Topics in Current Chem 97, 215

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H eparinoid M im eticsH a n s P e t e r W e s s e lPh arm a Divis ion, Preclinical Research, E Ho ffm ann -La Roche Ltd,CH-4070 Basel, Switzerland

Hepar ino id po lysacchar ides such as heparan su lfa te and hepar in a re ab le to in te rac t wi thnumerous proteins and inf luence vita l biological processes . Hepar inoid mimetics wereprepared to reduce the s t ruc tura l com plex i ty of hepar ino ids a nd to ob ta in se lec tivi ties . T hisa r t ic le sum m ar izes the deve lopm ent o f hepa r ino id mim et ics o f d if fe ren t c lasses inc lud ingrepresentative synthe ses and biologica l activities. La rgely s implif ied co m po un ds w ith regardto s t ruc ture and syn the t ic access a re descr ibed which main ta in or exceed the ac t iv i ty o fhepa r ino id p o lysacchar ides . One of the recipes to increase b ind ing or m odi fy pharm aco-k in e t ic p a r a m e te r s wa s th e in t r o d u c t io n o f h y d r o p h o b ic g r o up s .

Table of Co ntents1

22 . 12 . 2344 .14 .255.15 .26

7R e f e r e n c e s

I n t r o d u c t i o n . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 1 6B i o lo g ic a l P r o p e r t ie s o f H e p a r i n o i d s . . . . . . . . . . . . . . . . . . 2 18H e p a r i n o i d I n t e r a c t i o n s w i t h P r o t e i n s . . . . . . . . . . . . . . . . . 2 18S e l ec t ed B i o lo g ic a l P r o p e r t ie s o f H e p a r i n o i d s . . . . . . . . . . . . . 2 20D e r iv a t i v e s o f H e p a r i n S a c c h a r i d e s . . . . . . . . . . . . . . . . . . . 2 2 1S u l fa t ed O l i g o s a c c h a r i d e s . . . . . . . . . . . . . . . . . . . . . . . . 2 23S u lf a t ed L i n e a r O l i g o s a c c h a r id e s . . . . . . . . . . . . . . . . . . . . 2 23S u l fa te d C y c li c O l i g o s a c c h a r i d e s . . . . . . . . . . . . . . . . . . . . . 2 31S u l fa t ed S p a c e d S u g a r s . . . . . . . . . . . . . . . . . . . . . . . . . . 2 33S u lf a t ed S p a c e d O l i g o s a c c h a r i d e s . . . . . . . . . . . . . . . . . . . . 2 33S u lf a t ed S p a c e d O p e n C h a i n S u g a rs . . . . . . . . . . . . . . . . . . . 2 34N o n - C a r b o h y d r a t e C o m p o u n d s . . . . . . . . . . . . . . . . . . . . . 2 3 6C o n c l u d i n g R e m a r k s . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 3 6

. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2 3 7

Topics n C urrent Chem istry,Vol. 1879 SpringerVerlag Berlin Heidelberg 1997


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216 H.P. WesselList of Ab brev ia tions and SymbolsA ca F G F =AIDS ----A P T T =A T II I ----fl-CD ----bFGF ----B n - -C M V =C R S - h e p a r i n =D M F =D M S O =D S =E G F =E t =F G F =HBGF ----H C II ----H I V =H S V =Le v =M e =P D G F =P F 4 - -P h =r i - ---

RT ---S M C =T M U =to l =

a c e ty la c i d ic f i b r o b l a s t g r o w t h f a c t o ra c q u ir e d i m m u n e d e f ic i en c y s y n d r o m ea c t iv a t e d p a r t i a l t h r o m b o p l a s t i n t i m ea n t i t h r o m b i n I IIf l - c y c l o d e x t r i nb a s i c f ib r o b l a s t g r o w t h f a c t o rb e n z y lc y t o m e g a l o v i r u sc a r b o x y l - r e d u c e d s u l fa t e d h e p a r i nN , N - d i m e t h y l f o rm a m i d ed i m e t h y l s u l f o x i d ed e g r e e o f s u l f a t io ne p i d e r m a l g r o w t h f a c to re t h y lf i b r o b l a s t g r o w t h f a c t o rh e p a r i n - b i n d i n g g r o w t h f a c to rh e p a r i n c o f a c to r IIh u m a n i m m u n o d e f i c ie n c y v i ru sh e r p e s s i m p l e x v i r u sl e v u l i n o y lm e t h y lp l a t e l et d e r i v e d g r o w t h f a c t o rp la t e l e t f a c to r 4p h e n y lr e la t iv e in h i b i t o r y a c t iv i t y c o m p a r e d t o h e p a r i n i n a n S M Cc e ll p r o l i f e r a t i o n a s s a yr o o m t e m p e r a t u res m o o t h m u s c l e ce llt e t r a m e t h y l u r e at o l u e n e

1Int roduct ionH e p a r a n s u l fa t e a n d h e p a r i n a re s u l f a t ed p o l y s a c c h a ri d e s o f t h e g l y c o s a m i n o -g l y c a n fa m i ly . B o t h a r e b i o s y n t h e t i c a l l y r e la t e d b e i n g g e n e r a t e d f r o m a c o m m o npo lysa c c ha r ide wi th f l - D- g luc ur on ic a c id - o~ - ( 1 - - > 4) - N- a c e ty l - D- g luc osa m iner e p e a t s e q u e n c e o n a p r o t e o g l y c a n c h a i n [1 ]. T h e e n z y m a t i c m a t u r a t i o n p r o c e ssi n v o lv e s d e - N - a c e t y l a t i o n a n d N - s u l f a ti o n ; t h e f o l l o w i n g t r a n s f o r m a t i o n s , e p i -m e r i z a t i o n a t C - 5 o f t h e u r o n i c a c i d a n d O - s u l f a ti o n a t v a r i o u s s p e c i f ic p o s i -t i o n s , d e p e n d o n N - s u lf a ti o n . H e p a r i n is p r o c e s s e d t o a h i g h e r d e g r e e a n dc a r r i e s m o r e L - i d u r o n i c a c i d r e s i d u e s , m o r e N - s u l f a t e a n d O - s u l f a t e g r o u p s ,


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H e p a r in o i d M im e t ic s 217

R - - ~ . S 0 3 "1 - HOl~.~e , , - -iS O i O ~N H S ~I]O, 1 0 -2

Structures1. Characteristicstructural motifs in heparan sulfate and heparin

an d l e s s Noace ty l g roup s t han he pa ran su lfa te . Al tho ugh c ha rac t e r i s t i c r epea t -i ng un i t s (S t ruc ture s 1 ) can be a t t r i bu t ed t o hep a ran su l fa t e (1) an d h epa r in (2 ),t h e s e p o l y s a c c h a r id e s a r e c h a r a c t e r iz e d b y g r e a t s t r u c t u r a l h e t e r o g e n e i t y s t e m -m i n g f r o m d i f f e re n t s u b s t it u t io n p a t te r n s , a n d 19 d if f e r e n t n a t u r a l l y o c c u r r i n guro nic ac id - g luco sam ine d i saccha r ides have been iden t i f i ed so far . In add i t ion ,t h e p r e p a r a t io n s s h o w d i s p e r s io n o f m o l e c u l a r w e i g h t a f t e r cl ea v a ge f r o m t h ep r o t e o g l y c a n co r e. E m p h a s i z in g t h e i r c o m m o n f e a tu r e s, h e p a r i n a n d h e p a r a nsu l fa t e a re re fe r red t o a s "hepar ino ids" [2].S t a r t i ng f rom b ioac t ive p ro t e ins , pep t ides wi th s imi l a r ac t i v i t y o r b ind ingp r o p e r t i e s c o u l d f r e q u e n t l y b e i d e n ti fi e d ; t h e d e v e l o p m e n t o f p e p t i d e m i m e t i c sf rom those p ep t ides is a c l a s si ca l exe rc i se i n m edic ina l chem is t ry . The ana logou sa p p r o a c h f o r c a r b o h y d r a t e s, t h e d e v e l o p m e n t o f c a r b o h y d r a t e m i m e t i c s fr o mbioac t ive polysa ccha r ides , i s in i t s infancy.I n t h e c a s e o f h e p a r i n o i d s , t h e d e v e l o p m e n t o f m i m e t i c s i s d e s i r a b l e t or e d u c e t h e e n o r m o u s c o m p l e x i ty o f th e p o l y s a c c h a ri d e m i x t u r e s t o a r r iv e a tc o m p o u n d s o f lo w e r m o l e c u l a r w e i g h t t h a t c a n b e s y n t h e s i z ed in a r e a s o n a b len u m b e r o f s te p s . A s d e l in e a t e d i n m o r e d e t a il in t h e n e x t s e c t io n , h e p a r i n o i d sa r e a s s o c i a t e d w i t h a m u l t i tu d e o f b io l o g i ca l p r o p e r t i e s , a n d t h is r a is e s t h eq u e s t i o n o f s e le c ti v it y . M i m e t i c s c a n b e i n v e s ti g a t ed i n t h i s r e s p e c t a n d m a yb e v a lu a b l e t o o l s t o s t u d y h e p a r i n o i d - p r o t e i n i n t e r a c t i o n s a n d t h e i r i m p l i-c a t i o n s . A s w e l l e s t a b l i s h e d f o r p e p t i d e m i m e t i c s , h e p a r i n o i d m i m e t i c sm a y a ls o b e t a i lo r e d t o m o d i f y p h a r m a c o k i n e t i c p a r a m e t e r s . W h e r e a s h e p a r -i n o i d - p r o t e i n i n t e r a c t i o n s a r e m a i n l y b a s e d o n i o n i c f o rc e s , h y d r o p h o b i cg r o u p s i n h e p a r i n o i d m i m e t ic s m a y le a d t o n o n - c h a r g e d i n t e ra c ti o n sa n d e n h a n c e t h e b i n d i n g . I n a d d i t i o n , s i n c e h e p a r i n o i d s a r e i s o l a t e d f r o mn a t u r a l s o u r c e s , e . g . c o m m e r c i a l h e p a r i n p r e s e n t l y m a i n l y f r o m p o r c i n ei n te s ti n e , p r e p a r a t io n s m a y v a r y w i t h r e s p e c t to t h e i r h e t e r o g e n e i t y a n d t h u st h e i r c o m p o s i t io n ; c o n t a m i n a t i o n w i th v i r u s e s o r p r i o n s m a y a ls o b e c o m e a ni ss u e o f p u r it y . A h e p a r i n o i d m i m e t i c w o u l d i d e a ll y m a i n t a in o n l y o n e p h a r -m a c o l o g i c a l a c ti v it y , b e s t r u c t u r a l l y l es s c o m p l e x th a n h e p a r i n a n d b e e a s il yp r e p a r e d .

