U.S. patent application number 13/392623 was filed with the patent office on 2012-07-26 for improvement to trabeculectomy.
This patent application is currently assigned to THROMBOGENICS NV. Invention is credited to Ingeborg Stalmans, Jean-Marie Stassen.
Application Number | 20120189609 13/392623 |
Document ID | / |
Family ID | 41434741 |
Filed Date | 2012-07-26 |
United States Patent
Application |
20120189609 |
Kind Code |
A1 |
Stassen; Jean-Marie ; et
al. |
July 26, 2012 |
IMPROVEMENT TO TRABECULECTOMY
Abstract
The current invention relates to the improvement of
trabeculectomy surgery. The improvement more specifically resides
in an extended lifetime of the sclera-corneal drainage channel
created by trabeculectomy surgery. The improvement is obtained by
post-surgical administration of a plasmin or active derivative
thereof in the form of topical eye drops alone, by anterior chamber
injection alone, or by any combination of these.
Inventors: |
Stassen; Jean-Marie;
(Lubbeek, BE) ; Stalmans; Ingeborg; (Knokke,
BE) |
Assignee: |
THROMBOGENICS NV
B-3001 Heverlee
BE
|
Family ID: |
41434741 |
Appl. No.: |
13/392623 |
Filed: |
August 27, 2010 |
PCT Filed: |
August 27, 2010 |
PCT NO: |
PCT/EP2010/062584 |
371 Date: |
March 19, 2012 |
Related U.S. Patent Documents
|
|
|
|
|
|
Application
Number |
Filing Date |
Patent Number |
|
|
61237723 |
Aug 28, 2009 |
|
|
|
Current U.S.
Class: |
424/94.64 ;
435/217 |
Current CPC
Class: |
A61P 27/08 20180101;
A61P 27/02 20180101; A61K 38/484 20130101; A61K 9/0051 20130101;
A61K 45/06 20130101; A61P 31/12 20180101; A61P 23/02 20180101; A61K
47/12 20130101; A61K 9/0048 20130101; A61P 29/00 20180101 |
Class at
Publication: |
424/94.64 ;
435/217 |
International
Class: |
A61K 38/48 20060101
A61K038/48; A61P 31/12 20060101 A61P031/12; A61P 27/08 20060101
A61P027/08; A61P 27/02 20060101 A61P027/02; A61P 23/02 20060101
A61P023/02; C12N 9/68 20060101 C12N009/68; A61P 29/00 20060101
A61P029/00 |
Foreign Application Data
Date |
Code |
Application Number |
Aug 28, 2009 |
EP |
09168912.5 |
Claims
1. A plasmin or an active truncated variant thereof for treating
filtration failure after trabeculectomy surgery of an eye, or for
preventing, reducing or retarding the occurrence of filtration
failure after trabeculectomy surgery of an eye.
2. The plasmin or variant thereof according to claim 1 which is in
a pharmaceutically acceptable formulation capable of being
administered to an eye as topical eye drops.
3. The plasmin or variant thereof according to claim 1 which is in
a pharmaceutically acceptable formulation capable of being
administered by injection into the anterior chamber of an eye.
4. The plasmin or variant thereof according to claim 1 wherein said
treating of filtration failure after trabeculectomy surgery of an
eye, or said preventing, reducing or retarding of the occurrence of
filtration failure after trabeculectomy surgery of an eye results
from contacting said eye with an effective amount of topical eye
drops comprising said plasmin or variant thereof.
5. The plasmin or variant thereof according to claim 1 wherein said
treating of filtration failure after trabeculectomy surgery of an
eye, or said preventing, reducing or retarding of the occurrence of
filtration failure after trabeculectomy surgery of an eye results
from introduction into the anterior chamber of an eye of an
effective amount of said plasmin or variant thereof.
6. The plasmin or variant thereof according to claim 1 wherein said
treating of filtration failure after trabeculectomy surgery of an
eye, or said preventing, reducing or retarding of the occurrence of
filtration failure after trabeculectomy surgery of an eye results
from contacting said eye with an effective amount of said plasmin
or variant thereof via topical eye drops comprising said plasmin or
variant thereof, combined with introduction into the anterior
chamber of an eye of said plasmin or variant thereof.
7. The plasmin or variant thereof according to claim 1 wherein said
active truncated variant of plasmin is lacking one or more kringle
domains and/or lacking parts of one or more kringle domains.
8. The plasmin or variant thereof according to claim 1 wherein said
active truncated variant of plasmin is selected from the group
consisting of midiplasmin, miniplasmin, microplasmin or
deltaplasmin.
9. The plasmin or variant thereof according to claim 1 which is in
a pharmaceutically acceptable formulation further comprising one or
more of an agent for controlling the intra-ocular pressure, an
anti-inflammatory agent, an antiviral agent, an antibacterial
agent, an antiviral agent, an anti-angiogenic agent, an
anti-mitotic agent, an antihistamine, an anesthetic, an agent to
induce mydriasis and an agent to induce cycloplegia.
10. The plasmin or variant thereof according to claim 4 wherein
said eye is contacted further with one or more agents chosen from
an agent for controlling the intraocular pressure, an
anti-inflammatory agent, an antiviral agent, an antibacterial
agent, an antiviral agent, an anti-angiogenic agent, an
anti-mitotic agent, an antihistamine, an anesthetic, an agent to
induce mydriasis and an agent to induce cycloplegia.
Description
FIELD OF THE INVENTION
[0001] The current invention relates to the improvement of
trabeculectomy surgery. The improvement more specifically resides
in an extended lifetime of the sclera-corneal drainage channel
created by trabeculectomy surgery. The improvement is obtained by
post-surgical administration of a plasmin or active derivative
thereof in the form of topical eye drops alone, by anterior chamber
injection alone, or by any combination of these.
BACKGROUND OF THE INVENTION
[0002] Glaucoma is a multifactorial, neurodegenerative disease and
the second most important cause of irreversible blindness (Quigley,
1996, Br J Ophthalmol 80, 389-393). This disease is characterized
by progressive retinal ganglion cell apoptosis, resulting in visual
field loss. Current treatment of this disease is directed towards
the reduction of intraocular pressure (IOP), which is the main risk
factor for glaucoma (Collaborative Normal-Tension Glaucoma Study
Group, 1998, Am J Opthalmol 126, 487-497).
