GMI-1070 (rivipansel) inhibits E-selectin recognition of...
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μ"@ `)+$E('. 6NOCBH =(2% &'-0+1; 1(1 '+2 &)2#, 2%(/ ('%(0(-+'K ('1($&-'. 2%&2 (2 (/ '+2 ,#<3(,#1 *+, CB/#)#$-' >#1(&2#1 +32/(1#B(' /(.'&)('. +* ! G B('2#.,(' 1#4#'1#'2 ,+))('. 2+ &,,#/2@ R#/2#,' 0)+2 &'&);/(/ +* 6"7 );/&2#/ (>>3'+4,#$(4(2&2#1 &.&('/2 AW6B/#)#$-' ,#:#&)#1 2%&2 O"LBHUVU &2 ^@\ _" ('%(0(2/ AB/#)#$-' ,#$+.'(-+' +* /C# D +' CB/#)#$-' 0; ]VUa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`)+$E('. &,,#/2 &'1 &$-:&-+' +* 1+='/2,#&> &1%#/(+' ,#$#42+,/ +' )#3E+$;2#/ 0; O"LBHUVU 5,(:(4&'/#)8 >&; 4)&; &' (>4+,2&'2 ,+)# (' (2/ ,#4+,2#1 $)('($&) 0#'#X2 *+, 2%# 2,#&2>#'2 +* :&/+B+$$)3/(:# $,(/(/ (' /($E)# $#)) 4&-#'2/@ J0/2,&$2 GMI-1070 (rivipansel) inhibits E-selectin recognition of sialyl Lewis X expressed on L-selectin and blocks integrin activation and arrest of human neutrophils. Scott I Simon 1 , Vasilios A Morikis 1 , Shannon Chase 1 , and John L. Magnani 2 1 UC Davis Biomedical Engineering, 2 Glycomimetics Inc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b L>4+,2&'$# +* 2%# Y,&'/$#))3)&, T+32# 5GUH^8 !"#$%&'( *% +,,-%./.01 c&/$3)&, >(>#-$B$%(4 2+ <3&'-*; 6"7 ,#$,3(2>#'2 *,+> (/+)&2#1 6"7 3'1#, /%#&, 6"7 &,,#/2 &'1 2,&'/>(.,&-+' +' ('?&>#1 #'1+2%#)(3> (/ ('%(0(2#1 0; O"LBHUVU On Chip • HUU dC +* 4,(>&,; (/+)&2#1 %3>&' 6"7 +02&('#1 *,+> :#'(43'$23,# =&/ /%#&,#1 +:#, AB/#)#$-'WLMJ"BH &2 & =&)) /%#&, /2,#// +* G 1;'#/W$> G @ • L/+)&2#1 %3>&' 6"7/ =#,# 4#,*3/#1 +:#, LCBHF /->3)&2#1 efcAM >+'+)&;#,/ (' & >($,+?3(1($ ?+= $%&>0#, &2 & 4%;/(+)+.($&) /%#&, /2,#// T+))('. +' AB/#) 032 '+2 6B/#) &$-:&2#/ 6"7 &,,#/2 1#4#'1#'2 34+' ,#$+.'(-+' +* CB/#) MZHgWLMJ"BH 0+'1 $)3/2#, &'1 &1%#/(:# $+'2&$2 *+,>&-+' &,# ('%(0(2#1 0; O"LBHUVU 0)+$E('. +* CB/#)#$-' /(.'&)('. • O"LBHUVUK & .);$+>(>#-$ /C#D &'&)+. 1#/(.'#1 2+ ('%(0(2 )#3E+$;2# &1%#/(+' &'1 ('?&>>&-+' 2%,+3.% 0('1('. 2+ 2%# )#$-' 1+>&(' +* AB/#)#$-'K ('%(0(2/ ,#$+.'(-+' +* 6NOCBH &'1 2+ & .,#&2#, #D2#'2 CB/#)#$-'@ • AB/#)#$-' $)3/2#,('. +* CB/#)#$-' +' & ,+))('. 6"7 3'1#, /%#&, ?+= (/ '#$#//&,; &'1 /3S$(#'2 2+ /(.'&) &$-:&-+' +* CIJBH &'1 0+'1 *+,>&-+' 2+ LMJ"BH@ • O"LBHUVU &'2&.+'(Q#/ AB/#)#$-' ,#$+.'(-+' +* /C# D #D4,#//#1 +' CB/#)#$-' &'1 '+2 6NOCBH@ • 6NOCBH &'1 CB/#)#$-' $+B)+$&)(Q# +' 6"7 13,('. ,+))('. +' AB/#)#$-'K 032 CB/#)#$-' &)+'# 2,&'/13$#/ +32/(1#B(' /(.'&)('. 