Chloroquine Molecules Research Articles

Magnetic Mesoporous Silica for Targeted Drug Delivery of Chloroquine: Synthesis, Characterization, and In Vitro Evaluation.

Malaria is a dangerous tropical disease, with high morbidity in developing countries. The responsible parasite has developed resistance to the existing drugs; therefore, new drug delivery systems are being studied to increase efficacy by targeting hemozoin, a parasite paramagnetic metabolite. Herein, magnetic mesoporous silica (magMCM) was synthesized using iron oxide particles dispersed in the silica structure for magnetically driven behavior. The X-ray diffractogram (XRD) and Mössbauer spectra show patterns corresponding to magnetite and maghemite. Furthermore, Mössbauer spectroscopy revealed superparamagnetic behavior, attributed to single magnetic domains in particles smaller than 10 nm. Even in the presence of iron oxide particles, the hexagonal structure of MCM is clearly identified in XRD (low-angle region) and the channels are visible in TEM images. The drug chloroquine (CQ) was encapsulated by incipient wetness impregnation (magMCM-CQ). The N2 adsorption-desorption isotherms show that CQ molecules were encapsulated in the pores, without completely filling the mesopores. BET surface area values were 630 m2 g-1 (magMCM) and 467 m2 g-1 (magMCM-CQ). Encapsulated CQ exhibited rapid delivery (99% in 3 h) in buffer medium and improved solubility compared to the non-encapsulated drug, attributed to CQ encapsulation in amorphous form. The biocompatibility assessment of magMCM, magMCM-CQ, and CQ against MRC5 non-tumoral lung fibroblasts using the MTT assay after 24 h revealed no toxicity associated with magMCM. On the other hand, the non-encapsulated CQ and magMCM-CQ exhibited comparable dose-response activity, indicating a similar cytotoxic effect.

Ultrasonic-Assisted Fabrication of MIL-100(Fe) Metal-Organic Frameworks as a Carrier for the Controlled Delivery of the Chloroquine Drug.

Metal-organic framework materials (MOFs) are materials with an ordered crystalline structure and high porosity that have been intensively investigated for many applications, such as gas adsorption, catalysis, sensors, drug delivery, and so on. Among them, the MOF-based drug delivery system has received increasing interest from scientists worldwide. This work presented the preparation of the MIL-100(Fe) metal-organic framework from the organic ligand of trimesic acid and iron ions with ultrasonic assistance. Scanning electron microscopy (SEM), Brunauer-Emmett-Teller surface area (BET), X-ray diffraction (XRD), infrared spectroscopy (FTIR), and Raman spectroscopy were employed to characterize the prepared MIL-100(Fe) material. MIL-100(Fe) materials synthesized by the ultrasonic method have uniform particle morphology ranging from 100 to 300 nm with a surface area of 1033 m2/g. The prepared MIL-100(Fe) was employed as a carrier for delivering chloroquine drug with a maximal loading capacity of 220 mg/g. The MIL-100(Fe)@chloroquine system was also characterized in detail. The delivery system's slow drug release was studied, showing that nearly 80% of chloroquine molecules were released after 7.5 h of immersing time in PBS and simulated gastric solutions and completely detached from the MIL-100(Fe)@chloroquine system only after approximately 80 h. This result shows the ability to control chloroquine drug release of the material, reducing the possibility of drug shock.

Surfactant-free air bubble flotation-coagulation for the rapid purification of chloroquine.

A simple and rapid separation method based on surfactant-free air bubble flotation and coagulation was designed for the purification of chloroquine (CQ) from its crude product. An open glass column having a sintered glass filter (for column chromatography) was used as a flotation vessel. The flotation was conducted by pouring the crude CQ into the aqueous solution containing 0.1% (v/v) of 2-propanol followed by feeding air through the glass filter to generate air bubbles. At pH 12, CQ was enriched into the foam temporary generating on the surface of water to form the coagulates within 90s after the start of the air bubble flotation. On the other hand, reactants; 4,7-dichloroquinoline and 4-amino-1-diethylaminopentane, as well as generated impurities remained in the bulk aqueous solution. The result of dynamic surface tension measurement indicated that CQ molecules selectively adsorbed on the air-water interface and the coagulates more strongly adsorbed the interface. Adsorption and coagulation of CQ molecules on the air-water interface were also reproduced in the calculation results of a molecular dynamic simulation. The coagulates were collected from the surface of water by suction and then poured into another flotation vessel for conducting repeated separation. The time required for the respective separation process including air bubble flotation and collection by suction was within 5min. After three-times separation, highly purified (> 99.0%) CQ was obtained with a yield of 72 ± 8%. The amounts of reactants and other impurities reduced into undetectable levels.

