U.S. patent application number 10/086286 was filed with the patent office on 2002-08-01 for collagen fabrics.
Invention is credited to Eldridge, Stephen N., Maddalo, Francis B., Torgerson, Robert D., Uhoch, John.
Application Number | 20020103503 10/086286 |
Document ID | / |
Family ID | 22780002 |
Filed Date | 2002-08-01 |
United States Patent
Application |
20020103503 |
Kind Code |
A1 |
Torgerson, Robert D. ; et
al. |
August 1, 2002 |
Collagen fabrics
Abstract
A collagen fiber and methods for producing and using the
collagen fiber of the inventions to prepare hemostatic fabrics to
control bleeding are provided. The collagen particles (e.g.,
fibrils) of the fiber, preferably, have a hemostatic activity that
is equivalent to the hemostatic activity of the collagen particles
from which the fiber is formed. The collagen fibers of the
invention and hemostatic fabrics formed thereof optionally include
hemostatic agents and/or other therapeutic agents, to further
promote hemostasis and wound healing.
Inventors: |
Torgerson, Robert D.;
(Wakefield, RI) ; Uhoch, John; (Warwick, RI)
; Maddalo, Francis B.; (Needham, MA) ; Eldridge,
Stephen N.; (Cranston, RI) |
Correspondence
Address: |
Elizabeth R. Plumer
Wolf, Greenfield & Sacks, P.C.
600 Atlantic Avenue
Boston
MA
02210
US
|
Family ID: |
22780002 |
Appl. No.: |
10/086286 |
Filed: |
March 1, 2002 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10086286 |
Mar 1, 2002 |
|
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09209723 |
Dec 11, 1998 |
|
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6361551 |
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Current U.S.
Class: |
606/214 |
Current CPC
Class: |
A61L 2400/04 20130101;
D01F 4/00 20130101; D01D 5/06 20130101; A61L 15/325 20130101 |
Class at
Publication: |
606/214 |
International
Class: |
A61B 017/08 |
Claims
What is claimed:
1. A hemostatic fabric comprising collagen fibrils, wherein the
fibrils have not been subjected to acid dissolution
2. The fiber of claim 1, wherein the collagen fibrils comprise
microfibrillar collagen fibrils.
3. The fiber of claim 1, wherein the collagen fibrils comprise a
collagen flour.
4. The fiber of claim 1, wherein the collagen fibrils have a bulk
density in the range of from about 1.5 to about 3.5
lbs/ft.sup.3.
5. The fiber of claim 1, further comprising a hemostatic agent.
6. A fabric formed of a plurality of collagen fibers of claim
1.
7. A sterile package containing a fabric according to claim 6.
8. A collagen fiber formed of collagen fibrils, wherein the
collagen fibrils are not denatured.
9. The fiber of claim 8, wherein the collagen fibrils comprise
microfibrillar collagen fibrils.
10. The fiber of claim 8, wherein the collagen fibrils comprise a
collagen flour.
11. The fiber of claim 8, wherein the collagen fibrils have a bulk
density in the range of from about 1.5 to about 3.5
lbs/ft.sup.3.
12. The fiber of claim 8, further comprising a hemostatic
agent.
13. A fabric formed of a plurality of collagen fibers of claim
8.
14. A sterile package containing a fabric according to claim 13.
Description
RELATED APPLICATIONS
[0001] This application is a continuation of U.S. Ser. No.
09/209,723, filed Dec. 11, 1998, entitled "COLLAGEN HEMOSTATIC
FIBERS," the entire contents of which application incorporated
herein by reference.
FIELD OF THE INVENTION
[0002] This invention relates to the field of hemostatic devices
for controlling bleeding.
BACKGROUND OF THE INVENTION
[0003] Uncontrolled bleeding can result in shock and death. In
surgical patients and patients receiving anticoagulant medication,
the problem of rapid blood loss arising from, for example, a
hemorrhage of a blood vessel, body tissue, organ or bone can give
rise to a life threatening situation.
[0004] Biodegradable devices for controlling bleeding are
commercially available. However, many of these devices require the
impregnation of protein agents such as thrombin or fibrinogen to be
effective. Unfortunately, special storage conditions are required
to preserve the hemostatic activity of these protein agents. For
example, many of these devices must be stored under refrigeration
conditions to maintain the bioactivity of the hemostatic devices
into which the protein agents have been impregnated. Such
requirements prohibit certain field applications of the device,
where refrigeration facilities are unavailable. Another problem
with certain commercially available hemostatic devices is their
lack of flexibility in the dry state. Many hemostatic devices do
not conform easily to the shape of the body surface to which it is
applied. In addition, hemostatic devices which further include
hemostatic agents, such as thrombin, typically require that the
thrombin be reconstituted and added to the dry devices immediately
before use to provide a flexible hemostatic device having
sufficient hemostatic activity to control bleeding.
SUMMARY OF THE INVENTION
[0005] The invention provides a hemostatic collagen fiber which can
be processed into a hemostatic fabric. The collagen fibers and
hemostatic fabric of the invention are collectively referred to
herein as "hemostatic devices". The hemostatic devices of the
invention solve the above-described and other problems of the prior
art hemostatic fibers and fabrics. For example, the hemostatic
devices of the invention do not require exogenously added protein
agents to be effective. Accordingly, the hemostatic devices of the
invention can withstand elevated temperatures and do not require
refrigeration to retain hemostatic efficacy. In addition, the
hemostatic fabrics of the invention are easy to use and mold easily
to body contours. Accordingly, the hemostatic fabrics of the
invention are particularly useful for treating the problematic
hemorrhages of parenchymal organs, spine and brain.
[0006] According to one aspect of the invention, a hemostatic
collagen fiber is provided. Hemostasis is a term of art which
refers to cessation of bleeding. The collagen fiber contains
collagen particles (preferably, collagen fibrils) which,
preferably, have a hemostatic activity that is equivalent to the
hemostatic activity of the collagen fibrils from which the fiber is
formed. In the preferred embodiments, the fiber is prepared by a
process which involves extruding a collagen slurry containing
collagen fibrils into a dehydrating bath. An exemplary process is
summarized below and described in detail in the Examples.
