognosis, and response to therapy
• direct biological information regarding the "activity" of the expressed proteins specifically with regard to cellular metabolism and transformation of small molecules
• data on the efficacy, toxicity, carcinogenicity and teratogenicity of xenobiotics, including drugs and environmental pollutants

Experimental Workflows Available

• Protein identification
• Phosphoproteomics
• Glycoproteomics
• Protein-protein complexe identification
• Membrane proteomics
• Quantitative proteomics (label free, SILAC, ¹⁸O/¹⁶O)
• MS-based quantitative assay development (MRM assays)
• Biofluid proteomics (serum, CSF, urine…)
• Tissue proteomics (FFPE/fresh)
• MALDI-based profiling/imaging
• Bioinformatics/pathway analysis
• Metabolite Assays:
  • standard isotopic dilution mass spectrometry from biological samples for 16 estrogen metabolites and testosterone. Estriol (E3), estradiol (E2), estrone (E1), 16-epi E3, 17-epi E3, 16-keto E2, 16-α E1, 4-Me E1, 4-Me E2, 2-Me E1, 2-Me E, 3-Me E, 4OH E1, 4OH E2, 2OH E1, 2OH E2, and testosterone are separated by UPLC and quantitated using multiple reaction monitoring (MRM) of CID MS/MS.
  • β-nicotinamide adenine dinucleotide (NAD+) and related metabolites; adenosine 5′ -diphosphoribose (ADPR), β-nicotinamide mononucleotide (NMN), nicotinic acid mononucleotide (NAMN), nicotinic acid adenine dinucleotide (NAAD), adenosine 5′ -monophosphate (5′ AMP), nicotinamide (NAM) and nicotinic acid (NA) using 2-chloroadenosine as internal standard.
  • paclitaxel (Taxol®) from cell systems.
  • cyclosporin A quantitation from whole blood (cyclosproin D as IS).
  • Phenotyping of drug metabolizing enzymes (P450s) by the Pittsburgh Cocktail; a total of 7 drugs listed as follows used as probes to investigate in vivo activity and regulation of P450s: caffeine, chlorzoxazone, debrisoquine, mephenytoin, flurbiprofen, midazolam and dapsone.