L o w m o l e c u l a r w e i g h t h e p a r in s , h e p a r i n f r a ct io n s , o r o t h e r s u l fa t ed p o l y s a c-cha r ides s t il l have t he sam e or a s im i l a r com plex i ty a s he pa r in an d a re , t he re fo-r e , n o t r e g a r d e d a s h e p a r i n o i d m i m e t i c s ; o n l y c o m p o u n d s w i th o n e d e f i n e d c a r-bo hy dra t e backb on e se rv ing a s a t emp la t e fo r su l fa t es wi ll be d i scussed in t h i scontext .


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218 H.P. Wessel2B i o l o g i c a l P r o p e r t i e s o f H e p a r i n o i d sA s p o l y a n i o n i c s a cc h a ri d es , h e p a r i n o i d s i n t e ra c t w i t h a h i g h n u m b e r o f p r o -t e ins , t he so-ca l l ed hep a r in b ind ing pro t e ins . H epa r in o id - p ro t e in i n t e rac t i onshave usu a l ly bee n desc r ibed for i n v i t ro sys t ems , and the i r phys io log i ca l r e le -vance i s no t a lways clear . On the o th e r han d , hep a r ino ids can i n f luence b io log i-ca l sys t ems , bu t i n m os t ca ses t he m olecu la r bas is i s no t fu l ly und e rs too d . Forth i s r ea son , in t e rac t i ons wi th p ro t e ins a nd b io log ica l p rope r t i e s o f hepa r ino idswi l l be d i scussed sep a ra t e ly in t he fo l lowing . As hepa r ino id - p ro t e in i n t e rac -t i ons have been rev i ewed [3] the focus wi ll be i n ra the r s hor t fo rm o n those a reasi n w h i c h h e p a r i n o i d m i m e t ic s h a v e b e e n i n ve s ti ga te d .2 .1H e p a r in o id In t e r a c t io n s w i t h P r o t e in sI n t e r ac t io n s o f h e p a r in o i d s w i t h t h e m o s t d i v e r se p ro t e in s s u c h a s e n z y m e s a n de n z y m e i n h i b it o rs , c y t o k in e s , a n d a d h e s i o n m o l e c u l e s h av e b e e n d e s c r ib e d . T od a te , m a n y m o r e t h a n a h u n d r e d h e p a r i n b i n d i n g p r o te i ns a r e k n o w n . A n u m -b e r o f h e p a r i n b i n d i n g p r o t e i n s a r e m e m b e r s o f t h e s e r p i n f a m i l y o f s e r in e p r o -t ea se inh ib i to r s . The bes t d e sc r ibed exam ple is an t i t h ro m bin [4]. A nt i t h rom binIII (AT II I ) is able to in hibi t var ious ser in e proteases involved in the b loo d co a-gu la t i on process b y fo rm a t ion o f s tab le , equ imo la r complexes . B ind ing of hepa -f in t o AT I l l a cce l e ra t e s t he k ine t ic s o f th i s com plex forma t ion by seve ra l o rde rsof m agn i tude . Thi s has bee n the bas i s fo r t he success fu l c l in i ca l use o f hep a r inas an an t i coagu lan t fo r nea r ly s ix ty yea rs .D e g r a d a t i o n o f h e p a r i n f o ll o w e d b y a f f in i ty c h r o m a t o g r a p h y o n i m m o b i l iz e dAT I l l l ed t o t he i den t i f ica t i on of a pen t a sacch a r ide un i t wi th h igh an t i t h ro m bina f fin it y . Th i s p en t a saccha r ide 3 i s cha rac t e r i zed by a un iqu e h igh ly su l fa t ed cen-t ra l g lucosam ine un i t w i th a 3 -O-su lfa te g roup (S t ruc ture s 2 ).Thi s pen t a saccha r ide sequence i nduces a conforma t iona l change in AT I I Iw hich pro bably causes t he co m plex to be m ore access ib l e t o t he ac ti ve s it e o f thepro t eases . The m os t r e l evan t p ro t ease a f fec t ed by th e pen ta saccha r ide 3 i s f ac torXa, bu t fac tor XIIa an d p lasm a ka l l ikre in ac t ivi ties can a l so be poten t ia ted.Sequen ce 3 occurs i n he pa r in a s we ll a s i n va r ious hep a ran su lfa te p ro t eoglycanso f d i f f e r e n t o ri g i n i n c lu d i n g t h e v a s c u l ar e n d o t h e l i u m .Oth e r m em be rs o f t he coagu la t ion ca scade such a s fac tor IIa (al so re fe r red toas t h rom bin) , f ac tor IXa , an d fac tor XIa requ i re a l onge r hepa r in u n i t o f 13 or

HO NHS%'HO OH " % S O N HS%" HO (~O~ HO NHSO i3

Structures 2. Structure of the heparin pentasaccharidewith high antithrom binaffinity


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HepadnoidMimetics 21 9more sacchar ide un i t s i n add i t i on to t he pen tasacchar ide to acce l e ra t e t hea n t i t h ro mb i n - p ro t e i n r e a c ti o n . A t emp l a t e mo d e l w a s p o s t u l a t e d i n w h i c h ah e p a r i n o i d b in d s t o a n t i t h ro mb i n p l u s t h e s e c o n d p ro te i n i n a t e rn a ry c o mp l e x .To un der s t an d the con fo rm at iona l change o f AT I I I on a m o lecu la r leve l, thep ro te in - pen tasacch ar ide b ind ing s it e was inves tiga ted by va r ious m e tho ds [4 ]inc lud ing spec t ro scop ic [5 ] a s we l l a s m o lecu la r m ode l l i ng [6 ] s tud ies ; wherea sf ir s t m o d e l l in g e x p e r i me n t s w e re b a s e d o n a l - a n t i t r y p s i n , c ry s t a l s t r u c t u r e so f c l eaved AT I I I [7] an d o f ac t ive d ime r i c an t i t h rom bin [8 ] have s ince beenrepo r t ed . In add i t i on to t he AT I I I con fo rm at iona l change , t he co n fo rm at iona lf lex ib i l ity of the T.-iduronic ac id un i t o f hep ar in , a l so prese nt in th e h ep ar in pen -t a sacchar ide , has be en pos tu l a t ed to p l ay an im por t an t ro l e i n t he p ro t e in i n t e r -ac t ion [9] .A n o t h e r s e r in e p ro t e a s e i n h ib i to r o f th e a l - a n t i t r y p s i n f a mi l y ( s e rp in ) ishep a r in co fac to r II (HC I I ), wh ich a l so fo rm s a 1 : 1 com plex w i th t h rom bin , bu tdoes n o t reac t w i th fac to r Xa [4 , 10 ] . The ra t e o f inh ib i t i on o f t h rom bin is no to n l y i n c r e a s e d b y h e p a r i n o i d s b u t a ls o b y t h e r e la t e d g l y c o s a mi n o g l y c a n d e r -m a tan su lfa te . The iden t i f i ca t ion o f an inh ib i to r va r i an t an d s i t e -d i rec t ed m u ta -genes is s tud ies on HC I I cDNA led to t he u nde rs t an d ing tha t t he b ind ing s it esfo r h e p a r i n a n d d e rm a t a n s u lf at e m a y b e o v e r l a p p in g b u t n o t i d en t ic a l. Fu r t h e rp ro t e inase inh ib i to rs i n t e rac t ing w i th hepa r ino ids a re t i s sue fac to r pa thwayinh ib i to r and p ro t ease nex in -1 .Re levan t hep a r in -b ind ing enz ym es no t i nvo lved in t he co agu la t ion cascadeare , for exam ple , e las tase , ca thep sin G, supe roxide d ism utase , l ipopro te in l ipaseand o the r l ipases . The p l a sm a c l ea r ing p roper t i e s o f hep a r in a re a ssoc i a t ed w i thi ts b ind ing to l i pop ro te in l ipase and hepa t i c l ipase wh en the enz ym es a re re lea -sed f rom the su r face o f endo the l i a l cells [ 11 ] and have been s tud ied in v i ew o f apo ten t i a l imp ac t on the regu la t ion o f a the rosc l e ros i s .M em bers o f t he f i b rob las t g rowth fac to r (FGF) fami ly [12 ] b ind hep ar in w i thh i g h a f f in i ty a n d w e re a ls o t e rm e d " h e p a r i n -b i n d i n g g ro w t h f a c to r s " (H B G F).The m os t t ho ro ugh ly cha rac t e r i zed me m ber s a re ac id ic FGF (aFGF, FGF-1 ) andbasic FGF (bFGF, FGF-2); they are m i togen ic for a wide va r ie ty o f cell s and sup-po r t chem otac t i c m ig ra t ion an d in duc t ion o f p ro teases t ha t co u ld fac i li ta t e t is -sue remo de l l i ng . Hep ar ino ids can s t abi li ze t he g row th fac to rs and p reven t i nac -t i va tion b y p ro t eo lys i s. Acco rd ing to b ind ing s tud ies , t he b in d ing s i te o f h epa r i -n o i d s f o r bFG F a n d a FG F w a s e s ti ma t e d t o b e c o m p o s e d o f f o u r to e i g h t mo n o -sacchar ide un i t s r i ch in 2 -O-su lfa ted idu ron ic ac id [13 ] . The invo lvemen t inhep ar ino id b ind ing o f t he bFGF bas i c am ino ac ids i den t i f i ed by va r iousm e t h o d s h a s b e e n c o n f i rm e d v e ry r e c e n tl y b y c ry s t a l s t r u c t u r e s o f bFG F c o m -p lexes w i th hepa r in t e t ra - an d hexasac char ides , t he f i r st X- ray s t ruc tu res o fhep ar ino id f ragm en t s [ 14] . I t is i n t e res t ing to no te , however , t ha t i n a s tudy o f t hebFGF/hepar in i n t e rac t ion the amoun t o f non -e l ec t ro s t a t i c i n t e rac t ions , i . e .hyd roge n bon d ing a nd van d e r Waals pack ing , has bee n es t im a ted a t 70% [15 ].The h epa r ino id sequen ce requ i red to e l ic i t a b io log ica l re sponse ( s igna l t rans -d u c t i o n , m i t o g e n i c it y ) is l o n g e r t h a n t h e b i n d i n g s e q u e n c e m e n t i o n e d a b o v e,on ly dodeca sacchar ides be ing equ iva l en t to hep a r in . I t was h ypo th es i zed tha tt h e a d d i t io n a l s a c c h a r id e s a r e n e e d e d t o b i n d t h e r e c e p t o r o r a n o t h e r b FGFm olecu le t o fac il it a te recep to r d im er i za t ion [16 ].