[0003] Of all currently used treatments to lower IOP, filtration
surgery (trabeculectomy) was shown to be the most effective (Burr
et al., 2005, Cochrane Database Syst Rev 18(2):CD004399; Hitchings,
1998, Arch Ophthalmol 116, 241-242). A trabeculectomy creates a
"controlled" leak of fluid (aqueous humor) from the eye, which
percolates under the conjunctiva. During the operation a piece of
trabecular meshwork in the drainage angle of the eye is removed,
creating an opening. The opening is partially covered with a flap
of tissue from the sclera and conjunctiva. A small conjunctival
"bleb" (bubble) appears at the junction of the cornea and the
sclera (limbus) where this surgically produced valve is made.
[0004] In 30% of the cases, however, the constructed channel closes
due to excessive scar tissue formation, resulting in surgical
failure (Addicks et al., 1983, Arch Ophthalmol 101, 795-798). The 4
important processes contributing to post-operative conjunctival
scarring are: clot formation, inflammation, angiogenesis and
fibrosis (Lee et al., 1995, J Ocul Pharmacol Ther 11, 227-232; Lama
& Fechtner, 2003, Sury Ophthalmol 48, 314-346). Indeed,
increased conjunctival infiltration of inflammatory cells and Tenon
fibroblasts (Hitchings & Grierson, 1983, Trans Ophthalmol Soc
UK 103, 84-88; Skuta & Parrish, 1987, Sury Ophthalmol 32,
149-170), and higher levels of bleb vascularisation (Jampel et al.,
1988, Arch Ophthalmol 106, 89-94) are associated with surgical
failure. These processes are mediated by various cytokines (e.g.
IL-1 and INF-.alpha.2b) and growth factors (e.g. PDGF, FGF,
TGF-.beta.1 and VEGF (Lama & Fechtner, 2003; Gillies & Su,
1991, Aust NZ J Ophthalmol 19, 299-304)). Peroperative
anti-mitotics, such as mitomycin-C and 5-Fluorouracyl can improve
surgical outcome (Quigley, 1996; Katz et al., 1995, Ophthalmol 102,
1263-1269). However, these antimetabolites carry a risk of
vision-threatening complications such as scleral thinning and
infections (Lama & Fechtner, 2003; Hitchings & Grierson,
1983; Skuta & Parrish, 1987; Jampel et al., 1988; Gillies &
Su, 1991; Katz et al., 1995; Greenfield et al., 1998, Arch
Ophthalmol 116, 443-447). Furthermore, blocking TGF-.beta. seemed
promising in animal models (Cordeiro et al., 2003, Gene Ther 10,
59-71), but was not efficient in a clinical study (CAT-152 0102
Trabeculectomy Study Group, Kwah, Grehn, 2007, Ophthalmol 114,
1822-1830). The number of post-trabeculectomy interventions
expressed as the incidence of post-surgery "bleb manipulations" was
reported to be as high as 78% (King et al., 2007, Br J Ophthalmol
91, 873-877). Therefore, there is still a need for alternative
strategies to prevent filtration failure and, thus, to reduce the
incidence of bleb manipulations.
[0005] Microplasmin is a recombinant protein that dissolves blood
clots by degrading fibrin. Recently, microplasmin has been shown to
be efficient, well tolerated and safe for intra-ocular use in a
phase II clinical trial to study its efficacy to induce
non-surgical posterior vitreous detachment, PVD (Gandorfer, 2008,
Eye 22, 1273-1277; WO 2004/052228) and is currently under
investigation in phase III clinical trials. Plasmin was previously
shown to be able to induce PVD as well (e.g. U.S. Pat. No.
5,304,118). The mechanism by which PVD is induced by plasmin or
microplasmin is currently not fully understood. Unsupported by any
or any conclusive experimental data, WO 2009/073457 and WO
2009/067407 propose subconjunctival plasmin injection for rescuing
filtering blebs and the use of matrix metalloproteinase activating
proteases for reducing IOP, respectively.
SUMMARY OF THE INVENTION
[0006] The invention relates to (the use of) a plasmin or an active
truncated variant thereof (for the manufacture of a medicament) for
treating filtration failure after trabeculectomy surgery of an eye,
or for preventing, reducing or retarding the occurrence of
filtration failure after trabeculectomy surgery of an eye.
[0007] Said plasmin or active truncated variant thereof, or said
medicament, may be in a pharmaceutically acceptable formulation
capable of being administered to an eye as topical eye drops.
Alternatively, said plasmin or active truncated variant thereof, or
said medicament, may be a in pharmaceutically acceptable
formulation capable of being administered by injection into the
anterior chamber of an eye.
[0008] The treatment of filtration failure after trabeculectomy
surgery of an eye, or the prevention, reduction or retardation of
the occurrence of filtration failure after trabeculectomy surgery
of an eye may result from contacting said eye with an effective
amount of topical eye drops comprising said plasmin or active
truncated variant thereof. Alternatively, it may result from
introduction into the anterior chamber of an eye of an effective
amount of said plasmin or active truncated variant thereof. In a
further alternative, it may result from contacting said eye with an
effective amount of topical eye drops comprising said plasmin or
active truncated variant thereof, combined with introduction into
the anterior chamber of an eye of an effective amount of said
plasmin or active truncated variant thereof.
[0009] In any of the above, said active truncated variant of
plasmin may be lacking one or more kringle domains and/or lacking
parts of one or more kringle domains. More specifically, said
active truncated variant of plasmin may be selected from the group
consisting of midiplasmin, miniplasmin, microplasmin or
deltaplasmin.
[0010] The invention further covers the (use of) a plasmin or an
active truncated variant thereof (for the manufacture of a
medicament) for treating filtration failure after trabeculectomy
surgery of an eye, or for preventing, reducing or retarding the
occurrence of filtration failure after trabeculectomy surgery of an
eye, wherein said plasmin or active truncated variant thereof, or
said medicament, may be in a pharmaceutically acceptable
formulation further comprising one or more of an agent for
controlling the intra-ocular pressure, an anti-inflammatory agent,
an antiviral agent, an antibacterial agent, an antiviral agent, an
anti-angiogenic agent, an anti-mitotic agent, an antihistamine, an
anesthetic, an agent to induce mydriasis and an agent to induce
cycloplegia. Alternatively, when said further agent(s) is(are) not
included in the pharmaceutically acceptable formulation, or in the
medicament, said eye may be contacted further with one or more
agents chosen from an agent for controlling the intra-ocular
pressure, an anti-inflammatory agent, an antiviral agent, an
antibacterial agent, an antiviral agent, an anti-angiogenic agent,
an anti-mitotic agent, an antihistamine, an anesthetic, an agent to
induce mydriasis and an agent to induce cycloplegia.