[0007] In one embodiment of the invention, the method for forming a
collagen fiber of the invention involves suspending a plurality of
collagen particles (preferably, collagen fibrils) in water to form
a collagen slurry. The collagen fibrils have a bulk density
sufficient to form a suspension in water. In general, the bulk
density of the collagen fibrils is in the range of from about 1.5
to about 3.5 lbs/ft.sup.3 and, more preferably, from about 2 to
about 3 lbs/ft.sup.3. The fibrils are suspended in water to obtain
a collagen concentration in the range of about 3% to about 10%
(weight/volume). Preferably, the collagen fibrils of the fiber have
a hemostatic activity that is equivalent to the hemostatic activity
of the collagen fibrils from which the fiber is formed. In the
preferred embodiments, the collagen fibers are formed of collagen
fibrils that have not been subjected to acid dissolution or other
denaturing conditions.
[0008] According to the foregoing embodiment, the collagen slurry
is introduced into a first dehydrating bath to at least partially
dehydrate the collagen slurry and, thereby, form a collagen fiber.
The collagen fiber, optionally, is introduced into a second
dehydrating bath to further dehydrate the fiber. Exemplary
dehydrating baths that can be independently selected for use as the
first or second (if present) dehydrating baths include (1) an
ammonia bath comprising from about 10% to about 30% ammonia in
water; (2) an ammonia/acetone bath comprising from about 50% to
about 70% ammonia in acetone; (3) an acetone bath; (4) an ethanol
bath; (5) an isopropanol bath (containing about 70% isopropanol in
water); and (6) a propylene glycol bath (containing about 30% to
about 95% propylene glycol in water). Preferably, the first
dehydrating bath is an ammonia bath and the second dehydrating bath
is an acetone bath.
[0009] According to yet another aspect of the invention, a collagen
fiber prepared by the above-described process is provided.
According to yet another aspect of the invention, a collagen fabric
formed of the collagen fibers of the invention and methods of
preparing the collagen fabrics are provided. Such fabrics are also
referred to herein as "hemostatic fabrics" of the invention.
[0010] The collagen fibers and hemostatic fabrics of the invention
are referred to herein as "hemostatic devices" of the invention.
Such hemostatic devices can be sterilized and packaged in a sterile
package for pharmaceutical applications.
[0011] In certain embodiments, the hemostatic devices of the
invention further include a hemostasis-promoting amount of at least
one hemostatic agent. As used herein, a "hemostasis-promoting
amount" is the amount effective to accelerate clot formation at an
interface between a surface (e.g., of a wound or lesion) and the
hemostatic fabric. Exemplary hemostatic agents include a thrombin
molecule, a fibrinogen molecule, a source of calcium ions, an RGD
peptide, protamine sulfate, an epsilon amino caproic acid, and
chitin. In the preferred embodiments, the hemostatic agent is
thrombin. The hemostatic agents can be introduced into the
hemostatic devices at any stage during the preparation of these
devices.
[0012] In certain embodiments, the hemostatic devices of the
invention further include a therapeutically effective amount of at
least one therapeutic agent, such as agents which promote
wound-healing and or reduce pain (e.g., vascular pain). Agents
which promote wound-healing and/or reduce pain include
anti-inflammatory agents (steroidal and non-steroidal) such as
agents which inhibit leukocyte migration into the area of surgical
injury, anti-histamines; agents which inhibit free radical
formation; and bacteriostatic or bacteriocidal agents.
[0013] Various additives, optionally, can be incorporated into the
hemostatic devices of the invention without adversely affecting the
hemostatic activity of these devices. The term
"pharmaceutically-acceptab- le carrier" as used herein means one or
more compatible solid or liquid fillers, diluents or encapsulating
substances which are suitable for administration into a human. The
term "carrier" denotes an organic or inorganic ingredient, natural
or synthetic, with which the active ingredient is combined to
facilitate the application. The components of the pharmaceutical
compositions also are capable of being co-mingled with the collagen
fibrils of the present invention, and with each other, in a manner
such that there is no interaction which would substantially impair
the desired hemostatic activity.
[0014] The hemostatic devices of the invention are useful for
promoting hemostasis at a site of bleeding (e.g., reducing or
eliminating bleeding from a wound). Accordingly, a further aspect
of the invention involves a method for promoting hemostasis. In
general, such methods of the invention involve manually pressing a
hemostatic fabric formed of the collagen fibers of the invention
against a surface of a wound or a surface of a lesion on an organ,
such as a parenchymal organ (e.g., spleen, liver, lung or
pancreas), the spine, or the brain, for a period of time until
clotting has occurred at the interface between the hemostatic
fabric and the surface.
[0015] According to yet another aspect of the invention, a collagen
fiber is provided, wherein the collagen fibrils of the fiber have a
hemostatic activity that is equivalent to the hemostatic activity
of the collagen fibrils from which the fiber is formed. Hemostatic
fabrics formed of such collagen fibers also are provided. Although
not wishing to be bound to any particular theory or mechanism, it
is believed that avoiding contact between the collagen and an acid
solution and minimizing exposure of the collagen to a denaturing
condition such as, e.g., excess mechanical shear, high temperature,
or long water residence times, during the fiber- or fabric-forming
process results in a greater retention of hemostatic activity by
the collagen.
[0016] A number of embodiments of the invention are summarized
above. However, it should be understood that the various
limitations presented in each embodiment are not mutually exclusive
and, accordingly, the limitations can be combined to obtain further
aspects of the invention.
BRIEF DESCRIPTION OF THE DRAWINGS
[0017] FIG. 1 illustrates a process for forming a collagen fiber in
accordance with the invention.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS
[0018] According to one aspect of the invention, a collagen fiber
having hemostatic activity is provided. The collagen fiber of the
invention comprises a biodegradable fiber formed of a collagen,
such as a microfibrillar collagen (e.g., absorbable Avitene.RTM.
flour), which has not been subjected to acid dissolution or exposed
to denaturing conditions. The collagen fibers in the invention
possess a hemostatic activity. Although applicants do not wish to
be bound to one particular theory or mechanism, it is believe that
avoiding contact of the collagen with acid solution and minimizing
exposure of the collagen to denaturation steps prior to and during
the fiber-and-fabric-forming process, results in a greater
retention of the hemostatic activity by the collagen starting
material. In a preferred embodiment, the collagen fiber of the
invention is formed of Avitene.RTM. flour which has not been
subjected to acid dissolution or exposed to denaturing conditions,
such as, e.g., excess mechanical shear, high temperatures or long
water residence times. Accordingly, the invention provides a
hemostatic devices having unexpected improved hemostatic properties
compared to the hemostatic devices of the prior art that are formed
by processes which involve collagen dissolution in acid solution or
exposure to denaturing conditions.
