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Immunologic Research 1998; 17/3:313-32 7

The Intercellular Adhesion Molecule(ICAM) Fam ily of ProteinsNew Membersand Novel Functions ICOS Corporation, Bothell, WA.

AbstractMacro molecu lar adhesive associations between cel ls are

important for transmitting spatial and temporal informationthat is critical for imm une sy stem function. One such group ofproteins, the intercellular adhesion molecules (ICAMs), hasgrow n as newly identified mem bers are revealed. In addition,the functions of the ICAM s, in general, have begun to be betterunderstood, including intracel lular signal ing events. Thisinformation has led to the design of novel therapeutic agentsthat may prove ef fective in a variety of disease states.

Key WordsIntercellular adhesion mol-eculesSignalingTherapeutics

IntroductionIntercellular adhesion molecules (ICAMs),

are a family of related cell surface glycopro-teins as defined by the criteria of polypeptidesequence homology, conserved [~2 integrinligands, and, in most cases, genetic linkage.Each IC AM protein has extracel lular C-typeimmunog lobu l in - l i ke domains rang ing innumb er from two to nine making them mem -bers of the immun oglobulin superfamily (1) .Desp ite their structural similarity and integrin-binding capability, these proteins are expre ssedon a wide range of cell types, with very dis-tinct patterns of gene regulation and down-stream effector behaviors. Rece nt data reveal

that ICAM engage men t triggers outside-in sig-nals that effect cellular responses. This articlesumm arizes the current level of understandingregarding ICA M-triggered outside-in signalingevents and recent developments in the area ofnewly characterized ICAM proteins.

With the discovery of new mem bers of thisprotein family, we propose that a commonnaming system be adopted for the currentlyknown ICA M fam ily mem bers. These and anyadditional members should be given numeri-cal designations based on the chronology oftheir discovery. Given the numerical assign-ments to ICAM- 1, ICAM-2, and ICAM-3, webel ieve i t i s appropria te to des ignate the

Dr. Joel S. Hayt]ick 9 1998[COS Corp. Huma na Press Inc.22021 20th Ave. SE 0257-277X/98/Bothell, WA 98021 17/3:313-32 7/$11.75E-mail: [email protected]

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L a n d s t e i n e r - W e i n e r ( L W ) b l o o d g r o u p an ti -g e n a s IC A M - 4 a n d t e l e n c e p h a li n a s IC A M - 5 .ICAM- ! (CD54)

I C A M - I i s a 7 6 - 1 1 4 - k D a s u r f a c e g l y c o -p r o t e i n th a t h a s fi v e e x tr a c e l l u la r i m m u n o g l o -b u l i n - l i k e d o m a i n s ( 2 , 3 ) . A m o t i f w i th i nd o m a i n s 1 a n d 3 o f I C A M - 1 h a s b e e n s h o w nto be c r i t i ca l fo r b ind ing to the ~ 2 i n t egr insl y m p h o c y t e f u n c t i o n - a s s o c i a t e d a n t i g e n(LF A)- 1 (CD 1 I a /CD 18) and M ac- 1, r e spec -t iv e l y (C D 1 l b / C D I 8 ) (4,5). I n t e g r i n b i n d i n gt o I C A M - 1 i s r e g u l a t e d b y a f f i n i t y a n d a v i d i t yc h a n g e s . T h r o u g h t h e s e i n te r a c t i o n s w i thL FA - 1 a n d M a c - 1 , I C A M - 1 p l a y s a n i m p o r -t an t r o l e i n l e u k o c y t e - l e u k o c y t e , e p i t h e l i a l -l e u k o c y t e , a n d e n d o t h e l i a l - le u k o c y t e c o n ta c t.As p a r t o f b o t h t h e c e l l u l a r a n d h u m o r a l i m -m u n e r e s p o n e d u r i n g l e u k o c y t e - l e u k o c y t ec o n t a c t , IC A M- 1 se r v e s as an a c c e s so r y p r o -t e in f o r a n t i g e n r e c e p t o r a c t i v a t io n o n B a n d Tcel l s (6) (see R e c e n t D e v e l o p m e n t s i n O u t -s i d e -I n S i g n a l i n g a n d C y t o p l a s m i c F u n c t i o n so f I C A M P r o t e i n s ) . E x p r e s s e d b a s a ll y o nm a n y c e ll l i n e ag e s o f m e s y n c h y m a l o r ig i n , th el ev e ls o f s u r f a c e I C A M - 1 p r o t e i n a r e m a r k -e d l y u p r e g u l a t e d a t t h e t r a n sc r i p t i o n a l l e v e lb y i n f l a m m a t o r y s t im u l i l ik e t u m o r n e c r o s isf a c t o r ( T NF ) , i n t e r fe r o n y a n d l i p o p o l y sa c c h a -r i d e (L PS) . T h e p r o t e i n a l so p l a y s a m a j o r r o l ed u r i n g l e u k o c y t e tr a n sm i g r a t i o n f ro m t h e v a s -c u l ar c o m p a r t m e n t d u r in g i m m u n e s u rv e il -l a n c e . I n th i s c o n t e x t , l e u k o c y t e s a r e t e t h e r e dv i a sp e c i f ic c a r b o h y d r a t e m o i t i e s to s e l e c t i n s ,a s we l l a s ve ry l a t e an t igen-4 (VLA-4) b ind-i n g to V C A M - 1, b o t h o f w h i c h m e d i a t e le u k o -c y t e r o l l in g o n c y t o k i n e - a c t i v a t e d e n d o t h e l i a lc e ll s . L e u k o c y t e s b e a r i n g h i g h - a f f i n it y L FA - 1o r M a c - I t h e n a d h e r e f i r m l y t o t h e e n d o t h e -l i u m b y b i n d i n g t o I C A M - I t o i n it ia te t h ep r o c e s s o f tr a n s m i g r a t o r y e g r e s s f r o m t h ev a s c u l a t u r e (7). G i v e n t h e a f o r e m e n t i o n e dr o l e s o f I C AM - 1, m i c e d e f i c i e n t in i ts e x p r e s -s i o n t h r o u g h t a r g e t e d d i s r u p t i o n o f t h e g e n e

h a v e n u m e r o u s in f l a m m a t o r y r e s p o n s e a b n o r -m a l i t ie s , i n c l u d i n g i m p a i r e d n e u t r o p h i l e m i -g r a t i o n , a n i n a b i l i t y t o s t i m u l a t e T c e l lp r o l if e r at io n i n m i x e d a l lo g e n e i c l y m p h o c y t ec u l t u r e s , r e s i s t a n c e t o s e p t i c sh o c k , a n d ar e d u c e d su sc e p t i b i l i t y t o c e r e b r a l i s c h e m i a -r e p e r f u s i o n i n j u r y ( 8 - 1 0 ) .I n t e r e s t i n g l y , I C AM- I i s u t i l i z e d b y s e v -e r al p a t h o g e n s t o i n f e c t h o s t c e l ls a n d e v a d ei m m u n e d e t e c t i o n . I t w a s i d e n t i f i e d a s t h em a j o r c e l l- su r f a c e r e c e p t o r f o r r h i n o v i r u s a n dP l a sm o d i u m f a l c i p a ru m ( 4 ,1 1 ). HI V p a r t ic l e sh a v e b e e n f o u n d t o i n c o r p o r a t e IC A M - 1 i n t ot h e v ir a l e n v e l o p e d u r i n g t h e b u d d i n g p r o c e s s ,r e su l t ing in inc rease d in fec t iv i ty tha t is ICA M -1 / L F A - I d e p e n d e n t (12) .ICAM -2 (CD1021

T h e e x i s t en c e o f I C A M - 2 , a s e c o n d [3 2i n t egr in l i gand , was in fe r red by the inab i l i t yo f I C A M - 1 sp e c i f ic a n t i b o d i e s t o c o m p l e t e l yb l o c k t h e a d h e s i o n o f l e u k o c y t e s to e n d o t h e -l i a l m o n o l a y e r s (13) . I C A M - 2 i s e x p r e s s e dc o n s t i t u t i v e l y at l o w l e v e l s o n r e s t i n g l y m p h o -c y t e s a n d m o n o c y t e s a n d a t h i g h e r l e v e l s o nvascu la r endo the li a l ce ll s , bu t i s no t up regu la t edi n r e s p o n s e t o i n f l a m m a t o r y s t i m u l i a s i sI C A M - 1 (14) . I t s t wo e x t r a c e l l u l a r i m m u n o -g l o b u l i n - l i k e d o m a i n s a r e m o s t c l o se l y r e la t e dt o th e a m i n o t e r m i n a l d o m a i n s o f I C A M - l (15)(see T a b l e I f o r c o m p a r i so n s ) a n d v i a b i n d i n go f L F A - l , I C A M - 2 p l a y s an i m p o r t a n t r o le i ne n d o t h e l i a l - l e u k o c y t e i n te r a c ti o n s a s s o c ia t e dw i t h i n f la m m a t i o n (14) . E v i d e n c e a l so e x i s tst o su g g e s t th a t Ma c -1 is a li g a n d f o r I C AM - 2in a m a n n e r s i m i l a r t o t h a t p r e v i o u s l y d e m o n -s t ra t e d f o r I C AM - 1 (16) .