LEGENDS TO THE FIGURES
[0011] FIG. 1 shows the amino acid sequence with double numbering
of the amino acid positions of wild-type human Glu-plasminogen (1
to 791) and of the plasmin catalytic domain (1 to 230, amino acid
sequence and numbering in bold). Microplasminogen as used for
demonstrating the invention starts at amino acid position 543
(numbering relative to Glu-plasminogen). Kringle domains (as
derived from GenBank accession number AAA36451) are boxed and their
amino acid sequences typed alternating in normal and italic
letters. The catalytic triad amino acids are circled.
[0012] FIG. 2 schematically depicts an eye after trabeculectomy.
The arrow shaded with vertical lines indicates the flow of aqueous
liquid from the eye's anterior chamber to the outside through the
filtration channel created by trabeculectomy.
[0013] FIG. 3 depicts the results obtained after trabeculectomy
combined with (i) post-operative administration of topical drops
containing microplasmin (diamonds), (ii) post-operative anterior
chamber injection of microplasmin (triangles), or (iii) combined
post-operative administration of topical drops containing
microplasmin and anterior chamber injection of microplasmin
(squares). The data are normalized meaning that background values
obtained with placebo treatment have been deducted from the values
obtained with non-placebo treatment.
DETAILED DESCRIPTION OF THE INVENTION
[0014] The present invention is based on the effect of
administration of microplasmin on the clinical outcome of
trabeculectomy surgery, said effect being positive and resulting in
the prevention, reduction or retardation of the occurrence of
filtration failure. As known from clinical practice, each patient
that underwent trabeculectomy surgery is at significant risk to
develop filtration failure.
[0015] Therefore, the invention relates to (the use of) a plasmin
or an active truncated variant thereof (for the manufacture of a
medicament) for treating filtration failure after trabeculectomy
surgery of an eye, or for preventing, reducing or retarding the
occurrence of filtration failure after trabeculectomy surgery of an
eye.
[0016] "Plasmin", also known as fibrinolysin or lysofibrin, is a
serine-type protease which results from the activation of the
zymogen plasminogen. Activation is the result of a proteolytic
cleavage between amino acids 561 and 562 (numbering relative to
human Glu-plasminogen). Plasmin carries a heavy chain comprising 5
kringle domains and a light chain comprising the catalytic domain.
Plasminogen can be enriched from blood plasma, e.g., via lysine
affinity-chromatography (Deutsch & Mertz, 1970, Science 170,
1095-1096). Truncation of the plasmin molecule is possible as long
as the catalytic domain remains functional, such truncation thus
results in the formation of an "active truncated variant" of
plasmin As such, one or more of the 5 kringle domains can be
deleted wholly or partially. Truncated plasmins lacking one or more
kringle domains and/or lacking parts of one or more kringle domains
therefore are envisaged by the current invention. Examples of
truncated variants of plasmin include, but are not limited to,
"midiplasmin", "miniplasmin", "microplasmin", and "delta-plasmin".
Midiplasmin is basically lacking kringle domains 1 to 3 (e.g.
Christensen et al., 1995, Biochem J 305, 97-102). Miniplasmin was
originally obtained by limited digestion of plasmin with elastase
and is basically lacking kringle domains 1 to 4 (e.g. Christensen
et al., 1979, Biochim Biophys Acta 567, 472-481; Powell &
Castellino, 1980, J Biol Chem 255, 5329). Miniplasmin has
subsequently been produced recombinantly (WO 2002/050290).
Microplasmin was originally obtained by incubation of plasmin at
elevated pH and is basically lacking all kringle domains (e.g. WO
89/01336). Whereas the microplasmin obtained from incubation of
plasmin at elevated pH is containing the 30-31 carboxy-terminal
amino acids of the heavy chain, a recombinantly produced
microplasmin variant contains the 19 carboxy-terminal amino acids
of the heavy chain (WO 2002/050290). Delta-plasmin is a recombinant
version of plasmin in which kringle domain 1 is linked directly
with the catalytic domain (WO 2005/105990). The above described
truncated variants of plasmin are obtained by activation of
"midiplasminogen", "miniplasminogen", "microplasminogen" and
"delta-plasminogen", respectively. In order to be activatable, a
truncated plasminogen needs to comprise a minimum number of amino
acids of the linker between the kringle 5 domain and the catalytic
domain (see, e.g., Wang et al., 1995, Protein Science 4,
1758-1767). As alternative to plasmin or an active truncated
variant thereof, an activatable plasminogen or an activatable
truncated variant thereof can be used in the context of the current
invention (see, e.g. EP 0480906; U.S. Pat. No. 5,304,383; EP
0631786; U.S. Pat. No. 5,520,912; U.S. Pat. No. 5,597,800; U.S.
Pat. No. 5,776,452). "Plasminogen" refers to any form of
plasminogen e.g. Glu-plasminogen or Lys-plasminogen (starting with
Arg at position 68 or Lys at positions 77 or 78). When using
activatable plasminogen or an activatable truncated variant
thereof, the activation to plasmin may be delayed and will occur
after contacting it with an organ, tissue or body fluid. In yet
another alternative, the plasmin or an active truncated variant
thereof can be substituted in the context of the current invention
for an activatable plasminogen or an activatable truncated variant
thereof in conjunction with a plasminogen activator (such as tissue
plasminogen activator (tPA), urokinase, streptokinase or
staphylokinase; see, e.g. U.S. Pat. No. 6,733,750; U.S. Pat. No.