[0019] According to one aspect of the invention, a method for
forming a collagen fiber of the invention is provided. The method
involves: (a) suspending a plurality of collagen particles
(preferably, collagen fibrils) in water to form a "collagen
slurry", wherein the collagen fibrils have a bulk density
sufficient to form a suspension in water (preferably, in the range
of from about 1.5 to about 3.5 lbs/ft.sup.3) and wherein the
collagen slurry has a collagen concentration in the range of about
3% to about 10% (weight/volume); and (b) introducing the collagen
slurry into a first dehydrating bath to at least partially
dehydrate the collagen slurry and thereby form a collagen fiber. In
certain preferred embodiments, the collagen fibrils that are used
to form the collagen fibers of the invention have a bulk density in
the range of from about 2 to about 3 lbs/ft.sup.3. In these and
other certain preferred embodiments, the collagen slurry has a
collagen concentration in the range of about 5% to about 8%
(weight/volume).
[0020] In general, it is preferred that the process includes the
further step of degassing the collagen slurry prior to extruding
the slurring into the first dehydrating bath. The process can be
accomplished on a relatively small scale by employing a syringe as
an extruder. For example, the collagen slurry can be placed into a
syringe (e.g., 5 to 60 cc) which is used an "extruder" for
extruding the slurring into a dehydrating bath. A stopper is placed
on the luer-lock end and the syringe is placed into a centrifuge.
The "slurry" preferably is centrifuged to remove excess air bubbles
(de-gas). The syringe is then placed onto a screw driven syringe
pump with a needle of 14 to 40 gauge needle. Alternatively, the
process can be accomplished on a larger scale and, optionally,
automated, using the exemplary process illustrated in FIG. 2 and
described in the Examples. According to this embodiment, an
extruder (2) is used to deliver a controlled amount of collagen
slurry at a controlled rate of speed to a first dehydrating bath
(4) (e.g., an ammonia bath) and, optionally, a second dehydrating
bath (6). In a preferred embodiment, the slurry is extruded into an
ammonia bath and/or an ammonia/acetone bath at a rate of speed
sufficient to form a collagen fiber in the dehydrating bath.
Preferably, the process is performed in an environment which
includes an exhaust system (8) to remove noxious vapors that may
originate from the dehydrating baths. In the preferred process, a
draw frame (14) and a take-up element (12) provide the mechanism by
which the emerging collagen fiber is drawn through the process
steps. Optionally, the process further includes a dryer (14) to
provide a dried collagen fiber of the invention. Further examples
of a process in which a collagen dispersion is extruded into a
dehydrating bath or a coagulating bath to form a collagen fiber or
ribbon are shown in U.S. Pat. No. 3,114,593, entitled "Method of
Producing a Collagen Strand", issued to Griset; U.S. Pat. No.
3,114,591, entitled "Process for the Manufacture of Suture Material
from Animal Tendons", issued to Nichols et al.; U.S. Pat. No.
2,920,000, entitled "Collagen Article and the Manufacture Thereof",
issued to Hochstadt et al.; U.S. Pat. No. 2,637,321, entitled
"Shaped Article and Method of Producing It", issued to Cresswell;
and U.S. Pat. No. 2,598,608, entitled "Preparation of Collagenous
Materials", issued to Salo et al.
[0021] The process of the invention avoids dissolving the collagen
in acid solution and minimizes or avoids exposing the collagen to
other process steps which could denature the collagen and, thereby,
adversely affect its hemostatic activity. In the preferred
embodiments, the collagen is microfibrillar collagen; more
preferably, a collagen flour such as Avitene.RTM. flour.
Accordingly, in certain embodiments, the fibrils of the hemostatic
devices of the invention have a hemostatic activity that is about
the same as the hemostatic activity of Avitene.RTM. flour.
Avitene.RTM. flour is a microfibrillar collagen hemostat that is
indicated for all surgical specialties, including neurosurgery,
vascular, orthopaedic, urologic, and other general procedures.
Avitene.RTM. is available from Davol, Inc. (product numbers 101001,
101002, 101003, 101004, and 101034, Cranston, R.I.). The process
for preparing Avitene.RTM. flour is described in U.S. Pat. No.
3,742,955, issued to Battista et al.
[0022] As used herein, "hemostatic activity" refers to the ability
to stop bleeding and can be determined, e.g., in animal models that
are recognized as predictive of an in vivo effect by those of
ordinary skill in the art. Exemplary hemostasis animal models
include the pig and dog spleen animal models. A preferred animal
model for hemostasis activity is provided in the Examples.
[0023] Preferably the method for preparing the collagen fibers of
the invention further includes the step of (c) introducing the
collagen fiber into a second dehydrating bath. The first
dehydrating bath and the second dehydrating bath (if present) are
independently selected from the group consisting of: (1) an ammonia
bath comprising from about 10% to about 30% ammonia in water; (2)
an ammonia/acetone bath comprising from about 50% to about 70%
ammonia in acetone; (3) an acetone bath; (4) an ethanol bath; (5)
an isopropanol bath (containing about 70% isopropanol in water);
and (6) a propylene glycol bath (containing about 30% to about 95%
propylene glycol in water). Preferably, the first dehydrating bath
is an ammonia bath and the second dehydrating bath is an acetone
bath.
[0024] In certain embodiments, the fiber-forming processes
described herein, optionally, further include the step of drying
the collagen fiber. The collagen fibers, in wet or dry state, can
be formed into hemostatic fabrics using conventional
fiber-processing technology (e.g., weaving, knitting).
[0025] According to another aspect of the invention, the processes
for forming the hemostatic devices of the invention further
includes the step of introducing a hemostatic agent into the fiber
or fabric. The hemostatic agent can be introduced into the collagen
fiber or hemostatic fabric of the invention at any stage in the
process, including before the fiber-formation step (e.g., by adding
the hemostatic agent to the slurry) and after fabric formation step
(e.g., by soaking the hemostatic fabric in a solution containing
one or more hemostatic agents).
