Research On The Inhabitation Of Small Molecule Compounds MLN4924 Target NEDD8 NAE Of The Proliferation Of Non-small Cell Lung Cancer

Background and objectiveNEDD8 is a kind of ubiquitin-like protein is similar to the ubiquitination means, and during the physiological process of the cells, it mainly participates in such functions as stabilizing protein, regulating the substrate protein activities and transcription regulation. It has been verified by research that the inhibition of the NEDD8 pathway will be a new anti-tumor method and means. MLN4924 is a kind of effective and selective small molecule inhibitor of the NEDD8 NAE and can participate in the combination with NEDD8 NAE to form the compound of NAE-MLN4924, inhibit the activities of the NEDD8 NAE, control the ubiquitin pathways that are partially dependent on the CRL enzyme and affect the degradation of the relevant protein. It was discovered by research that NAE is expected to become a new target for the development of anti-tumor medicines, and as the inhibitor of NAE, MLN4924 features the hope of becoming the new type anti-tumor medicines.This research took the non-small cell lung cancer cells as the research model, and observations were made on the inhabitation functions of MLN4924 on cell growth, the proliferative activities of the non-small cell lung cancer cells after the interventions of MLN4924 and the expressions of the cycle distribution and cycle regulation proteins analyzed, the regulations of the relevant signal pathways related to the cell proliferations explored and the molecular mechanism of the MLN4924 inhabitations of the proliferation of non-small cell lung cancer cells determined.Experiment methodIn vitro culture of H1975 cells of human non-small cell lung cancer was conducted, and MTT experiment conducted for the analysis of the effects of the MLN4924 working cells of various concentrations on their proliferations at 24,48and 72h; cluster cloning experiment was conducted to analyze the effects of MLN4924 on the cell’s proliferation capacities; PI single dyeing method was adopted for the analyses pf the cycle distributions of cells; and Western blot experiment was conducted for the determinations of the expressions and the protein expressions of the relevant signal pathways and the phosphorylation levels of the cycle regulatory proteins CDK4, cyclin D1, CDKs natural inhibiting protein p21, p27.ResultMLN4924 presents the dosage-time dependent inhibition of the growths of non-small cell lung cancer cells, among which the H1975 cells are rather sensitive to MLN4924, and after the 48h functions of MLN4924 of various concentrations (5,10, 20,40μM), the survival rates were respectively 91.3±2.6%,86.0±4.2%,82.4±4.1%, and 79.8±3.4%, after 72h functions, the survival rate were respectively 39.3±5.7%, 39.7±4.1%,35.9±2.8%, and 36.9±2.1%. H1975 cells were treated with the 5,10,20 and 40μM of MLN4924 for 7 days, H1975 cells were significantly inhibited the formation of colony. H1975 cells were treated with the 10 and 40μM of MLN4924 for 48hours, the cell proportions of the G0/G1 phase were substantially increased, while the expressions of the cycle regulatory proteins CDK4 and cyclin D1 were obviously down-regulated, and CDKs natural inhibiting protein p21, p27 were obviously up-regulated,ConclusionMLN4924 features the function effects of inhibiting the proliferations of the non-small cell lung cancer cells, and its the molecular mechanism is through regulate expression of cell cycle related proteins and it blocks the cells in the G0/G1 phase.