Detection of PD-L1 MRNA expression in circulating tumor cells (CTCS) from patients with metastatic NSCLC, HNSCC and melanoma using a novel highly sensitive commercially available CE-IVD KIT

Abstract Background: We have previously reported the development of an RT-qPCR assay for PD-L1 expression in CTCs (Strati et al, Ann Oncol, 2017). This assay is now commercially available as Oncolipsy PD-L1 kit (CE-IVD, Pharmassist, Greece) and we report here it’s application in size-based enriched CTCs from patients with metastatic NSCLC, HNSCC and melanoma, before and during PD-1/PD-L1 blockade therapies. Patients and Methods: The Oncolipsy PD-L1 kit was analytically validated using Mini-Genes as positive controls (IDT, USA). The analytical sensitivity was tested by spiking known numbers of H1975 cells in peripheral blood (PB) samples from healthy donors (HD). In total 95 patients were enrolled in the study: 69 patients with metastatic NSCLC, 19 patients with metastatic HNSCC and 7 patients with metastatic melanoma. 40 PB samples from HD were used as a control group. 10 ml of PB was collected at baseline (V0), two months after immunotherapy (V1) and at the time of disease progression (PD) or 18 months after initiation of immunotherapy (V2). PD-L1 and B2M transcripts were quantified in cDNAs obtained from CTCs isolated with the size-based PARSORTIX (ANGLE, UK) device. A sample was considered as CTC-positive when cDNAs were positive for CK-8, CK-18 or CK-19 expression. Results: Spiking experiments have shown that 10 H1975cells/10ml PB can be detected using the kit. In CTC-positive samples, overexpression of PD-L1 was detected in 15/58(25.8%) patients at V0, in 6/34(17.6%) patients at V1 and in 3/11(27.3%) at V2. Patients with elevated expression of PD-L1 at V1 in respect to V0 and detectable CTCs had longer PFS (18.5mo vs. 13.2mo, p=0.033) and longer OS (24.8mo vs. 12.8mo, p = 0.071) after initiation of immunotherapy. Patients with elevated PD-L1 at V1 compared to V0 benefited from immunotherapy, with a longer DFS (17.8mo vs. 13.4mo) and OS (29.4mo vs. 26.9mo). Conclusions: The commercially available Oncolipsy PD-L1 kit can be used for the quantification of PD-L1 transcripts in CTCs. Expression of PD-L1 in size-based enriched CTCs can predict response to PD-1/PD-L1 blockade therapy. Acknowledgements: This study has been financially supported by the European Union and Greek national funds through the Operational Program Competitiveness, Entrepreneurship and Innovation, under the call RESEARCH - CREATE - INNOVATE (project code: T1RCI-02935). Trial registration: ClinicalTrials.gov Identifier: NCT04490564 Citation Format: Areti Strati, Martha Zavridou, Stavroula Smilkou, Victoria Tserpeli, Eleni Tzanikou, Dimitra Stergiopoulou, Eleni Efthimiadou, Emily Tsaroucha, Amanda Psyrri, Aggeliki Sfika, Evangelos Bournakis, Ioanna Balgkouranidou, Stylianos Kakolyris, Ioannis Boukovinas, Christos Papadimitriou, Loukas Kaklamanis, Ioanna Koukli, Evi Lianidou. Detection of PD-L1 mRNA expression in circulating tumor cells (CTCs) from patients with metastatic NSCLC, HNSCC and melanoma using a novel highly sensitive commercially available CE-IVD kit [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7488.