U.S. patent application number 13/883521 was filed with the patent office on 2014-05-15 for treatment method for lung remodeling diseases.
The applicant listed for this patent is Kenneth E. Lipson. Invention is credited to Kenneth E. Lipson.
Application Number | 20140134181 13/883521 |
Document ID | / |
Family ID | 45044713 |
Filed Date | 2014-05-15 |
United States Patent
Application |
20140134181 |
Kind Code |
A1 |
Lipson; Kenneth E. |
May 15, 2014 |
Treatment Method For Lung Remodeling Diseases
Abstract
The present disclosure relates to methods and medicaments useful
for pretreatment, treatment, or amelioration of lung remodeling
disease. Methods and medicaments for reducing, preventing, or
reversing increased lung density, improving lung function, and
increasing survivability in subjects having lung remodeling disease
are also provided.
Inventors: |
Lipson; Kenneth E.; (San
Mateo, CA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Lipson; Kenneth E. |
San Mateo |
CA |
US |
|
|
Family ID: |
45044713 |
Appl. No.: |
13/883521 |
Filed: |
November 7, 2011 |
PCT Filed: |
November 7, 2011 |
PCT NO: |
PCT/US11/59589 |
371 Date: |
January 29, 2014 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
61456370 |
Nov 5, 2010 |
|
|
|
Current U.S.
Class: |
424/158.1 |
Current CPC
Class: |
C07K 2317/24 20130101;
A61K 45/06 20130101; C07K 2317/76 20130101; A61K 39/3955 20130101;
C07K 16/22 20130101; A61P 11/00 20180101; A61K 2039/505
20130101 |
Class at
Publication: |
424/158.1 |
International
Class: |
C07K 16/22 20060101
C07K016/22; A61K 39/395 20060101 A61K039/395; A61K 45/06 20060101
A61K045/06 |
Claims
1. A method of treating a lung remodeling disease in a subject, the
method comprising administering to the subject an anti connective
tissue growth factor (anti-CTGF) agent, wherein the lung remodeling
disease is selected from the group consisting of asthma, chronic
bronchitis, chronic obstructive pulmonary disease (COPD), cystic
fibrosis and emphysema, thereby treating the lung remodeling
disease.
2-11. (canceled)
12. The method of claim 1, wherein the anti-CTGF agent is selected
from the group consisting of anti-CTGF antibodies, antibody
fragments, antibody mimetics, antisense oligonucleotides, siRNA,
miRNA, ribozymes, aptamers and small molecules.
13. The method of claim 12, wherein the anti-CTGF agent is an
anti-CTGF antibody that binds specifically to CTGF.
14. The method of claim 13, wherein the anti-CTGF antibody has the
same amino acid sequence as the antibody produced by the cell line
identified by ATCC Accession No, PTA-6006.
15. The method of claim 13, wherein the anti-CTGF antibody binds to
CTGF competitively with an antibody produced by the cell line
identified by ATCC Accession No PTA-6006.
16-20. (canceled)
21. The method of claim 1, further comprising measuring a lung
function parameter prior to treatment.
22. The method of claim 1, further comprising measuring a lung
function parameter after treatment.
23. The method of claim 22, wherein the measured lung function
parameter is compared to a standard lung function parameter and
wherein the administration of the anti-CTGF agent is repeated if
measured lung function parameter is outside of the standard lung
function parameter.
24. The method of claim 22, wherein the standard lung function
parameter is selected from the group consisting of vital capacity
(VC), residual volume (RV), forced expiratory volume (FEV), forced
vital capacity (FVC), forced expiratory flow (FEF), maximum flow
(Vmax), peak expiratory flow rate (PEFR), inspiratory reserve
volume (IRV), functional residual capacity (FRC), inspiratory
capacity, total lung capacity (TLC), expiratory reserve volume
(ERV), tidal volume and maximum voluntary ventilation (MVV).
25. The method of claim 1, wherein the anti-CTGF agent is
administered systemically, aerosolized or nebulized.
26. The method of claim 1, wherein the method improves the
subject's lung function.
27. The method of claim 26, wherein the improvement in lung
function is determined by a decrease in lung density in the
subject.
28. The method of claim 26, wherein the improvement in lung
function is an increase in a lung function parameter selected from
forced expiratory volume, forced expiratory flow, peak expiratory
flow, tidal volume, forced vital capacity, inspiratory capacity,
inspiratory vital capacity, vital capacity, inspiratory reserve
volume and total lung capacity.
29. The method of claim 26, wherein the improvement in lung
function produces an increase in the subject's PaO.sub.2 or DLCO
value.
30-31. (canceled)
32. The method of claim 1, further comprising the administration of
an additional therapeutic agent.
33. The method of claim 32, wherein the additional therapeutic
agent is selected from the group consisting of corticosteroids,
bronchodilators, anticholinergics, vasodilators, diuretics,
anti-hypertensive agents, acetazolamide, antibiotics,
immunosuppressive drugs, surfactants, supplemental oxygen and
mechanical ventilation.
34. (canceled)
35. The method of claim 1, wherein treating a lung remodeling
disease comprises reducing lung inflammation.
Description
FIELD OF THE INVENTION
[0001] The present invention relates to methods and medicaments
useful for treatment of lung remodeling diseases. Methods and
medicaments for improving lung function, reducing lung
inflammation, and increasing survivability in subjects having a
lung remodeling disease are also provided.
BACKGROUND
[0002] Lung remodeling diseases (LRD) are a group of disorders that
lead to progressive loss of function in the alveoli. Although the
disease typically involves an initial acute inflammatory response,
many patients do not seek treatment until the disease has
progressed to a more advanced chronic phase. LRD may be due to a
number of different underlying factors. Exposure to occupational or
environmental inhalants, including inhalation of organic dust,
inorganic dust, cigarette smoke or noxious gases can often result
in LRD. First line treatment requires identification and removal of
the causative agent from the patient's environment. LRD can also be
caused by exposure to certain drugs or ionizing radiation, as may
occur during chemo- or radiation therapy in cancer patients. LRD
may also result from an exaggerated immune response, such as in
sarcoidosis, or part of a more systemic collagen vascular disorder.
In many cases, the underlying cause of LRD remains unknown.
[0003] Although the initiating agent(s) or circumstances may vary,
the immunopathogenic response of lung tissues generally involves a
similar course of events. The initial response is inflammation of
the air spaces and alveolar walls, causing an acute alveolitis. If
the condition persists, inflammation spreads to the interstitium
and vasculature of the alveoli. At early stages, the alveolar and
adjacent capillary endothelial cells become leaky, leading to
alveolar and septal edema, and the number of immune cells found in
bronchoalveolar lavage (BAL) fluid and/or sputum increases. In
particular, the number of polymorphonuclear leukocytes (PMNs),
which normally comprise about 1-3% of the cellular component of BAL
and/or sputum, can increase to 20% or more. Persistence in the
inflammatory response leads to desquamation of the wall of the
alveoli and compensatory proliferation of fibroblast in the
interstitium. The resultant scarring of lung tissue leads to
significant alterations in gas exchange and ventilatory function.