[0026] It is believed that the hemostatic fabrics of the invention
do not require a hemostatic agent to function effectively to
control bleeding, e.g., hemorrhage of a parenchymal organ. As a
result, the hemostatic devices of the invention which do not
further contain a hemostatic agent have good thermal stability and
can be stored for months to a few years without refrigeration and
losing effectiveness. Such embodiments of the invention are useful
for various medical situations and are particularly useful for
field and emergency use, since each may be stored in a ready-to-use
state for a lengthy period, even in the absence of refrigeration.
Such devices of the invention also are less expensive to make
and/or use compared to hemostatic devices which contain a further
hemostatic agent to achieve a comparable level of hemostatic
activity.
[0027] One advantage of the hemostatic fabric of the invention is
its flexibility compared to hemostatic devices such as
Gelfoam.RTM., that is, the hemostatic fabrics of the invention can
be provided in a form that easily conforms to the contours of an
organ or biological surface, making the manipulation of applying
the foam quicker to perform. As a result, there is less overall
blood loss to the patient and less time is spent in surgery.
Further, the hemostatic fabrics of the invention can be applied, in
wet or dry state, to a bleeding site and do not require pre-wetting
with a sterile solution prior to use.
[0028] The collagen fibers of the invention are formed of an
absorbable collagen from any source, e.g., corium collagen, tendon
collagen, and preferably is a microfibrillar collagen. More
preferably, the fiber preferably is formed of a collagen flour,
such as Avitene.RTM. flour. The fibers can be fabricated into
hemostatic fabrics having predictable hemostatic activities, based
on the activities of the collagen fibers of the invention. The
effectiveness of fabrics of the present invention in promoting clot
formation is further enhanced by their lattice structures, which
are selected to be of a sufficient weave size to promote enzyme
substrate interactions. In particular, the weave size and structure
of the hemostatic fabrics of the invention are selected to enhance
contact between thrombin that, optionally, is provided exogenously
in the fabric with endogenous fibrinogen present in the blood
exuding from a wound or lesion of, e.g., a parenchymal organ, a
spine or a brain.
[0029] In certain embodiments, at least one hemostatic agent can be
included in the hemostatic devices of the invention. Because
certain combinations of hemostatic agents can act synergistically,
the amount of each hemostatic agent can be less than that which
would be required to improve the hemostatic activity of the
hemostatic devices of the invention if the agents were used
individually. Accordingly, the collective amount of the hemostatic
agent(s) which are included in the collagen fiber or hemostatic
fabric of the invention is a "hemostasis-promoting amount", i.e.,
the amount of at least one hemostatic agent effective to accelerate
clot formation at an interface between a surface (e.g., of a wound,
of a lesion on a parenchymal organ, the spine or the brain) and the
hemostatic devices of the invention.
[0030] Exemplary hemostatic agents that can be applied to the
hemostatic devices of the invention in amounts effective for
stimulating hemostasis, include, but are not limited to: thrombin,
an enzyme which converts fibrinogen to fibrin; calcium, sodium,
magnesium or other ions that stimulate hemostasis; protamine
sulfate, an epsilon amino caproic acid, fibrinogen, and chitin.
Epsilon amino caproic acid and its analogs which possess a similar
chemical structure and hemostatic activity for use in a hemostatic
device are described in U.S. Pat. No. 5,645,849, assigned to
Clarion Pharmaceuticals. In terms of ion additives, calcium
chloride is generally a preferred additive for introducing a
calcium ion into the foam.
[0031] Thrombin is an active ingredient found in other hemostatic
devices. It is believed that, in general, the collagen fibrils of
the hemostatic devices of the invention have a hemostatic activity
that is equivalent to the hemostatic activity of the collagen
fibrils from which the fiber is formed. Thus, the invention
(without thrombin) advantageously provides a device having enhanced
hemostatic activity compared to the hemostatic devices of the prior
art. A further increase in the hemostatic activity of the
hemostatic devices of the invention can be achieved by, optionally,
including a hemostatic agent in the hemostatic devices of the
invention.
[0032] As used herein, the term "equivalent" with respect to
hemostatic activity means that the hemostatic activity is
substantially the same when measured in the same activity assay. An
exemplary hemostatic activity assay, a pig spleen hemostasis assay,
is provided in the examples. The assay can be used to measure the
hemostatic activity of the devices of the invention and can also be
used to measure the hemostatic activity of the collagen particles,
e.g., collagen flour, from which the hemostatic device is formed
by, for example, by placing powder over the incision, overlaying
the powder with a sterile gauze, and applying pressure to the wound
in the same manner as described in the example for a device of the
invention. The experimental results for the pig spleen assay are
reported in terms of the number of tamponades necessary to achieve
hemostasis at an incision in the pig spleen. The number of
tamponades for multiple samples is determined to obtain a
distribution of the number of tamponades. The distribution of
tamponades is a measure of the hemostatic activity for the device
or flour that is being tested. Accordingly, devices which have a
similar distribution of tamponades have "equivalent" hemostatic
activity. For example, if 80 of 100 samples of a first device
require one tamponade to achieve hemostasis, and 70 of 100 samples
of a second device require one tamponade to achieve hemostasis, the
hemostatic activity of the second device is considered to be within
10% of the hemostatic activity of the first device. Equivalent
hemostatic activity means that the hemostatic activity for two
samples are within at least 50%, more preferably, within 60%, 70%,
80%, 90% and, most preferably, within 95%.
[0033] The preferred hemostatic agent is thrombin (e.g., human or
bovine thrombin). Preferably, the thrombin is a recombinant
thrombin to avoid viral or other contamination from the organism
from which the thrombin is derived. The molecules "thrombin" and
"fibrinogen", as defined herein, are meant to include natural
thrombin and fibrinogen molecules derived from an animal or human
origin, a synthetic form or a recombinant form of the molecules,
including functionally active analogs that effectively maintain the
enzyme's clot promoting activity in an animal or human. The species
of animal from which the molecule is derived can vary and depends
on the intended use of the foam. For example, a foam intended for
human use for safety reasons preferably contains recombinant human
thrombin or non-human thrombin, e.g, bovine thrombin. By avoiding
use of human fibrinogen isolated from a human tissue or using viral
deactivated human thrombin, risks associated with viral
contamination of purified blood products are minimized.
[0034] Additionally or alternatively, the tripeptide RGD, composed
of arginine, glycine and aspartic acid, and optionally serine
"RGDS," can be incorporated into the hemostatic devices of the
invention as a hemostatic agent. RGD is the active site of
fibrinogen and fibronectin. RGD accelerates wound healing and is
believed to stimulate fibroblast migration The RGD additive is also
much less expensive than fibrinogen because it can be synthesized
using solid phase chemistry.
