Biomedical Engineering Reference
In-Depth Information
high affi nity.
196
Hirolog is constructed from covalent linkage of an irreversible thrombin active
site inhibitor to residues 53-65 of the hirudin C-terminal.
197
Studies have suggested that hirudin
can inhibit clot-bound thrombin since no rebound of clot growth occurred after hirudin had been
cleared.
198
Unfortunately, intravenous half-lives of hirudin
199
and hirolog
200
are not signifi cantly lon-
ger than that of UFH, and coagulation does not appear to be terminated since markers of thrombin
generation increase over time even after large hirudin doses.
201
PPACK is an irreversible, selective,
direct thrombin inhibitor that can effi ciently neutralize thrombin on fi brin clots.
202
Similar to the
hirudin derivatives, PPACK is rapidly cleared
203
and has exhibited excessive bleeding risk associ-
ated with doses necessary to achieve plasma concentrations required for treatment.
204
Ximelagatran
(which is metabolically converted to its active melagatran form
in vivo
) has shown promise since
it had treatment outcomes rivaling heparins and could be taken orally.
195
Sadly, the compound has
just been withdrawn by the manufacturer because of reports of serious hepatoxicity in a subpopu-
lation of patients.
205
As this short survey illustrates, issues of effi caciousness, bioavailability, and
hemorrhage in heparinoids have not been clearly overcome in newly developed agents. Thus, our
laboratory and other researchers have attempted to address heparin's limitations by pursuing a more
radical direction in heparinoid modifi cation.
18.4.2 P
OTENTIAL
A
DVANTAGES
OF
C
OVALENT
A
NTITHROMBIN
-H
EPARIN
C
OMPLEXES
Given that many of the deleterious effects of heparin occur when it is in the free state, dissociated
from its target serpin, concepts were brought forward for permanently stabilizing AT complexes
with heparin. A number of theoretical concepts that might logically ensue from the linkage of
serpin to GAG incited the design of covalent ATH compounds. A discourse follows on this line of
thought.
Many of the likely advantages of AT attachment to heparin revolve around the prevention of
heparin dissociation from the inhibitor. Since AT in ATH cannot dissociate from the heparin chain,
the serpin will always be in the activated state. Therefore, wherever the conjugated AT is located
in vivo
, it will always be in the most reactive form for inhibition of thrombin, FXa, etc. Furthermore,
reaction with this highly activated anticoagulant will not depend on conditions within the fl uid
phase or tissue environment that might lead to disruption of the noncovalent AT-UFH complex. An
added benefi t associated with the lack of dissociation of heparin in ATH is that ATH should be read-
ily able to inhibit fi brin clot-bound thrombin. Obviously, ATH heparin cannot detach to generate
thrombin-heparin-fi brin complexes
177
that are resistant to inhibition by ATH.
178
Additionally, any
attraction of thrombin-fi brin to heparin will necessarily also locate the activated AT moiety in the
conjugate close to the bound thrombin for an inhibition reaction to take place. In conjunction with
colocalization of ATH to the sites of coagulant insult, permanent complexation of AT with heparin
should maintain the active heparin within the vascular space. Since heparin is fi xed to AT in ATH,
the heparin chains cannot be lost through glomerular fi ltration as the ATH is vastly increased in
size.
141,206
Moreover, loss of activity from the circulation due to plasma or cell surface protein bind-
ing is likely to be curtailed because of masking of a signifi cant proportion of the heparin chain
that is forcibly associated with the AT polypeptide. Such perpetual interaction of a segment of the
heparin will prevent other proteins to approach it because of steric hindrance. A primary advance
in heparin application resultant from ATH preparation depends on the synthetic methodology
employed. If AT is allowed to interact noncovalently prior to permanent bonding, selection could
take place so that only heparin molecules with high-affi nity pentasaccharide sequences appear in
the ATH product. Thus, in adducts from such protocols, all serpins within the ATH molecules will
be activated by a potent pentasaccharide, as opposed to the starting commercial UFH in which
only one in three chains have the active sequence.
141,207
Once made, direct enhancement of hepa-
rin as an anticoagulant would be observed. Previous studies have indicated that binding of AT to
heparin is the rate-determining step in heparin-catalyzed AT inhibition of thrombin.
208
Of course,
linkage of heparin to AT eliminates this requirement. Other studies have revealed that a surfeit of
