Biological Function And Molecular Mechanism Of MicroRNA-2in Inhibiting Lung Cancer Cell Proliferation, Invasion And Migration

MicroRNAs (miRNAs) are evolutionarily conserved, endogenous, small, noncoding RNA molecules of about22nucleotides in length that are discovered in C.elegans about20years ago. MiRNAs spread wide in multicellular organisms and develop in a step-wise process through hairpin precursors cleaved by the dsRNA-specific endonucleases Drosha, Dicer and Argonaute. MiRNAs interact with target messenger RNA at specific sites to induce cleavage or inhibit translation. MiRNAs have been implicated in various important biological processes including embryogenesis, differentiation, organogenesis, cell cycle progression, apoptosis, stress response and metabolism. A number of investigations have demonstrated that altered expression of miRNAs are closely relevant to the clinical malignancies in both solid organs and hematological system, several miRNAs have been implicated in non-small cell lung carcinoma (NSCLC).In previous clinical research, we examined the miRNA expression profiles in human small cell lung cancer(SCLC), and found that downregulation of miRNA-2expression was correlated with poor survival among patients with SCLC.Compared with the patients in low miRNA-2expression group, the patients in high expression group had longer overall survival.NCI-H446, NCI-H520, A549and Anip973were transient transfected with chemically synthesized mature miRNA-2mimic oligonucleotides, introduction of miRNA-2clearly led to a significant decrease in proliferation, invasion and migration in NCI-H446A549and Anip973cell line. Analysis with TargetScan4.0indicates that there is a miRNA-2binding site in the3’UTR regions of the PLK1and TGF-β1.We also measured the mRNA and protein levels of these miRNA-2targeted genes and found that PLK1and TGF-β1expression were substantially reduced following the introduction of miRNA-2into the cells. We further investigated the suppressive role of miRNA-2in SCLC malignancy using animal model. The miRNA-2expression vector was stably transfected into NCI-H446cells and pooled clones were obtained. These pooled clones (NCI-H446-miRNA-2) expressed much higher levels of miRNA-2, exhibited slow-growing capabilities and reduced levels for PLK1and TGF-β1in both mRNA and protein expression. NCI-H446-miRNA-2and NCI-H446-control cells were implanted subcutaneously into the right upper back of nude mice, we measured the volumes of the implanted tumors weekly and found that expression of miR-886-3p significantly suppressed the growth of SCLC tumors in vivo. Interestingly, NCI-H446-miR-886-3p cells and NCI-H446-control cells differed dramatically in their invasive and metastatic potentials. In seeking the the mechanism of migration reduction in lung cancer cell lines, We found that miRNA-2expression downregulated of TGFβ1, which can inhibit epithelial mesenchymal transition in NCI-H446cells. To examine whether dysregulated expressions of miRNA-2targeted genes were linked to clinical outcome of SCLC. Totally,40SCLC specimens with known states of miRNA-2expression were analyzed for PLK1protein staining using an immunohistochemical approach. In the low miRNA-2expression group, strong cytoplasmic staining of PLK1. In contrast, among20samples from high miRNA-2expression group, all most all samples revealed weak or negative staining of PLK1. The difference of Plkl expression between two groups is statistically significant.In this report, we choose miRNA-2which was correlated with survival among patients with SCLC. As to the molecular mechanism(s) by which miRNA-2exerts its role as tumor suppressor, we have shown that miRNA-2negatively regulates PLK.1and TGF-β1, which function in promoting cell proliferation, cell migration, tumor invasive growth and metastasis. Furthermore, we demonstrated that miRNA-2may act as a tumor-suppressor to inhibit SCLC cell proliferation, migration and invasion in both cell and animal models. Therefore, it is reasonably interpreted that during the malignant development of SCLC. The better understanding of SCLC biology would substantially improve the effective therapies available for this challenging disease and promote the development of novel therapeutic agents as well. Objective To analyze the feasibility and the practicability of combining two-course treatment plans of one patient who was localized in different positions during two-course. Methods Ten patients with small cell lung cancer were chosen in the present study. According to the clinical requirement, all patients were localized twice with different positions in total course. Using the Cadplan R3.1.2treatment planning system, conformal planning was designed for each patient in two-course. Total radiation dose was50Gy.The reproductive plans can be applied to evaluate the dosimetric distribution to target volume and the surrounding organs at risk in the total course. Results1.The movements of carina in three-dimensional directions were closely correlated between two courses (P<0.05).2.No significant disparities in target volumes and normal tissue volumes were found except for the volume of PTV2. Except for maximum dose of spinal cord (r=0.337, P=0.341), positive correlations of dosimetric parameters were displayed between reproductive plans and original plans. Conclusions To assure the accuracy of isocenter’s coordinate in combined treatment plans and the positive correlations of dosimetric parameters between reproductive plans and original plans, it is feasible and the practicable to combine two-course treatment plans of one patient who was localized in different position during two-course.