[0035] Thrombin-containing hemostatic devices of the inventions can
be prepared in a variety of ways to result in fibers and fabrics in
which the thrombin is dispersed within the hemostatic device or
applied to a surface of the hemostatic device in an amount
effective for inhibiting fibrinolysis and, thereby, stimulating
clot formation. Thus, according to one embodiment, a hemostatic
device of the invention is made by applying to the hemostatic
device, an amount of thrombin effective for promoting (stimulating)
hemostasis.
[0036] The hemostatic agent can be introduced into the hemostatic
device of the invention at any stage in the process, including
before the device-formation step (e.g., by adding the hemostatic
agent to the slurry) and after device formation step (e.g., by
soaking the hemostatic device in a solution containing one or more
hemostatic agents). Thrombin and/or other hemostatic agents or
additives described as components of a hemostatic device according
to the invention, can be applied to the hemostatic device by any of
several methods which all would be performed most advantageously
under sterile conditions. Thrombin can be introduced into the
collagen slurry prior to extrusion or applied as a layer to a
particular surface or side of a hemostatic device of the invention,
which surface is then designated as the wound-contacting surface.
For example, this can be accomplished by spraying thrombin in
powder form onto a hemostatic device of the invention.
Alternatively, a solution of thrombin can be coated onto a
hemostatic device of the invention and dried by lyophilization or
by conventional means. In another method of applying thrombin, a
hemostatic device of the invention is dipped completely or
partially into a sterile solution of thrombin such that a
sufficient amount of thrombin accumulates within the hemostatic
device effective to inhibit fibrinolysis in a mammal. Preferably,
the thrombin solution contains 1000 I/U of thrombin dissolved in 1
ml saline. The amount of thrombin applied in the solution can vary.
Preferably, the total amount of thrombin applied to a hemostatic
device of the invention or surface thereof is 100-1000
units/cm.sup.3. It is understood that alternative methods of
applying the hemostatic agents and additives to a hemostatic device
of the invention in addition to the methods described herein also
can be used.
[0037] The collagen fiber or hemostatic fabrics of the invention
that have been soaked in thrombin solution or other solution
containing a hemostatic agent optionally can be dried. The drying
step can be accomplished by lyophilization, preferably. Other
drying procedures appropriate for a material containing an active
protein ingredient can also be employed, so long as the drying
treatment does not denature the proteins or render them inactive.
Alternatively, the fiber or fabric can be dried by maintaining it
at room temperature for a period of 1-3 hours, followed by
refrigeration overnight.
[0038] In yet other embodiments, hemostatic agents other than, or
in addition to, thrombin can be incorporated, partially or fully,
into the hemostatic devices of the invention. For example,
protamine sulfate can be added to the hemostatic devices of the
invention in an amount that is effective to neutralize heparin in
the local environment of the device. Protamine sulfate can be added
in an amount between about 1-15 mg/cm.sup.2 of the hemostatic
device, preferably in an amount between 2-5 mg/cm.sup.2 of a wound
contacting surface of the hemostatic device.
[0039] Likewise, RGD or RGDS peptide can be dissolved in double
distilled water and sprayed onto a wound-contacting surface of a
hemostatic device of the invention. Preferably, such embodiments of
the invention contain an amount of RGD effective to enhance clot
formation. For example, RGD or RGDS can be applied to a hemostatic
device of the invention in an amount between about 110-130
mg/cm.sup.2. Thus, a standard size hemostatic device that is a
fabric would contain about 1-10 mg/fabric or about 5-7 mg/fabric of
RGD or RGDS.
[0040] The hemostatic agents described herein can be applied to a
fabric as a layer, for example, by spraying them onto the
wound-contacting surface of the fabric in dry forms. Alternatively,
the hemostatic fabrics of the inventions of the invention can be
soaked in a solution containing the hemostatic agent. Accordingly,
the invention embraces hemostatic fabrics of the inventions in
which the hemostatic agent(s) are absorbed into the interstices of
the fabric, as well as fabrics in which such agents are layered on
a surface of the fabric. In certain embodiments, the hemostatic
agents and additives are coated on only one surface (the
wound-contacting surface) to minimize the likelihood of hemostasis
between the wound and a non-wounded tissue in the vicinity of the
fabric. In yet other embodiments intended for packing a void in
body tissue, the fabric can be coated with hemostatic agent(s)on
all surfaces.
[0041] In certain embodiments, the hemostatic devices of the
invention further include a therapeutically effective amount of at
least one therapeutic agent, such as agents which promote
wound-healing and or reduce pain (e.g., vascular pain). Agents
which promote wound-healing and/or reduce pain include
anti-inflammatory agents (steroidal and non-steroidal) such as
agents which inhibit leukocyte migration into the area of surgical
injury, anti-histamines; agents which inhibit free radical
formation; and bacteriostatic or bacteriocidal agents. In general,
a therapeutically effective amount means that amount necessary to
delay the onset of, inhibit the progression of, or halt altogether
the particular condition being treated. Generally, a
therapeutically effective amount will vary with the subject's age,
condition, and sex, as well as the nature and extent of the
condition in the subject, all of which can be determined by one of
ordinary skill in the art. The dosage of therapeutic agent
contained in the hemostatic devices of the invention may be
adjusted to accommodate the particular subject and condition being
treated.
[0042] As used herein, the phrase, "agents which promote
wound-healing" refers to agents, the administration of which,
promote the natural healing process of a wound. Agents that promote
wound-healing include anti-inflammatory agents, agents which
inhibit free radical formation, and bacteriostatic or bacteriocidal
agents.
[0043] Anti-inflammatory agents are agents which inhibit or prevent
an immune response in vivo and include: (i) agents which inhibit
leukocyte migration into the area of surgical injury ("leukocyte
migration preventing agents"), and anti-histamines. Representative
leukocyte migration preventing agents include silver sulfadiazine,
acetylsalicylic acid, indomethacin, and Nafazatrom. Representative
anti-histamines include pyrilamine, chlorpheniramine,
tetrahydrozoline, antazoline, and other anti-inflammatories such as
cortisone, hydrocortisone, beta-methasone, dexamethasone,
fluocortolone, prednisolone, triamcinolone, indomethacin, sulindac,
its salts and its corresponding sulfide, and the like.