An o u t s t a n d i n g i s su e y e t t o b e r e so l v e d i swh y e n d o t h e l i a l c e l l s e x p r e s s t wo d i f f e r e n tI C AM p r o t e i n s t h a t h a v e t h e s a m e i n t e g r i nl ig a n d , L F A - I . C o u l d t h e i r r e l a ti v e s i z es b e ac l u e t o t h e i r ro l e s i n a m u l t i - s t e p p r o c e s s t h a tr e su l t s i n f i r m l e u k o c y t e a t t a c h m e n t t o a c t i -v a te d e n d o t h e l i u m ? T h e s h a p e o f t h e I C A M - 1

314 Hayf l i ck , Ki lgannon , and Gal la t in


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Table ! , Hum an ICAM fami ly Domains I and 2 (% amino ac id i den ti t y )"

ICAM-1 34 77 26 64I C A M - 2 3 4 38 27 42ICAM -3 38 36 27 64ICAM -4 30 30 30 28ICA M-5 25 33 31 36

aDomain 1, boldface; do ma in 2, italic.

e x t r a c e l l u l a r r e g i o n w a s f o u n d t o b e a nextended rod of approx 16.6 nm (4). Eachimm uno glob ul in dom ain therefore forms sub-regions of 3-3.5 nm. ICAM -2 sho uld measure6.5 nm putt ing i ts LFA-1 binding si te muchcloser to the nega t ive ly charged plasma m em-brane. In a hypothetical stepwise model, LFA - 1wou ld engage ICAM-1 in it s fi rs t domain a tabout 16 nm fro m the l ipid surface. This bind-ing could displace negatively charg ed surfacemolecules on the oppo sing cells and el ici t bend-ing of the ICAM bringing dom ain 3, at approx9 nm from the surface, w ithin proximity of Mac- 1for binding to occur. As the cells are drawntogether and i f repuls ion by the nega t ive lycharged glycocalyx on each cel l ' s surface isreduced in this manner , then LFA-1 binding toICAM -2 at about 6 nm from the surface couldthen occur resulting in firm attachment.ICAM- (CDSO, CAM-R)

ICAM-3 was soug ht by us and others basedon da ta tha t ICAM-l-spec i f ic and ICAM-2-specif ic antibody treatment par t ia l ly blockedLFA- l -dep ende nt homoty pic T ce l l aggrega-tion (14). U t il iz in g p r i m e r d e g e n e r a c y a n dt h e a m i n o a c i d s e q u e n c e c o n s e r v a t i o nb e t w e e n I C A M - 1 , - 2 a n d o t h e r C 2 - t y p eimmu noglo bul in domains , a polymerase cha inreac t ion product was genera ted and used top r o b e a c D N A l i b r a r y f r o m t h e h u m a nmyelo mon ocyt ic ce ll l ine , HL60. The deduce damino ac id sequence revea led tha t ICAM-3,

l ike ICAM-1, was com pos ed of five ext ra -c e l l u la r i m m u n o g l o b u l i n - l i k e d o m ai ns(17-19). Dom ain- spec i f i c sequence hom ol -ogy wi th ICAM-1 ranged from 36-77% , wi thdomain 2 be ing most h ighly conserved (seeTable 1 fo r com par i son o f the f ir s t twodomains) and the region of leas t homologybe ing the cy top la sm ic t a i l , which m ay beref lected by differences in outside - in signalst r iggered by engagement of ICAM-3 versusICAM-1 (see Recent Developments in Out-s ide-In Signa l ing and Cy toplasmic Funct ionsof ICAM Proteins) .

The integr in-binding propert ies and expres-sion pattern of ICAM -3 p ar t ial ly over lap withthose o f ICAM -1 and -2 . ICAM -3 b indsLFA - l , but apparently not Mac- 1 (20,21). TheICAM -3 binding s i te wi thin LFA-1 has beenlocalized to regions within the evolution ar i lyconserved I d omain of the ~-cha in (CDI l a )that over laps with, but is dist inct f rom, the si tefor ICAM - 1 binding . This obse rvation is use-ful for those laborator ies t rying to developI C A M - s p e c i f i c f u n c t i o n a l i n h i b i t o r s (seeN o v e l A r e a s o f T h e ra p e u t i c D e v e l o p m e n tInvo lv ing ICAM Bio logy) (22,23). In addi-t ion to LFA- 1 binding, ICAM-3 has been iden-t if ied as the prefered l igand for O~d~2, a recentlyd i scove red m em ber o f t he [~2 integr in family(24). In humans , th is in tegr in i s expressedhighly on cer tain t issue leukocytes, par t icu-la r ly macrophages , and on subse ts of b loodleukocytes (24). Given tha t the no rmal expres-

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s i o n p a t t e r n o f I C A M - 3 is h i g h o n a l l le u k o -c y t e s , b u t l a c k i n g o n e n d o t h e l i u m , it is l i k e lytha t i t p l ays an im por t an t ro l e in ea r ly even t sd u r in g l e u k o c y t e - l e u k o c y t e c o n t a c t , f o re x a m p l e B c e l l a c ti v a t i o n m e d i a t e d b y T c e l lh e l p o r T c e ll a c t i v a t i o n b y a n t i g e n - p r e se n t i n gc e ll s ( A P C s ) . M o n o c l o n a l a n t i b o d ie s ( M A b s )t o I C AM - 3 h a v e b e e n sh o w n to i n h i b i t p e r i p h -e ra l b lood lym ph ocy te p ro l i f e ra tion in re spon set o p h y t o h e m a g g l u t i n i n , a ll o g e n e ic s t im u l a t o rce l l s , and sp ec i f i c an t igen (25,26). I C A M - 3 ish i g h l y e x p r e s s e d o n e p i d e r m a l L a n g e r h a n sc e l ls w h e r e p r e s e n t a t i o n o f a l l o a n t i g e n o r p e p -t ide an t igen by thes e ce l l s t o CD4 + T ce l l s m ayr e q u i r e a n i n t e r a c t i o n b e t we e n I C AM- 3 a n dL F A - 1 ( 2 7 - 30 ) . T h u s , I C A M - 3 h a s an i m p o r -t a n t r o l e i n T c e l l s t i m u l a t i o n d r i v e n b yL a n g e r h a n s c e l l s . H o w e v e r , w h e n a c h r o n i ci n f l a m m a t o r y r e a c t i o n i n t h e s k i n o c c u r s ,T c e l l s t i m u l a t i o n l e a d s t o p a t h o l o g i c a l c o n -d i t i o n s s u c h a s p s o r i a s is , s o b l o c k a d e o fI C A M - 3 / L F A - 1 b i n d i n g m a y b e a n a t t r a c -t i v e t h e r a p e u t i c t a r g e t ( see N o v e l A r e a s o fT h e r a p e u t i c D e v e l o p m e n t I n v o l v i n g I C A MB i o l o g y ) ( 31 ) . U n l i k e i t s c l o s e r e l a t i v e s ,I C A M - 3 is n o t e x p r e s s e d o n n o r m a l o ra c t iv a t e d e n d o t h e l i u m p r e c l u d i n g i ts i n v o l v e -m e n t i n l e u k o c y t e - e n d o t h e l i a l a d h e s i o nu n d e r n o r m a l c i r c u m s t a n c e s . I n t e r e s t i n g l y ,d u r i n g n e o v a s c u l a r i z a t i o n a s s o c i a t e d w i t ht u m o r g r o w t h , I C A M - 3 e x p r e s s i o n is i n d u c e do n t h e v a s c u l a r e n d o t h e l i u m w h e r e i ts r o l e isy e t to b e d e f i n e d (32 ,33) .

M e sse n g e r R NA b l o t a n a ly s i s d e m o n s t r a t e dt h e p r e se n c e o f t r a n sc ri p t s f o r I C AM - 3 i n le u -k o c y t e s i s o l at e d f ro m n u m e r o u s p r im a t e s s p e -c i e s , dogs , an d rabb i t s , bu t no t i n roden t ce l l s( P K i lg a n n o n , u n p u b l i s h e d d a ta ). C o n s i s t e n tw i t h t h e se r e su l t s, a t t e m p t s to c l o n e t h e r o d e n tI C A M - 3 h o m o l o g h a v e b ee n u n s u c c e s s fu l an di t is l ike ly tha t roden t s do no t hav e the ge ne t i ch o m o l o g o f I C A M - 3 . A l t h o u g h th i s s it u a ti o ni s unusua l , i t i s no t wi thou t p receden t . In t e r -l e u k i n 8 ( I L 8 ) i s a n o t h e r c a s e w h e r e i n a n

i m p o r t a n t i m m u n e c e ll m e d i a to r f o u n d inh u m a n s i s a b se n t fr o m t h e r o d e n t g e n o m e (34) .S i n c e t h e m a j o r i n t e g r i n r e c e p t o r s f o rI C A M - 3 a re c o n s e r v e d i n r o d e n t s p e ci e s, w eh y p o t h e s i z e t h a t a n o t h e r p r o t e i n p e r f o r m st h e f u n c t i o n s o f IC A M - 3 , t h u s c o m p e n s a t i n gf o r t h e a b se n c e o f t h e g e n e .IC A M - 4 (IC A M - L W , L W B lo o d G r o u pGlycoprote in)

