214P - DNA-damaging therapies trigger transcriptome and metabolism changes in peripheral NK cells of SCLC patients

Background

Small cell lung cancer (SCLC) is currently treated with a combination of chemotherapy and immunotherapy. Unfortunately, not all patients benefit from this regimen and few alternative therapies are available. Based on recent transcriptomic data defining an “inflamed” subtype of SCLC patients who benefit from immunotherapy, we hypothesised that innate immune activation and natural killer (NK) cells may provide an opportunity to evaluate immune-responsive patients.

Methods

SCLC patients receiving two cycles of chemo-immunotherapy were divided into best responders (BR) and non-responders (NR). NK cells from SCLC patients (n=10) were isolated from peripheral blood-collected PBMCs and subjected to RNA-seq and gene set enrichment analysis (GSEA). Isolated NK cells were in vitro treated with DNA-dependent protein Kinase (DNA-PK) inhibitor. RT-PCR, Seahorse assay and flow cytometry were performed to investigate immune checkpoint expression.

Results

GSEA analysis of RNAseq data on NK cells from BR patients of SCLC cohorts showed high levels of IFN-related pathways (IFN_GAMMA_RESPONSE NES=2.77; p<0.0001, IFN_ALPHA_RESPONSE NES=2.49; p<0.0001) and Influenza infection (NES=3.57; p<0.0001) enrichment in BR patients derived NK cells in comparison to NR patients. In addition, NK cells from BR patients showed enrichment in oxidative phosphorylation genes (NES=2.14; p<0.0001) as well as DNA repair pathway (NES=2.07; p<0.0001). ISMARA analysis revealed that BR patients-derived NK cells showed high Interferon regulatory factor 3 (IRF3)-binding motif (Pearson corr. coeff. = 0.76; p<0.0001) compared to the NR cohort. In vitro treatment of NK cells with DNA-PKi significantly increased multiple innate immunity cytosolic DNA/RNA sensors, namely cGAS-STING (p < 0.0001), MAVS (p < 0.0001) and IFI16 (p < 0.0001). Finally, metabolic analysis showed an increase in ATP production Rate and phenotypic characterization by flow cytometry showed an increase in ICOS (CD278) expression in DNA-PKi- treated NK cells.

Conclusions

Comprehensive transcriptomic analysis of SCLC-isolated NK cells revealed significant positive modulation of innate immune pathways, suggesting a potential role for DNA-PKi in metabolic and phenotypic remodelling.

Legal entity responsible for the study

The authors.

Funding

Associazione Italiana per la Ricerca sul Cancro (AIRC).

Disclosure

All authors have declared no conflicts of interest.