Professor David Warhurst BSc PhD DSc FRCPath
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David Warhurst's Background
My main research has been on the Chemotherapy of Malaria which is described in 200 publications on that subject from 1965 to date.
In 1963 when I started work in the area of chloroquine (CQ) action and resistance in malaria, under grant support from WHO, the mode of action of CQ was accepted to be through binding to DNA, and the evidence, in particular from Hahn's group in the USA, was thought to be conclusive. At Mill Hill, with David Hockley (electron microscopist) we questioned this, concluding that the drug attacked the parasite lysosome. [ Ref.3. Warhurst,D.C. and Hockley,D.J. (1967) The mode of action of chloroquine on Plasmodium berghei and P.cynomolgi. Nature 214: 935‑6]. This stimulated much further work which has so far supported our view. Lysosomal drug accumulation had first been noticed in mammalian cells at NIMR, and our paper [Ref.11.Homewood,C.A. et al , (1972) Lysosomes, pH and the antimalarial action of chloroquine. Nature 235 : 50‑52] expanded the concept, explaining why the basic drugs became trapped by protonation in the acidic lysosome. It was, however, clear that this was not the full story, and the likelihood of a specific lysosomal receptor that was responsible for selective toxicity on malaria compared with mammalian cells was suggested by our group and others.
Work by Coy Fitch, in USA, on the competitive effects of other blood schizontocides like quinine on CQ uptake made it likely that a whole range of drugs which could be described as "blood schizontocides" were attacking the same site. Before Trager's cultivation studies in 1976, we used the rodent malaria P. berghei in short-term cultures, and were able to show that quinine, and some other agents, showed competitive inhibition of CQ-induced morphological changes in that parasite. The most potent drug tested was WR 142,490, later to be developed as mefloquine.[ Ref.20. Warhurst, D.C. and Thomas, S.C. (1975) Pharmacology of the malaria parasite. Biochemical Pharmacology 24: 2047‑56]. Although haematin, released from haemoglobin in the lysosome of the intraerythrocytic malaria parasite, had been suggested as a possible drug target early on, Fitch produced convincing evidence for this in 1980. After confirming binding of quinine and mefloquine to haemin in vitro using spectrophotometry, I modelled the molecular interaction between drug and ligand through a study of structure-activity relationships [Ref. 33. Warhurst, D.C. (1981) The quinine‑haemin interaction and its relationship to antimalarial activity. Biochemical Pharmacology 30:3323‑3327]. At last it was possible to explain why 9-epiquinine, which differs from quinine only in the configuration of asymmetric carbon 9, is inactive both in haematin binding and as an antimalarial, whilst other stereoisomers such as quinine and quinidine which differed in both carbons 8 and 9 were active. It has been chemically confirmed that ring/ring interactions take place between the drug quinoline nucleus and the porphyrin, together with the stereospecific interaction of haematin iron with hydroxyl on the drug side chain, in the case of the cinchona alkaloids. We have recently developed a descriptive formula to explain the effects of structure on function in these agents. [Ref 190 below].
We were fortunate to be one of the first western laboratories to have access to artemisinin from China. One of our several studies demonstrated that combinations of artemisinin with mefloquine, showed a marked synergism in vivo in rodent malaria. [Ref.73 Chawira et al, (1987) The effect of combinations of qinghaosu (Artemisinin) with standard antimalarial drugs in the suppressive treatment of malaria in mice. Transactions of the Royal Society of Tropical Medicine and Hygiene 81 :554‑558]. Artemisinin and its derivatives are now a boon in treatment of multidrug-resistant malaria. Use of mefloquine or a related drug together with artemisinin avoids the problem of recrudescence. Both the new drug combination, artemether/benflumetol (Co-artemether: introduced for treatment by Novartis) and mefloquine /artemisinin show the same type of synergism for the resistant strains of P. falciparum as they did for P. berghei. In the 90s we were able to examine drug-resistance-related genetic polymorphisms in P. falciparum in samples from field studies. The genetic determinants for CQ-resistance were still not agreed. However we were able to confirm the association of a mutation, Asn86Tyr , in the Pfmdr1 gene, with resistance to CQ in W.Africa [Ref.140 Adagu, et al. (1996). Guinea Bissau: association of chloroquine resistance of Plasmodium falciparum with the Tyr86 allele of the multiple drug‑resistance gene Pfmdr1. Trans.R. .Soc.Trop.Med.Hyg. 90:90‑91], and for the first time to demonstrate selection for this allele in the field. [Ref.148 Duraisingh, et al. (1997) Evidence for selection for the tyrosine‑86 allele of the pfmdr 1 gene in Plasmodium falciparum by chloroquine and amodiaquine.Parasitology 114:205‑211].
