Abstract 1815: Extracellular vesicles derived from lung cancer cell may contribute to the formation of malignant pleural effusion via activating Stat3/Tissue factor/VEGF

Abstract Malignant pleural effusion (MPE) is a poor prognostic sign for patients with non-small-cell lung cancer (NSCLC) and is noted around 15% at the time of initial diagnosis and 50% in their later courses. Previously, we have shown that IL-6/Stat3/tissue factor (TF)/VEGF pathway enhances lung cancer angiogenesis, metastasis and the generation of MPE. In the current study, we demonstrated that EVs derived from lung cancer cells may also contribute MPE formation. We first used ultra-filtration (UF) method as well as the classical ultra-centrifugation (UC) method to isolate EV from lung cancer cell line and demonstrated that the EVs isolated by UF method were characteristic with exosome marker in Western Blotting (Alix, CD63, and Tsg101), morphology at Transmission electron microscopy (TEM) and size in Nanosight system (30 to 150 nm). After labeling these EVs with the green fluorescent dye Cellvue, EVs uptake in AS2 cell can be imaged by the fluorescent microscopy. After the EV uptake of recipient cell, the oncogenic signal such as Stat3 and AKT were activated with increasing phosphorylation at Western blotting comparing to those adding serum free medium. Using the ELISA method, we found the cargos in these EV were enriched in IL6. We also found these cargo derived from EVs, containing high VEGF and TF, induced more prominent extravasation than serum free medium did in Miles assays. In clinical sample, we also isolated cancer cell from MPE and the purity of the cells was confirmed by immunofluorescent staining with specific TTF-1expression of lung adenocarcinoma. After short-term culture, EV from MPE cancer cell have the characteristic of exosome marker and induced more prominent extravasation than serum free medium as EV from AS2 cell did. And we also isolated EV direct from MPE and pleural effusion from congestive heart failure (CHF) patients. And we found EV derived from MPE induced more prominent extravasation than those derived from CHF in Miles assays. In conclusion, we demonstrated the UF is clinical assessable method for isolating EV from body fluid and EV derived from lung cancer cell and MPE may contribute oncogenic and angiogenic signal transmission accelerating the formation of MPE. Citation Format: Chien-Chung Lin, Wei-Lun Huang, Wu-Chou Su. Extracellular vesicles derived from lung cancer cell may contribute to the formation of malignant pleural effusion via activating Stat3/Tissue factor/VEGF [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1815. doi:10.1158/1538-7445.AM2017-1815