Autoimmunity in chagas disease cardiomyopathy: Fulfilling the criteria at last?

Reviews . . . . .......

40 Shimokawa, H. et al. (1988) Prostacyclin releases endotheliumderived relaxing factor and potentiates its action in coronary arteries of the pig. Br. ]. Pharmacol.95, 1197-1203 41 Betz,M. and Fox, B. (1991) Prostaglandin E2 inhibits production of Thl lymphokines but not of Th2 lymphokines. ]. hnmunol. 146,108-113 42 Goodwin, J.S., Messner, R.P. and Peake, G.T. (1978) Prostaglandin suppression of mitogen-stimulated lymphocytes in vitro. Changes with mitogen dose and preincubation. Clin. Invest. 62, 753-760 43 Tilden, A.B. and Balch, C.M. (1982) A comparison of PGE2 effects on human suppressor cell function and on interleukin 2 function. J. hnmunol. 129, 2469-2473 44 Hasler, F. et al. (1983) Analysis of the defects responsible for the impaired regulation of EBV-induced B cell proliferation by rheumatoid arthritis lymphocytes. II. Role of monocytes and the i,acreased sensitivity of rheumatoid arthritis lymphocytes to prostaglandin E. I. hmmmol. 131,768-772 45 Lict~tenstein, L.M. and DeBarnardo, R. (1971) The immediate allergic response: in vitro action of cyclic AMP-active and other drugs on the two stages of histamine release. ]. hmmmol. 107, 1131-1136 46 Ham, E.A. et al. (1983) Inhibition by prostaglandins of leukotriene B4 release from activated neutrophils. Proc. Natl. Acad. Sci. U.S.A. 80, 4349-4353 47 Andersen, N.H. el al. (1980) On the multiplicity of platelet prostaglandin receptors. I. Evaluation of competitive antagonism by aggregometry. Pwstaglandin:~19, 711-735 48 Williams, T.J. and Peck, M.J. (1977) Role of prostaglandinmediated vasodilation in inflammation° Nature 270, 530-532 49 Davies, P. et al. (1984)The role of arachidonic acid oxygenation products in pain and inflammation. Amul. Rev. Immmtol. 2, 335-357

50 McMillen, M.A. and Sumpio, B.E. (1995) Endothelins: polyfunctional cytokines. J. Am. Coll. Surg. 180,621-637 51 Yakubu, M.A., Shibata, M. and Leffler,C.W. (1995) Hematomainduced enhanced cerebral vasoconstrictions to leukotriene C4 and endothelin-1 in piglets: role of prostanoids. Pediatr. Res. 38, 119-123 52 Silldorff, E.P., Yang, S. and Pallone, T.L. (1995) Prostaglandin E2 abrogates endothelin-induced vasoconstriction in renal outer medullary descending vasa recta of the rat. J. Clin. Invest. 95, 2734-2740 53 Kemp, D.H. et al. (1983) Comparison of cutaneous hyperemia in cattle elicited by larvae of Boophilus microplus and by prostaglandins and other mediators. Experientia39, 725--727 54 Wikel, S.K. (1996) Host immunity to ticks. Annu. Rev. Entomol. 41, 1-22 55 Ramachandra, R.N. and Wikel, S.K. (1992) Modulation of hostimmune responses by ticks (Acari: Ixodidae): Effects of salivary gland extracts on host macrophages and lymphocyte cytokine production. I. Med. Entomol. 29, 818-826 56 Goodwin, J.S. and Ceuppens, J. (1983) Regulation of the immune response by prostaglandins. ]. Clin. hnmunol. 3, 295-315 57 Krause, D.S. and Deutsch, C. (1991) Cyclic AMP directly inhibits IL-2 receptor expression in human cells. Expression of both p55 and p75 subunits is affected. ]. lmmunol. 146,2285-2294 58 Bergman, D.K.. Ramachandra, R.N. and Wikel, S.K. (1996) Dermacentor andersoni: Salivary gland proteins suppressing T-lymphocyte responses to concanavalin A in vitro. Exp. Parasitol. 81,262-271 59 Chinery, W.A. and Ayitey-Smith, E. (1977) Histamine blocking agent in the salivary gland homogenate of the tick Rhipicephalus sanguineus sanguineus. Nature 265, 366-367 60 Ribeiro,J.M.C. and Spielman, A. (1986) Ixodes dammini: salivary anaphylatoxin inactivating activity. Exp. Parasitol.62, 292-297

