Protection of cardiac myocardium
United States Patent: 7,504,379
Issued: March 17, 2009
Inventors: Edelberg; Jay
(New York, NY), Xaymardan; Munira (Toronto, CA)
Assignee: Cornell Research
Foundation, Inc. (Ithaca, NY)
Appl. No.: 11/129,076
Filed: May 13, 2005
Web Seminars -- Pharm/Biotech/etc.
The invention provides compositions and
methods for protecting vascular tissues from injury that occurs, for
example, during occlusion of one or more arteries. In some embodiments,
the injury is myocardial infarction. The compositions of the invention
include combinations of platelet derived growth factor, vascular
endothelial growth factor, and angiopoietin-2.
Description of the
SUMMARY OF THE INVENTION
The invention provides compositions and methods for protecting vascular
tissue in a patient from damage, for example, during occlusion of an artery.
In some embodiments, the compositions and methods are used to prevent or
treat tissue injuries resulting from myocardial infarction.
The methods of the invention involve administering to the patient a
composition comprising a therapeutically effective amount of
platelet-derived growth factor, angiopoietin-2, and vascular endothelial
growth factor. The composition can also include a pharmaceutically
In one embodiment, the invention provides a method of promoting vascular
health in a patient by administering to the patient a therapeutically
effective amount of platelet derived growth factor, angiopoietin-2, and
vascular endothelial growth factor.
In another embodiment, the invention provides a method of protecting cardiac
tissue in a patient from damage during myocardial infarction by
administering to the patient a therapeutically effective amount of platelet
derived growth factor, angiopoietin-2, and vascular endothelial growth
In yet another embodiment, the invention provides a method of improving
survival of transplanted tissue in a patient by administering to the patient
a therapeutically effective amount of platelet derived growth factor,
angiopoietin-2, and vascular endothelial growth factor.
Administration can be parenteral, as by intravascular, intravenous,
intraarterial, intraperitoneal, intraventricular infusion, stent, infusion
catheter, balloon catheter, bolus injection, topical administration as by
direct application to tissue surfaces during surgery, or oral.
Alternatively, administration can be directly into the heart or into the
vasculature of the patient.
The platelet-derived growth factor employed can, for example, be platelet
derived growth factor AB, platelet derived growth factor A, platelet derived
growth factor B, or mixtures thereof. In some embodiments, the
platelet-derived growth factor can have a sequence comprising SEQ ID NO:1,
SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6 or a mixture
thereof. The angiopoietin-2 can, for example, have a sequence comprising SEQ
ID NO:7, SEQ ID NO:8, or a mixture thereof. The vascular endothelial growth
factor can, for example, have a sequence comprising SEQ ID NO:9, SEQ ID
NO:10, SEQ ID NO:11, SEQ ID NO:12 or a mixture thereof.
The size of necrosis, vascular injury or myocardial infarction in the
patient can be reduced by such compositions and methods of the invention.
DETAILED DESCRIPTION OF THE INVENTION
The present invention provides pharmaceutical compositions comprising an
effective amount of platelet-derived growth factor (PDGF), angiopoietin-2
(Ang-2) and vascular endothelial growth factor (VEGF) that are effective for
preventing damage to cardiac tissues as a result of vascular conditions or
diseases, for example, those conditions and diseases that can lead to
occlusion of an artery. Such administration is given before, during or as
soon as possible after arterial occlusion.
The vascular diseases treated by the present invention are vascular diseases
of mammals. The word mammal means any mammal. Some examples of mammals
include, for example, pet animals, such as dogs and cats; farm animals, such
as pigs, cattle, sheep, and goats; laboratory animals, such as mice and
rats; primates, such as monkeys, apes, and chimpanzees; and humans.
According to the invention, endothelial cells within normal vascular tissues
change as they grow older, exhibiting reduced angiogenesis and losing their
ability to communicate with other cells by secreting signaling agents. These
changes can lead to a diminished capacity for blood vessel formation, a
reduction in blood flow to the associated organ or system, and an inability
to recover from injuries or diseases that can block or adversely affect
Many pathological conditions can lead to vascular diseases that can block
blood vessels such as atherosclerosis, preeclampsia, peripheral vascular
disease, erectile dysfunction, cancers, renal failure, heart disease, and
stroke. Such conditions are associated with alterations in the normal
vascular condition of the affected tissues and/or systems.
