Study On N-containing Heterocyclic HIV-1 NNRTIs And Ebola Virus Entry Inhibitors

Acquired immune deficiency syndrome(AIDS)has severely threatened the health and life of human being.It has caused more than 35 million deaths since it was first reported in 1981.The most recent data from WHO shows the AIDS epidemic is still a serious public health problem and the prominent social problem worldwide.Human immunodeficiency virus(HIV)is the pathogenic factor of AIDS.The virus first destroys the human immune system,then causes opportunistic infections or cancer,and finally the death of the infected patient.As the retrovirus,there're two subtypes of HIV.Among them,HIV-1 is the main pathogen of AIDS.The complicated life cycle of HIV-1 mainly includes the attachment,fusion,reverse transcription,integration,transcription,and so on,which provides the opportunity to interrupt the replication of virus by medication.Due to the it's critical role played in the life cycle of HIV-1,reverse transcriptase(RT)has become one of the most important target for the inhibitors of HIV.And now,Non-nucleoside reverse transcriptasc inhibitors(NNRTIs)arc widely used in the clinial for their high potency,low toxicity and high selectivety.Diarylpyrimidine(DAPY)analogues was proved to be one of the most important and promising class of HIV-1 NNRTIs.As the second-generation NNRTIs,DAPYs demonstrate extremely excellent potency against both the wild-type(WT)virus and mutant strains.So far,two drugs belonging to this series TMC125(Etravirine,ETV)and TMC278(Rilpivirine,RPV),have been approved by FDA.However,the usage of these existing drugs has been compromised by the emergence of drug-resistant viral strains and imperfect pharmacokinetic properties.Over the past few years,considerable efforts have been devoted to the structural modification of DAPYs,leading to the discovery of highly potent drug candidates.The available crystal structure of DAPY/RT complex demonstrated the binding mode of these series of NNRTIs.They share a 'U' shape in the NNRTI binding pocket in RT and have four important regions for the crucial or additional interactions with the amino acid residues of the pocket.And recently,pyrrolopyrimidine derivatives RDEA427 and RDEA640 have been disclosed to possess excellent HIV antiviral activity against wild-type and a broad panel of NNRTI-resistant mutant viruses and better PK profiles.Based on these two important findings,in Chapter ? part ?,a novel class of 5,7-disubstituted pyrazolo[1,5-a]pyrimidine derivatives(Series A,18 compounds)were rationally designed,synthesized and evaluated for their anti-HIV activities in MT4 cell cultures.Biological results showed that most of the tested compounds displayed excellent activity against wild-type HIV-1 with a wide range of EC50 values from 5.98?M to 0.07 ?M.Among the active compounds,A5a was found to be the most promising analogue with an EC50 of 0.07 ?M against wild-type HIV-1 and very high selectivity index(SI,3999).Compound A5a was more effective than the reference drugs nevirapine(by 2-fold)and delavirdine(by 2-fold).But it didn't show any potency against the common K103N/Y181C mutant strain.In order to further confirm their binding target,an HIV-1 RT inhibitory assay was also performed.Furthermore,SAR analysis among the newly synthesized compounds was discussed and the binding mode of the active compound A5a was rationalized by molecular modelling studies.In our previous efforts to identify novel potent HIV-1 NNRTIs,a dozen of novel classes of NNRTIs with different cores were rationally designed,synthesized and evaluated for their anti-HIV activities.Among them,we picked the best three rings of them and two additional privileged structures as the central ring of our newly designed NNRTIs.And we also chose the cyanovinyl which presents in the structure of RPV and played an important role for the high potency.In the Chapter ? part ?,based on the understanding of crystal structure information,we combined these two optimized parts into one molecule and designed 5 subseries of a total of 21 compounds with N-containing aromatic rings(Series B).Biological results showed that 15 compounds were with EC50 against WT HIV-1 below 200 nM which are better than the first generation of marketed NNRTI drugs.Nine of them exhibited single nanomolar EC50 values,which are equally potent or more potent than the second generation NNRTIs ETR(EC50 ?B = 3.6 nM).B10a with an EC50 of 1.5 nM and SI value of 90730,was proved to be the best one against