Effect of pH and structural orientation on supramolecular complexation of chloroquine in β-cyclodextrin medium

The supramolecular interaction of host-guest structure between Chloroquine (CQ) plus nano-hydrophobic cavity beta-cyclodextrin (β-CD) and the complex has observed by employing techniques such as UV and fluorescence spectroscopy. Here, the compound formed is of 1: 2 stoichiometric ratio. The chloroquine molecule has extremely caught into the cavity of β-cyclodextrin that is established by an in-depth analysis of spectral changes and matching to the absorption and fluorescence spectrometric interpretations. The binding constant for the inclusion complex at room temperature has evaluated by the use of Benesi-Hildebrand plot. The monocation and neutral forms of CQ with moist along with β-CD platform have discussed for the determination of pKa and pKa⁎. The thermal stability for the inclusion complex of CQ with β-CD has been examined by utilizing DSC technique. The change of the crystal arrangement of the substantial complex has substantiated by powder XRD technique. The mechanism of the inclusion complex and its orientation is done by utilizing Patch-Dock server. The cytotoxic exploration has again done to pure CQ along with its inclusion complex on MDA MB 231 cell line; then it is presented that the activity is subtle for the two materials.

Fluorescence quenching of chloroquine by Cu2+ in micelles

Abstract Fluorescence quenching, very often, has been employed as a suitable technique for characterization of the immediate environment of a fluorophore. This paper reports the absorption and fluorescence characteristics of an antimalarial drug, chloroquine, in presence of the salt copper chloride (CuCl2) in aqueous and micellar systems. Chloroquine fluorescence was quenched considerably by the copper ion (Cu2+); quenching ability of Cu2+ in micellar solution of the cationic surfactant cetyltrimethylammonium chloride (CTAC) is higher than that in water while for the nonionic surfactant polyoxyethylene (20) cetyl ether (Brij 58) it has much lower quenching efficiency. Downward deviations in Stern-Volmer (SV) plots were observed for all the systems. Possibly, the chloroquine molecules are inserted somewhat deeper inside the micelles of Brij 58 as compared to the CTAC micelles and in Brij 58 micelles the fluorophore might be less exposed to the aqueous phase to become available for the metal ion quenching.

Integrated computational investigation to develop molecular design of quinazoline scaffold as promising inhibitors of plasmodium lactate dehydrogenase

An integrated computational approach has been applied to a series of quinazoline derivatives to identify a potentially efficacious agent for malarial therapy. The best QSAR model has a good predictive correlation coefficient (r2) of 0.7908 and a significant Leave-one-out cross-validated correlation coefficient (q2) of 0.7232. Pharmacophore mapping results highlighted that 2-4- di substituted quinazoline derivative is an essential feature for the antimalarial activity. New molecules were designed based on above results and finally screened through docking tool and binding affinities of designed molecules were studied on the plasmodium lactate dehydrogenase enzyme. Docking studies revealed that carbonyl group act as a hydrogen acceptor region and 4-amino group of designed molecules act as a hydrogen donor region which facilitates hydrogen bonding with ASP-53, TYR-85 and GLU-122. Further, three complexes, i.e. chloroquine and best two docked complexes M9 and M14 were selected for molecular dynamics simulation studies. RMSD and RMSF analysis revealed that all complexes were quite stable throughout 20ns time period below 2 Å. Additionally, MM-GBSA analysis computes binding free energy suggested that ligand M14 has the highest binding energy (ΔGtotal = −34.34 kcal/mol) towards plasmodium lactate dehydrogenase as compared to a reference chloroquine molecule (ΔGtotal = −29.29 kcal/mol). In silico ADME studies revealed that the designed molecules were potential orally bioavailable drug like molecules. Thus, these studies suggest that established models have good predictive ability and designed molecules may elicit potent antimalarial activity.