Accordingly, this invention relates to methods for treating endothelial
dysfunction, such as a vascular condition or a circulatory condition
associated with loss, injury or disruption of the vasculature within an
anatomical site or system. The term "vascular condition" or "vascular
disease" refers to a state of vascular tissue where blood flow is, or can
become, impaired or even blocked.
Examples of vascular conditions or vascular disease to which the methods of
the invention apply are those in which the vasculature of the affected
tissue or system is injured, senescent, or otherwise altered in some way, so
that blood flow to the tissue or system is reduced or is in danger of being
reduced. Vascular, circulatory or hypoxic conditions to which the methods of
the invention apply are those associated with, but not limited to,
myocardial infarction, maternal hypoxia (e.g., placental hypoxia,
preeclampsia), abnormal pregnancy, peripheral vascular disease (e.g.,
arteriosclerosis), transplant accelerated arteriosclerosis, deep vein
thrombosis, erectile dysfunction, cancers, renal failure, stroke, heart
disease, sleep apnea, hypoxia during sleep, female sexual dysfunction, fetal
hypoxia, respiratory disorders due to smoking, anemia, hypovolemia, vascular
or circulatory conditions which increase risk of metastasis or tumor
progression, hemorrhage, hypertension, diabetes, vasculopathologies,
disorders due to surgery (e.g., per-surgical hypoxia, post-operative
hypoxia), Raynaud's disease, endothelial dysfunction, regional perfusion
deficits (e.g., limb, gut, renal ischemia), stroke, thrombosis, frost bite,
decubitus ulcers, asphyxiation, poisoning (e.g., carbon monoxide, heavy
metal), altitude sickness, pulmonary hypertension, sudden infant death
syndrome (SIDS), asthma, chronic obstructive pulmonary disease (COPD),
congenital circulatory abnormalities (e.g., Tetralogy of Fallot) and
erythroblastosis (blue baby syndrome). In particular embodiments, the
invention provides methods of treating loss of circulation or endothelial
dysfunction in a mammal such as a human patient.
Thus, the invention is directed to methods of promoting vascular health by
preventing or treating diseases or conditions like those listed above. In
some embodiments, the disease or condition is stroke, atherosclerosis, acute
coronary syndromes including unstable angina, thrombosis and myocardial
infarction, plaque rupture, both primary and secondary (in-stent) restenosis
in coronary or peripheral arteries, transplantation-induced sclerosis,
peripheral limb disease, intermittent claudication and diabetic
complications (including ischemic heart disease, peripheral artery disease,
congestive heart failure, retinopathy, neuropathy and nephropathy), or
In some embodiments, the vascular condition or vascular disease arises from
injured or damaged myocardium. As used herein injured or damaged myocardium
refers to myocardial cells that have been exposed to ischemic conditions.
These ischemic conditions may be caused by a myocardial infarction,
transplant injury, heart surgery or cardiovascular disease. The lack of
oxygen causes the death of the cells in the surrounding area, leaving an
infarct that can eventually scar.
Preferably, injured or damaged myocardium is treated with the methods and
compositions of the invention before damage occurs (e.g. when damage is
suspected of occurring or a possibility exists of potential vascular
injury), during injury or as quickly as possible after injury or damage
occurs. Hence, the methods and compositions of the invention are
advantageously employed on tissues that may be experiencing, are
experiencing, have just experienced or are in danger of ischemia, myocardial
infarction, heart attack or loss of blood flow. The methods and compositions
of the invention are also advantageously employed on recently damaged
vascular tissues (e.g. myocardium) and on not so recently damaged vascular
tissues (e.g. myocardium).
As used herein "recently damaged vascular tissues" refers to vascular
tissues (or myocardium) that has been damaged within zero to one day of
treatment being started. In a preferred embodiment, the tissue (or
myocardium) has been damaged within zero to twelve hours of the start of
treatment. In a further preferred embodiment, the tissue or myocardium has
been damaged within 6 hours of the start of treatment.