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220 H.P. W esselO t h e r c y t o k i n e s b i n d i n g t o h e p a r i n o i d s i n c l u d e p la t el e t f a c to r 4 ( P F 4) a n din t e r l e u k in - 8 [ 1 7 ] .

2.2S e l e c t e d B i o l o g i c a l P r o p e r t i e s o f H e p a r i n o i d sH e p a r i n a n d h e p a r a n s u lf at e i n h i b i t t h e m i g r a t i o n a n d p r o l i f e ra t io n o f s m o o t hm u s c l e ce ll s (S M C ) i n v i t r o a n d i n t i m a l t h i c k e n i n g i n v i v o [ 1 8]. T h e h i g h a n t i -p r o l i f e ra t i v e a c t iv i t y o f h e p a r a n s u l fa t es [1 9 ] p a r t i c u l a r l y s u g g e s t e d t h a t e n d o -g e n o u s h e p a r a n s u lf at es f u n c t i o n a s s p e c if ic re g u l a t o rs o f S M C g r o w t h , a n do t h e r c e ll t y p e s w e r e o n l y m i n i m a l l y in h i b i t e d . T h e a n t i p r o l i f e ra t i v e e ff e c t o fh e p a r i n o i d s w a s n o t d e p e n d e n t o n t h e A T I I I - m e d i a t e d a n t ic o a g u l a n t a c ti v i ty[2 0]. S iz e f r a c t io n a t i o n o f h e p a r i n d e m o n s t r a t e d t h a t a d o d e c a s a c c h a r i d e w a sr e q u i r e d t o o b t a i n t h e f u l l h e p a r i n e f fe c t, a n d o v e r s u l f a t io n o f t h e f r a c t i o n ss l ig h t ly i n c re a s e d t h e a c t iv i ty [2 1]. T h e m o l e c u l a r m e c h a n i s m f o r t h e h e p a r i n o -i d a n t i p r o l i f e r a ti v e a c t i v it y is n o t k n o w n y e t ; i t s e e m s t h a t t h e i n h i b i t o r y a c t i o nis n o t e x e r t e d b y b i n d i n g o f s e r u m m i t o g e n s s u c h a s p l a te l et d e r i v e d g r o w t h f a c-t o r ( P D G F ) o r e p i d e r m a l g r o w t h f a c t o r ( E G F ) . W h i l e i t h a s b e e n s h o w n t h a th e p a r i n i s i n t e r n a l i z e d b y S M C [2 2] i t i s n o t c l e ar w h e t h e r h e p a r i n o i d s a c t i n t r a -c e l l u l a r l y o r t h r o u g h a sp e c i f i c s i t e o n t h e c e l l su r f a c e [ 2 3 ] . E x c e s s iv e SMCg r o w t h p l a y s a n i m p o r t a n t r o le i n d i s e as e s s u c h a s a r t e r io s c l e r o s i s [ 2 4]. C l i n i c alt r i a l s w i t h h e p a r i n s a g a i n s t v a s c u l a r r e s t e n o s i s , a r e n a r r o w i n g o f t h e a r t e r i a ll u m e n w h i c h o c c u r s w i t h h i g h i n c i d e n c e a f te r a n g io p l a st y , a n o f t e n u s e d p r o c e -d u r e t o r e s to r e b l o o d f l o w i n s t e n o t i c c o r o n a r y a r te r ie s , h a v e n o t s h o w n c o n v i n -c i n g e f fi ca cy , h o w e v e r , o n l y l o w d o s a g e s w e r e p o s s i b l e d u e t o i m m i n e n t b l e e d i n gc o m p l i c a t i o n s .M o s t m a m m a l i a n c e lls c ar r y h e p a r a n s ul fa te s as p l a s m a m e m b r a n e s u b s ti tu -e n t s . C e l l s u r f a c e h e p a r a n s u l fa t es h a v e b e e n s h o w n t o m e d i a t e t h e i n i t i a l b i n -d i n g o f e n v e l o p e d v i r u s e s s u c h a s h e r p e s s i m p l e x v i r u s ( H S V ) , c y t o m e g a l o v i r u s( C M V ) , a n d p s e u d o r a b i e s v i r u s t o t h e c e ll [ 25 ]. E q u a ll y , i t w a s s h o w n t h a t h e p a -r a n s u lf at e p r o te o g l y c a n m e d i a te s h u m a n i m m u n o d e f i c i e n c y v i r u s ( H I V ) i n f ec -t i o n o f T - c el l l i n e s [ 2 6]. T h u s , h e p a r a n s u l fa t e a n d h e p a r i n , b u t a l so o t h e r p o l y -su l f a t e d s a c c h a r id e s su c h a s d e x t r a n su l f a t e , sh o w a n t i - v i r a l a c t i v i t y [ 27 , 2 8 ] .H e p a r i n f r a g m e n t s w e r e s i g n if ic a n t ly l es s a ct iv e t h a n h e p a r i n , a n d f r a g m e n t sb e l o w a m e a n m o l e c u l a r w e i g h t o f 7 k D l o s t a c t iv i t y b y m o r e t h a n a f a ct o r o f t e n[ 2 7 ] . H I V r e p l i c a t i o n i s b e l i e v e d t o b e i n h i b i t e d b y t h e i n h i b i t i o n o f v i r u sa d s o r p t i o n t o t h e c e ll s u rf a ce h e p a r a n s u lf at es , t h e m e c h a n i s m b e i n g b i n d i n g o fs u l fa t e d p o l y s a c c h a ri d e s t o t h e v i r u s t o i n h i b i t t h e b i n d i n g o f t h e v i r io n s t oC D 4 + c el ls a n d s u b s e q u e n t g i a n t c e ll ( s y n c y t i u m ) f o r m a t i o n . S t u d i e s w i t h s p e c i -f ic a n t i b o d i e s c o n f i r m e d t h a t p o l y s u l fa t e d m o l e c u l e s in t e r a c t w i t h t h e p o s i ti v e -l y c h a r g e d a m i n o a c i d s c o n c e n t r a t e d i n t h e V 3 l o o p o f g p 1 20 o f H IV -1 . S u l fa t e dp o l y s a c c h a r i d e s a r e a l s o i n h i b i t o r y t o H IV -1 r e v e r s e t r a n s c r i p t a s e , b u t o n l y a tc o n c e n t r a t i o n s t h a t a r e a p p r o x i m a t e l y 1 0 0 -f ol d h i g h e r t h a n t h o s e r e q u i r e d t oi n h i b i t v i r u s - c e ll b i n d i n g [2 9]. D i s a p p o i n t i n g l y , n e i t h e r o f th e p o l y s a c c h a r i d e sd e x t r a n s u l fa t e n o r t h e p e n t o s a n s u l f a t e w e r e c l in i c a ll y e f fi c a c io u s a f te r o r a l o rp a r e n t e r a l d o s a g e a l t h o u g h h i g h p l a s m a l e v e l s w e r e a c h i e v e d b y c o n t i n u o u si n t r a v e n o u s i n f u s i o n [ 3 0].