LRD can also involve the bronchioles, and patients may present with
bronchiolitis.
[0004] Current treatment options for LRD are limited and do not
provide long-term improvement in most patients. Corticosteroids
such as prednisone are often provided to reduce the inflammation
associated with LRD. However, immunosuppressant therapy can lead to
increased infection in the compromised lung and a worsening of the
condition. As the treatment options for patients with lung
remodeling disease are inadequate, new methods and medicaments for
the treatment of lung remodeling disease are therefore
desirable.
SUMMARY OF THE INVENTION
[0005] In one embodiment, the present invention provides a method
of treating a lung remodeling disease in a subject, the method
comprising administering to the subject an anti-connective tissue
growth factor (anti-CTGF) agent, thereby treating the lung
remodeling disease. In another embodiment, the present invention
provides a method for pre-treating a subject at increased
probability of being afflicted with a lung remodeling disease to
prevent or reduce a resulting pathological feature of the lung
remodeling disease, the method comprising administering to the
subject an anti-CTGF agent, thereby preventing or reducing a
resulting pathological feature of lung remodeling disease. In some
embodiments, the anti-CTGF agent is selected from the group
consisting of antibodies, antibody fragments, antibody mimetics,
antisense oligonucleotides, siRNA, miRNA, ribozymes, aptamers and
small molecules. In a preferred embodiment, the anti-CTGF agent for
use in these methods is an antibody that binds specifically to
connective tissue growth factor (CTGF). In particular embodiments,
the anti-CTGF agent is an antibody that has the same amino acid
sequence as the antibody produced by the cell line identified by
ATCC Accession No. PTA-6006, or is an antibody that binds to CTGF
competitively with an antibody produced by the cell line identified
by ATCC Accession No. PTA-6006.
[0006] Evidence is now presented that use of an anti-CTGF agent is
effective to treat various lung remodeling diseases. The results
from a radiation-induced lung injury model, typified by an acute
inflammatory response and a later chronic response that features a
progressive increase in lung density and lung remodeling (e.g.,
septal thickening), show that treatment with anti-CTGF agents is
effective to halt and, in some circumstances, reverse the increases
in lung density. Results from a neonatal hyperoxemia model,
typified by reduced alveolarization and vascularization in the
lungs, show that treatment with anti-CTGF agents is effective to
prevent or attenuate a reduction in alveolarization and
vascularization. Both models exhibit pathological remodeling in the
lung, although the remodeling manifests differently with different
results. The method of the present invention using anti-CTGF agents
is effective to treat these different manifestations of lung
remodeling. In further embodiments, the methods of the invention
are effective in preventing, reducing or reversing destruction of
alveoli in a subject in need thereof by administering to the
subject an effective amount of an anti-connective tissue growth
factor (anti-CTGF) agent, thereby preventing, reducing or reversing
destruction of alveoli. In additional embodiments, the methods of
the invention are effective in increasing alveolarization in a
subject in need thereof by administering to the subject an
effective amount of an anti-connective tissue growth factor
(anti-CTGF) agent, thereby increasing alveolarization. In other
embodiments, the methods of the invention are effective in
increasing or preserving pulmonary vascularization in a subject in
need thereof by administering to the subject an effective amount of
an anti-connective tissue growth factor (anti-CTGF) agent, thereby
increasing or preserving pulmonary vascularization.
[0007] The present invention provides methods and medicaments for
treatment of lung remodeling disease. The present invention also
provides methods and medicaments for pre-treating an individual
that has an increased probability of being afflicted with lung
remodeling disease, thereby preventing or reducing the severity of
a subsequent lung remodeling disease. In some embodiments the lung
remodeling disease is selected from the group consisting of asthma,
chronic bronchitis, chronic obstructive pulmonary disease (COPD),
sarcoidosis, hypersensitivity pneumonitis, collagen-vascular
disease, eosinophilic granuloma, nonspecific interstitial
pneumonitis, respiratory bronchiolitis-associated interstitial lung
disease, desquamative interstitial pneumonitis, lymphocytic
interstitial pneumonitis, acute interstitial pneumonitis,
bronchopulmonary dysplasia, hypersensitivity pneumonitis,
asbestosis, bauxite fibrosis, beryliosis, byssinosis, coal worker's
pneumoconiosis, cork worker's lung, farmer's lung, pigeon breeder's
lung, silo filler's disease, mushroom worker's lung, Labrador Lung
disease, siderosis, silicosis and silicosiderosis, Caplan's
syndrome, high-altitude pulmonary edema, aspiration pneumonia,
drug-induced pulmonary toxicity, lymphangioleiomyomatosis,
bronchiolitis obliterans (BOS), pediatric respiratory distress
syndrome, acute respiratory distress syndrome, hyperoxia-induced
bronchopulmonary dysplasia, cystic fibrosis and lung remodeling
associated with pulmonary hypertension and pulmonary venoocclusive
disease.
[0008] In one embodiment, the present invention provides methods
for treatment of lung remodeling disease in a subject, wherein the
method comprises administering to the subject an anti-connective
tissue growth factor (anti-CTGF) agent. In another embodiment, the
present methods can be used to pre-treat a subject at increased
probability of being afflicted with lung remodeling disease. The
methods of the present invention prevent, reduce, stabilize,
reverse or normalize various pathological features of lung
remodeling disease. Such pathological features include, but are not
limited to, decreased lung volume, decreased potential air space,
increased lung density, presence of remodeled lung tissue,
increased septal thickness, decreased alveolarization, decreased
pulmonary vascularization, increased medial wall thickness,
decreased formation or presence of secondary septa, increased right
ventricular systolic pressure (RVSP), increased right ventricular
hypertrophy (RVH), decreased PaO.sub.2 (arterial pressure of
oxygen), increased lung volume, increased presence of
polymorphonuclear leukocytes (PMNs) in bronchoalveolar lavage (BAL)
fluid and/or sputum, the presence of inflammation and the presence
of pulmonary edema. Other pathological features of lung remodeling
diseases are well known and are included here.
[0009] Thus, in one embodiment, the present methods provide a
method of reducing, stabilizing, reversing or normalizing a
pathological feature associated with lung remodeling disease in a
subject, the method comprising administering to the subject an
anti-CTGF agent, thereby reducing, stabilizing, reversing or
normalizing the pathological feature of lung remodeling disease. In
another embodiment, the present invention provides a method for
pre-treating a subject at increased probability of being afflicted
with lung remodeling disease to prevent or reduce a resulting
pathological feature of lung remodeling disease, the method
comprising administering to the subject an anti-CTGF agent, thereby
preventing or reducing a resulting pathological feature of lung
remodeling disease. In various embodiments, the pathological
feature of lung remodeling disease is selected from the group
consisting of decreased lung volume, decreased potential air space,
increased lung density, presence of remodeled lung tissue,
increased septal thickness, decreased alveolarization, decreased
pulmonary vascularization, increased medial wall thickness,
decreased formation or presence of secondary septa, increased right
ventricular systolic pressure (RVSP), increased right ventricular
hypertrophy (RVH), decreased PaO.sub.2, increased lung volume,
increased presence of polymorphonuclear leukocytes (PMNs) in
bronchoalveolar lavage (BAL) fluid and/or sputum, the presence of
inflammation and the presence of pulmonary edema.
