Potent HIV-1 Protease Inhibitors Research Articles

Simple, Rapid and Validated LC Determination of Lopinavir in Rat Plasma and Its Application in Pharmacokinetic Studies

Lopinavir is a new specific and potent HIV-1 protease inhibitor. A simple and rapid Reverse Phase High-Performance Liquid Chromatographic method using UV detection was developed and validated for the analysis of lopinavir in rat plasma under isocratic conditions. The method involves a single step protein precipitation technique. The detector response was linear over the concentration range of 250 to 4000 ng mL −1. High recovery ranging from 97.5 to 101.2 percent was obtained which precludes the use of internal standard. The developed method was validated as per standard guidelines. Validation of the developed method demonstrated accuracy, precision and selectivity of the proposed method. The drug was found to be stable under various processing and storage conditions. This rapid and cost-effective method was successfully applied in the estimation of lopinavir and determination of various pharmacokinetic parameters during post intravenous bolus administration of the drug in rats. The developed method can be suitably employed in preclinical pharmacokinetic evaluation of new formulations designed to improve the bioavailability of lopinavir.

Fuzzy ARTMAP Prediction of Biological Activities for Potential HIV-1 Protease Inhibitors Using a Small Molecular Data Set

Obtaining satisfactory results with neural networks depends on the availability of large data samples. The use of small training sets generally reduces performance. Most classical Quantitative Structure-Activity Relationship (QSAR) studies for a specific enzyme system have been performed on small data sets. We focus on the neuro-fuzzy prediction of biological activities of HIV-1 protease inhibitory compounds when inferring from small training sets. We propose two computational intelligence prediction techniques which are suitable for small training sets, at the expense of some computational overhead. Both techniques are based on the FAMR model. The FAMR is a Fuzzy ARTMAP (FAM) incremental learning system used for classification and probability estimation. During the learning phase, each sample pair is assigned a relevance factor proportional to the importance of that pair. The two proposed algorithms in this paper are: 1) The GA-FAMR algorithm, which is new, consists of two stages: a) During the first stage, we use a genetic algorithm (GA) to optimize the relevances assigned to the training data. This improves the generalization capability of the FAMR. b) In the second stage, we use the optimized relevances to train the FAMR. 2) The Ordered FAMR is derived from a known algorithm. Instead of optimizing relevances, it optimizes the order of data presentation using the algorithm of Dagher et al. In our experiments, we compare these two algorithms with an algorithm not based on the FAM, the FS-GA-FNN introduced in [4], [5]. We conclude that when inferring from small training sets, both techniques are efficient, in terms of generalization capability and execution time. The computational overhead introduced is compensated by better accuracy. Finally, the proposed techniques are used to predict the biological activities of newly designed potential HIV-1 protease inhibitors.

Design and synthesis of potent HIV-1 protease inhibitors incorporating hexahydrofuropyranol-derived high affinity P(2) ligands: structure-activity studies and biological evaluation.

The design, synthesis, and evaluation of a new series of hexahydrofuropyranol-derived HIV-1 protease inhibitors are described. We have designed a stereochemically defined hexahydrofuropyranol-derived urethane as the P2-ligand. The current ligand is designed based upon the X-ray structure of 1a-bound HIV-1 protease. The synthesis of (3aS,4S,7aR)-hexahydro-2H-furo[2,3-b]pyran-4-ol, (-)-7, was carried out in optically active form. Incorporation of this ligand provided inhibitor 35a, which has shown excellent enzyme inhibitory activity and antiviral potency. Our structure-activity studies have indicated that the stereochemistry and the position of oxygens in the ligand are important to the observed potency of the inhibitor. Inhibitor 35a has maintained excellent potency against multidrug-resistant HIV-1 variants. An active site model of 35a was created based upon the X-ray structure of 1b-bound HIV-1 protease. The model offers molecular insights regarding ligand-binding site interactions of the hexahydrofuropyranol-derived novel P2-ligand.

