Potential antimalarial substances. Antimetabolites of pantothenic acid

a deficiency in pantothenic acid (la).3 These observa- tions stimulated a search for antimetabolites that might interfere with the utilization of pant...
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Yeptemher 19tiS

SOTE:,

activity of both acetylcholine and carbachol. The dose-response curves of acetylcholine ( -11) antagonized b) 1 or 2 were identical within experimental error. The ED,, for both 1 and 2 waq 0.07 mg'ml (ea. -I x 10-4 -11). Previous workers have studied a~etylcarbocholine~ and ticetylsilicocholiiiej and have concluded that these two compounds act as indirect, nonnicotinic, cholinergic :igents. Thiq stud) implies that both 1 and 2 have :in affinity for the mu-c:irinic qite hut (10 not powes:.; intriii4c [ictivitj..

at least five times duriiig and at the eiid of the assays of the test compoiirids. The average of the3e valries wa$ takeii as the 100% ctrritraction valiie for 10-7 .IT acetylcholiiie for the particular miiscle. The t,est compokinds were assayed in essentially the Sam? manner. The initial bath volume was 28 ml. T o this was added 1.0 ml of the test, solution followed within about 15 sec by 1.0 ml of the acetylcholine solution (final bath concent,ration, 10-7 I\. ACh). The process was repeated three times and the average of the peii deflections wab taken as the value for that particiilar coiiceritrat,ion of test rompoiiiid. This value was then expressed as a per cent of the c-oittractioii valiie for lo-' J I ACh. At least foiir animal, were iised for each cvmpoiuid. The data that were obtaiiied are siimmarized i n Figiire I . The test rompiiiiiids aloiie gave i i o niiisc-le respoiise in the dose i'aiige of 0.30 to 2..i x 10-3 mg nil.

Experimental Section The procediire described hy TiiriierGwas followed for bioassay piirposes. The bath tempemtiire was maintained at 3 i i l o , arid ox>-geiiated Tyrode's soliitioii was iised as the perfiiriiig fliiid. IIexamethoiiiiini bromide (10 nig 1.) was added to the Tyrode'- soliitioii t o preveiit gaiiglioiiica respoiise. The filial bath voliime for each rim was 30 ml aiid chemical concentrations were rnlciilsted accordiiigly. The teht rompoiiiids coiild only be dissolved iii Tyrode's soliitioii with ihe aid of ethaiiol. Ilelivery of 1 ml of the test >oliiiiiiii to 20 ml of tiath gave a final concetitratioii of l.6yc ethaiiol. This did i i o t have a sigiiificaiit iiifliieiice oii the miiscle respoiise iiidiic-ed hy 10-' J I aretylcholine, A typiral rtiii was carried oiit i l l the following manlier. The ileiim was disw'ted, then washed with Tyrode's solution. -4pproximately .i cam of the niriwle was attached to a transducer in the miiscle bath (29 nil) aiid allowed to stand iiiitil the spontaiieoii: cwntract.ioiis had siihsided. The transducer was connected to a strip chart recorder. ilcetylcholiiie chloride sollition (1 ml) (final bath roiicaeiitratioii lo-' JI, 1.6% ethanol) was delivered to the bath. The amplitiide of the strip chart recorder was adjiisted b o that the resiiltirig coiitract.ioii gave a i 0 deflectioii. The mwcle was then washed coiitiniioi Tyrode's soliitioii iiiitil the miiscle had relaxed. The wash soliitioii was drained, f fresh Tyrode's rolritioli was added t o the bath, mid the was allowed t u eqiiilibrate. After the pen tleflecatioii was adjiistedl the c.alibratioii was repeated

DOSE RESPONSE CURVE FH3

A CH,FCH,CH,OCNH,

R

CH,

0

1071

Potential Antimalarial Substances. Antimetabolites of Pantothenic Acid' ED^

IRD

F. E L ~ L L G CX R ~ R, L - \ \ D P HLTT, \ ? D L C ~ L I 11. E ~~ERl3EL

Research 1,ahoialorzes. P a i k e . Daiic and Company, Ann .-libor, Michiqan .@lo6 Recezced llarch 2.5. 1968

I n 1943 it was observed that the survival of the erythrocyt'ic stages of Plasmodium l o p h u m e , maintained intracellularly in duck red cell suspensions in citro, was favored by the addition of calcium pantothenate t o the medium.? lloreover, the development of Plasinoclium qallinaceunz in chickens is inhibited by a deficiency in pantothenic acid These observations stimulated a search for antimetabolites that might interfere with the utilization of pantothenic acid by plasmodia. 3 - 7 Among the antipailtothenates that were synthesized and tested earlier,3 - 7 D-( +)-phenylpantothenone (Ib) vias equiactive with quinine against P.galliiraceunz and twice as potent as quinine against P . lophurae in the c h i c l ~ ~ 1In 7 m:m, I b exhibited slight act'ivit>y against blood-induced vivax malaria and was tolerated well in doses of 2 g daily for 4 days.5 The pantoyltnuriiie derivatives T-13 nnd YIb n-ere the CH? ~IOCII,CCHOHCO?;II(CTT,)ili I