T h e f o u r t h m e m b e r o f t h e I C A M f a m i ly , a sd e t e r m i n e d b y t h e c h r o n o l o g y o f i ts d i s c o v e r ya s su c h , is a b l o o d g r o u p a n t i g e n k n o w n a s t h eL a n d s t e i n e r W e i n e r ( L W ) g l y c o p r o t e i n (35).I t wa s d i s c o v e r e d a t t h e s a m e t i m e a s , a n d f o rm a n y y e a rs c o n f u s e d w i th , t h e R h b l o o d g r o u psy s t e m , s i n c e b o t h R h a n d L W d e f i c i e n c i e sa r e p h e n o t y p i c a l l y l i n k e d (36) . Ge n e t i c a l l y ,h o w e v e r , t h e se l o c i a re d i s t i n c t (37,38) . T h i s4 2 - k D a g l y c o p r o t e i n h a s b e e n f o u n d o n l y o ne r y t h r o c y t e s w h e r e i t i s l i n k e d t o t h es u b m e m b r a n o u s c y t o s k e l e t o n . R e c e n t l y , t h ed e d u c e d p o l y p e p t i d e s e q u e n c e s h o w e d t h a tI C A M - 4 c o n t a i n s t w o e x t r ac e l lu l a r i m m u n o -g l o b u l i n - l i k e d o m a i n s w i t h s i g n i f i c a n t h o m o l -o g y t o t h e f i r s t t w o d o m a i n s o f t h e o t h e rm e m b e r s o f t h e fa m i l y (see Table 1 ) . Cons i s -t e n t w i t h t h i s h o m o l o g y , I C A M - 4 h a s b e e nsh o w n t o b i n d t o L FA - 1 a n d t o M a c - 1 i n s t at ica d h e s i o n a s sa y s (39) . T h i s i n t e r a c t i o n m i g h tb e i n v o l v e d in e r y t h r o c y t e t u r n o v e r w i t h i n t h esp l e e n , g i v e n t h e s e l e c ti v e e x p r e s s i o n o f ~ d ~ 2o n m a c r o p h a g e s i n sp l e n i c r e d p u l p c o rd s a n dI C AM - 4 o n e r y t h ro c y t e s (24).ICAM -$ (Te lencepha lin )

R e l a t e d t o th e o t h e r I C AM p r o t e i n s p r e v i -o u s l y d e s c r i b e d b y p o l y p e p t i d e s e q u e n c ec o n s e r v a t io n , I C A M - 5 is th e m o s t r e c e n t l yi d e n ti f ie d a n d p e r h a p s t h e m o s t u n i q u e m e m -b e r o f t h e I C A M f a m i ly . T h e d e d u c e d p r o t e ins e q u e n c e h a s n i n e i m m u n o g l o b u l i n - l i k ed o m a i n s a n d is e x p r e s se d i n r a ts a n d h u m a n s

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o n l y o n t e l e n c e p h a l i c n e u r o n s w h e r e i t i sl o c a l iz e d to so m a t i c a n d d e n d r i ti c m e m b r a n e s ,h e n c e i ts i n it i a l n a m e t e l e n c e p h a l i n (40,41) (PK i l g a n n o n , u n p u b l i s h e d da ta ). T h e f i ve a m i n ot e rm i n a l i m m u n o g l o b u l i n d o m a i n s o f I C A M - 5a re h o m o l o g o u s t o th e f iv e d o m a i n s o fI C A M - 1 a n d - 3 w h e r e a s d o m a i n s 6 - 8 a p p e a rt o h a v e a r is e n f r o m e x o n d u p l i c a t io n o f d o m a i n5 . T h e n i n t h d o m a i n i s le a s t l ik e t h o se o f t h eo t h e r I C A M s , r e s e m b l i n g m o r e c l o s e l y t h eC - t y p e i m m u n o g l o b u l i n d o m a i n s o f o t h e ra d h e s i o n m o l e c u l e s e x p r e s s e d i n th e c e n t r a ln e r v o u s s y s t e m s u c h a s c h i c k a x o n i n - 1 a n dn e u r o f a s c in . D o m a i n 1 o f I C A M - 5 p r e s e r v e st h e b i n d i n g m o t i f s h o w n t o b e r e q u i r e d f o ro t h e r I C A M / i n t e g r i n i n te r a c ti o n s a n d r e c e n tr e s u l t s f o u n d t h a t I C A M - 5 b o u n d t o L F A - 1a n d M a c - 1 (41,42) ( P K i l g a n n o n , u n p u b l i s h e dd a t a ) . F r o m t h e s e d a t a , i t i s p r o p o s e d t h a tI C A M - 5 i s i n v o l v e d i n n e u r o n a l - g l i a l c el li n t e r a c t i o n s u n d e r n o r m a l c o n d i t i o n s o r n e u -r o n a l - l e u k o c y t i c i n t e r a c t i o n s i n p a t h o l o g i cs e t t i n g s w h e r e l y m p h o c y t e i n f l u x i n t o t h i s" i m m u n o l o g i c a l l y p r i v i l e d g e d " s i te d o e s n o tn o r m a l l y o c c u r.Sim ilarities and Differences Withinthe ICAM Family

St ruc tu ra l , gene t i c , and func t iona l s imi l a r i -t ie s a m o n g t h e I C A M p r o t e in s s e r v e to g r o u pt h e se p r o t e i n s t o g e t h e r . I t i s li k e l y t h a t o n e o ft h e I C A M g e n e s is t h e p r im o r d i a l g e n e f r o mw h i c h a l l o t h e r s we r e d e r i v e d . T h e g e n e s f o rI C A M - 1, - 3, - 4 , a n d - 5 h a v e a l l b e e n m a p p e dt o h u m a n c h r o m o so m e 1 9 p l 3 .2 - p 1 3 .3 , wh e r e a sI C A M - 2 r e si d es o n h u m a n c h r o m o s o m e1 7 q 2 3 - 2 5 (37 ,43-46) ( P K i l g a n n o n , W MGa l l a ti n , u n p u b l i sh e d d a ta ) . S i n c e f o u r o f th ef iv e k n o w n I C A M g e n e s in h u m a n s a re c lu s -t e re d , t h i s su g g e s t s t h a t l o c a l i z e d g e n e d u p l i -c a t i o n e v e n t s a r e r e sp o n s i b l e f o r g e n e r a t i n gd ive rs ity , a s i t ua tion tha t is qu i t e co m m on fo rre l a ted gen es w i th in a g iven fam i ly , fo r i ns t ance ,c h e m o k i n e s a n d t h e ir r e c e p to r s (47-49).

A s t h e b a s i s f o r I C A M g e n e d u p l i c a t i o n ,t h e i n d iv i d u a l i m m u n o g l o b u l i n - l i k e d o m a i n sa r e e a c h f o u n d w i t h i n d i f f e r e n t e x o n s r e f l e c t -i n g w h a t i s t h e sm a l l e s t f u n c t i o n a l u n i t t h a t h a sb e e n m a i n t a i n e d b y e v o l u t io n a r y c o n s e r v a t i o n(1,50). A s d i s c u s s e d p r e v i o u s l y , t h e I C A Mp r o t e in s d i f fe r i n th e n u m b e r o f i m m u n o g l o -b u l i n - l i k e d o m a i n s t h a t a re l i n k e d t o g e t h e r a n dw h e t h e r t h e s e d o m a i n s c o n t a i n a n i n t e g r i nb i n d i n g s it e. T h e i m p o r t a n c e o f m u l t i p l ei m m u n o g l o b u l i n d o m a i n s l in k e d t o g e t h ermeans tha t seve ra l spa t i a l ly d i s t inc t i n t egr in -b i n d i n g s i te s ( I B Ss ) m a y e x i s t w i t h i n a s in g l ep o l y p e p t i d e , a s i s t h e c a se f o r I C AM- 1 . Ass u c h , I C A M - 1 / i n t e g r i n - b i n d i n g i n t e r a c t i o n sm a y b e o c c u r r i n g s i m u l t a n e o u s l y t o b r i n go p p o s i n g c e l ls m o r e t i g h t ly t o g e t h e r , a s m i g h tb e r e q u i r e d f o r f i r m a d h e s i o n u n d e r s h e a rs t re s s o r t h e y c o u l d b e t i m e d t o o c c u r i n a r e g u -l a te d m a n n e r a s d i s c u s s e d p r e v i o u s ly . I n s u c hc a se s , t h e r e su l t i n g o u t s i d e - i n s i g n a l s d e l i v -e r e d b y t h e I C A M m a y d i f f e r. Ye t t h e fa c t th a tt h e b a s i c u n i t i s r e p l i c a t e d m u l t i p l e t i m e swi t h i n a s i n g l e p o l y p e p t i d e , r e g a r d l e s s o f t h ep r e se n c e o r a b se n c e o f a n I B S , i n d i c a t e s t h a ti ts s t ruc tu re i s c r i t ica l t o i t s func t ion . The re -f o re , th e f o r m o f t h e d o m a i n w h i c h m a k e s u pa s t a l k m a y p r o j e c t t h e I B Ss a wa y f r o m t h ep l a s m a m e m b r a n e .