Synthesis of Baylis–Hillman-Derived Phosphonated 3-(Benzylaminomethyl)coumarins

Treatment of Baylis–Hillman-derived 3-(chloromethyl)coumarins with benzylamine has afforded benzylamino derivatives, sequential chloroacetylation and Michaelis–Arbuzov phosphonation of which have provided access to a series of phosphonate esters as potential HIV-1 protease inhibitors.

Towards the synthesis of coumarin derivatives as potential dual-action HIV-1 protease and reverse transcriptase inhibitors

3-(Chloromethyl)coumarins, obtained via acid-catalysed cyclisation of salicylaldehyde-derived Baylis–Hillman adducts, have been treated with propargylamine; reaction of the resulting 3-alkynylmethylcoumarins with azidothymidine (AZT) in the presence of a Cu(I) catalyst has afforded a series of cycloaddition products for evaluation, in their own right, as potential dual-action HIV-1 protease and non-nucleoside reverse transcriptase inhibitors, and as scaffolds for further structural elaboration.

ChemInform Abstract: Stereoselective Synthesis of N-Boc-O-Benzyl-(4S,5S)-5-amino-4-hydroxy- 6-phenylhexanoic Acid, the Hydroxyethylene Isosteric Moiety of Potent HIV-1 Protease Inhibitor.

ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.

ChemInform Abstract: Synthesis of Novel Steroidal Inhibitors of HIV-1 Protease.

The design and synthesis of potential steroidal HIV-1 protease inhibitors is described. Compounds derived from 11-amino-12-keto-cholanic acid derivatives show modest activity.

Indolizine Studies, Part 5: Indolizine-2-carboxamides as Potential HIV-1 Protease Inhibitors[ 1

1,1-Carbonyldiimidazole-promoted coupling of Baylis–Hillman-derived indolizine-2-carboxylic acids with a range of amine and amino acid derivatives has provided access to the corresponding carboxamides in moderate to excellent yield.

ChemInform Abstract: A Conformationally Restricted β-Strand HIV Protease Inhibitor

The use of cyclopropanes as a conformationally restricted subunits in biological systems has been the subject of intense study by our group and others (1-10). Our recent efforts have focused on the use of 1, 2, 3-trisubstituted cyclopropanes as novel [-NH-Cα-] or [-CO-Cα-] bond replacements in pseudopeptides to restrict both side-chain orientation and enforce backbone secondary structures. To test these assumptions, the cyclopropane containing analog 1 (Fig. 1) was modeled after the potent HIV protease inhibitor 2, which together with a series of related derivatives was developed at Abbott Laboratories (11). This pseudopeptide contains a symmetrical diamino diol motif 8 (Fig. 2) flanked by Cbz-protected valine residues and is known to bind in a β-strand fashion at the enzyme-active site (12). Our analog 1 was designed to restrict the orientation of the valine residues and to mimic this extended backbone conformation. Comparison of enzyme inhibition constants for both compound 1 and the parent inhibitor 2 will then elucidate the efficacy of the cyclopropane as a conformationally restrictive subunit. Fig. 1. HIV protease inhibitors. Fig. 2. Synthesis of the cyclopropane containing inhibitor 1.

ChemInform Abstract: Synthesis of N-Alkyl-(α-aminoalkyl)phosphine Oxides and Phosphonic Esters as Potential HIV-Protease Inhibitors, Starting from α-Amino Acids.

ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.

Micelle-like nanoparticles of block copolymer poly(ethylene oxide)- block-poly(methacrylic acid) incorporating fluorescently substituted metallacarboranes designed as HIV protease inhibitor interaction probes

We prepared nanoparticles differing in morphology from double-hydrophilic block copolymer poly(ethylene oxide)- block-poly(methacrylic acid), PEO–PMA, and two types of fluorescein–[3-cobalt(III) bis(1,2-dicarbollide)] conjugates, GB176 and GB179, in alkaline buffer. GB176 molecule consists of fluorescein attached to the metallacarborane anion. In GB179 molecule, the fluorescein moiety connects two metallacarborane anions. The self-assembly is based on the unusual interaction of metallacarborane clusters with PEO blocks which form insoluble micellar cores. The GB176 containing nanoparticles are loose and irregular, while the GB179 ones are rigid and spherical. The structure of nanoparticles depends to some extent on a procedure of preparation. The micelles were studied by static and dynamic light scattering, fluorometry and atomic force microscopy. Since the metallacarborane conjugates act as potent inhibitors of HIV protease, the presented system is important from the point of view of drug delivery.