2%&2 )#&1/ 2+ CIJBH 1#4#'1#'2 &,,#/2@ • AB/#)#$-' 0('1('. 2+ 6NOCBH >#1(&2#/ 2#2%#,('. &'1 ,+))('.K 2%#,#0; ('$,#&/('. 2%# 6"7 h &:&()&0)# 2+ 0# &$-:&2#1 0; CB/#)#$-' &'1 &,,#/2 :(& CIJBHWLMJ"BH 0+'1/@ • e(.% &S'(2; MZHg $)3/2#,/ &'1 $+'2&$2 &,#& &,# /(.'(X$&'2); ,#13$#1 =%#' CB/#)#$-' /(.'&)('. (/ 0)+$E#1@ M+'$)3/(+'/ H@ R3' Y@ #2 &)@K 5GUHi8@ j6%&/# H /231; +* 2%# AB/#)#$-' ('%(0(2+, O"LBHUVU (' 4&-#'2/ =(2% /($E)# $#)) &'#>(&@k 6C+N P'# l5HU8b#HHH^lU G@ M%&/#K N@ Z@K #2 &)@ 5GUHG8@ mAB/#)#$-' )(.&'1/ &/ >#$%&'+/#'/(-:# ,#$#42+,/ +' '#32,+4%()/ (' %#&)2% &'1 1(/#&/#@m J'' `(+>#1 A'. iU5i8b gilBg\l@ n@ N2&12>&''K J@K #2 &)@ 5GUHn8@ mY%# 6NOCBHBCB/#)#$-' /(.'&)('. $+>4)#D ,#.3)&2#/ '#32,+4%() &1%#/(+' 3'1#, ?+=@m o AD4 "#1 GHU5HH8b GHVHBGHgU@ i@ N(>+'K N@ L@K #2 &)@ 5GUUU8@ m7#32,+4%() 2#2%#,('. +' AB/#)#$-' &$-:&2#/ 0#2& G ('2#.,(' 0('1('. 2+ LMJ"BH 2%,+3.% & >(2+.#'B&$-:&2#1 4,+2#(' E('&/# /(.'&) 2,&'/13$-+' 4&2%=&;@m o L>>3'+) H^i5g8b inigBin\g@ Y%(/ =+,E =&/ *3'1#1 (' 4&,2 0; JLiVGli 2+ NLN T#*#,#'$#/WJ$E'+=)#1.>#'2/ L>>3'+4,#$(4(2&-+' +* CB/#)#$-'K 032 '+2 6NOCBHK (/ 0)+$E#1 0; O"LBHUVU J$-:&-+' +* MZHg :(& pB)('E('. AB/#)#$-' (/ 0)+$E#1 (' 4,#/#'$# +* O"LBHUVU =>0-/-()" 5$%00/."?-$ • Y,#&2>#'2 +* AB/#)#$-' =(2% O"LBHUVU &'2&.+'(Q#/b ! CB/#)#$-' ,#$+.'(-+' q $)3/2#,('.K 032 '+2 6NOCBH ! e(.% &S'(2; 5eJ8 MZHg $)3/2#, h &'1 $+'2&$2 &,#& *+))+=('. &,,#/2 :(& CIJBH • @2,0- !A!! (/.".(,/ #$.,/ 0'78-(#B0 0-$'C D+!>EFGF (%"(-"#$,)%" ,H-$ ."I'0.%" N($E)# M#)) Z(/#&/# 4&-#'2/ ('*3/#1 =(2% O"LBHUVU /%+= 1#$,#&/#1 6"7 ,#$,3(2>#'2 0 2 4 6 8 0 200 400 600 800 1000 Arrest TEM Time (min) PMN/mm 2 0 12 26 38 54 66 0 500 1000 1500 Arrest TEM GMI-1070 Concentration (μ M) PMN/mm 2 !"#$% '"%( )*)()+, -./01232 )*4#%)"* 5+()6*( 1 78.9 5+()6*( : 78.9 5+(6)*( ; 78.9 5+(6)*( < 78.9 2 ="#$% 2>22 2>22 2>22 2>22 2>? ="#$% 12@>:? A;>AA A;>A; @B>?: : ="#$% 3B>1< ??>@< @A>?: @;>1< < ="#$% ?3>B; <@>2? <@>:A <1>2A B ="#$% <3>A: ;1>2< :?>A; ;B>A? :< ="#$% ;:>:; 1;>:B 12><@ ;@>12 <B ="#$% ?><3 1>;3 2>B< A>;2 0 20 40 60 0 100 200 300 400 500 600 700 Patient 1 Patient 2 Patient 3 Patient 4 y = (-4.46± 1.21)x+290.2 y = (-6.15± 1.94)x+472.3 y = (-8.27± 4.09)x+627.4 y = (-4.24± 0.95)x+304.4 Serum GMI-1070 ( μ M) Arrested PMN (PMN/mm 2 ) 20 30 40 50 -200 -100 0 100 200 300 J-$'C D+!>EFGF ,# K 2$0 5d"8 :.L2 MN".#; 54EK 5a 0&/#)('#8 • L':#,/# $+,,#)&-+' 1#2#$2#1 0#2=##' rO"LBHUVUsK 6"7 &,,#/2K &'1 ('13$-+' +* %(.