Drug Design Against HIV-1 Targeting Viral Entry Process

In 1983, Barre-Sinousi et al. identified the human immunodeficiency virus (HIV-1) as the source of infection of acquired immunodeficiency syndrome (AIDS) for the first time. Since the first patient of AIDS was discovered in June 1981, this fatal disease has taken over 20 million people away. Although the application of current drugs and highly active antiretroviral therapy has achieved great successes, there still exist a number of difficulties, such as the unavailable eradication of the virus, the emergence of drug resistant virus and the strong side effects. Up till now, the medical research on vaccine also failed to break through. Therefore, it is of great urgency to design the novel drugs with safety and effectiveness against the new targets in the life cycle of HIV-1. The Fuzeon (T-20) which came into the market as an anti-HIV-1 drug in 2003 proved the entry process as potential and effective targets. Recently, it also becomes the focus of the medical research for the purpose to discover safe and effective inhibitors dealing with the entry stage of HIV-1 infection. It is well known that the discovery of new drugs is quite a costly and time consuming task. The application of computer-aided drug design (CADD) in the process of drug discovery is expected to save both money and time. In recent years, it is widely used in current drug discovery research. CADD is commonly separated into two general categories: Structure-Based Drug Design and Ligand-Based Drug Design. In this thesis, a series of research work has been done based on the ligand and receptor structures by using the CADD methods to study the entry process of the virus infection, with the aim to design the lead compounds against AIDS. The content of the thesis contain the following major aspects:(1) Study on binding mode between BMS-378806 and HIV-1 envelope protein - gp120BMS-378806 is a newly discovered small molecule that effectively blocks the binding of CD4 with gp120. The binding mode of this type of inhibitors remains unknown. AutoDock 3.0 in conjunction with molecular dynamics (MD) simulation was used to explore the binding mode between BMS-378806 and gp120. Two structures, Mode I and Mode II, with the lowest docking energy were selected as different representative binding modes. The analysis for the MD simulation data of the complex indicates that the binding of BMS-348806 with gp120 in Mode II is more stable. The average structure of Mode II was analyzed and compared with the experimental data. The sensitive of C-4 substitution on the azaindole ring may result from its occurence in the vicinity of Trp427. The Large side chain mutations inside the Phe43 cavity, such as S375W and T257R mutation, make steric hindrance for the binding. Other reisdues in the cavity such as Ser256, Thr257, Asn425, Met426 and VAL430 have considerable interactions with the small molecule. The average structure of the complex is consistent with the experimental data of mutation. The binding mode between BMS-378806 and the bounded state gp120 is similar to the binding of Phe43 in CD4 with gp120. The simulation results give an atomic insight to the possible binding mode of this kind of inhibitors.(2) Inhibitor design targeting to HIV-1 transmembrane protein - gp41(a) Pharmacophore model construction based on ligands'structuresThe software of GASP was used to generate ligand-based pharmacophore models on the basis of the identified structure information of active inhibitors interfering with the six helix bundle formation of gp41. The selected model - Model 01 was comprised of two hydrophobic features, two pairs of hydrogen bond acceptor features and one pair of hydrogen bond donor feature. The mapping of active molecules to the model showed that the hydrophobic features represented the phenyl group or pyrrole group, and the hydrogen bond features represented the hydroxyl group or carboxyl group.(b) Binding modes research and pharmacophore model construction based on receptor structuresThe binding modes of the known inhibitors with the 5-helix gp41 were obtained by using the molecule docking methods. The acceptor-based pharmacophore model was constructed according to the docking results. It is composed of four hydrophobic features, three hydrogen bond acceptor features and one hydrogen bond donor feature.(c) Pharmacophore model validation By comparison of the ligand-based pharmacophore model (Model 01), the binding modes of the inhibitors and the acceptor-based pharmacophore model, they all turned out a sort of consistency. The ligand-based pharmacophore model seemed to be a subset of the acceptor-based pharmacophore model, which is necessary for the activity. The filter property of the model was validated by a test database which was composed by molecules randomly selected from the ACDSC database and the known inhibitors. The results show that the model could filter out the active molecules efficiently with an enrichment factor of 5.04.(d) Virtual screening target to gp41A virtual screening strategy was designed to combine both the ligand and the receptor structure information. About 2,500,000 molecules from CMC, ACDSC, NCI, MDDR and CHDD were screened. The toxicity and drug-like properties were also estimated. Thirty six molecules were selected finally and two of them were synthesized. Both of the compounds showed the ability to inhibit the infection of the HIV-1 IIIB virus in vitro.(e) Structural optimization based on NB-2The de novo drug design method ( LeapFrog ) was used to optimize structure of NB-2 targeting the 5-helix structure of gp41. The docking method was adopted to recalculate the binding modes and the binding free energy of the new compounds. It was found that the binding free energy of new molecules were low than that of NB-2.The synthesis experiments are undertaken on the way.(3) Inhibitor design targeting to CCR5(a) Construction of predicable pharmacophore modelA three dimensional pharmacophore model was developed for a considerable number of pyrrolidine based and butane-based chemokine (C-C motif) receptor 5 (CCR5) antagonists. The most predictive pharmacophore model (Hypo 1), consisting of two positive ionizable points and three hydrophobic groups, had a correlation of 0.924 for the training set, and a cost difference of 63.67 bits between the null cost and the total cost. The model was applied to predict the activity of 74 compounds as a test set with a correlation factor of 0.703. The results indicated that the model was able to provide accurate activity prediction for novel antagonist design. It has been employed to filter a combinatorial database, and the molecules with good predicted activities were selected into the synthesis experiments.(b) Construction of CoMFA and CoMSIA modelA series of 1,3,4-trisubstituted pyrrolidine-based CCR5 receptor antagonists was taken as our target to construct CoMFA (Comparative Molecular Field Analysis) and CoMSIA (Comparative Molecular Similarity Indices Analysis) models. The training set and test set consisting of 72 and 39 selected molecules, respectively. For the best CoMFA model (CoMFA2), the conventional correlation coefficient r2 = 0.952, the cross-validated coeffient q2 = 0.637, and the correlation factor on the test set prediction R = 0.785, while for the best CoMSIA model (CoMSIA2), r2 = 0.958, q2 = 0.677, and R = 0.806. Furthermore the contour map also provides a visual representation for the contributions of steric, electrostatic, hydrogen-bond and hydrophobic fields. The study could provide useful information for structure optimization of this kind of compounds.In this paper, a series of research work, including binding mode prediction, pharmacophore model construction and QSAR (Quantitative structure-activity relationship) analysis has been done against the key proteins (gp120, gp41, CCR5) involved in the entry of HIV-1. A binding mode of BMS-378806 with gp120 which is consistent with the experimental data as well as effective pharmacophore models based on gp41 inhibitors and predictable QSAR models based on CCR5 antagonists were achieved. The new compounds targeting to different proteins were designed and a part of them were synthesized and proved to be active. These works may benefit the development of anti-AIDS drug candidates.