Antimalarial activity of thioacridone compounds related to the acronycine alkaloid

A series of thioacridone compounds that were previously shown to have DNA binding interaction, were screened for antimalarial activity. The new compounds were assessed for in vitro antimalarial activity against a chloroquine sensitive (D10) strain of the malaria parasite Plasmodium falciparum, using a lactate dehydrogenase (PfLDH) assay. In the series, the IC(50) values ranged from 0.4 to 27 microg/ml. 1-(2-Dimethylaminoethylamino)-9(10H)-thioacridone was found to be the most potent against P. falciparum (D10) with an IC(50) value of 0.4 microg/ml. This compound was also evaluated against a South African chloroquine resistant (RSA 11) P. falciparum strain and was found to have an IC(50) value of 1 microg/ml, compared with 0.16 microg/ml for chloroquine. Quantitative structure-activity relationships of this series were also investigated and a multiple linear regression r(2) of 0.58 was found for the best fit equation. The most potent compound, 1-(2-dimethylaminoethylamino)-9(10H)-thioacridone, was docked into the chloroquine binding site of PfLDH and it was found that the slightly lower activity of this compound, compared with chloroquine, is likely due to steric interference within a restricted binding pocket.     

Synthesis and bioevaluation of hybrid 4-aminoquinoline triazines as a new class of antimalarial agents

The emergence and rapid spread of chloroquine resistant strains of Plasmodium falciparum has dramatically reduced the chemotherapeutic options. Towards this goal, a series of new class of hybrid 4-aminoquinoline triazines were synthesized and screened against CQ sensitive strain 3D7 of P. falciparum in an in vitro model. Compounds 65 and 69 exhibited more than 99% suppression on day 4 and on day 6 post treatment, compound 69 showed impressive 99.11% suppression against CQ resistant strain N-67 of P. yoelii in an in vivo assay.     

Novel oxazine skeletons as potential antiplasmodial active ingredients: Synthesis, in vitro and in vivo biology of some oxazine entities produced via cyclization of novel chalcone intermediates

A novel series of 6-(2-chloroquinolin-3-yl)-4-substituted-phenyl-6H-1,3-oxazin-2-amines were synthesized and evaluated for in vitro antimalarial efficacy against chloroquine sensitive (MRC-02) as well as chloroquine resistant (RKL9) strains of Plasmodium falciparum. The activity tested was at nanomolar concentration. β-Hematin formation inhibition activity (BHIA(50)) of oxazines were determined and correlated with antimalarial activity. A reasonably good correlation (r?=?0.49 and 0.51, respectively) was observed between antimalarial activity (IC(50)) and BHIA(50). This suggests that antimalarial mode of action of these compounds seems to be similar to that of chloroquine and involves the inhibition of hemozoin formation. Some of the compounds were showing better antimalarial activity than chloroquine against resistant strain of P. falciparum and were also found to be active in the in vivo experiment.     

Synthesis and evaluation of phenylequine for antimalarial activity in vitro and in vivo

Synthesis of the potent antiplasmodial 4-aminoquinoline, phenylequine (PQ), is reported for the first time. PQ and the two analogues show increased efficacy in moving from the chloroquine sensitive D10 to the chloroquine resistant K1 strain in vitro. The in vivo efficacy of PQ, and salts thereof, have been determined in Plasmodium berghei ANKA and Plasmodium yoelii. Phenylequine hydrochloride has shown an ED₅₀ of 0.81 in P. yoelii (cf chloroquine ED₅₀ = 1.31).     

Synthesis and antimalarial activity of new chloroquine analogues carrying a multifunctional linear side chain

We report the synthesis and in vitro antimalarial activity of several new 4-amino- and 4-alkoxy-7-chloroquinolines carrying a linear dibasic side chain. Many of these chloroquine analogues have submicromolar antimalarial activity versus HB3 (chloroquine sensitive) and Dd2 (chloroquine resistant strain of Plasmodium falciparum) and low resistance indices were obtained in most cases. Importantly, compounds 11–15 and 24 proved to be more potent against Dd2 than chloroquine. Branching of the side chain structure proved detrimental to the activity against the CQR strain.     

