Journal of Mtdiunul ChcniLstry, 1970, L-oZ. l J , ,\o. 3 411
4-'~Hlo-6-sUBsTlTU'r~D PURINES
Preparation and Activity of the 4'-Thio Derivatives of Some 6- Substituted Purine Nucleosides' MIROSLAV BOBEK,ROYL. WHI~TLEK., Dcparlrritnt of Bzochemzstry, Purdue Cniversily, Lufaytlte, I n d i a n a AND
47907
ALEXANDER BLOCH
Dcpartmtnt of E'xptr cintntul Thcruptutics, Roswtll P a i k Mcniorial Instifutc, Bu$alo, A Y t k'orh ~
14203
Recetved Octobcr 28, 1969 (i-Chloro-9-(4-thio-~-~-ribofuranosyl)purine was prepared by cotidetisation of Z,S,.i-tri-O-acelyl-4-thio-D-ribi)furaiiosyl chloride with the chloromercuri derivative of 6-chloropurine, followed by ammoiiol Me*S, or SH, and dehalogenation furnished the 4'-thio substitution was used to replace the 6-chloro with "2, analog of 9-0-D-ribofuranosylpurine (nebularine). Replacemerit of the 0 in the carbohydrate skeletoii with S led to marked changes in the potency of the compounds, as determined in vitro Kith Streptococcus , f ~ c c z u ~ r ~ , Escherichia coli, Leukemia L-1210, and Ehrlich ascites cells. Depending on the test system used, the potency of the thioribosyl nucleosides was greater or smaller t,han that of the corresponding ribosyl analog. These differences in activity are likely related to differences in the metabolic disposition of the compounds. For example, the corresponding thio-D-ribosyl arialog did not undergo enayurilike 6-mercapt~o-9-(@-~-ribofuranosyl)purine, matic cleavage of its glycosyl bond. As a result, a mutant strain of S. fuecium resistant to the inhibitory effect was still sensitive to the action of both Gmercaptopuriiie and of 6-rnercapto-9-(4-thio-p-~-ribofura1~osyl)purine 6-mercapto-9-(p-~-ribofuranosyl)puririe.When used in combination with the corresponding ribosyl analogs, the thioribosyl derivatives tested interfered with the growth of S. faecilcm in a synergiatic mariner. 111view of this synergism and t,he observed activity against resist,arlt strains, these compounds deserve evaluation in vivo.
In the past, numerous chemotherapeutically useful compounds of the nucleoside variety have been obtained through structural modification of either the base or the sugar moiety. Compounds of this nature include, for instance, 6-mercapto-9-(/3-~-ribofuranosyl)purine2 and l-(p-~-arabinofuranosyl)cytosine.~The simultaneous modification of both the base and the sugar has also been done, as, for example, g-(p-D-arabinofurano~yl)-6-mercaptopurine.~Such double alterations offer the possibility for additional changes in the potency and selectivity of singly modified compounds, resulting, possibly, in their improved chemotherapeutic effectiveness. This paper reports the preparation and activity of a number of adenosine analogs, modified both at the 6 position of the heterocycle and in the 4 position of the carbohydrate (Figure 1). From the available synthetic routes for the preparation of the desired purine nucleoside^,^-^ direct glycosylation of the chloromercuri derivative of 6-chloropurine was chosen. Treatment of 2,3,5-tri-O-acetyl-4thio-D-ribofuranosyl chloride with the chloromercuri derivative of 6-chloropurine gave a 3774 yield of the acetylated syrupy nucleoside (Ia). Deacetylation of I a gave crystalline 6-chloro-9-(4-thio-~-~-ribofuranosy1)purine (Ib). Its structure was established by conversion into 9-(4-thio-P-D-ribofuranosyl)adenine through treatment in ,IleOH-SH3 to produce com-
(1) This investigation was supported by Public Health Serivce Research Grant KO.R01 hM11463, from t h e National Institutes of Health, Bethesda. l l d . , and by Grant T-436 from t h e American Cancer Society. Journal Paper S o . 3868. Agriculture Experiment Station, Purdue University, (2) J. A. Johnson, Jr., and H. J. Thomas, J . Amer. Chem. Soc., 7 8 , 3863 (1956). ( 3 ) E. R . Walwick, C. A. Dekker, and W.K . Roberts, Proc. Chem. Soc., 84 (1959). (4) E. .J. Reist. A. Benitea, L. Goodman, R . R. Raker. and \V. \V. Lee, J . Org. Chem.. 27, 3274 (1962). ( 5 ) G . n. Brown and V. S. Weliky. J . B i d . Chem., 204, 1019 (1953). (6) J. J. Fox. I . Wempen, A. Hampton, and I. L. Doerr, J . Amer. Chem. Yoc., 80, 1669 (1958). (i) H. Iwainma and T. Flashisurne, J . Ory. Chem.. 93, 1786 (1968). ( 8 ) H. G. Garg, J . S c i . India72 Res. Sect. C , 2 6 , 404 (1966).