Molecular docking and 3D-QSAR studies of HIV-1 protease inhibitors

HIV-1 protease is an obligatory enzyme in the replication process of the HIV virus. The abundance of structural information on HIV-1PR has made the enzyme an attractive target for computer-aided drug design strategies. The daunting ability of the virus to rapidly generate resistant mutants suggests that there is an ongoing need for new HIV-1PR inhibitors with better efficacy profiles and reduced toxicity. In the present investigation, molecular modeling studies were performed on a series of 54 cyclic urea analogs with symmetric P2/P2' substituents. The binding modes of these inhibitors were determined by docking. The docking results also provided a reliable conformational superimposition scheme for the 3D-QSAR studies. To gain insight into the steric, electrostatic, hydrophobic and hydrogen-bonding properties of these molecules and their influence on the inhibitory activity, comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) were performed. Two different alignment schemes viz. receptor-based and atom-fit alignment, were used in this study to build the QSAR models. The derived 3D-QSAR models were found to be robust with statistically significant r(2) and r(2)(pred) values and have led to the identification of regions important for steric, hydrophobic and electronic interactions. The predictive ability of the models was assessed on a set of molecules that were not included in the training set. Superimposition of the 3D-contour maps generated from these models onto the active site of enzyme provided additional insight into the structural requirements of these inhibitors. The CoMFA and CoMSIA models were used to design some new inhibitors with improved binding affinity. Pharmacokinetic and toxicity predictions were also carried out for these molecules to gauge their ADME and safety profile. The computational results may open up new avenues for synthesis of potent HIV-1 protease inhibitors.

In situ click chemistry: probing the binding landscapes of biological molecules

Combinatorial approaches to the discovery of new functional molecules are well established among chemists and biologists, inspired in large measure by the modular composition of many systems and molecules in Nature. Many approaches rely on the synthesis and testing of individual members of a candidate combinatorial library, but attention has also been paid to techniques that allow the target to self-assemble its own binding agents. These fragment-based methods, grouped under the general heading of target-guided synthesis (TGS), show great promise in lead discovery applications. In this tutorial review, we review the use of the 1,3-dipolar cycloaddition reaction of organic azides and alkynes in a kinetically-controlled TGS approach, termed in situ click chemistry. The azide-alkyne reaction has several distinct advantages, most notably high chemoselectivity, very low background ligation rates, facile synthetic accessibility, and the stability and properties of the 1,2,3-triazole products. Examples of the discovery of potent inhibitors of acetylcholinesterases, carbonic anhydrase, HIV-protease, and chitinase are described, as are methods for the templated assembly of agents that bind DNA and proteins.

Pharmacokinetic study and comparative bioavailability of two nelfinavir tablet formulations in Iranian healthy volunteers after a low-dose administration

The objective of this study is to evaluate pharmacokinetic parameters, bioavailability of a potent HIV protease inhibitor, nelfinavir mesylate (NFV), following a single oral administration of 2 tablet formulations. A randomized, 2-way, crossover, bioequivalence study was conducted in 24 healthy male volunteers to compare 2 brands of nelfinavir 250 mg tablets, Nelfabiovir (Bakhtar Bioshimi, Iran) as test and Viracept (Roche, Germany) as reference product. Blood samples were collected at selected times during 12 h and plasma concentrations were determined with a sensitive and validated HPLC method involving a simple protein precipitation step. Individual pharmacokinetic parameters, t1/2, t1/2(abs), K, Ka, tmax, Cmax, Vd/F, Cl/F, AUC0-12 and AUC0- yen were determined from plasma concentration-time profiles for both formulations and were compared statistically. The analysis of variance (ANOVA) did not show any significant difference between the two formulations and 90% confidence intervals (CI) fell within the acceptable range, satisfying the bioequivalence criteria of the FDA. In vitro parameters of mean dissolution time (MDT) and time for 70% dissolution (T70) were also determined. There was no significant difference between these parameters for two dosage forms (p > 0.05). It was concluded that the two nelfinavir products are bioequivalent with respect to the rate and extent of absorption.