Identification of distinct accumulation sites of 4-aminoquinoline in chloroquine sensitive and resistant Plasmodium berghei strains

We report the synthesis of an analogue of chloroquine (CQA) which can be used as a probe to visualize accumulation of 4-aminoquinoline by electron microscopy. A mouse monoclonal antibody against CQA was raised and used for immunodetection by the protein-A gold method on ultrathin cryosections, of CQA treated parasites. We demonstrate that in a P. berghei chloroquine(CQ)-sensitive strain (N strain) the chloroquine analogue used accumulates in the endocytic vacuoles where hemoglobin (Hb) degradation is occurring. In contrast, in a P. berghei CQ-resistant strain (RC strain) the probe was found scattered all over the cytoplasm of the parasite. This result suggests that endocytic vacuoles of the parasite could constitute the site of antimalarial action of CQ.     

Effect of chloroquine phosphate and toxic concentrations of lead acetate on Ca2+-ATPase activity in isolates and clones of Plasmodium Falciparum

The basal activity of Ca2+-ATPase in two isolates (NL56, UNC) and two clones (D6, W2) of P.falciparum was assessed. The effects of various concentrations of chloroquine phosphate and toxic concentrations of lead acetate were also evaluated in the clones and strains of P.falciparum. The Ca2+-ATPase activity was measured by monitoring the rate of release of inorganic phosphate from the gamma-position of ATP on spectrophotometer at 820nm wavelength. The various concentrations of chloroquine (3, 6, 9, 12, 18μg/ml) and lead acetate (5, 10, 20, 30, 40μg/ml) on Ca2+-ATPase activity were measured respectively. Chloroquine phosphate inhibited Ca2+-ATPase activity in both the isolates and the cloned strains of P.falciparum in concentration dependent manner. Median Inhibitory concentration of chloroquine (MIC50) estimated from the plot of activity against chloroquine concentration was found to be 2.6mg/ml at pH 7.4 for both the isolates and cloned strains examined. Lead acetate at concentrations 5-20μg/ml inhibited Ca2+-ATPase activity in concentration dependent manner in clone W2 (Chloroquine resistant strain) while the same range of concentrations of lead acetate stimulated the activity of the enzyme in clone D6 (Chloroquine sensitive strain).The inhibitory effect of lead acetate on the enzyme in clone D6 was observed at concentrations above 20μg/ml. The result also suggests that lead ions could modulate and moderate calcium ion homeostasis in P. falciparum via its effect on Ca2+-ATPase activity. Also sufficient influx of lead ions into P. falciparum may transform the biochemical or bioenergetics nature of chloroquine sensitive strain of P. falciparum (D6) to that similar to chloroquine resistant strain (W2). In conclusion, inhibition of Ca2+-ATPase activity of P.falciparum may be part of the mechanism of action of chloroquine in its use as chemotherapy for malaria. The study implies that populations simultaneously exposed to lead pollution and malaria infection may experience failure in chloroquine therapy.     

Antiplasmodial activity of novel keto-enamine chalcone-chloroquine based hybrid pharmacophores

A series of novel keto-enamine chalcone-chloroquine based hybrids were synthesized following new methodology developed in our laboratory. The synthesized compounds were screened against chloroquine sensitive strain (3D7) of Plasmodium falciparum in an in vitro model. Some of the compounds were showing comparable antimalarial activity at par with chloroquine. Compounds with significant in vitro antimalarial activity were then evaluated for their in vivo efficacy in Swiss mice against Plasmodium yoelii (chloroquine resistant N-67 strain), wherein compounds 25 and 27 each showed an in vivo suppression of 99.9% parasitaemia on day 4. Biochemical studies reveal that inhibition of hemozoin formation is the primary mechanism of action of these analogues.     

