Abstract 2292: Novel method for the detection and evaluation of disease biomarkers on white blood cells from liquid biopsies
Authors: Bayer Erin , Ladd Jon , Nordberg Joshua , Ramirez Arturo B. , Baker Jessica A. , Daly Thomas 
Published: June 2023

Abstract
Purpose: Blood based liquid biopsies of circulating tumor cells (CTCs) are a non-invasive tool with proven effectiveness in the prognosis, treatment selection, and stratification of cancer patients. In this study we demonstrate a novel application of the RareCyte platform, which was developed to find and analyze rare CTCs from blood samples, in evaluating white blood cell (WBC) sub-populations for disease biomarkers.
Experimental Procedures: Using AccuCyte®, an unbiased density-based method for collecting nucleated cells from whole blood, we transferred the nucleated cells to slides and stained for CD14, CD66b, and CK/EpCAM to identify monocytes (CD14+/CD66b-), granulocytes (CD14-/CD66b+), and CTCs (CK/EpCAM+), as well as a drug-target biomarker to evaluate its expression across all cells. These slides were then imaged with CyteFinder®, an automated multiparameter immunofluorescent (IF) microscopy system that applies machine learning algorithms for cell identification. Each slide contains millions of cells, so we developed novel scanning and analysis algorithms to quantify WBC sub-populations in addition to rare CTCs.
Results: In addition to detection and enumeration of CTCs, the RareCyte system was able to detect individual white blood cells and quantify monocyte and granulocyte subpopulations. Our results correlated very well with standard CBC (Complete Blood Count) readings from multiple donors. Additionally, the monocyte and granulocyte populations were evaluated for expression of a drug-target biomarker. While expression was observed on both granulocytes and monocytes, monocytes had a significantly higher expression of the drug-target biomarker, a critical finding for future implementation of this drug.
Conclusions: Our results show that through modifications to the RareCyte platform we can identify rare and non-rare cell populations simultaneously and evaluate expression of biomarkers and drug-targets across multiple cells of interest. This new application combines the benefits of high throughput data analysis of flow cytometry with the specificity and sensitivity of rare cell detection. Additionally, high-resolution multiplexed immunofluorescent imaging provides cell morphology and biomarker localization information. Our ability to add multiple biomarkers in conjunction with CTC and other rare cell detection provides multiple avenues for future research. One possible application is characterizing white blood cells that cluster with CTCs, which are known to portend a worse prognosis for patients. Our modified workflow could be implemented to precisely characterize these WBCs that contribute to the metastatic process. Utilization of the RareCyte platform for both rare and non-rare cell analysis presents an exciting opportunity for a deeper evaluation of patients and their prognosis through liquid biopsies.
Citation Format: Erin Bayer, Jon Ladd, Joshua Nordberg, Arturo B. Ramirez, Jessica A. Baker, Thomas Daly. Novel method for the detection and evaluation of disease biomarkers on white blood cells from liquid biopsies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2292.