Ov e r a l l p r o t e i n s e q u e n c e i d e n t i ti e s b e t w e e nf a m i l y m e m b e r s i n th e a m i n o t e r m i n a l d o m a i nr a n g e s f r o m 2 1 - 3 8 % (see T a b l e 1 ). H o w e v e r ,a c o n s e r v e d p e n t a p e p t i d e s e q u e n c e k n o w n t obe c r i t ica l fo r IC A M bin d in g to ]32 in t egr ins ,d e n o t e d a s t h e I B S , I ( L ) - D ( E ) - S ( T ) - P ( X ) -L ( X ) , o r i g i n a l l y id e n t i f i e d in I C A M - 1 b yS t a u n t o n a n d c o l l e a g u e s (4,20), i s foun d in a lli m m u n o g l o b u l i n - l ik e d o m a i n s o f I C A M p r o-t e in s t h a t b i n d t o i n t e g r i n s . A l t h o u g h t h e p r e -d i c t e d s e c o n d a r y s t r u c tu r e s c a f f o l d o f t h ed o m a i n s r e m a i n s s i m i l a r , t h e r e l a t i v e l y l o wl e v e l o f s e q u e n c e c o n s e r v a t i o n m a y a f f e c tb i n d i n g a f f in i ty b e t w e e n d i f f er e n t I C A M p r o -t e ins and [~2 i n t e g r i n s . F o r e x a m p l e , L F A - 1

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b i n d s t o I C AM - 1 w i t h g r e a t e r a v i d i t y th a n i td o e s t o 1 C AM - 3 a n d c o n v e r se l y , (~d[~2 b i n d s t oI C A M - 3 p r e f e r e n t i a l l y o v e r I C A M - I ( 2 4 ) .No w t h a t a c r y s t a l s t r u c t u r e f o r i m m u n o g l o -b u l in d o m a i n s o f I C A M - 2 h a s b e e n s o l v e d ,t h e s t r u c t u r e s c a n b e m o r e r e a d i l y c o m p a r e da n d c o m p a r i s o n s w i th o t h e r IC A M s m a y b eins igh t fu l wi th re spec t t o the i r r e l a t ive in t e -g r i n - b in d i n g p r e f e r e n c e s (51) .

Dr a m a t i c d i ff e r e n c e s ex i s t in I C A M e x p r e s -s i o n p a t t e r n s a n d t h e r e g u l a t i o n o f t h e i r g e n e sb y i n f l a m m a t o r y c y t o k i n e s . I C A M - 1 , - 2, - 3,a n d - 4 e x p r e s s i o n i s f o u n d o n c e ll s o f h e m a t o -p o i e t i c li n e a g e s , h o w e v e r , n o n e a r e e x p r e s se do n a l l l i n e a g e s . Fo r i n s ta n c e , I C A M - 4 i s f o u n do n l y o n e r y t h r o c y t e s w h e r e a s n o n e o f th e o t h -e r s a r e. D i sp a r a t e t i s su e e x p r e s s i o n p a t t e rn s o fthese p ro te ins i s a f ea tu re tha t i s the re su l t o fd i f f e r e n t ia l t r a n sc r i p t i o n f a c t o r e x p r e s s i o n o rr e c e p t o r / e f f e c t o r p a t h w a y r e g u l a t i o n . K e y d i f -f e r e n c e s a l so e x i s t i n wh e t h e r t h e l e v e l o fI C A M e x p r e s s i o n o n l e u k o c y t e s ar e r e g u la t e do r n o t . Fo r e x a m p l e , I C AM- 1 i s i n d u c e d t oh i g h l e v e l s o n l e u k o c y t e a c t i v a t i o n b y c y t o -k i n e s s u c h as T N F , w h e r e a s I C A M - 3 is c o n -s t i t u t i v e l y e x p r e s s e d a t h i g h l e v e l s . T h e s ed i f f e r e n c e s r e g u l a t e t h e t i m i n g w i t h w h i c hI C A M / in t e g r in e n g a g e m e n t ca n o c c u r a n dm a n i f e s t t h e m se l v e s i n t h e m a n n e r i n wh i c hi n te g r in e n g a g e m e n t t r ig g e r s o u t s i d e - in s i g n a lsand ce l lu l a r behav io rs tha t a re ICA M spec i f ic .Recent Dev elopm ents in O utside- InSignal ing an d Cytoplasm ic Funct ionsof ICA M Pro te ins

A c r o s s - s p e c ie s c o m p a r i s o n o f th e a m i n oa c id s e q u e n c e s c o m p r i s i n g th e p r e d i c t e d c y t o -p l a s m i c r e g io n s o f th e f iv e k n o w n I C A M f a m -i l y m e m b e r s c a n b e s u m m a r i z e d i n t h ef o l l o wi n g wa y . T h e r e i s g r e a t e r a m i n o a c i dc o n s e r v a t i o n a m o n g s t s e q u e n c e s o f a p a r ti c u -l ar I C A M p r o t e i n f r o m d i f f e r e n t sp e c i e s t h a nt h e r e i s b e t w e e n d i f f e r e n t I C A M p r o t e i n swi t h i n a s in g l e sp e c i es . Fo r e x a m p l e , 7 1 - 9 3 %

o f r e s i d u e s a r e c o n se r v e d i n t h e c y t o p l a sm i cd o m a i n s o f I C A M - I f r o m m o u s e , d og , a n dh u m a n s . F o r I C A M - 3 f r o m d o g , c o w , a n dh u m a n s , 6 8 - 9 0 % o f c y t o p la s m i c d o m a i n r es i-d u e s a r e c o n s e r v e d . H o w e v e r , o n l y 1 3 % o fr e s id u e s a re c o n s e r v e d b e t w e e n I C A M - 1 a n dI C A M - 3 f ro m h u m a n s . I C A M - s p e c i f i cs e q u e n c e c o n s e r v a t i o n o f t h e c y t o p l a s m i cr e g i o n s i m p l i e s t h a t e v o l u t i o n h a s m a i n t a i n e dthem for spec i f i c func t ions invo lv ing spec i f iccy top la sm ic p ro te in -p ro te in in t e rac t ions , w hicha re p robab ly e ssen ti a l fo r ICA M func t ion .

M e c h a n i s ti c a ll y , s t u d ie s o f t h e I C A M p r o -t e i n s a r e n o t a s f a r a l o n g a s t h o se o f o t h e rp l a s m a m e m b r a n e r e c e p to r s s u c h a s t h e e p i-de rma l g row th fac to r o r p l a te l e t -de r ived g rowthf a c t o r r e c e p t o r s , h o w e v e r s o m e t h e m e s a r eb e g i n n i n g t o e m e r g e . G r o w t h f a c to r r e c e p t o rd i m e r i z a t i o n f o l l o w i n g l i g a n d b i n d i n g i s am e c h a n i s m k n o w n f o r r e c e p t o r a c t i v a t i o n .R e c e n t o b s e r v a t i o n s s u g g e s t t h a t I C A M - If o r m s a n o n c o v a l e n t d i m e r a t t h e c e ll su r f a c ea n d t h a t t h is f o r m e x h i b i t s h i g h e r a v i d i t y f o rL F A - 1 b i n d i n g th a n d o e s th e m o n o m e r f o r m( 5 2 , 5 3 ) . U n d e r t h e c o n d i t i o n s t h a t y i e l d e dI C A M - l d i m e r s , I C A M - 2 w a s f o u n d to r em a i nm o n o m e r i c a lt h o u g h l o c a ll y e n r ic h e d p l a s m am e m b r a n e r e g i o n s o f I C A M - 2 ( s e e b e l o w )wo u l d p o t e n t i a l l y e x h i b i t g r e a t e r a v i d i t y f o rL F A - 1 b i n d i n g d e s p i t e i t s a p p a r e n t m o n o -m e r i c f o r m . T h e r e q u i r e m e n t f o r c h e m i c a lc r o s s l i n k e r s d u r i n g t h e e x p e r i m e n t a l i so l a t io na n d c h a r a c t e r iz a t io n o f t h e I C A M - 1 d i m e r su g -ges t s t ha t t h i s is a l ab i l e fo rm of the p ro te in andtha t un ide n t i f i ed cofac to r s o r po s t t r ans l a t iona lm o d i f i c a ti o n s su ch a s p h o sp h o r y l a t i o n m a y b erequ i red to s t ab i l i ze the in t e rac t ion in s i tu .I n t e re s t in g l y , IC A M - 5 h a s b e e n i so la t e d f ro mce l l ex t rac t s a s a t e t r amer i c complex o f h ighm o l e c u l a r w e i g h t an d I C A M - 3 h a s b ee n f o u n dt o f o r m a h i g h m o l e c u l a r we i g h t c o m p l e x t h atp r o b a b l y re p r ese n ts a h o m o d i m e r (54) (J Hay-f li c k , u n p u b l i sh e d d a ta ). I f m u l t i m e r i c I C A Mm o l e c u l e s a r e i m p o r t a n t f o r f u n c t i o n , t h e n

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he te romer ic complexes m ay form on those ce l lstha t express more than a s ing le ICA M polypep-t ide . This type of comb ina tor ia l d ive rs i ty mayregula te in tegr in / ICAM binding in te rac t ionsand the resul t ing intrace l lular events .

