The potential use of plasma-derived small extracellular vesicles (sEV) as predictors of response to therapy and clinical outcome in chemotherapy-naïve patients with non-small-cell lung cancer (NSCLC) was explored. sEV were isolated by size-exclusion chromatography from the plasma of 79 chemotherapy-naïve NSCLC patients and 12 healthy donors (HD). sEV were characterized with regard to protein content, particle size, counts by qNano, morphology by transmission electron microscopy, and molecular profiles by Western blots. PD-1 and PD-L1 expression on circulating immune cells was analysed by flow cytometry. Pre-treatment levels of total sEV protein (TEP) were correlated with overall (OS) and progression-free survival (PFS). The sEV numbers and protein levels were significantly elevated in the plasma of NSCLC patients compared to HD (p = 0.009 and 0.0001, respectively). Baseline TEP levels were higher in patients who developed progressive disease compared to patients with stable disease (p = 0.007 and 0.001, stage III and IV, respectively). Patient-derived sEV were enriched in immunosuppressive proteins as compared to proteins carried by sEV from HD. TEP levels were positively correlated with CD8+PD-1+ and CD8+PD-L1+ circulating T cell percentages and were independently associated with poorer PFS (p < 0.00001) and OS (p < 0.00001). Pre-therapy sEV could be useful as non-invasive biomarkers of response to therapy and clinical outcome in NSCLC.

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e20633 Background: Recently, the antiPD-L1 agent, durvalumab, was approved in first line treatment in combination with chemo. However, there is not a clear benefit from this regimen for the majority of patients. In this study, we explored the predictive role of regulatory T-cells in response of patients with ES-SCLC treated with the aforementioned regimen. Moreover, we evaluated the potential role of inflammatory hematological indices such as Neutrophil to Lymphocyte ratio (NLR) and CRP levels to predict responders. Methods: Blood was collected before initiation of immunotherapy from 24 patients with ES-SCLC. PBMCs were isolated with Ficoll density gradient centrifugation from patients and 1O healthy donors (HD). Immunophenotyping was performed by multi-color flow cytometry using antibodies against CD3,CD8,CD45RA,CD45RO,CCR7,PD-1, for CD8 T-cells and CD3,CD4,FoxP3,CD25,CD127,CTLA-4,CD39 for Tregs. Results: High percentages of CD39+ (p = 0.0061) and CTLA-4+ (p = 0.012) Tregs were detected in circulation of ES-SCLC compared to healthy individuals. Additionally, higher percentages of CD8+ Teff were detected in SCLC patients (p = 0.027). Patients with high numbers of Teff had significant higher PFS (p = 0.021, median 183 and 97 days) and better OS (p = 0.075, median 318 and 202 days) in response to immunotherapy compared to those with low numbers. Spearman analysis revealed a significant negative correlation between Tregs and Teff (Spearman r2 = -0.39,p = 0.043) that was accompanied with significant higher PFS (p = 0.041, median 207 and 130 days ) and OS (p = 0.029, median 318 and 202 days ) in patients with high levels of Teff and low Tregs comparing to the rest. Further analysis on inflammatory clinical signatures demonstrated that patients with better clinical response to immunotherapy have significantly lower NLR (p = 0.034) while no significant difference was revealed for CRP (p = 0.401) levels before therapy initiation. Conclusions: Combinatorial analysis of Tregs and Teff cells may be used as a predictive tool for response to immunotherapy in ES-SCLC patients.