Antiplasmodial Activity and In Vivo Bio-Distribution of Chloroquine Molecules Released with a 4-(4-Ethynylphenyl)-Triazole Moiety from Organometallo-Cobalamins

We have explored the possibility of using organometallic derivatives of cobalamin as a scaffold for the delivery of the same antimalarial drug to both erythro- and hepatocytes. This hybrid molecule approach, intended as a possible tool for the development of multi-stage antimalarial agents, pivots on the preparation of azide-functionalized drugs which, after coupling to the vitamin, are released with a 4-(4-ethynylphenyl)-triazole functionality. Three chloroquine and one imidazolopiperazine derivative (based on the KAF156 structure) were selected as model drugs. One hybrid chloroquine conjugate was extensively studied via fluorescent labelling for in vitro and in vivo bio-distribution studies and gave proof-of-concept for the design. It showed no toxicity in vivo (zebrafish model) as well as no hepatotoxicity, no cardiotoxicity or developmental toxicity of the embryos. All 4-(4-ethynylphenyl)-triazole derivatives of chloroquine were equally active against chloroquine-resistant (CQR) and chloroquine-sensitive (CQS) Plasmodium falciparum strains.

In silico modeling of Plasmodium falciparum chloroquine resistance transporter protein and biochemical studies suggest its key contribution to chloroquine resistance

Chloroquine (CQ) has been used for decades as the primary chemotherapeutic drug for the treatment of malaria. The emergence of drug resistance in Plasmodium falciparum has been considered to be because of the excessive use of antimalarial drugs worldwide. Moreover, the intense distribution and prevalence of chloroquine-resistant strains in endemic regions has aided the incidence of more complications to malaria treatment and control. Due to the lack of literature that portrays evident molecular mechanisms of drug resistance, it has been difficult to understand the drug resistance conferred by Plasmodium species. Intensive research on CQ drug resistance has identified the association of P. falciparum chloroquine resistance transporter protein (PfCRT), which belongs to the drug/metabolite transporter and EamA-like superfamily. Additionally, it has shown that K76 T mutation in PfCRT protein has mainly attributed to CQ resistance than other mutations. This study deals with the development of an in silico model of the PfCRT protein and its interaction with the CQ ligand molecule as well as the biochemical and biophysical characterization of the transmembrane domain 1 (TMD 1) peptide of the PfCRT protein. The physiochemical analysis of the PfCRT protein identified basic differences between the wild and mutant forms of the protein, as well as identifying the high hydrophobic nature of the mutant-type protein. The tertiary structure of the PfCRT protein was predicted and interaction with CQ revealed different active pocket binding regions in both the wild and mutant form of PfCRT proteins. The CQ2+ molecule interacts with TMD 10 of the wild-type PfCRT protein, whereas it interacts with TMD 1 of the mutant-type protein. Studies on the TMD 1 peptide revealed the insertion of the peptide in the micelles adopting stable alpha-helical structure. Binding studies with the CQ molecule detected high binding affinity toward the mutant-type TMD 1 peptide rather than the wild-type, thus confirming that the TMD 1 peptide is involved in substrate selectivity. Our findings help to characterize the structure of the PfCRT protein and the role played by the TMD 1 region in CQ resistance using in silico and biochemical approaches. Molecular docking and ligand binding studies confirm that TMD 1 is involved in substrate selectivity and aids in CQ efflux, thereby contributing to the parasite's CQ drug resistance mechanism.

