Virtual Screening And Mechanism Studying Of Novel RT(NNRTI)/IN Dual Inhibitors Against HIV From Traditional Chinese Medicine Component Database

Objectives: Currently,the multi-target drugs have become a new hot spot in the research and development of anti-HIV-1 drugs.Dual inhibitors,abbreviated RT(NNRTI)/IN,are targeting the NNRTI-binding pocket and active site of strand transfer inhibitors of integrase.In this study,the co-crystallized structures of the inhibitors and the target enzymes are unknown,we are going to find potential dual inhibitors of RT(NNRTI)/IN from TCMD and anti-HIVTCMCD and study their interaction mechanism upon molecular level through molecular simulation and virtual screening technology.Methods:1 Though literature review,dual inhibitors of HIV-1 RT(NNRTI)/IN which were chemically synthesised were summarized,and using module of MOE Creating and Editing Molecular Databases,database of dual inhibitors of HIV-1 RT(NNRTI)/IN was built.3-OH HEPT derivatives of dual inhibitors of HIV-1 RT(NNRTI)/IN were selected to be studied first,also a database was built,and some of the compounds were used to bulid a pharmacophore model.Anti-HIV traditional Chinese medicine components which had been collected before by our laboratory were input the database of anti-HIVTCMCD,the database was further sorted and improved.2 Based on the co-crystallized structure of RT and NNRTI TNK-651 which its chemical structure was similar to 3-OH HEPT derivatives of dual inhibitors of HIV-1 RT(NNRTI)/IN,and the co-crystallized structure of IN and INSTI 5Cl TEP with double Mg2+ which was simulated by our laboratory,The binding mode and interaction mechanism between inhibitors and the target enzymes were studied using molecular simulation and and graphic display function.With MOE2009.10 Dock and SYBYL8.1 Surflex-Dock,ligandswere extracted from the active pockets and then re-docked separately.By observing whether the docking conformations could reproduce the space position of the inhibitors in the complex crystal structures and the interactions between the active pockets and the ligands,we could confirm the best docking parameter settings.3 Through conformation analyzing and molecular docking,the binding mode and important residues between 3-OH HEPT derivitives of dual inhibitors of RT(NNRTI)/IN and enzymes were studied,also the reasons for the poor inhibition activity against target enzymes of some inhibitors were discussed upon molecular level.4 5 compounds of 3-OH HEPT derivitives of dual inhibitors of HIV-1RT(NNRTI)/IN were selected with the good rigidity and high activity,using conformation searching,conformation folding and pharmacophore elucidation,pharmacophore model of 3-OH HEPT derivitives of dual inhibitors of HIV-1RT(NNRTI)/IN was built through module of MOE Pharmacophore Query Editor.In order to validate the model,database of 3-OH HEPT derivatives of dual inhibitors of HIV-1 RT(NNRTI)/IN was searched,the lowest energy conformations and the optimal docking conformations of the molecules were superimposed,and the pharmacophore model and the optimal docking conformations were also superimposed.5 Based on the pharmacophore model and model of active pocket of HIV-1 NNRTI and active site of INSTI,two combination strategies were formulated with some methods of virtual screening,including similarity search,pharmacophore query,screening of Lipinski “5 rules” and molecular docking to filter TCMD and anti-HIVTCMCD to find potential dual inhibitors of HIV-1 RT(NNRTI)/IN.6 The final molecular docking scores were sorted and compounds with docking scores which were both greater than 6 docking with RT and IN were selected.Also,structure types and sources of hit compounds were analyzed.In order to reveal the material basis of traditional Chinese medicines of multi-target inhibitors against HIV-1,the interaction mechanism of hitcompounds were studied upon molecular level.Results:1 There were 42 compounds in the database of dual inhibitors of