% &S'(2; MZHg 0 20 40 60 Extended ($) High (#) E-Selectin Crosslinker GMI-1070 (100 μM) IL-8 (10 μM) - - - - - - - + + + - - + + + - + - - - $# $# $ $ CD18 Affinity (Receptor x 10 3 ) IgG PSGL-1 L-sel PSGL-1+L-sel 0 20 40 60 80 100 # ## $$ $$ E-selectin Antibody Blocking PMN Adhesion (# FOV/minute) IgG PSGL-1 L-sel PSGL-1+L-sel 0 20 40 60 80 100 ### Rolling (#) Arrest ($) ### P-selectin Antibody Blocking PMN Adhesion (# FOV/minute) • T+))('. +' AB/#)#$-'WLMJ"BH (/ /344+,2#1 0; 0+2% CB/#)#$-' &'1 6NOCBH • T+))('. +' 6B/#)#$-' (/ 6NOCBH 1#4#'1#'2 &'1 1+#/ '+2 ('13$# &,,#/2 • J$-:&-+' +* &,,#/2 (/ ('%(0(2#1 0; &'-BCB/#) &'1 '+2 &'-B6NOCBH Vehicle GMI-1070 GSnP-6 Untreated GMI-1070 GSnP-6 0 500 1000 1500 2000 Treatment: *** ** *** * L-selectin Content (Relative Density) Vehicle GMI-1070 GSnP-6 Untreated GMI-1070 GSnP-6 0 5000 10000 15000 20000 *** ** *** *** Treatment: PSGL-1 Content (Relative Density) Treatment: 69 kDa - 120 kDa - 210 kDa - - GMI-1070 GSnP-6 - GMI-1070 GSnP-6 IP: E-selectin P-selectin L-selectin Monomeric PSGL-1 Dimeric PSGL-1 • AB/#)#$-' (>>3'+4,#$(4(2&2#/ 0+2% CB/#)#$-' &'1 6NOCBH • O"LBHUVUK 032 '+2 ON'6B^K 0)+$E/ AB/#)#$-' 43)) 1+=' +* CB/#)#$-' • ON'6B^ 0)+$E/ 6B/#)#$-' 43)) 1+=' +* 6NOCBH 2+ & .,#&2#, #D2#'2 2%&' O"LBHUVU O"LBHUVU ('%(0(2/ &$-:&-+' +* 6"7 &,,#/2 13,('. ,+))('. +' (>>+0()(Q#1 AB/#)#$-'WLMJ"BH 0.1 6.5 19.5 100 0.1 12 36 120 0 20 40 60 80 100 120 Arrest Rolling GMI-1070 GSnP-6 *** * Treatment (μ M): PMN Adhesion (% Total Interaction) 0 2 4 6 8 10 12 0 20 40 60 80 100 Vehicle GMI-1070 GSnP-6 * ** Rolling Velocity (μ m/s) PMN Rolling (%) • CB/#)#$-' /(.'&)('. (/ '#$#//&,; &'1 /3S$(#'2 2+ ('13$# F G B('2#.,('WLMJ"BH >#1(&2#1 X,> &,,#/2 +* 6"7 ,+))('. +' AB/#)#$-' • 6"7 ,+))('. :#)+$(2; (/ ('$,#&/#1 0; O"LBHUVU +, ON'6B^ 0)+$E('. 1(/-'$2 /(2#/ +' 2%# )#$-' 1+>&(' +* AB/#)#$-' • ON'6B^ *3'$-+'/ &/ & 6NOCBH >(>#-$ 2+ &'2&.+'(Q# ,+))('.K =%#,#&/ O"LBHUVU 0)+$E/ ,#$+.'(-+' +* CB/#)#$-' &'1 F G B('2#.,(' >#1(&2#1 &,,#/2 Vehicle GMI-1070 GSnP-6 0 5 10 15 ** Rolling to Arrest Treatments: HA CD18 Clusters (# Sites/PMN) Vehicle GMI-1070 GSnP-6 0 1 2 3 4 *** Arrested Treatments: CD18 Area ( μ m 2 /PMN) • ZL >#>0,&'# 1;# ('2#'/(2; 4,+:(1#1 >#&/3,# +* %#(.%2 • CB/#)#$-' &'1 6NOCBH *++24,('2 (>&.#1 =(2% 1(/$,#2# ?3+,+4%+,#/ 3/('. GB)('# <ZI
Transcript of GMI-1070 (rivipansel) inhibits E-selectin recognition of...
