ライフサイエンスコーパス: FXIa

1 FXIa and its isolated light chain (FXIa-LC) cleave S-236

2 FXIa binds to heparin (Kd approximately 0.7 x 10(-9) M)

3 FXIa contains four apple domains (A1-A4) and a catalytic

4 FXIa(G193E) activated FIX with approximately 300-fold re

5 FXIa(G193E) inhibition by diisopropyl fluoro-phosphate w

6 FXIa-PN1 complexes are shown to be internalized and degr

7 Factor XIa and 1/2-FXIa activate the substrate factor IX, with similar kine

8 gh an intermediate with one active site (1/2-FXIa).

9 intermediate formation was detected with 1/2-FXIa, factor XIa with one inhibited active site, or a re

10 ery of a potent FXIa clinical candidate, 55 (FXIa Ki = 0.7 nM), with excellent preclinical efficacy i

11 Ia series, exemplified by compound 16, had a FXIa Ki = 0.16 nM with potent anticoagulant activity in

12 sed to block FXIIa-dependent FXI activation, FXIa-dependent factor IX (FIX) activation, or platelet-d

13 In addition, FXIa can activate factor FXII, which could contribute to

14 t SPGG's sulfation level moderately affected FXIa inhibition potency and selectivity over thrombin an

15 itor 6 displayed a potency of 551 nM against FXIa, which was at least 200-fold more selective than ot

16 ate that exhibits equivalent potency against FXIa.

17 izing p-aminobenzamidine, was intact for all FXIa mutants.

18 ected to open up a major route to allosteric FXIa anticoagulants with clinical relevance.

19 n FXIa/S557A but not in FXIa/C362S/C482S and FXIa/S557A/C362S/C482S.

20 57A, FXIa-HC, FXIa-LC, FXIa/C362S/C482S, and FXIa/S557A/C362S/C482S.

21 xhibited near normal reactivity with FXa and FXIa in the absence of cofactors and in the presence of

22 , we examined binding of factor XI (FXI) and FXIa to heparin.

23 lusion, the ability of plasma kallikrein and FXIa to activate pro-HGF in vitro raises the possibility

24 The concentrations of kallikrein and FXIa to cleave 50% (EC(50)) of (125)I-labeled pro-HGF du

25 tely resistant to cleavage by kallikrein and FXIa.

26 inhibitory complexes formed between PN1 and FXIa are stable when subjected to reducing agents, SDS,

27 able rates of carbamylation for FXIa(WT) and FXIa(G193E), suggesting that the occupied active site ha

28 hrombin flux after 500 s was blocked by anti-FXIa antibody (O1A6), consistent with thrombin-feedback

29 of tetrahydroquinoline (THQ) derivatives as FXIa inhibitors.

30 vestigate the molecular interactions between FXIa and the small synthetic substrate (S-2366), the mac

31 Unlike other known activators, both FXIa and kallikrein processed pro-HGF by cleavage at two

32 es the activation of factor IX by cell bound FXIa.

33 a-LC) cleave S-2366 at comparable rates, but FXIa-LC is a very poor activator of FIX, possibly becaus

34 PAB inhibited the hydrolysis of S-2366 by FXIa-LC in a classically competitive fashion.

35 nionic surface but not for FIX activation by FXIa in solution.

36 eavage products resulting from activation by FXIa or FXIa-LC.

37 e rate of factor IXa generation catalyzed by FXIa was unaffected by the presence of surfaces; however

38 potentiation in the rate of FIX cleavage by FXIa-LC.

39 into the mechanisms of activation of FIX by FXIa, we have investigated the kinetic properties of FXI

40 as FXIa/S557A inhibited FIX activation of by FXIa, FXIa-HC did not.