Selective antimalarial activity of tetrandrine against chloroquine resistant Plasmodium falciparum

Antimalarial activity of tetrandrine was studied using a continuous in vitro culture of Plasmodium falciparum. Experimental results showed that tetrandrine has potent antimalarial effect on both chloroquine sensitive and resistant strains of Plasmodium falciparum. Interestingly, tetrandrine is about three times more potent against the chloroquine resistant strain than it is against the sensitive strain based on their IC50 values, which were 5.09 x 10(-7) M for the sensitive strain and 1.51 x 10(-7) M for the resistant strain. In addition, reversal experiments revealed that tetrandrine cannot reverse chloroquine-resistance, although it has verapamil-like, calcium-channel-blocker activity.     

The susceptibility of the Indonesian I/CDC strain of Plasmodium vivax to chloroquine.

A strain of Plasmodium vivax from Indonesia was adapted to splenectomized Aotus and Saimiri monkeys and tested for its susceptibility to chloroquine. Animals were infected by intravenous inoculation of heparinized parasitized blood and subsequently treated with 8 or 15 mg (base) of chloroquine by oral intubation. Recrudescence of infection occurred in 4 of 4 Aotus and 5 of 6 Saimiri monkeys treated with 15 mg base of chloroquine, indicating a level of resistance between that of the standard Chesson strain of P. vivax and the recently reported resistant strains from Papua New Guinea.     

Antimalarial drug susceptibility testing of Plasmodium falciparum in Brazil using a radioisotope method

From March 1996 to August 1997, a study was carried out in a malaria endemic area of the Brazilian Amazon region. In vivo sensitivity evaluation to antimalarial drugs was performed in 129 patients. Blood samples (0.5 ml) were drawn from each patient and cryopreserved to proceed to in vitro studies. In vitro sensitivity evaluation performed using a radioisotope method was carried out with the cryopreserved samples from September to December 1997. Thirty-one samples were tested for chloroquine, mefloquine, halofantrine, quinine, arteether and atovaquone. Resistance was evidenced in 96.6% (29/30) of the samples tested for chloroquine, 3. 3% (1/30) for quinine, none (0/30) for mefloquine and none for halofantrine (0/30). Overall low sensitivity was evidenced in 10% of the samples tested for quinine, 22.5% tested for halofantrine and in 20% tested for mefloquine. Means of IC 50 values were 132.2 (SD: 46. 5) ng/ml for chloroquine, 130.6 (SD: 49.6) ng/ml for quinine, 3.4 (SD: 1.3) ng/ml for mefloquine, 0.7 (SD: 0.3) ng/ml for halofantrine, 1 (SD: 0.6) ng/ml for arteether and 0.4 (SD: 0.2) ng/ml for atovaquone. Means of chloroquine IC 50 of the tested samples were comparable to that of the chloroquine-resistant strain W2 (137.57 ng/ml) and nearly nine times higher than that of the chloroquine-sensitive strain D6 (15.09 ng/ml). Means of quinine IC 50 of the tested samples were 1.7 times higher than that of the low sensitivity strain W2 (74.84 ng/ml) and nearly five times higher than that of the quinine-sensitive strain D6 (27.53 ng/ml). These results disclose in vitro high resistance levels to chloroquine, low sensitivity to quinine and evidence of decreasing sensitivity to mefloquine and halofantrine in the area under evaluation.     

Design, synthesis and antimalarial activity of a glyoxylylhydrazone library

Synthesis of a new family of quinolylhydrazone derivatives and evaluation of their activity against a chloroquine-resistant strain of Plasmodium falciparum are described. The best compound displayed an activity 6-fold higher than chloroquine. None of the active compounds were found to inhibit beta-hematin formation in vitro in the same range as chloroquine and five among them displayed lower calculated vacuolar accumulation ratios, suggesting the implication of a different mechanism of action.     

