Design, Synthesis And Bioactivity Studies Of Novel Ligustrazine Derivatives

Cerebrocardiac Vascular Diseases (CVDs) are common diseases that seriously threaten people's health and more than half of the deaths of the world are caused by CVDs. Therefore, the research and development of cerebrocardiac vascular drugs are important strategies for the medical therapy in the world.Atherosclerosis is the pathological basic of cerebrovascular diseases and plays a very important role in its incidence. The endothelial cell damage is a common initiating process in atherosclerosis, and the following platelet aggregation eventually lead to atherosclerosis thrombosis. The thrombus blocks the vessels and consequently leads to ischemic heart diseases and cerebrovascular diseases. So, two main approaches for treatment of ischemic diseases are inhibition of platelet aggregation and to protection of the ischemic tissue.Ligustrazine is an active component of Chinese traditional medicine Chuanxiong (Ligusticum wallichii Franchat). Currently ligutrazine is an important cerebrocardiac vascular drug for the treatment of coronary atherosclerotic cardiovascular disease and ischemic cerebrocardiac vascular disease in China. However, Ligustrazine was found dissatisfactory to be applicated in clinical practice because of its moderate activity, rapid metabolism and short half life time, therefore accumulated toxicity often appeared in the patients for keeping an effective plasma concentration by the frequent administration. Thus, it is conspicuous to develop new generation of the Ligustrazine drugs for treatment of cerebrocardiac vascular diseases from molecular modification.Structure-activity relationship studies indicated that pyrazine ring of Ligustrazine might largely be the determinant of its pharmacodynamics, while the substituted groups might primarily govern its pharmacokinetics and toxicity. One of the frequently used and efficient anti-platelet aggregation agents is Ozagrel, whose structure is composed by two parts:an imidazole ring and a cinnamic acid. In accordance with the structure feature of Ozagrel, we designed and synthesized 20 compounds. We use the ligustrazine ring as a isostere of the imidazole ring and combined it with a series of hydroxy cinnamic acids/cinnamates such as Ferulic acid through ether linkage. Besides Ozagrel, there are still some commonly used anti-ischemia agents such as Piracetam, salicylic acids and coumarins. Based on the principle of combination, we combined the structure of ligustrazine with these agents and obtained 10 novel liagustrazine derivatives. By introducing structures of effective anti-platelet agents into the TMP, we hope to get improved anti-CVD activity along with good ADME character.All the newly synthesized compounds have not been reported in literature, and their chemical structures were confirmed by IR,]H NMR and ESI-MS.Activity assays:(1) Anti-platelet aggregation assay:The inhibition of platelet aggregation of the compounds was tested using a newly developed measuring method i.e. microplate-reader-nephelometry. The positive control drug is Ozagrel. The results were as following:Several compounds derived from cinnamic acid:A'2, A'3, A'4, A'5, A'6 showed higher inhibiting activity than Ozagrel (IC50=0.360 mM), with IC50 values of 0.176,0.157,0.182,0.161 and 0.054 mmol/L, respectively. Especially, compound A'6 displayed excellent activity (IC50=0.054 mmol/L),6.7 fold better than Ozagrel. Structure activity relationships were summarized as follows:(i) carboxyl group at the structure of cinnamic acid is important for keeping the anti-platelet activity, (ii)ligustrazine-cinnamic acid derivatives with one methoxy on the benzene ring of cinnamic acid exhibited high activity, while two methoxys decrease the activity, (iii) when electron withdrawing groups such as -C1,-Br at the benzene ring of the cinnamic acid, the inhibitory activities of compounds were decreased.(2) Protecting vascular endothelial cells against hyperoxic acute injury assay: These Ligustrazine derivatives have been tested for protecting vascular endothelial cells against hyperoxic acute injury. The viability of injured endothelial cells is assessed by methylthiazolyltetrazolium(MTT) assay. The biological results obtained from our experiments have demonstrated that compounds A'2, A'5, D1 present excellent cerebrocardiac vascular activities with EC50 values of 0.020,0.046 and 0.004 mM, respectively, being more active than positive control drug Ligustrazine (EC50=0.600 mM).In conclusion,30 novel Ligustrazine derivatives have been designed and synthesized. The preliminary biological results have demonstrated that some Ligustrazine derivatives were found to have better activities than positive control drug, which is significant in developing new generation of the Ligustrazine drugs for treatment of cerebrocardiac vascular diseases.