[0010] Therefore, in one embodiment, the present invention provides
a method of preventing or stabilizing lung density in a subject
having pathologically changed lung density due to lung remodeling
disease, the method comprising administering to the subject an
anti-CTGF agent, thereby preventing or stabilizing lung density in
the subject. In another embodiment, the present invention provides
a method of reducing or normalizing a pathological change in lung
density in a subject at increased probability of being afflicted
with lung remodeling disease, the method comprising administering
to the subject an anti-CTGF agent, thereby reducing or normalizing
a pathological change in lung density in the subject. Lung density
may be measured by any method known to one of skill in the art. In
one particular aspect, the lung density is measured using lung
images from computed tomography (CT) scans; more particularly, from
high resolution CT (HRCT) scans. In some embodiments, the subject
having increased lung density has a lung density of -500 to +100
Hounsfield Units (HU). In other embodiments, the subject having
increased lung density has a lung density of -500 to -100 HU. In
still other embodiments, the subject having increased lung density
has a lung density of -500 to -300 HU. Normal healthy human lung
density varies but typically is in the range of -800 to -900 HU.
Subjects having a LRD may have a lung density that varies
substantially from the normal and may be either more or less dense
than the normal. For example, a subject having a radiation-induced
lung disorder, e.g., radiation pneumonitis, may have a lung density
that is higher than normal e.g., a LD of >-800 HU, or >-750
HU, or >-700 HU, while a subject having a LRD that is
characterized by airway remodeling, e.g., COPD or emphysema, may
have a LD that is lower than normal, e/.g a LD of <-900 HU, or
<-910 HU, or <-950 HU. The methods of the present invention
are effective to normalize the lung density (i.e., return the LD
closer to the normal range or to within the normal range) in a
subject having abnormal lung density as a result of a LRD. In some
cases, such normalizing will bring about a decrease in lung density
in a subject with increased lung density. In other cases, such
normalizing will bring about an increase in lung density in a
subject with decreased lung density.
[0011] In some embodiments, the patient's lung density is measured
prior to treatment. The measured lung density can be compared to a
standard lung density range and if the lung density is denser than
the standard lung density range, treatment with an anti-CTGF agent
can be initiated. In other embodiments, the patient's lung density
is measured after treatment. The measured density can be compared
to a standard lung density range and if the lung density is denser
than the standard density range, treatment can be repeated as
necessary to normalize the patient's lung density.
[0012] In another embodiment, the present invention provides a
method of reducing, stabilizing, reversing or normalizing lung
remodeling in a subject having lung remodeling disease, the method
comprising administering to the subject an anti-CTGF agent, thereby
reducing, stabilizing, reversing or normalizing lung remodeling in
the subject. In another embodiment, the present invention provides
a method of preventing or reducing lung remodeling in a subject at
increased probability of being afflicted with lung remodeling
disease, the method comprising administering to the subject an
anti-CTGF agent, thereby preventing or reducing lung remodeling in
the subject. Lung remodeling may be measured by any method known to
one of skill in the art. In some embodiments, lung remodeling is
measured by lung imaging from CT scans; more particularly, from
HRCT. In other embodiments, lung remodeling is measured by lung
biopsy and histology. In some embodiments, lung remodeling is
measured as a change in potential airspace in lung tissue. In
particular embodiments, the subject having a lung remodeling
disease has decreased potential airspace in lung tissue relative to
normal. In other embodiments, lung remodeling is measured by
percentage of lung showing honeycomb changes or fibroblastic foci
on lung images. In particular embodiments, the subject having lung
remodeling disease has increased percentage of honeycomb change or
fibroblastic foci in lung images.
[0013] In some embodiments, the presence and extent of lung
remodeling is determined after measuring at least one functional
lung parameters selected from the group consisting of vital
capacity (VC), residual volume (RV), forced expiratory volume
(FEV), forced vital capacity (FVC), forced expiratory flow (FEF),
maximum flow (Vmax), peak expiratory flow rate (PEFR), inspiratory
reserve volume (IRV), functional residual capacity (FRC),
inspiratory capacity, total lung capacity (TLC), expiratory reserve
volume (ERV), tidal volume and maximum voluntary ventilation (MVV).
The patient's values can then be compared against standard lung
function parameters for diagnosing the presence and extent of the
lung remodeling disease. In other embodiments, treatment with an
anti-CTGF agent improves one or more functional lung
parameters.
[0014] The measurement of a subject's PaO.sub.2, diffusing capacity
of the lung for CO (DLCO), or percent oxygen saturation of blood
are also useful in assessing lung function because typically
reduced values for functional lung parameters result in lower
PaO.sub.2, DLCO or percent oxygen saturation of blood values.
Treatment with an anti-CTGF agent can increase the PaO.sub.2, DLCO
or percent oxygen saturation of blood values of a patient with a
lung remodeling disease. In particular embodiments, improved lung
function increases or normalizes a patient's PaO.sub.2. In various
embodiments, the methods of the present invention are used to treat
a subject having a PaO.sub.2 of below 80 mmHg, particularly below
75 mmHg, and more particularly below 70 mmHg.
[0015] Measurement of one or more lung function parameters,
PaO.sub.2, DLCO or percent oxygen saturation of blood values can be
measured prior to treatment in a patient suspected of having a lung
remodeling disorder and compared to one or more standard values to
form the basis of a diagnosis. Treatment can be initiated if the
measured lung function parameters, PaO.sub.2, DLCO or percent
oxygen saturation of blood values are appreciable outside a
standard range of values, i.e., at least 5%, 10%, 15%, 20%, 25%,
30%, 40% or 50%. Further, treatment response can be monitored by
measuring one or more lung function parameters of PaO.sub.2, DLCO
or percent oxygen saturation of blood values after treatment and
comparing the measured lung function parameters to standard lung
function parameters. An increase or normalization in one or more
lung function parameters, PaO.sub.2, DLCO or percent oxygen
saturation of blood values indicate that the treatment is
efficacious. The administration of the anti-CTGF agent can be
repeated if the measured lung function parameters, PaO.sub.2, DLCO
or percent oxygen saturation of blood values are below a desired
range or appreciable outside of the standard range of values for
the lung function parameters, i.e., at least 5%, 10%, 15%, 20%,
25%, 30%, 40% or 50%.
[0016] In additional embodiments, the presence and extent of a lung
remodeling disease is measured by the cell count and cellular
composition of BAL fluid and/or sputum. In some embodiments, the
methods of the invention reduce or normalize the cellular make-up
of BAL fluid and/or sputum. In various embodiments, the methods of
the present invention are used to treat a subject having an
elevated percentage of PMNs in BAL fluid and/or sputum with the
methods of the invention reducing the percentage of PMNs in BAL
fluid and/or sputum. In particular embodiments, the subject has
greater than 5% PMNs in BAL fluid and/or sputum, particularly
greater than 10%, and more particularly greater than 15%. In
particular embodiments, the cell count and cellular composition of
BAL fluid and/or sputum is normalized over the treatment time
course.
