Circulating CD28-KLRG1+CD8+ T cells as prognostic indicators in advanced NSCLC chemoimmunotherapy.

Abstract

8553 Background: Chemoimmunotherapy has become the standard first-line treatment for advanced non-small cell lung cancer (NSCLC), but patient responses vary. Consequently, identifying predictive biomarkers is crucial to optimize therapeutic strategies. Circulating T cells are a promising focus due to the convenience of blood sampling and the feasibility of repeated monitoring. Deciphering the specific T-cell subsets that respond to chemoimmunotherapy is critical for personalizing treatment and improving outcomes in advanced NSCLC patients. Methods: We conducted unsupervised analysis using multi-color flow cytometry on peripheral blood samples from 30 NSCLC patients enrolled in a phase 2 clinical study (ClinicalTrials.gov NCT04836728) to explore correlations between immune cell subsets with therapeutic outcomes. We integrated single-cell RNA and T cell receptor sequencing data from peripheral blood, tumor, and non-tumor tissues of 8 NSCLC patients to study the transcriptional state of key cell types involved in these correlations. Results: Flow cytometry analysis revealed that a higher proportion of CD28-KLRG1+ CD8+ T cells was found in the peripheral blood of patients with durable clinical benefit (DCB) and improved overall survival (OS). Within these T cells, the CD57+ subset was positively correlated with OS at baseline, while the CD57- subset was negatively correlated. However, during treatment, both subsets showed a positive association with OS, highlighting the predictive value of CD28-KLRG1+ CD8+ T for chemoimmunotherapy response. Further phenotypic and functional analyses demonstrated that CD28-KLRG1+ CD8+ T cells are highly proliferative (Ki67) and produce anti-tumor cytokines (IFN-γ, IL-2, and TNF-α) upon TCR stimulation, indicating their immune-responsive role. Although these cells expressed relatively low levels of exhaustion markers such as PD-1 and TIGIT, they exhibited high expression of TCF1 and TOX, pointing to a progenitor exhausted T cell state characterized by reduced exhaustion and enhanced functional potential. Single-cell transcriptomic and TCR profiling revealed that CD28-KLRG1+ CD8+ T cells underwent significant clonal expansion in the peripheral blood during chemoimmunotherapy, evidenced by the higher clonality index and lower Inverse Simpson index, indicating their superior clonal expansion capacity upon activation. Longitudinal analysis showed that these cells had the highest proportion of expanded clones during treatment, primarily distributed in the effector T cell clusters, suggesting their anti-tumor activity during chemoimmunotherapy. Conclusions: Circulating CD28-KLRG1+ CD8+ T cells are a valuable biomarker for predicting outcomes in first-line chemoimmunotherapy for patients with advanced NSCLC. These findings highlight their functional activity, clonal expansion, and role in antitumor immunity during treatment.