HIV-1RT(NNRTI)/IN which were chemically synthesised.Among them,HEPT derivatives were the most.What's more,pyrimidine-2,4-diones yields with3-OH which was scaffold of 3-OH HEPT derivatives had been proved as a promising scaffold for dual inhibitors of HIV-1.Database of 3-OH HEPT derivatives of dual inhibitors of HIV-1 RT(NNRTI)/IN which contained 16 compounds was built,named 3-OHHEPT.mdb.5 compounds in3-OHHEPT.mdb with good rigidity and high activity were selected to build a pharmacophore model.With further improving,there were 651 compounds which were from Forsythia(96),Astragalus mongholicus(98),Scutellaria baicalensis(80),Glycyrrhiza(258)and Honeysuckle(119)in the database of anti-HIVTCMCD.2 Using MOE-Dock,when Placement was set as Triangle Matcher,Rescoring 1 was set as London dG,Refinement was set as Forcement,Rescoring 2 was set as London dG,docking conformations of TNK-651 with RT and 5ClTEP with IN could reproduce the interactions between the active pockets and the ligands.Optimal docking conformation of TNK-651 superposed well with conformation in the co-crystallized structure,while there were a few differences on the space position of 5ClTEP?3 When TNK-651 was docked with RT,and 5ClTEP was docked with IN by Surflex-Dock,docking parameters were set as follows: Max conformations per Fragment was 20,Maximum Number of Poses per Ligand was 20,and Minimum RMSD Between Final Poses was 0.05?,the docking conformations would well simulate the interactions between TNK-651,5ClTEP and the active sites of enzymes.Among them,the optimal docking conformation of TNK-651 and RT was scored 10.60,while the optimal docking conformation of 5ClTEP and IN was scored 3.98.4 Based on the results of MOE-Dock experiments,the binding mode between 3-OH HEPT derivatives of dual inhibitors of RT(NNRTI)/IN and RTwere the same as TNK-651,but hydrogen bonds between N-3 hydroxy in the pyridine ring of most of the compounds and Lys101 were lost,which was in contradiction with the activity test results.While the docking results of SYBYL Surflex-Dock showed the binding mode with RT of 3-OH HEPT derivatives of dual inhibitors of RT(NNRTI)/IN was the same as TNK-651.There was hydrogen bond between N-3 hydroxy in the pyridine ring and Lys101,among them,N-3 hydroxy in the pyridine ring was a hydrogen bond donor.Moreover,substituted benzene in N-1 and C-6 both had interactions with active region consisted of the amino acid residues in the active pocket.The docking results were in agreement with the test results.We could forecast that the binding mode with RT of 3-OH HEPT derivatives of dual inhibitors of RT(NNRTI)/IN would be the same as docking results of SYBYL Surflex-Dock.Furthermore,compound 6,10,12,and 16 whose inhibitory activity against RT were not good lost the hydrogen bonds between Lys101 and N-3 hydroxy.5 The docking results of SYBYL Surflex-Dock showed the metal ion chelation with IN of 3-OH HEPT derivatives of dual inhibitors of RT(NNRTI)/IN was generally weak.Most of the compounds merely chelated single Mg2+,and only residue Asp64 was involved,which was in contradiction with the activity test results.While the molecular docking results based on MOE-Dock showed the binding mode of 3-OH HEPT derivatives of dual inhibitors of RT(NNRTI)/IN and IN active site was the same as 5ClTEP.Besides,2,4-carbonyl group and 3-OH in pyridine ring in most of the compounds chelated double Mg2+,2,4-carbonyl group and 3-OH in pyridine ring were Metal ligators.It was the docking results based on MOE-Dock that would accurately reflected the binding mode with IN of 3-OH HEPT derivatives of dual inhibitors of RT(NNRTI)/IN.What's more,compound 7,8and 14 which inhibitory activity against IN were not good could only chelate single Mg2+ and could not bind tightly with active pocket in IN.6 The pharmacophore model of 3-OH HEPT derivitives of RT(NNRTI)/IN with six features was built.The model contained two Metalligators,two Aromatic centers,one Hydrophobic centroid and another