Antifolic combinations in the treatment of malaria*

Bull. Org. mond. Sant) 1974, 50, 223-230Bull. Wld Hlthi Org. Antifolic combinations in the treatment of malaria* L. DONNO1 For the treatment of malaria, combinations of drugs with antifolic action have the great advantage, compared with other drug associations, of synergic action, which increases the effectiveness of the preparation, limits its toxicity, and reduces the risk of resistance. The associations of a sulfonamide (sulfalene, S) with a diaminopyrimidine derivative (pyrime- thamine, P, or trimethoprim, T) have given good immediate clinical results. An analysis of the pharmacodynamic andpharmacokinetic characteristics ofthe three components explains the relative ease with which relapse is encountered with the association ST. The association SP is, from this point of view, more logical and effective and its importance in prophylaxis is surveyed in the light ofa critical discussion of the arguments that are at present throwing doubt on the usefulness of its wider employment. The resistance of Plasmodiumfalciparum to chloro- quine in some areas of the world (1, 2, 3) has revolu- tionized a standard scheme of treatment and given rise to a proliferation of drug associations of an empi- rical nature, the use of which has been justified perhaps only by the disturbing situation and the urgency of tackling it. It is noteworthy that in these combinations quinine-which seemed to have lost favour with the arrival of the 4-aminoquinolines- has regained prominence. The subject has been dealt with extensiveJy in the literature, and in particular by Peters (1). In any case, such associations have only an additive effect. Nevertheless, the practical importance of syner- gism in an antibacterial or antiparasitic drug association is well known. It is sufficient to mention the enhanced therapeutic activity, the reduced risk of toxicity, and the weakening of resistance. The association between sulfonamides or sulfones on the one hand and diaminopyrimidine derivatives on the other is characterized precisely by a high degree of synergism due to the well-known phenomenon of sequential blocking. PHARMACODYNAMIC AND PHARMACOKINETIC CONSIDERATIONS Not all sulfonamides have the same antiplasmodial activity, as Chin et al. (4) and Walker et al. (5) have * Presented at the Symposium on Malaria Research, Rabat, Morocco, 1-5 April 1974. 1 Pneumological Clinic, University of Milan, Italy. Pres- ent address: Department of Pneumology and Respiratory Physiopathology, Cuasso al Monte Hospital, Varese, Italy. observed. Furthermore, according to Struller (6), the antibacterial action of a sulfonamide in vitro is proportional to its biological half-life, so that, in equal concentrations, long-acting sulfonamides would appear to be more active than short-acting sulfona- mides. Diaminopyrimidine derivatives, whose me- chanism of action depends on the inhibition of dihydrofolate dehydrogenase, also show significant differences in their capacity to combine with enzym- ic receptors of different species. Thus Burchall & Hitchings (7) demonstrated that trimethroprim, pyrimethamine, and amethopterine have a binding affinity with the dihydrofolate dehydrogenase in bacteria, plasmodia, and animals, respectively. It should be remembered, however, that the therapeutic activity of any drug-and particularly that of an antibacterial or antiparasitic drug- depends not only on its pharmacodynamic action but also on its pharmacokinetic action. It therefore seems opportune to recall briefly the pharmacokinetic characteristics of sulfalene, pyrimethamine, and trimethoprim, which the author used in the treatment of P. falciparum malaria. Sulfalene is a long-acting sulfonamide with the following principal characteristics. (1) High invasion constant with oral administration. Intestinal ab- sorption is rapid, the highest concentrations in the blood being reached between the fourth and sixth hours. (2) Low renal elimination constant. The drug remains in the circulation for a long time owing to its high liposolubility, which gives rise to a strong tubular reabsorption. The biological half-life of sulfalene is approximately 65 h. (3) Low protein 3182 - 223 L. DONNO binding (about 60 %), particularly in comparison with other long-acting sulfonamides. There is thus a high concentration