Abstract 4584: Verification of ovarian cancer biomarker candidates by nanoparticle-capture MRM

Abstract

Abstract Background: Detection of early stage ovarian cancer offers the best opportunity to reduce mortality. In the attempt to improve Ovarian Cancer prognosis there is a sheer need for new sensitive and specific early detection biomarkers. We have recently proposed a new workflow to overcome the limitations that characterize serum protein biomarker discovery. A new class of hydrogel (NIPAm) nanoparticle, with the incorporation of an affinity bait (AAC), had been developed and tested, showing to successfully separate from albumin, harvest, concentrate, and protect from degradation low abundance and LMW potential protein biomarkers, amenable for downstream mass spectrometry (LTQ-Orbitrap) analysis. Applying such workflow to a set of 40 serum samples from ovarian cancer patients and benign controls a list of 163 differentially expressed peptides was generated. Our novel nanoparticle technology provides a rapid and near 100% efficient means to capture and greatly amplfy the resting concentration of LMW protein analytes. Since most of these markers do not have well performing specific antibodies commercially available, in this work we utilized MRM to verify, in an independent study set of 20 cancer samples and 24 benign controls, their differential expression as first identified by Orbitrap-MS/MS. Method: NIPAm/AAC type of core-shell hydrogel nanoparticle was used for both orbitrap-LTQ based discovery and MRM verification such that the same analyte fractionation workflow was used but different downstream mass spectrometry. MRM was performed on a Thermo Quantum triple quadropole-MS using the nanoparticle trapped analyte archive as the input for analysis. On the basis of their biological significance and following manual verification of the raw LTQ-Orbitrap analysis data, a list of 35 biomarker candidates was selected for MRM verification. Among which i.e.: gelsolin isoform b, integrin α 2b preproprotein, apolipoprotein B precursor, fibronectin 1 isoform 2 preproprotein, LPS-binding protein precursor, ceruloplasmin, α-1apolipoprotein L1 isoform a precursor, leucine-rich α-2-glycoprotein 1, apolipoprotein C-IV, thrombospondin 1 precursor, pro-platelet basic protein precursor, serum amyloid A4, apolipoprotein H precursor, gelsolin isoform b, UNC-112 related protein 2 long form, H4 histone familymember J, cadherin 1, apolipoprotein AV, H2A histone family, member D, apolipoprotein C-III precursor, heterogeneous nuclear ribonucleoprotein AB isoform a, ras suppressor protein 1 isoform 2, cofilin 1 (non-muscle), a disintegrin & metalloproteinase domain 7, transgelin 2, S100-A7, mucin 16. Conclusions: Several ovary candidate biomarkers were verified by Nanoparticle capture-MRM, thus providing data supporting the use and continued improvement of this novel biomarker discovery system for the identification of protein biomarker panels for early disease detection and therapeutic monitoring. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4584.