feature which was not only Metal ligator but also H-bond acceptor.Based on the pharmacophore model,6 compounds which matched all of the six features were hit after pharmacophore searching from 3-OHHEPT.mdb,and the detection rate was 37.5%.Matching five out of the six features,16 compounds were all hit,and the detection rate was 100%.The optimal docking conformations of the molecules were superimposed well with the lowest energy conformations and the pharmacophore model.7 After database searching from TCMD,35 molecules which were potential dual inhibitors of HIV-1 RT(NNRTI)/IN were hit.The detection rate was 0.38%,and most of the hit compounds were flavonoids.8 53 molecules which were potential dual inhibitors of RT(NNRTI)/IN were hit after database searching from anti-HIVTCMCD and the detection rate was 8.14%,and it was 21.3 times of the detection rate based on the TCMD.The hit compounds of dual inhibitors derived from Honeysuckle(21)and Forsythia(21)accounted for 79.2%.Also,most of the hit compounds were flavonoids.Conclusions:1 With MOE-Dock and SYBYL Surflex-Dock,active pocket of HIV-1NNRTI and active site of INSTI were simulated,and the docking parameter settings were confirmed.After verification,the model and the current parameter settings were relatively reliable.The models were suitable for studying the interaction mechanism between inhibitors and enzymes and for molecular docking screening.2 Docking results forecast that the binding mode and interaction mechanism between 3-OH HEPT derivatives of dual inhibitors of RT(NNRTI)/IN and RT may be the same as TNK-651.Interactions with RT were mainly hydrogen bonds,hydrophobic interactions,?-? interactions and electrostatic interactions.Amomg which,hydrogen bond between 3-OH in pyridine ring and Lys101 was important.While it were the hydrophobic interactions and chelating interactions with metal ions occupied most betweenligands and IN.We guess that through cheating double Mg2+,3-OH HEPT derivatives of dual inhibitors of RT(NNRTI)/IN would then combined with the catalytic site of IN to inhibit the activity of IN.Furthermore,the reason why inhibitory activity of compound 6,10,12,and 16 against RT was not good may be loss of hydrogen bonds between Lys101 and N-3 hydroxy.And the reason why inhibitory activity of compound 7,8 and 14 against IN was not good may chelate single Mg2+ and could not bind tightly with active pocket in IN.3-OH in pyridine ring which was not only Metal ligator but also H-bond acceptor was a common feature of both NNRTI and INSTI.Pyrimidine-2,4-diones yields was Metal ligator which was a feature of INSTI.3 The pharmacophore model of 3-OH HEPT derivitives of RT(NNRTI)/IN with six features was built.Matching five out of the six features,the detection rate was 100%,and the optimal docking conformations of the molecules were superimposed well with the lowest energy conformations and the pharmacophore model,which proved the reliability of the pharmacophore model.The model contained features which were not only unique for NNRTIs and INSTIs,but also common for NNRTIs and INSTIs.Moreover,the model accurately reflected the interactions between dual inhibitors of RT(NNRTI)/IN and enzymes.4 Based on virtual screening model,potential dual inhibitors of RT(NNRTI)/IN were hit after database searching from anti-HIVTCMCD and TCMD.Most of the hit compounds were flavonoids,which proved that flavonoids were potential dual inhibitors of RT(NNRTI)/IN.5 The hit compounds of dual inhibitors derived from Honeysuckle and Forsythia accounted for 79.2% screening from anti-HIVTCMCD.Therefore,we could draw chemical compositions containing in Honeysuckle and Forsythia were likely to find potential dual inhibitors of RT(NNRTI)/IN.6 The detection rate of searching from anti-HIVTCMCD was 21.3 times of the detection rate based on the TCMD.With the construction of anti-HIVTCMCD,the detection rate increased,and it was more advantageous to find potential dual inhibitors against HIV-1.