in the tissues or organic liquids since only the fraction of the drug that is freely dissolved in the aqueous part of the plasma is able to spread through the tissues. In other words, the antibacterial (or, in this case, the antiplasmodial) activity of the sulfonamide does not depend on its total concentration but only on the fraction that is not bound to the plasma proteins. Trimethoprim is a diaminopyrimidine derivative that acts as an inhibitor of dihydrofolate dehydro- genase and has the following pharmacokinetic characteristics. (1) High invasion constant reaching the highest concentrations in the blood within the second hour. (2) A constant of elimination that is relatively low though appreciably higher than that of sulfalene; its biological half-life is in fact about 13 h. (3) Protein binding: 31 %. Pyrimethamine is another diaminopyrimidine derivative that acts as an inhibitor of dihydrofolate dehydrogenases but has poorly defined pharmaco- kinetic characteristics. Smith & Ihrig (8) have shown it to have a high invasion constant, the highest concentrations in the blood being reached within the fourth hour, and a biological half-life only slightly longer than that for sulfalene (about 72 h). There seems to be no information in the literature on the protein binding value. The concentration levels in the blood observed by Brooks et al. (9) are questionable because their behaviour is in conflict not only with those of Smith & Ihrig but also with the process of renal excretion, which normally follows an exponen- tial regression curve. This type of curve is exemplified in Fig. 1, which shows the blood concentrations observed during the course of time after a single, but not simultaneous, dose of each of the following drugs has been ad- ministered: sulfalene (1 g), pyrimethamine (0.1 g), and trimethoprim (0.5 g). The curves, which were plotted on semilogarithmic paper, do not record the real values as there are considerable differences in the blood concentrations obtained with the doses of the three drugs administered. Thus they have been plotted taking the maximum plasma peaks to be equal to 100 (S=81 mg/1; T=6.9 mg/I; P=l.1 mg/l), the other values being expressed as percentages of the peak values. In this way comparison is easier and shows more clearly, on the one hand, the similar be- haviour of sulfalene and of pyrimethamine and, on the other, the early disappearance of trimethoprim from the circulation, thereby interrupting the synergic action. The pharmacodynamic interaction of sulfonamide and diaminopyrimidine derivatives is well known and, as mentioned above, it results in synergism. It would also be interesting to know the pharmaco- kinetic interaction-that is, the mutual influences of, for example, the protein bond, the invasion constant, and the elimination constant. The study of the inter- action between drugs began only recently, but has already yielded results with considerable practical implications. In the field of antimalarial drugs, for instance, mepacrine and pamaquine establish a firm bond with liver proteins. Consequently, when pamaquine is administered shortly after mepacrine, binding with hepatic proteins is extremely limited. In these conditions, as noted by Morelli et al. (10), a therapeutic dose of pamaquine, which usually is well tolerated, may exert toxic effects on the gastrointestinal tract and haematopoietic system. Blount (11) demonstrated competition between quinine and pyrimethamine in regard to binding with plasma proteins. As a result, severe toxic effects, Fig. 1. Blood concentration curves after a single dose each of sulfalene (S), pyrimethamine (P), and trimethoprim (T) . 224 ANTIFOLIC DRUGS AGAINST MALARIA such as cinchonism and neutropenia, may be induced by the quinine. No studies on the interaction of these combinations appear to have been reported in the literature. Moreover, with regard to the dosage of pyrimethamine, the chemical methods so far avail- able are not sufficiently sensitive to reveal the low concentrations obtained in man with the therapeutic doses employed, whereas microbiological methods give misleading results owing to the simultaneous presence of the sulfonamide. The author has only recently learned of an extremely sensitive gas chromatographic method and his research based on this method will be reported