6,585,972; U.S. Pat. No. 6,899,877; WO 03/33019). In yet a further
alternative, a mixture of any of (i) plasmin or an active truncated
variant thereof, (ii) activatable plasminogen or an activatable
truncated variant thereof, and (iii) a plasminogen activator can be
used in the context of the current invention (see, e.g. US
2004/0081643). In order to ensure stability of the plasmin (or
plasminogen), it will generally be stored at lowered temperatures
(e.g. 4 degrees Celsius or -20 degrees Celsius) in a stabilizing
composition such as low pH (pH 4 or lower; obtained by e.g. 1 mM to
250 mM of an acid such as citric acid, see, e.g. Castellino &
Sodetz, 1976, Methods Enzymol 45, 273-286; WO 01/36608; WO
01/36609; WO 01/36611) or high glycerol content (30-50% v/v, e.g.,
Castellino & Sodetz, 1976, Methods Enzymol 45, 273-286),
alternatively in or in conjunction with one or more further
stabilizers such as an amino acid (e.g. lysine or an analogue
thereof), a sugar (e.g. mannitol) or any stabilizer as known in the
art (e.g. dipeptides, WO 97/01631). Further included in the genus
"plasmin" is any active derivative thereof (or of an active
truncated plasmin variant), or similar derivative of activatable
plasminogen (or of activatable truncated variant thereof). Such
derivates include e.g. labeled plasmin or plasminogen (or truncated
variants thereof) such as Tc.sup.99-labeled plasmin (Deacon et al.,
1980, Br J Radiol 53, 673-677) or pegylated or acylated plasmin or
plasminogen (or truncated variants thereof; EP 9879, WO 93/15189).
Said derivatives further include hybrid or chimeric plasmin or
plasminogen molecules comprising e.g. a truncated plasmin or
plasminogen according to the invention fused with e.g. a
fibrin-binding molecule (such as kringle 2 of tPA, an
apolipoprotein kringle, the finger domain of tPA or fibronectin and
the Fab domain of a fibrin-binding antibody).
[0017] Many assays exist to determine whether or not a plasmin
species is proteolytically active. Easy and straightforward assays
are based on the digestion of a chromogenic substrate by plasmin
present in a sample; chromogenic substrates include S-2403 and
S-2251 which release p-nitroaniline (pNA) upon proteolytic
cleavage. The amount of pNA formed can be measured by light
absorbance at 405 nm. An alternative assay for determining plasmin
activity is a potentiometric assay. Colorimetric (using a
chromogenic substrate) and potentiometric assays are described in
e.g., Castellino & Sodetz (1976, Methods Enzymol 45, 273-286).
A further alternative assay for determining plasmin activity is a
caseinolytic assay (e.g., Robbins & Summaria, 1970, Methods
Enzymol 19, 184-199; Ruyssen & Lauwers, 1978, Chapter
IX--Plasmin, In "Pharmaceutical Enzymes", Story-Scientia, Gent,
Belgium, pp. 123-131). Yet another alternative assay for
determining plasmin activity is a fibrinolytic assay (e.g., Astrup
& Mullertz, 1952, Arch Biochem Biophys 40, 346-351). Any
suitably labeled natural substrate of plasmin can in fact be used
by the skilled person to design a plasmin activity assay.
[0018] The "trabecular meshwork (TM)" is a mesh-like structure
inside the eye at the iris-scleral junction of the anterior chamber
angle. The TM filters the aqueous fluid and controls its flow into
the canal of Schlemm prior to its leaving the anterior chamber.
Increased resistance in the TM leads to reduced aqueous fluid
outflow and thus increased intra-ocular pressure (IOP).
[0019] When left untreated, this elevated IOP leads to glaucomatous
damage to the optic nerve and retinal nerve fibers, and leads to
loss of vision. This vision loss can be prevented or halted by
administering medication, an "agent for controlling the
intra-ocular pressure", which controls the intra-ocular pressure.
Such medicaments include adrenergic blocking agents (beta blockers
or sympatholytic drugs such as betaxolol, carteolol, levobunolol,
metipanolol and timolol), adrenergic stimulating agents
(sympathomimetic drugs such as aproclonidine, epinephrine,
hydroxyamphetamine, phenylephrine, naphazoline and
tetrahydrozaline), carbonic anhydrase inhibitors (such as systemic
acetozolamide, and topical brinzolamide and dorzolamide), miotics
(cholinergic stimulating agents, parasympathomimetic drugs such as
carbachol and pilocarpine), osmotic agents (such as glycerin and
mannitol), prostaglandin and prostaglandin analogues (prostamides,
bimatoprost, unoprostone isopropyl, travoprost, latanoprost,
natural prostaglandin, prostaglandin F2.alpha., and FP prostanoid
receptor agonists). When such medicaments are not efficient (or not
anymore), then filtration surgery is a viable treatment.
[0020] "Trabeculectomy", "trabeculectomy surgery" or "filtration
surgery", is defined as a surgical procedure on the eye wherein
part of the trabecular meshwork is removed whereby a filtration
site (a sclera-corneal drainage channel) is created that increases
the outflow of aqueous fluid from the eye; this type of filtering
procedure is commonly used in the treatment of glaucoma, and more
specifically to reduce the IOP in an eye subject to/suffering from
glaucoma. FIG. 2 is a schematical representation of the result of
trabeculectomy surgery.
[0021] "Filtration failure" is a condition reversing the clinically
desired effect of trabeculectomy surgery, i.e., reversing the
desired drop in IOP. The initial post-operative time is crucial in
the sense that eye-healing activities are highest in this period.
This period of high eye-healing capacity is dependent upon the
species and spans about 2 weeks for rabbits and up to 1- to
2-months in humans. Upon contacting plasmin or an active truncated
variant thereof (or any alternative therefore as described above)
with an eye according to the current invention, the frequency of
occurrence of filtration failure over a given period of time is
lowered. Plasmin or an active truncated variant thereof (or any
alternative therefore as described above) used according to the
current invention thus results in the prevention, reduction or
retarding of the occurrence of filtration failure.
[0022] The plasmin or active truncated variant thereof of the
invention, or the medicament containing one or more of them, for
treating filtration failure after trabeculectomy surgery of an eye,
or for preventing, reducing or retarding the occurrence of
filtration failure after trabeculectomy surgery of an eye may be in
a pharmaceutically acceptable formulation capable of being
administered to an eye as topical eye drops. Alternatively, the
plasmin or active truncated variant thereof of the invention, or
the medicament containing one or more of them, is in a
pharmaceutically acceptable formulation capable of being
administered by injection into the anterior chamber of an eye. The
composition of the eye drop formulation and the formulation for
injection into the anterior chamber of an eye may be the same or
different. To obtain optimal clinical outcomes, the compositions of
the formulations may need to be adjusted to their mode of
application and may thus need to be different.