Focus

Autoirnrnunity in Chagas Disease Cardiornyopathy: Fulfilling the Criteria at Last? J. Kalil and E. Cunha-Neto Here, ]orge Kalil and Eddcio Cunha-Neto review the recent evidence for autoi,t,tunity i, chronic Chagas cardhmtyopathy (CCC) involving molecularly defined antigens and im,tunopathoh~gical mechanisms. They also discuss the criteria for assign,tent of CCC as an organ-specific autoimmune disease. Chronic C h a g a s disease c a r d i o m y o p a t h y (CCC) is an i n f l a m m a t o r y c a r d i o m y o p a t h y afflicting about 30% of the 20 million i n d i v i d u a l s infected with the protozoan Trypailosoma cruzi in the Americas 1,2. 1 h e finding of a T-cell-rich i n f l a m m a t o r y m o n o n u c l e a r cell infiltrate ( p r e s u m a b l y the ultimate effector of tissue damage) in the presence of extremely few parasites in the heart lesions cast d o u b t on the direct participation of Trypanosoma cruzi in CCC heart tissue lesions, and suggested the possible involvement of a u t o i m m u n i t y 2. Such an anti-heart a u t o i m m u n e response could be i

]orge Kalil and Ed6cio Cunl rd-Neto are at theTransplantation Immunology Laboratory, Instituto do Cora(;~o Faculdade de Medicina, Universidade de S~o Paulo, S~o Paulo, SP, Brazil. Tel: +55 I I 282 9350, Fax: +55 I I 282 2354, e-mail: [email protected] 396

triggered either by molecular m i m i c r y with s o m e T. cruzi antigens h o m o l o g o u s to heart proteins, or b y the d i s p l a y of intracellular sequestered heart proteins secondary to parasite-driven myocarditis d u r i n g acute infection. The early attempts to identify an autoi m m u n e c o m p o n e n t in CCC (reviewed in Ref. 2) involved the induction of heart lesions in rabbits b y repeated i m m u n i z a t i o n w i t h T. cruzi h o m o g e n a t e s 3. A major drawback on the study of a u t o i m m u n i t y in C C C w a s the use of relatively crude antigen p r e p a r a t i o n s a n d assays and the absence of an adequate a n i m a l m o d e l 4. The inability to find a defined relevant selfantigen (and crossreactive T. cruzi molecule) h i n d e r e d the fulfillment of W i t e b s k y ' s criteria 5 for a u t o i m m u n e pathogenesis. In search of d e f i n e d a n t i g e n i c targets T i s s u e - d a m a g e - i n d u c e d a u t o i m m u n i t y can be an i m p o r t a n t i m m u n o p a t h o l o g i c a l m e c h a n i s m in some a u t o i m m u n e diseases. However, in h u m a n disease, it is often hard to discern w h e t h e r d a m a g e - i n d u c e d a u t o i m m u n i t y bears causal relationship w i t h disease or is just an e p i p h e n o m e n o n . Moreover, in a n infectious disease, one has to exclude the possibility that

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Parasitology Today, vol. 12, no. I O, 1996

FOCUS Table I. Molecular mimicry in Chagas diseasea T. cruz/molecule

Host c o m p o n e n t

Host

Disease-related

Molecular definition

Ref.

c~-Gal residues

~-Gal, EVI

A

No

Sugar moieties

7, 8

SRA

Skeletal muscle, SRA

A

NT

AS

9

Neurons, liver, kidney, testis

A

NT

mAb

10

Neurons

A

NT

mAb

II

Sulphated glycolipids

Neurons

A

NT

mAb

12

FI-160

47 kDa neuronal protein

A

No

RDNA, AS

13

Ribosomal P protein

Ribosomal P protein (HeLa cell line)

H

Yes

RDNA, Ab, SP

14

23 kDa ribosomal protein

23 kDa ribosomal protein (HeLa cells reticulocytes)

H

No

Ab

IS

Microtubule-associated proteins

Microtubule-associated proteins (brain)

A, H

No

RDNA, AS

16

Ribosomal P0 protein

13I-adrenoreceptor

H

No

RDNA, Ab, SP

17

BI 3 protein

Human cardiac myosin heavy chain (human ventricle)

H

Yes

RDNA, Ab, SP

24

?