It now appears that changes in PGH-1, the product of pfmdr1 modulate but do not determine CQ-resistance (but it is much more important for mefloquine [ Ref 168: Duraisingh et al. Increased sensitivity to the antimalarials mefloquine and artemisinin is conferred by mutations in the Pfmdr1 gene of Plasmodium falciparum. Molecular Microbiology 2000; 36: 9]. CQ-resistant Plasmodium falciparum parasites take up less drug, apparently due to drug release due to a K76T change in a transmembrane domain of lysosomal protein PfCRT. Although this residue change is critical, other changes are always present in one or more of the adjacent residues 72, 74 and 75, and these vary with geographical origin of the isolate. CQ resistance is partially reversed in vitro by verapamil (VE) and other weakly basic hydrophobic agents. Although amodiaquine (AQ) is a 4-aminoquinoline like CQ, it is effective in CQ-resistance in Africa. We found that resistance-reversing drugs and also AQ and DAQ, its metabolite, were more hydrophobic than CQ. The relative activity in CQ-resistant parasites of a series of antimalarial 4-aminoquinolines correlated well with measured hydrophobicity [Ref 191]. Brazilian CQ-resistant isolates with 72-SVMNT-76 compared with 72-CVIET-76 in Africa showed poor VE reversal in vitro and AQ was clinically ineffective. In clones transfected with different PfCRT alleles, mean hydrophobicity of residues 72-76 correlated with activity of DAQ and CQ-VE but not CQ. [Ref. 189]. With this information we can predict the effect of PfCRT allele variations (in the absence of variation in PGH-1 the product of pfmdr1) on the effectiveness of AQ in the clinic. Allele SVMNT was also recently detected in Papua New Guinea and in the Indian subcontinent and is predicted to be refractory to AQ treatment like the same allele in Brazil [Ref 189]. The scheme we published in 2002 [Ref 180] explains the rationale. Crucial changes in residues 72-76 of wild type (CVMNK) PfCRT remove a positive charge (K76T) and render the transmembrane domain more hydrophobic, allowing exit of positively charged drug (CQ2H+).Verapamil is also trapped within the lysosome, forming a monocation (VEH+) which is much more hydrophobic than the CQ dication, and binds to the hydrophobic channel formed by the modified transmembrane domain, reducing the exit of CQ2H+ by its bulk and by its positive charge. 4-aminoquinolines like AQ and its metabolite DAQ which are more hydrophobic than CQ are significantly more active than CQ against CQ-resistant strains. Hydrophobic 4-aminoquinolines are likely to have a VE-like effect and prevent their own exit.
Selected Malaria Publications on chloroquine-resistance 2000-2003
169: Omar SA, Adagu IS, Gump DW, Ndaru NP, Warhurst DC. Plasmodium falciparum in Kenya: high prevalence of drug-resistance-associated polymorphisms in hospital admissions with severe malaria in an epidemic area. Ann Trop Med Parasitol. 2001; 95: 661-9.
171: Warhurst DC, Duraisingh MT. Rational use of drugs against Plasmodium falciparum. Trans R Soc Trop Med Hyg. 2001; 95: 345-6.