Jus t a s the d imer ic form o f ICAM - 1 inc reasesb ind ing a f f in i ty for LFA- 1 , th i s com plex m aybe the bas ic func t iona l un i t for t r ansmi t t ingICA M -tr iggered s igna ls in to ce ll s . Ear ly s ig-n a li n g w o r k u s i n g c o - i m m o b i l iz e d I C A M - IMA b a nd a subop t ima l a moun t o f a n t i - C D 3MAb to l iga te sur face pro te ins demonst ra tedtha t I C A M- l i s a c o - s t imu la to r y mo le c u le .M ore recent ly , l iga t ion of ICA M - 1 was obse rvedto t r i gge r a n a c c um ula t ion o f t y r os ine phos -phopr o te in s , i nc lud ing c dc 2 in b lood - de r ive dle ukoc y te s a nd the huma n T l e uk e mic c e l l li ne ,Mol t - 3 (55), a nd c o r t a c t in i n immor ta l i z e dr a t b r a in e ndo the l i a l c e l ls (56). A d d i t i o n a l l y ,l i ga tion o f I C A M - 1 by a n t ibody t r e a tme n t o fSV-40 la rge T an t igen (Ag) t r ansform ed syn-ovia l ce l l s indu ced ac t iva t ion of the A P- 1 t r an-s c r i p t i o n f a c t o r a s m e a s u r e d b y n u c l e a rt rans loca t ion and DN A binding (57). Upregula -t i on o f IL -113 m e ssa ge a nd c h lo r a m phe n ic o lace ty l t r ansfe rase (CAT) ac t iv i ty f rom pro-m ote r dr iven cons t ruc ts tha t conta in the IL- 1 [ 3AP-1 b inding s i te was a l so obse rved . Fur the re v ide nc e o f t he c a pa c i ty f o r s igna l ing m e d i -a t e d by I C A M - 1 l i ga tion on b loo d l e ukoc y te sw a s f ound us ing M A b t r e a tme n t o f pe r iphe ra lb lood m ono nuc le a r c el l s (P B M C ) in t he p re s -e n c e o f a s u b o p t i m a l d o s e o f N - f o r m y l -meth ionyl - leucyl -phenyla lan ine FML P) whichtoge the r i nduc e d a n ox ida t ive bu r s t r e sponse(58 ) . T h e s e o b s e r v a t i o n s t a k e n t o g e t h e r ,sugge s t a mo de l fo r IC A M - I f unc t ion in thed e v e l o p m e n t o f c e l l u l a r i m m u n e r e s p o n s e sw h e r e b y e a r l y a c t i v a t i o n e v e n t s s u c h a scytok ine b inding to receptors or T ce l l an t igenr e ce p t or b i n d i n g to a n t i g e n / m a j o r h i s t o c o m -p a t i b i l i t y c o m p l e x ( M H C ) c o m p l e x e s t r i g -ge rs ICAM-1 sur face express ion a long wi tho t h e r c o m p o n e n t s o f a s i g n a l t r a n s d u c i n g

c o m p l e x . T h e s e c o m p l e x e s m i g h t i n c l u d ekinase/ph osph atase act ivi t ies s ince no intrinsicenzymat ic ac t iv i t ie s have been demonst ra tedf o r a n y I C A M c y t o p l a s m i c r e g i o n . S u c ha s s e m b l a g e s m a y f o r m s t a b i l i z e d I C A M - 1d ime r s a nd c ou ld b ind to a c t ivat e d LF A -1 onoppo s ing ce l l s wi th h igher av id i ty . This t igh tc e l l - c e l l c on ta c t c ou ld p r e se n t o the r r e c e p -tor / l igand pa i r s wi th an op por tuni ty for in te r -a c t ion , pe r ha ps l i ga t ing I C A M- 1 in to e ve nlar ge r m ol ec ul ar assemblages, eventual ly t rig-ge r ing ICA M -1-spec i f ic ac t iva t ion events .

The o the r me m be r o f t he IC A M f a mi ly mos tf requently associa ted with s ignal t ransductionand ac t iva t ion events i s ICAM -3. Th e fac t tha tICA M -3 was con s t i tu t ive ly expressed a t h ighleve ls on a ll ce l l s o f the hem atopoie t ic l ineagesinvo lve d in c e l lu la r imm une r e sponse s im p l i e dthat i t has a role dur ing the ini t ia l phase ofa c t iva t ion by ma in t a in ing c e l l - c e l l c on ta c t .R e c e n t o b s e r v a t i o n s w i t h p e r i p h e r a l b l o o dT ce l ls and T leukem ic ce ll l ines s t imula ted byc o - imm obi l i z e d I C A M - 3 M A b a nd subop tima la moun t s o f C D 3 M A b p r ov ide d support f o r a near ly ac t iva t ion ro le of ICAM-3 (26,59,60).Som e have obse rved tha t on pe r iphera l b loodleukocy tes and T ce l l lines , ICAM -3 an t ibodyt rea tment t r iggered tyros ine phosphory la t ion o fspecif ic prote ins and f luxed intracel lular ca l-c ium concent ra t ions (55,61,62). Others how-ever , ourse lves inc luded , have fa i led to seethese responses. Th e reasons for these dispar i-t ie s m ay be caused by an t ibody ep i tope spec i-f ic i ty or to the inhe rent dif ferences in ce l l l inesor ac t iva t ion s ta tes r e su l t ing f rom iso la t ionp r oc e dure s . I C A M - 3- de pe nde n t a c t iva t ion o fother surface prote ins has a lso been rep or ted. Inparticular, upregulation of [31 and [32 integrinfunc t ion on PBMC blas ts has been obse rved .H o w e v e r t he b ioc he mic a l s igna ling pa thw a ysl e a d ing to t he se e f f e c t s a r e a s ye t unc l e a r(63,64). ~ in tegr in upregula t ion by a l igandfor ]32 integ rins su gge sts that an intere sting cir-cular ac t ivat ion pathw ay co uld exist.

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T o u n d e r t a k e a d e sc r i p t io n o f h o w i n tr a c e l-l u l a r e v e n t s a r e t r ig g e r e d b y I C A M - 3 , a f u n c -t io n a l m a p o f t h e c y t o p l a s m i c r e g io n o f t h ep r o t e in w a s d e v e l o p e d u s in g a n IC A M - 3 -d e f i c i e n t Ju r k a t c e l l l i n e a n d g e n e t r a n s f e rt e c h n iq u e s . H u m a n I C A M - 3 c y t o p l a s m i c ta ilt r u n c a t i o n s a n d s i n g l e a m i n o a c i d m u t a n t sw e r e t e s t e d w i t h r e s p e c t t o C D 3 / I C A M - 3 -d e p e n d e n t c y t o k i n e s ec r e ti o n , h o m o t y p i ca g g r e g a t i o n , a n d c e l l u l a r sp r e a d i n g ( 65 ) . T h em a j o r i t y o f a m i n o a c i d s i n t h e c y t o p l a s m i cr e g i o n we r e r e q u i r e d f o r C D 3 c o - s t i m u l a t i o n .D e l e t io n o f t h e m e m b r a n e d i s ta l o r p r o x i m a lt h i rd s o f t h e c y t o p l a sm i c r e g i o n s i g n i f i c a n t lya f f e c t e d a g g r e g a t i o n a n d sp r e a d i n g r e sp o n se s .Po i n t m u t a t i o n o f s e r in e 4 8 9 o r s e r i n e 5 1 5 t oa l an i n e ( l o c a te d i n t h e m e m b r a n e p r o x i m a l o rd i s t a l r e g i o n s , r e s p e c t i v e l y ) a l s o b l o c k e dI C A M - 3 f u n c t io n . P h o s p h o r y l a t io n o f s e ri n e4 8 9 i n J u r k a t c e l l s w a s e n h a n c e d b y C D 3c r o s s l i n k i n g o r b y d i r e c t ly s t i m u l a t i n g p r o t e i nk i n a se C ( PKC ) i so f o r m s w i t h p h o r b o l e s t e r .F u r t h e r m o r e , s e r i n e 4 8 9 w a s p h o s p h o r y l a t e ds p e c i fi c a ll y b y p u r i f ie d i s o f o r m s o f P K C s u g -g e s t i n g t h a t th i s e n z y m e m a y b e a s i g n i f i c a n tc o n t r i b u t o r to I C A M - 3 - m e d i a t e d s i g n a l tr a ns -d u c t i o n e v e n t s .

E f f i c i e n t a c t i v a t i o n o f T c e l l s i s t h o u g h t t or e q u i r e s i g n a l i n g a t o r g r e a te r t h a n a t h r e sh o l dl ev e l f r o m b o t h a n t i g e n r e c e p t o r c o m p l e x e sa n d n u m e r o u s o t h e r s u r f ac e p r o t e in s f o r w h i c hl i g a n d s e x i s t o n t h e AP C su r f a c e w i t h i n a sp e -c i a l iz e d c o n t a c t z o n e b e t w e e n t h e se c e l ls . A tt h is a d h e s i v e i n t e r fa c e , I C AM - 3 p r o b a b l y p e r -f o r m s i m p o r t a n t f u n c t i o n s in s u p p o r t o f a n d t oa m p l i f y s i g n a l i n g e v e n t s . Fo r n a i v e T c e l l s( C D4 5 R A + ) , t h e s i g n a l i n g t h r e sh o l d r e q u i r e df o r a c t i v a ti o n h a s b e e n o b se r v e d t o b e g r e a t e rt h a n i t i s f o r m e m o r y T c e l l s ( C D 4 5 R O +) ( 66 ) .T h i s m a y b e t h e r e su l t o f in c r e a se d l e v e ls i ns u r f a c e e x p r e s s i o n o f a c t i v a t i o n - p r o m o t i n gp r o t e i n s o n m e m o r y v s n a i v e T c e l l s o rd e c r e a s e d l e v e ls o f m o r e n e g a t i v e ly c h a r g e dp r o t e i n s , su c h a s t h e l a r g e r i so f o r m s o f C D4 5 ,

p r o v i d i n g l e s s r e p u l s i o n b e t we e n c e l l s . On em e a n s f o r n a i v e T c e ll s to a c h i e v e p r o d u c t i v ea c t i v a t io n i s f o r t h e m t o f o r m g r e a t e r c e l l - c e l lc o n t a c t z o n e s w i t h a n APC a l l o wi n g f o r su r -v e i l la n c e o f c o g n a t e a n t i g e n i c p e p t i d e s o r t oi n c r e a se t h e d u r a t i o n o f t h e c o n t a c t . I n t e r e s t -i n g l y , I C A M - 3 , b u t n o t IC A M - 1 , t r i g g e r e d t h ei n d u c t i o n o f T c e l l sp r e a d in g . T h i s l e n d s su p -p o r t t o t h e n o t i o n t h a t o n e o f t h e f u n c t i o n sI C A M - 3 i s t o i n c r e a s e t h e c o n t a c t a r e ab e t we e n c e l l s , f o r e x a m p l e , d u r i n g t h e e a r l ye v e n t s a t t e n d a n t T c e l l / A P C e n g a g e m e n t .T h u s , h i g h c o n s t i t u t i v e e x p r e s s i o n l e v e l s o f Tc el l I C A M - 3 c o m p l e x e d w i th L F A - 1 m a ya l l o w p r o d u c t i v e a c t i v a t i o n t o o c c u r b yi n c r e a s i n g t h e p r o b a b i l i t y o f T C R / A g i n t e r ac -t i o n s o r m a i n t a i n i n g t h a t i n t e r a c t io n i n c a se o fn a i v e T c e l l c o n t a c t o r l o w a n t i g e n a f f i n it y .