The methods and compositions of the invention can be used to prevent or to
treat vascular conditions such as those listed above. These methods involve
administering an effective amount of a combination of platelet-derived
growth factor (PDGF), angiopoietin-2 (Ang-2) and vascular endothelial growth
factor (VEGF). Such an effective amount is effective when it reduces the
extent of damage due to vascular injury, vascular occlusion or myocardial
Platelet-Derived Growth Factor
Naturally occurring, platelet-derived growth factor ("PDGF") is a
disulfide-bonded dimer having two polypeptide chains, namely the "A" and "B"
chains, with the A chain being approximately 60% homologous to the B chain.
Naturally occurring PDGF is found in three dimeric forms, namely PDGF-AB
heterodimer, PDGF-BB homodimer, or PDGF-AA homodimer. Hannink et al., Mol.
Cell. Biol., 6, 1304-1314 (1986). PDGF-AB has been identified as the
predominant naturally occurring form. However, some data indicates that the
PDGF-BB homodimer may be effective for wound healing. Each monomeric subunit
of the biologically active dimer, irrespective of whether it is an A chain
monomer or a B chain monomer, contains eight cysteine residues. Some of
these cysteine residues form interchain disulfide bonds that hold the dimer
together. As used herein, the term PDGF means any PDGF polypeptide or
protein, including PDGF A, PDGF B, PDGF AB, PDGF BB, and PDGF AA.
The A polypeptide of human PDGF can be any mammalian PDGF A polypeptide
including, for example, human, mouse, rat, rabbit, goat, bovine, horse,
sheep and any other mammalian PDGF A polypeptide. The following sequence is
one example of an amino acid sequence of a human PDGF A polypeptide (SEQ ID
-- see Original Patent.
The following sequence is an
example of a mouse PDGF A sequence (SEQ ID NO:2)
-- see Original Patent.
Other sequences for PDGF A can readily be obtained by one of skill in the
art, for example, from the GenBank database of sequences. Variability in
these and other sequences is permitted so long as the PDGF A polypeptide can
dimerize with PDGF B and/or function in cell-to-cell communication.
The PDGF B polypeptide found in human platelets has been identified as a 109
amino acid cleavage product (PDGF-B.sub.109) of a 241 amino acid precursor
polypeptide Johnsson et al., EMBO Journal, 3(5), 921-928 (1984). An example
of a human sequence for the PDGF B polypeptide is provided below (SEQ ID
-- see Original Patent.
The following sequence is an example of a mouse PDGF B sequence (SEQ ID
-- see Original Patent.
A 109 amino acid PDGF B polypeptide is believed to be the mature form of
PDGF in humans and constitutes a cleavage product of the PDGF-B precursor
protein. Homology with the precursor protein begins at amino acid 82 of the
241 amino acid precursor protein and continues for 109 amino acids yielding,
for example, a polypeptide with the following sequence (SEQ ID NO:5)
-- see Original Patent.
Another form of PDGF-B
(PDGF-B.sub.119), corresponds to the first 119 amino acids of the PDGF-B
precursor protein (SEQ ID NO:6)
-- see Original Patent.
This PDGF-B.sub.119 form has
also been identified as a major cleavage product of the precursor protein
when the entire gene is expressed in a transfected mammalian host. See U.S.
Pat. No. 5,149,792.
Human platelet-derived growth factor is believed to be the major mitogenic
growth factor in serum for connective tissue cells. PDGF can positively
affect mitogenesis in arterial smooth muscle cells, fibroblast cells lines,
and glial cells. Deuel et al., J. Biol. Chem., 256(17), 8896-8899 (1981).
See also, e.g., Heldin et al., J. Cell Physiol., 105, 235 (1980) (brain
glial cells); Raines and Ross, J. Biol. Chem., 257, 5154 (1982) (monkey
arterial smooth muscle cells). PDGF is also believed to be a chemoattractant
for fibroblasts, smooth muscle cells, monocytes, and granulocytes.
An angiogenic factor, which was originally called TIE-2 ligand-1 (TL1), but
which is also referred to as angiopoietin-1 (Ang1) is required for normal
vascular development in the mouse. Angiopoietin-2 (also called TIE-2
ligand-2, TL2) is believed to be a naturally occurring antagonist for Ang1
and the angiopoietin-2 receptor. A description of the cloning and sequencing
of TL1 (Ang1) and TL2 (Ang2) as well as for methods of making and uses
thereof is provided in PCT International Publication No. WO 96/11269
published Apr. 18, 1996, PCT International Publication No. WO 96/31598
published Oct. 10, 1996 and S. Davis, et al., Cell 87: 1161-1169 (1996) each
of which is hereby incorporated by reference.
