Synthesis And Biological Evaluation Of Novel Substituted Dihydro-Alkylthio-Benzyl-Oxopyrimidines (S-DABOs) As Potent HIV-1 NNRTIs

Human immunodeficiency virus type 1 (HIV-1) is the main cause of the acquired immunodeficiency syndrome (AIDS), which target at the most important T4 lymphoid tissue of human immune system. Currently, AIDS has developed into a worldwide pandemic. HIV-1 is a retro virus which encodes a reverse transcriptase (RT) that is required for viral replication. RT is responsible for the conversion of the single-stranded RNA viral genome into double-stranded DNA that integrates into the host DNA.HIV-1 RT is one of the main targets for the action of anti-AIDS drugs. Two main classes of RT inhibitors have been hitherto discovered:nucleoside/nucleotide reverse transcriptase inhibitors (NRTIs/NtRTIs) and non-nucleoside reverse transcriptase inhibitors (NNRTIs). NNRTIs have gained a definitive and important place for their unique antiviral potency, low toxicity and high specificity. NNRTIs inhibit the enzyme by a non-competitively allosteric mechanism through binding to a hydrophobic pocket adjacent to the dNTP binding site of the enzyme, altering the conformation of RT. NNRTIs currently in clinical use have shown some problems, such as poor bioavailability and emergence of drug resistance. Therefore, the need for novel NNRTIs active against drug-resistant mutants selected by current therapies is of paramount importance. To date more than 50 different classes of NNRTIs have been reported, among the most representative classes, DABOs (Dihydro-alkylthio-benzyl-oxopyrimidines) occupy a relevant position.Since DABOs were disclosed as potent HIV-1 NNRTIs in 1992, much effort has been paid for the structural modifications with a view to increase potency and decrease drug resistance. Based on the Structure-Activity Relationship (SAR) of DABOs and the general principle of bioisosteric replacement, we have designed and synthesized two series of novel DABO derivatives:A,5-alkyl-2-arylthio-6-((3, 4-dihydroquinolin-1(2H)-yl) methyl) substituted S-DABOs and B, 5-alkyl-2-arylthio-6-(3-indolmethyl) substituted S-DABOs.In order to investigate the interaction of newly designed compound with RT, molecular model was established by docking method, which provides clear guideline and activity predicitions for novel HIV RT inhibitors. Based on docking results of virtual screening, some selected target molecules (A6a1-14, A6b1-14, A6c1-14 and B4d1-10) were synthesized.The key intermediateβ-ketoesters A4a-c and B2 were prepared with a simple method reported by Clay et al. through the reaction of 2-(3, 4-dihydroquinolin-1(2H)-y1) acetic acid/1H-indole-3-acetic acid with N, N'-carbonyldiimidazole (CDI) followed by treatment with different ethyl potassium malonates in the presence of anhydrous MgCl2 and Et3N. Next, the cyclization reaction ofβ-ketoesters A4a-c and B2 with thiourea in the presence of EtONa in refluxing EtOH gave the substituted uracil A5a-c and B3. Treatment of A5a-c and B3 with appropriate arylcarbonylmethyl halides or benzyl halides in the presence of K2CO3 in anhydrous DMF afforded the desirable target compounds A6a1-14, A6b1-14, A6c1-14 and B4d1-10. Both analytical and spectral data of all the compounds are in full agreement with the proposed structures.Evaluation of the antiviral activity of the compounds against HIV-1 strain IIIB and HIV-2 strain (ROD) in MT-4 cells was performed using the MTT assay. The results were expressed as EC50, CC50 and SI (selectivity, given by the CC50/EC50 ratio). This results indicated that most of the compounds of series A exhibited evident inhibition activity of HIV-1. Among the evaluated compounds, the most potent HIV-1 inhibitors were compounds 6c1,6c6 and 6b1 (EC50=0.24±0.05,0.38±0.13,0.39±0.05μM, respectively), which possess improved or similar HIV-1 inhibitory activity compared with nevirapine (NVP) (EC50=0.21μM) and delavirdine (DLV) (EC50= 0.32μM). None of the newly synthesized derivatives exhibited inhibitory activity against HIV-2. Moreover, the anti-HIV activities against some NNRTI-resistant strains of the compounds are being tested, from which we are expecting to get some useful informations.Of the newly synthesized S-DABO analogues, SAR studies have shown that:the activity of series C5-Et (A6c1-14) was obviously better than C5-Me series (A6b1-14), and, followed by C5-H series (A6a1-14), confirmed that the steric bulkiness of C5 substituent is favored to maintain inhibitory activity against HIV-1 replication; the nature of the substituents at the para substitution at the phenyl ring of the C2 moiety essentially influences the anti-HIV-1 activity. Introduction of substituted arylcarbonylmethyl groups at the C2 side chain led to compounds 6c1-7 with much better activity and selectivity index than the corresponding benzyl substituted compounds A6c8-14. Compared to the previous S-DABOs with 1-naphthylmethyl at C6 position, the new compounds showed modest antiviral activity. Based on these results and the fact that charge-transfer interactions between theπ-stacking aromatic rings at C6 position of S-DABOs and Tyr188 and Tyr181 in HIV-1 RT probably play an important role in keeping anti-HIV activity, it is suggested that the nature of the 3, 4-dihydroquinolin-1(2H)-y1)methyl group did not accommodate the chemical environment well in this region of RT.In summary, through the method of computer adied drug design, we designed and synthesized a new series of S-DABO analogues with prominent anti-HIV-1 activities, which are worth further investigation and development. This is an useful information for the development of new NNRTIs with more potent anti-HIV-1 activity and selectivity.