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H e p a r i n o i d M i m e t i e s 2 2 1

C o n s i d e r i n g t h e m a n y i n t e r a c t io n s o f h e p a r i n o i d s w i t h t h e v a r i o u s p r o t e i n sa n d t h e m u l t i t u d e o f r e s u l t i n g b i o l o g i c a l a c t i v i t i e s , i n c l u d i n g e f f e c t s o nt u m o u r c e ll m e t a s ta s i s , a n g i o g e n e s is , a s t h m a , a n d i m m u n e c e ll m i g r a t i o n i ni n f l a m m a t i o n , i t i s c le a r th a t h e p a r i n o i d s a n d e s p ec ia l ly h e p a r i n o i d m i m e t i c sm a y b e u s e f u l n o t o n l y i n a n t i c o a g u l a t i o n b u t a ls o i n a s e r ie s o f o t h e r i n d i c a -t i o n s [3 I ] .3Derivatives of H eparin SaccharidesA n a l o g u e s o f t h e h e p a r i n p e n t a s a c c h a ri d e 3 h a v e b e e n p r e p a r e d b y v a r i o u s a u t -h o r s [3 2] w i t h t h e m a i n c o n t r ib u t i o n s f r o m t h e S a n o f i a n d O r g a n o n g r o u p s ;t h e i r a c h i e v e m e n t s h a v e b e e n c o m p e t e n t l y r e v i e w e d r e c e n t l y [3 3]. T h i s w o r kh a d t h e m a i n g o a l o f d e v e l o p i n g s e le c ti v e i n h i b i t o r s o f f a c to r X a a s a n t i t h r o m -b o t i c s . I m p o r t a n t f o r t h e d e v e l o p m e n t o f s y n t h e t i c a l l y s i m p l i f i e d d e r i v a t i v e sw e r e t h e f i n d i n g s th a t a n a - m e t h y l g l y c o s id e h a d t h e s a m e b i o l o g ic a l a ct i v it y a s

. . 3 "O a , ,/ C S O ~ ~ , / C S O ~H O e ' " " , O = , " O i , . . . , l O i , , , - O i , , . - , , iO M eH O N I '. L ., ~" H O O H "O 3 S O N H S C b " X O O S O ~ " O ~ S O N t .'I ,. ., ~"4.(o c oo - f ( : ~ o 3 c oo - f o 6 o ~O 0 0 " O 0R O t, . - . , , lO t , . O f , , " . ,l O t, . . " 0 . , . . . . . tO M eRo ( :~o 3- ueO O M e~ SO OSO3" MoO ( :~O3" ~ S O OSO3"5 R= CH36 R = (CH2)3CH3

9 I 0 1 r O l i , o . - . , I O M e( ~ o ~ M o o o s o ~ - o ~ s o o s o ~ - M o o (:~ eo ~- ~ s o o s o = -

7

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8S t r u c t u r e s 3 . Derivativesof heparin pentasaccharides


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222 H.P. W esselT a b l e 1 . Activities of hepa rin pentasaccharide 3 an d derivatives

3 4 5 6 7 8

anti-Xa activ itiy [U m g ~] 700 1250 1323 1529 1159 1611KD [nM ] 22 16 2 139 13

t h e p a r e n t p e n t a s a c c h a r id e 3 w i t h f r e e a n o m e r i c c e n te r, a n d t h a t r e p l a c e m e n t o fN-su l fa tes b y O-sul fa tes d id n ot d ecrease the ac t iv i ty .A s e r ie s o f m o d i f ic a t io n s w a s b a s e d o n t h e p e n t a s a c c h a ri d e 4 ( S t r u c t u r e s 3 ),w h ich ca r r i e s an add i t i ona l O -su l fa t e g roup a t t he reduc ing t e rminus and w asm o r e a c ti v e t h a n t h e r e f er e n c e c o m p o u n d 3 . T h e m e t h y l a te d a n a l o g u e 5 w a s s y n -the s i zed and p roved to have a s imi l a r a c t i v i t y . Rep lacement o f t he t e rmina lm e thy l g roup s b y n -b u ty l e the r s (6 ) i nc rea sed the an t i coagu lan t a c t iv i ty ,w h e rea s l on ge r a lky l cha ins l ed t o a r edu c t ion in a c t iv i ty . M oreove r , t he hy dro -pho b ic g roup s i nc rea sed the a f f i n i ty t ow ards A T I I I (Table 1 ); an i nc rea se i na f f in i ty (a l ow er K D ) w as , how ev e r , no t nece ssa r i l y l inked to h ighe r an t i-X aac t iv it y : h igh a f f i n i ty hep a r in saccha r ide de r iva tive s w i th on ly l ow an t i-X aac t iv i t y a re kn ow n [33].T h e v e r y l o n g s y n t h e s e s ( t h e f i rs t s y n t h e s is o f p e n t a s a c c h a r i d e 3 r e q u i r e d 7 5s t e p s ) c o u l d b e f a c il it a te d w i t h t h e i n t r o d u c t i o n o f t w o d i s a c c h a r i d e u n i t s t h a td i f fe r o n l y in t h e c o n f i g u r a t i o n o f th e u r o n i c a c i d a s i n 7 a n d 8 ( S t r u c t u r e s 3 ) .T h e i n v e r s i o n o f a n L - i d u r o n i c a c i d d i s a c c h a r id e i n to a D - g l u c u r o n ic a c i d a n a -l o g u e s a v e d t h e i n d e p e n d e n t c o n s t r u c t i o n o f a n o t h e r d i s a cc h a r id e a n d t h u s a napp rec i ab l e n um be r o f syn the t i c s t eps (Sch em e 1 ) [34]. I t i s i n t e re s t i ng to no t et h a t t h e s u b s t i t u t i o n o f th e 2 - O - s u l fa t e i n t h e L - i d u ro n i c a c i d m o i e t y b y a nO -m e thy l g ro up aga in i nc re a sed th e an t i -X a ac t iv i t y (Tab le 1 ), i nd i ca t ing tha th y d r o p h o b i c i n t e ra c t i o n s c o n t r i b u t e t o A T I I I b i n d i n g a n d c o m p e n s a t e f o r t h el o s s o f a n e l e c t r o s t a t ic s u l fa t e i n t e r a ct i o n . T h e h i g h n u m b e r o f h y d r o p h o b i cg r o u p s l e d t o a n i n c r e a s e i n e l i m i n a t i o n h a l f- li fe i n r a t o f u p t o m o r e t h a n 1 0 hin t he ca se o f pen ta sacc ha r id e 8 [34].F o r t h e i n h i b i t io n o f A T I I I - m e d i a t e d t h r o m b i n a c ti v it y , d e r iv a t iv e s o f 8w e r e p r e p a r e d c o n t a i n i n g a fl e xi b le s p a c e r o f a r o u n d 5 0 a t o m s p l u s a s u lf a t e dm a l t o o l i g o s a c c h a r i d e o r a n o t h e r A T I I I b i n d i n g p e n t a s a c c h a r i d e [ 35 ].Whi l e a l l t he se hepa r in saccha r ide de r iva t ive s a re s t ruc tu ra l l y c lose t o t hena tu ra l a c t ive pen ta saccha r ide , a d isadvan tage i s the co m plex syn the s i s com pr i -s ing m ore t han 30 st eps even fo r t he s imp l i fi ed ana logues (Scheme 1 ) w h ich ren -de rs a fu ll dev e lopm ent o f t he se co m po un ds l e s s likely .


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Hepar ino idM i m e t i c s 2 2 3

1 s t e p f ro m s t m 'c h

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4S u l f a t e d O l i g o s a c c h a r i d e s4.1Sulfated Linear 01igosaccharidesT h e w o r k o n t h e f i r s t s u l f a t e d o l i g o s a c c h a r i d e s w a s b a s e d o n i n v e s t i g a t i o n s o fthe polym eric heparinoids: derivat ives o f heparin had bee n prepared to obtaininform ation on the s tructural requirements of he parin for SM C ant|proli ferati -ve a ctivity. A clue for further studies w as brought about by carboxTl-reduced sul-fated hep arin (C RS-heparin, 9) in w hich al l carboxy1 groups were reduced and


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2 2 4 H . E Wessel

t h e r e s u l t i n g p r i m a r y h y d r o x y l g r o u p s w e r e s e l e ct iv e l y s u l f at e d ( S t r u c t u r e s 4 ) .C R S - H e p a r i n s h o w e d h i g h a n t i p r o l i f e r a t i v e a c t i v i t y b u t b a s i c a l l y n o A T I I I -m e d i a t e d a n t i c o a g u l a n t e f fe c t [3 6].C o n s e q u e n t l y , s u l f a t e d n o n - u r o n i c , m a i n l y ( 1 - - > 4 ) - l i n k e d o l i g o s a c c h a r i d e sw e r e p r e p a r e d . T r e s t a t in A s u l f at e ( 10 , S t r u c t u r e s 4 ) e m e r g e d a s h i g h l y a c t i v ec o m p o u n d d e v o i d o f A T I I Io m e d i a t e d a n ti c o a g u l a n t a c t iv i t y [ 36 ]. T r e s ta t in A , ap s e u d o - n o n a s a c c h a r i d e o b t a i n e d f r o m s t ra i ns o f Streptom yces dimorphogenes,i s a p o t e n t a - a m y l a s e i n h ib i to r . A s t h e m o l e c u l e a s s u m e s a n a m y l o s e - li k e h e li c alt e r t i a r y s t r u c t u r e i t w a s o b v i o u s t o c o m p a r e t h e a c t iv i t y o f s u l fa t ed m a l to - o l i-g o s a c c h a r i d e s. V e r y s u rp r i si n g l y , t h e s u l fa t ed a m y l o s e s u b s t r u c t u r e s h a d o n l yl o w a n t i p r o l i f e r a t i v e a c t i v i t y . M o l e c u l a r m o d e l l i n g s t u d i e s t h e n i n d i c a t e d t h a tt h e t r e h a l o s e m o i e t y o f T r e st a ti n A b e n d s o u t f r o m t h e h e li c al c o n f o r m a t i o n , s ot h a t T r e s t a ti n s u b s t r u c t u r e s c o n t a i n i n g t h e t r e h a l o s e e n d w e r e s y n t h e s i z e d . T h ea l l - a - l i n k e d o l i g o s a c c h a r i d e s w e r e a v a il a b le i n b l o c k s y n t h e s e s f r o m a t r e h a l o s eg l y c o s y l a c c e p t o r a n d a s u i t a b le m a l t o o l i g o o s y l d o n o r [ 37 ], t h e c r i t i c a l a -D - g l u -f ( = o , ( = o , ]9 O O