One example of a sequence of a human angiopoietin-2 precursor polypeptide is
as follows (SEQ ID NO:7)
-- see Original Patent.
Another example of a sequence of a human angiopoietin-2 precursor
polypeptide is as follows (SEQ ID NO:8)
-- see Original Patent.
Vascular Endothelial Growth
Vascular endothelial growth factor (VEGF) (Ferrara et al., Endo. Rev., 13:
18-32 ) is a potent angiogenic factor that acts via the endothelial
cell-specific receptor tyrosine kinases fins-like tyrosine kinase (Flt1) and
fetal liver kinase (Flk1) (also designated KDR). Shibuya et al., Oncogene,
5: 519-524 (1990); Devries et al., Science, 255: 989-991 (1992); Quinn et
al., Proc. Natl. Acad. Sci. USA, 90: 7533-7537 (1993); Millauer et al.,
Cell, 72: 835-846 (1993); Matthews et al., Proc. Natl. Acad. Sci. USA, 88:
9026-9030 (1991); Terman et al., Biochem. Biophys. Res. Commun., 187:
1579-1586 (1992); Terman et al., Oncogene, 6: 1677-1683 (1991); Oelrichs et
al., Oncogene, 8: 11-18 (1993). These two VEGF receptors and a third orphan
receptor, Flt4 (Pajusola et al., Cancer Res., 52: 5738-5743 ; Galland
et al., Oncogene, 8: 1233-1240 ; Finnerty et al., Oncogene, 8:
2293-2298 ) constitute a subfamily of class III receptor tyrosine
kinases that contain seven extracellular immunoglobulin-like domains and a
split intracellular tyrosine kinase domain. Mustonen and Alitalo, J. Cell.
Biol., 129: 895-898 (1995). See also WO 94/10202 published May 11, 1994 and
PCT/US93/00586 filed Jan. 22, 1993 (Avraham et al.). These three receptors
have 31-36% amino acid identity in their extracellular ligand-binding
VEGF is a homodimeric, cysteine-rich protein that can occur in at least four
forms due to alternative splicing of its mRNA. Ferrara et al., Endo. Rev.,
13: 18-32 ). One example of a VEGF polypeptide that may be used in the
invention is a polypeptide of the following sequence (SEQ ID NO:9)
-- see Original Patent.
Another example of a VEGF polypeptide that may be used in the invention is
vascular endothelial growth factor B precursor, which has the following
sequence (SEQ ID NO:10)
-- see Original Patent.
Another example of a VEGF
polypeptide that may be used in the invention is vascular endothelial growth
factor C preproprotein, which has the following sequence (SEQ ID NO:11)
-- see Original Patent.
Another example of a VEGF polypeptide that may be used in the invention is
vascular endothelial growth factor D precursor, which has the following
sequence (SEQ ID NO:12)
-- see Original Patent.
As recognized by one of skill in the art, each of the PDGF, angiopoietin-2
and VEGF polypeptides from different mammalian species have similar amino
acid sequences. According to the invention any PDGF, angiopoietin-2 and VEGF
polypeptide from any mammalian species can be utilized in the practice of
the invention so long as the PDGF, angiopoietin-2 and VEGF polypeptide can
stimulate endothelial cells to promote angiogenesis.
The polypeptides of the invention, including their salts, are preferably
administered so as to achieve a reduction in at least one symptom associated
with a myocardial infarction, or a decrease in the degree or amount of
tissue injury associated with myocardial infarction or heart attack.
To achieve the desired effect(s), the polypeptides or variants thereof, may
be administered as single or divided dosages, for example, of at least about
0.01 mg/kg to about 500 to 750 mg/kg, of at least about 0.01 mg/kg to about
300 to 500 mg/kg, at least about 0.1 mg/kg to about 100 to 300 mg/kg or at
least about 1 mg/kg to about 50 to 100 mg/kg of body weight, although other
dosages may provide beneficial results. The amount administered will vary
depending on various factors including, but not limited to, the polypeptides
chosen, the disease, the weight, the physical condition, the health, the age
of the mammal, whether prevention or treatment is to be achieved, and if the
peptide is chemically modified. Such factors can be readily determined by
the clinician employing animal models or other test systems that are
available in the art.
