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P G Bray

Publications and source records attributed to P G Bray.

At least 19 recordsLinked to original sources
PubMed2005

Quinolines and artemisinin: chemistry, biology and history.

Plasmodium falciparum is the most important parasitic pathogen in humans, causing hundreds of millions of malaria infections and millions of deaths each year. At present there is no effective malaria vaccine and malaria therapy is totally reliant on the use of drugs. New drugs are urgently needed because of the rapid evolution and spread of parasite resistance to the current therapies. Drug resistance is one of the major factors contributing to the resurgence of malaria, especially resistance to the most affordable drugs such as chloroquine. We need to fully understand the antimalarial mode of action of the existing drugs and the way that the parasite becomes resistant to them in order to design and develop the new therapies that are so urgently needed. In respect of the quinolines and artemisinins, great progress has been made recently in studying the mechanisms of drug action and drug resistance in malaria parasites. Here we summarize from a historical, biological and chemical, perspective the exciting new advances that have been made in the study of these important antimalarial drugs.

Animals↗
PubMed2003

Artemisinins target the SERCA of Plasmodium falciparum.

Artemisinins are extracted from sweet wormwood (Artemisia annua) and are the most potent antimalarials available, rapidly killing all asexual stages of Plasmodium falciparum. Artemisinins are sesquiterpene lactones widely used to treat multidrug-resistant malaria, a disease that annually claims 1 million lives. Despite extensive clinical and laboratory experience their molecular target is not yet identified. Activated artemisinins form adducts with a variety of biological macromolecules, including haem, translationally controlled tumour protein (TCTP) and other higher-molecular-weight proteins. Here we show that artemisinins, but not quinine or chloroquine, inhibit the SERCA orthologue (PfATP6) of Plasmodium falciparum in Xenopus oocytes with similar potency to thapsigargin (another sesquiterpene lactone and highly specific SERCA inhibitor). As predicted, thapsigargin also antagonizes the parasiticidal activity of artemisinin. Desoxyartemisinin lacks an endoperoxide bridge and is ineffective both as an inhibitor of PfATP6 and as an antimalarial. Chelation of iron by desferrioxamine abrogates the antiparasitic activity of artemisinins and correspondingly attenuates inhibition of PfATP6. Imaging of parasites with BODIPY-thapsigargin labels the cytosolic compartment and is competed by artemisinin. Fluorescent artemisinin labels parasites similarly and irreversibly in an Fe2+-dependent manner. These data provide compelling evidence that artemisinins act by inhibiting PfATP6 outside the food vacuole after activation by iron.

Animals↗
PubMed2001

P-Glycoprotein and transporter MRP1 reduce HIV protease inhibitor uptake in CD4 cells: potential for accelerated viral drug resistance?

BACKGROUND: The multidrug transporters P-glycoprotein (P-gp) and MRP1 are functionally expressed in several subclasses of lymphocytes. HIV-1 protease inhibitors interact with both; consequently the transporters could reduce the local concentration of HIV-1 protease inhibitors and, thus, influence the selection of viral mutants. OBJECTIVES: To study the effect of the expression of P-gp and MRP1 on the transport and accumulation of HIV-1 protease inhibitors in human lymphocytes and to study the effects of specific P-gp and MRP1 inhibitors. METHODS: The initial rate and the steady-state intracellular accumulation of radiolabelled ritonavir, indinavir, saquinavir and nelfinavir was measured in three human lymphocyte cell lines: control CEM cells, CEM-MDR cells, which express 30-fold more P-gp than CEM cells, and CEM-MRP cells, which express fivefold more MRP1 protein than CEM cells. The effect of specific inhibitors of P-gp (GF 120918) and MRP1 (MK 571) was also examined. RESULTS: Compared with CEM cells, the initial rates of uptake and the steady-state intracellular concentrations of all protease inhibitors are significantly reduced in CEM-MDR cells. The intracellular concentrations of the protease inhibitors are increased upon co-administration with GF 120918, in some cases to levels approaching those in CEM cells. The intracellular concentrations of the protease inhibitors are also significantly reduced in CEM-MRP cells. Co-administration with MK -571 can partially overcome these effects. CONCLUSIONS: The overexpression of multidrug transporters significantly reduces the accumulation of protease inhibitors at this major site of virus replication, which, potentially, could accelerate the acquisition of viral resistance. Targeted inhibition of P-gp may represent an important strategy by which this problem can be overcome.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2001

Diamidine compounds: selective uptake and targeting in Plasmodium falciparum.

