Abstract A038: Investigating PD-L1 status in circulating tumor cells isolated from blood samples of lung cancer patients
Authors: Spode Morgan , Goodwin Chloe , Varatharajah Aarabhi , Cottingham Aaron , Duhig Elisha , Kaja Laura , Greaves David , Ciccioli Mariacristina , Pailhes-Jimenez Anne-Sophie 
Published: November 2024

Abstract
Background: This research study aimed to assess the performance of a research use only (RUO) immunofluorescence (IF) assay targeting programmed death-ligand 1 (PD-L1) evolution in epithelial and/or mesenchymal circulating tumor cells (CTCs) isolated from blood using Parsortix® microfluidic technology. Upregulation of PD-L1 enables cancer cells to evade the host immune response. Assessment of PD-L1 status in the tumor, as determined by traditional tissue biopsy, can indicate if immunotherapy has the potential to be an effective treatment. However, PD-L1 expression in tumor biopsies may not reflect metastatic sites in their entirety and can become outdated during tumor evolution, as the process cannot be repeated due its invasive nature. Liquid biopsy offers a minimally invasive option for dynamic testing of PD-L1 in CTCs as patients undergo treatment. Methods: Analytical linearity, and specificity and sensitivity were assessed by spiking HCC1954 and Hs 578T cancer cell lines into the blood samples of 34 healthy volunteers. The assay was then evaluated in samples from 47 metastatic lung cancer patients, among which 32 had a known PD-L1 tissue status (19 positive, 13 negative). Up to six successive draws were taken for each patient, collected into Streck Cell-Free DNA blood collection tubes. ANGLE’s Parsortix® instruments were used to isolate spiked cancer cells or CTCs from blood samples based on their size and deformability. The CTC- enriched harvests were processed onto ANGLE’s CellKeepTM slides and IF stained with ANGLE’s Portrait® PD-L1 RUO assay for PD-L1 identification on CTCs. Slides were imaged on a BioView automated imaging system. Results: Analytical specificity of PD-L1 was 98% and analytical sensitivity was 81%. Analytical linearity (R2=0.91) was shown over a 0–500 cells range. In metastatic lung cancer patients, ≥1 CTC was identified in at least one draw for 91% of donors, with 55% of those donors exhibiting ≥1 PD-L1 positive CTC. A majority (81%) of donors exhibited a positive correlation between PD-L1 tissue status and CTC expression. However, the remaining 19% of donors exhibited discordance with a PD-L1 negative tissue status but PD-L1 positivity identified in ≥1 CTC. Of the PD-L1 positive CTCs identified, 98% expressed mesenchymal markers. Dynamic change was shown in all but one donor, with PD-L1 status of the CTCs changing over time. Conclusions: This study demonstrated the ability to determine PD-L1 status in CTCs from the blood of metastatic lung cancer patients and, subject to further study, the future potential to develop dynamic PD-L1 testing for advancement of more personalized cancer treatments. Patients with a PD-L1 negative tissue status exhibited PD-L1 positive CTCs, demonstrating the feasibility of a more repeatable and accurate assessment of PD-L1 status than can be achieved through a traditional tissue biopsy.
Citation Format: Morgan Spode, Chloe Goodwin, Aarabhi Varatharajah, Aaron Cottingham, Elisha Duhig, Laura Kaja, David Greaves, Mariacristina Ciccioli, Anne-Sophie Pailhes-Jimenez. Investigating PD-L1 status in circulating tumor cells isolated from blood samples of lung cancer patients [abstract]. In: Proceedings of the AACR Special Conference: Liquid Biopsy: From Discovery to Clinical Implementation; 2024 Nov 13-16; San Diego, CA. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(21_Suppl):Abstract nr A038.