[0017] In other embodiments, the presence and extent of a lung
remodeling disease is measured by the presence and extent of
pulmonary edema or cellular infiltration of lung tissue. In certain
instances, edema and/or cellular infiltration is an acute event,
while in other instances the edema and/or cellular infiltration is
a chronic event. Typically, cellular infiltration is associated
with the proliferation of leukocytes. In some embodiments, the
methods of the present invention prevent, reduce or reverse
pulmonary edema, cellular infiltration and/or cellular
proliferation.
[0018] In yet another embodiment, the present invention provides a
method of increasing the likelihood of survival in a subject having
lung remodeling disease, the method comprising administering to the
subject an anti-CTGF agent, thereby increasing the likelihood of
survival in the subject. In various embodiments, the subject is an
individual, preferably a mammal, more preferably a human, who has
lung remodeling disease. In other embodiments, the subject is an
individual, preferably a mammal, more preferably a human, who is at
increased probability of being afflicted with lung remodeling
disease. In certain embodiments, the subject's probability of being
afflicted with lung remodeling disease is due to an occupational
exposure to an inhalant or particulate suspected of causing lung
remodeling disease. Such inhalants or particulates may include, but
are not limited to, asbestos, organic dust, inorganic dust or smoke
inhalation. In certain embodiments, the subject's probability of
being afflicted with lung remodeling disease is due to
occupational, environmental, or therapeutic exposure to ionizing
radiation. Such exposure may be due to, e.g., radiation therapy or
nuclear medicine treatments.
[0019] In some embodiments, the methods are initiated upon
diagnosis of lung remodeling disease in a subject. In various
embodiments, the methods may be initiated prior to an event
associated with increased probability of being afflicted with lung
remodeling disease. Such an event may be an occupational event,
e.g., potential exposure to asbestos as part of an asbestos
abatement project; or a medical event, e.g., exposure to ionizing
radiation as part of radiation therapy. The method may be applied
at appropriate intervals to achieve improvement in a pathological
feature of lung remodeling disease and/or lung function as measured
by any of the parameters provided herein. In particular
embodiments, the methods may be applied 1, 2, 3, 4, or 5 time(s) or
more per month. In some embodiments, the methods are applied 1 time
per week or 1 time every other week. In various embodiments, the
methods are continued until the pathological feature or functional
parameter is essentially normalized or the subject is no longer
considered at risk.
[0020] The methods of the invention are accomplished by
administering to the subject in need an anti-CTGF agent or a
medicament comprising an anti-CTGF agent. In particular embodiments
the anti-CTGF agent is an antibody that binds specifically to CTGF,
or a polynucleotide inhibitor of CTGF expression (for example, an
antisense oligonucleotide, siRNA, shRNA, or miRNA). In a preferred
embodiment, the anti-CTGF agent is an antibody that binds
specifically to CTGF. In various embodiments, the antibody is an
antibody described and claimed in U.S. Pat. No. 7,871,617. In
particular embodiments, the antibody has the amino acid sequence of
the antibody produced by the cell line identified by ATCC Accession
No. PTA-6006. In other embodiments, the antibody competitively
binds to CTGF with an antibody produced by ATCC Accession No.
PTA-6006. A particular antibody for use in the present methods is
CLN1 or mAb1 as described in U.S. Pat. No. 7,405,274, or an
antibody substantially equivalent thereto or derived therefrom.
[0021] In another aspect, the present invention provides
medicaments for treating lung remodeling disease. In one
embodiment, the present invention provides the use of an anti-CTGF
agent in preparing a medicament for preventing, stabilizing or
reversing a pathological change in lung density in a subject having
lung remodeling disease. In another embodiment, the present
invention provides the use of an anti-CTGF agent in preparing a
medicament for preventing, stabilizing or reversing decreased
alveolarization in a subject having lung remodeling disease. In
another embodiment, the present invention provides the use of an
anti-CTGF agent in preparing a medicament for attenuating lung
remodeling in a subject having lung remodeling disease. The
medicament may be used to treat, prevent, reduce, reverse,
normalize, and/or stabilize various pathological features of lung
remodeling disease. Such features include, but are not limited to,
decreased lung volume, increased lung density, remodeled lung
tissue, decreased alveolarization, decreased vascularization in the
lung. Any anti-CTGF agent that directly inhibits the expression or
activity of CTGF may be used in formulating the present
medicaments. In particular embodiments the anti-CTGF agent is an
antibody that binds specifically to CTGF, or a polynucleotide
inhibitor of CTGF expression (for example, an antisense
oligonucleotide, siRNA, shRNA, or miRNA). In a preferred
embodiment, the anti-CTGF agent is an antibody that binds
specifically to CTGF. Any antibody that binds specifically to CTGF
may be used in formulating the present medicaments. In various
embodiments, the antibody for use in the present medicaments is an
antibody described and claimed in U.S. Pat. No. 7,871,617, which
application is incorporated in its entirety by reference herein. In
particular embodiments, the antibody has the amino acid sequence of
the antibody produced by the cell line identified by ATCC Accession
No. PTA-6006. In other embodiments, the antibody competitively
binds to CTGF with an antibody produced by ATCC Accession No.
PTA-6006. A particular antibody for use in the present medicaments
is CLN1 or mAb1 as described in U.S. Pat. No. 7,405,274, or an
antibody substantially equivalent thereto or derived therefrom.
[0022] The anti-CTGF agent can be administered systemically or
directly to the lungs as a particulate, aerosol or nebulized
solution. In further embodiments, in addition to the anti-CTGF
agent, the subject also administered a therapeutic agent selected
from the group consisting of corticosteroids, bronchodilators,
anticholinergics, vasodilators, diuretics, anti-hypertensive
agents, acetazolamide, antibiotics, immunosuppressive drugs,
surfactants, supplemental oxygen and mechanical ventilation.
[0023] These and other embodiments of the present invention will
readily occur to those of skill in the art in light of the
disclosure herein, and all such embodiments are specifically
contemplated.
BRIEF DESCRIPTION OF THE DRAWINGS
[0024] FIG. 1 sets forth various aspects of an animal model of one
type of lung remodeling disease. FIG. 1A provides a schematic of
the different dosing schedules exemplified for the present methods
and medicaments following an initial insult, i.e., exposure to
ionizing radiation. FIG. 1B sets forth the typical course of
leukocyte infiltration following initial insult. FIG. 1C sets forth
the typical course of edema and fibrosis following initial insult.
FIG. 1D sets forth the change in lung density following initial
insult.
[0025] FIG. 2 sets forth increased potential airspace in subjects
treated with the methods and medicaments of the invention. FIG. 2A
sets forth data for improved potential airspace when administration
of the present methods or medicaments was initiated 2 days before
or 2 days after the initial insult and continued for 8 weeks. FIG.
