title: Rapid label-free profiling of oral cancer biomarker proteins using nano-UPLC-Q-TOF ion mobility mass spectrometry.
abstract: OBJECTIVE
  It has become quite clear that single cancer biomarkers cannot in general provide high sensitivity and specificity for reliable clinical cancer diagnostics. This paper explores the feasibility of rapid detection of multiple biomarker proteins in model oral cancer samples using label-free protein relative quantitation.
  
  
  METHODS
  MS-based label-free quantitative proteomics offer a rapid alternative that bypasses the need for stable isotope containing compounds to chemically bind and label proteins. Total protein content in oral cancer cell culture conditioned media was precipitated, subjected to proteolytic digestion, and then analyzed using a nano-UPLC (where UPLC is ultra-performance liquid chromatography) coupled to a hybrid Q-Tof ion-mobility mass spectrometry (MS).
  
  
  RESULTS
  Rapid, simultaneous identification and quantification of multiple possible cancer biomarker proteins was achieved. In a comparative study between cancer and noncancer samples, approximately 952 proteins were identified using a high-throughput 1D ion mobility assisted data independent acquisition (IM-DIA) approach. As we previously demonstrated that interleukin-8 (IL-8) and vascular endothelial growth factor A (VEGF-A) were readily detected in oral cancer cell conditioned media(1), we targeted these biomarker proteins to validate our approach. Target biomarker protein IL-8 was found between 3.5 and 8.8 fmol, while VEGF-A was found at 1.45 fmol in the cancer cell media.
  
  
  CONCLUSIONS
  Overall, our data suggest that the nano-UPLC-IM-DIA bioassay is a feasible approach to identify and quantify proteins in complex samples without the need for stable isotope labeling. These results have significant implications for rapid tumor diagnostics and prognostics by monitoring proteins such as IL-8 and VEGF-A implicated in cancer development and progression. The analysis in tissue or plasma is not possible at this time, but the subsequent work would be needed for validation.
authors.affiliation: Department of Internal Medicine, School of Medicine, Yale University, New Haven, CT, USA.|Department of Chemistry, University of Connecticut, Storrs, CT, USA.
authors.forename: Ala F
authors.initials: AF
authors.lastname: Nassar
authors.affiliation: Waters Corporation, Milford, MA, USA.
authors.forename: Brad J
authors.initials: BJ
authors.lastname: Williams
authors.affiliation: Waters Corporation, Milford, MA, USA.
authors.forename: Dustin C
authors.initials: DC
authors.lastname: Yaworksy
authors.affiliation: Cancer Research Initiatives Foundation (CARF), Sime Darby Medical Centre, Subang Jaya, Malaysia.
authors.forename: Vyomesh
authors.initials: V
authors.lastname: Patel
authors.affiliation: Department of Chemistry, University of Connecticut, Storrs, CT, USA.|Neag Comprehensive Cancer Center, University of Connecticut Health Center, Farmington, CT, USA.|Department of Cell Biology, University of Connecticut Health Center, Farmington, CT, USA.|Institute of Material Science, University of Connecticut, Storrs, CT, USA.|School of Chemistry, National University of Ireland, Galway, Ireland.
authors.forename: James F
authors.initials: JF
authors.lastname: Rusling
mesh_terms: D014408:Biomarkers, Tumor / Q000032:analysis*; D002851:Chromatography, High Pressure Liquid; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D009062:Mouth Neoplasms / Q000175:diagnosis* / Q000378:metabolism*; D036103:Nanotechnology; D009363:Neoplasm Proteins / Q000032:analysis*; D014407:Tumor Cells, Cultured
chemical_list: D014408:Biomarkers, Tumor; D009363:Neoplasm Proteins
keywords: Clinical proteomics; Ion mobility mass spectrometry; Label-free profiling; Oral cancer; Screening