pound I11 identical with that previously preparedg from chloroniercuri 6-benzamidopurine and 2,3,5-tri-Oacetyl-4-thio-~-i-ibofuranosy~chloride. Treatment of I b with thiourea in EtOH produced G-mercapto-9-(4thio-P-D-ribofuranosy1)purine (11). Sucleophilic displacement of the 6-C1 in I b with H S M e 2in AIeOH gave 6-dirnethylami1io-(4-thio-~-~-ribofuranosyl)purine (IV). Compounds I1 and IV possessed uv spectra similar to 6-mercapto-9-(P-~-ribofuranos~ 1)purine2B6and 6-dimethylamino-9-(p-D-ribofuratiosS-l)purine, respectively.roprl Dehalogenation of I b with H? and I'd catalyst gave 9-(4-thio-P-D-ribofuranosyl)purine (4'-thionebularine). ,4n attempt was made to synthesize 4'-thionebularine by condensation5 of 2,3,4-tri-O-acety1-4-thio-~-ribofuranosy1 chloride with the chloromercuri derivative of purine, but after the usual work-up and ammonolysis, tlc showed no evidence of compound V. The effects of the thionucleosides on the i?i vitro growth of 4 cell systems is shown in Table I. T o evaluate the contribution which the ring S niakes to the potency of the compounds, the corresponding nucleosides mere examined in parallel, with the exception of Gdimethylamino-9- (P-D-ribof uranosyl) purine. The introduction of S as the sugar ring heteroatom led to marked changes both in potency and in selectivity of the compounds. Thus, the G-chloro-9-(4-thio-P-Dribofuranosy1)purine was approximately 300 times more active against S . faeciunz than was the 6-chloro-Y-(P-~ribofuranosyl) purine. Similarly, the 9-(4-thio-P-Dribofuranosyl) purine was an effective inhibitor of the growth of this organism, whereas g-(p-D-ribofuranosyl)purine was inactive. This change in potency occurred also in the reverse direction, as shown by the fact that 6-mercapto-9-(P-~-ribofuranosy~)purine was approxi(9) E. J . Reist, D. E. Gueffroy, and L. Goodman, J . Arner. Chem. Soc.. 8 6 , 5658 (1964). (10) J. 4. Johnson, H. J. Thomas, and H. J. Scliaeffer, ibid., 80, 699 (1958). (11) L. 13. Townsend, It. K . Kobins, R . N. Loeggky, and N. J. Leonard, ibid.. 86, 5320 (1864).
CI I
Ib. R = Cl 11,R = SH 111. R = NH. IV.R = N(CH,)V,R=H Vigiire I
o -
Compound .5 A
TABLE I1 CROSS-I~ESISTANCS O F Streptococcus fuecium CELLSRESISTANT To 6-hIERChPTOPURINES 6-hIercaptopurine
+ +
S . faecium 6-mercaptopurine 8. fuccium,/6-mercapto-9-(p-o-r~bofuraiiosy~)pur~1ie S. fucc~un~~6-mercapto-9-(4-th~o-~-~-r~bofuranosy~)pur~ne (+) Resistant, ( - ) not reistarit. of the Hg derivat,ive'J of 6-chloropuriiie and Celite and 5 g of molecular sieve (Fisher Scientific Co., 4 -4)in 50 ml of dry PhMe was added 3.15 g (0.01 mol) of 2,3,5-tri-O-acetyl-4-thio-cu,8-~ribofiiranosyl chloride14 (prepared from 1,2,3,5-tri-O-acetyl-4thio-p-ribof~vatiosein 30 ml of dry P h l I e ) . The reaction mixture was stirred with exclusion of nioist,ure at 80-85" for 38 hr. The solids were removed by filtration and washed with 80 nil of EtOAc. This solution was washed with 30% K I ( 2 X 80 ml), once with H 2 0 (100 ml), and dried over aiihyd Na2HO4. T h e up remainiiig after removal of t8he EtOAc was purified by chumri chromatography oii bilica gel with hexaiie-EtOAc ( 3 : 2 , v , v ) as the eluent. After removal of the solvent, the syrup was dirsolved in 30 ml of NeOH and evaporated to yield 1.62 g (37.SC/l); uv max (MeOH) 264 m p . 6-Chloro-9-(4-thio-p-o-ribofuranosyl)purine (Ib).-Ia (1.27 g, 0.003 mol) was dissolved in 50 ml of methanolic NH, (batitrated at O"), allowed to statid at 3' overiiight, aiid was concentrated to drynesb at 40-45". The product was dissolved in 20 ml of hut NeOH from which it crystallized on staiidiug overnight. The colorless needles were removed by filtration, washed with EtOH, arid dried; yield 6x0 mg; mp 190-192" dec. Partial evaporatioii of the filtrate yielded an additional 65 mg; total yield 829;. Recrystallized from MeOH, mp 190-192" dec; [ C U ] ~ ~-41.8'; D IIV nlax (H>O) 264 mp ( e 10,200). dnal. (CioH~ClN403Sj C, H. CI, K, S. 6-'Mercapto-9-(4-th~o-p-~-r~bofuranosy~)pur~ne (II).-.4 mixture of 0.605 g (0.002 mol) of Ib, 0.2 g of t,hiourea, and 60 ml of 9.5.6(,7 EtOH was refliixed 1.6 hr on a steam bath. Charcoal was added to the warm solution and the mixture filtered. The residiie remaining after conceiitration of the filtrate was dissolved in 5 ml of hot H2O and 10 ml of hot EtOH was added. Crystallization occurred after staiidiiig of the soliitioii overnight at 2.i"; yield 300 mg; mp 233-233" dec. Partial coiicentratioii of the filtrate yielded ail additioiial 200 my. I