Administration of the combination of therapeutic agents in accordance with
the present invention may be in a single dose, in multiple doses, in a
continuous or intermittent manner, depending, for example, upon the
recipient's physiological condition, whether the purpose of the
administration is therapeutic or prophylactic, and other factors known to
skilled practitioners. The administration of the polypeptides of the
invention may be essentially continuous over a pre-selected period of time
or may be in a series of spaced doses. Both local and systemic
administration is contemplated.
To prepare the composition, polypeptides are synthesized or otherwise
obtained, purified as necessary or desired and then lyophilized and
stabilized. A given polypeptide can then be adjusted to the appropriate
concentration, and optionally combined with other agents. The absolute
weight of a given polypeptide included in a unit dose can vary widely. For
example, about 0.01 to about 2 g, or about 0.1 to about 500 mg, of the
combination of polypeptides of the invention. Alternatively, the unit dosage
can vary from about 0.01 g to about 50 g, from about 0.01 g to about 35 g,
from about 0.1 g to about 25 g, from about 0.5 g to about 12 g, from about
0.5 g to about 8 g, from about 0.5 g to about 4 g, or from about 0.5 g to
about 2 g.
Daily doses of the polypeptide combination of the invention can vary as
well. Such daily doses can range, for example, from about 0.1 g/day to about
50 g/day, from about 0.1 g/day to about 25 g/day, from about 0.1 g/day to
about 12 g/day, from about 0.5 g/day to about 8 g/day, from about 0.5 g/day
to about 4 g/day, and from about 0.5 g/day to about 2 g/day.
Thus, one or more suitable unit dosage forms comprising the therapeutic
polypeptides of the invention can be administered by a variety of routes
including oral, parenteral (including subcutaneous, intravenous,
intramuscular and intraperitoneal), rectal, dermal, transdermal,
intrathoracic, intrapulmonary and intranasal (respiratory) routes. The
therapeutic polypeptides may also be formulated for sustained release (for
example, using microencapsulation, see WO 94/07529, and U.S. Pat. No.
4,962,091). The formulations may, where appropriate, be conveniently
presented in discrete unit dosage forms and may be prepared by any of the
methods well known to the pharmaceutical arts. Such methods may include the
step of mixing the therapeutic agent with liquid carriers, solid matrices,
semi-solid carriers, finely divided solid carriers or combinations thereof,
and then, if necessary, introducing or shaping the product into the desired
When the therapeutic polypeptides of the invention are prepared for oral
administration, they are generally combined with a pharmaceutically
acceptable carrier, diluent or excipient to form a pharmaceutical
formulation, or unit dosage form. For oral administration, the polypeptides
may be present as a powder, a granular formulation, a solution, a
suspension, an emulsion or in a natural or synthetic polymer or resin for
ingestion of the active ingredients from a chewing gum. The active
polypeptides may also be presented as a bolus, electuary or paste. Orally
administered therapeutic polypeptides of the invention can also be
formulated for sustained release, e.g., the polypeptides can be coated,
micro-encapsulated, or otherwise placed within a sustained delivery device.
The total active ingredients in such formulations comprise from 0.1 to 99.9%
by weight of the formulation.
By "pharmaceutically acceptable" it is meant a carrier, diluent, excipient,
and/or salt that is compatible with the other ingredients of the
formulation, and not deleterious to the recipient thereof.
Pharmaceutical formulations containing the therapeutic polypeptides of the
invention can be prepared by procedures known in the art using well-known
and readily available ingredients. For example, the polypeptide can be
formulated with common excipients, diluents, or carriers, and formed into
tablets, capsules, solutions, suspensions, powders, aerosols and the like.