Extensive drug resistance in Plasmodium falciparum emphasizes the urgent requirement for novel antimalarial agents. Here we report potent antimalarial activity of a number of diamidine compounds. The lead compound pentamidine is concentrated 500-fold by erythrocytes infected with P. falciparum. Pentamidine accumulation can be blocked by inhibitors of hemoglobin digestion, suggesting that the drug binds to ferriprotoporphyrin IX (FPIX). All of the compounds bound to FPIX in vitro and inhibited the formation of hemozoin. Furthermore, inhibitors of hemoglobin digestion markedly antagonized the antimalarial activity of the diamidines, indicating that binding to FPIX is crucial for the activity of diamidine drugs. Pentamidine was not accumulated into uninfected erythrocytes. Pentamidine transport into infected cells exhibits an initial rapid phase, nonsaturable in the micromolar range and sensitive to inhibition by furosemide and glibenclamide. Changing the counter-ion in the order Cl(-) < Br(-) < NO(2)(-) < I(-) <SCN(-) markedly stimulated pentamidine transport. These data suggest that pentamidine is transported although a pore or ion channel with properties similar to those of the recently characterized 'induced permeability pathway' on the infected red cell membrane. In summary, the diamidines exhibit two levels of selectivity against P. falciparum. The route of entry and molecular target are both specific to malaria-infected cells and are distinct from targets in other protozoa. Drugs that target the hemoglobin degradation pathway of malaria parasites have a proven record of accomplishment. The employment of induced permeability pathways to access this target represents a novel approach to antiparasite chemotherapy and offers an additional level of selectivity.

Animals↗
PubMed2000

Potent enhancement of the sensitivity of Plasmodium falciparum to chloroquine by the bisbenzylisoquinoline alkaloid cepharanthin.

Cepharanthin is a proprietary extract of Stephania cepharantha, widely used in Japan for the treatment of inflammatory diseases. Cephranthin, its component alkaloids, and the standard resistance modulator verapamil were tested against Plasmodium falciparum for capacity to modulate sensitivity to chloroquine. Cepharanthin enhanced the activity of chloroquine against resistant clones by a factor of 15 at a concentration of only 200 nM (1.2 ng/ml). It is 50 times more potent than verapamil and 3 times more potent than the sum of its individual alkaloids. Combinations of component alkaloids acted synergistically to sensitize the parasite to chloroquine, possibly explaining the enhanced potency of Cepharanthin. Cepharanthin differed from verapamil in that it further sensitized clones that are considered to be fully susceptible, improving the baseline activity of chloroquine. Potent sensitization of parasites to chloroquine in vitro coupled with low toxicity suggests that coadministration of Cepharanthin might extend the clinical utility of chloroquine.

Alkaloids↗
PubMed1999

Cellular uptake of chloroquine is dependent on binding to ferriprotoporphyrin IX and is independent of NHE activity in Plasmodium falciparum.

Here we provide definitive evidence that chloroquine (CQ) uptake in Plasmodium falciparum is determined by binding to ferriprotoporphyrin IX (FPIX). Specific proteinase inhibitors that block the degradation of hemoglobin and stop the generation of FPIX also inhibit CQ uptake. Food vacuole enzymes can generate cell-free binding, using human hemoglobin as a substrate. This binding accounts for CQ uptake into intact cells and is subject to identical inhibitor specificity. Inhibition of CQ uptake by amiloride derivatives occurs because of inhibition of CQ-FPIX binding rather than inhibition of the Na+/H+ exchanger (NHE). Inhibition of parasite NHE using a sodium-free medium does not inhibit CQ uptake nor does it alter the ability of amilorides to inhibit uptake. CQ resistance is characterized by a reduced affinity of CQ-FPIX binding that is reversible by verapamil. Diverse compounds that are known to disrupt lysosomal pH can mimic the verapamil effect. These effects are seen in sodium-free medium and are not due to stimulation of the NHE. We propose that these compounds increase CQ accumulation and overcome CQ resistance by increasing the pH of lysosomes and endosomes, thereby causing an increased affinity of binding of CQ to FPIX.

Amiloride↗
PubMed1999

Chloroquine uptake and activity is determined by binding to ferriprotoporphyrin IX in Plasmodium falciparum.

The selective antimalarial activity of chloroquine and related compounds stems from the extensive saturable uptake of these drugs into malaria parasites. Chloroquine resistant strains of Plasmodium falciparum have evolved a mechanism to reduce the saturable uptake. The molecular mechanism of saturable chloroquine uptake is controversial and attention is currently focused on mutually exclusive models of active chloroquine uptake and intracellular chloroquine binding. We sum up recent evidence which conclusively proves that the saturable accumulation of chloroquine is due to intracellular binding to ferriprotoporphyrin IX rather than active transport into the parasite via the sodium/hydrogen exchanger. We discuss recent findings that the affinity of chloroquine binding to ferriprotoporphyrin IX is reduced in resistant parasites. The mechanism responsible for reduced binding affinity can be overcome by verapamil and various lysosomotropic agents, and is thought to be the basis of chloroquine resistance.

