Research Of Target-based HIV-1Inhibitors And Fragment-based Methionyl-tRNA Synthetase Inhibitors

There are three chapters in this thesis:the first is HIV-1RT based drug design for S-DABOs; the second is multivalent inhibitors that target HIV gp120; the third is fragment-based drug design for human African trypanosomiasis methionyl-tRNA Synthetase.HIV-1RT based drug design for S-DABOs:Human Immunodefeciency Virus type1(HIV-1) is the etiologic agent responsible for the onset of the Acquired Immunodeficiency Syndrome (AIDS). AIDS has developed into a worldwide pandemic of disastrous proportions. HIV-1is a retrovirus which encodes a reverse transcriptase (RT) that is required for viral replication. Because of the crucial function of RT in the HIV-1life cycle, it becomes one of the important targets of antiviral therapy. Two main classes of RT inhibitors have been hitherto discovered:nucleoside/nucleotide reverse transcriptase inhibitors (NRTIs/NtRTIs) and non-nucleoside reverse transcriptase inhibitors (NNRTIs). Especially, NNRTIs have gained a definitive and important place for their unique antiviral potency, high specificity and low toxicity. Recently NNRTIs have shown problems, such as poor bioavailability, less antiviral specificity, and failure due to resistance mutations in the virus.Therefore, the need for novel NNRTIs active against drug-resistant mutants selected by current therapies is of paramount importance. To date more than50different classes of NNRTIs have been reported, among the most representative classes of them, DABOs (Dihydro-alkylthio-benzyl-oxopyrimidines) ossupy a relevant position. Owing to their structures, DABOs can adapt to changes in the binding pocket due to mutations. So DABOs can inhibit the mutation of the target and also inhibit the emergence of drug resistance.Based on computer aided drug design (CADD), we have designed and synthesized nove DABO derivatives:2-(N-phenylacetamide) substituted S-DABO analogues. We have also used molecular docking to study the relationship between compounds and HIV-1RT, which provide clear guideline and actual activity predicitions for novel HIV-1RT inhibitors. The selected target molecules based on docking results of virtual screening were synthesized.The preliminary activity and cytotoxicity screening of the newly designed and synthesized target compounds S-DABOs were tested for inhibition of HIV-1(strain ⅢB), HIV-2(strain ROD) and the selected compounds were also tested against a panel of HIV-1mutant virus strains and HIV-1RT. Most of these new congeners exhibited moderate to good activities against wild-type virus with an EC50value ranging from1.40-0.19μM. The most active S-DABO derivative was compound4b6with an EC50value of0.19±0.005μM, which are much better than those of NVP, DDC, and DDI. The selected compounds retained in part their activities against the E138K and Y181C mutant strains, with EC50values at the low micromolar level. Among the tested compounds,4d6was identified as the most active compound, along with moderate activity against mutant strains E138K (EC501.05±0.24μM with a2-fold resistance ratio with respect to wt, which are much better than that of TMC125) and Y181C (EC502.38±0.13μM with a4.5-fold resistance ratio, which are much better than that of PNU-90152T). Furthermore, enzyme inhibitory assays were performed with selected derivatives against HIV-1wtRT, confirming that the main target of these compounds is the HIV-1RT and these new S-DABO analogues are acting as NNRTIs.In summary, by the method of target-based drug design, we successfully designed and synthesized a series of S-DABO new analogues with prominent and high broad spectrum HIV-1inhibitory, which are worth further investigation and development. This is a useful information for the development of new NNRTIs with more potent anti-HIV-1activity and selectivity.The novel S-DABO derivatives were evaluated for their activity against influenza virus. Four DABO derivatives (4d2,4e2,4e3,4e4) showed potent activity against influenza virus. Compound4e4was the most promising with broad activity against influenza A and influenza B virus, which is better than the the reference compounds oseltamivir carboxylate,amantadine, and rimantadine those only have activity against influenza A (H1N1, H3N2).4e4was superior to the reference compounds ribavirin, amantadine, and rimantadine in anti-influenza A (H1N1).4e4was also superior to the reference compound ribavirin in selectivity index for influenza B virus. The antiviral mechanism of action of these DABO derivatives must be quite different from that of the currently approved anti-influenza virus drugs which target the viral M2or neuraminidase proteins. The S-DABO derivatives presented here represent a new avenue for further optimization and development of DABO compounds as novel anti-influenza virus agents.Multivalent inhibitors that target HIV gp120-SAP:We pioneered the concept of modular design of multivalent inhibitors. Optimization of individual modules of a multivalent ligand allows the final assembled multivalent inhibitors to improce potency dramatically, and therefore enhance the overall success rate of drug discovery. Multivalent inhibitor design is an attractive approach in modern drug discovery, and can ideally be applied to anti-HIV drug development. The current project is based on our previous work of template-mediated hetero-bivalent inhibitors.Targeting the key HIV viral-surface invasion protein gp120and using an abundant human serum protein SAP as the multivalent template, our work will apply computer-aided design,focused chemical synthesis, and anti-HIV assays to identify gp120inhibitors with potent activitiesFragment-based drug design for human African trypanosomiasis methionyl-tRNA synthetase inhibitors:Human African trypanosomiasis (HAT) continues to be an important public health threat in extensive regions of sub-Saharan Africa. Treatment options for infected patients are unsatisfactory due to toxicity, drug resistance, and poor efficacy of available drugs. Therefore, there is an urgent need for the design and development of high-efficiency, low toxicity, and anti-drug resistance drug that are specifically active against human African trypanosomiasis. Recently, fragment-based drug design has been developed as a very important approach for drug discovery. The methionyl-tRNA synthetases (MetRS) were selected as the drug target for human African trypanosomiasis. We use the method of fragment screening (Thermal shift) to screen more than3000small molecules in order to evaluate the interaction between small molecules and protein. The result of screening showed that2-amino-8-hydroxyquinoline could bind to methionyl-tRNA synthetases. On the bases of the structure of2-amino-8-hydroxyquinoline, we design and synthesis of a series of new compounds. A series of new2-(3-substituted-benzylamino)propylamino) quinolin-8-ols have been synthesized and evaluated for anti-T. brucei activity in cell and enzyme cultures. Most of these new congeners exhibited moderate to good activities against Trypanosoma brucei (T. brucei) with an EC50value ranging from7000-900nM. Among them,2-(3-(3-chloro-5-methoxybenzylamino)propylamino) quinolin-8-ol (4b) was one of the compounds endowed with the highest activity, with an EC50=900nM against the T. brucei. Furthermore, enzyme inhibitory assays were performed with selected derivatives against methionyl-tRNA synthetase (MetRS), confirming that the main target of these compounds is the MetRS enzyme. Pronounced shifts in melting temperatures were observed (ΔTms,9.5,8.6℃, respectively). The data indicated that the compounds were tightly bound by the recombinant MetRS enzyme. We have also used molecular docking to study the the new compounds. In summary, by the method of fragment-based drug design, we successfully designed and synthesized a series of new analogues with prominent anti-T. brucei activity, which are worth further investigation and development. The research is an exciting first step toward our goal of developing safe and effective new treatments for Human African trypanosomiasis (HAT).