Immunomagnetic isolation of circulating melanoma cells and detection of PD-L1 status

Joseph W Po,Daniel Brungs,Yafeng Ma,Farhad Azimi,Bavanthi Balakrishna,Paul De Souza,Therese M Becker,Nikolas K Haass

doi:10.1371/journal.pone.0211866

Abstract

Personalised medicine targeted to specific biomarkers such as BRAF and c-Kit has radically improved the success of melanoma therapy. More recently, further advances have been made using therapies targeting the immune response. In particular, therapies targeting the PD-1/PD-L1 or CTLA-4 axes alone or in combination have shown more sustained responses in 30–60% of patients. However, these therapies are associated with considerable toxicities and useful biomarkers to predict responders and non-responders are slow to emerge. Here we developed a reliable melanoma circulating tumor cell (CTC) detection method with PD-L1 evaluation on CTCs. A set of melanoma cell surface markers was tested as candidates for targeted melanoma CTC isolation and a melanoma specific immunostaining-based CTC identification protocol combined with PD-L1 detection was established. In vitro testing of the effect of exposure to blood cells on melanoma cell PD-L1 expression was undertaken. Immunomagnetic targeting isolated melanoma CTCs in up to 87.5% of stage IV melanoma patient blood samples and 3 8.6% of these had some PD-L1 expressing CTCs. Our in vitro data demonstrate PD-L1 induction on melanoma cells in the blood.This study established a robust, reliable method to isolate melanoma CTCs and detect expression of PD-L1 on these cells.

Highlights

Improved technology for the capture of circulating tumor cells (CTCs) is increasing the utility of CTCs to predict prognosis and patient survival
The aim of the current study is to demonstrate that screening PD-L1 from liquid biopsies (CTCs) is feasible with the use of an efficient protocol to isolate melanoma CTCs
The most comprehensive previous investigation of melanoma antigens suitable for immunomagnetic isolation of melanoma CTCs was reported by Freeman et al Their study chose antigens either relatively highly expressed on melanoma cells (MCAM and MCSP) or expressed on subpopulations of melanoma cells proposed to be aggressive and competent to initiate metastasis (CD271 and ACBC5)

Summary

Introduction

Improved technology for the capture of circulating tumor cells (CTCs) is increasing the utility of CTCs to predict prognosis and patient survival. CTCs are a non-invasive biosource for molecular biomarker detection that can inform precision therapy and together with analysis of circulating tumor nucleic acids (ctRNA and ctDNA) are emerging with high potential for widespread clinical utility (reviewed by [1,2,3]). One challenge for biomarker testing from common tissue biopsies is tumor heterogeneity. It is widely accepted that a single tissue biopsy is poorly representative for a patient’s cancer. This is particular relevant in advanced malignancies, where biopsies of the primary tumor provide limited information at a time of therapy resistance and tumor progression [4].

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