2B sets forth data for improved potential airspace when
administration of the present methods or medicaments was initiated
20 days after the initial insult and continued for 8 weeks. FIG. 2C
sets forth data for improved potential airspace when administration
of the present methods or medicaments was initiated 112 days after
the initial insult and continued for 8 weeks.
[0026] FIG. 3 sets forth reduction, prevention, or reversal in
increased lung density in subjects treated with the methods and
medicaments of the invention. FIG. 3A sets forth data for reducing
increased lung density when administration of the present methods
or medicaments was initiated 2 days before or 2 days after the
initial insult and continued for 8 weeks. FIG. 3B sets forth data
for preventing increased lung density when administration of the
present methods or medicaments was initiated 20 days after the
initial insult and continued for 8 weeks. FIG. 3C sets forth data
for preventing and/or reversing increased lung density when
administration of the present methods or medicaments was initiated
112 days after the initial insult and continued for 8 weeks.
[0027] FIG. 4 sets forth improved blood gas parameters in subjects
treated with the methods and medicaments of the invention. FIG. 4A
sets forth blood partial oxygen pressure (PaO.sub.2) at week 30
following the initial insult for animals that were untreated or
treated with the methods and medicaments of the present invention.
The striped box in the figure defines the normal range for
PaO.sub.2 in this model organism and demonstrates that treatment
with an anti-CTGF agent can normalize PaO.sub.2 values in treated
animals. FIG. 4B plots blood PaO.sub.2 and lung density at week 30
following the initial insult for animals that were untreated or
treated with the methods and medicaments of the present invention,
showing that improved lung function correlates inversely with lung
density. The shaded region in the figure defines the normal oxygen
saturation range in this model organism.
[0028] FIG. 5 sets forth the survival rates in subjects treated
with the methods and medicaments of the invention. FIG. 5A sets
forth data for increased survivability when administration of the
present methods or medicaments was initiated 2 days before or 2
days after the initial insult and continued for 8 weeks. FIG. 5B
sets forth data for increased survivability when administration of
the present methods or medicaments was initiated 20 days after the
initial insult and continued for 8 weeks. FIG. 5C sets forth data
for increased survivability when administration of the present
methods or medicaments was initiated 112 days after the initial
insult and continued for 8 weeks.
[0029] FIG. 6 illustrates the ability of an anti-CTGF antibody
(Ab3149) to increase secondary septal count in animals exposed to
hyperoxia. n=5/group. *P<0.001 and ** P<0.01 compared to
normoxia lungs.
[0030] FIG. 7 illustrates the ability of an anti-CTGF antibody
(Ab3149) to reduce the increase in lung macrophages induced by
hyperoxia.
DESCRIPTION OF THE INVENTION
[0031] It is to be understood that the invention is not limited to
the particular methodologies, protocols, cell lines, assays, and
reagents described herein, as these may vary. It is also to be
understood that the terminology used herein is intended to describe
particular embodiments of the present invention, and is in no way
intended to limit the scope of the present invention as set forth
in the appended claims.
[0032] Each of the limitations of the invention can encompass
various embodiments of the invention. It is, therefore, anticipated
that each of the limitations of the invention involving any one
element or combinations of elements can be included in each aspect
of the invention. This invention is not limited in its application
to the details of construction and the arrangement of components
set forth in the following description or illustrated in the
drawings. The invention is capable of other embodiments and of
being practiced or of being carried out in various ways.
[0033] It must be noted that as used herein and in the appended
claims, the singular forms "a," "an," and "the" include plural
references unless context clearly dictates otherwise. Thus, for
example, a reference to "a fragment" includes a plurality of such
fragments; a reference to an "antibody" is a reference to one or
more antibodies and to equivalents thereof known to those skilled
in the art, and so forth.
[0034] Unless defined otherwise, all technical and scientific terms
used herein have the same meanings as commonly understood by one of
ordinary skill in the art to which this invention belongs. Although
any methods and materials similar or equivalent to those described
herein can be used in the practice or testing of the present
invention, the preferred methods, devices, and materials are now
described. All publications cited herein are incorporated herein by
reference in their entirety for the purpose of describing and
disclosing the methodologies, reagents, and tools reported in the
publications that might be used in connection with the present
invention. Nothing herein is to be construed as an admission that
the present invention is not entitled to antedate such disclosure
by virtue of prior invention.
[0035] The practice of the present invention will employ, unless
otherwise indicated, conventional methods of chemistry,
biochemistry, molecular biology, cell biology, genetics, immunology
and pharmacology, within the skill of the art. Such techniques are
explained fully in the literature. See, e.g., Gennaro, A. R., ed.
(1990) Remington's Pharmaceutical Sciences, 18th ed., Mack
Publishing Co.; Colowick, S. et al., eds., Methods In Enzymology,
Academic Press, Inc.; Handbook of Experimental Immunology, Vols.
I-IV (D. M. Weir and C. C. Blackwell, eds., 1986, Blackwell
Scientific Publications); Maniatis, T. et al., eds. (1989)
Molecular Cloning: A Laboratory Manual, 2nd edition, Vols. I-III,
Cold Spring Harbor Laboratory Press; Ausubel, F. M. et al., eds.
(1999) Short Protocols in Molecular Biology, 4th edition, John
Wiley & Sons; Ream et al., eds. (1998) Molecular Biology
Techniques: An Intensive Laboratory Course, Academic Press); PCR
(Introduction to Biotechniques Series), 2nd ed. (Newton &
Graham eds., 1997, Springer Verlag).
[0036] In one aspect, the present invention provides methods for
treating lung remodeling disease. As used herein, "lung remodeling
disease" or "LRD" refers to a group of lung diseases affecting the
alveoli and interstitium that result in progressive scarring and
loss of function. LRD may be caused by exposure to a number of
toxins or pollutants including, but not limited to, inorganic dust
such as silica or hard metal dust; asbestos fibers, noxious gases
such as ammonia or chlorine gas; organic particles from various
sources such as grain, sugar cane, and bird and animal droppings,
moldy hay, and bacterial or fungal growths. LRD may also be caused
by thoracic exposure to radiation or certain drugs such as
chemotherapy drugs, various heart medications, certain psychiatric
medications, and some antibiotics. In many cases, the cause of LRD
is unknown and LRD is identified as idiopathic. Diagnosis of LRD is
based on symptoms including dyspnea, nonproductive cough, fatigue,
excessive mucus and/or sputum production, chest tightness, wheezing
and low-grade fever; and generally involves blood tests, e.g.,
measurement of partial oxygen saturation of the blood; pulmonary
function tests, e.g., measurement of total lung volume, residual
volume, and vital capacity; and computed tomography (CT) scans of
the thorax, e.g., to measure lung density and monitor lung
remodeling.