WT HIV-1 strain.Selected compounds were also tested for their activity against a panel of common clicinal resistant mutants.B4c,one of the compounds with lowest EC50 against IIIB strain,also exhibited superb EC50 of 1.8 nM and 6.8 against K103N and E138K strains respectively,which is better than that of all control drugs.Against L1001 and Y181C mutants,B4c showed the same outstanding EC50 of 10 nM.B17a is another compound with remarkable potencies against these mutants(EC50 L1001 = 11 nM,EC50 K103N = 1.6 nM,EC50 E138K = 2.8 nM,EC50 Y181C = 29 nM,EC50 Y188L = 160 nM,EC50 F227L+V106A = 84 nM.),and thanks to its low cytotoxicity it had satisfactory SI.And further mouse liver microsomal stability study showed that B17a had better T1/2(60.8 min)than RPV and similar to ETR,which indicated B17a has the potential for further development.And some more evaluations are undergoing.Ebola virus(EboV)is a highly pathogenic enveloped virus and the transmission of Ebola virus(EboV)from infected primates to humans causes outbreaks of severe and often fatal hemorrhagic fever(lethality up to 50-90%).Very recently,from 2014 to2016,the historic Ebola virus epidemic in West Africa resulted in at least 28,616 Ebolacases and 11,310 confirmed deaths which captured close attention globally.Although several vaccines entered the clinical trial and some compounds are under the research or development,there's no therapeutic method is approved by the FDA so far.Therefore,the necessity for the development of anti-EboV drugs has resulted in a wealth of antiviral research on Ebola virus.Previous study in Dr.Cunningham's lab in Harvard Medical School identified aseries of adamantane dipeptide piperazine derivatives as specific Ebola virus entry inhibitors.Among them,3.47 represents the most potent compound against EboV infection.However,the poor physicochemical properties,which resulted in unacceptable in-vitro pharmacokinetic profiles diminished its potential for clinical use.Focused on these issues,several rounds of pharmacokinetic property-driven optimization have been conducted in our lab.In Chapter ? part ?,we first systematically modified or blocked the potential metabolic sites on the right benzyl(Series C1),central dipeptide linker(Series C2)and left admantane group(Series C3)of the 3.47 structure.Three classes and totally 26 compounds were designed and synthesized.After the biological evaluation and microsomal stability tests,no compounds with retained activity showed better T1/2.Nevertheless,we have systematically extended the SAR information of 3.47 and several crucial pharmacophores were identified.The pharmacokinetic profiles of compounds are closely related to their physicochemical properties.Some well-know rules in medicinal chemistry,like Lipinski's 'Rule of Five' or Gleeson 4/400 Rule reveal that good candidate with better PK profiles usually have the molecular weight(MW)of about 400 and logP of 2-3.Besides,other parameters,like Ligand efficiency(LE),lipophilic ligand efficiency(LLE)or LELP which reflect the efficiency of the ligand utilizes its size and lipophilicity allow the evaluation of druglikeness of compounds.With the information we got from the above study,guided by the useful rules of drug development,we tried to simplify the two ends of 3.47 by multiple medicinal chemistry strategies to lower the MW and logP of 3.47,therefore to improve its PK properties.In Chapter ? part ?,6 series of compounds(86 totally,Series C4-C6)were designed and synthesized.Several promising compounds,like C7a and C7f,with retained activity and remarkably improved pharmacokinetic profiles were identified by the biological and pharmacokinetic evaluation.Further deuteration and chiral separation afforded better compounds C8p,C9g and C9n.More comprehensive PK evaluation exhibited their much more improved PK profiles than these of 3.47.And in-vivo mouse oral PK evaluation indicated the potential for C9n to serve as an orally bioavailable candidate for treatment of EBoV infection.More pharmacology and pharmacodynamics study are undergoing.In summary,in this thesis,to solve the issues of drug-resistant and unsatisfactory PK profiles of current marketed NNRTIs and to optimize the poor PK property of compound 3.47 as a lead of Ebola Virus entry inhibitors,hundreds of compounds were rationally designed and synthesized,respectively.Then through a series of biological evaluation and pharmacokinetic studies,several promising compounds were identified as good candidates for further studies.