41 ysis of a sensitive fluorogenic substrate by FXIa in the presence of PN-2 to ascertain the kinetic ra

42 FXIa and its isolated light chain (FXIa-LC) cleave S-2366 at comparable rates, but FXIa-LC

43 the kinetic properties of FXIa-light chain (FXIa-LC) with its active site occupied by either a rever

44 Specific and saturable zinc-dependent FXIa binding was demonstrated to 250 +/- 48 sites per ac

45 activated FIX similarly to wild-type dimeric FXIa.

46 the concentration of NaCl required to elute FXIa from heparin-Sepharose.

47 Factor XI (FXI) is the zymogen of an enzyme (FXIa) that contributes to hemostasis by activating facto

48 When converted to the active enzyme, FXIa, all the monomeric mutants activated FIX similarly

49 ted in comparable rates of carbamylation for FXIa(WT) and FXIa(G193E), suggesting that the occupied a

50 tive serine proteases, was 4-fold faster for FXIa(G193E).

51 d peptide substrate, the K(assoc) of PN1 for FXIa was determined to be 7.9 x 10(4) m(-)(1) s(-)(1) in

52 e (S1 site probe) was 6-fold weaker than for FXIa(WT).

53 red its activity to wild type-activated FXI (FXIa(WT)).

54 t has also been reported that activated FXI (FXIa) binds to 1.5 x 10(6) sites per HUVEC and promotes

55 tivation; and inactivation of activated FXI (FXIa) by serine protease inhibitors.

56 The primary substrate of activated FXI (FXIa) is FIX, leading to FX activation (FXa) and thrombi

57 iring factor IX activation by activated FXI (FXIa).

58 physiologic site for reactions involving FXI/FXIa.

59 of prekallikrein (PK) or factor XI/XIa (FXI/FXIa) binding to endothelial cells (HUVECs).

60 ng to an exosite on the heavy chain of FXIa (FXIa-HC) required for optimal cleavage rates of the two

61 a/S557A inhibited FIX activation of by FXIa, FXIa-HC did not.

62 examined FIX binding to FXIa/S557A, FXIa-HC, FXIa-LC, FXIa/C362S/C482S, and FXIa/S557A/C362S/C482S.

63 (6) as a highly selective inhibitor of human FXIa.

64 mportance, Arg15, Phe34, Pro13, and Arg20 in FXIa inhibition by PN2KPI.

65 n conclusion, enzyme activity is impaired in FXIa when Gly193 is replaced by a non-Gly residue, and r

66 avy and light chains are disulfide-linked in FXIa/S557A but not in FXIa/C362S/C482S and FXIa/S557A/C3

67 re disulfide-linked in FXIa/S557A but not in FXIa/C362S/C482S and FXIa/S557A/C362S/C482S.

68 other SERPIN known to significantly inhibit FXIa.

69 completely abolished its ability to inhibit FXIa.

70 C38 were tested for their ability to inhibit FXIa.

71 Zn2+ augments the ability of PN-2 to inhibit FXIa.

72 uman and murine plasma, and PN2KPI inhibited FXIa activity in both human and murine plasma in vitro.

73 smooth muscle cells was active in inhibiting FXIa.

74 ets and in plasma of pregnant women inhibits FXIa and tissue-type plasminogen activator-induced clot

75 with a saturating concentration of isolated FXIa-HC did not result in any potentiation in the rate o

76 yme-substrate interaction, then the isolated FXIa-HC should inhibit the rate of FIX activation by dep

77 FIX binding to FXIa/S557A, FXIa-HC, FXIa-LC, FXIa/C362S/C482S, and FXIa/S557A/C362S/C482S.

78 A novel series of macrocyclic FXIa inhibitors was designed based on our lead acyclic p

79 The macrocyclic FXIa series, exemplified by compound 16, had a FXIa Ki =

80 crocycles, which were double-digit nanomolar FXIa inhibitors, were further optimized with assistance

81 ome of these inhibitors showed low nanomolar FXIa inhibitory activity with >1000-fold FXa selectivity

82 Addition of FXIa to plasma was able to reverse the ability of TFPI t

83 ntration-dependent decrease in the amount of FXIa bound to HUVEC.