I n l i g h t o f t h e p r e v i o u s d i s c u s s i o n s , a m o d e lo f t h e r e l a ti o n sh i p b e t we e n I C AM - 3 f u n c t i o nand ac t iva t ion can be p rop osed . At in i ti a l con-t ac t be tw een an AP C and a T ce ll , su r face adhe -s i v e p r o t ei n s su c h a s I C AM - 3 / L FA - 1 i n te r ac t.Un d e r c o n d i t i o n s o f h i g h - a f fi n i ty i n t e g r in b i n d -i n g , wh i c h wo u l d b e t h e c a se i f t h e APC i sa c t i v a t e d , t h e se c e l l - c e l l c o n t a c t s wo u l d b es t ab li li zed . T ce l l r ecep to r (TCR ) eng age m en twi t h c o g n a t e a n t i g e n / MH C c o m p l e x e s l e ad s t ok i n a se a c t iv a t io n a n d p h o sp h o r y l a t i o n o f f u n c -t i o n a l l y e s s e n t i a l c y t o p l a s m i c r e s i d u e s o fI C AM - 3 ( i .e ., s e r in e 4 89 ) . D i m e r s o f I C A M - 3form or a re s t ab i l i zed , poss ib ly a s a r e su l t o fp h o sp h o r y l a t i o n a n d r e c r u i t m e n t o f c y t o so l icfac to r s , add ing s t ab il it y to the ce l lu l a r con tac tzone an d the p rogram for T ce l l ac t iva t ion andc lona l p ro l i f e ra t ion ensues . Seve ra l ob jec t ivesa re a c h i e v e d b y e n g a g e m e n t o f I C AM - 3 i n t h isc o n t e x t. T h e n u m b e r o f a n t i g e n - r e c e p t o r i n te r -ac t ions a re inc reased in the en la rged con tac tzone . Th e con tac t is s t ab i l i zed a l low ing the s ig -na l ing th re sho ld to be m e t in the case o f a na iveT ce l l . Al so , i t b r ings the con tac t a rea be twe ent h e t wo c e ll s i n t o t ig h t j u x t a p o s i t i o n a l l o wi n go t h e r r e c e p t o r p a i rs ( i.e ., C D 4 0 - C D 4 0 L ) w i t h

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an opportunity to interact a lso contr ibuting toa t ta inment of the ac t iva t ion threshold . Withc lona l pro l i f e ra t ion comes the need for d is -soc ia tion of the T ce l l f rom the APC, and th isc ou ld be a c he ive d by de - phosphor y la t ion o fI C A M - 3 , w h ic h w ou ld l e a d to lo s s o f su r fa c ed i m e r s , l o w e r L F A - 1 b i n d i n g a v i d i t y , a n ddestabil ized adhesive interact ions.

R e - o r ga n iz a t ion o f t he a c t in c y to ske le tonin T c e l l s is ne c e s sa r y f o r t he de ve lo pm e n t o fa n t ige n - induc e d a c t iva t ion , w h ic h inc lude sr e c r u i tme n t o f im por t a n t c y to ske le ton - a s soc i-a ted pro te ins to the cor t ica l ac t in ne tw ork andm o r p h o l o g i c a l c h a n g e s a c c o m p a n y i n g c e l l -ce l l contac t (67) . M e c ha n i s t i ca l ly , the po ly -me r i z a t ion o f a c tin ha s be e n hy po the s i z e d a sthe d r iv ing f o r c e f o r p r o tr u s ion o f t he p l a sm ame mb r a ne l e a d ing e dge du r ing c e l l sp re a d ingas occurs w hen a T ce l l engul f s an AP C or itmig r a t e s du r ing imm une su r ve i l l a nc e w i th inT c e l l z one s o f s e c onda r y lympho id o r ga ns(68) . S i n c e I C A M - 3 l i g a t i o n t r i g g e r s c e l lspreading , change s in ce l lu la r f -ac tin conten tw o u l d b e e x p e c t e d t o o c c u r . J u r k a t c e l l st re a te d w i t h IC A M - 3 M A b r e s p o n d ed w i t h as low and p ro long ed inc rease in ce l lu la r f -ac t inand a dec rease in g-ac t in . Also , ICA M -3- t r ig-ge red sp read ce l l s exhib i ted f luorescen t s ta in-ing for f -ac tin a t the edges an d in a punc ta ted i s tr ibu tion , w h ic h c o loc a l i z e d w i th I C A M - 3(J Hayf l ick , unpubl ished da ta ) . In cont ras t ,an t i -CD3 MAb e l ic i ted a rap id , bu t t r ans ien tinc rease in to ta l ce l lu la r f -ac t in wh ich w as notc o l o c a l i z e d i n s p r e a d c e l l s ( 6 9 - 7 1 ) (JHay f l ick , unpubl ish ed da ta ). In te res t ing ly , inhuma n lymphoc y te s , P K C a c t iv i ty t r i gge r e db y T C R e n g a g e m e n t h a s b e e n i m p l i c at e d inr e gu la t ing c e l l sha pe a nd mo t i l i t y by mod-u la t ing the c e l lu l a r c on te n t o f f - a c t in a nda c t in - b ind ing p r o te in f unc t ion ( 72 - 74 ) . A na d d i t io n a l c o n s e q u e n c e o f r e - o rg a n i z a t io n o fthe c y tosk e le ton i s it s e f f e c t on ge ne e xp r e s -s ion by a c t iva t ion o f t r a nsc r ip t ion f a c to r s o ft h e N F - k B f a m i l y (75) . I C A M - 3 - t r i g g e r e d

c y toske le t a l r e o r ga n iz a t ion a nd g e ne a c t iva-t ion is no t l imi t e d to T c e l l s , s inc e im m ob i -l i z e d M A b e n g a g e m e n t o f I C A M - 3 o n P B M Cand the mon ocy t ic ce l l l ine , THP -1 t r iggeredc e l l u l a r s p r e a d i n g a n d a d o s e - d e p e n d e n ti n cr ea s e i n c h e m o k i n e g e n e e x p r e s s i o n a n ds e c r e t i o n (75a) .The consequences of ICAM-1 and -3 t r ig -ge r ing on c e l lu l a r be ha v io r a r e mos t l i ke lymedia ted by d i rec t in te rac t ions wi th p lasmamembrane-assoc ia tedcytoskele ta l comp onents .ICA M - 1 has been o bse rved in assoc ia tion wi thmicrof i lament-r ich, detergent-extracted f ibro-blasts (76) . Chan ges in ICA M -3 sur face loca l -i z a ti o n b y a n t i b o d y - i n d u c e d c a p p i n g a n dcoinc iden t inc rease in res is tance to de te rgente x t r a c t i o n h a v e b e e n f o u n d ( S R o s e n m a n ,M Gal la tin , unpub l ished data ). Ant i - ICAM -3M A b t r e a t m e n t l e d t o c a p f o r m a t i o n a n di n c r e a s e d d e t e r g e n t r e s i s ta n c e i n a n e p i t o p e -de p e nd e n t ma n ne r ind ic a ting tha t l inka ge o fICAM -3 to the cy toske le ton is a r egula ted eventp e r h a p s m e d i a t e d b y i n t e g r i n b i n d i n g o rI C A M - 3 d i m e r i z a ti o n . D i r e c t cy t o s o l i c p r o-t e in - p r o te in in t e r a c t ions a r e p r e sum a b ly theu n d e r l y i n g m e c h a n i s m f o r t h e s e f i n d i n g s ,a nd se ve r a l suc h b ind ing in t e r a c t ions w i tht h e I C A M s h a v e a l r e a d y b e e n r e p o r t e d .A m o n g t h e c y t o s k e l e t o n - a s s o c i a te d p r o te i n st h a t h a v e b e e n o b s e r v e d b y i n v i t ro b i n d i n gt o I C A M - 1 c y t o p l a s m i c d o m a i n a r e a c t i n ,(x-ac t in in , ~- tubul in , and g lyce ra ldehyde-3-p h o s p h a t e d e h y d r o g e n a s e , a p r o t e i n w i t hm i c r o t u b u l e - b u n d l i n g a c t i v i t y ( 7 6 - 7 8 ) .T h e s e i n t e r a c ti o n s c o u l d l i n k IC A M - 1 w i t hthe c y toske le ton a nd pe r ha ps pa r t i c ipa t e i nregula t ing cy toske le ta l o rganiza tion . ICA M -2h a s a l s o b e e n o b s e r v e d t o b i n d d i r e c t l y t o~- a c t in in in v i t r o (79) . In fac t , the b indingsi tes on both ICAM-1 and -2 for oc-act ininshare fea tures in co m m on w i th o the r o~-ac tininb ind ing p r o te in s tha t spa n the p l a sma me m-b r a n e , n a m e l y a p r e p o n d e r a n c e o f b a si c a n dh y d r o p h o b i c a m i n o a c i d s .



