Exosomes Cargo Analysis As An Approach To Identify New Biomarkers In Nsclc

Non-small cell lung cancer (NSCLC) is one of the cancer types with the highest incidence and mortality. More than 60% of patients are diagnosed in advanced stages, and tissue biopsy is unavailable in many cases. Exosomes are membranous vesicles (40-150 nm) that carry biological information to distant tissues, being able to regulate several tumor processes. The objective of this study was to analyze NSCLC exosomes cargo for searching new biomarkers that could improve NSCLC clinical management. Biomarker screening was carried out in exosomes secreted by tumor cell cultures (cell lines and primary cultures derived from resected NSCLC patients). Secreted exosomes were isolated by ultracentrifugation, and characterized through nanovesicles tracking analysis (NTA), electron microscopy and immunoblotting. Exosomal DNA mutations were determined by Digital PCR, and gene expression was carried out with Transcriptomic microarrays (p≤0.01). Expression of selected biomarkers was evaluated in (TCGA) database and assessed by RTqPCR in our resected NSCLC training cohort (paired tumor/normal tissue). Prognostic value was determined by Kaplan-Meier curves, p<0.05. Exosomes size was around 130 nm and exhibited specific surface markers (TSG101, CD9). Mutational analysis of EGFR, RAS and ALK genes showed the same pattern in exosomes and the cells of origin. Transcriptomic analysis revealed a significant differential expression between adenocarcinoma (ADC) vs squamous cell carcinoma (SCC) derived exosomes. Concretely, XAGE1B and AQP4 were overexpressed in ADC, whereas CABYR and RIOK3 were overexpressed in SCC (p<0.01). These results were confirmed in TCGA (n=706) and in our NSCLC patient cohort (n=186) (p<0.05). Furthermore, our cohort revealed an association between XAGE1B expression and prognosis in ADC group (n=74, Overall Survival: NR months vs 49.6, p=0.013). Exosomes are a source of biomarkers for the study of NSCLC, which provide valuable information of tumor characteristics, especially in cases where it is not possible to obtain a tissue sample. Supported by GV/2018/026, PI18/00266, AMACMA & AECC Valencia.