[0023] The treatment of filtration failure after trabeculectomy
surgery of an eye, or the prevention, reduction or retardation of
the occurrence of filtration failure after trabeculectomy surgery
of an eye may result from contacting said eye with an effective
amount of topical eye drops comprising said plasmin or active
truncated variant thereof. In other words, for treatment of
filtration failure after trabeculectomy surgery of an eye, or for
prevention, reduction or retardation of the occurrence of
filtration failure after trabeculectomy surgery of an eye, the
effective amount of plasmin or active truncated variant thereof may
be or is to be administered in the form of topical eye drops.
[0024] Alternatively, the treatment of filtration failure after
trabeculectomy surgery of an eye, or the prevention, reduction or
retardation of the occurrence of filtration failure after
trabeculectomy surgery of an eye may result from introduction into
the anterior chamber of an eye of an effective amount of said
plasmin or active truncated variant thereof. In other words, for
treatment of filtration failure after trabeculectomy surgery of an
eye, or for prevention, reduction or retardation of the occurrence
of filtration failure after trabeculectomy surgery of an eye, the
effective amount of plasmin or active truncated variant thereof may
be or is to be administered by introduction or injection into the
anterior chamber of an eye.
[0025] In a further alternative, the treatment of filtration
failure after trabeculectomy surgery of an eye, or the prevention,
reduction or retardation of the occurrence of filtration failure
after trabeculectomy surgery of an eye may result from contacting
said eye with an effective amount of topical eye drops comprising
said plasmin or active truncated variant thereof, combined with
introduction into the anterior chamber of an eye of an effective
amount of said plasmin or active truncated variant thereof. The
effective amount of plasmin or active truncated variant thereof may
in this case be reached only by the combined administrations. In
other words, for treatment of filtration failure after
trabeculectomy surgery of an eye, or for prevention, reduction or
retardation of the occurrence of filtration failure after
trabeculectomy surgery of an eye, the effective amount of plasmin
or active truncated variant thereof is to be administered in the
form of topical eye drops combined with introduction or injection
into the anterior chamber of an eye. In the above, the amount or
concentration of active substance in the eye drop formulation and
in the formulation for anterior chamber intracameral injection may
be the same or different. The amounts or concentrations of active
substance may need to be adjusted such as to the mode of
application or such as to minimize eventual side effects that may
occur when e.g. administering a high amount or concentration of
active substance by either one of the administration routes. In the
latter case, the effective amount of active substance can still be
reached by compensation of a low amount or concentration of active
substance via one administration route by a higher amount or
concentration of active substance via the other administration
route.
[0026] In an embodiment to any of the above, said active truncated
variant of plasmin may be lacking one or more kringle domains
and/or lacking parts of one or more kringle domains. More
specifically, said active truncated variant of plasmin may be
selected from the group consisting of midiplasmin, miniplasmin,
microplasmin or deltaplasmin.
[0027] The invention further covers the (use of) a plasmin or an
active truncated variant thereof (for the manufacture of a
medicament) for treating filtration failure after trabeculectomy
surgery of an eye, or for preventing, reducing or retarding the
occurrence of filtration failure after trabeculectomy surgery of an
eye, which is in a pharmaceutically acceptable solution that may
further comprise one or more of an agent for controlling the
intra-ocular pressure, an anti-inflammatory agent, an antiviral
agent, an antibacterial agent, an antiviral agent, an
anti-angiogenic agent, an anti-mitotic agent, an antihistamine, an
anesthetic, an agent to induce mydriasis and an agent to induce
cycloplegia. Alternatively, when said further agent(s) is(are) not
included in the pharmaceutically acceptable solution or medicament
containing said plasmin or an active truncated variant thereof,
said eye may be contacted further with one or more agents chosen
from an agent for controlling the intra-ocular pressure, an
anti-inflammatory agent, an antiviral agent, an antibacterial
agent, an antiviral agent, an anti-angiogenic agent, an
anti-mitotic agent, an antihistamine, an anesthetic, an agent to
induce mydriasis and an agent to induce cycloplegia.
[0028] Methods of treatment of filtration failure after
trabeculectomy surgery of an eye, and in particular methods of
preventing, reducing or retarding the occurrence of filtration
failure after trabeculectomy surgery of an eye are also envisaged.
These methods comprise the step of contacting said eye after
trabeculectomy surgery with a medicament comprising plasmin or an
active truncated variant thereof wherein said contacting results in
said treatment of filtration failure, or in said preventing,
reducing or retarding of the occurrence of filtration failure.
Modalities of said medicament, plasmin or an active truncated
variant thereof according to the invention, and contacting are as
described above.
[0029] In any of the above-described medical uses and methods, the
plasmin can be substituted for plasminogen, plasminogen activators
or any possible combination (whether or not in the same formulation
or as separate solutions or medicaments) of plasmin (or any active
truncated variant thereof), plasminogen, plasminogen activators,
etc. as described earlier in the definition of "plasmin".
[0030] "Contacting" means any mode of administration that results
in interaction between a composition such as a medicament and an
object (such as conjunctiva or subconjunctival tissue) with which
said composition is contacted. The interaction between the
composition and the object can occur starting immediately or nearly
immediately with the administration of the composition, can occur
over an extended time period (starting immediately or nearly
immediately with the administration of the composition), or can be
delayed relative to the time of administration of the composition.
More specifically the "contacting" may result in delivering an
effective amount of the medicament to the object.