" AS, antisecum; Ab, antibody (fi-om patient): mA.b, monoclonal antibody: RDNA, recombinant DNA: SP, synthetic peptides: A, animal model: H, human: NT, not tested: EVI, endothelium and vascular inter~titium antibodies.

autoimmunity was induced by molecular mirnicry invo' ¢ing pathogen components. Molecular mimicry is taken as the most established autoimmunityinducing mechanism °. Table I lists the antibody-based molecular mimicry systems involving defined target antigens. Antibodies binding to rodent endothelium and vascular interstitium (EVI antibodies) present in sera from Chagas disease patients could be absorbed with T. cruzi epimastigotes7; it was later shown that EVI antibodies recognize o~-galactosyl glycoproteins absent from human tissue ~. Anti-SRA antibody identiffed in CCC sera crossreactively recognized a conserved ion-translocating enzyme present in T. cruzi and striated muscle membranes, which is also recognized by sera from patients with degenerative muscle diseased J. in the 1980s, several groups developed murine monoclona] antibodies (]nAbs) crossreactively recog-, and neural tissl,] e 10- 12 . nizing components of T. cru,.i Molecular m i m i c r y was also s h o w n to occur between recombinant flagellar T. cruzi molecule (F1-160) and an undefined 47 kDa neuronal protein. A 12-residue epi-

~l-adrenergic receptor 17. However, there was no proof that antibodies that recognize the epitope shared between P0 pcotein and ~l-receptor are more prevalent among CCC than in asymptomatic T. cruzi seropositive patients (ASY). The lack of specific recognition by CCC patients as compared to ASY individuals, as well as the lack of organ specificity of tile selfantigen recognized, seemed to indicate that most of the crossreactive antibodies disclosed had limited clinical significance, not unlike the innocuous 'natural' autoantibodies found in sera from normal individuals is. The finding that CD4 + T cells from chronically infected BALB/c mice apparently can induce the production of heart lesions l~ suggested a possible role for T-cell-mediated autoimmunity in murine models of CCC. However, the establishment of human CCC as an organ-specific autoimmune disease depends on matching several of the criteria for autoimmune disease recently revisited by Rose and Bona 2° (Box 1). Myosin as major autoantigen

Myosin, the most abundant heart protein, is a major tope from Fl-160 was found to be capable of competiantigen in several instances of heart-specific autoimtively inhibiting the binding of anti-Fl-lo0 antibodies reunify; moreover, immunization with cardiac myosin to neural tissue 13. However, antibodies neither to the in Freund's adjuvant induces severe myocarditis in whole Fl-160 recombinant protein nor to the competing peptide were associated with CCC or with chronic digestive forms Box 1. Fulfilling Rose and Bona's Criteria 2° for Autoimmunity? of Chagas disease. Other defined crossreactive molecules included Circumstantial evidence: evolutionarily conserved molecules • Scarcity of parasites in heart lesions 2,3° such as ribosomal proteins 14,Is or • Lymphocytic infiltration of target organ (heart) 27 cytoskeletal MAP proteins 16 which • Restricted T-cell receptor variable gene usage in situ 2s have an ubiquitous distribution, thus being unsuitable targets of a Indirect evidence: heart- or neuron-specific immune • Passive transfer of disease in murine models by CD4 + T cells l~ • Molecular mimicry between T. cruzi and target organs (see Table 1) attack. A recent i'epert showed mol• Heart disease-associated molecular mimicry between heart-specific ecular mimicry at the epitope level epitope and T. cruzi antigen 25 between ribosomal protein P0 of • isolation of self reactive T cells infiltrating the lesions in target organ 31'32 T. cruzi and an epitope at the external domains of the cardiovascular Parositolop~y Today, vol. 12, no. I O, 1996

397

Focus mice 2t. Cardiac myosin is a major heart-specific autoantigen for CD4 + T cells 22 in murine models of CCC. Anti-cardiac myosin antibodies are associated with cardiomyopathy in T. cruzi infected mice 23. Taken together, these Ieports suggest the possible relevance of myosin recognition in human CCC. We recently identified a heart-specific epitope (residues 1442-1447, AAALDK) of cardiac myosin heavy chain, displaying molecular mimicry with a secondary epitope (hexapeptide AAAGDK from 12-mer tandem repeats) of the immunodominant recombinant prot,~'tn B13 from T. cmtzi 24. Cardiac myosin-B13 cross: eactive antibodies were present in 100% of CCC sera tested but only 14% of ASY sera tested, a statistically significant difference (P = 2.3 x 10-6). The peculiar patient distribution clearly distinguish antibodies directed to the B13 crossreactive epitope in cardiac myosin, from natural anti-myosin antibodies present in normal controls, ASY and CCC patients 24,25. Furthermore, myosin-lY 3 ,::'ossreactive antibodies were absent in sera from ca :diomyopathic Duchenne's muscular dystrophy patients, which suggests that such antibodies may not be formed as a consequence of heart damage alone. However, that heart damage in a T. cruzi-infected individual might raise such crossreactive antibodies cannot be ruled out. In er,y case, the association of myosin-B13 crossreactive antibodies with CCC uncovered a heart-specific, T. cruzi crossreactive epitope in molecular mimicry with an immunodominant defined protein which may be pathogenetically relevant. Heart-infiltrating T cells