Animals↗
PubMed1999

Phenotypic and genotypic characteristics of recently adapted isolates of Plasmodium falciparum from Thailand.

The drug sensitivity characteristics and Plasmodium falciparum pfmdr1 status of five isolates of P. falciparum recently isolated from patients presenting for treatment from the Thailand/Myanmar border have been investigated. The aim of the study was to avoid the criticisms of some earlier studies by focusing on newly collected isolates from a specific geographic location. Three of the isolates studied exhibited clear resistance to chloroquine similar to that observed in the K1 Thai standard isolate obtained in the 1970s, and the other two isolates were of intermediate sensitivity to chloroquine with concentrations of drug that inhibit parasite growth by 50% of 50 and 43 nmol. The sensitivity of all isolates was enhanced by verapamil but we found no clear association between chloroquine sensitivity and gene copy number or intra-allelic variation of pfmdr1. In contrast, clear cross-resistance was seen between mefloquine and halofantrine, with the most sensitive isolates carrying the K1 mutation in pfmdr1.

Animals↗
PubMed1998

A comparison of the phenomenology and genetics of multidrug resistance in cancer cells and quinoline resistance in Plasmodium falciparum.

Plasmodium falciparum is the causative agent of the most deadly form of human malaria. Chemotherapy traditionally has been the main line of defense against this parasite, and chloroquine, the drug of choice, has been one of the most successful drugs ever developed. Unfortunately, the evolution and spread of resistance to chloroquine and other quinoline-containing drugs means that these compounds are now virtually useless in many endemic areas. Future prospects for the use of quinoline compounds improved considerably when it was demonstrated that chloroquine resistance could be circumvented in vitro by a number of structurally and functionally unrelated compounds such as verapamil and desipramine. The phenomenon of resistance reversal by compounds such as verapamil is also a key feature of drug resistance in mammalian cells, and this has raised the possibility that the underlying mechanisms of drug resistance of the two cell types could be similar. This hypothesis has prompted a large number of studies into the genetics and biochemistry of resistance to quinoline-containing drugs in P. falciparum. Both the genetic and the biochemical studies have raised issues of controversy and stimulated much debate. These issues are discussed in this review, in the context of a comparison with the genetics and biochemistry of multidrug resistance in mammalian cells.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1998

4-Aminoquinolines--past, present, and future: a chemical perspective.

The 4-aminoquinoline chloroquine (1) can be considered to be one of the most important synthetic chemotherapeutic agents in history. Since its discovery, chloroquine has proved to be a highly effective, safe, and well-tolerated drug for the treatment and prophylaxis of malaria. However, the emergence of chloroquine-resistant strains of the malarial parasite has underlined the requirement for a synthetic alternative to chloroquine. This review describes structure-activity relationships for the 4-aminoquinolines, along with views on the mechanism of action and parasite resistance. A description of drug metabolism and toxicity also is included, with a brief description of potential approaches to the design of new synthetic derivatives.

Aminoquinolines↗
PubMed1998

Access to hematin: the basis of chloroquine resistance.

The saturable uptake of chloroquine by parasites of Plasmodium falciparum has been attributed to specific carrier-mediated transport of chloroquine. It is suggested that chloroquine is transported in exchange for protons by the parasite membrane Na+/H+ exchanger [J Biol Chem 272:2652-2658 (1997)]. Once inside the parasite, it is proposed that chloroquine inhibits the polymerization of hematin, allowing this toxic hemoglobin metabolite to accumulate and kill the cell [Pharmacol Ther 57:203-235 (1993)]. To date, the contribution of these proposed mechanisms to the uptake and antimalarial activity of chloroquine has not been assessed. Using sodium-free medium, we demonstrate that chloroquine is not directly exchanged for protons by the plasmodial Na+/H+ exchanger. Furthermore, we show that saturable chloroquine uptake at equilibrium is due solely to the binding of chloroquine to hematin rather than active uptake: using Ro 40-4388, a potent and specific inhibitor of hemoglobin digestion and, by implication, hematin release, we demonstrate a concentration-dependent reduction in the number of chloroquine binding sites. An equal number of chloroquine binding sites are found in both resistant and susceptible clones, but the apparent affinity of chloroquine binding is found to correlate with drug activity (r2 = 0.93, p < 0.0001). This completely accounts for both the reduced drug accumulation and activity observed in resistant clones and the "reversal" of resistance produced by verapamil. The data presented here reconcile most of the available biochemical data from studies of the mode of action of chloroquine and the mechanism of chloroquine resistance. We show that the activity of chloroquine and amodiaquine is directly dependent on the saturable binding of the drugs to hematin and that the inhibition of hematin polymerization may be secondary to this binding. The chloroquine-resistance mechanism regulates the access of chloroquine to hematin. Our model is consistent with a resistance mechanism that acts specifically at the food vacuole to alter the binding of chloroquine to hematin rather than changing the active transport of chloroquine across the parasite plasma membrane.