[0031] The term "effective amount" refers to the dosing regimen of
the medicament according to the invention, in particular of the
active ingredient of the medicament according to the invention,
i.e., plasmin or an active truncated variant thereof (or any
alternative therefore as described above). The effective amount
will generally depend on and will need adjustment to the mode of
contacting or administration. The effective amount of the
medicament, more particular its active ingredient, is the amount
required to obtain the desired clinical outcome or therapeutic or
prophylactic effect without causing significant or unnecessary
toxic effects. To obtain or maintain the effective amount, the
medicament may be administered as a single dose or in multiple
doses. The effective amount may further vary depending on the
severity of the condition that needs to be treated or the expected
severity of the condition that needs to be prevented or treated;
this may depend on the overall health and physical condition of the
patient and usually the treating doctor's or physician's assessment
will be required to establish what is the effective amount. The
effective amount may further be obtained by a combination of
different types of contacting or administration. In the context of
the present invention the effective amount may more particularly be
obtained by either one or more of administration of topical eye
drops, administration by injection into the anterior chamber of an
eye or administration by subconjunctival injection. A typical dose
of a single administration of the medicament of the invention may
comprise 10 .mu.g to 1 mg of the active compound (i.e., a plasmin
or an active truncated variant thereof, or any alternative thereto
as described higher). Administration of the medicament of the
invention by means of injection typically is kept to a minimum,
i.e., the frequency of repeat injections is kept to a minimum.
Administration of the medicament of the invention by means of
topical eye drops can be done more frequently, e.g., once per hour,
or e.g. 1 to 6 times a day. As the first weeks or months
post-trabeculectomy (species dependent as described higher) are
crucial in the sense that eye-healing activities are highest in
this period, the duration of treatment with a medicament according
to the present invention should be adjusted to this period. Initial
dosage and administration frequency may thus be relatively high and
may be gradually decreased when the risk of the occurrence of
filtration failure is decreasing.
[0032] In general, the medicament or composition of the invention
comprising a plasmin (or any variant or derivative thereof or
alternative thereto) according to the invention may, depending on
its ultimate use and mode of administration, comprise one or more
further active ingredients such as an agent controlling the
intra-ocular pressure (see higher), an anticoagulant, a
thrombolytic agent, an anti-inflammatory agent, an antiviral agent,
an antibacterial agent, an antifungal agent, an anti-angiogenic
agent, an anti-mitotic agent, an antihistamine or anesthetic.
[0033] "Anticoagulants" include hirudins, heparins, coumarins,
low-molecular weight heparin, thrombin inhibitors, platelet
inhibitors, platelet aggregation inhibitors, coagulation factor
inhibitors, anti-fibrin antibodies and factor VIII-inhibitors (such
as those described in WO 01/04269 and WO 2005/016455).
[0034] "Thrombolytic agents" include urokinase, streptokinase,
tissue-type plasminogen activator (tPA), urokinase-type plasminogen
activator (uPA) and staphylokinase or any variant or derivative of
any thereof such as APSAC (anisoylated plasminogen streptokinase
activator complex), alteplase, reteplase, tenecteplase, and scuPA
(single chain uPA).
[0035] "Anti-inflammatory agents" include steroids (e.g.
prednisolone, methylprednisolone, cortisone, hydrocortisone,
prednisone, triamcinolone, dexamethasone) and non-steroidal
anti-inflammatory agents (NSAIDs; e.g. acetaminophren, ibuprofen,
aspirin).
[0036] "Antiviral agents" include trifluridine, vidarabine,
acyclovir, valacyclovir, famciclovir, and doxuridine.
[0037] "Antibacterial agents" or antibiotics include ampicillin,
penicillin, tetracycline, oxytetracycline, framycetin,
gatifloxacin, gentamicin, tobramycin, bacitracin, neomycin and
polymyxin.
[0038] "Anti-mycotic/fungistatic/antifungal agents" include
fluconazole, amphotericin, clotrimazole, econazole, itraconazole,
miconazole, 5-fluorocytosine, ketoconazole and natamycin.
[0039] "Anti-angiogenic agents" include antibodies (or fragments
thereof) such as anti-VEGF (vascular endothelial growth factor) or
anti-P1GF (placental growth factor) antibodies and agents such as
macugen (pegaptanib sodium), trypthophanyl-tRNA synthetase (TrpRS),
anecortave acetate, combrestatin A4 prodrug, AdPEDF (adenovector
capable of expressing pigment epithelium-derived factor),
VEGF-trap, inhibitor of VEGF receptor-2, inhibitors of VEGF, P1GF
or TGF-.beta., Sirolimus (rapamycin) and endostatin.
[0040] "Anti-mitotic agents" include mitomycin C and
5-fluorouracyl.
[0041] "Antihistamine" includes ketitofen fumarate and pheniramine
maleate.
[0042] "Anesthetics" include benzocaine, butamben, dibucaine,
lidocaine, oxybuprocaine, pramoxine, proparacaine, proxymetacaine,
tetracaine and amethocaine.
[0043] Other adjunct agents or drugs that can be used in
conjunction with the plasmin or active variant thereof (or any
alternative thereto as described above) according to the invention
include scopoloamine, atropine or tropicamide, to induce mydriasis
(pupillary dilation) and/or cycloplegia (paralysis of the eye
focusing muscle).
[0044] In addition to plasmin or active truncated variant thereof
(or any of the alternatives therefor as described above), each of
the above listed agents as well as antihistamine and anesthetics is
to be considered as an "active ingredient".
[0045] A "pharmaceutically acceptable formulation" is, in the
context of the current invention more particular an
"ophthalmologically acceptable formulation". A formulation in
general is a composition comprising a carrier, diluent or adjunvant
compatible with the one or more active ingredients to be
formulated, the whole formulation being compatible with the
intended use in the intended tissue or organ, etc. Examples of
pharmaceutically acceptable formulations as well as methods for
making them can be found, e.g., in Remington's Pharmaceutical
Sciences (e.g. 20.sup.th Edition; Lippincott, Williams &
Wilkins, 2000) or in any Pharmacopeia handbook (e.g. US-, European-
or International Pharmacopeia).
[0046] "Topical eye drops" typically contains an active ingredient
(such as plasmin or an active truncated variant thereof or any
alternative thereto as described higher) or a combination of active
ingredients in a saline solution, and optionally one or more
lubricants.
[0047] "Lubricants" include propylene glycerol, glycerin,
carboxymethylcellulose, hydroxypropylmethylcellulose, soy lecithin,
polyvinyl alcohol, white petrolatum, mineral oil, povidone,
carbopol 980, polysorbate 80, dextran 70.
EXAMPLES
[0048] The Examples included hereafter demonstrate the invention
and are not construed to be limiting the scope of the invention in
any way.
