# Analyzed PD-L1-positive subpopulations by dual-labeling TSA-IF-FISH predicts immunotherapy efficacy in advanced lung cancer

## TL;DR

A new dual-labeling method helps predict immunotherapy response in lung cancer by analyzing PD-L1-positive cells in biofluids.

## Contribution

A novel dual-labeling TSA-IF-FISH assay enables concurrent detection of PD-L1 and chromosomal changes in rare cell populations.

## Key findings

- PD-L1+/Vimentin CTCs correlate with improved immunotherapy response.
- PD-L1+ CTECs are linked to treatment resistance and tumor markers.
- PD-L1+ WBC levels correlate with systemic inflammation markers like C-reactive protein.

## Abstract

The evaluation of predictive biomarkers in advanced lung cancer requires methods that can comprehensively profile rare cell populations. We developed a dual-labeling assay integrating tyramide signal amplification immunofluorescence (TSA-IF) with fluorescence in situ hybridization (FISH) to concurrently detect protein expression and chromosomal aberrations. This approach was used to analyze PD-L1-positive circulating tumor cells (CTCs), circulating tumor endothelial cells (CTECs), and white blood cells (WBCs) from multiple biofluid types. Our assay improved signal integrity and revealed distinct clinical associations: specific PD-L1+ CTC phenotypes were linked to metastasis and correlated with improved immunotherapy response, whereas PD-L1+ CTECs were associated with treatment resistance and serum tumor markers. Furthermore, PD-L1+ WBC levels were strongly correlated with C-reactive protein, connecting them to systemic inflammation. This integrated liquid biopsy strategy enables a multifaceted view of the tumor microenvironment and host immune status, presenting a conceptual advance for monitoring treatment efficacy and inflammatory activity in advanced lung cancer.

•PD-L1+/Vimentin CTCs are correlate with immunotherapy response•PD-L1+ CTECs are associated with immunotherapy resistance•PD-L1+ WBCs correlate with inflammatory response

PD-L1+/Vimentin CTCs are correlate with immunotherapy response

PD-L1+ CTECs are associated with immunotherapy resistance

PD-L1+ WBCs correlate with inflammatory response

Health sciences