Expedient solid-phase synthesis of both symmetric and asymmetric diol libraries targeting aspartic proteases

C 2-symmetric diols have been shown to be highly potent against HIV-1 protease (PR). However, gaining access to these compounds has been hampered by the need of multistep solution-phase reactions which are often tedious and inefficient. In this Letter, we have disclosed a solid-phase strategy for rapid preparation of small molecule-based, symmetric and asymmetric diols as potential HIV-1 protease inhibitors. Upon biological screening, we found one of them, SYM-5, to be a potent and selective inhibitor ( K i = 400 nM) against HIV-1 protease.

Prenylisoflavonoids fromErythrina senegalensisas Novel HIV-1 Protease Inhibitors

Eight compounds were isolated from the CH (2)Cl (2) extracts of ERYTHRINA SENEGALENSIS to assess HIV-1 protease (PR) activity inhibition. The prenylated isoflavone structures, identified by spectroscopic analysis, were 8-prenylluteone ( 1), auriculatin ( 2), erysenegalensein O ( 3), erysenegalensein D ( 4), erysenegalensein N ( 5), derrone ( 6), alpinumisoflavone ( 7), and 6,8-diprenylgenistein ( 8). The constituents showed dose-dependent inhibitory activities on HIV-1 PR with IC (50) values from 0.5 to 30.0 muM. Compounds 1 - 5 possessing two hydroxy groups in the 2' and 4' positions of the B ring, potently inhibited HIV-1 PR activity. In addition, 6,8-diprenylgenistein ( 8) with two prenyl groups in the 6 and 8 positions of the A ring and one hydroxy group in the 4' position of B-ring was the most potent HIV-1 PR inhibitor.

Interpretation of Protein/Ligand Crystal Structure using QM/MM Calculations: Case of HIV-1 Protease/Metallacarborane Complex

Deltahedral metallacarborane compounds have recently been discovered as potent, specific, stable, and nontoxic inhibitors of HIV-1 protease (PR), the major target for AIDS therapy. The 2.15 A-resolution X-ray structure has exhibited a nonsymmetrical binding of the parental compound [Co(3+)-(C2B9H11)2](-) (GB-18) into PR dimer and a symmetrical arrangement in the crystal of two PR dimer complexes into a tetramer. In order to explore structural and energetic details of the inhibitor binding, quantum mechanics coupled with molecular mechanics approach was utilized. Realizing the close positioning of anionic inhibitors in the active site cavity, the possibility of an exchange of structural water molecules Wat50 and Wat128 by Na+ counterions was studied. The energy profiles for the rotation of the GB-18 molecules along their longitudinal axes in complex with PR were calculated. The results show that two Na+ counterions are present in the active site cavity and provide energetically favorable and unfavorable positions for carbon atoms within the carborane cages. Eighty-one rotamer combinations of four molecules of GB-18 bound to PR out of 4 x 10(5) are predicted to be highly populated. These results lay ground for further calculations of interaction energies between GB-18 and amino acids of PR active site and will make it possible to interpret computationally the binding of similar metallacarborane molecules to PR as well as to resistant PR variants. Moreover, this computational tool will allow the design of new, more potent metallacarborane-based HIV-1 protease inhibitors.