Histiocytes and endothelial cells display increased expression of human leukocyte antigen (HLA) class I and class II molecules; CCC cardiomyocytes only display increased levels of HLA class 1 molecules (Ref. 26). The inflammatory infiltrate is rich in C D 4 ' and CD8 + T cells with a 2:1 predominance of the CD8 ~ Tcell subset 27, The demonstration of restricted heterogeneity of T-cell receptor Va transcripts in heart biopsies from CCC patients 28 is in line with similar findings in established atttoimmune diseases 2'J, Recently, the use of sensitive immunoperoxidase techniques disclosed loci of inflammation around scarce T. cruzi parasites in post-mortem studies of CCC hearts 3°. Authors speculated that the T cells recognizing parasite antigens in situ could be the ultimate effectors of heart tissue damage. However, this hypothesis fails to accommodate their own results showing that the overwhelming majority of microscope fields in CCC hearts display myocarditis in the absence of T. cruzi. In a recent study addressing this issue, T-cell clones have been obtained from heart lesions of a CCC patient. While some CD4 + T-cell clones crossreactively recognized cardiac (but not skeletal) myosin heavy chain and T. cruzi protein B13, like the crossreactive antibodies described above, the clones responded neither to T. cruzi lysate nor to the immunodominant recombinant T. cruzi antigens CRA, FRA, JL5 or B12 (Ref. 31). As heart macrophages are known constitutively to present antigenic myosin peptides on HLA class II molecules 21, myosin-loaded macrophages, rather than HLA class II-negative cardiomyocytes, could trigger the pathogenic CD4 + T cells. If confirmed, such results may indicate that auto398

immunity/ molecular mimicry targets, rather than the direct antigenic stimulus of T. cruzi, are the primary stimuli of the heart tissue-damaging T-cell infiltrate. Conclusion

Human CCC fulfills many of the revisited criteria for autoimmune diseases postulated by Rose and Bona 2°. Were it not for the clear infectious etiology, CCC wo~dd most possibly be assigned as an autoimmune disease, because it matches as many criteria as, for instance, insulin-dependent diabetes mellitus or rheumatoid arthritis. However, the ultimate validation of the candidate target antigens must involve the induction of heart lesions after immunization or passive transfer of antigen-specific T cells. The insights obtained from assigning post-infectious autoimmunc disease status to CCC may help unravel the pathogenesis of classical autoimmune diseases in search of a triggering infectious agent. As in such diseases 32, the identification of the relevant autoepitope in CCC may allow the use of antigen-specific tolerance induction therapy to block the heart lesion process without interfering with the protective immune response against T. crttzi. References

1 Mac6do, V. (1982) in Cecil Textbook of Medicine, 16th edn (Wyngaarden, J.B.and Smith, L.H.,eds), pp 1728-1731,W.B.Saunders 2 Schmufiis,G.A. (1987) Autoimmunity in Chagas' disease. Mere. hist. Oswaldo Cruz 82 (Suppl. 1), 287-310 3 Teixeira, A., Teixeira, M.!:. and qantos-Buch, C. (1975) The Immunology of experimental Chagas' disease: IV. Production of lesions in rabbits similar to those of chronic Chagas' disease in man. Am. J. Pathol. 80, 163-180 4 Kierszembaum, F. (1985) Autoimmunity in Chagas disease: cause or symptom?Parasitol. Today 1, 4-6 5 Witebsky, E, et al. (1957) Chronic thyroiditis and autoimmunization. ]. Am. Med. Assoc. 164, 1439-1447 0 Klein,J. (19t)0)hmmmology, BlackwellScientific

