Abstract P1-02-07: Delineation of breast cancer based on circulating protein biomarker profile using the OLINK proteomics multiplexed proximity extension assay
Authors: Jayachandran G , Cohen EN , Gao H , Alvarez RH , Valero V , Lim B , Woodward WA , Ueno NT , Reuben JM 
Published: March 2017

Abstract
Background: Expression of cancer related genes and proteins in clinical specimens are the mainstay of personalized targeted therapy. In this study we investigated a blood-based non-invasive and sensitive technique to map biomarkers in breast cancer patients at the protein level instead of expression of cancer related genes. Other multi-platform assays require a large amount of clinical material, multi-step sample processing and complicated data analysis. Proximity Extension Assay performs a harmonious blending of immunoassay and PCR to amplify protein expression signal, thereby enabling multiplexing with small sample input (1 µl).
Material and methods: Plasma samples (n=25) from patients with inflammatory breast cancer (IBC) or non-IBC, metastatic and non-metastatic, collected prior to starting a new therapy (treatment naive) or a new line of therapy were analyzed using the Proseek Multiplex Oncology I v2 panel (Olink Proteomics, Uppsala, Sweden) for simultaneous detection of 92 human protein biomarkers. In the assay, each protein biomarker is detected by a matched pair of antibodies coupled to unique DNA-tags. Upon binding to the proteins, the correctly hybridized DNA-tags form an amplicon that is measured by a digital PCR. The data was subjected to a dynamic Principal Component Analysis (PCA) with a T-test filter using Multid GenEx software allowing the identification of a specific protein signature with the greatest inter-group differences. Plasma from seven healthy normal donors (HD) was also included in the analysis as comparison.
Results: A plasma protein signature consisting of MK, elf-4B, VIM biomarkers delineated all breast cancers from normal healthy donors. IL-8, CD40L, GDF-15, MCP-1, PARK7, CXCL11, FADD, CAIX, CD69, MIC-A, VEGF-D, EGFR, elf-4B, VIM, and PRSS8 distinguished patients with metastatic breast cancer from normal healthy donors. PCA also identified differences between IBC patients, non-IBC patients, and healthy donors. Clustered association of several protein biomarkers (MCP-1, VIM, HGF, PRSS8 and HER2) distinguished IBC from healthy donors. Plasma from IBC and non-IBC patients differed in their distribution of TNFSF14, CAIX, KN1A, CDHE4 and EGFR. All protein comparisons have p values &lt; 0.05.
Conclusion: These preliminary data suggest that it is possible to distinguish between cancer patients and healthy normal donors, and IBC and non-IBC patients based on a plasma protein profile using the Proseek Multiplex Oncology I v2 panel from OLINK with just 1 µl of plasma. This pilot study will lead to the establishment of a training set for a protein signature to be applied in a subsequent validation in a larger cohort to assess treatment induced biomarker profile changes.
Citation Format: Jayachandran G, Cohen EN, Gao H, Alvarez RH, Valero V, Lim B, Woodward WA, Ueno NT, Reuben JM. Delineation of breast cancer based on circulating protein biomarker profile using the OLINK proteomics multiplexed proximity extension assay [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P1-02-07.