A novel multiplexed immunoassay identifies CEA, IL-8 and prolactin as prospective markers for Dukes' stages A-D colorectal cancers

Abstract

Background: Current methods widely deployed for colorectal cancers (CRC) screening lack the necessary sensitivity and specificity required for population-based early disease detection. Cancer-specific protein biomarkers are thought to be produced either by the tumor itself or other tissues in response to the presence of cancers or associated conditions. Equally, known examples of cancer protein biomarkers (e.g., PSA, CA125, CA19-9, CEA, AFP) are frequently found in plasma at very low concentration (pg/mL-ng/mL). New sensitive and specific assays are therefore urgently required to detect the disease at an early stage when prognosis is good following surgical resection. This study was designed to meet the longstanding unmet clinical need for earlier CRC detection by measuring plasma candidate biomarkers of cancer onset and progression in a clinical stage-specific manner. EDTA plasma samples (1 μL) obtained from 75 patients with Dukes' staged CRC or unaffected controls (age and sex matched with stringent inclusion/exclusion criteria) were assayed for expression of 92 human proteins employing the Proseek® Multiplex Oncology I proximity extension assay. An identical set of plasma samples were analyzed utilizing the Bio-Plex Pro™ human cytokine 27-plex immunoassay.

Results: Similar quantitative expression patterns for 13 plasma antigens common to both platforms endorsed the potential efficacy of Proseek as an immune-based multiplex assay for proteomic biomarker research. Proseek found that expression of Carcinoembryonic Antigen (CEA), IL-8 and prolactin are significantly correlated with CRC stage.

Conclusions: CEA, IL-8 and prolactin expression were found to identify between control (unaffected), non-malignant (Dukes' A + B) and malignant (Dukes' C + D) stages.

Keywords: Colorectal cancer; Multiplex immunoassay; Plasma biomarker.

Figure 1

Panels (P) (A, B, C, D & E) representing protein abundance data for CEA (A), IL-8 (B) prolactin (C), amphiregulin (D) and PDGF-BB (E) in individual Dukes’ stage A, B, C & D with E controls; pooled group (defined control, non-malignant, malignant groups). Values were determined by Proseek PEA using CRC patient EDTA plasmas and was based on z-scores (log2 scale).

Figure 2

SOMs trends (A-F) of control, non-malignant and malignant CRC groups normalised median Proseek biomarker protein expression scores, showing these can be clustered into six distinct trends.

Figure 3

SOMs trends (A-F) of control, non-malignant and malignant CRC groups normalised median Bio-Plex biomarker protein expression scores showing these can clustered into six distinct patterns/trends.

Figure 4

Spearman correlation scatter plots for 13 individual common biomarkers (PDGF-BB, IL-8, MCP1, IL-6, IL-7, VEGFA, IFNG, IL-2, GM-CSF, IL-4, IL-1ra, IL-12, and TNF) common to the Proseek and Bio-Plex multiplexed immunoassays.