Examples of excipients, diluents, and carriers that are suitable for such
formulations include buffers, as well as fillers and extenders such as
starch, cellulose, sugars, mannitol, and silicic derivatives. Binding agents
can also be included such as carboxymethyl cellulose, hydroxymethylcellulose,
hydroxypropyl methylcellulose and other cellulose derivatives, alginates,
gelatin, and polyvinyl-pyrrolidone. Moisturizing agents can be included such
as glycerol, disintegrating agents such as calcium carbonate and sodium
bicarbonate. Agents for retarding dissolution can also be included such as
paraffin. Resorption accelerators such as quaternary ammonium compounds can
also be included. Surface-active agents such as cetyl alcohol and glycerol
monostearate can be included. Adsorptive carriers such as kaolin and
bentonite can be added. Lubricants such as talc, calcium and magnesium
stearate, and solid polyethyl glycols can also be included. Preservatives
may also be added. The compositions of the invention can also contain
thickening agents such as cellulose and/or cellulose derivatives. They may
also contain gums such as xanthan, guar or carbo gum or gum arabic, or
alternatively polyethylene glycols, bentones and montmorillonites, and the
For example, tablets or caplets containing the polypeptides of the invention
can include buffering agents such as calcium carbonate, magnesium oxide and
magnesium carbonate. Caplets and tablets can also include inactive
ingredients such as cellulose, pre-gelatinized starch, silicon dioxide,
hydroxy propyl methyl cellulose, magnesium stearate, microcrystalline
cellulose, starch, talc, titanium dioxide, benzoic acid, citric acid, corn
starch, mineral oil, polypropylene glycol, sodium phosphate, zinc stearate,
and the like. Hard or soft gelatin capsules containing at least one
polypeptide of the invention can contain inactive ingredients such as
gelatin, microcrystalline cellulose, sodium lauryl sulfate, starch, talc,
and titanium dioxide, and the like, as well as liquid vehicles such as
polyethylene glycols (PEGs) and vegetable oil. Moreover, enteric-coated
caplets or tablets containing one or more polypeptides of the invention are
designed to resist disintegration in the stomach and dissolve in the more
neutral to alkaline environment of the duodenum.
The therapeutic polypeptides of the invention can also be formulated as
elixirs or solutions for convenient oral administration or as solutions
appropriate for parenteral administration, for instance by intramuscular,
subcutaneous, intraperitoneal or intravenous routes. The pharmaceutical
formulations of the therapeutic polypeptides of the invention can also take
the form of an aqueous or anhydrous solution or dispersion, or alternatively
the form of an emulsion or suspension or salve.
Thus, the therapeutic polypeptides may be formulated for parenteral
administration (e.g., by injection, for example, bolus injection or
continuous infusion) and may be presented in unit dose form in ampoules,
pre-filled syringes, small volume infusion containers or in multi-dose
containers. As noted above, preservatives can be added to help maintain the
shelve life of the dosage form. The active polypeptides and other
ingredients may form suspensions, solutions, or emulsions in oily or aqueous
vehicles, and may contain formulatory agents such as suspending, stabilizing
and/or dispersing agents. Alternatively, the active polypeptides and other
ingredients may be in powder form, obtained by aseptic isolation of sterile
solid or by lyophilization from solution, for constitution with a suitable
vehicle, e.g., sterile, pyrogen-free water, before use.
These formulations can contain pharmaceutically acceptable carriers,
vehicles and adjuvants that are well known in the art. It is possible, for
example, to prepare solutions using one or more organic solvent(s) that
is/are acceptable from the physiological standpoint, chosen, in addition to
water, from solvents such as acetone, ethanol, isopropyl alcohol, glycol
ethers such as the products sold under the name "Dowanol," polyglycols and
polyethylene glycols, C.sub.1-C.sub.4 alkyl esters of short-chain acids,
ethyl or isopropyl lactate, fatty acid triglycerides such as the products
marketed under the name "Miglyol," isopropyl myristate, animal, mineral and
vegetable oils and polysiloxanes.
It is possible to add, if necessary, an adjuvant chosen from antioxidants,
surfactants, other preservatives, film forming, keratolytic or comedolytic
agents, perfumes, flavorings and colorings. Antioxidants such as t-butylhydroquinone,
butylated hydroxyanisole, butylated hydroxytoluene and .alpha.-tocopherol
and its derivatives can be added.