Impedance Method for Detecting HIV-1 Protease and Screening For Its Inhibitors Using Ferrocene−Peptide Conjugate/Au Nanoparticle/Single-Walled Carbon Nanotube Modified Electrode

A highly sensitive screening assay based on electrochemical impedance spectroscopy (EIS) has been developed for detecting HIV-1 protease (PR) and subsequent evaluation of its corresponding inhibitors at picomolar levels. The assay format was based on the immobilization of the thiol terminated ferrocene(Fc)-pepstatin conjugate on a single-walled carbon nanotube/gold nanoparticle (SWCNT/AuNP) modified gold electrode. The alteration of the interfacial properties of electrodes upon HIV-1 PR and Fc-pepstatin conjugate interaction was traced by EIS. On the basis of the charge transfer resistance data obtained and using a mixed kinetic and diffusion model, this procedure was capable of detecting picomolar HIV-1 PR owing to the specific binding of this enzyme to Fc modified pepstatin. A competitive inhibition assay format was then performed using four potent HIV-1 PR inhibitors. The estimated inhibition constant ( K i) attested that lopinavir/ritonavir ( K i = 20 +/- 3 pM) and saquinavir ( K i = 57 +/- 8 pM) even at 10 pM competed strongly with pepstatin for effective binding to HIV-1 PR. Indinavir ( K i = 630 +/- 22 pM) only competed well with pepstatin at a much higher concentration (1 nM). No significant inhibitory effect was observed for the fosamprenavir ( K i =11 +/- 0.5 nM) as expected from this pro-drug. Such results agreed well with the values reported in the literature. This assay format is a definite asset for the expedited development of effective HIV-1 PR inhibitors with low molecular weights.

3D‐QSAR Studies on Chromone Derivatives as HIV‐1 Protease Inhibitors: Application of Molecular Field Analysis

Three-dimensional quantitative structure-activity relationship (3D-QSAR) models were developed for chromone derivatives against HIV-1 protease using molecular field analysis (MFA) with genetic partial least square algorithms (G/PLS). Three different alignment methods: field fit, pharmacophore-based, and receptor-based were used to derive three MFA models. All models produced good predictive ability with high cross-validated r(2) (r(2) (cv)), conventional r(2), and predictive r(2)(r(2)(pred)) values. The receptor-based MFA showed the best statistical results with r(2) (cv) = 0.789, r(2)= 0.886, and r(2)(pred) = 0.995. The result obtained from the receptor-based model was compared with the docking simulation of the most active compound 21 in this chromone series to the binding pocket of HIV-1 protease (PDB entry 1AJX). It was shown that the MFA model related well with the binding structure of the complex and can provide guidelines to design more potent HIV-1 protease inhibitors.

A Potent HIV Protease Inhibitor, Darunavir, Does Not Inhibit ZMPSTE24 or Lead to an Accumulation of Farnesyl-prelamin A in Cells

HIV protease inhibitors (HIV-PIs) are key components of highly active antiretroviral therapy, but they have been associated with adverse side effects, including partial lipodystrophy and metabolic syndrome. We recently demonstrated that a commonly used HIV-PI, lopinavir, inhibits ZMPSTE24, thereby blocking lamin A biogenesis and leading to an accumulation of prelamin A. ZMPSTE24 deficiency in humans causes an accumulation of prelamin A and leads to lipodystrophy and other disease phenotypes. Thus, an accumulation of prelamin A in the setting of HIV-PIs represents a plausible mechanism for some drug side effects. Here we show, with metabolic labeling studies, that lopinavir leads to the accumulation of the farnesylated form of prelamin A. We also tested whether a new and chemically distinct HIV-PI, darunavir, inhibits ZMPSTE24. We found that darunavir does not inhibit the biochemical activity of ZMPSTE24, nor does it lead to an accumulation of farnesyl-prelamin A in cells. This property of darunavir is potentially attractive. However, all HIV-PIs, including darunavir, are generally administered with ritonavir, an HIV-PI that is used to block the metabolism of other HIV-PIs. Ritonavir, like lopinavir, inhibits ZMPSTE24 and leads to an accumulation of prelamin A.