CH3 D-(

Ia, R b, R

= =

+)

coori

COC6Ilj

most active compounds synthesized; both were approximately ten times as pot'ent as quinine against P. gallirruceuin in chicks.jSfi More recently it was shown that pantothenic acid has no effect 011 the erythrocytic stages of P. lophume 0 10

0 05

0 01

BATH CONCENTRATION (mg/ml)

Figure 1. (4) (a) .J. Raaister a n d V. P. Whittaker. .\-atwe, 167, GO5 (1951); (11) A . S. Y. Biiram, Brit. .I. Plinrmncol., 25, 4 (1965). ( 5 ) P. T i l . Henderson. E. J. .\riens, B. TT-. J . Ellenbrock, a n d A . .\I. Simonie, .I. i ' l t i t r m . i'liarmacol., 20, 26 (1968). (6) K . A Turner, "Screening Methods in Pharmacology," Academic Press Inc., Ke,v Y'urk, N . Y.,1965, p p 42-M.

(1) This investigation was supported 11y U a n d Development Command Contract D. Contrihution KO.360 to t h e h r m y Research Program on hlalaria. (2) W , Trager. J . E x p . M e r ! . , 77, i l l (1943). (3) S. Brackett, E. Waletaky, and .\I. Baker, J . F u r o ~ i l o l . ,32, 453 (1945). (4) J. F. Mead, 11. 11. Rapport, .I. E. Senear. J . T . AIaynard. and B i d . Chem., 163, 16.5 (1946). selople, ".I Sur\-ey of .Intimalarial Driigs, 19-11-1945," Vol. I , J. T. Edwards, .\nn Arlmr, 5Iich.. 19.16, p p 138-140. (6) R . \Vinterl)ottom, ,J. \V. Clapp, 11.. H . lliller. J . P. English, a n d R . 0 . Roblin, ,Jr., J . A m . Chem. S O C . , 69, 1393 (1947). ( 7 ) R . E. L u t z , 3. JV. \Vilson, 111, h. J . Deinet, G . H. Harnest, T. -i. Martin, and J. A. Freek, J . U r g . Chrm., 12, 96 (1947).

'1'\lil,I,,

s

I11

v

VI, D - ( + ) form a, X, Y = H b,X,Y=4-CI C, X, Y = 3,4-C12 d , X, Y 3-CF3

e, X , Y =4-CF? f,X,Y=3,5-(CFz)Z g, X, Y = 4-SCH7

h, X, Y = 4-OCHACsHs

lined previousl! l 1 were gerierall\ satisfactory. However. the Iiydraziriolysii of 11-d (X, T = H, 3-CF1) (Table I ) gave a poor yield of "amiiio-R'-(trifluoronncthj ljethanesulfo~~a~~ilide (\7d) (Table 11) and the sequence TI i i i not completed. I:urther, the coridensat i o i t of 1-f jX, 1 = :3,>-(C13)2] with I)-( -)-pantolactone (1

I

September 196s

1073

SOTER

TABLE I1 2 - A M I N n E T H . i ~ E S v L F O N . ~ ~ I L I D E(T') S

No.

x, Y

Yield punned, Mp,

70

O C

Vb 4-C1 160-162a 66 VC 3,4-C1, 157- 139 64 VJ 3-CF1 134-136 3 1.e 4-CF3 163-167 76 262-263 der 44 \'i 3 , ,?-( CFa)? c1.g 4-SCH3 162-16.5 \h 4-0CIT2c61Ij 211-214 94 a Lit.6 mp 160.3-161.5". These intermediates were riot analyzed. next step. Free base, mp 178-181".

I

HOCH2-C-CHOHCONH(CH2)2S02NH

I

CH?

x. Y

SO.

TIb

Mp, "C

4-C1

103-104h TIC 3,4-C12 141-143 \ Ie 4-CF3 144-146 4-SCHa 112-1 14 VIg T Ih 4-OCH2CsHj 142-144 Lit.6 mp 101-103° from C6fI6. c 1, 95yo EtOH.

Yield purihed.