as soon as possible. CLINICAL RESULTS Much clinical research has been carried out so far on the two combinations sulfalene with pyrimeth- amine (SP) and sulfalene with trimethoprim (ST). The results of the studies known to the author are listed in Tables 1 and 2, which show the immediate therapeutic effects, based on a briefperiod of observa- tion (5-7 days). As may be seen, the results with both forms of treatment were satisfactory, parasitaemia being cleared in a relatively short period (2.24 days on the average) in approximately 98 % of patients. In the field of antimalarial drugs, speed of action is a very important factor in view of the damage that may be caused by the intensity and persistence of plasmodia in the human body, particularly when immunity is lacking. Fig. 2, based on research carried out in Nigeria by Donno et al. (12), gives the average percentage reduction in the number of parasites in circulation and the percentage of patients cleared of parasitaemia observed daily in three groups of patients suffering from malaria caused by P. falciparum and treated with chloroquine (CL), the association SP, or the association ST. The results were similar; the para- sitaemia fell to practically insignificant levels within 2 days and the proportion of patients cured of parasitaemia by the end of the second day was 56.7% in the chloroquine group, 70% in the SP group, and 60% in the ST group. By the fifth day these per- centages had risen to 90, 93.3, and 100, respectively. Table 1. Results with sulfalene-pyrimethamine treatment No. of Recovery No. of days until Author Country Plasmodium patients Failure disappearance No. % fever parasites Mazzoni (26) Somalia P. falciparum 49 49 100 0 1.88 2.65 Catarinella (27) Cameroon P. falciparum 31 31 100 0 2.64 2.48 (P. falciparum Rey et al. (28) Senegal 21 21 100 0 1.40 1.80 P. malariae (P. falciparum Rey et al. (29) Senegal 49 45 92 4 2.00 2.00 P. malariae Donno et al. (12) Nigeria P. falciparum 52 48 92 4 1.64 2.16 Donno et al.a (30) Nigeria P. falciparum 20 18 90 2 2.97 3.16 Cambodia Mazaud et al.b (31) P. fakciparum 100 97 97 3 3.00 2.90 Cameroon Catarinella & Donno (32) Cameroon P. falciparum 140 140 100 0 1.88 2.16 ( P. falciparum Togo Donno & Ricciardi (33) P. malariae 136 136 100 0 2.04 1.72 Congo P. ovale Storey et al.C Nigeria P. falciparum 213 213 100 0 - - Total or average 7 3 811 798 98.3 13 2.15 2.24 a Nonimmune patients. b Seven nonimmune patients. c Unpublished observations, 1972. 225 Table 2. Results with sulfalene-trimethoprim treatment No. ofRecovery ~~~~~No.of days untilAuthor Country Plasmodium patients Failure disappearance No. % fever parasites Martin & Arnold (34)a USA P. falciparum 19 18 95 1 2.59 2.14 Donno et al. (30)b Nigeria P. falciparum 22 22 100 0 2.96 2.98 Donno et al. (12) Nigeria P. falciparum 30 30 100 0 1.67 2.29 P. fakciparum Rey et al. (28) Senegal 28 26 93 2 2.80 1.60 P. ovale Donno et al. (16) Cameroon P. falciparum 51 51 100 0 2.04 2.20 Martin & Arnold (17)C USA P. vivax 9 9 100 0 1.20 2.50 Canfield et al. (13)C Viet Nam P. falciparum 36 35 97 1 - - Chin et al. (14)d Thailand P. falciparum 45 44 97.8 1 2.40 2.20 Total or average 6 3 240 235 97.8 5 2.28 2.23 a Nonimmune volunteers. b Nonimmune patients. c Nonimmune volunteers, repeated doses. d Four delayed relapses ( ?). e Nonimmune patients, nine relapses. Yet a statistical comparison of the three types of treatment showed no significant differences. However, as regards the results presented in the tables, it is noteworthy that both Canfield et al. (13) and Chin et al. (14) noted a certain number of 100, 90O 80- 70- 60- 50- 40. 30. 20. 10. 0 relapses after treatment with ST, thus confirming the observations of Clyde (15) in a group of volun- teers. There have been no conclusive reports of such relapses in subjects treated with SP, despite follow-up by certain authors. 1_ I I1 2 3 4 5 II 2 3 4 5 Days Fig. 2. Daily rate of parasitaemia clearance (A) and recoveries (B) in patients treated with chloroquine (CL), SP, or ST. A B /0111 /-O I~~~~~~~~~~~~~I 'A ~~~~~~~~~~~~~~~~~~~~~~~~I! I . I I~~~/ I I, 1.