Also contemplated are combination products that include one or more
polypeptides of the present invention and one or more other anti-microbial
agents. For example, a variety of antibiotics can be included in the
pharmaceutical compositions of the invention, such as aminoglycosides (e.g.,
streptomycin, gentamicin, sisomicin, tobramycin and amicacin), ansamycins
(e.g. rifamycin), antimycotics (e.g. polyenes and benzofuran derivatives),
.beta.-lactams (e.g. penicillins and cephalosporins), chloramphenical
(including thiamphenol and azidamphenicol), linosamides (lincomycin,
clindamycin), macrolides (erythromycin, oleandomycin, spiramycin),
polymyxins, bacitracins, tyrothycin, capreomycin, vancomycin, tetracyclines
(including oxytetracycline, minocycline, doxycycline), phosphomycin and
Additionally, the polypeptides are well suited to formulation as sustained
release dosage forms and the like. The formulations can be so constituted
that they release the active polypeptide, for example, in a particular part
of the intestinal or respiratory tract, possibly over a period of time.
Coatings, envelopes, and protective matrices may be made, for example, from
polymeric substances, such as polylactide-glycolates, liposomes,
microemulsions, microparticles, nanoparticles, or waxes. These coatings,
envelopes, and protective matrices are useful to coat indwelling devices,
e.g., stents, catheters, peritoneal dialysis tubing, draining devices and
For topical administration, the therapeutic agents may be formulated as is
known in the art for direct application to a target area. Forms chiefly
conditioned for topical application take the form, for example, of creams,
milks, gels, dispersion or microemulsions, lotions thickened to a greater or
lesser extent, impregnated pads, ointments or sticks, aerosol formulations
(e.g., sprays or foams), soaps, detergents, lotions or cakes of soap. Other
conventional forms for this purpose include wound dressings, coated bandages
or other polymer coverings, ointments, creams, lotions, pastes, jellies,
sprays, and aerosols. Thus, the therapeutic polypeptides of the invention
can be delivered via patches or bandages for dermal administration.
Alternatively, the polypeptide can be formulated to be part of an adhesive
polymer, such as polyacrylate or acrylate/vinyl acetate copolymer. For
long-term applications it might be desirable to use microporous and/or
breathable backing laminates, so hydration or maceration of the skin can be
minimized. The backing layer can be any appropriate thickness that will
provide the desired protective and support functions. A suitable thickness
will generally be from about 10 to about 200 microns.
Ointments and creams may, for example, be formulated with an aqueous or oily
base with the addition of suitable thickening and/or gelling agents. Lotions
may be formulated with an aqueous or oily base and will in general also
contain one or more emulsifying agents, stabilizing agents, dispersing
agents, suspending agents, thickening agents, or coloring agents. The active
polypeptides can also be delivered via iontophoresis, e.g., as disclosed in
U.S. Pat. Nos. 4,140,122; 4,383,529; or 4,051,842. The percent by weight of
a therapeutic agent of the invention present in a topical formulation will
depend on various factors, but generally will be from 0.01% to 95% of the
total weight of the formulation, and typically 0.1-85% by weight.
Drops, such as eye drops or nose drops, may be formulated with one or more
of the therapeutic polypeptides in an aqueous or non-aqueous base also
comprising one or more dispersing agents, solubilizing agents or suspending
agents. Liquid sprays are conveniently delivered from pressurized packs.
Drops can be delivered via a simple eye dropper-capped bottle, or via a
plastic bottle adapted to deliver liquid contents dropwise, via a specially
The therapeutic polypeptide may further be formulated for topical
administration in the mouth or throat. For example, the active ingredients
may be formulated as a lozenge further comprising a flavored base, usually
sucrose and acacia or tragacanth; pastilles comprising the composition in an
inert base such as gelatin and glycerin or sucrose and acacia; and
mouthwashes comprising the composition of the present invention in a
suitable liquid carrier.
The pharmaceutical formulations of the present invention may include, as
optional ingredients, pharmaceutically acceptable carriers, diluents,
solubilizing or emulsifying agents, and salts of the type that are available
in the art. Examples of such substances include normal saline solutions such
as physiologically buffered saline solutions and water. Specific
non-limiting examples of the carriers and/or diluents that are useful in the
pharmaceutical formulations of the present invention include water and
physiologically acceptable buffered saline solutions such as phosphate
buffered saline solutions pH 7.0-8.0.
The polypeptides of the invention can also be administered to the
respiratory tract. Thus, the present invention also provides aerosol
pharmaceutical formulations and dosage forms for use in the methods of the
invention. In general, such dosage forms comprise an amount of at least one
of the agents of the invention effective to treat or prevent the clinical
symptoms of a specific infection, indication or disease. Any statistically
significant attenuation of one or more symptoms of an infection, indication
or disease that has been treated pursuant to the method of the present
invention is considered to be a treatment of such infection, indication or
disease within the scope of the invention.