Animals↗
PubMed1998

Central role of hemoglobin degradation in mechanisms of action of 4-aminoquinolines, quinoline methanols, and phenanthrene methanols.

We have used a specific inhibitor of the malarial aspartic proteinase plasmepsin I and a nonspecific cysteine proteinase inhibitor to investigate the importance of hemoglobin degradation in the mechanism of action of chloroquine, amodiaquine, quinine, mefloquine (MQ), halofantrine, and primaquine. Both proteinase inhibitors antagonized the antiparasitic activity of all drugs tested with the exception of primaquine. An inhibitor of plasmepsin I, Ro40-4388, reduced the incorporation of radiolabelled chloroquine and quinine into malarial pigment by 95%, while causing a 70% reduction in the incorporation of radiolabelled MQ. Cysteine proteinase inhibitor E64 reduced the incorporation of chloroquine and quinine into malarial pigment by 60 and 40%, respectively. This study provides definitive support for the central role of hemoglobin degradation in the mechanism of action of the 4-aminoquinolines and the quinoline and phenanthrene methanol antimalarials.

Amodiaquine↗
PubMed1998

Relationship between antimalarial drug activity, accumulation, and inhibition of heme polymerization in Plasmodium falciparum in vitro.

We have investigated the contribution of drug accumulation and inhibition of heme polymerization to the in vitro activities of a series of antimalarial drugs. Only those compounds exhibiting structural relatedness to the quinolines inhibited heme polymerization. We could find no direct correlation between in vitro activity against chloroquine-susceptible or chloroquine-resistant isolates and either inhibition of heme polymerization or cellular drug accumulation for the drugs studied. However, in vitro activity against a chloroquine-susceptible isolate but not a chloroquine-resistant isolate showed a significant correlation with inhibition of heme polymerization when the activity was normalized for the extent of drug accumulation. The importance of these observations to the rational design of new quinoline-type drugs and the level of agreement of these conclusions with current views on quinoline drug action and resistance are discussed.

Animals↗
PubMed1997

Synthesis, antimalarial activity, and molecular modeling of tebuquine analogues.

Tebuquine (5) is a 4-aminoquinoline that is significantly more active than amodiaquine (2) and chloroquine (1) both in vitro and in vivo. We have developed a novel more efficient synthetic route to tebuquine analogues which involves the use of a palladium-catalyzed Suzuki reaction to introduce the 4-chlorophenyl moiety into the 4-hydroxyaniline side chain. Using similar methodology, novel synthetic routes to fluorinated (7a, b) and a dehydroxylated (7c) analogue of tebuquine have also been developed. The novel analogues were subjected to testing against the chloroquine sensitive HB3 strain and the chloroquine resistant K1 strain of Plasmodium falciparum. Tebuquine was the most active compound tested against both strains of Plasmodia. Replacement of the 4-hydroxy function with either fluorine or hydrogen led to a decrease in antimalarial activity. Molecular modeling of the tebuquine analogues alongside amodiaquine and chloroquine reveals that the inter-nitrogen separation in this class of drugs ranges between 9.36 and 9.86 A in their isolated diprotonated form and between 7.52 and 10.21 A in the heme-drug complex. Further modeling studies on the interaction of 4-aminoquinolines with the proposed cellular receptor heme revealed favorable interaction energies for chloroquine, amodiaquine, and tebuquine analogues. Tebuquine, the most potent antimalarial in the series, had the most favorable interaction energy calculated in both the in vacuo and solvent-based simulation studies. Although fluorotebuquine (7a) had a similar interaction energy to tebuquine, this compound had significantly reduced potency when compared with (5). This disparity is possibly the result of the reduced cellular accumulation (CAR) of fluorotebuquine when compared with tebuquine within the parasite. Measurement of the cellular accumulation of the tebuquine analogues and seven related 4-aminoquinolines shows a significant relationship (r = 0.98) between the CAR of 4-aminoquinoline drugs and the reciprocal of drugs IC50.

Aminoquinolines↗