%~~~~~ 9~ I 4 / 226 L. DONNO 10, aY IE ANTIFOLIC DRUGS AGAINST MALARIA DISCUSSION To assess the effectiveness of an antimalarial product it is useful and necessary to consider various aspects of the pathogenic agent (variety of plasmodial species and of their strains), the patient (different degrees of immunity), and the drug (side-effects, methods of administration, pharmacokinetic charac- teristics). As regards the pathogenic agent, both the com- binations considered here were mainly employed against malaria caused by P. falciparum; however, their effectiveness was satisfactory also in cases caused by other plasmodial species, although slightly less so than that of chloroquine in that defervescence and parasitaemia clearance took longer. The thera- peutic efficacy did not vary with the plasmodial strain but remained constant in the many countries in which research was carried out. It should be recalled in this context that, whereas strains resistant to pyrimethamine are particularly frequent in Africa, those resistant to chloroquine are more frequent in Asia. As far as the patient is concerned, the importance of the state of immunity on the effect of an anti- malarial drug is well known, as also the fact that in populations exposed to the disease there exists a certain ratio between the degree of immunity and the age of the patient. Both the drug associations under discussion have been used in patients whose ages ranged from 1 month to over 50 years, and children under the age of 2 years are well represented in almost all the studies. Even in definitely non- immune subjects treated with the same dosages as those used for semi-immune subjects, the results have been favourable, with a high proportion of clinical and parasitological cures. In the few cases in which the parasitaemia was not completely cleared within the brief observation period, the number of parasites in circulation was strongly reduced. With regard to the drug, all workers who have used SP and ST have stressed that these drug com- binations were well tolerated and did not give rise to serious side-effects. Storey et al. (unpublished report to WHO, 1972), in particular, state that SP gave better results from this point of view than did chloroquine combined with pirimethamine. If the pharmacodynamic and pharmacokinetic characteristics of the components of these drug combinations are analysed, it becomes evident that SP is a more logical combination than ST. It is likely that the special affinity of pyrimethamine with plasmodial dihydrofolate dehydrogenases and the fact that sulfalene and pyrimethamine have the same biological half-life ensure greater activity and an enduring and constant synergic action, with all the implications that this involves as regards both effectiveness and resistance. An explanation for the tendency towards relapse observed in treatment with ST (without taking into account differences in the sensitivity of various strains) may lie in the observa- tions of Martin & Arnold (17). These authors used ST in the treatment of malaria caused by P. vivax, obtaining poor results with a single administration and good results with repeated administration -probably because the latter method prolonged the synergic action. It is likely-though as yet unconfirmed-that the antifolic combination exerts a plasmodicidal effect analogous to the bactericidal effect of the sulfa- methoxazole-trimethoprim combination. On the other hand it is known that a sulfonamide alone, which is bacteriostatic in low concentrations, becomes bactericidal in high concentrations. The antifolic combination certainly affects the primary tissue forms and the asexual erythrocytic forms; its sporontocidal activity, although still debated, is probable. Apart from the 8-amino- quinolines, whose use is limited by their high toxicity, this type of combination has a wider spectrum of action than has any other antimalarial drug in use. The fundamental characteristic of SP-the possibility of curing the patient with a single dose-is highly important because malaria occurs chiefly in countries in which the social, economic, and health conditions make it necessary to give out-patient treat- ment. In such conditions, treatment of several days' duration fails because the patient, benefiting from the first dose, does not return for further treatment. He then often becomes a carrier of plasmodia, is subject to relapses, and causes the disease to spread further. The advantages of treatment with SP are thus obvious, and in this context the figures of Michel (18) are encouraging. By