Example 1
Rabbit Model for Glaucoma Filtration Surgery
[0049] Female New-Zealand rabbits, aged 12 to 14 weeks and weighing
2 to 3 kg, underwent a filtration surgery (trabeculectomy) in both
eyes in the same way as in human eyes, except that in rabbits a
much more aggressive postoperative fibrosis occurs, resulting in a
filtration failure after 10 to 14 days (Miller et al., 1985, Trans
Ophthalmol Soc UK 104, 893-897).
[0050] General anesthesia was induced with an intramuscular
injection of Ketalar (50 mg/ml) and Rompun (2%). Before the
operation the IOP was measured in both eyes by using the
Tono-Pen.RTM. tonometer (Medtronic Solan) under topical anesthesia
(Unicain, 4 mg/ml).
[0051] Briefly, for the trabeculectomy a Vicryl 9/0 corneal
traction suture was placed superiorly and the eye was pulled down.
A limbus-based conjunctival flap was raised after which a blunt
dissection of subconjunctival space was performed. After a scleral
flap of 5 mm to 5 mm was formed, a piece of the trabecular meshwork
was removed and an iridectomy was performed. The conjunctival and
scleral flaps were closed by using a Nylon 10-0 suture. At the end
of the operation a bleb was formed.
[0052] Postoperative follow up of the rabbits took place daily
during the first week and two-daily until they were scarified.
Examination of both eyes was done and all measurements were
performed under topical anesthesia. The IOP-recordings were
performed by using a Tono-Pen.RTM. tonometer. Bleb characteristics
including the bleb area (width and length) and the conjunctival
vascularity were investigated according to the Moorfields bleb
grading system. During the first week there was an assessment of
the anterior segment and of the presence of blood clots around the
filtration channel by slit lamp examination.
Example 2
Immunohistochemical Investigation
[0053] On day 30 after surgery, rabbits were killed using a lethal
intravenous injection of Rompun under general anesthesia. Both eyes
were enucleated, fixed overnight in 4% PFA and embedded in
paraffin. Seven-.mu.m thin slides were (immuno-)stained for CD45 to
evaluate inflammation and for Sirius red and Trichrome to evaluate
fibrosis.
A. Collagen Deposition
[0054] Sirius Red and Trichrome staining were used to demonstrate
collagen deposition. After Sirius red staining collagen is colored
red; after Trichrome staining collagen is colored blue (aniline
blue, 5 minutes), nuclei black (Weigert hematoxyline, 10 minutes),
and cytoplasm red (Biebrich scarlet fuchsin, 2 minutes).
B. Inflammation
[0055] A CD45 staining was performed to study inflammatory cells.
After a 20 minutes incubation with methanol and 45 minutes with PIR
(1/5; Dakocytomation) the samples were incubated overnight with
mouse anti-rabbit CD45 antibody (1/3, 10-50 .mu.g/ml; MCA808;
AbDSerotec). The next day the samples were incubated for 45 minutes
with RAM-B (1/300; Dakocytomation). The staining was finished by
using the Perkin Elmer kit (Renaissance TSA.TM. Indirect; NEL700).
The DAB (Fluka) is giving the tissue a brown color by adding
H.sub.20.sub.2. The counterstaining was performed by using Harris
hematoxyline (Merck).
Example 3
Effect of Microplasmin on Blebs after Trabeculectomy on Rabbit
Eyes
[0056] Group 1: Filtration surgery followed by injection in the
anterior chamber of microplasmin on day 0: immediately after the
trabeculectomy operation 10 rabbits got an anterior chamber
injection of microplasmin (200 .mu.l of a solution of 2,5 mg
microplasmin/mL in 5 mM citric acid, 6 mg/mL mannitol, pH 3.1). Ten
other rabbits underwent trabeculectomy followed by control
injections of the same volume of 0.9% NaCl.
[0057] Group 2: Filtration surgery followed by administration of
topical eye drops containing microplasmin: immediately after the
trabeculectomy operation on 3 rabbits, microplasmin was
administered in the form of topical eye drops (4 mg microplasmin/mL
in 5 mM citric acid, 6 mg/mL mannitol, pH 3.1; 1 drop of ca. 50-55
.mu.L was administered 4 times per day during a period of 14 days).
Three other rabbits underwent trabeculectomy followed by
administration of control eye drops of 0.9% NaCl.
[0058] Group 3: Filtration surgery followed by injection in the
anterior chamber of microplasmin and administration of topical eye
drops containing microplasmin: immediately after the trabeculectomy
operation 5 rabbits got an anterior chamber injection of
microplasmin (as in Group 1) combined with administration of
topical eye drops (as in Group 2). Five other rabbits underwent
trabeculectomy followed by control injections as in Group 1 and
administration of control eye drops as in Group 2.
[0059] Group 4: similar to Group 1 except that 100 .mu.L
microplasmin is administered subconjunctivally (instead of 200
.mu.L in the anterior chamber) in the eyes of 5 rabbits. The
control group consists of 5 rabbits.
[0060] Group 5: similar to Group 4 except that an additional 100
.mu.L microplasmin is administered subconjunctivally (repeat
administration) 1 week after the initial administration. The
control group consists of 5 rabbits.
[0061] In any of the above outlined experiments the acidic
microplasmin solution may alternatively be neutralized prior to
contacting with the eye.
[0062] Results: As illustrated in FIG. 3 (normalized data),
microplasmin significantly augmented the bleb area and survival in
a rabbit model of trabeculectomy. All depicted treatments had an
initial more or less equal positive effect on the bleb survival
(diamonds: topical administration; triangles: anterior chamber
injection; squares: combined topical administration and anterior
chamber injection). The anterior chamber injection of microplasmin
and the combined administration of eye drops and the anterior
chamber injection had a positive effect on bleb survival over a
longer period of time. Collagen deposition was borderline reduced
after microplasmin administration compared to control. No
significant changes in inflammation were observed in the anterior
chamber or in the conjunctiva. Contrary to the eye drops and/or
anterior chamber injection, subconjunctival injection of
microplasmin did not result in enhanced bleb survival.
