Design, Synthesis And Biological Evaluation On Inhibiting The Melanoma Cell Proliferation Of Dihydropyrazole Sulfonamide Derivatives As Potential COX-1/COX-2 Inhibitors

Objective: Through studying the structure of cyclooxygenase, we have designed and synthesized a series of dihydropyrazole sulfonamide derivatives as potential COX-1/COX-2 inhibitors. And we concentrate on selecting the most effective cyclooxygenase inhibitors. After that, through observing its influence of melanoma cells’ growth and proliferation, we intend to provide the experimental evidence for future medical application of the drug in the anti-melanoma therapy.Methods: Through the computer simulations and free energy calculation, we can select much better active molecular target skeleton and re-transform their structures from the known non-steroidal anti-inflammatory drugs(such as SC-558, Celecoxib, etc.from Chart 1). Firstly, we studied the molecular structure of Celecoxib and found that the middle heterocyclic structure and sulfonamide structure are the most important structure of cyclooxygenase inhibitors. We reserved the structure of that part, and reformed its lead drug. Then, based on the structure of the lead compounds, we began to expand the structural modification and combined the structure-activity relationships. And we use computer simulation techniques to optimize the further structure. At the end, we synthesized 27 new dihydropyrazole sulfonamide derivatives, using 1H NMR and MS to identify the structure. We used X-ray to test the fostered single crystal structure of compounds 36 and 54. Next, we selected Celecoxib and Rofecoxib as the positive control drugs and tested the 27 target compounds’ selective COX-1/COX-2 inhibitory activities and antitumor proliferative activities on tumor cells(F10, MCF-7, He La). With the melanoma cell lines F10 as the object of observation, we observed the cell morphology, proliferation and apoptosis in vitro after the best drug intervention. Calculated the cell proliferation rate by MTT, a semi-quantitatively investigate the effects on proliferation. We calculated its half proliferation inhibitory concentration(IC50) and tested the 27 target compounds’ side effects on normal cell(such as kidney epithelial cells 293T) at the same time.Results: All of the synthesized compounds showed bad effects of COX-1 activity, the median inhibitory concentration IC50>59.36 μM, and some even greater than 100 μM. That is much larger than the positive control drug Celecoxib(IC50=32.6 μM). However, all were proved to be potent and selective inhibitors of COX-2, the median inhibitory concentration IC50 was 0.33 μM~32.26 μM. Compound 48 showed the most potent COX-2 inhibitory activity, which COX-2’s IC50 is equal to 0.33 μM. That was very close to the positive control drug Celecoxib(IC50=0.052 μM). This series of compounds had some anti-proliferative effect on the three cell lines, especially showed significant anti-proliferative effect on F10. Compound 35 showed the most restrained effect on F10 cell, and its median inhibitory concentration is 1.78 μM, while the positive control drug Celecoxib and Gefitinib respectively 4.56 μM and 2.20 μM. The results of cell toxicity test showed that this series of new compounds to human kidney epithelial cells’(293T) half cytotoxic concentration(CC50) was 52.64 μM ~ 83.17 μM, while the positive control drugs Gefitinib and Celecoxib were 52.03 μM and 72.42 μM, indicating that the cytotoxicity of synthesized novel dihydropyrazole sulfonamide derivatives are significantly better than the positive control drug or equivalent. Especially the compound 48, it showed the lowest cell toxicity, whith CC50 was 83.17 μM, significantly better than the positive control drug.Conclusions: The synthesized of novel dihydropyrazole sulfonamide derivatives showed well selective COX-2 inhibitory activity and low cell toxicity to normal human cell. Compound 48 is a kind of the best selective COX-2 inhibitor with the lowest cell toxicity, which may become a new type of promising and potential anti-malignant melanoma drug in the future.