PO-279 Effect of the in vitro A549 neuroendocrine differentiation on the activity cytotoxic T lymphocytes

Abstract

IntroductionLung cancer is one of the leading causes of deaths by cancer in both sexes worldwide. A primary concern of this disease is the presence of neuroendocrine (NE) phenotype which has been correlated to a decreased survival, an increased number of peripheral tumour cells and an increased percentage of metastasis. A key aspect of NE phenotype is the formation of secretory granules with the fundamental property of secreting a great variety of factors such as hormones and neurotransmitters. In recent years, there has been an increasing interest in the role of this factors as immunomodulators, a primary concern is their role in the bidirectional communication within the tumour microenvironment.It remains unclear the immunomodulator effect of factors secreted in NE phenotype of lung cancer. The objective of this work was to test the effect of factors secreted by the lung adenocarcinoma cell line (A549NE) with NE phenotype on the activity of cytotoxic T lymphocytes (CTL) in vitro.Material and methodsHuman lung adenocarcinoma A549 cells were treated with [cAMP] increasing agents up to 120 hours (isobutyl-1-methylxanthine (IBMX, 0.5 mM), forskolin (FSK, 0.5 mM) or both IBMX+FSK) to observe the changes in the phenotype acquisition: morphology and neurite number (LM), Chromogranin A (CgA by FACS).Co-cultures and fluorescence release were used to evaluate the interaction of cytotoxic activity of T lymphocytes (C: Jurkat) against target cells (T: A549/GFP) and to observe the effect of the neuroendocrine differentiation on the cytotoxic activity, using as target cells original A549 cells and transdifferentiated (A549NE).Results and discussionsChanges in the A549 cell morphology were observed (size and presence of neurite-like projections), decreased proliferation rate and the overexpression of neuroendocrine marker chromogranin A were also observed in all treatments since 72 hours (except for CgA overexpression with IBMX treatment).The fluorescence release of A549/GFP after C: T co-cultures showed an increasing cytolytic time-dependent effect and C:T ratio. The NE phenotype acquisition diminishes the fluorescence release of target cells in 24 hour co-culture suggesting a decreased cytolysis.ConclusionThe current data support the generation of a neuroendocrine phenotype from A549 cell line (A549NE), stable for 48 hours after cAMP stimuli withdrawal. In co-cultures with CTL, there is evidence of diminished cytolysis by means of the acquisition of the NE phenotype.