Abstract C27: Temporal proteomic characterization of spontaneous ovarian cancer in the chicken
Authors: Hawkridge Adam M. , Wysocky Rebecca B. , Petitte James N. , Anderson Kenneth E. , Mozdziak Paul E. , Horowitz Jonathan M. , Muddiman David C. 
Published: May 2012

Abstract
Epithelial ovarian cancer (EOC) remains the most lethal gynecological cancer in the Western world due to a combined lack of effective therapeutics and screening strategies. The inability to identify effective EOC screening strategies has been attributed to the heterogeneous pathogenesis of EOC, difficulty in obtaining significant numbers of human EOC tumors at the initial stages of development, and the lack of in vitro and in vivo experimental models that recapitulate the onset of EOC. The chicken has emerged as a promising animal model for studying spontaneous EOC. There are significant factors that support this claim including 1.) a disease prevalence of 5–35% between 2–7 years of age and 2.) molecular level similarities to human EOC including CA-125 expression and the mutational frequency of the p53 and HER-2/neu genes. Finally, with regard to biomarker discovery (in blood), 1–3 ml of blood can be drawn from the chicken without sacrificing the animal. This allows for longitudinal sampling (vide infra) and the assessment of nonpathological variability of biological constituents (e.g., proteins) versus changes specifically related to the onset of disease. The overall goal of this research is to identify candidate early stage EOC biomarker(s) using advanced proteomics technology to characterize longitudinal plasma samples from the chicken.
A total of 250, age-matched 2 year old birds were obtained in May 2007. Approximately 2 ml of blood was drawn from each bird every 3 months for one year starting at 2.5 years of age (5 time points total). The blood samples were processed immediately to plasma and stored at −80°C. At the conclusion of the study in October 2008 (3.5 years of age), the 106 birds that remained alive were euthanized and necropsied. Healthy and cancerous tissues from these birds were preserved by snap freezing in LN2, RNAlater, and formalin for proteomic, genomic, and pathological characterization respectively. The gross pathology of all 106 birds showed that 40% had neoplasms of which 17% had early stage EOC, 7% had late-stage EOC, 38% had advanced EOC/oviductal cancer (cancer origin unknown), and the remaining 38% had tumors present on tissues other than the ovary.
Longitudinal plasma samples from 2 birds were chosen for initial quantitative proteomic characterization. One bird was considered “healthy” with no visible neoplasms and the second bird had advanced EOC/oviductal cancer that had metastasized to neighboring tissues. The longitudinal plasma samples were separated by 1D gel electrophoresis, subjected to in-gel tryptic digestion, and fractionated for proteomic analysis. Samples were analyzed using a splitless nanoLC coupled to an LTQ-Orbitrap mass spectrometer. Protein identification and label-free quantification were accomplished using the Mascot and ProteoIQ software platforms respectively. Method development, analytical reproducibility, and the quantitative changes of 2 plasma proteomes as a function of time and health status (with and without cancer) will be presented.
Citation Information: Cancer Res 2009;69(23 Suppl):C27.