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Structures4 . Struc tures o f CRS-h epar in , Tres ta t in A su l fa te, subs t ru c tu re s the re of , an d f l - l inkeda n a lo g u e s . D S d e n o te s t h e d e g re e o f su l f a ti o n , d e f in e d a s a v e ra g e n u m b e r o f su lf at e g ro u p s p e rm o n o s a c c h a r i d e u n i t


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HeparinoidMimetics 225M a l t o t r i o s e T r e h a l o s e

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n) O 3 - N M e 3 ,D M F ; - H'O01~ 0N a O A c ,H 2 0 HO~,,~OH1-1

1 3.Scheme2. Synthesisof sulfated Trestatin A pen tasaccharide substructure 13

c o s y l l i n k a g e b e i n g a c h i e v e d w i t h h i g h s t e re o s p e c i fi c it y u s i n g t ri fl ic a n h y d r i d ea s a n e w p r o m o t e r ( S c h e m e 2) [ 38 ]. W h i l e th e t ri - a n d t e t r a sa c c h a r i d e s u b s t r u c -t u r e s 1 1 a n d 1 2 e x h i b i t e d n o a p p r e c i a b l e a c t iv i ty , t h e h i g h l y s u l f a t e d m a l t o t r i o -s y l t r e h a l o s e p e n t a s a c c h a r i d e 1 3 ( S t r u c t u re s 4 ) , a p e n t a s a c c h a r i d e s u b s t r u c t u r eo f T r e s ta t in w i t h a n a d d i t i o n a l t e r m i n a l h y d r o x y l g r o u p , h a d a n a n t i p ro l i fe r a ti v ea c t i v it y c o m p a r a b l e t o h e p a r i n [3 9].I n a n i n v e s t i g a t i o n o f a n a l o g u e s o f t h e s e s u b s t r u c t u r e s a ls o t h e c o r r e s p o n -d i n g / 3 - D - l i n k e d o l i g o s a c c h a r i d e s w e r e p r e p a r e d ; i n t h i s s e r i e s , t h e s u l f a t e df l-D - (1 --> 4 ) - l i n k e d m a l t o s y l t r e h a l o s e t r i s a c c h a r i d e 1 4 h a d a l r e a d y a p p r e c i a b l eS M C a n t i p r o l if e r a t i v e a c t iv i t y , a n d t h e a c t i v i t y o f te t r a s a c c h a r i d e 1 5 w a s c o m p a -r a b l e t o h e p a r i n ; t h e a c t i v i t y o f th e / 3 - D - ( 1 - - > 4 ) - li n k e d p e n t a s a c c h a r i d e 1 6 w a so n l y s l i g h t l y i n c r e a s e d o v e r t h e t e t r a s a c c h a r i d e a c t i v i t y [ 39 ]. T e t r a s a c c h a r i d e 1 5i s t h e s m a l l e s t s u lf a te d c a r b o h y d r a t e w i t h h i g h a n t i p r o l if e r at i v e a c t i v it y k n o w nso f ar . T h e f l - D - l i n k a g e o f g luco-con f igura ted b u i l d i n g b l o c k s i s a c h i e v e d e a s i ert h a n a n a - I s -l in k a g e a s i n 1 3 s in c e t h e s t e r e o d i r e c t i n g i n f u e n c e o f n e i g h b o u r i n gg r o u p s c a n b e e x p l o i t e d in t h e g l y c o s id e s y n th e s i s ; t h u s t h e s y n t h e s i s o f 1 5 w a ss h o r t (9 s te p s ) a n d p r o c e e d e d w i t h h i g h y i e l d s ( S c h e m e 3) [4 0].W i t h r e g a r d t o t h e d i s t i n c t ly d i f fe r e n t a c ti v i ti e s o f t h e a - a n d f l- D - li n k ed m a l -t o s y l t r e h a l o s e t e t r a s a c c h a r i d e s , i t i s i n t e r e s t i n g t o n o t e t h a t h i g h l y s u l f a t e da - i s -l i n k e d g l u c o s i d e s w e r e c o n f o r m a t i o n a U y s t a b le w h e r e a s t h e f l- is - li n k e d g lu -


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226 H.P. WesselM a l t o s e

I 2 s t e p s

3 0 r a i n , 9 9 % ;h ) H 2 , P d / C , E t O H / H 2 0 , R T , 2 h , 10 0 % .H O " ~ O H , ~ O H O HH O . . I k . . . ~ _ ~ , . , f -

O OS O 3 - N M e 3 ,D M F ; [ - ~ O Hn ) N a O A c .H 2 0 1 HO ~O OH1 5

Scheme 3 . Synthes i s o f su l fa ted ma hos y l f l -D-( l - - ) 4 ) -a , cc -t rehalose 15

T r e h a l o s e3 steps

O B n O ~

f ) S i lv e r r if la t e , T M U , C H 2 C I , o ~ .~0 ~ --e R T , 2 6 h , 9 1 % ; ]g ) N a O M e , M e O H / c y c l o h e x a n e , R T , gn

cos ides occu r red in con fo rm at iona l equ i l ib r ium, wh ich was , fo r example , shownfo r D-g lucu ron ic ac id s and me thy l D-g lucopyranos ides a s mode l compounds[41]. This i s a para l le l to the con form at ion al equi l ib r iu m of L-iduronic ac id inhep ar in wh ich was h ypo thes i zed to p l ay a ro l e i n b io log ica l ac tiv i ty ( see above) .On the o the r han d , t he d i f fe ren t ac t iv i ti e s o f t he a - and f l-D- linked ma l to sy lt reha lose t e t ra sacchar ides h in t ed a t a spec if ic i n t e rac t ion o f t he su l fa t ed ca rbo -hydr a t e w i th o the r b iomolecu les ; t he requ i rem en t fo r t he l oca li za tion o f su lfa te sa t spec i fi c pos i t ions was su ppo r t ed in an inves t iga t ion on t e t ra sacchar ide ana -logues o f 15 [42 ], in w h ich the ma l to sy l m o ie ty was rep l aced by d i f fe ren t equa to -r ia l ly l inked d isacchar ides . W hereas on ly the confo rm at ional ly s imi lar i somal toseana logue h ad a c om parab le ac t iv i ty , o the r mo d i f i ca tions , such as t he /3 -mal to serep la cem ent by f l-ceUobiose, led to a decrea se in ac t iv i ty of up to 70 % [43].The su l fa ted o l igosacchar ides d i scussed above w ere h igh ly , bu t no t comp le te -ly , su l fa t ed so tha t m ix tu res o f co m poun ds w i th a s l igh t ly d if fe ren t su l fat ion pa t -t e rn re su l ted . An a t t emp t to u nd ers t a nd w h ich su l fa t ion s it e was imp or t an t fo rthe b io log ica l ac t iv i ty o f t e t ra sacchar ide 15 was m ade b y the syn thes is o f mo no-and o l igodeox ygena ted ana logues o f 15. These t e t ra sacchar ides were p repa redin a b lock syn thes i s app roa ch co m parab le t o t he syn thes i s o f 15 , and the deoxyfunc t ions w ere e ssen t i a ll y i n t rodu ced o n the d i - o r t e t ra sacchar ide leve l. A re -p resen ta t ive syn the t i c exam ple is dep ic t ed in Schem e 4 [44 ].Indee d , som e s ing le deoxygena t ions - and thus t he absence o f a su l fa t e g roupa t t h is ve ry pos i ti on - l ed to a d ec rease in an t ip ro li fe ra t ive ac tiv i ty o f m ore th an


13/25

Hep arinoid Mimetics 2 2 7M a l t o s e , t r e h a l o s e

~ ~ o \ ~OA= s tep s' el" S eh ern e 3

g ) A c O H 8 0 & , 8 0 ~ 1 h , 9 1 % / r ~ ~ ~ - o _h ) 1 2 , P P h 3 , I m , C H 3 C N / to , , 7 0 ~ 1 h , 9 5 % ,t ~ O I r " O ~ ' - ' 'L .i ) H 2 , 1 0 % P d / C , d i m e t h o x y e th a n e /E t O H , ~ ~ 1 'N E t 3 , R T , 4 h , 8 5 % lo b

- - ' 1AcO I : t l~ 0

8.0j ) N a O M e , M e O H / E t 2 0 , R T , 4 h , 8 6 % , o - ~ o lk ) H 2 , 1 0 % P d / C , E t O H / H 2 0 , R T , 6 h , 9 5 % I t ~ " ~I ) S O 3 - N M e 3 , D M F ; N a O A c , H 2 0 ~ H ~ O H

~ /O R _O R _O Rx " 4 r R O . O RO 0 O :R O ' - R O ~ R O ~ = R O ~O ~ ' - ~ C H O RO" O R O P " O R 'q i ,, . .,IO R 3

R = S O z N a o r H , ~ - 8 3 % s u l f a t io nS d t e m e4 . Blo ck syn thes i s o f 6H - f f -ma l tosy l - (1 - -~ 4 ) -o t, o t -t rehaJose , a rnonodeox Tgena ted ana-l o g u e o f ] 53 0 % ( S c h e m e 5 ) [ 4 5] . T h e s u lf a te s c o n t r i b u t i n g t h e m o s t t o t h e a n t ip r o l if e r a ti v ea c t i v i ty s e e m t o b e l o c a t e d a t t h e b o t h e n d s o f th e t e t r a s a c c h a r i d e m o l e c u l e .