Abstract 178: Blocking the Acetylcholine Activated Inward Rectifier Potassium Current but not the Rapid Delayed Rectifier Potassium Current Restores Sinus Rhythm in the Chronically Fibrillating Sheep Heart

Background: The acetylcholine activated inward rectifier potassium current (IKACh) was shown to be constitutively active in chronic atrial fibrillation. Its blockade has been proposed to be a possible antifibrillatory pharmacotherapy. We therefore tested the hypothesis that blocking IKACh with the bee venom peptide tertiapin, or the small molecule chloroquine, terminates chronic AF in the sheep heart. Methods and Results: We tested our hypothesis using biochemistry, electrophysiology and molecular modeling. In patch clamp, the IC50 of IKACh block by tertiapin, and chloroquine was 60 nM, and 710 nM respectively, while dofetilde, a currently used class III antiarrhythmic, did not block the current. On the other hand, dofetilide and chloroquine blocked the rapid delayed rectifier potassium current (IKr) with an IC50 of 50nM, and 2.3 microM respectively, while tertiapine had no effects. Furthermore, molecular modeling indicated that 1 chloroquine molecule blocks the intracellular ion permeation vestibule of the tetrameric Kir3.1, a molecular correlate of IKACh, by interacting with amino acids important for the channel’s rectification. Dofetilide did not interact with Kir3.1. In vitro fluorescence measurements of chloroquine titration into the purified Kir3.1 intracellular domain protein confirmed the computational results and indicated that chloroquine directly binds Kir3.1 with a stoichiometry of 1 chloroquine molecule per Kir3.1 tetramer. Finally, optical mapping of the chronically fibrillating sheep atria showed that while tertiapine, chloroquine, and dofetilide slowed down chronic AF and prolonged the action potential, only tertiapine and chloroquine (the IKACh blockers) restored normal sinus rhythm. Conclusions: IKACh could be a more powerful anti atrial fibrillation target compared to IKr. Pharmacological blockade of IKACh with a peptide, or a small molecule similar to chloroquine could terminate chronic AF and restore normal sinus rhythm.

A whole cell pathway screen reveals seven novel chemosensitizers to combat chloroquine resistant malaria.

Due to the widespread prevalence of resistant parasites, chloroquine (CQ) was removed from front-line antimalarial chemotherapy in the 1990s despite its initial promise of disease eradication. Since then, resistance-conferring mutations have been identified in transporters such as the PfCRT, that allow for the efflux of CQ from its primary site of action, the parasite digestive vacuole. Chemosensitizing/chemoreversing compounds interfere with the function of these transporters thereby sensitizing parasites to CQ once again. However, compounds identified thus far have disappointing in vivo efficacy and screening for alternative candidates is required to revive this strategy. In this study, we propose a simple and direct means to rapidly screen for such compounds using a fluorescent-tagged CQ molecule. When this screen was applied to a small library, seven novel chemosensitizers (octoclothepin, methiothepin, metergoline, loperamide, chlorprothixene, L-703,606 and mibefradil) were quickly elucidated, including two which showed greater potency than the classical chemosensitizers verapamil and desipramine.

Reversed Chloroquine Molecules as a Strategy to Overcome Resistance in Malaria

This short review tells the story of how Reversed Chloroquine drugs (RCQs) were developed. These are hybrid molecules, made by combining the quinoline nucleus from chloroquine (CQ) with moieties which are designed to inhibit efflux via known transporters in the membrane of the digestive vacuole of the malaria parasite. The resulting RCQ drugs can have potencies exceeding that of CQ, while at the same time having physical chemical characteristics that may make them favorable as partner drugs in combination therapies. The need for such novel antimalarial drugs will continue for the foreseeable future.

Development of ELISA-based methods to measure the anti-malarial drug chloroquine in plasma and in pharmaceutical formulations.

BackgroundIn Central and South America and Eastern and Southern Africa, Plasmodium vivax infections accounts for 71-81% and 5% of malaria cases, respectively. In these areas, chloroquine (CQ) remains the treatment of choice for P. vivax malaria. In addition, CQ has recently proven to be an effective HIV-1 therapeutic agent. There is a dire need to continue monitoring quality of CQ as there is a major influx of substandard and fake formulations into malaria-endemic countries. The use of fake/substandard drugs will result in sub-therapeutic levels endangering the patient and possibly select for parasite resistance. The aim of this study was to develop an inexpensive, simple antibody-based ELISA to measure CQ concentrations in tablets and in plasma.MethodsA monoclonal antibody (MAb) that reacts with the N-side chain of the CQ molecule was prepared by use of a CQ analogue. A specific and reliable ELISA for detection of CQ was developed. The developed assay was validated by measuring CQ in tablets sold in Denmark, India and Sudan. Furthermore, kinetics of CQ concentrations in plasma of four volunteers, who ingested two tablets of Malarex® containing, 250 mg CQ base, were measured before drug intake, three hours later and thereafter at days 1, 3, 7, 14, 21 and 28. The same plasma samples were simultaneously measured by high performance liquid chromatography (HPLC).ResultsThe ELISA proved an easy-to-handle and very sensitive tool for the detection of CQ with a lower limit of detection at 3.9 ng/ml. ELISA levels of CQ in plasma showed high agreement with the levels obtained by HPLC (r = 0.98). The specificity in the negative control group was 100%.ConclusionThe developed ELISA can be used for quality screening of CQ in pharmaceutical formulations and for drug monitoring in malaria and in other infectious diseases, such as HIV, where CQ proved to be an effective therapeutic agent. The methodology has been exploited to develop monoclonal antibodies for the drugs used in artemisinin-based combination therapy (ACT).