administering a single dose every 2 weeks, he obtained a remarkable reduction in the parasite and gametocyte indexes. The extensive prophylactic and therapeutic use of the antifolic combination in the field is open to question, and many authors have adopted a cau- tious attitude towards it. But until the ideal anti- malarial drug appears, the characteristics of which have been most clearly described by Bruce- Chwatt (19), the arguments against a wider use of 227 L. DONNO this drug combination might be reconsidered in the light of the following remarks. Risk of toxicoallergic manifestations Particularly in the case of prolonged administra- tion, the risk of such side-effects is certainly not high. Pyrimethamine has been widely used in the pro- phylaxis of malaria, at doses higher than those envisaged in this combination. Moreover, its side- effects are easily controlled by folinic acid. The great danger in the use of long-acting sulfonamides is said to be the possible occurrence of the Stevens- Johnson and Lyell syndromes. However, without going too far into discussions that are not relevant to this paper, it may be recalled that the International Congress of Chemotherapy, held in Vienna in 1967, showed the error of this belief. On the other hand, alkali treatment considerably speeds up the elimina- tion of sulfonamides. According to research carried out by Flatz et al. (20) both in vitro and in vivo, the use of sulfalene does not seem to entail the possibility of haemolitic crises in subjects who are deficient in glucose-6-phosphate dehydrogenase. Moreover, any form of treatment with drugs in- volves some risk of undesirable consequences. Chloroquine itself-to quote an example from the field of antimalaria treatment-was initially rejected because it was believed to be toxic, and it is still not exempt from suspicion. Everyone knows of its retinotoxic action in the course of massive, prolonged treatment of some forms of mesenchymopathy. Less well known, perhaps, is the fact that weekly anti- malarial prophylaxis, when prolonged for a few months, can cause an accumulation of chloroquine in the pigmentary structure of the human eye-an accumulation that is still evident years later, as has been observed by Lawill et al. (21). Risk ofplasmodial resistance There is no doubt that such resistance may arise easily and rapidly when the sulfonamide and di- hydrofolate dehydrogenase inhibitor are administered separately. However, as has been demonstrated, among others, by Thompson et al. (22) in the experi- mental infection with P. berghei and by Lucas et al. (23) in natural human infection by P. falciparum, resistance appears much more slowly and weakly when the two components are administered together. The alarming observations of Verdrager et al. (24) are hardly surprising, since resistance to harmful agents in general and drugs in particular is still the rule among living beings and has caused the survival of species through the ages. It will perhaps be possible to reduce the risk still further by adding acridinic derivatives to the association, according to the promising research of Peters (25), but it must be iemembered that the phenomenon of resistance has affected the 4-aminoquinoline derivatives; neverthe- less, they continue-rightly-to be used in chloro- quine-sensitive cases. Risk of bacterial resistance Sulfonamides are still widely used in under- developed countries, particularly against cerebro- spinal meningitis. Research into other diseases has shown bacterial resistance to sulfonamides to be a real risk. Thus, during the above-mentioned study (23), the strains of Escherichia coli selected were sulfonamide-resistant in about 30% of cases. In the therapy of meningococcal infection, also, the use of sulfonamides is declining in favour of peni- cillin; on the other hand, it occupies an important place in the prophylaxis not only of malaria but also of numerous other sulfonamide-sensitive diseases, such as trachoma, urinary and intestinal infections, and meningitis itself. The results of my experience in Africa suggest that the widepread prophylactic action of sulfon- amides will compensate for the risk of resistance. Moreover, many of the antibiotics now available are equally, if not more, active and are cheap enough to use in emergencies. Furthermore, the extended use