84 Therefore, the binding of FXIa to activated platelets was compared with FXIa bindi

85 binding to an exosite on the heavy chain of FXIa (FXIa-HC) required for optimal cleavage rates of th

86 vy chain and the other on the light chain of FXIa is required to mediate the formation of the Michael

87 absence of either Ca2+ or the heavy chain of FXIa there was substantial accumulation of the inactive

88 essential for the structure-based design of FXIa-selective inhibitors.

89 the complex between the catalytic domain of FXIa and the Kunitz protease inhibitor (KPI) domain of a

90 CHWA-D Abeta caused a >5-fold enhancement of FXIa inhibition by PN-2/AbetaPP.

91 f FIX binding via the heavy chain exosite of FXIa determines the affinity of the enzyme-substrate int

92 ction of anionic SAMs to the cationic HBS of FXIa that forms a locked complex through tight interacti

93 esidues of the heparin-binding site (HBS) of FXIa introduced a nearly 5-fold loss in inhibition poten

94 odeling also indicates that the inability of FXIa(G193E) to bind antithrombin/APPI or activate FIX is

95 ation and Zn2+ increases the inactivation of FXIa.

96 n and are required for optimal inhibition of FXIa and FXa.

97 Inhibition of FXIa by ATIII in the presence of heparin was decreased 4

98 c rate constants governing the inhibition of FXIa by PN-2.

99 exert opposite effects on the inhibition of FXIa by PN-2.

100 Inhibition of FXIa by PN2 requires interactions between the two protei

101 Thus, inhibition of FXIa by PN2KPI is a promising approach to antithrombotic

102 mined the effect of heparin on inhibition of FXIa by the inhibitors C1-inhibitor (C1-INH) and antithr

103 fect was also observed for the inhibition of FXIa by ZPI mutants.

104 evidence suggests that direct inhibition of FXIa can block pathologic thrombus formation while prese

105 pound 16b, a potent, reversible inhibitor of FXIa (Ki = 0.3 nM) having in vivo antithrombotic efficac

106 (-)(1), making PN1 a far better inhibitor of FXIa than C1 inhibitor, which is the only other SERPIN k

107 2) is a potent, highly specific inhibitor of FXIa, suggesting its possible role in the inhibition of

108 and a physiologically important inhibitor of FXIa.

109 highly selective small molecule inhibitor of FXIa.

110 Novel inhibitors of FXIa containing an (S)-2-phenyl-1-(4-phenyl-1H-imidazol-

111 tent, selective peptidomimetic inhibitors of FXIa were designed and synthesized.

112 efforts at identifying potent inhibitors of FXIa with a focus on discovering an acute antithrombotic

113 XIa activity, including direct inhibitors of FXIa, have demonstrated good antithrombotic efficacy wit

114 hanism of FXI activation, the interaction of FXIa with the substrate factor IX, and the binding of FX

115 tanding of substrate binding interactions of FXIa, the structural information essential for the struc

116 the release of PN-2, limits the lifetime of FXIa activity within the locus of activated platelets.

117 ch predominates and prolongs the lifetime of FXIa after platelet activation.

118 Inhibitor 6 reduced V(MAX) of FXIa hydrolysis of chromogenic substrate without affecti

119 We conducted sensitive measurements of FXIa activity in the presence of human platelets before

120 To investigate the potential of FXIa inhibitors as safe anticoagulants, a series of pote

121 have investigated the kinetic properties of FXIa-light chain (FXIa-LC) with its active site occupied

122 Partial reduction of FXIa/S557A to produce heavy and light chains resulted in

123 results suggest that the hemostatic role of FXIa may be attributed not only to activation of FIX but

124 indicate that the unoccupied active site of FXIa(G193E) is incompletely formed, and the amide N of G

125 ue structural features in the active site of FXIa.

126 conformational changes in the active site of FXIa.

127 that 6 bound in the heparin-binding site of FXIa.