Alternatively, for administration by inhalation or insufflation, the
composition may take the form of a dry powder, for example, a powder mix of
the therapeutic agent and a suitable powder base such as lactose or starch.
The powder composition may be presented in unit dosage form in, for example,
capsules or cartridges, or, e.g., gelatin or blister packs from which the
powder may be administered with the aid of an inhalator, insufflator, or a
metered-dose inhaler (see, for example, the pressurized metered dose inhaler
(MDI) and the dry powder inhaler disclosed in Newinan, S. P. in Aerosols and
the Lung, Clarke, S. W. and Davia, D. eds., pp. 197-224, Butterworths,
London, England, 1984).
Therapeutic polypeptides of the present invention can also be administered
in an aqueous solution when administered in an aerosol or inhaled form.
Thus, other aerosol pharmaceutical formulations may comprise, for example, a
physiologically acceptable buffered saline solution containing between about
0.1 mg/ml and about 100 mg/ml of one or more of the polypeptides of the
present invention specific for the indication or disease to be treated. Dry
aerosol in the form of finely divided solid polypeptide that are not
dissolved or suspended in a liquid are also useful in the practice of the
present invention. Polypeptides of the present invention may be formulated
as dusting powders and comprise finely divided particles having an average
particle size of between about 1 and 5 .mu.m, alternatively between 2 and 3
.mu.m. Finely divided particles may be prepared by pulverization and screen
filtration using techniques well known in the art. The particles may be
administered by inhaling a predetermined quantity of the finely divided
material, which can be in the form of a powder. It will be appreciated that
the unit content of active ingredient or ingredients contained in an
individual aerosol dose of each dosage form need not in itself constitute an
effective amount for treating the particular infection, indication or
disease since the necessary effective amount can be reached by
administration of a plurality of dosage units. Moreover, the effective
amount may be achieved using less than the dose in the dosage form, either
individually, or in a series of administrations.
For administration to the upper (nasal) or lower respiratory tract by
inhalation, the therapeutic polypeptides of the invention are conveniently
delivered from a nebulizer or a pressurized pack or other convenient means
of delivering an aerosol spray. Pressurized packs may comprise a suitable
propellant such as dichlorodifluoromethane, trichlorofluoromethane,
dichlorotetrafluoroethane, carbon dioxide or other suitable gas. In the case
of a pressurized aerosol, the dosage unit may be determined by providing a
valve to deliver a metered amount. Nebulizers include, but are not limited
to, those described in U.S. Pat. Nos. 4,624,251; 3,703,173; 3,561,444; and
4,635,627. Aerosol delivery systems of the type disclosed herein are
available from numerous commercial sources including Fisons Corporation
(Bedford, Mass.), Schering Corp. (Kenilworth, N.J.) and American Pharmoseal
Co., (Valencia, Calif.). For intra-nasal administration, the therapeutic
agent may also be administered via nose drops, a liquid spray, such as via a
plastic bottle atomizer or metered-dose inhaler. Typical of atomizers are
the Mistometer (Wintrop) and the Medihaler (Riker).
Furthermore, the active ingredients may also be used in combination with
other therapeutic agents, for example, pain relievers, anti-inflammatory
agents, antihistamines, bronchodilators and the like, whether for the
conditions described or some other condition.
The present invention further pertains to a packaged pharmaceutical
composition for controlling microbial infections such as a kit or other
container. The kit or container holds a therapeutically effective amount of
a pharmaceutical composition for controlling cardiac damage and instructions
for using the pharmaceutical composition for reducing the effects of
myocardial infarction. The pharmaceutical composition includes at least one
polypeptide of the present invention, in a therapeutically effective amount
such that the effects of myocardial infarction are minimized.
Claim 1 of 32 Claims
1. A method of increasing survival of
blood vessels by reducing apoptosis of blood vessel cells during or after
vascular injury in a patient consisting of administering to the patient a
therapeutically effective amount of platelet derived growth factor,
angiopoietin-2, and vascular endothelial growth factor, and thereby
increasing survival of blood vessels by reducing apoptosis of blood vessel
cells in the patient during or after vascular injury.
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