[0037] Lung remodeling diseases (LRD) include diseases, disorders
and pathological conditions in which pathological or maladaptive
remodeling occurs in the lung tissues, including bronchi,
bronchioles, alveoli, airways, interstitium and pulmonary
vasculature. LRD can include airway diseases like asthma and
chronic bronchitis and lung tissue diseases like sarcoidosis. Other
lung remodeling diseases include interstitial lungs diseases, e.g.,
hypersensitivity pneumonitis, collagen-vascular disease,
eosinophilic granuloma, nonspecific interstitial pneumonitis,
respiratory bronchiolitis-associated interstitial lung disease,
desquamative interstitial pneumonitis, lymphocytic interstitial
pneumonitis, acute interstitial pneumonitis. Numerous lung
remodeling diseases are associated with occupational exposure to
particulates including asbestosis, bauxite fibrosis, beryliosis,
byssinosis, coal worker's pneumoconiosis, cork worker's lung,
farmer's lung, mushroom worker's lung; Labrador Lung disease,
pigeon breeder's lung, siderosis, silicosis, silo filler's disease
and silicosiderosis. Certain lung remodeling diseases are
associated with autoimmune diseases including, collagen-vascular
diseases, e.g., systemic lupus erythematosus, rheumatoid arthritis
(Caplan's syndrome), progressive systemic sclerosis, scleroderma,
dermatomyositis, polymyositis, ankylosing spondylitis, Sjogren
syndrome and mixed connective-tissue disease, or genetic diseases
e.g., cystic fibrosis and alpha-1 antitrypsin deficiency. Other
lung remodeling diseases include bronchiectasis, emphysema, COPD;
diseases associated with barotrauma and mechanical ventilation;
high-altitude pulmonary edema, aspiration pneumonia, drug-induced
pulmonary toxicity, lymphangioleiomyomatosis, lung transplantation
rejection, e.g., bronchiolitis obliterans (BOS), pediatric and
adult respiratory distress syndrome, hyperoxia-induced
bronchopulmonary dysplasia, and remodeling associated with
pulmonary hypertension and pulmonary venoocclusive disease.
[0038] In some embodiments, the subject in need of treatment is an
individual, preferably a mammal, more preferably a human, who has
been diagnosed with a lung remodeling disease. In other
embodiments, the subject is an individual, preferably a mammal,
more preferably a human, who is at increased probability or
likelihood of being afflicted with a lung remodeling disease. In
certain embodiments, the subject's probability or likelihood of
being afflicted with a lung remodeling disease is due to exposure
to an inhalant suspected of causing lung remodeling disease, as
described supra. In other embodiments, the subject's probability or
likelihood of being afflicted with lung remodeling disease is due
to a treatment with a drug such as those described infra. In
certain embodiments, the subject's probability or likelihood of
being afflicted with lung remodeling disease is due to an
underlying condition associated with LRD, e.g., systemic lupus
erythematosus.
[0039] Lung remodeling diseases can be classified into three
categories: lung remodeling diseases associated with a decrease in
lung volume or a restriction in lung expansion; lung remodeling
diseases associated with an increase in lung volume; and lung
remodeling diseases associated with constricted or blocked airways.
Frequently, a lung remodeling disease will feature both a
pathological change in lung volume plus constricted or blocked
airways.
[0040] In some embodiments, the subject in need of treatment has a
lung remodeling disease that is associated with in a decrease in
lung volume or a restriction in lung expansion. Non-limiting
examples of lung remodeling diseases that decrease a subject's lung
volume or restrict the expansion of the subject's lung include:
asthma, chronic bronchitis, COPD, sarcoidosis, interstitial lungs
diseases, e.g., hypersensitivity pneumonitis, collagen-vascular
disease, eosinophilic granuloma, nonspecific interstitial
pneumonitis, bronchiolitis obliterans with organizing pneumonia,
respiratory bronchiolitis-associated interstitial lung disease,
desquamative interstitial pneumonitis, lymphocytic interstitial
pneumonitis, acute interstitial pneumonitis, bronchopulmonary
dysplasia, restrictive lung disease; occupational lung diseases,
e.g., asbestosis, bauxite fibrosis, beryliosis, byssinosis, coal
worker's pneumoconiosis, hypersensitivity pneumonitis: including
cork worker's lung, farmer's lung, pigeon breeder's lung, silo
filler's disease and mushroom worker's lung; Labrador Lung disease,
siderosis, silicosis and silicosiderosis; Caplan's syndrome,
diseases associated with barotrauma and mechanical ventilation;
high-altitude pulmonary edema, aspiration pneumonia, drug-induced
pulmonary toxicity, lymphangioleiomyomatosis, lung transplantation
rejection, e.g., bronchiolitis obliterans (BOS), pediatric and
adult respiratory distress syndrome, acute respiratory distress
syndrome, hyperoxia-induced bronchopulmonary dysplasia, and
remodeling associated with pulmonary hypertension and pulmonary
venoocclusive disease and cystic fibrosis.
[0041] In some embodiments, a lung remodeling disease that is
associated with in a decrease in lung volume or a restriction in
lung expansion is not idiopathic pulmonary fibrosis. In other
embodiments, a lung remodeling disease that is associated with in a
decrease in lung volume or a restriction in lung expansion is not
bleomycin pulmonary toxicity.