128 To determine the structure of FXIa, we derived a recombinant catalytic domain of FXI,

129 with kinetic parameters similar to those of FXIa.

130 Factor XI (FXI) is the zymogen of FXIa, which cleaves FIX in the intrinsic pathway of coag

131 molecular weight kininogen had no effect on FXIa binding to platelets, but revealed a concentration-

132 Similar stimulatory effects on FXIa inhibition by PN-2/AbetaPP were also observed with

133 e than one anion-binding, allosteric site on FXIa.

134 ylglucopyranosyl scaffold may be the optimal FXIa inhibitor for further preclinical studies.

135 K preferentially bound to HUVECs over FXI or FXIa.

136 roducts resulting from activation by FXIa or FXIa-LC.

137 tic activation of FXI by FXIIa, thrombin, or FXIa either in solution or on an anionic surface but not

138 trations, PK (450 nM) abolished FITC-FXI or -FXIa binding to HUVEC suspensions in the absence or pres

139 ons without added Zn2+, whereas FITC-FXI or -FXIa binding to HUVEC suspensions required 10 microM add

140 the absence of HK, the level of FITC-FXI or -FXIa binding was half that seen in its presence.

141 ormal substrate binding compared with plasma FXIa; however, all except E98A and K192A had impaired va

142 Herein we describe the discovery of a potent FXIa clinical candidate, 55 (FXIa Ki = 0.7 nM), with exc

143 in the FXIa active site, resulting in potent FXIa inhibitors.

144 Predictably, FXIa activation of FIX(PCEGF1) was normal, whereas it wa

145 the reported FXIa Glu193 mutant, we prepared FXIa with Asp (short side chain) or Lys (opposite charge

146 nly the activated platelet surface protected FXIa from inhibition by protease nexin 2.

147 As in the purified system, HK protects FXIa from inactivation and Zn2+ increases the inactivati

148 HK protects FXIa from inactivation in a dose dependent and saturable

149 193E), we expressed and purified recombinant FXIa(G193E), activated it to FXIa(G193E), and compared i

150 dies using a variety of approaches to reduce FXIa activity, including direct inhibitors of FXIa, have

151 r (KPI) domain of a physiologically relevant FXIa inhibitor, protease nexin 2 (PN2).

152 th venous and arterial thrombosis, rendering FXIa a potential target for the development of antithrom

153 For comparison to the reported FXIa Glu193 mutant, we prepared FXIa with Asp (short sid

154 fore, we examined FIX binding to FXIa/S557A, FXIa-HC, FXIa-LC, FXIa/C362S/C482S, and FXIa/S557A/C362S

155 Since FXIa proteolytically modifies the amyloid precursor prot

156 Six FXIa catalytic domain residues (Glu(98), Tyr(143), Ile(1

157 To better understand the SPGG-FXIa interaction, we utilized eight SPGG variants and a

158 the extrinsic pathway followed by late-stage FXIa contributions, with fibrin localizing thrombin via

159 s emerges as a novel anticoagulant targeting FXIa under conditions in which the coagulation activatio

160 We found that FXIa neutralized both endothelium- and platelet-derived

161 These results imply that FXIa is the primary in vivo target for Desmolaris at ant

162 activation of FXI in plasma and suggest that FXIa may provide a link between tissue factor-initiated

163 f crystal structures for zymogen FXI and the FXIa catalytic domain have enhanced our understanding of

164 e conclude that substrate recognition by the FXIa exosite(s) requires disulfide-linked heavy and ligh

165 This demonstrates proof of concept for the FXIa mechanism in animal models with a reversible, small

166 rogen bond with the carbonyl of Leu41 in the FXIa active site, resulting in potent FXIa inhibitors.

167 ed extensive pharmacologic evaluation of the FXIa mechanism up to the ID90 for thrombus inhibition.

168 Occupancy of the active site of the FXIa-LC by S-2366 also resulted in noncompetitive inhibi

169 alysis of the X-ray crystal structure of the FXIa/11h complex.