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J0/2,&$2!
GMI-1070 (rivipansel) inhibits E-selectin recognition of sialyl LewisX expressed on L-selectin and blocks integrin activation and arrest of human neutrophils.
Scott I Simon1, Vasilios A Morikis1, Shannon Chase1, and John L. Magnani2 1UC Davis Biomedical Engineering, 2Glycomimetics Inc. MD
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6"7!&,,#/2!&'1!2,&'/>(.,&-+'!+'!('?&>#1!#'1+2%#)(3>!(/!('%(0(2#1!0;!O"LBHUVU!
Glass Coverslip
VCAM-1
PDMS Chamber
Reservoir Syringe Pump
CD14++CD16- CD14++CD16+ CD14+CD16++
CD14:FACS
% o
f Max
CD14: Calibration CD14: On Chip
CD14+
CD14++
FACS MFI
On
Chi
p M
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FACS MFI
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N($E)#!M#))!Z(/#&/#!4&-#'2/!('*3/#1!=(2%!O"LBHUVU!/%+=!1#$,#&/#1!6"7!,#$,3(2>#'2!
0 2 4 6 80
200
400
600
800
1000 ArrestTEM
Time (min)
PMN/
mm
2
0 12 26 38 54 660
500
1000
1500
ArrestTEM
GMI-1070 Concentration (µM)
PMN/
mm
2
!"#$%&'"%(&)*)()+,&-./01232&)*4#%)"* 5+()6*(&1&78.9 5+()6*(&:&78.9 5+(6)*(&;&78.9 5+(6)*(&<&78.9
2&="#$% 2>22 2>22 2>22 2>222>?&="#$% 12@>:? A;>AA A;>A; @B>?::&="#$% 3B>1< ??>@< @A>?: @;>1<<&="#$% ?3>B; <@>2? <@>:A <1>2AB&="#$% <3>A: ;1>2< :?>A; ;B>A?:<&="#$% ;:>:; 1;>:B 12><@ ;@>12<B&="#$% ?><3 1>;3 2>B< A>;2
0 20 40 600
100
200
300
400
500
600
700
Patient 1Patient 2Patient 3Patient 4
y = (-4.46±1.21)x+290.2y = (-6.15±1.94)x+472.3y = (-8.27±4.09)x+627.4y = (-4.24±0.95)x+304.4
Serum GMI-1070 (µM)
Arre
sted
PM
N(P
MN
/mm
2 )
20 30 40 50-200
-100
0
100
200
300201 8hr203 8hr204 8hr205 8hr
p = 0.0558r2 = 0.8916
8hr
Serum GMI-1070 at 8 hrs [uM]
Activ
ated
CD
18 [%
bas
elin
e]
J-$'C1D+!>EFGF1,#1K12$015d"8!
:.L2
1MN
".#;
154E
K11
5a!0&/#)('#8!