[0042] In further embodiments, the drug associated with
drug-induced pulmonary toxicity that results in a reduction of lung
volume or lung function is selected from the list consisting of
5-fluorouracil (5FU), abacavir, abciximab, acebutolol,
acetaminophen, acetazolamide, acetylcysteine, acetylsalicylic acid,
acrylate, acyclovir, adalimumab, adenosine and derivatives,
albumin, alemtuzumab, allopurinol, almitrine, alteplase,
amifostine, aminoglutethimide, aminoglycosid antibiotics, aminorex,
amiodarone, amitriptyline, amlodipine, amphotericin B, ampicillin,
amrinone, anagrelide, anastrozole, angiotensin converting enzyme
inhibitors, antazoline, anti TNF-alpha, non-steroidal
anti-inflammatory drugs, anti-lymphocyte (thymocyte) globulin,
anticoagulants, antidepressants, aprotinin, arsenic trioxide,
atenolol, atociban, aurothiopropanosulfonate, azacitidine,
azapropazone, azathioprine, azithromycin, balsazalide,
barbiturates, basiliximab, BCG therapy, beclomethasone, benfluorex,
benzbromarone, benzocaine, benzylthiouracil, bepridil,
beta-agonists, beta-blockers, betahistine, betaxolol, bevacizumab,
bicalutamide, blood transfusions, bortezomib, botulinum toxin,
bromocriptine, bucillamine, bumetanide, buprenorphine, bupropion,
busulfan, cabergoline, calcium salts, camptothecin, captopril,
carbamazepine, carbimazole, carmustine (BCNU), carvedilol,
cefotiam, celecoxib, celiprolol, cephalexin, cephalosporins,
cetuximab, chlorambucil, chlorhexidine, chloroquine, chlorozotocin
(DCNU), chlorpromazine, chlorpropamide, cilazapril, ciprofloxacin,
cisapride, cisplatin, citalopram, cladribine, clarithromycin,
clindamycin, clofazimine, clofibrate, clomiphene, clomipramine,
clonidine, clopidogrel, clozapine, codeine, colchicine,
contraceptives (oral), cotrimoxazole, cromoglycate,
cyclophosphamide, cyclosporine, cyproterone acetate, cytarabine
(cytosine arabinoside), cytokines, D-tubocurarine, danazol,
dantrolene, dapsone, daptomycin, darbepoetin, dasatinib,
daunorubicin, deferoxamine, desipramine, dexamethasone, dextran,
diclofenac, diflunisal, dihydralazine, dihydro-5-azacytidine,
dihydroergocristine, dihydroergocryptine, dihydroergotamine,
diltiazem, dimethylsulphoxide, dipyridamole, disopyramide,
divalproex sodium, docetaxel, dothiepin, doxorubicin, duloxetine,
efavirenz, enalapril, epoprostenol, epinephrine, ergometrine,
ergotamine, erlotinib, erythromycin, estrogens, etanercept,
ethambutol, ethchlorvynol, ethosuximide, etoposide, etretinate,
everolimus, Factor VIIa, famotidine, febarbamate, fenbufen,
fenfluramine/dexfenfluramine, fenoprofen, fentanyl, fibrinolytics
(including rTPA), flecainide, floxuridine, fludarabine,
fluoresceine, fluoxetine, flurbiprofen, flutamide, fluticasone,
fluvastatin, folinic acid, fosinopril, fotemustine, furazolidone,
G-CSF, GM-CSF, gefitinib (ZD1839), gemcitabine, gemtuzumab,
glafenine, glibenclamide, gliclazide, gold salts, gonadotrophin,
goserelin, haloperidol, heparin, heroin, hexamethonium,
hydralazine, hydrochlorothiazide, hydrocortisone,
hydroxyquinoleine, hydroxyurea, ibuprofen, ifenprodil, ifosfamide,
imatinib, imidapril, imiglucerase, imipramine, immunoglobulins
(IV), indinavir, indomethacin, infliximab, insulin, interferon
alfa, interferon beta, interferon gamma, interleukin 2, iodine and
iodinated radiographic contrast media, irinotecan, isoflurane,
isoniazide, isotretinoin, itraconazole, ketamine, ketorolac,
L-asparaginase, L-ryptophan, labetalol, lansoprazole, latanoprost
leflunomide, lenalidomide, leukotrien receptor ntagonists,
leuprorelin (leuprolide), levodopa, levofloxacin, levomepromazine,
lidocaine, lipids, lisinopril, lisuride, lomustine (CCNU),
losartan, loxoprofen, maprotiline, mazindol, mecamylamine,
medroxyprogesterone, mefloquine, meloxicam, melphalan, mephenesin,
mephenytoin, mercaptopurine, mesalamine (mesalazine, 5-ASA),
mesulergine, metamizole (noramidopyrine), metapramine, metformine,
methadone, methenolone, methimazole, methotrexate, methyldopa,
methylphenidate, methylprednisolone, methysergide, metipranolol,
metoclopramide, metoprolol, metronidazole, mexiletine, miconazole,
midazolam minocycline, minoxidil, misoprostol, mitomycin C,
mitoxantrone, montelukast, morphine, moxalactam, mycophenolate
mofetil, nabumetone, nadolol, naftidrofuryl, nalbuphine, nalidixic
acid, naloxone, naphtazoline, naproxen, nevirapine, nicardipine,
nicergoline, niflumic acid, nilutamide, nimesulide, niridazol,
nitric oxide, nitrofurantoin, nitroglycerin, nitrosoureas,
nomifensine, OKT3, olsalazine, omalizumab, omeprazole, ornipressin,
oxaliplatin, oxprenolol oxycodone, oxyphenbutazone, oxytocin,
paclitaxel, pamidronate, para-(4)-aminosalicylic acid, paraffin
(mineral oil), paraldehyde, parenteral nutrition, paroxetine,
patent blue, pemetrexed, penicillamine, penicillins, pentamidine,
pergolide, perindopril, phenazopyridine, phenylbutazone,
phenylephrine, phenylpropanolamine, phenytoin (diphenylhydantoin),
pindolol, pioglitazone, piperacilline, piroxicam, pituitary snuff,
platinum salts, polyethylene glycol, practolol, pranlukast,
pranoprofen, pravastatin, praziquantel, procainamide, procarbazine,
progesterone, propafenone, propofol,
propoxyphene/dextropropoxyphene, propranolol, propylene glycol,
propylthiouracil, prostacyclin, prostaglandin E1, prostaglandin
F2-alpha, protamine, pyrimethamine-dapsone,
pyrimethamine-sulfadoxine, quinidine, quinine, raloxifene,
raltitrexed, ranitidine, retinoic acid, rifampicin, riluzole,
risedronate, risperidone, ritodrine, rituximab, rofecoxib,
rosiglitazone, roxithromycin, salbutamol (Albuterol), serrapeptase,
sertraline, sildenafil, simvastatin, sirolimus, sorafenib, sotalol,
statins, steroids, streptokinase, streptomycin, sufentanil,
sulfamides-sulfonamides, sulfasalazine (Salazopyrine), sulindac,
sulprostone, tacrolimus (FK506), tamoxifen, telithromycin,
telmisartan, temozolomide, temsirolimus, tenidap, terbutaline,
tetracycline, thalidomide, thiazolidinediones, thiopental,
tiaprofenic acid, ticlopidine, tiopronin, tirofiban, tizanidine,
TNF-alpha, tocainide, tocilizumab, tolazamide, tolfenamic acid,
topiramate, topotecan, tosufloxacin, tramadol, tranexamic acid,
trastuzumab, trazodone, triazolam, trimipramine, triptans,
trofosfamide, troglitazone, troleandomycin, trokinase, valproic
acid, valsartan, vancomycin, vasopressin, venlafaxine, verteporfin,
vinblastine, vindesine, vinorelbine, vitamin D, zafirlukast,
zanamivir, zinostatin and zomepirac.
[0043] In some embodiments, the method for treating a lung
remodeling disease that is associated with a decrease in lung
volume or a restriction in lung expansion is capable of increasing
the effective lung volume by at least 5%, 10%, 15%, 20%, 25%, 30%,
40%, 50%, 60% or 75% compared to a baseline measurement taken
before treatment with an anti-CTGR agent. In further embodiments,
the method for treating a lung remodeling disease associated with a
decrease in lung volume or a restriction in lung expansion is
capable of increasing a patient's FEV.sub.1 or FEV.sub.6 value by
at least 5%, 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60% or 75% compared
to a baseline measurement of the patient's FEV.sub.1 or FEV.sub.6
before treatment with the anti-CTGF agent.
[0044] In other embodiments, the subject in need of treatment has a
lung remodeling disease that results in an increase in lung volume
(airspace enlargement). Non-limiting examples of lung remodeling
diseases that feature an increase in a subject's lung volume
include: emphysema, COPD; hyperoxia-induced bronchopulmonary
dysplasia and remodeling associated with pulmonary hypertension and
pulmonary venoocclusive disease. In some embodiments, the
enlargement of the air spaces occurs across the entire respiratory
acinus. In other embodiments, the enlargement of the air spaces
occur from the respiratory bronchiole to the terminal bronchioles.