A par t f ro m i ts po t e n t an t i p ro l i f e ra t i ve ac t iv i t y , t e t r a s ac cha r i de 15 wa s e ff ec -t iv e in b l o c k i n g h u m a n c o m p l e m e n t i n v it r o a n d i n h i b i te d t h e r e le a s e o f h e p a -r a n s u lf a te f r o m c a r d i a c m i c r o v a s c u l a r e n d o t h e l i a l c e ll s. T o o v e r c o m e h y p e r a c u -t e r e j e c t i o n , t h e t e t r a s a c c h a r i d e h a s b e e n i n v e s t ig a t e d i n a g u i n e a p i g to r a t c a r -d i a c x e n o t r a n s p l a n t a ti o n m o d e l a n d s ig n i fi c a n tl y p r o l o n g e d t h e s u r v iv a l o fh e a r t r e c i p i e n ts w h e n c o m p a r e d t o c o n t r o l a n d h e p a r i n t r e a t e d g r o u p s [4 6].W h i l e t h e s i z e r e q u i r e m e n t f o r S M C a n t i p r o li f e ra t i v e a c t iv i t y h a d b e e n i n v e -s t i ga t ed fo r he pa r i n ( cf . Sec t. 2 .2 ), a s i m i l a r s t udy on f r a c t i ons o f h i gh l y ac ti veh e p a r a n s u lf a te h a s n o t b e e n c a r r i e d o u t . A f t e r th e d i s c o v e r y o f a c t i v e s u l fa t e dt e t r a s a c c h a r i d e s i t s e e m e d p o s s i b l e t h a t r e l a ti v e ly s m a U h e p a r a n s u lf a te s u b -s t r u c t u r e s w i t h t h e a n t i p r o l i f e r a t i v e a c t i v i t y o f h e p a r i n c o u l d b e i d e n t i f i e d .M o d e l h e p a r a n s u l f a t e o l i g o s a c c h a r i d e s w e r e d e v i s e d i n w h i c h t h e N - s u l f a t eg r o u p s w e r e r e p l a c e d b y O - su lfa te s a n d t h e g l u c u r o n i c a c i d s w e r e r e p l a c e d b yg l u c o s e s, f o r m a l l y c a r b o x y l - r e d u c e d a n d s u l fa t e d g l u c u r o n i c a c i d s - i n p a r a l l e lt o C R S - h e p a r i n ( se e a b o v e ). W i t h t h i s a p p r o a c h t h e h e p a r a n s u lf a te b a c k b o n e


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228 H. E W essel|

0.7/ 0 . 9

4 ", 6"H : r~ = 0.73 " ,3 " H : r~ = 0 .8 / 12 , 3 , 2 ' , 3 ' H : r = = 0 .6 0 .93 ,3 'H : r~= 0 .5 0 ~

S c h e m e 5. S u m m ary o f an t i p ro l i f e r a t i v e ac t i v i t i e s o f su l f a t ed m o n o - an d o l i g o h y d ro x y l a t edf l -m a l t o sy l - ( l - - ) 4 ) - t r eh a l o se t e tr a sacch a r i d e s . A r ro w s i n d i ca t e p o s i t i o n s o f m o n o d e o x y g en a -t i o n , a c t i v it i e s a r e g i v en a s r i ; v a l u e s , t h e r e l a ti v e i n h i b i t i o n co m p ared t o h ep a r i n . S ite s o nw h i c h m o n o d e o x y g e n a t i o n l e d t o m o r e t h a n 3 0 % l o ss o f a c t i v i ty ar e c i r cl ed

was reduced to an o~, f l - (1 - - - )4 ) -g lucan , i . e . a l te rna t ing a -D- and f l - D - ( 1 - - - > 4 ) -l i n k e d g l u c o s e s . S u l f a t e d m o n o - t o h e x a s a c c h a r i d e s u b s t r u c t u r e s w e r e s y n -t h e s i z e d a s m e t h y l g l y c os i de s ; t w o s e ri e s o f c o m p o u n d s w e r e i n v e s ti g a te d ,n a m e l y m e t h y l C ~ -D -g lu co sid es s u c h a s 1 7 a n d 1 8, a n d t h e f r a m e - s h i f t e d m e t h y lf l- D - g l u c o s id e s r e p r e s e n t e d b y 1 9 - 2 1 ( S t r u c t u r e s 5 ) .G i v e n t h o s e s i m p l i f i c a t i o n s t h e o l i g o s a c c h a r i d e s u b s t r u c t u r e s w e r e r e a d i l ya v ai la b le i n b l o c k s y n t h e s e s u s i n g g l u c o sy l o r m a l to s y l b u i l d i n g b l o c k s a s e x e m -p l i fi e d f o r t h e s y n t h e s is o f p e n t a s a c c h a r i d e 2 0 ( S c h e m e 6 ); t h e a n a l o g o u s[ 2 + 2 + 2 ] b l o c k s y n t h e s i s f u r n i s h e d t h e h e x a s a c c h a r i d e s u b s t r u c t u r e 2 1 [ 4 7 ] .P e n t a s a c c h a r i d e 2 0 a n d h e x a s a c c h a r i d e 2 1 r e a c h e d t h e a n t i p r o l i f e r a ti v e a c ti v i t yo f h e p a r i n ( T a b le 2 ) [ 4 8]. T h e d a t a s u g g e s t e d t h a t t h r e e f l - D - li n k e d g l u c o s y l u n i t sa r e r e q u i r e d t o e l ic i t t h e f u l l h e p a r i n a c t iv i ty . T h e s e h e p a r a n s u l fa t e m o d e l s a c-c h a r i d e s w e r e t h u s m o r e a c t i ve t h a n o v e r s u l fa t e d h e p a r i n f r a g m e n t s o f e q u iv a -l e n t s i ze .A s e r ie s o f a lk y l m a l t o - a n d l a m i n a r o a - o l i g o s a c c h a r i d e s h a s b e e n i n v e s t i g a t e df o r a n t i - H I V a c t iv i ty . A n a c t i v e r e p r e se n t a t i v e ( a n t i H I V E Cs0 - - 0 .2 m g m l -z ) isTabl e 2. R el a t i v e an t i p ro l i f e r a t i v e ac t i v i t i e s o f h ep a r i n an dh e p a r a n s u lf a te m o d e l o l ig o s a c c h a ri d e sCo m pd. 2 17 18 19 20 21r i 1.0 0.7 0.7 0.8 1.0 1.0


15/25

H e p a r i n o i d M i m e t i c s 2 2 9< o .

R O l l O i l . . .o l O I s . O i l ' " .I n O C H 3

1 7 R = S O a N a o r H , D S = 2 . 8

. ~ . _ OR ~ OR / O R ~ O R

R O l l 9 I C l l , , O I i , o)11Oil Oi l , ,l lOC~el

1 8 R = S O a N a o r H , D S = 2 . 8, ~ , / O R O R _ OR, : kOI , IO O ~O OCH~

1 9 R = S O a N a o r H , D S = 2.7

OR OR OR OR ~ OR

RO O= , i O O . O OCHa

2 0 R = S O 3 N a o r H , D S = 2 . 7

= . ,O R _OR . ,,OR _OR . ,.ORR O , ) O , I O O , I O O C H ~2 1 R = S O 3 N a o r H , D S = 2 . 7

Structures 5 . S t r u c tu r e s o f h e p a r a n s u l f at e m o d e l o l ig o s a c c h a r i d e s . D S = d e g r e e o f s u l f a ti o n ,c f . S t r u c t u r e s 4

th e s u lfat ed d od e c y l l am i n ar op e n taos id e 22 w i th c on c om i tan t ly l ow an t ic oagu l an tactivi ty. Th ese com po un ds probably have surface-active pro per ties [49] .S u c r os e oc tas u l fa t e (23 , S t r u c tu r es 6 ) b ou n d to F GF an d w as ab l e to i n d u c e ,l ik e h e p ar i n , a c on for m at i on a l c h an ge i n th e p e p t i d e , b u t s h o w e d ve r y l ow m i to -ge n i c ac t i v i ty for an F 32 c e l l l in e . T h e b as i c a l u m i n i u m s a lt o f s u c r os e oc tas u l fa -te , wh ich i s used as a dru g (Sucralfate , Carafate ) for the treatm ent o f duod ena lu l c e r s w as c l a i m e d to e xe r t p ar t o f i t s ac t iv i ty th r ou gh th e s tab i li z a t ion o f F GF


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230 H.P. W esselM al tose M ethy l l~ma i tos ide

1 2 s t e p s I 3 s te p sQAc OBn

~ ~ B~J f ) S i lver t r if la te , TM U , CH 2CI2,0 --> 35 ~ 5 h , 64 % ;g ) N a O M e , M e O H / E t 2 0 , R T ,. . . O HH ^ ~ \ _ O 16h , 7 8 %' ~ o . . . ~ . ~ o -

h ) P h C H O , Z n C I2, RT , 18 h , 79 %i) N a i l , DM SO , BnC I , RT , 3 h , 67 %j ) NaBH 3(CN , TH F, HC I /E t20 , 0 ~~o__~~ ol h , 8 2 %OR1+ - ~ 1 - - ~ ' ~ o ~ o ~

~ ~ e e H 3BnOI k ) S i lver td f la te , TM U , CH 2CI2,-oH | 0 -q> 35 ~ 7 h , 57 % ;H n - - ~ . ~ O x"O S n ~ I) N a O M e , M e O H / T H F ,H O ~ ~ n O ~ Oen R T , 1 6 h , 9 6 %

. o - - - og , o ~ o--'X --x~ o ,n n ~ r ~ I B r r u ~'n s

n) S'~O;-'~l'ge3,D M F ; N a O A c , H 2 0 1 B ~2 0

Scheme6 . [ 2 + 2+ 1 ] B lock syn thes i s o f hepa r an su l fa t e mode l pen tasacchar Jde 20

i n t he s tom ach [50 , 51 ]. A n X -ray o f a suc rose oc t a su l fa te - aFG F com plex w asp u b l i s h e d w i t h e i g h t d i ff e r en t a F G F m o l e c u l e s in t h e a s y m m e t r i c u n i t c el l i nw h i c h t h e s u lf a te g r o u p s a n d s u g a r r i n g s o f s u c r o s e o c ta s u lf a te a d o p t e d d i f fe r e n tc o n f o r m a t i o n s [ 52 ].