170 resence of an exosite for FIX binding on the FXIa-LC remote from its active site.

171 from P5 to P2' in ecotin was mutated to the FXIa substrate sequence, and the structures of the rhFXI

172 a-branching causes steric conflicts with the FXIa 140-loop, which could perturb the local tertiary st

173 toward macromolecular interactions with the FXIa mutants.

174 Therefore, FXIa is a potential target for anti-thrombosis therapy.

175 Thus FXIa, generated on contact phase activation, cleaves che

176 Mz-IIa catalyze factor (F) XI activation to FXIa, which sustains alpha-thrombin production through a

177 or protein Kunitz domain inhibitor (APPI) to FXIa(G193E) was impaired approximately 8000- and approxi

178 FXI undergoes autoactivation to FXIa in the presence of heparin.

179 Therefore, we examined FIX binding to FXIa/S557A, FXIa-HC, FXIa-LC, FXIa/C362S/C482S, and FXIa

180 ucture of tetrahydroquinoline 3 complexed to FXIa.

181 ied recombinant FXIa(G193E), activated it to FXIa(G193E), and compared its activity to wild type-acti

182 Wild type FXIa eluted from the column at 320 mM NaCl, whereas FXIa

183 However, whereas FXIa/S557A inhibited FIX activation of by FXIa, FXIa-HC

184 uted from the column at 320 mM NaCl, whereas FXIa with multiple substitutions (A252-254 or A250-255)

185 ovel model for factor IX activation in which FXIa binds to activated platelets by one chain of the di

186 Treatment of cultured endothelial cells with FXIa increased the generation of FXa and promoted TF-dep

187 XIa to activated platelets was compared with FXIa binding to HUVEC and HEK293 cells immobilized on mi

188 ly elevated stoichiometry of inhibition with FXIa in the absence of heparin.

189 strates its unique binding interactions with FXIa.

190 nce heparin potentiates activated factor XI (FXIa) inhibition by protease nexin-2 by providing a temp

191 Activated Factor XI (FXIa) is a key enzyme in the amplification phase of the

192 ZPI) inhibition of factors Xa (FXa) and XIa (FXIa) by a template mechanism.

193 sic pathway of blood coagulation factor XIa (FXIa) activates factor IX (FIX) by cleaving the zymogen

194 Factor XIa (FXIa) cleaved chem163S, generating a novel chemerin form

195 Factor XIIa (FXIIa) and factor XIa (FXIa) contribute to thrombosis in animal models, whereas

196 ic agents that are inhibitors of factor XIa (FXIa) have the potential to demonstrate robust efficacy

197 Factor XIa (FXIa) is a blood coagulation enzyme that is involved in

198 Inhibition of factor XIa (FXIa) is a novel paradigm for developing anticoagulants

199 Factor XIa (FXIa) is a serine protease important for initiating the

200 an exosite on the heavy chain of factor XIa (FXIa) is essential for the optimal activation of FIX.

201 o select residues in coagulation factor XIa (FXIa) potentially important for substrate and inhibitor

202 Human coagulation factor XIa (FXIa), a serine protease activated by site-specific clea

203 Moreover, the activity of factor XIa (FXIa), but not FXIIa, was higher in APP-KO mice compared

204 teases (such as Factor Xa (FXa), Factor XIa (FXIa), urokinase-type plasminogen activator (uPA), throm

205 g proteins (GBPs), such as human factor XIa (FXIa), we screened a library of 26 synthetic, sulfated q

206 t with Ala or Val in coagulation factor XIa (FXIa).

207 lasma kallikrein and coagulation factor XIa (FXIa).

208 potent inhibitor of coagulation factor XIa (FXIa).

209 hibitor of the blood coagulation Factor XIa (FXIa).

210 e inhibitor of blood coagulation factor XIa (FXIa).

211 G) as an allosteric inhibitor of factor XIa (FXIa).