Purifn

L?c

so11 ent

.j9

C6156 CHCL

[m]23D,'c

der

Formula'

+40

CI4H,iC1?;,OaS C14H&l?K,O,S Ci,H,iF,NzO,S Cl ,Hu?;jO,S2

+ + +

66 38 43 Hz0 36 67 cicrtcEr,ci +40 28 ClCII,CH,Cl 33 compounds weie analvLed for C, H, N.

Escherichia coli (T'ogel), Streptococcus pyogenes (C203), Proteus mil-abilis (ALGH-l), Salmonella typhimuriunz (V-31), and Shigella son?zei (C-10). Among them, VIh suppressed T . vayinalis i n viti.0 a t a concentration of 25 pg/ml and completely inhibited the growth of S. ~ J O qenes ((3203) a t 1.25 pg/ml. Experimental SectionlSrl9

1,3-Dioxo-2-isoindolineethanesulfonic acid monopotassium salt (11) was prepared by the method of IIiller and Roblinh11 in 83YGyield. 1,3-Dioxo-2-isoindolineethanesulfonylchloride (111) was obtained from I1 by t h e method of Miller and Roblin6111 in 80% yield, mp 158-162'.

CPIHBS?OBS

+

D-( )- 2,4 -Dihydroxy- 3,3-dimethyl-N- [2-(phenylsulfamoyl)ethyllbutyramides (VI, Table III).-A mixtiire of 0 Old mole of the reqiii4te 2-aminoethanesiilfonanilide (I-)and 3 9 g ( 0 03 mole) of D-( -)-pantolactone was heated in a melt a t 100-ll.io for 2 hr The melt was cooled and crystallized from the rolvents indicated.

Acknowledgments.-The authors are indebted to Dr. Leo Rane of the University of AIiami and to Dr. Paul E. Thompson and Dr. AI. W. Fisher of Parke, Davis and Company for the biological testing. We also wish to thank A h . C. E. Childs and associates for the microanalyses, and Dr. J . AI. Vandenbelt and coworkers for determination of the spectral data reported herein.

1,3-Diox0-2-isoindolineethanesulfonanilides (IV, Table I).T o a stirred solution of 0.1 mole of the substituted aniline in 75 ml of pyridine, cooled with an ice bath, was added slowly 30.1 g (0.11 mole) of 1,3-dioxo-2 .isoindolineethanesdfonyl chloride Derivatives of 5-Phenyl-2,4-pentadienoicAcid (111). After the reaction mixture was stirred for 1 hr with cooling, the ice bath was removed and stirring was continued for 1.25 hr. as Potential Antimalarial Agents' The reaction mixture was poured into 500 ml of H2O with vigorous stirring, and the crude product was isolated by filtration. ReLESLIE31.WERREL,N ~ S C YHE \DEN, crystallization from glacial or dilute AcOH gave the product. .WD EDW IRD F. E L S L ~ G E R 2-Aminoethanesulfonanilides (V, Table II).-A mixture of 0.02 mole of the appropriate 1,3-dioxo-2-isoindolineethanesulfonR w a r c h Laboratories, Parke, Davis and Company, anilide, 1.2 g (0.02 mole) of 85% hydrazine hydrate, and 100 ml Ann Arbor, Mzchzgan 48106 of EtOH was heated imder reflux for 3 hr. The reaction solution was homogeneous when refluxing began, but after 15 min a Received dfay 9, 1968 precipitate appeared. The mixture was concentrated to dryness and the residue was suspended in 200 ml of H,O and made acidic to congo red with 4 .V HCI. This slurry was heated on a The reported effectiveness of 5-(p-chlorophenyl)-Ssteam bath for 10 min, cooled in an ice bath, and filtered. The isopropy1-2,4-pentadienaniide (Ia) against Plasnzodiunz filtrate was neutralized with concentrated ?;H,OH to give the gallinaceunz in the chick2 prompted the synthesis of prodiict. The compounds were recrystallized from the indicated solvents when neceesary. (1) This investigation v a s supported by t h e U. S.Army Medical Research (18) Melting points (corrected) mere taken in open capillary t u b e s in a Thomas-Hoover capillary melting point a p p a r a t u s . (19) IVhere analyses a r e indicated only Iiy symbols of t h e elements or functions, analytical results obtained for those d e m e n t s or funrtions were witliin r0.4yoof tlie theoretical values.

a n d Development Command under Contract DA-49-193-.\ID-2754. This is Communication S o . 382 to tlie Army Research Program on 1Ialaria. (2) G . R . Coatney, W.C. Cooper, N. B. E d d y , a n d J . Greenllerp, "Survey of Antimalarial .%gents," Pulilic Health Service Pullliration No. 193, \Vasl~ington, D . C . . 1953, p p 98, 134, 262, 2 i G .