Synthesis of acridine derivatives of amino acid hydrazines and their antimalarial activity

2-Methoxy-6-chloroacridine-9-yl- and 2-ethoxy-6-nitroacridine-9-yl-hydrazides of glycine, alpha- and beta-alanines, gamma-aminobutiric acid, epsilon-aminocaproic acid have been synthesized and their antimalarial activity has been investigated. The compounds were found to inhibit the growth of malaria parasite P. falciparum in in vitro cultures. Fifty per cent inhibitory concentrations ranged from 2 x 10(-7) to 6 x 10(-7) M and corresponded to therapeutic concentrations of known quinoline and acridine antimalarial drugs. The beta-alanine and gamma-aminobutiric acid derivatives were the most active and showed high activity against a chloroquine resistant strain of P. falciparum.     

Potentiation of antimalarial drug action by chlorpheniramine against multidrug-resistant Plasmodium falciparum in vitro

Chlorpheniramine, a histamine H1 receptor antagonist, was assayed for in vitro antimalarial activity against multidrug-resistant Plasmodium falciparum K1 strain and chloroquine-resistant P. falciparum T9/94 clone, by measuring the 3H-hypoxanthine incorporation. Chlorphenirame inhibited P. falciparum K1 and T9/94 growth with IC50 values of 136.0+/-40.2 microM and 102.0+/-22.6 microM respectively. A combination of antimalarial drug and chlorpheniramine was tested against resistant P. falciparum in vitro. Isobologram analysis showed that chlorpheniramine exerts marked synergistic action on chloroquine against P. falciparum K1 and T9/94. Chlorpheniramine also potentiated antimalarial action of mefloquine, quinine or pyronaridine against both of the resistant strains of P. falciparum. However, chlorpheniramine antagonism with artesunate was obtained in both P. falciparum K1 and T9/94. The results in this study indicate that antihistaminic drugs may be promising candidates for potentiating antimalarial drug action against drug resistant malarial parasites.     

Effect of artemisinin (qinghaosu) and chloroquine on drug-sensitive and drug-resistant strains of Plasmodium falciparum malaria: use of [2,8-3H]adenosine as an alternative to [G-3H]hypoxanthine in the assessment of in vitro antimalarial activity

Using [G-3H]hypoxanthine uptake as a radioactive indicator for the growth of malarial parasites, we measured the antimalarial activity of artemisinin (Qinghaosu, QHS) against FCMSU1/Sudan strain (chloroquine-sensitive strain) and FCB K+ strain (chloroquine-resistant strain) of Plasmodium falciparum in continuous culture in vitro. The 50% inhibitory concentrations (IC50) for QHS against FCMSU1/Sudan strain and FCB K+ strain were 2.8 X 10(-8) and 3.0 X 10(-8) M, respectively. On the contrary, the response of the two strains to chloroquine was quite different. The IC50 for chloroquine against FCMSU1/Sudan strain was 5.6 ng/ml, whereas that for the FCB K+ strain was 65.6 ng/ml. Therefore, QHS did not appear to exhibit any cross-resistance with chloroquine. If [2,8-3H]adenosine was used as a radioactive precursor instead of [G-3H]hypoxanthine for the determination of antimalarial activity, virtually identical results were obtained. Therefore, [2,8-3H]adenosine can be used as an alternative to [G-3H]hypoxanthine for the assessment of antimalarial action.     

Variation of incubation time in an in vitro drug susceptibility test of Plasmodium falciparum isolates studied in the Solomon Islands

A study on chloroquine resistance of falciparum malaria was conducted in the Solomon Islands. Both in vitro and clinical tests were performed. In our regular studies of in vitro chloroquine susceptibility tests on Plasmodium falciparum from non-immuners in Japan, the threshold point to differentiate resistant and susceptible isolates was set at a 0. 114 microM chloroquine in the semi-micro culture system, and this point was also applicable in the study of the malaria parasite taken in the highly endemic malarious area with good coincidence with clinical observation. Variation in the incubation time (24-63) to reach the schizont stage of the isolated parasites were noted. It appeared that chloroquine resistant P. falciparum showed traits to reach the schizont stage within a shorter incubation period.     

