The design, synthesis, and biological evaluation of novel substituted purines as HIV-1 Tat-TAR inhibitors

A series of novel substituted purines containing a side chain with a terminal amino or guanidyl group were designed and synthesized as HIV-1 Tat-TAR inhibitors. All the compounds could effectively block the TAR transactivation in human 293T cells with the CAT expression percentage ranging from 34.4% to 65.7% and showed high antiviral effects with low cytotoxicities in inhibiting the formation of SIV-induced syncytium in CEM174 cells. Molecular modeling studies by Auto-dock process suggest that the compounds bind to TAR RNA in two different modes.     

Discovery of a new HIV-1 inhibitor scaffold and synthesis of potential prodrugs of indazoles

A new oxazole scaffold showing great promise in HIV-1 inhibition has been discovered by cell-based screening of an in-house library and scaffold modification. Follow-up SAR study focusing on the 5-aryl substituent of the oxazole core has identified 4k (EC₅₀ = 0.42 μM, TI = 50) as a potent inhibitor. However, the analogues suffered from poor aqueous solubility. To address this issue, we have developed broadly applicable potential prodrugs of indazoles. Among them, N-acyloxymethyl analogue 11b displayed promising results (i.e., increased aqueous solubility and susceptibility to enzymatic hydrolysis). Further studies are warranted to fully evaluate the analogues as the potential prodrugs with improved physiochemical and PK properties     

A cell-penetrating helical peptide as a potential HIV-1 inhibitor

The capsid domain of the human immunodeficiency virus type 1 (HIV-1) Gag polyprotein is a critical determinant of virus assembly, and is therefore a potential target for developing drugs for AIDS therapy. Recently, a 12-mer α-helical peptide (CAI) was reported to disrupt immature- and mature-like capsid particle assembly in vitro; however, it failed to inhibit HIV-1 in cell culture due to its inability to penetrate cells. The same group reported the X-ray crystal structure of CAI in complex with the C-terminal domain of capsid (C-CA) at a resolution of 1.7 Å. Using this structural information, we have utilized a structure-based rational design approach to stabilize the α-helical structure of CAI and convert it to a cell-penetrating peptide (CPP). The modified peptide (NYAD-1) showed enhanced α-helicity. Experiments with laser scanning confocal microscopy indicated that NYAD-1 penetrated cells and colocalized with the Gag polyprotein during its trafficking to the plasma membrane where virus assembly takes place. NYAD-1 disrupted the assembly of both immature- and mature-like virus particles in cell-free and cell-based in vitro systems. NMR chemical shift perturbation analysis mapped the binding site of NYAD-1 to residues 169-191 of the C-terminal domain of HIV-1 capsid encompassing the hydrophobic cavity and the critical dimerization domain with an improved binding affinity over CAI. Furthermore, experimental data indicate that NYAD-1 most likely targets capsid at a post-entry stage. Most significantly, NYAD-1 inhibited a large panel of HIV-1 isolates in cell culture at low micromolar potency. Our study demonstrates how a structure-based rational design strategy can be used to convert a cell-impermeable peptide to a cell-permeable peptide that displays activity in cell-based assays without compromising its mechanism of action. This proof-of-concept cell-penetrating peptide may aid validation of capsid as an anti-HIV-1 drug target and may help in designing peptidomimetics and small molecule drugs targeted to this protein.     

Design and synthesis of novel pyrimidone analogues as HIV-1 integrase inhibitors

A series of novel pyrimidone analogues have been designed and synthesized as HIV-1 integrase (IN) inhibitors. This study demonstrated that introducing a substituent in the N1-position of the pyrimidone scaffold does not significantly influence IN inhibitory activity. Molecular docking studies showed these compounds could occupy the IN active site and form pi–pi interactions with viral DNA nucleotides DC16 and DA17 to displace reactive viral DNA 3′OH and block intasome activity.     

Synthesis and assay of isoquinoline derivatives as HIV-1 Tat-TAR interaction inhibitors

Four new isoquinoline derivatives bearing guanidinium group or amino group-terminated side chain were synthesized to target the HIV-1 TAR element. Their abilities to bind TAR RNA and inhibit Tat-TAR RNA interaction were determined by CE analysis, a Tat-dependent HIV-1 LTR-driven CAT assay and SIV-induced syncytium evaluation.     