[0044] Representative agents which inhibit free radical formation
include antioxidants that inhibit the formation and/or action of
oxide products, superoxide dismutase (SOD), catalase, glutathione
peroxidase, b-carotene, ascorbic acid, transferrin, ferritin,
ceruloplasmin, and desferrioxamine .alpha.-tocophenol.
[0045] Representative bacteriostatic or bacteriocidal agents
include antibacterial substances such as .beta.-lactam antibiotics,
such as cefoxitin, n-formamidoyl thienamycin and other thienamycin
derivatives, tetracyclines, chloramphenicol, neomycin, gramicidin,
bacitracin, sulfonamides; aminoglycoside antibiotics such as
gentarnycin, kanamycin, amikacin, sisomicin and tobramycin;
nalidixic acids and analogs such as norfloxican and the
antimicrobial combination of fluoroalanine/pentizidon- e;
nitrofurazones, and the like.
[0046] The hemostatic devices of the invention can contain one or
more therapeutic agents, alone or in combination with one or more
hemostatic agents.
[0047] Various additives, optionally, can be incorporated into the
hemostatic devices of the invention without adversely affecting the
hemostatic activity of these devices. The term
"pharmaceutically-acceptab- le carrier" as used herein means one or
more compatible solid or liquid fillers, diluents or encapsulating
substances which are suitable for administration into a human. The
term "carrier" denotes an organic or inorganic ingredient, natural
or synthetic, with which the active ingredient is combined to
facilitate the application. The components of the pharmaceutical
compositions also are capable of being co-mingled with the collagen
fibrils of the present invention, and with each other, in a manner
such that there is no interaction which would substantially impair
the desired hemostatic activity.
[0048] According to certain embodiments, a hemostatic device of the
invention is contained within a sealed sterile package which
facilitates removal of the hemostatic device without contamination.
Such a package, for example, can be an aluminum foil pouch or other
material that is easily sterilized. Radiation, e.g., gamma
radiation, can be applied to sterilize the hemostatic device and
packaging material together. In yet other embodiments, a container
having dual compartments is provided in which a first compartment
contains distilled water, sterile saline or a sterile buffer, and a
second compartment contains a hemostatic device of the invention.
The hemostatic device of the second compartment can be readily
dipped into an opened first compartment and subsequently applied to
the wound.
[0049] According to yet another aspect of the invention, a product
prepared by the above-described process is provided. A particular
embodiment of this process in provided in the Examples. The
process, optionally, further includes the step of cross linking the
collagen within the hemostatic devices of the invention, e.g., by
heating the collagen fibers of the invention at a temperature and
for a period of time sufficient to form crosslinks without
adversely affecting the hemostatic activity of the collagen
fiber.
[0050] According to still another aspect of the invention, a method
for promoting hemostasis is provided. The method involves the steps
of pressing a hemostatic fabric of the invention against a surface
of a wound or a surface of a lesion on an organ, tissue, or other
bleeding surface, e.g., a parenchymal organ, the spine or the
brain, for a period of time until clotting has occurred at the
interface between the hemostatic fabric of the invention and the
surface. The fabric may be applied to the surface in a dry state
or, alternatively, may be soaked in sterile saline solution or a
sterile hemostatic agent-containing solution prior to use. Use of a
hemostatic fabric of the invention according to the invention,
without first soaking in saline solution permits quick and simple
application of the fabric in various situations, including field
situations such as may be encountered by an emergency medical
technician. In certain embodiments, the hemostatic fabric is soaked
in a thrombin solution prior to use to introduce a therapeutically
effective amount of thrombin into the sponge. Thus, a collagen
fabric of the invention of the invention can be used by applying a
"wound-contacting" surface of the fabric, a surface intended to
contact the wound and containing hemostatic agent(s) and,
optionally, additives, with or without prior soaking in a sterile
solution, to a surface of a bleeding wound or lesion. Then, the
fabric is maintained in contact with the surface for a period of
time sufficient for clotting to occur at the interface between the
hemostatic fabric of the invention and the surface and for bleeding
to be substantially arrested. Preferably, the fabric is maintained
in contact with the surface for a period of about 3-20 minutes,
advantageously 3-10 minutes, and more advantageously, 3-5
minutes.
[0051] Where thrombin and/or other hemostatic agents also are
present on/in the hemostatic fabric, the time period is preferably
about 5 minutes. The hemostatic fabric is held in place against the
biological surface, preferably with light pressure, e.g., by means
of a sterile saline soaked sponge. Alternatively, the hemostatic
fabric may be held in place simply by applying pressure to the
hemostatic fabric by means of a gauze or other dry sterile
material. Depending on the location of the wound, a bandage can be
wrapped around the hemostatic fabric to provide light pressure on
the wound surface.
[0052] The efficacy of the hemostatic fabrics of the invention can
be assessed in art-recognized animal models that are believed to be
predictive of an in vivo hemostatic effect in humans. For example,
surgical lesions induced in parenchymal organs of pigs provide a
good model system for hemostasis in the analogous human organs as
evidenced by preclinical studies which employ pig models. See e.g.,
SWINE AS MODELS IN BIOMEDICAL RESEARCH, Swindle, M., Iowa State
Univ. Press (1992).
[0053] A preferred use of a hemostatic fabric according to the
present invention is to inhibit or completely stop bleeding of a
parenchymal organ, such as the liver, kidney, spleen, pancreas or
lungs. Other preferred uses are to inhibit or completely stop
bleeding of a wound or lesion on the spine or brain. Additional
uses for the hemostatic fabrics of the invention include inhibiting
bleeding during surgery, e.g., internal/abdominal, vascular
(particularly for anastomosis), urological, gynecological,
thyroidal, neurological, tissue transplant uses, dental,
cardiovascular, cardiothoracic, ENT (ear, nose, throat), and
orthopedic surgeries.
[0054] Another use of a hemostatic fabrics of the invention is
topical treatment, such as for burn or tissue transplants or dura
replacement and/or substitution. A hemostatic fabric of the
invention for topical use preferably contains additives, such as
anti-infection medicaments, bactericides, flimgicides and wound
healing agents, for example, neomycin and bacitracin.