7 Cossio, P.M. et al. (1~)74)Chagasic cardiopathy. Demonstration of a serum gamma globulin factor which reacts with endocard ium and vascular structures. Circulation 50, 1252-1259 8 Khoury, E.I,.et al. (1983) Heterophil nature of EVI antibody in "rt'l/pclllOSOHItl fl'll.?,[ infection. Cli,. Immu,ol. hmmmopathol. 27, 283-288 9 Santos-Bttd~,C.A.et al. (1985)Primary muscle disease: Definition I0

12 13

14

15

16

17

18

of a 25 kDa polypeptide myopathic specific Chagas antigen. Clin. hmmmol, hmmmopathol. 37, 334-350 Wood, I.N. et al. (1982) A monoclonal antibody defining antigenic determinants on subpopulations of human neurons and Trypanosoma cruzi parasites. Natm'e 296, 34-38 Snarv, D. et al. (1983) A monoclonal antibody with specificity for Trypanosoma cnlzi, central and peripheral neurons and glia. Clin. Exp. hmmmol. 54, 617-624 Petry, K. and Eisen, H. (1989) Chagas disease: a model for the study of autoimmune diseases. Parasitol. Today 5, 111-116 Van Voorhis,W., Schelekewy, L. and Trong, H. (1991) Molecular mimicry by Trypanosoma cruzi: the Fl-160 epitope that mimics mammalian nerve can be mapped to a 12-aminoacid peptide. Proc. Natl. Acad. Sci. U.S.A. 88, 5993-5997 Levin, M.J. et al. (1989) Identification of major Trypanosoma cru:i antigenic determinants in chronic Chagas' heart disease. Am. ]. Trop. Med. Hyg. 41,530-538 Bonf,i, E. et al. (1993) Autoantibodies in Chagas' disease: an antibody cross-reactive with human and Trypanosoma cruzi ribosomal proteins. ]. hmmmol. 150,3917-3923 Kerner, N. et al. (199l) Trypanosoma cruzi: antibodies to MAP-like protein in chronic Chagas' disease cross-react with mammalian cytoskeleton. Exp. ParasitoL 73, 451--459 Ferrari, I. et al. (1995)Molecular mimicrybetween the immunodominant ribosomal protein P0 of Trypanosoma cruzi and a functional epitope on the Human I~l-adrenergic receptor. ]. Exp. Med. 182,59-65 Sen6cal, J-L. et al. (1993) Autoantibodies to nuclear lamins and to intermediate filament proteins: natural, pathologic or Parasitolo~ Today, vol. 12, no. I O, 1996

Focus pathogenic? J. Rheumatol. 20, 211-219 19 Ribeiro-dos-Santos, R. et al. (1992) Anti-CD4 treatment abrogates rejection and reestablishes long-term tolerance to syngeneic newborn hearts grafted in mice chronically infected with Trypanosoma cruzi. J. Exp. Med. 175, 29-39 20 Rose, N.R. and Bona, C. (1993) Defining criteria for autoimmune diseases (Witebsky's postulates revisited), hmmmol. Today 14, 426-430 21 Smith, S. and Allen, P. (19~2) Expression of myosin-class II major histocompatibility complexes in the normal myocardium occurs before induction of autoimune myocarditis. Proc. Natl. Acad. Sci. U.S.A. 89, 9131-9135 22 Rizzo, L.V., Cunha-Neto, E. and Teixeira, A. (1989) Specific inhibition of reactivity of CD4 + T cells against myosin in mice chronically infected with Trypanosoma cruzi. Infect. hmmm. 57, 2640-2644 23 Tibbetts, R.S. et al. (1994) Cardiac antigen-specific autoantibody production is associated with cardiomyopathy in Trypanosoma cruzi-infected mice. J. hmmmol. 152, 1493-1499 24 Cunha-Neto, E. et al. (1995) Autoimmunity in Chagas' disease cardiopathy: biological relevance of a cardiac myosin-specific epitope crossreactive to an immunodominant Trypanosoma cruzi antigen. Proc. Natl. Acad. Sci. U.S.A. 92, 2591-2595 25 Unterkircher, C., Avrameas, S. and Ternynck, T. (1993) Autoantibodies in the sera of Tndpanosoma cruzi-infected individuals