Progress in the development of Reversed Chloroquine molecules as antimalarial therapy

Background Chloroquine (CQ) resistance in P. falciparum is strongly linked to mutations in the gene pfcrt that gives rise to the protein, PfCRT (P. falciparum chloroquine resistance transporter), located in the parasite’s digestive vacuole (DV) membrane [1]. In CQ resistant (CQR) strains, accumulation of CQ is reduced in the DV due to increased efflux [2], relative to CQ sensitive (CQS) malaria.

Reversal Agent and Linker Variants of Reversed Chloroquines: Activities against Plasmodium falciparum

We have shown that "reversed chloroquine molecules" constructed from chloroquine-like and resistance "reversal-agent"-like cores can be powerful drugs against malaria ( J. Med. Chem. 2006 , 49 , 5623 - 5625 ). Several reversed chloroquines are now presented that probe parameters governing the activities against chloroquine-resistant and chloroquine-sensitive malaria strains. The design is tolerant to linker and reversal agent changes, but a piperazinyl group adjacent to the quinoline, at least for the group of compounds studied here, may be detrimental.

Solvent-Induced Effects: Self-Association of Positively Charged π Systems

Antimalarial cationic drugs, such as chloroquine (CQ) and ferroquine (FQ), form stable dimer structures not only in the solid state but also in solution. The short distances (3.3-3.5 A) observed between the positively charged quinolinium rings suggest that this self-association process is driven by pi/pi stacking interactions. Nevertheless, the strength of these dispersive forces is likely not sufficient to overcome the strong repulsive +/+ electrostatic effects. The question of the exact role of the environment, particularly the solvent, is clearly raised here. Characterization of these unconventional stabilizing nonbonding interactions which we have named +-pi/+-pi is therefore of great importance. In the present work, we describe theoretical calculations and NMR experiments undertaken to probe the nature and the strength of +-pi/+-pi interactions occurring upon self-association of FQ and CQ molecules in water.

The interaction of the heme-octapeptide, N-acetylmicroperoxidase-8 with antimalarial drugs: Solution studies and modeling by molecular mechanics methods

The equilibrium constants for the coordination of quinine (log K = 2.55 ± 0.02) and 9-epiquinine (log K = 2.42 ± 0.05) to form a 1:1 complex with the monomeric ferric hemeoctapeptide from cytochrome c, N-acetylmicroperoxidase-9 have been determined in aqueous solution (pH 6.25, 0.1 M phosphate buffer) at 25°C. The electronic spectra of the complexes suggest that coordination to Fe(III) occurs through the 9-alkoxide group, in agreement with previous NMR work on the interaction of quinine with urohaemin-I [I. Constantinidis and J. D. Satterlee, J. Amer. Chem. Soc. 110, 927 (1988)]. By contrast, two chloroquine molecules bind sequentially to N-acetylmicroperoxidase-8 under the same conditions (log K 1 = 3.08 ± 0.04; log K 2 = 1.56 ± 0.10) in an apparently φ − φ cofacial manner. Molecular mechanics techniques have been applied to a study of the structure of these species with an Fe(III)-porphine nucleus, and show that coordination of the 9-alkoxide group to Fe(III) is possible for both quinine and 9-epiquine, with the quinoline ring system virtually parallel to the porphyrin plane. The energy-minimized structure for the interaction of a single chloroquine molecule with Fe(III)-porphine has the quinoline rings parallel to the periphery of the porphyrin ring.