Design and SAR of new substituted purines bearing aryl groups at N9 position as HIV-1 Tat-TAR interaction inhibitors

Twenty-four purine derivatives bearing aryl groups at N9 position were designed and synthesized as HIV-1 Tat-TAR interaction inhibitors. All the compounds showed high antiviral activities in inhibiting the formation of SIV-induced syncytium in CEM174 cells. Ten of them with low cytotoxicities were evaluated by Tat dependent HIV-1 LTR-driven CAT gene expression colorimetric enzyme assay in human 293T cells at a concentration of 30 microM, indicating effective inhibitory activities of blocking the Tat-TAR interaction. The aryl groups at N9 position affected the binding affinities between compounds and TAR RNA, showing some specificities of aryl groups to TAR RNA.     

Design and evaluation of sifuvirtide, a novel HIV-1 fusion inhibitor

Enfuvirtide (T20) is the first and only HIV-1 fusion inhibitor approved for clinical use, but it can easily induce drug resistance limiting its practical application. A novel anti-HIV peptide, termed sifuvirtide, was designed based on the three-dimensional structure of the HIV-1 gp41 fusogenic core conformation. Here we report its in vitro anti-HIV potency, its mechanism of action, as well as the results from Phase Ia clinical studies. We demonstrated that sifuvirtide inhibited HIV-1-mediated cell-cell fusion in a dose-dependent manner and exhibited high potency against infections by a wide range of primary and laboratory-adapted HIV-1 isolates from multiple genotypes with R5 or X4 phenotypes. Notably, sifuvirtide was also highly effective against T20-resistant strains. Unlike T20, sifuvirtide could efficiently block six-helix bundle formation in a dominant negative fashion. These results suggest that sifuvirtide has a different mechanism of action from that of T20. Phase Ia clinical studies of sifuvirtide (FS0101) in 60 healthy individuals demonstrated good safety, tolerability, and pharmacokinetic profiles. A single dose regimen (5, 10, 20, 30, and 40 mg) by subcutaneous injection once daily at abdominal sites was well tolerated without serious adverse events. Pharmacokinetic studies of single and multiple administration of sifuvirtide showed that its decay half-lives were 20.0 +/- 8.6 h and 26.0 +/- 7.9 h, respectively. In summary, sifuvirtide has potential to become an ideal fusion inhibitor for treatment of HIV/AIDS patients, including those with HIV-1 strains resistant to T20.     

Design and synthesis of novel dihydroquinoline-3-carboxylic acids as HIV-1 integrase inhibitors

Previously, we discovered linomide analogues as novel HIV-1 integrase (IN) inhibitors. Here, to make possible structure–activity relationships, we report on the design and synthesis of a series of substituted dihydroquinoline-3-carboxylic acids. The crystal structure of the representative compound 2c has also been solved. Among the eight new analogues, 2e showed a potency in inhibiting IN strand transfer catalytic activity similar to the reference diketo acid inhibitor L-731,988 (IC₅₀ = 0.9 μM vs. 0.54 μM, for 2e and L-731,988, respectively). Furthermore, none of the compounds showed significant cytotoxicity in two tested cancer cell lines. These compounds represent an interesting prototype of IN inhibitors, potentially involved in a metal chelating mechanism, and further optimization is warranted.     

Design and synthesis of a novel series of HIV-1 protease inhibitors.

The synthesis and the SAR study of novel pseudo symmetric inhibitors of HIV-1 protease are described. Michael addition of amino acid derivatives to vinyl ketones was utilized to derive a potent (nM) series of HIV-1 protease inhibitors.     

Design,synthesis and biological evaluation of novel pyrrole derivatives as potential ClpP1P2 inhibitor against Mycobacterium tuberculosis

In an effort to discover novel inhibitors of M. tuberculosis Caseinolytic proteases (ClpP1P2), a combination strategy of virtual high-throughput screening and in vitro assay was employed and a new pyrrole compound, 1-(2-chloro-6-fluorobenzyl)-2, 5-dimethyl-4-((phenethylamino)methyl)-1H-pyrrole-3-carboxylate was found to display inhibitory effects against H₃₇Ra with an MIC value of 77 µM. In order for discovery of more potent anti-tubercular agents that inhibit ClpP1P2 peptidase in M. tuberculosis, a series of pyrrole derivatives were designed and synthesized based on this hit compound. The synthesized compounds were evaluated for in vitro studies against ClpP1P2 peptidase and anti-tubercular activities were also evaluated. The most promising compounds 2-(4-bromophenyl)-N-((1-(2-chloro-6-fluorophenyl)-2, 5-dimethyl-1H- pyrrolyl)methyl)ethan-1-aminehydrochloride 7d, ethyl 4-(((4-bromophenethyl) amino) methyl)-2,5-dimethyl-1-phenyl-1H-pyrrole-3-carboxylate hydrochloride 13i, ethyl 1-(4-chlorophenyl)-4-(((2-fluorophenethyl)amino)methyl)-2-methyl-5-phenyl-1H-pyrrole-3-carboxylate hydrochloride 13n exhibited favorable anti-mycobacterial activity with MIC value at 5 µM against Mtb H₃₇Ra, respectively.     