with or without clinical Chagas disease. ]. Clin. Lab. Anal. 7, 60-69 26 D'Avila Reis, D. et al. (1993) Expression of major histocompatibility complex antigens and adhesion molecules in hearts of patients with chronic Chagas" disease. Am. J. Trop. Med. Hyg. 49, 192-200 27 Higuchi, M.L. et al. (1993) Immunohistochemicai characterization of infiltrating cells in human chronic chagasic myocarditis: comparison with myocardial rejection process. Virchows Arch. A. Pathol. Anat. 423,157-160 28 Cunha-Neto, E. et al. (1994) Restricted heterogeneity of T-cell receptor variable alpha chain transcripts in hearts of Chagas' disease patients. Parasite hnmunol. 16,171-179 29 Heber-Katz, E. and Acha-Orbea, H. (1989) The V region hypothesis: evidence from autoimmune encephalomyelitis, hnmunol. Today 10,164-169 30 Higuchi, M.L et al. (1993) Correlation between T. cruzi parasitism and myocardial inflammatory infiltrate in human chronic chagasic myocarditis: light microscopy and immunohistochemical findings. Cardiovasc. Pathol. 2,101-106 31 Cunha-Neto, E. et al. Autoimmunity in Chagas' disease: identification of cardiac myosin-B13 Trypanosoma cruzi protein crossreactive T-cell =lones in heart lesions of a chromic Chagas' cardiomyopathy patient J. Clin. Invest. (in press) 32 Tretham, D.E. et al. (1993) Effects of oral administration of type II collagen on rheumatoid arthritis. Science 261,1727-1730

Strategies for the Prevention of Antimalarial Drug Resistance: Rationale for Combination Chemotherapyfor

Nalaria

N.J. White and P.L. Olliaro Among the several 'tropical' diseases that affect humans, malaria poses special control problems due to the i,creasing population at risk from the disease, the difficulties in eradicating the mosquito vector in the tropics and the emergence amt spread of parasite resista,ce to commonly used antimalarial drugs, Their is both clinical experience amt experimental evidence that, however effective when first introduced, the lifespan of drugs is inevitably curtailed by the emergence of resistant parasites. Resistance is the most important factor in determining the useful lifespan of antimalarial drugs. In this review, Nick White and Piero Olliaro discuss the rationale for combination chemotherapy. With the possible exception of artemisinin derivatives (only recently introduced for widespread use in Southeast Asia) 1,2, resistance has emerged to every antimalariaP -7. Crossresistance between compounds of the same class magnifies the problem. Antifolate resistance in both Plasmodium falciparum and P. vivax developed rapidly following the introduction of pyrimethamine in many parts of the tropics. Mefloquine resistance was reported even before the drug i m

Nick White is at Wellcome-Mahidol University, Oxford Tropical Medicine Research Programme, Faculty of Tropical Medicine, Mahidol University, Bangkok 10400, Thailand, and Wellcome Trust Clinical Research Unit, Centre for Tropical Diseases, Cho Quan Hospital, i90 Ben Ham Tu, Quan 5, Ho Chi Minh City, Vietnam. Piero Olliaro is at UNDP/World Bank/WHO Special P,-o~rarnme for Research and Training in Tropical Diseases, 20 Avenue Appia, 1211 Geneva 27, Switzerland. Tel: +41 22 191 3734, Fax: +41 22 791 4854, e-maih [email protected] PardSitolo~ Toddy, vol. 12, no. I0, • 9 9 6

copings-©1996. ElsevrerSoenceLtd

had been routinely used 8,'~, and has now severely curtailed the usefulness of the drug in some parts of Southeast Asia m,11. Halofantrine resistance has emerged concomitantly 12-14. When atovaquone was used alone ill preliminary clinical trials, 28% of patients had recrudescent infections associated with the development of resistance 15. Quinine and chloroquine have enjoyed longer periods of effective use, but, in large parts of the world, chloroquine is no longer effective treatment for faiciparum malaria 5, a n d P. vivax has developed resistance in some parts of Oceania 1°,17. Quinine, too, is threatened in some parts of the world 18. While waiting for new drugs with novel mechanisms of action to be discovered, developed and hopefully deployed, appropriate measures should be taken to safeguard the few compounds available to us. The development of resistance Operational strategies for preventing resistance to insecticides and antibacterials ~eg. the regular rotation of agents to reduce the selection pressure) have not been employed in malaria because of the paucity of drugs and the difficulties in regulating their u:~e. The operational factors that may affect the development of resistance include appropriate deployment, sensible prescribing, and patient treatment-seeking behaviour and compliance with the prescribed regimen 19-21. Ideally, drugs should not be made available until they are needed. In practice, it is not possible to regulate the use of drugs that are available freely for purchase in the rnarket-place. All nghts reserved 016q- 475~196,'$1500

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399