I n k e e p i n g w i t h a h i g h a m o u n t o f n o n - i o n i c c o n t a c t i n t h e b F G F / h e p a r i n i n t er -ac t i on i s t he recen t and in t e re s t i ng f i nd ing tha t non-su l fa t ed hepa ran-de r ived


17/25

Hepar ino id Mim et ics 231

2 2ORR O I I ' , ~

mOR

'~BO OR

OR

RO~ O- (C . ,~ 11C . ~-- '0 OR

R = SO3Na or H, DS = 3.0

2 3

Structures 6. Structure of a laminaro ol igosaccharide and sucrose octasul fate

d i - a n d t r i s a c c h a r i d e s c o n t a i n i n g o n e o r t w o u r o n i c a c id s a l so e l i ci t e d h e p a r i n o i da c t i v i t y b i n d i n g t o F G F a n d a c t i v a t e d t h e F G F s i g n a l l in g p a t h w a y [ 5 1].4 . 2S u l f a t e d C y c l ic O l i g o s a c c h a r i d e sC y c l o d e x t r in s a r e c y c li c m a l t o o l i g o s a c c h a r i d e s a va il ab l e b y e n z y m a t i c s y n t h e s e sf r o m s t a rc h . D u e t o t h e i r e a s y a cc e ss , s u l fa t e d c y c l o d e x t r i n s h a d b e e n i n v e s t ig a t e db a c k i n t h e e a r ly 1 9 6 0 s f o r t h e i r p l a sm a c l e a r in g a c t i v i t y [ 53 ]. M o r e r e c e n t ly , t h ea c t i v i t ie s o f f l - c y c lo d e x t r i n ( c y c lo h e p ta m a l t a o se , f l- CD ) t e t r a d e c a su l f a t e ( 2 4) w e r ee v a l u a t e d i n m o r e d e t ai l. I t w a s p r e p a r e d b y s u l f a t io n o f f l- C D w h i c h l e d t o t h es u l fa t io n o f th e p r i m a r y a n d o n e o f t h e s e c o n d a r y h y d r o x y l g r o u p s p e r p y r a n o s er i n g s o t h a t a m i x t u r e o f d i ff e r e n tl y s u l fa t e d c o m p o u n d s r e s u l t e d ( S t r u c t u r e s 7 ) .f l- C D T e t r ad e c a su l fa t e i n h i b i t e d t h e m i g r a t i o n o f s m o o t h m u s c l e c e lls a t h i g hc o n c e n t r a t i o n s , a n d t h e e f fe c t o n t h e p r o l i f e r a ti o n o f h u m a n a n d r a b b i t S M C w a sc o m p a r a b l e t o t h a t o f h e p a r i n [5 4]. T h e s a m e p r e p a r a t i o n w a s a ls o a c ti v e i n a na n g i o p l a s ty m o d e l i n r at s u p o n c o n t i n u o u s i n f u s io n , a l t h o u g h t h e h e p a r i n e ff ec ti s n o t r e a c h e d e v e n a t v e r y h i g h d o s e s ( 50 m g k g -1 d -1) [55] . S ulfa ted f l -CD ex hi-b i t e d p r a c ti c a l ly n o a n t i- X a a c t i v it y b u t s o m e A P T T ( a c ti v a te d p a r t i a l t h r o m b o -p l a s t in t i m e ) c l o t t in g a c ti v it y , a l t h o u g h r e d u c e d v s h e p a r i n . V e r y r e m a r k a b l y ,s u l f a t e d f l- C D w a s a l s o a c t iv e i n t h e r a b b i t m o d e l a f t e r o r a l d o s a g e [5 6 ]. " P o l y -m e r i c " s u l f a te d f l- C D w a s s h o w n t o h a v e a n a f f i n i ty fo r b a s i c F G F [5 7].F u r t h e r m o r e , su l fa te d fl- C D m i m i c k e d h e p a r i n i n t h e a u g m e n t a t i o n o f a n t i-a n g i o g e n i c a c ti v i ty o f a n g i o s ta t i c s te r o i d s, b u t u s e d a l o n e p r o m o t e d b l o o d v e s -s e l c e ll g r o w t h ( a n g i o g e n e s i s ) [5 8 ].W h e r e a s s u l f a t ed f l- C D a n d y -C D e x e r t e d c o n s i d e r a b l e a c t iv i t y a g a in s t t h eh u m a n i m m u n o d e f i c i e n c y v i ru s e s H I V -1 a n d H I V -2 [5 9], th e m o d i f i c a ti o n w i thh y d r o p h o b i c g r o u p s c o u l d i n cr ea s e t h e i n v i tr o a c t i v it y d e t e r m i n e d b y t h e i n h i -


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232 H.P. Wessel

2 4 R = S O a " o r H , 5 0 % su lf at io n o nsecondaryhydroxy ls

! ~.- OI . - , l l O

R

2 5 R = S O 3 K o r H( n u r n b e r o t S O 3 K = 1 6 )mCDS11

I l l . . 0

2 6 R = S O 3 N amCDS71

Structures 7. Structures of fl-CD tetradecasulfate and of modified cyclodextrins

b i t i o n o f t h e c y t o p a t h i c e f f ec t i n M T - 4 c ells . U s e f u l s u b s t i t u t i o n s w e r e t h e i n t r o -d u c t i o n o f t h r e e S - b e n zy l g r o u p s i n t h e p r i m a r y p o s i t i o n s o f fl-C D t e tr a d e c a s u l-f a t e t o g iv e 2 5 ( mCD S1 1 , S t r u c tu r e s 7 ) [ 60 ] o r , p a r t i c u l a r ly , O - b e n z y la t i o n o f al l2 - p o s i t i o n s t o r e s u l t in 2 6 ( m C D S 7 1 , S t r u c t u r e s 7 ) [ 61 ]. T h i s c o m p o u n d w a s p r e -p a r e d f r o m f l- C D b y s il yl p r o t e c t i o n o f th e p r i m a r y h y d r o x y l g r o u p s a n d r e g io -s e le c ti v e b e n z y l a t i o n o f t h e s e c o n d a r y h y d r o x y l g r o u p s i n p o s i t i o n 2 f o ll o w e d b yd e s i l y l a t io n a n d s u l f at i o n . A s a p e r s u l f a t i o n w a s p o s s i b l e , 2 5 t u r n e d o u t t o b e ac h e m i c a l ly d e f i n e d c o m p o u n d . A d v a n t a g eo u s ly , t h e s u b s t a n c e w a s o r a ll y a v ai l-a b l e a n d l e d t o a p p r e c i a b l e p l a s m a l ev e ls in t h e r a t [6 1].

27

Structure 8. Structure of calixarene 27


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Hepadnoid M imet ics 233The su l fona ted ca lix [8 ]a rene 27 (S t ruc tu res 8 ) , wh ich can be v i ew ed as a s im -p l i fi ed y -CD wi th th e ca rbo hyd ra t e backbo ne rep laced by an a ry l mo ie ty ,m i mi c k e d h e p a r i n i n t h e s t imu l a t i o n o f h e p a ra n s u lf at e sy n t h e s is f r o m c u l t u r e dend othel ia l ce l l s [62].

5Su l fa ted Spaced Sugars5.1S u l f a t e d S p a c e d O l i g o s a c c h a r i d e sBased on the f i nd ing tha t su l fat e s i n t he c en te r pa r t o f t he t e t ra saccha r ide 15 m ayno t be o f u tm os t im por t a nce fo r SMC an t ip ro l ife ra tive ac tiv ity , su l fa ted b i s-g ly -c o s id e s o f a ro m a t i c b i s-h y d ro x y l c o m p o u n d s h a v e b e e n p r e p a re d ; t h e y w e re c a l-l ed su l fa t ed ' spaced o l igosacchar ides ' a s fo rm a l ly a p a r t o f an o l igosacchar idewas rep laced by a space r g roup . The advan tage o f t hese s t ruc tu res was tha t t heno n-ch a rge d space rs avo ided the ch a rge in te rac t ions wh ich in t he o l igosaccha-r ides d id n o t a l low fu ll su lfa ti on ; a s exp ec ted f rom su l fa tion s tud ies on d i sac -cha r ides t he spaced o l igosacchar ides cou ld be pe rsu l fa t ed , i f t he saccha r idem oie t i e s were no t l a rge r t han d i saccha r ides , and thus y i e lded chem ica l ly de f i-ned com pou nds . H igh an t ip ro l ife ra tive ac tiv it ie s cou ld be ach ieved in t h i s se r ie so f com poun ds , e.g . com po un d 28 (S t ruc tu res 9) was a s ac t ive a s hepa r in , and2 9 w a s e v e n 2 0 % m o re a c ti ve t h a n h e p a r i n , b u t n o A T I I I - m e d i a t e d a n t ic o a g u -l an t e f fec ts we re obse rved [63 ]. These h igh ac t iv i ti e s po in t ed a t a b ind ing con -t r ibu t ion o f the hyd rophob ic space r.