Discovery of a new isomannide-based peptidomimetic synthetized by Ugi multicomponent reaction as human tissue kallikrein 1 inhibitor

Human kallikrein 1 (KLK1) is the most extensively studied member of this family and plays a major role in inflammation processes. From Ugi multicomponent reactions, isomannide-based peptidomimetic 10 and 13 where synthesized and showed low micromolar values of IC₅₀ for KLK1 The most active compound (10) presented competitive mechanism, with three structural modifications important to interact with active site residues which corroborates its KLK1 inhibition. Finally, the most active compound also showed good ADMET profile, which indicates compound 10 as a potential hit in the search for new KLK1 inhibitors with low side effects.     

Design and synthesis of N1-aryl-benzimidazoles 2-substituted as novel HIV-1 non-nucleoside reverse transcriptase inhibitors

A series of novel N₁-aryl-2-arylthioacetamido-benzimidazoles were synthesized and evaluated as inhibitors of human immunodeficiency virus type-1 (HIV-1). Some of them proved to be effective in inhibiting HIV-1 replication at submicromolar and nanomolar concentration acting as HIV-1 non-nucleoside RT inhibitors (NNRTIs), with low cytotoxicity. The preliminary structure–activity relationship (SAR) of these new derivatives was discussed and rationalized by docking studies.     

Design and synthesis of a novel BODIPY-labeled PSMA inhibitor

Prostate-specific membrane antigen (PSMA) is a zinc-bound metalloprotease which is highly expressed in metastatic prostate cancer. It has been considered an excellent target protein for prostate cancer imaging and targeted therapy because it is a membrane protein and its active site is located in the extracellular region. We successfully synthesized and evaluated a novel PSMA ligand conjugated with BODIPY_650/665. Compound 1 showed strong PSMA-inhibitory activity and selective uptake into PSMA-expressing tumors. Compound 1 has the potential to be utilized as a near infrared (NIR) optical imaging probe targeting PSMA-expressing cancers.     

Design and synthesis of photoactivatable aryl diketo acid-containing HIV-1 integrase inhibitors as potential affinity probes

Aryl diketo acids (ADKs) represent an important new class of HIV-1 integrase (IN) inhibitors. In order to facilitate examination of the structural basis underlying IN?ADK interaction, biphenyl ketone and phenyl azide photophores were incorporated into ADK structures. Of particular note is the novel dual utilization of azide and phenyketone moieties for both enzyme recognition and for crosslinking. The resulting analogues maintained low micromolar inhibitory potency against IN in recombinant in vitro assays. These potential HIV-1 integrase photoaffinity labels may provide useful tools for studying enzyme interactions of the ADK inhibitor class.     

Design and synthesis of novel HIV-1 protease inhibitors incorporating oxyindoles as the P2'-ligands

A series of novel oxyindole-derived HIV-1 protease inhibitors were designed and synthesized based upon our X-ray crystal structure of inhibitor 2 (TMC-114) bound to HIV-1 protease. The effects of substituents, spirocyclic rings, and ring sizes have been investigated. A number of inhibitors exhibited low nanomolar inhibitory potencies against HIV protease.     

Design and synthesis of potent HIV-1 protease inhibitors incorporating hydroxyprolinamides as novel P2 ligands

A series of new HIV-1 protease inhibitors with the hydroxyethylamine core and different hydroxyprolinamide P2 ligands were designed and synthesized. Variation of substitutions at the P2 significantly affected the enzyme inhibitory potency of the inhibitors. Compounds 2a and 2d showed excellent enzyme inhibitory activity with IC₅₀ values in the nanomolar range. An active site binding model for inhibitors 2a and 2d was suggested based upon the computational-docking results of the ligand with HIV-1 protease. This model offers molecular insights regarding ligand-binding site interactions of the hydroxyprolinamide-derived novel P2-ligand.