| Flavivirus are transmitted by blood-sucking arthropods(such as ticks or mosquitoes)and are the pathogens of serious human diseases such as dengue fever,yellow fever,West Nile fever,Japanese encephalitis and Tick-borne encephalitis.More than 400 million clinical infections caused by Flavivirus are reported worldwide every year,and various epidemics caused by Flavivirus have become a serious public health safety problem faced by the international community.At present,no approved treatment can be used to treat infection caused by Flavivirus.It is extremely important to develop new and effective antiviral drugs and treatment strategies for new and recurring Flavivirus.In the face of new problems caused by the massive movement of people around the world,the loss of animal habitat and the increasing number of zoonotic diseases,as well as the increasingly spread of virus vectors due to climate change.The genome of the Flavivirus can be encoded a polyprotein precursor that is cut into three structural proteins and seven non-structural proteins under the action of viruses and host cell enzymes,among them,the structural protein E and non-structural NS1、NS3 and NS5 are the most likely targets for antiviral drugs.NS5 protein has the maximum molecular weight and is the most conserved protein in Flavivirus,which has high homology: The homology of NS5 between Zika virus and West Nile virus,Japanese encephal virus,Dengue virus are 72%,70%,and 68%,respectively.The C-terminal of NS5 has RNA-dependent RNA Polymerase(Rd Rp)structure,which has the natural advantage as an antiviral inhibitor target: Rd Rp is highly conserved in Flavivirus;It is an essential enzyme for the viral replication cycle;In addition,due to host cells lack Rd Rp,drugs targeting Rd Rp have less inhibitory effect on host cells.Therefore,Rd Rp Inhibitors may have broad-spectrum and low-toxicity antiviral effects.The inhibitors that act on Rd Rp include nucleoside/tide analogues and non-nucleoside/tide analogues,among them,the triphosphate active metabolites of nucleoside/tide analogues can compete with natural nucleotides to bind to the catalytic active site of Rd Rp,block viral RNA replication and transcription by competing with natural nucleotides to bind the active sites of the enzyme to play an antiviral effect,thus becoming attractive candidate drugs for the treatment of diseases caused by various RNA virus infections.At present,most of the nucleoside inhibitors reported in the literature have been proved to be active in vivo and in vitro and have entered clinical trials.Although most nucleoside analogues have failed in the late stage of clinical development,the successful marketing of Sofosbuvir and Favipiravir still proves the effective of the Rd Rp.In the process of searching for new drugs for the treatment of Chronic hepatitis C,a ribonucleoside analogue 2’-C-methylcytidine(NM107)was found.NM107 was also the first anti-RNA inhibitor found to act through the mode of nucleoside analogues.In view of the reported studies of various nucleoside inhibitors,NM107 was selected as the lead compoud to carry out follow-up research.Many literature have reported that NM107 has good antiviral activity against a variety of Flavivirus in vivo and in vitro,indicating that NM107 has a broad spectrum of anti-Flavivirus activity,but the oral bio-availability of NM107 is only 11%.In view of this deficiency,it is reported that NM107 was modified to obtain 3’-OH amino acid ester prodrugs and 5’-OH phosphoramide prodrugs.Two kinds of esterified prodrugs can both improve the apparent oral bio-availability of NM107.In addition,the antiviral activity of various esterified prodrugs reported in the literature is basically the same as that of NM107 or even better than that of NM107,indicating that the structural modification of NM107 can not only improve the bio-availability of the compounds through the prodrug principle,but also change the antiviral activity of the compounds by changing the membrane permeability of the compounds.Therefore,In this paper,we modified the5’-OH ester of NM107 and explored its effect on the medicinal effect of NM107.First of all,a new synthetic route for the lead compound NM107 was designed with several key steps were optimized and the total yield of the nine-step reaction was11%.In the new synthetic route,when 2-position secondary alcohol was oxidized to ketone,the oxidation system of TEMPO/TCCA was used to react at room temperature,and the post-treatment only needed to remove the by-products,compared with the low temperature of-78 ℃ and high toxicity of the reaction reagents used in the Swern oxidation system reported in the literature,the reaction condition was mild and the post-treatment was simple.In the step of methylation with Grignard reagent,The yield of chiral configuration methylation product was increased from 11% to 46%,thus the overall yield of the target compound was increased from 2% to 11%.In the new route,several reaction routes to get intermediates were mild;in addition,the raw materials used were cheap with high economic benefit.finally,we confirmed the configuration of the compound prepared by the new route,which was same with the target compound NM107,and its purity was more than 99.5%.Therefore,the designed new route is suitable for the derivation of new compounds and may also become a potential industrial production route.Secondly,on the basis of the constant preparation of NM107,three synthetic routes of new compounds were designed and implemented,and 14 5’-OH esterification products of NM107 were synthesized with the structures of which were confirmed,among them,there were 9 chain esters,2 cyclic esters and 3 benzoyl esters.The evaluation results of antiviral activity in vitro of three compounds were obtained.From the existing biological activity results,the inhibitory activities of the three new compounds to ZIKV were similar to that of NM107,compound ZXX-5-5 had higher activity to DENV than NM107,but less activity to JEV,and the activities of long chain compounds ZXX-5-15 and ZXX-5-41 to JEV were similar to that of NM107,but the activities to YFV were poor.The analysis of the anti-Flavivirus activity of the NM107 esterified modified compounds reported in the literature and the evaluation of the activities of the compounds we obtained showed that the esterified modification of the hydroxyl groups at different positions of NM107 could not only improved the bio-availability of NM107 through prodrug principle,but also enhanced the antiviral activity of the compounds by increasing the permeability of the membrane and changing the efficacy of the compounds.Since the evaluation results of antiviral activity of only three compounds were obtained,the antiviral effect of the compounds could not be accurately evaluated,the evaluation of the activity of the compounds and the study of drug metabolism in vivo would be completed in order to complete the overall evaluation of the efficacy of the compounds. |
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