of sulfonamides would probably limit the indiscri- minate use of antibiotics. It might also help to ward off, in those countries, the reversal of the bacterial ecology that has resulted in the emergence of poly- resistant strains and of Gram-negative micro- organisms of low sensitivity-the cause of so many problems in the developed countries. RESUME ASSOCIATIONS D'ANTIFOLATES DANS LE TRAITEMENT DU PALUDISME Les associations de sulfamides et d'inhibiteurs de la dihydrofolate r6ductase dans le traitement du paludisme offrent l'avantage, par rapport aux autres types d'asso- ciation, d'agir de facon synergique. L'analyse des pro- prietes pharmacodynamiques et pharmacocinetiques de trois preparations antipaludiques indique que l'emploi 228 ANTIFOLIC DRUGS AGAINST MALARIA 229 de l'association sulfal&ne-pyrimethamine (SP) est plus logique que celui de I'association sulfalene-trimethoprime (ST). La pyrimethamine, en effet, fait preuve d'une affi- nite plus grande pour les dihydrofolate reductases plasma- tiques et sa demi-vie biologique est d'une dur&e tres voisine de celle du sulfalene. Les resultats cliniques obtenus avec les deux types d'association sont tres satisfaisants, le sang etant debar- rasse des parasites dans 98% environ des cas traites. Dans le groupe des malades recevant l'association ST, cependant, on note une certaine tendance aux rechutes; ce fait doit probablement etre attribue a l'interruption de l'action synergique par suite de l'e1imination precoce de la trimethoprime de la circulation. L'emploi de l'association sulfalene-pyrimethamine pour le traitement de l'acces de paludisme est maintenant lar- gement admis. I1 persiste un leger doute en ce qui regarde son utilisation prophylactique. I1 semble neanmoins que les craintes emises a ce sujet ont ete exagerees et que les avantages de son emploi gen&alise l'emportent sur les inconvenients eventuels. REFERENCES 1. PETERS, W. Chemotherapy and drug resistance in malaria. London and New York, Academic Press, 1970, p. 528. 2. PETERS, W. Transactions of the Royal Society of Tropical Medicine and Hygiene, 63 (1): 25 (1969). 3. YOSHNAGA, T. et al. Arzneimittel Forschung, 20 (9): 1206 (1970). 4. CHIN, W. et al. American journal of tropical medi- cine and hygiene, 15 : 823 (1966). 5. WALKER, A. J. & LOPEZ ANTUNANO, F. I. Trans- actions ofthe Royal Society of Tropical Medicine and Hygiene, 62: 654 (1968). 6. STRULLER, T. Progress in drug research, 12: 389 (1968). 7. BURCHALL, J. J. & HITCHINGS, G. H. Molecular pharmacology, 1: 126 (1965). 8. SMITH, C. C. & IHRIG, J. American journal of tropical medicine and hygiene, 8 (1): 60 (1959). 9. BROOKS, M. H. et al. Clinical pharmacology and therapeutics, 10: 85 (1969). 10. MORELLI, H. F. & MELMON, K. L. Californian medi- cine, 8: 248 (1961). 11. BLOUNT, R. E. Archives of internal medicine, 119: 537 (1967). 12. DONNO, L. et al. American journal of tropical medicine and hygiene, 18 (2): 182 (1969). 13. CANFIELD, C. J. et al. American journal of tropical medicine and hygiene, 20 (4): 524 (1971). 14. CHIN, W. et al. American journal of tropical medi- cine and hygiene, 22 (3): 308 (1973). 15. CLYDE, D. F. Journal of the American Medical Association, 209 (4) : 563 (1969). 16. DoNNO, L. & CATARINELLA, G. Journal of tropical medicine and hygiene, 74: 246 (1971). 17. MARTIN, D. C. & ARNOLD, J. D. Journal of clinical pharmacology, 9 (3): 155 (1969). 18. MNCHEL, R. Medicine tropicale, 28 : 488 (1968). 19. BRUCE-CHWATT, L. J. Transactions of the Royal Society of Tropical Medicine and Hygiene, 61 (3): 412 (1967) 20. FLATZ, G. et al. Klinische Wochenschrift, 48 (2): 2 (1970). 21. LAWILL, T. et al. American journal of ophthalmology, 65: 530 (1968). 22. THOMPSON, P. E. et al. American journal of tropical medicine and hygiene, 14: 198 (1965). 23. LucAs, A. 0. et al. Transactions of the Royal Society of Tropical Medicine and Hygiene, 63: 216 (1969). 24. VERDRAGER et al. Medecine tropicale, 28: 663 (1968). 25. PETERS, W. Nature, 223: 858 (1969). 26. MAZzoNI, P. L. Archivio Italiano di Science Mediche Tropicali e di Parassitologia, 7-8: 215 (1967). 27. CATARINELLA, G. Gazzetta Medica Italiana, 126 (12): 478 (1967). 28. REY, M. et al. Bulletin de la Societt me'dicale d'Afrique noire de langue franCaise, 13 (2) : 366 (1968). 29. REY, M. et al. Bulletin de la Socie'te medicale d'Afrique noire de langue fran9aise, 13 (4) : 959 (1968). 