Redetermination of the crystal and molecular structure of the antimalarial chloroquine bis(dihydrogenphosphate) dihydrate.

N4-(7-Chloro-4-quinolinyl)-N',N'-diethyl-1,4-pentanediamine bis(dihydrogenphosphate) dihydrate, C18H28ClN2+(3).2H2PO-4-2H2O, Mr = 551.8, monoclinic, P2(1)/c, a = 9.830 (2), b = 16.879 (3), c = 15.783 (4) A, beta = 105.51 (2) degrees, V = 2523.2 A3, Z = 4, Dx = 1.452 g cm-3, Mo K alpha, lambda = 0.71073 A, mu = 2.78 cm-1, F(000) = 1168, room temperature, final R = 5.5% for 2431 reflections with [F0] greater than 3 sigma. The chloroquine molecule is a dication with a hydrogen atom from each of the phosphate moieties residing on the quinoline and the terminal chain nitrogen atoms. Neighboring phosphate chains are bridged by chloroquine molecules via hydrogen bonding. Each hydrogen atom on each nitrogen atom, on each phosphate oxygen atom, and in each water molecule participates in hydrogen bonding. The helical manner in which the side chains of the chloroquine molecules wrap around phosphate chains and the stacking interval of the quinoline rings between the phosphate groups may be indicative of the interaction of chloroquine molecules with cellular constituents important to antimalarial action.

A study of interactions of chloroquine with ethanol, sugars and glycerol using ultraviolet spectrophotometry

The percentage of unprotonated aromatic amino grouping (p K a 8.4) in the chloroquine molecule has been determined spectrophotometrically from measurements at 329 nm and 343 nm in suitable buffer solutions from pH 2.20 to 9.00. It is shown that ethanol, glycerol, lactose and dextrose can reduce the extent of protonation of chloroquine over this pH range, and could therefore possibly promote a more rapid absorption of chloroquine from the gastrointestinal tract. Complexation effects in the presence of these compounds cause chloroquine spectral shifts even in 0.01 M hydrochloric acid and in 0.01 M sodium hydroxide solutions. Errors can be minimised in the direct spectrophotometric assay of chloroquine by making measurements at 343 nm in 0.01 M hydrochloric acid. The ionisation of many compounds used in medicine that are weak acids or bases could be subject to effects similar to those shown by chloroquine.

An ELISA test for detecting chloroquine in urine

Two ELISA tests for detecting chloroquine in urine have been developed using polyclonal and monoclonal antibodies which react with the 7-chloro-4-aminoquinoline part of the chloroquine molecule and thus recognize chloroquine, its metabolites, and amodiaquine. The ELISAs were sensitive and specific and did not cross-react with other commonly used antimalarials. In a field trial the chloroquine ELISA performed better than the Dill Glazko or Haskins colorimetric tests. A small proportion of urines gave an apparently false positive reaction when tested at a dilution of 1:10, but not when tested at higher dilution.

Studies of the mechanism of chloroquine binding to synthetic dopa-melanin

In order to elucidate the mechanism of drugs binding to melanin, effects of pH, ionic stregth and organic solvent on the interaction of chloroquine with synthetic dopa-melanin were studied. The results indicate that electrostatic, hydrophobic and van der Waals' forces participate in the formation of the chloroquine-melanin complex. Binding analysis by the Scatchard method showed that two classes of binding sites take part in the complex formation: strong binding sites with the association constant K 1 ∼ 10 5 and weak binding sites with K 2 ∼ 10 4 Experiments with chemically modified melanin yielded some information about binding sites of this biopolymer. The obtained results suggest that strong binding involves both hydrophobic interaction and electrostatic attraction between the protonated ring system of chloroquine and the ortho-semiquinone groups of melanin. However, the weakly reacting sites can be identified as ionic bonds between protonated aliphatic nitrogen of chloroquine molecule and carboxyl groups of melanin. Van der Waals' forces occuring at the conjunctions of the aromatic rings of the drug and the aromatic indole-nuclei of the melanin probably take part in the weak binding too.