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Ch a nge s i n c e l l - su r f a c e p r o t e i n l oc a l iz a t i ona r e c o m m o n l y o b s e r v e d d u r i n g o r r e s u l t i n gf r o m c e l l u l a r a c t i v a t i o n o r d i f f e r e n t i a t i o ns t at e , w h i c h a r e l i ke l y to be m e d i a t e d by d i r e c ti n t er a c t io n s b e t w e e n t h e t ra n s m e m b r a n e p r o -t e i n s t h e m s e l v e s a n d t h e p e r i p h e r a l a c t i nc y t o s k e l e t o n . A s n o t e d p r e v i o u s l y , e v i d e n c ef o r d i r e c t c y t o p l a s m i c d o m a i n i n t e r a c t i o n sw i t h i n t r a c e l lu l a r p r o t e i n s ha s be e n f ou nd f o rt h e c y t o p l a s m i c d o m a i n s o f I C A M - 1 a n d - 2.I n a dd i t i on , h i gh - l e ve l e xp r e s s i on o f c e r t a i nc y t o s k e l e t o n - a s s o c i a t e d p r o t e i n s h a s b e e nf o u n d t o p r o f o u n d l y a f f e c t s u r f a c e I C A Ml o c a l i z a t i o n . F o r e x a m p l e , w h e n e z r i n w a sove r e xp r e s se d i n a na t u r a l k i ll e r ( N K ) t a r ge tce l l va r i an t tha t was res i s t an t to ce l l k i l l ing , i tl o c a li z e d p r e d o m i n a n t l y t o u r o p o d i a l p r o t r u -s i o n s ( 8 0 ) . C o i n c i d e n t r e - d i s t r i b u t i o n o fI CA M - 2 i n t o t he s a m e s t r uc tu r e s w a s a lso se e na nd t h i s c on t r i bu t e d t o t he c e ll s a c qu i r i ng se n -s i t iv i ty to k i l l ing by NK ce l l s . Thi s cy to tox ic -i ty w as a l so inh ib i t e d by ~32 in teg r in M A bt r e a t m e n t . T h e s e d a t a s u g g e s t t h a t N Kc y t o l y t i c a c t i v i t y i s de pe nde n t on r e - l oc a l -i z a ti o n o f I C A M - 2 a n d P 2 i n t e g r i n f unc t i on .E z r in , a m e m b e r o f th e e z r i n - ra d i x i n - m o e s i n( E R M ) f a m i l y o f c y t o s k e l e t a l - m e m b r a n ep r o t e i n l in k e r s , i s t h o u g h t t o c o n t a i n c r y p t i cf - a c t i n b i nd i ng s i t e s t ha t a r e e xpose d du r i ngm i c r o v i l l u s o r u r o p o d f o r m a t i o n ( 81 ) . E z r i nb i nd i ng t o m i c r o f i l a m e n t s m i gh t t he n l a t e ra l lyr e d i s t r i bu t e m i c r o f i l a m e n t bound c ~ - a c t i n i n /I C A M - 2 c o m p l e x e s . C o r t i c a l f l o w o f a c t i nf i l a m e n t s a n d t he i r a s soc i a t e d p r o t e i n s ( e . g .,ez r in an d c~-act in in) has bee n p os tu la ted as theb a s i s f o r g e n e r a t i n g h i g h l y e n r i c h e d m e m -b r a n e s u b d o m a i n s li k e th o s e f o u n d i n u r o p o ds t ruc tures ( 82 ) . S u c h e n r i c h e d d o m a i n s c o u l da l so l e a d t o I C A M m u l t i m e r s th a t w o u l d f o s t e rh i g h a v i d i t y i n t e g r i n - b i n d i n g i n t e r a c t i o n sw h i c h ha s be e n r e po r te d f o r d i m e r s o f I CA M - 1 .

L o c a l i z a t i o n t o u r o p o d s i s a f e a t u r e o fI C A M - 2 t h a t i s s h a r e d b y o t h e r I C A M s .I CA M - 1 ha s a l so be e n f ound t o l oc a l i z e t o

u r opod s t r uc t u r e s i n JY B c e l l s w he n se e de don t o pu r i f i e d L F A - 1 b i l a ye r s ( 83 ) . U r o p o dpr o t r u s i ons on a c t i va t e d pe r i phe r a l b l ood Tl y m p h o b l a s t s a n d J u r k a t T c e l l s h a v e b e e nf o u n d t o c o n t a i n I C A M - 3 ( 6 3 , 8 4 , 8 5 ) . In te r -e s t ing l y , I CA M - 3 r e d i s t ri bu t i on on T c e l ls w a sf o u n d t o o c c u r w h e n t h e s e c e ll s w e r e s e e d e do n t o m o n o l a y e r s o f T N F - a c t i v a t e d e n d o t h e -l ia l ce l l s , a s we l l as on to pur i f i ed ICA M -1 orva sc u l a r c el l a dhe s i on m o l e c u l e - 1 ( V CA M - 1 ,i n th e p r e s e n c e o f t h e c h e m o k i n e s R A N T E Sa nd m o noc y t e c he m ot a c t i c p r o t e in - 1 (M C P - 1 ),sugge s t i ng t ha t m u l t i p l e s i gna l i ng pa t hw a ysa r e i n v o l v e d i n t h e m e m b r a n e d i s t r i b u t i o nc h a n g e s . A s p r o p o s e d f o r I C A M - 2 , a h i g h l yc o n c e n tr a te d m e m b r a n e d o m a i n o f I C A M - 1or - 3, p r oba b l y a s a d i m e r , w ou l d be e xpe c t e dt o e x h i b i t g r e a t e r b i n d i n g a v i d i t y f o r P 2i n t e g ri n s . T h e o c c u r a n c e o f s i m u l t a n e o u sl o c al iz a t io n o f d i f fe r e n t I C A M s t o t h e u r o p o do r w h e t h e r a te m p o r a l s e q u e n c e o f o n e I C A Mf o l l ow e d by a no t he r i n to suc h s t r uc t u r e s w i l lr e qu i r e f u r t he r s t ud i e s .N o v e l A r e a s o f T h e r a p e u tic D e v e l o p m e n tI n v o lv i n g IC A M B i o lo g y

T h e n o t i o n t h a t t h e r a p e u t i c i n t e r v e n t i o nd u r i n g c h r o n i c a n d a c u te i m m u n e re s p o n s e s a tt he l e ve l o f I CA M / i n t e g r i n i n t e r a c t i ons ha sg a i n e d m u c h f a v o r a n d i s b e i n g p u r s u e d v i g o r -o u s l y b y t h e p h a r m a c e u t i c a l i n d u s t r y . T h ef o l l ow i ng s t r at e g i es a r e be i ng e xp l o r e d :

1. Physical blockag e of extracellular bind ingdomain interactions;2. Phys ical block age of intracellular interac-

tions; and3. Interruption of protein expression.

T h e i d e a t h a t p h y s i c a ll y b l o c k in g a n I C A M /i n t e g r i n i n t e r a c t i on by c om pe t i t i ve a n t i bodyb i n d i n g h a s l e d to t h e d e v e l o p m e n t o f n u m e r -ous t he r a pe u t i c c a nd i da t e s t ha t ha ve e n t e r e dc l in ica l t ri a l s fo r a va r i e ty of ind ica t ions . Ana l te r n at iv e a p p r o a c h s t e m m e d f r o m t h e o b s e r-

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va t ions of na tura l ly occur in g so luble forms ofse ve ra l I C A M pr o te in s (86-88). In te res t ing ly ,s e r um l e ve l s o f bo th so lub le I C A M - 1 a nd - 3f luc tua te , appear ing to cor re la te wi th d is t inc tpa tho log ie s suc h a s ne u t rope n ic pne um on ia ,m ul t ip le sc le ros is , and as thma , leav ing ava i l -ab le the poss ib i l ity tha t the i r de tec t ion ma y beo f d i a gnos t i c va lue (89-91). P r e c l in i c a l ly ,a dmin i s t r a t ion o f so lub le I C A M- 1 ha s be e nf ound to be e f f e c t ive in p r e ve n t ion o f r h ino -v i ra l i n f e c t ion in c h impa n z e e s (92). The r o l eof so luble ICAM prote ins in v ivo i s unc lea r ,bu t t he y m a y se r ve to dow nr e gu la t e i n te g r inbinding and the re fo re ce l l - ce l l contac ts a f te ra n imm une r e sponse by spe c i f i c a lly she dd ingthe in t e g r in -b ind ing dom a in o r t o r e l e a se a na n t a g o n i s t i c f o r m t h a t c o u l d c o m p e t e f o ri n t e g r i n b i n d i n g . L o w e r i n t e g r i n - b i n d i n ga f fin ity o f mo nom e r ic I C A M - 1 c om pa r e d tothe d im er form sugges ts tha t the so luble pro-t e in w ou ld ha ve to be in a d im e r to e f f e c t ive lyc omp e te . The se obse r va tions ha ve g ive n r i seto the ide a tha t a n t ibody F c / I C A M f us ion p r o -te ins which possess two respec t ive in tegr in-b i n d i n g s i t e s m i g h t p r o v e t o b e a m o r ee f fec t ive so luble an tagonis t . An ex tens ion o fth is i s the proposa l tha t pept ides or s imi la rspace f i l l ing , low molecula r we ight en t i t ie s ,m i g h t c o m p e t i v e l y i n h i b i t b i n d i n g t h e r e b yproving to be e f f icac ious .