[0055] In addition to inducing hemostasis, the hemostatic fabrics
of the inventions of the invention can be used to hermetically
sealing body tissue. For example, when air leaks from a wound in
the lungs, a hemostatic fabric of the invention can be applied to
the surface surrounding the wound, held in place for a period of
time sufficient to induce hemostasis and allow a hermetic seal to
form.
[0056] The hemostatic devices of the invention also are useful for
treating animals, preferably humans or other mammals, including
domestic mammals and livestock.
[0057] The hemostatic fabrics of the invention can be provided in a
variety of sizes and shapes, depending upon its intended use.
Typically, the hemostatic fabrics of the invention are provided in
a standard size rectangular fabric, e.g., 2".times.14";
1".times.1"; 4".times.8"; 1".times.3.5"; 2".times.3"; 3".times.4";
0.5".times.2"; and 6".times.9". The hemostatic fabrics may be cut
to size with a pair of scissors. The hemostatic fabrics of the
invention may be folded, bundled, wrapped, or prefabricated into
small squares, such as for packing into a body cavity, such as a
dental cavity following a tooth extraction. The collagen fibers of
the invention may be knitted into various structures.
Alternatively, the hemostatic fabric can be shaped for epistaxis
(profusely bleeding nostril) or insertion into a cavity. The
hemostatic fabrics of the invention that are intended for topical
applications can be applied with an adhesive tape, as a band-aid
form, where the hemostatic fabric is adhered to an adhesive
backing. One or more additional layers of wound dressing material,
preferably a layer which aids in absorption of blood or other
exudants, can be applied to or incorporated into the hemostatic
fabrics of the invention to form a stronger bandage. Alternatively,
the layer may be applied as a supplement to the backside (non-wound
contacting surface) of a fabric according to the invention.
Particularly for topical use, the layer(s) can contain
superabsorbents to wick exudant solution from the wound site. For
hemostatic fabrics of the inventions intended for internal-surgical
applications, where an added layer(s) is integral with the fabric,
the layer(s) should be both biodegradable and pharmaceutically
acceptable.
[0058] The hemostatic fabrics of the invention can designed to
facilitate its application to fuse ends of a blood vessel or other
body lumen having been severed, e.g., surgically. To apply a
hemostatic fabric for anastomosis, a rectangular fabric, for
example, is wrapped around the external surface of the ends of a
Dacron.RTM. graft and the graft is positioned into place. The
hemostatic fabric portion of the graft accelerates fibrin growth
into the graft to seal the graft in place (hemostatically and
hermetically). According to certain embodiments of the invention, a
kit is provided for this application. The kit contains a graft and
a hemostatic fabric of the invention that is designed for fitting
with the ends of the graft. Alternatively, a kit is provided having
a hemostatic fabric of the invention pre-fitted onto at least one
end of a graft.
[0059] According to still other aspects of the invention, various
specialized kits can be provided. The kits contain any of the
hemostatic device embodiments disclosed herein and a package,
wherein the hemostatic device of the invention is contained within
a sealed sterile package which facilitates removal of the fabric
without contamination. The kit can contain multiple hemostatic
devices of the inventions, preferably wherein each hemostatic
device is contained within a separate sealed sterile package. A kit
that is designed for autonomous use, e.g., for field/military use
can, in addition to a hemostatic fabric of the invention, further
include disposable pre-sterilized surgical instruments and/or
agents that can be incorporated into the fabric, e.g., thrombin,
calcium chloride.
[0060] Unless defined otherwise, all technical and scientific terms
used herein have the same meaning as commonly understood by one of
ordinary skill in the art of the invention. Although any methods
and materials similar or equivalent to those described herein can
be used in the practice of the invention, the preferred methods and
materials have been described. Unless mentioned otherwise, the
techniques employed or contemplated herein are standard
methodologies well known to one of ordinary skill in the art. The
materials, methods and examples are illustrative only and not
limiting.
EXAMPLES
Example 1
Preparation of a Collagen Fiber from a Collagen Slurry
[0061] Avitene.RTM. flour (microfibrillar collagen) is mixed with
water at about a 2 to 10% mixture (by weight to volume) with
optimum being 5% and allowed to swell in water for a 4 to 72 hour
period. Avitene.RTM. flour is available from Davol, Inc. (product
numbers 101001, 101002, 101003, 101004, and 101034, Cranston,
R.I.). The "slurry" (collagen swelled by water) is filled into a
syringe of 5 to 60 cc. A stopper is placed on the luer-lock end and
the syringe is placed into a centrifuge. The "slurry" is
centrifuged to remove excess air bubbles (de-gas). The syringe is
then placed onto a screw driven syringe pump with a needle of 14 to
40 gauge needle. The slurry is extruded into an ammonia bath and/or
an ammonia/acetone bath. In one embodiment, the bath is sixteen
feet long to mildly dehydrate the fiber. The fiber is then further
dehydrated in an acetone bath before being dried in a dynamic
drying oven. The fiber is wound onto a spool in accordance with
conventional procedures, using conventional equipment and further
processed into a knitted or woven fabric. The fiber can be coated
with a 20% glycerin and ethanol coating solution to aid the fibers
in further processing (knitting or weaving).
[0062] Optionally, the "slurry" can be homogenized and/or filtered
to remove any fibrils or impurities that may cause defects in the
fibers. The fibers can be either mono- or multi filament.
[0063] The collagen fibers of the invention are processed into a
hemostatic fabric of the invention in accordance with standard
procedures for forming a fabric, e.g., quilting or weaving. (See,
e.g., U.S. Pat. Nos. 3,114,593; 3,114,591; 2,920,000; 2,637,321;
2,598,608 (noted above), as well as U.S. Pat. No. 5,378,469,
entitled "Collagen Threads"; U.S. Pat. No. 5,256,418, entitled
"Collagen Constructs"; PCT/US92/08520, entitled "Collagen
Constructs" (publication no. WO 93/06791, priority claimed to U.S.
Ser. No. 772,529, now U.S. Pat. No. 5,378,469); PCT/US95/03455,
entitled "Three-dimensional Bioremodelable Collagen Fabrics"
(publication no. WO 95/25482, priority claimed to U.S. Ser. No.
08/215,760); and PCT/US95/03525, entitled "Biocompatible Prosthetic
Devices" (publication no. WO 95/25550, priority claimed to U.S.
Ser. No. 08/216,527).)