Effects of anti-protozoal drugs and histopathological studies on trypanosome species

The trypanosomostatic and trypanosomicidal effects of four anti-protozoal drugs, namely halofantrine hydrochloride, chloroquine phosphate, benzoylmetronidazole and pyrimethamine, on species of trypanosomes, viz. Trypanosoma brucei brucei (MBOS/NG/94/NITR) Bassa strain, T. congolense (MBOS/NG/93/NVRI) Zaria strain and T. brucei gambiense (MHOM/NG/92/NITR) Abraka strain, were investigated. In vitro and in vivo studies on these drugs vis-a-vis the parasites were carried out. The histopathological changes in organs and tissues of experimentally infected rats were also studied. Results from the in vitro studies indicated that halofantrine hydrochloride, chloroquine phosphate, benzoylmetronidazole and pyrimethamine appeared to be effective trypanosomicidal agents against T. brucei brucei (Bassa strain), T. congolense (Zaria strain) and T. brucei gambiense (Abraka strain). The in vivo studies showed that these drugs were sub-curative by prolonging the survival period of the trypanosome-infected rats, but not necessarily curing the infection. Histopathological findings indicated inflammatory reactions characterised by infiltration to variable degrees in the majority of tissues, mostly in the lungs and liver. The most consistent lesions were interstitial pneumonia, multifocal necrosis and oedema. Pathological findings showed the T. brucei brucei and T. brucei gambiense strains studied to be both intravascular and extravascular parasites. These results suggest that halofantrine hydrochloride, chloroquine phosphate, benzoylmetronidazole and pyrimethamine could be used as supportive, suppressive and/or synergistic/additive drugs in the treatment of African trypanosomiasis. Their effects on species of trypanosomes have been studied and are reported for the first time.     

Synthesis of ring-substituted 4-aminoquinolines and evaluation of their antimalarial activities

A simple two-step synthesis method was used to make 51 B-ring-substituted 4-hydroxyquinolines allowing analysis of the effect of ring substitutions on inhibition of growth of chloroquine sensitive and resistant strains of Plasmodium falciparum, the dominant cause of malaria morbidity. Substituted quinoline rings other than the 7-chloroquinoline ring found in chloroquine were found to have significant activity against the drug-resistant strain of P. falciparum W2.     

Synthesis and evaluation of the antiplasmodial activity of novel indeno[2,1-c]quinoline derivatives

With the aim to explore the potentiality of new chemical scaffolds for the design of new antimalarials, a set of new indeno[2,1-c]quinolines bearing different basic heads has been synthesized and tested in vitro against chloroquine sensitive (CQ-S) and chloroquine resistant (CQ-R) strains of Plasmodium falciparum. Most of the synthesized compounds exhibited a moderate antiplasmodial activity, inhibiting the growth of both CQ-S and CQ-R strains of P. falciparum with IC₅₀ ranging from 0.24 to 6.9 μM and with a very low resistance index. The most potent compounds (1.2–1.3-fold the CQ on the W-2 strain) can be considered as promising ‘lead compounds’ to be further optimized to improve efficacy and selectivity against Plasmodia.     

Synthesis and in vitro activities of ferrocenic aminohydroxynaphthoquinones against Toxoplasma gondii and Plasmodium falciparum

Fourteen ferrocenyl aminohydroxynaphthoquinones, analogues of atovaquone, were synthesized from the hydroxynaphthoquinone core. These novel atovaquone derivatives were tested for their in vitro activity against two apicomplexan parasites of medical importance, Toxoplasma gondii and Plasmodium falciparum, including resistant strains to atovaquone (T. gondii) and chloroquine (P. falciparum). Three of these ferrocenic atovaquone derivatives composed of the hydroxynaphthoquinone core plus an amino-ferrocenic group and an aliphatic chain with 6-8 carbon atoms were found to be significantly active against T. gondii. Moreover, these novel compounds were also effective against the atovaquone-resistant strain of T. gondii (Ato(R)).