XIII. The Synthesis of an Active- Site-Directed Irreversible Inhibitor of Adenosine Deaminase'
Enzyme Inhibitors.
Previoiis stiicliea have wggesled that fhrre is :t hydropliot)ic. region on :ideriositie tleaniiiitise which i.. iI1lpUlt:lllt for binding the alkyl groiip of some O-:dkyladenines. 111 an at tempt, to take advantage of t,hi,qhydropliohic region ( I C adenosine deaminase for the preparation of some inhibitor.; of t,his enzyme, a series of 9-( para-substituted benzyl)6-sttbstituted purines was prepared. In addition t o studyiiig reversible inhibition of adenosine deaminase, it ~ ~ y found that 9-(pbromoacetamidoberizyl)adenine ( X I I ) was ai1 irreversible inhibitor of adenosine deaminase. ( ) i i the basis of kinetic studies and because of the lack of iriactivation of adenosine deaminase by iodoacetamide, i t is suggested that the inactivation of the enzyme by XI1 IWIYI~S throiigh an E-T comples nnd does not, proceed throriyh :I random biIno1ei~ttl:n.process.
I'reviously it hasl been suggested that L: hydrophobica region exists on adenosine deaminase in the region where the 9-substituent of some 9-alkyladenines bind.2 In ihe hope of taking advantage of this hydrophobic. region for the preparation of good reversible inhibitor& of adenosine deaniinaie, we decided to prepare soriic~ 9-(para-substituted benzyl)-6-substituted purines. If' the benzyl moiety of these purine derivatives were capable of binding to the hydrophobic region, it shoul(1 be possible to utilize ;Lrubbtitueiit at the para position which is c*apablr of forming ;t c~ov:~lcntb o d with 1 1 1 ~ en z y ir it'. of \ o m Y-(pa,nThis p:iper describes thc syiitht substituted benzyl)-6-subqtitutcd puriiirb and thcw enzymic evaluation as reversible irihibitors of aderiosiiw deamiriase. In addition, :L kinetic* evaluation of 9(p-bronioac.etaiiiidobeiizy1)adrriirir :I\ :HI irreversihlr inhibitor of adenosine deaininabc i5 presented. For the preparation of this hcri(1:. of inhibitors, me *elecatcd S-(p-nitrobeiizyl)-G-(,liloropuriiie a\ the key intermediate. The gencral method of synthesis I < b a d on L: inodific~ationof the procedurc of Montgomery :md and is outliried in Chart I. Conderisatiori of 6-chloropuri~ie (I) with p-riitrobenzyl bromide (11) gave L: mixture of the 9- and 7-p-riitrobenzyl-6-chloropurines (I11 and IV) whicbh was separated by chromatogr:Lphy 011alumimi. Treatment of I11 with aninionia. niethylnmine, dimethylamine, hydrochloric3 arid, or ihiouren gave thc rorresponding (i-substitutecl isomers (\'-IX). Catalytic reduction of V using ti palladiunioil-carbon catalyst gave the p-:iniino derivatiw (X) i n good yield. When X wab a l l o ~ e dt o rcacat it h plieriyl cl-iloroformatc, bromoacetyl bromide, or avetyl chloritlc 1 tie corresponding para-iubstitut et1 benzyl derivai ivc. (XI-XIII) were obtained. The ashigiiiiieiit of structure to XI, XII, aiid XI11 is bawd o n the obbervation that S6-:xylatioii of adenine clerivai ives shifts the ultraviolrt rriaxirria t o longer wavelength*. For examplr, it has h t w 1 found that N,O~'-cl~acetyldeoxyadenylic~ 5':wid exhibited nil ultraviolet maximum a t 273 111p at (1) This investigation uti5 supported 1,) a Public Iiealtlr beri Ice research Yrant (Ch-06388-03) and a research csreei program a u a r d (5-K3-C L18i18-04) from t h e h'ational Cancer Institute, a training grant (5-Tl-GlI5.55) from t h e Division of AIedical Sriences, C . Y. Public Health Servires.. Hethesda. Md.. a n d bv w a n t iT-33i) from the American Cancer Yoriet!. .a. chaeffer and L). Tugel. J . Mcd. Chem., 8, 507 (1965). (3) .I. : I . Montgomery and C. Temple, .Jr.. .J. Ani. C h ~ n r ,So?., . 83, 630
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