[0064] The hemostatic fabrics of the invention are intended for use
as an adjunct for hemostasis. The process for forming Avitene flour
involves swelling collagen in acidified alcohol, but does not
involve acid dissolution of collagen. It is believed that the
avoidance of acid dissolution in the Avitene.RTM. flour fabrication
process is responsible for its enhanced hemostatic activity.
Because the process for forming the Avitene.RTM. flour into fibers
and then interlacing the fibers into a fabric also does not involve
contacting the collagen with acid or exposing the collagen to other
denaturing conditions, it is believed that the hemostatic fabrics
of the invention exhibit enhanced hemostatic properties compared to
known collagen fabrics.
Example 2
AUTOMATED PROCESS FOR PREPARATION OF A COLLAGEN FIBER
[0065] The following procedure is contemplated for automating the
process for preparation of a collagen fiber. The procedure
illustrative and is not intended to limit the invention in any
way.
[0066] Mixing/Centrifuge of Slurry:
[0067] The collagen slurry is mixed at a concentration of about 3
to 10% w/v of Avitene Powder to water. The mixture is passed
through a continual operation of a Horizontal Shaft Model D-8
Stainless Steel Versator (The Cornell Machine company, Springfield,
N.J.), thereby eliminating the need to manual mix the slurry. The
slurry can immediately be fed into the extruder for extrusion or it
can be stored up to 72 hours before being extruded.
[0068] Extruder:
[0069] The collagen slurry is fed into the extruder feed throat and
extruded at room temperature to about 50.degree. C. through a 3/4"
diameter barrel with a 25:1 L/D ratio, air cooled extruder with a
2:1 compression ratio screw (C.W. Brabender Instrument, Inc.,
Hackensack, N.J.). The slurry passes through the barrel into the
block with a metering pump assembly (0.6 to 1.2 ml/min.), which
forces the slurry through the die assembly at a constant rate. The
slurry can be extruded through a single hole die to a multifilament
die of 30 holes.
[0070] Troughs (Dehydration Baths):
[0071] The slurry exits the die and enters the first dehydration
bath (10 to 15 feet long.times.5 inches wide.times.5 inches deep,
stainless steel) containing the first dehydration agent (preferably
ammonia). The fiber(s) then passes into the second dehydration bath
(5 feet long.times.5 inches wide.times.5 inches deep, stainless
steel) containing the second dehydrating agent (preferably
acetone). The baths are from C.W. Brabender Instruments, Inc.,
Hackensack, N.J.
[0072] Draw/Drying Frame:
[0073] The fiber(s) are wound through a draw frame (Dienes
Laboratory Draw Winder--Single Position, Comoli DWI-2000, Dienes
Apparatus, Inc., Pineville, N.C.) containing heated rollers to aid
in the drying of the fiber while it is being drawn to specification
of mechanical strength. The fiber can then be further dried (if
necessary) by passing by a "hot gun" (blow dryer).
[0074] Winder:
[0075] The fiber(s) are then collected onto a spool (preferably
perforated stainless steel), which is being spun by a winder
(Leesona Style 50 Winder, Standard Mill Machinery Corp., West
Warwick, R.I.).
[0076] Knitter:
[0077] The spool(s) of fiber can then be twisted together to form a
multifilament or a single filament end can be used to form a fabric
using a knitting machine (Circular Weft Knitting Machine, Lamb
Knitting Machine Corp., Chicopee, Mass.).
Example 3
PROTOCOL FOR COMPARISON BETWEEN A HEMOSTATIC FABRIC OF THE
INVENTION AND SURGICEL.RTM.
[0078] The following procedure is contemplated for comparing the
hemostatic activity of a fabric of the invention and a
representative, commercially available fabric. The procedure
illustrative and is not intended to limit the invention in any
way.
[0079] Hemostatic Activity Assay
[0080] The hemostatic fabrics of the invention contain Avitene.RTM.
flour and water. The hemostatic response time of a sample
hemostatic fabric of the invention ("TEST SAMPLE") with and without
thrombin, is compared to Surgicel.RTM. (Johnson & Johnson
Medical Inc., Arlington, Tex., with and without thrombin) in a pig
spleen model (J&J Hemostasis protocol) as described below.
Small incisions are made in the retracted spleen of anesthetized
juvenile Yorkshire pigs. The number of cuts per spleen ranges from
8 to 18. Eight pigs are required. Thrombin is added by soaking the
sample (TEST SAMPLE or Surgicel.RTM. sample) in a thrombin solution
until fully saturated. The test article (approximately
0.5".times.0.5") is placed on the wound, tamponaded with finger
pressure for 20 seconds, then the pressure is removed and the site
is observed for re-bleed for two minutes. If re-bleed is observed
within two minutes, pressure is reapplied for 20 seconds and the
cycle is repeated. The endpoint is the number of tamponades to
achieve no re-bleed. The following samples are paired during
testing (20 pairs each): TEST SAMPLE versus Surgicel.RTM., TEST
SAMPLE versus -Surgicel.RTM. thrombin, TEST SAMPLE-thrombin versus
Surgicel.RTM. -thrombin. A pair is defined as two samples tested
one after the other and adjacent to one another on the spleen. For
each pair, the first sample tested is alternated from pair to pair.
Each pair is tested at least once, usually twice, and sometimes 3
times on each animal to better characterize animal to animal
variability.
[0081] The frequency of the number of tamponades for each product
type within the paired group, is analyzed using the Fisher's exact
test and The Stuart-Maxwell test (both one-tailed) at alpha 0.05.
These paired groups are analyzed separately. Therefore, a one-sided
test based on expected results is appropriate.
[0082] It is expected that TEST SAMPLE without thrombin would need
fewer tamponades than Surgicel.RTM. with or without thrombin
because the collagen in the TEST SAMPLE has not been subjected to
acid dissolution or exposed to other denaturing conditions.
Accordingly, it is believed that clinical users may choose to use
the hemostatic fabrics of the invention in the dry state without
thrombin, saving time and money, since only one product would be
used instead of two. The soft and flexible handling characteristics
of the hemostatic fabrics of the invention will allow it to be used
in the dry state.
[0083] All of the references, patents and patent publications
identified or cited herein are incorporated, in their entirety, by
reference.
[0084] Although this invention has been described with respect to
specific embodiments, the details of these embodiments are not to
be construed as limitations. Various equivalents, changes and
modifications may be made without departing from the spirit and
scope of this invention, and it is understood that such equivalent
embodiments are part of this invention.
* * * * *