172: Adagu IS, Warhurst DC. Plasmodium falciparum: linkage disequilibrium between loci in chromosomes 7 and 5 and chloroquine selective pressure in Northern Nigeria. Parasitology. 2001; 123: 219-24.
177: Warhurst DC. A molecular marker for chloroquine-resistant falciparum malaria. N Engl J Med. 2001; 344: 299-302.
178: Sutherland CJ, Alloueche A, Curtis J, Drakeley CJ, Ord R, Duraisingh M, Greenwood BM, Pinder M, Warhurst D, Targett GA. Gambian children successfully treated with chloroquine can harbor and transmit Plasmodium falciparum gametocytes carrying resistance genes. Am J Trop Med Hyg. 2002; 67: 578-85.
180: Warhurst DC, Craig JC, Adagu IS. Lysosomes and drug resistance in malaria. Lancet. 2002; 360 : 1527-9.
189: Warhurst DC. Polymorphism in the Plasmodium falciparum chloroquine-resistance transporter protein links verapamil enhancement of chloroquine sensitivity with the clinical efficacy of amodiaquine. Malar J. 2003; 2:31.
190: Warhurst DC, Craig JC, Adagu IS, Meyer DJ, Lee SY. The relationship of physico-chemical properties and structure to the differential antiplasmodial activity of the cinchona alkaloids. Malar J. 2003; 2: 26.
191: Warhurst DC, Steele JC, Adagu IS, Craig JC, Cullander C. Hydroxychloroquine is much less active than chloroquine against chloroquine-resistant Plasmodium
Warhurst DC, Craig JC, Adagu IS, Guy RK, Madrid PB, Fivelman QL, Activity of piperaquine and other 4-aminoquinoline antiplasmodial drugs against chloroquine-sensitive and resistant blood-stages of Plasmodium falciparum.Role of b-haematin inhibition and drug concentration in vacuolar water- and lipidphases, Biochemical Pharmacology (2007) In Press, Accepted Manuscript, Available online 19 March 2007
David Warhurst's Affiliation
David Warhurst's Research
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- Molecular biology
Disease and Health Conditions
Mutation in the Plasmodium falciparum CRT protein determines the stereospecific activity of the antimalarial Cinchona alkaloids.
Griffin, C.E. ; Hoke, J.M. ; Samarakoon, U. ; Duan, J. ; Mu, J. ; Ferdig, M.T. ; Warhurst, D.C. ; Cooper, R.A. ;
Antimicrob Agents Chemother, 2012;
Dynamics of pfcrt alleles CVMNK and CVIET in chloroquine-treated Sudanese patients infected with Plasmodium falciparum.
Gadalla, N.B.; Elzaki, S.E.; Mukhtar, E.; Warhurst, D.C.; El-Sayed, B.; Sutherland, C.J.;
Malar J, 2010; 9:74
Oral activated charcoal prevents experimental cerebral malaria in mice and in a randomized controlled clinical trial in man did not interfere with the pharmacokinetics of parenteral artesunate.
de Souza, J.B.; Okomo, U.; Alexander, N.D.; Aziz, N.; Owens, B.M.; Kaur, H.; Jasseh, M.; Muangnoicharoen, S.; Sumariwalla, P.F.; Warhurst, D.C.; Ward, S.A.; Conway, D.J.; Ulloa, L.; Tracey, K.J.; Foxwell, B.M.; Kaye, P.M.; Walther, M.;
PLoS One, 2010; 5(4):e9867
Monitoring for multidrug-resistant Plasmodium falciparum isolates and analysis of pyrimethamine resistance evolution in Uige province, Angola.
Menegon, M.; Pearce, R.J.; Inojosa, W.O.; Pisani, V.; Abel, P.M.; Matondo, A.; Bisoffi, Z.; Majori, G.; Ord, R.; Warhurst, D.C.; Roper, C.; Severini, C.;
Trop Med Int Health, 2009; 14(10):1251-7
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