F rom obse rva t ions rev iew ed in the sec t ione n t it l ed R e c e n t D e v e lopme n t s i n O u t s ide - I nS igna l ing and Cytoplasmic Func t ions of ICAMProte ins regard ing the ro les of the ICA M s inintracel lular events such as d irect prote in inter-a c t ions o r ge ne r a t ion o f s e c ond me sse nge reffector pathw ays, i t is plausible tha t inhibi t inge i the r of these may prove to be b enef ic ia l . S imi-lar to the ex tracel lular binding interact ions, tar-geted drug screens against ICAM -1/o~-actinininteract ion, for example , could serve the pur-pose of deve loping drug leads tha t in te r ruptcytoskele ta l associa t ion. Interruption of suchassoc ia t ions in end othe l ia l ce l l s , fo r ins tance ,

m a y b e e f f e c t i v e i n r e d u c i n g l y m p h o c y t einf i l t ra t ion in disease se t tings such as rheu m a-toid ar thr i t is or Crohns disease . An intr iguinga l te rna t ive approach w here the d rug ta rge ts a rek inase /phospha tase e f fec tor molecu les tha t a ret r igge r e d on c e l lu l a r a c t iva t ion l e a d ing topos t tr a nsl a tiona l m od i f i c a t ion o f I C A M pr o -te ins o r that are act ivated as a result of ICA Ml iga t ion . C a ta ly t i c s i t e i nh ib i to r s o f t he seac t iv i tie s ma y b e re la t ive ly nonspec i f ic how -ever , unless their targets are suff ic iently l im-i ted in their dis tr ibution. Accessabil i ty to theinternal mil ieu is a fur ther hurdle to be over-com e, s ince these ta rge ts r e s ide w i th in the ce l l,the re fore sm al l mo lecule inh ib itor s wi th favor -able l ipid transpor t quali t ies m ust be soug ht .Final ly, a s tra tegy based on reducing synthe-sis of specif ic ICA M prote ins or important cyto-p lasmic components , such as second messengereffector proteins, utilizing main ly antisense tech-nology have been repor ted (93-96). Impor tan tfactors impinging on the effect ivness of thisapproach includ e the turnover ra te of the targetprotein itself , lysos om al degrad ation ofantisensenucle ic ac ids, and target ing to the messengerRN A. Th is s tra tegy has a lso yie lded drug candi-dates that are presen tly in clinic al tr ials.

I n t h e f u t u r e , t h e m e c h a n i s m s o f h o wI C A M - t r igge r e d in t r a c e llu l a r eve n t s a r e r e gu -l a te d by in t e g rin e nga ge m e n t shou ld be c o m eclar if ied. Fur therm ore , the re la tionship be tw eenI C A M/c y toske le ton in t e r a c t ions a nd I C A M/in tegr in av id i ty change s may lead to the e luc i -da t ion o f nove l the rapeu t ic ta rge ts .Acknowledgments

T h e a u t h o r s t h a n k t h e i r c o l l e a g u e s f o ra l low ing the m to inc lude unpub l i she d da t a a nda p o l o g i z e t o t h o s e w h o s e w o r k w e d i d n o tinc lude be c a use o f spa c e l imi t a tions . W e a l sot h a n k L o u i s e B a n d a n d A l i c e D e r s h a m f o rc r i t i c a l r e a d i n g a n d h e l p i n p r e p a r i n g t h emanuscr ip t , r e spec t ive ly .

T h e I C A M P r o t e in F a m i l y 3 2 3


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79 Heiska L, Kantor C, Parr T, Crit-chley DR, Vil ja P, Gahmberg CG ,Carpen O: Binding of the cyto-plasmic domain of in te rce l lu la radhesion molecule-2 (ICAM -2) toalpha-actinin. J Biol Chem 1996;271 (42):26,214-26,219.80 Helander TS, Carpen O, TurunenO, Kovanen PE, Vaheri A, Tim-onen T: ICAM-2 redistributed byezrin as a target for killer cells.Nature (Lond) 1996;382(6588):265-268.81 Jockusch BM, Rudiger M: Cross-talk between cell adhesion mol-ecules: viniculin as a paradigm forreguation by conformation. CellBiology 1996;6:311-315.82 Bray D, W hite JG: Cortical flowin animal cel ls . Science (WashDC) 1988;239(4842):883-888.83 Dustin ML, Carpen O, SpringerTA: R egulation o f locomotion andcell-cell contact area by the LFA - Iand IC AM -I adhesion receptors. JImm unol 1992;148(9):2654-2663.84 Campanero MR , de l Pozo MA,Arroy o AG , Sanchez-Mateos P,Hernandez-Caselles T, Craig A,P u l i do R , S a nc he z - Ma dr i d F :ICAM-3 interacts with LFA - 1 andregulates the LFA - I /ICAM - 1 celladhes ion pa thway. J Ce l l Biol1993;123(4):1007-1016.85 del Pozo MA, Sanchez-Mateos P,N i e t o M, S a nc he z - Ma dr i d F :Che moki ne s r e gu l a t e c e l l u l a rpolarization and adhesion receptorredistribution during lymphocyteinteraction with endothelium andextracellular matrix. Involvem entof cAMP signaling pathway. J CellBiol 1995;131(2):495-508.86 del Pozo MA, Pulido R, Munoz C,A I va r e z V , H umbr i a A , Ca m-panero MR, Sanchez-Madrid F:

Regula t ion of ICAM-3 (CD50)membrane expression on humanneutrophils through a proteolyticshedding mechanism. Eur J Im-munol 1994;24(11):2586-2594.87 Pino-Otin MR , Vinas O, de la Fu-ente MA, Juan M, Font J, Torra-def lot M, Pallares L, Lozano F,Alb erola-I la , Martorel l J , YagueJ, Vives J, Gaya A: Existence of aso l ub l e f o r m o f CD 50 ( i n t e r -c e l l u l a r a dhe s i on mol e c u l e - 3 )p r oduc e d upon huma n l ymph-oc y t e a c t i va t i on . J I mmunol1995; 154:3015-30 24.88 Martin S, Rieckman n P, M elchersI, W agner R, Bertrams J, VoskuylAE, Roep BO , Zielasek J, Hei-denthal E, Weichselbraun I, et a:C i r c u l a t i ng f o r ms o f I CA M- 3( c I CA M- 3) . E l e va t e d l e ve l s i nautoimm une diseases and lack ofassociation with cICAM-1. J Im-munol 1995;154(4): 1951-1955.89 Sudhoff T, Wehm eier A, ArningM, Bauser U, S chlomer P, Aul C,Schneider W: Increases o f sICAM -I during neutropenic pneum onia inleukemic patients. Leukem ia 1997;11(3):346-351.90 Rieckm ann P, Altenhofen B, Rie-gel A, Baudewig J, Felgenhauer K:S o l ub l e a dhe s i on mol e c u l e s( sVCAM -1 and s ICAM -1) incerebrospina l f lu id and se rumcor re la te wi th MRI ac t iv i ty inmul t ip le sc le ros is . Ann Neurol1997;41 (3):326-333.91 Gonokami Y, Konno S, KurokawaM, Kawazu K, Ueno K, Tomita K,I ke M, N yu i M, A da c h i M:Circulating intracellular adh esionmol e c u l e - I c onc e n t r a t i ons f o l -lowing bronchial provocation inatopic asthma, lnt Arch AllergyImmunol 1997;112(4):386-391.

92 Huguenel ED, Coh n D, DockumDP, Greve JM, Fournel MA, Ham -mond L , I r w i n R , Ma hone y J ,McClelland A, M uchmore E, OhlinAC, Scuderi P: Prevention ofrhinovirus infection in chimpanzeesby soluble intercellular adhesionmolecule-1. Am J Respir Crit CareM ed 1997;155(4): 1206-1210.93 Stepkowski SM, Yu Y, CondonTP , Be nne t t CF : B l oc k i ng o fhe a r t a l l og r a f t r e j e c t i on byintercellular adhesion molecule-Iantisense oligonucleotides aloneor in combina t ion wi th otheri mmunosuppr e s s i ve moda l i t i e s(published erratum appears in JI mm unol 1995 F e b 1 :154( 3) :1521). J Immunol 1994;153(11):5336-5346.94 Stepkowski SM, Tu Y, CondonTP, Benne t t CF: Induc t ion oftransplantation tolerance by treat-me n t w i t h I CA M- I a n t i s e nseoligonucleotides and anti-LFA-1monoc l ona l a n t i bod i e s . Tr a ns -plant Proc 1995;27(1):113.95 Katz SM, Browne B, Phan T,Wa ng ME, Be nne t t CF , S t e p -kowski SM , Kahan BD: Eff icacyofICAM -1 antisense oligonucleo-tide in pancreatic islet transplan-tation. Transplant Proc 1995;27(6):3214.96 Kumasaka T, Quinlan WM , DoyleNA, Condon TP, Sligh J, Takei F,Beaude t A, Benne t t CF, Doer -schuk CM: Role of the in te r -c e l l u l a r a dhe s i on mol e c u l e - I(ICAM-1) in endotoxin-inducedpne umoni a e va l ua t e d us i ngICAM-1 ant i sense ol igonuc leo-t ides, anti- ICAM-1 monoclonalantibodies, and ICAM-I mutantmice. J Clin Invest 1996;97(10):2362-2369.


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