2 8. O R ~ O R A

. o " . o " - -- o R

2 9(3~OR . ,,OR / / \ \ RO OR~ " o . " ~ Ro... o~ _ . . .

o o. o -X_o .R = SOaNa

Structures9. Struc tures of sulfated spaced oligosaccharides


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23 4 H.P. W essel5 .2S u l f a t e d S p a c e d O p e n C h a i n S u g a r sO p e n c h a in C -6 s u g a rs i n e x t e n d e d c o n f o r m a t i o n h a v e a p p r o x i m a t e ly th e s a m espa t ia l ex t ens ion a s a d i saccha r ide such a s m a l tose . Su lfa ted spaced open cha insuga rs w e re t he re fo re i nves t i ga t ed i n ana logy to t he su l fa t ed spaced o l igosac -cha r ide s . G lycamines , and pa r t i cu l a r ly commerc i a l l y ava i l ab l e g lucamine(1 -amino- l -deoxy-D -g luc i to l ) , cou ld be read i ly l i nked to va r ious a roma t i cspace rs v i a l i nke r g roups such a s amines , su l fonamides , u rea s , and amides a se x e m p l i f ie d b y c o m p o u n d s 3 0 - 3 3 ( S t r u c tu r e s 1 0) . O t h e r l in k e r g r o u p s i n c l u d e de the rs , t h iourea s , and inve r t ed am ides . A ll com po un ds w e re comp le t e ly su l fa tedand thus chem ica lly w e l l de f ined . The s yn the t i c a cce ss t o t h is t yp e o f com po un dw as s t ra igh t fo rw a rd and ex t reme ly shor t ; i n t he s imple s t c a se unpro t ec t edg lucam ine co u ld be l inked to a b i s - fun t iona l i z ed space r an d b e su l fat ed , and insom e ca se s t he i n t e rmed ia t e p ro t ec t ion o f hydrox y l g rou ps w as adv i sab l e t o fac i-l i ta te pur i f ica t ion .

3 0OR (~)R O " ~ ' ~ S ~ NO- OR OR

OR OR OR OR

31OR OR OR OR

OR OR OR OR

3 2OR ?R r f ~ OR Q R

R O - ~ ~ . ~ _ _ . ." N H - C O ' N ~ N - C O - H N ~ ~ OROR OR OR OR

3 3O R 01:1

OR oR ~ I ~- ORR O ~ NH . C ~ " - ' / ~ - - CO.HN ~ ~ - /~- - ~ ~ OR OROR ORR = SOaNa

Structures 10. Struc tures o f su lfated space d glucamines w ith arom atic spacers

The inves t iga t ion o f t he an t ip ro l if e ra ti ve a c t i v i t y on ra t SMC cu lmin a t ed int h e i d e n ti f ic a t io n o f h ig h l y ac ti v e c o m p o u n d s w i t h 5 0 - 6 0 % m o r e a c t i v i ty t h a nheparin (Table 3) [63].In a n A PT T c lo t t i ng a ssay , w h ich re f l e c ts no t on ly t he A T I I I - m ed ia t ed even t sb u t a l s o H C I I a n d o t h e r f a c to r s o f t h e c o a g u l a ti o n c a s ca d e b e s id e s t h r o m b i nand fac to r X a , su l fa ted spaced o pen cha in suga rs re ach ed se l ect i v it ie s aga ins t


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Hep arinoid Mimetics 2 3 5Table 3 . A c t i v it i es o f s u l fa t e d s p a c e d o p e n c h a i n s u g a r sC o m p o u n d 2 3 0 3 1 3 2 3 3Rela t ive an t ip ro l i fe ra t iv e 1 .0 1 .0 1 .0 1 .5 1 .6a c t iv i t y r iA n t i c o a g u l a n t a c t i v i tyan ti- II a/ an ti- X a 2.2/2.7 > 1000/ > 1000/ > 1000/ > 1000/ICso [lag m1-1] > 1000 > 1000 > 1000 > 1000

Table 4 . A P T T s d e c t i v i t i e s o f d i f f e r e n t s u lf a te d s u g a r d e r iv a t iv e s . IC so v a l u e s a r e c o m p o u n dc o n c e n t r a t i o n s l e a d i n g to a c l o t t i n g t i m e o f t w i ce t h e c o n t r o lC o m p o u n d C o m p o u n d c l as s r i A P T T IC so [ la g m l q ]2 Poly sacc har ide 1 .0 1 .39 Po lysacch . der iv a t ive 1 .2 111 0 N o n a s a c c h a r id e 1 .2 715 Te t ras acch ar ide 0 .9 122 8 S p a c e d s a c c h a r id e 1 .0 7 73 3 S p a c e d o p e n c h a in s u g a r 1 .5 > 1 0 00

_ c s o i _ o s o 3 " o

( : :~O i = (:3S0~

% 3

a ) L e w a t i t - S - 1 0 0b ) N H 2 ( C H 2 ) 3N H 2 , D M Fc ) S O 3 . P y r i d i n ed ) 3 0 % N a O H

oso~osoi9 9..osoi_~oi- 0 3 S O ~ N~ N ~ C 6 0 ~

: -- H H 9 9O S O ~ : O S O ~ O S O ~ 0 , , 0 , , o ~ _ _

3 4 ~ ~ceo ". -~-" u~oi" 0 3 S O " " O S O ~-03S 3"O SO ~ 5SO="Scheme7 . S y n t h e s is a n d s t r u c t u r e o f A p r o s u la t e


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236 H.P. Wesselh e p a r i n o f mo re t h a n t h r e e o rd e r s o f m a g n i t u d e d e p e n d i n g o n t h e c h o i ce o fspa cer (Table 4).Fro m a ser ies of su l fa ted b is-a ldonic ac id am ides wi th d i f feren t a lky l spacerl eng th , com po und 34 was chosen fo r fu r the r eva lua t ion a s an an t i t h romb o t i c .This c om po un d was synthe t ica l ly avai lab le in fo ur s teps f rom lac tobion ic ac id(Schem e 7) . Co m poun d 34 had re la t ive ly h igh AP TT va lues (42 U /m g) and an t i-t h ro m b o t i c a c ti vi ty , b o t h o f w h i c h d e c r e a s e d g r a d u a ll y w h e n t h e n u m b e r o fm e thy len e g roups in t he space r w as inc reased . I t was thoug h t t o ac t v i a HC I Ian d m ul t ip le s ites in the in t r ins ic pa thw ay of the co agula t ion cascade [64].U n d e r t h e n a m e o f A p ro s u la te c o m p o u n d 3 4 u n d e rw e n t p h a se I c li ni ca lt r ia ls . W herea s the d rug was wel l to lera ted and eff icac ious , t was asso cia ted wi thinc reased t ran sam inase leve ls [65 ]. The deve lopm en t o f Aprosu la te so d ium wassusp end ed in March 1995.6Non-Carbohydrate C ompoundsFor obv ious reasons o f s t ruc tu ra l ana logy to hepa r ino ids t he focus o f th i s rev i ewis on su l fa ted carboh ydr a te der iva t ives .Whi le it is no t in a ll cases c lear tha t thesecom pou nds rea l ly m imic the phys io log ica l ac tiv i ty o f hepa r ino ids , i t is evenless so for n on -car bo hyd ra te su lfa tes or su l fonates. Exam ples of the la t te r c lassinc lude sur am in an d the s im ple 1 ,3-propanedio l d isu l fa te . Su ram in is a su l fonat -ed b i s -naph tha l ene de r iva tive u sed as a d r ug to t rea t Afr i can t rypano som ias i san d o ncho cerc ias i s (a f ila ria l in fec t ion) ; i t was a l so tes ted in a nu m be r of o th erind ica t ions i nc lud ing ad renoc o r t i ca l ca rc inomas a nd AIDS. A w ide r u se is ,howev er , res t r ic te d b y var ious tox ic effec ts [66] . 1 ,3-Propanedio l d isu lfa te redu c-ed in f l am m at ion -assoc ia t ed am ylo id p rog ress ion in v ivo a f t e r o ra l admin i s t ra -t i on w h ich m ay be re l evan t t o t he t rea tm en t o f A lzhe imer ' s d i sease [67 ].7Concluding Rem arksThi s com pi l a t ion o f hep a r ino id m imet i c s t ha t have evo lved ove r t he l a s t yea rsdem ons t ra t e s t ha t t he re i s a t enden cy to l a rge ly s imp l if i ed com pou nds w i threga rd to s t ruc tu re and syn the t i c access ; a min imum requ i remen t fo r fu r the rp h a rm a c e u t i c a l d e v e lo p me n t s e e ms t o b e c h e mi c a l h o mo g e n e i t y . O n e o f t h er e c ip e s to i n c re a s e b i n d i n g a n d m o d i fy p h a rma c o k i n e t i c p a r a me t e r s w a s t h ei n t ro d u c t i o n o f h y d ro p h o b i c g ro u p s.In s om e o f the c a rboh ydra t e mim et i c s d i scussed above the g lycos id i c l inkagewas rep lace d by o th er l inkers such as, for exam ple , an am ide grou p (cf . Sect . 5 .2).In an in t e res t ing app roach towards new mimet i c s , ca rbohydra t e amino ac idshave been employed as bu i ld ing b locks wh ich can be eas i ly coup led us ingpep t ide t echno logy , i nc lud ing so lid phase syn thes i s. These m imet i c s p rov idet empla t e s fo r t he pos i t i on ing o f su l fa te s and m a in t a in pa r t s o f t he ca rbohy dra t eepi tope [68].W i th rega rd to d ru g d eve lopmen t , t he ques t ion o f se l ec tiv i ty w i ll r em a in acr i t ica l i ssue and the m ain chal lenge to avoid und esi re d s ide effec ts . However ,


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Hepar ino id Mim et ics 237t h e h i g h s e l e c t i v i ty a c c o m p l i s h e d a l re a d y , f o r e x a m p l e , w i t h t h e s u l f a t e d s p a c e do p e n c h a i n s u g a r 3 3 a g ai n st th e n u m e r o u s h e p a r i n o i d - b i n d i n g p r o te i n s i n v ol -v e d i n t h e c o a g u l a t i o n p r o c e s s, m a y s i g n a l th a t a c h i e v e m e n t o f s e le c t iv i t y w i l ln o t b e a n i n s u r m o u n t a b l e t a s k . T h e fi n a l a n s w e r s w i l l b e g iv e n i n th e c o m p l e x i nv i v o s i tu a t i o n .References

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