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0
20
40
60Extended ($)High (#)
E-Selectin Crosslinker
GMI-1070 (100 µM) IL-8 (10 µM)
----
---+
++--
+++-
+---
$#
$#
$ $
CD
18 A
ffini
ty(R
ecep
tor x
103 )
IgG
PSGL-1L-se
l
PSGL-1+L-se
l0
20
40
60
80
100
#
##
$$
$$
E-selectin
Antibody Blocking
PMN
Adhe
sion
(# F
OV
/min
ute)
IgG
PSGL-1L-se
l
PSGL-1+L-se
l0
20
40
60
80
100
###
Rolling (#)Arrest ($)
###
P-selectin
Antibody Blocking
PMN
Adhe
sion
(# F
OV
/min
ute)
•! T+))('.!+'!AB/#)#$-'WLMJ"BH!(/!/344+,2#1!0;!0+2%!CB/#)#$-'!&'1!6NOCBH!•! T+))('.!+'!6B/#)#$-'!(/!6NOCBH!1#4#'1#'2!&'1!1+#/!'+2!('13$#!&,,#/2!•! J$-:&-+'!+*!&,,#/2!(/!('%(0(2#1!0;!&'-BCB/#)!&'1!'+2!&'-B6NOCBH!
Vehi
cle
GM
I-107
0
GSn
P-6
Unt
reat
ed
GM
I-107
0
GSn
P-6
0
500
1000
1500
2000
Treatment:
***
**
***
*
L-se
lect
in C
onte
nt(R
elat
ive
Den
sity
)
Vehi
cle
GM
I-107
0
GSn
P-6
Unt
reat
ed
GM
I-107
0
GSn
P-6
0
5000
10000
15000
20000
***
** ***
***
Treatment:
PSG
L-1
Con
tent
(Rel
ativ
e D
ensi
ty)
E.
Treatment:
D.
69 kDa -
120 kDa -
210 kDa -
- GMI-1070 GSnP-6 - GMI-1070 GSnP-6IP: E-selectin P-selectin
L-selectinMonomeric PSGL-1Dimeric PSGL-1
•! AB/#)#$-'!(>>3'+4,#$(4(2&2#/!0+2%!CB/#)#$-'!&'1!6NOCBH!•! O"LBHUVUK!032!'+2!ON'6B^K!0)+$E/!AB/#)#$-'!43))!1+='!+*!CB/#)#$-'!•! ON'6B^!0)+$E/!6B/#)#$-'!43))!1+='!+*!6NOCBH!2+!&!.,#&2#,!#D2#'2!2%&'!O"LBHUVU!
O"LBHUVU!('%(0(2/!&$-:&-+'!+*!6"7!&,,#/2!13,('.!,+))('.!+'!(>>+0()(Q#1!AB/#)#$-'WLMJ"BH!
0.1 6.5 19.5 100 0.1 12 36 1200
20
40
60
80
100
120
ArrestRolling
GMI-1070 GSnP-6
****
Treatment (µM):PM
N Ad
hesi
on(%
Tot
al In
tera
ctio
n)
0 2 4 6 8 10 120
20
40
60
80
100
VehicleGMI-1070GSnP-6
***
Rolling Velocity (µm/s)
PMN
Rolli
ng(%
)
•! CB/#)#$-'! /(.'&)('.! (/! '#$#//&,;! &'1! /3S$(#'2! 2+! ('13$#! FGB('2#.,('WLMJ"BH!>#1(&2#1!X,>!&,,#/2!+*!6"7!,+))('.!+'!AB/#)#$-'!
•! 6"7!,+))('.!:#)+$(2;!(/! ('$,#&/#1!0;!O"LBHUVU!+,!ON'6B^!0)+$E('.!1(/-'$2!/(2#/!+'!2%#!)#$-'!1+>&('!+*!AB/#)#$-'!
•! ON'6B^! *3'$-+'/! &/! &! 6NOCBH! >(>#-$! 2+! &'2&.+'(Q#! ,+))('.K! =%#,#&/! O"LBHUVU!0)+$E/!,#$+.'(-+'!+*!CB/#)#$-'!&'1!FGB('2#.,('!>#1(&2#1!&,,#/2!
Vehicle GMI-1070 GSnP-60
5
10
15
**
Rolling to Arrest
Treatments:
HA C
D18
Clu
ster
s(#
Site
s/PM
N)
Vehicle GMI-1070 GSnP-60
1
2
3
4
***
Arrested
Treatments:
CD
18 A
rea
(µm
2 /PM
N)
•! ZL! >#>0,&'#! 1;#! ('2#'/(2;!4,+:(1#1!>#&/3,#!+*!%#(.%2!
•! C B /# ) # $-'! &'1! 6NOC BH!*++24, ('2! (>&.#1! =( 2%!1(/$,#2#! ?3+,+4%+,#/! 3/('.!GB)('#!<ZI!