Sequence CWU 1
1
11791PRTHomo sapiens 1Glu Pro Leu Asp Asp Tyr Val Asn Thr Gln Gly
Ala Ser Leu Phe Ser1 5 10 15Val Thr Lys Lys Gln Leu Gly Ala Gly Ser
Ile Glu Glu Cys Ala Ala 20 25 30Lys Cys Glu Glu Asp Glu Glu Phe Thr
Cys Arg Ala Phe Gln Tyr His 35 40 45Ser Lys Glu Gln Gln Cys Val Ile
Met Ala Glu Asn Arg Lys Ser Ser 50 55 60Ile Ile Ile Arg Met Arg Asp
Val Val Leu Phe Glu Lys Lys Val Tyr65 70 75 80Leu Ser Glu Cys Lys
Thr Gly Asn Gly Lys Asn Tyr Arg Gly Thr Met 85 90 95Ser Lys Thr Lys
Asn Gly Ile Thr Cys Gln Lys Trp Ser Ser Thr Ser 100 105 110Pro His
Arg Pro Arg Phe Ser Pro Ala Thr His Pro Ser Glu Gly Leu 115 120
125Glu Glu Asn Tyr Cys Arg Asn Pro Asp Asn Asp Pro Gln Gly Pro Trp
130 135 140Cys Tyr Thr Thr Asp Pro Glu Lys Arg Tyr Asp Tyr Cys Asp
Ile Leu145 150 155 160Glu Cys Glu Glu Glu Cys Met His Cys Ser Gly
Glu Asn Tyr Asp Gly 165 170 175Lys Ile Ser Lys Thr Met Ser Gly Leu
Glu Cys Gln Ala Trp Asp Ser 180 185 190Gln Ser Pro His Ala His Gly
Tyr Ile Pro Ser Lys Phe Pro Asn Lys 195 200 205Asn Leu Lys Lys Asn
Tyr Cys Arg Asn Pro Asp Arg Glu Leu Arg Pro 210 215 220Trp Cys Phe
Thr Thr Asp Pro Asn Lys Arg Trp Glu Leu Cys Asp Ile225 230 235
240Pro Arg Cys Thr Thr Pro Pro Pro Ser Ser Gly Pro Thr Tyr Gln Cys
245 250 255Leu Lys Gly Thr Gly Glu Asn Tyr Arg Gly Asn Val Ala Val
Thr Val 260 265 270Ser Gly His Thr Cys Gln His Trp Ser Ala Gln Thr
Pro His Thr His 275 280 285Asn Arg Thr Pro Glu Asn Phe Pro Cys Lys
Asn Leu Asp Glu Asn Tyr 290 295 300Cys Arg Asn Pro Asp Gly Lys Arg
Ala Pro Trp Cys His Thr Thr Asn305 310 315 320Ser Gln Val Arg Trp
Glu Tyr Cys Lys Ile Pro Ser Cys Asp Ser Ser 325 330 335Pro Val Ser
Thr Glu Gln Leu Ala Pro Thr Ala Pro Pro Glu Leu Thr 340 345 350Pro
Val Val Gln Asp Cys Tyr His Gly Asp Gly Gln Ser Tyr Arg Gly 355 360
365Thr Ser Ser Thr Thr Thr Thr Gly Lys Lys Cys Gln Ser Trp Ser Ser
370 375 380Met Thr Pro His Arg His Gln Lys Thr Pro Glu Asn Tyr Pro
Asn Ala385 390 395 400Gly Leu Thr Met Asn Tyr Cys Arg Asn Pro Asp
Ala Asp Lys Gly Pro 405 410 415Trp Cys Phe Thr Thr Asp Pro Ser Val
Arg Trp Glu Tyr Cys Asn Leu 420 425 430Lys Lys Cys Ser Gly Thr Glu
Ala Ser Val Val Ala Pro Pro Pro Val 435 440 445Val Leu Leu Pro Asp
Val Glu Thr Pro Ser Glu Glu Asp Cys Met Phe 450 455 460Gly Asn Gly
Lys Gly Tyr Arg Gly Lys Arg Ala Thr Thr Val Thr Gly465 470 475
480Thr Pro Cys Gln Asp Trp Ala Ala Gln Glu Pro His Arg His Ser Ile
485 490 495Phe Thr Pro Glu Thr Asn Pro Arg Ala Gly Leu Glu Lys Asn
Tyr Cys 500 505 510Arg Asn Pro Asp Gly Asp Val Gly Gly Pro Trp Cys
Tyr Thr Thr Asn 515 520 525Pro Arg Lys Leu Tyr Asp Tyr Cys Asp Val
Pro Gln Cys Ala Ala Pro 530 535 540Ser Phe Asp Cys Gly Lys Pro Gln
Val Glu Pro Lys Lys Cys Pro Gly545 550 555 560Arg Val Val Gly Gly
Cys Val Ala His Pro His Ser Trp Pro Trp Gln 565 570 575Val Ser Leu
Arg Thr Arg Phe Gly Met His Phe Cys Gly Gly Thr Leu 580 585 590Ile
Ser Pro Glu Trp Val Leu Thr Ala Ala His Cys Leu Glu Lys Ser 595 600
605Pro Arg Pro Ser Ser Tyr Lys Val Ile Leu Gly Ala His Gln Glu Val
610 615 620Asn Leu Glu Pro His Val Gln Glu Ile Glu Val Ser Arg Leu
Phe Leu625 630 635 640Glu Pro Thr Arg Lys Asp Ile Ala Leu Leu Lys
Leu Ser Ser Pro Ala 645 650 655Val Ile Thr Asp Lys Val Ile Pro Ala
Cys Leu Pro Ser Pro Asn Tyr 660 665 670Val Val Ala Asp Arg Thr Glu
Cys Phe Ile Thr Gly Trp Gly Glu Thr 675 680 685Gln Gly Thr Phe Gly
Ala Gly Leu Leu Lys Glu Ala Gln Leu Pro Val 690 695 700Ile Glu Asn
Lys Val Cys Asn Arg Tyr Glu Phe Leu Asn Gly Arg Val705 710 715
720Gln Ser Thr Glu Leu Cys Ala Gly His Leu Ala Gly Gly Thr Asp Ser
725 730 735Cys Gln Gly Asp Ser Gly Gly Pro Leu Val Cys Phe Glu Lys
Asp Lys 740 745 750Tyr Ile Leu Gln Gly Val Thr Ser Trp Gly Leu Gly
Cys Ala Arg Pro 755 760 765Asn Lys Pro Gly Val Tyr Val Arg Val Ser
Arg Phe Val Thr Trp Ile 770 775 780Glu Gly Val Met Arg Asn Asn785
790
* * * * *