30. DoNNo, L. & SANGUNETI, V. Chemotherapy, 15 (2): 118 (1970). 31. NMAZAUD, R. Medicine tropicale, 30 (6): 759 (1970). 32. CATARINELLA, G. & DONNO, L. Journal of tropical medicine and hygiene, 74: 243 (1971). 33. DoNNO, L. & RICciARDI, M. L. In: Proceedings of the Intemational Seminar on Parasitic Diseases of Social Importance in Latin America, Rome, 18-21 October 1971. Rome, Istituto Italo-Latinoamericano, p. 213. 34. MARTIN, D. C. & ARNOLD, J. D. Journal of the American Medical Association, 203 (7): 134 (1968). DISCUSSION DoNNo: The focal point of this discussion is the possible widespread use of antifolic combinations in the treatment of malaria. My opinion on this is stated in the conclusions of my paper. When penicil- lin was discovered, resistance to sulfonamides was frequently reported, and for this reason sulfonamides L. DONNO fell out of favour. But resistance to antibiotics is now increasing, whereas that to sulfonamides is decreasing. In Africa there is no chloroquine resistance at present. It may however appear in the near future. Many doctors in Africa have reported a decrease in the effectiveness of chloroquine against falciparum malaria. Antifolic combinations should be used also in Africa because they are very practical (one dose per month in prophylactic mass campaigns) and because their use would decrease the demand for chloroquine and therefore perhaps delay the appear- ance of chloroquine resistance. FERNEX: The synergistic effect of sulfonamides with antifols was first described in the late 1940s. Since 1968, six or seven different drugs based on this mode of action have been available on the market. For these combinations, the choice of the indication is important: some are specifically coccidiostatic and others are specifically active against bacteria or plasmodia. The combination of sulfamethoxazole and tri- methoprim, which is the most active antibacterial drug of this group, is relatively less effective against malaria. The dosage must be high or administration must be repeated over many days. Its cost is there- fore many times higher than more specific anti- malarial combinations that can be given as a single dose. The value of sulfadoxine-pyrimethamine for the oral or parenteral treatment of acute drug-resistant falciparum malaria is undenied. This combination has potential value as a suppressive drug, as is shown by data accumulated in South-East Asia and Africa on the prophylactic use of " Fansidar" among 4 487 subjects. With varying dosage schedules, rang- ing from 1 tablet of " Fansidar " weekly to 3 tablets monthly for periods ranging from a few weeks to two years, ten investigators observed a total pro- tection rate of 98 %. Positive slides were found mostly after 6 weeks, when one or several doses had not been taken. My conclusion is that " Fansidar " is an effective prophylactic in areas where chloroquine resistance is present. LUZZATO: Several hundred patients with various underlying chronic medical problems requiring long- term protection from malaria have been followed up at regular intervals at a haematological clinic in Ibadan. Observations accumulated on approximately 10 000 patient-years show the use of daily proguanil to have been extremely effective. When rare break- throughs occur, the infections respond promptly to a standard curative regimen of chloroquine. MICHEL: In Senegal, the efficacy of mass prophylac- tic treatment of children aged 1-14 years with sulfalene-pyrimethamine has proved to be similar to that of chloroquine. In areas where strains are resistant to proguanil or the injectable repository drug 501, sulfalene-pyrimethamine treatment has given good results. BRUCE-CHWATT: I am surprised to note from Profes- sor Donno's paper that no occurrence of haemolytic crisis was noted following the use of sulfalene in patients with G6PD deficiency. EDESON: No indication of the density of parasit- aemia is given in Table 2, which shows the clearance time following treatment with sulfalene-trimetho- prim as reported by various investigators. DoNNo: Complete data on density are reported in the original papers cited. In some of the cases observed, a parasitaemia level of 400 000 per mm3 was cleared within 24 h, whereas, in other cases with lower parasitaemia, the clearance time was longer. 230