In additional embodiments, the enlargement of the air spaces is
associated with the destruction of the air space walls. In further
embodiments, the enlargement of air spaces is associated with a
reduction in lung elasticity.
[0045] In some embodiments, the method for treating a lung
remodeling disease that is associated with an increase in lung
volume is capable of decreasing the effective lung volume by at
least 5%, 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60% or 75% compared to
a baseline measurement taken before treatment with an anti-CTGF
agent. In further embodiments, the method for treating a lung
remodeling disease that is associated with an increase in lung
volume is capable of increasing a patient's FEV.sub.1 or FEV.sub.6
value by at least 5%, 10%, 15%, 20%, 25%, 30%, 40%, 50%, 60% or 75%
compared to a baseline measurement of the patient's FEV.sub.1 or
FEV.sub.6 before treatment with the anti-CTGF agent.
[0046] In further embodiments, the subject in need of treatment has
a lung remodeling disease that is associated with constricted or
blocked airways. Typically, constricted or blocked airways produce
a shortness of breath. Frequently, a lung remodeling disease
associated with constricted or blocked airways features excess
mucus production that in severe cases occlude the small bronchi.
Non-limiting examples of lung remodeling diseases associated with
constricted or blocked airways include: astluna, COPD and chronic
bronchitis. Typically, patients with a lung remodeling diseases
associated with constricted or blocked airways have abnormally low
FEV.sub.1 or FEV.sub.6 values. In some embodiments, the method for
treating a lung remodeling diseases associated with constricted or
blocked airways is capable of increasing a patient's FEV.sub.1 or
FEV.sub.6 value by at least 5%, 10%, 15%, 20%, 25%, 30%, 40%, 50%,
60% or 75% compared to a baseline measurement of the patient's
FEV.sub.1 or FEV.sub.6 before treatment with the anti-CTGF
agent.
[0047] Frequently, lung remodeling diseases have an acute and/or
chronic inflammatory phase. In many instances the lung inflammation
is not associated with infection (pneumonitis). Usually,
inflammation is associated with edema and an influx of immune cells
such as mast cells, eosinophils, neutrophils and T lymphocytes.
Typically, lung remodeling is associated with chronic inflammation
that may affect different parts of the lung, including the walls of
the bronchioles (bronchiolitis); the walls and air spaces of the
alveoli (alveolitis); the small blood vessels (vasculitis).
Remodeling associated with inflammation may include thickening of
the lamina reticularis (the subepithelial reticular basement
membrane surrounding airways), thickening of the airway wall,
epithelial denudation, hypertrophy of the large bronchi, goblet
cell hyperplasia, bronchial wall edema and narrowed small
airways.
[0048] In some embodiments, the methods of the invention increase
the elasticity of lung alveoli. In other embodiments, the methods
of the invention prevent or reduce the incident of bullae
formation. In other embodiments, the methods of the invention
increase bulk air flow exchange. In further embodiments, the
methods of the invention prevent, reduce or revert partial airway
collapse. In additional embodiments, the methods of the invention
prevent, reduce or revert, bronchial hypertrophy. In other
embodiments, the methods of the invention preserve or increase
alveoli density. In further embodiments, the methods of the
invention prevent or reduce airway hyperresponsiveness (AHR). COPD
and emphysema are lung remodeling diseases that are characterized
by loss of alveolar elasticity and obstruction of bronchi and
bronchioles. It has been shown that cigarette smoke, a known cause
of these diseases, induces CTGF expression (Churg et al. (2006) Am
J Resp Crit Care Med 174:1327-1334). The fact that early events in
hyperoxia-induced neonatal lung disease resembles the lung
remodeling that occurs in COPD and emphysema, as well as in other
airway disease (asthma), suggest that anti-CTGF agents can be used
effectively to treat lung remodeling diseases generally.
Anti-CTGF Agents
[0049] The methods of the invention are accomplished by
administering to a subject an anti-CTGF agent. As used herein, the
term "anti-CTGF agent" refers to any agent, molecule,
macromolecule, compound, or composition that inhibits, reduces, or
stops the activity, function, production or expression of
connective tissue growth factor. Anti-CTGF agents include
antibodies, antibody fragments, antibody mimetics, antisense
oligonucleotides, siRNA, miRNA, ribozymes, aptamers and small
molecules. The anti-CTGF agent is preferably one that is specific
for CTGF and exerts its effect directly and specifically on the
CTGF protein or on the CTGF gene or mRNA, rather than a
non-specific inhibitor (e.g., a non-specific protease or
transcription inhibitor) or an indirect inhibitor (e.g., an
inhibitor of an upstream or downstream signaling pathway). As used
herein, "connective tissue growth factor" and "CTGF" refer to a
matricellular protein belonging to a family of proteins identified
as CCN proteins. CTGF may also be referred to within the art as
"hypertrophic chondrocyte-specific protein 24," "insulin-like,
growth factor-binding protein," and "CCN2."
[0050] In some embodiments, an antibody that binds specifically to
CTGF may be used in the present methods. In some embodiments, the
antibody for use in the present methods is obtained from the same
species as the subject in need. In other embodiments, the antibody
is a chimeric antibody wherein the constant domains are obtained
from the same species as the subject in need and the variable
domains are obtained from another species. For example, in treating
a human subject the antibody for use in the present methods may be
a chimeric antibody having constant domains that are human in
origin and variable domains that are mouse in origin. In preferred
embodiments, the antibody for use in the present methods binds
specifically to the CTGF endogenous to the species of the subject
in need. Thus, in certain embodiments, the antibody is a human or
humanized antibody, particularly a monoclonal antibody, that
specifically binds human CTGF (GenBank Accession No.
NP.sub.--001892.1). In particular embodiments, the antibody is the
antibody described and claimed in U.S. Pat. No. 7,871,617. In some
embodiments, the antibody has the amino acid sequence of the
antibody produced by the cell line identified by ATCC Accession No.
PTA-6006. In other embodiments, the antibody competitively binds to
CTGF with an antibody produced by ATCC Accession No. PTA-6006. A
particular antibody for use in the present methods is CLN1 or mAb1
as described in U.S. Pat. No. 7,405,274, or an antibody
substantially equivalent thereto or derived therefrom. An antibody
for use in the present methods may also be a fragment such as a
Fab, F(ab)2, Fv, or single chain variable fragment (scFV) of any
antibody described above. An antibody for use in the present
methods may also be derived from any antibody described above. Such
derivatives may include any suitable antibody derivation known to
those of skill in the art and include, but are not limited to,
diabodies, triabodies, and minibodies.
[0051] Additionally, antibodies for use in the present methods also
include antibody mimetics. Antibody mimetics are proteins,
typically in the range of 3-25 kD that are designed to bind an
antigen with high specificity and affinity like an antibody, but
are structurally unrelated to antibodies. Frequently, antibody
mimetics are based on a structural motif or scaffold that can be
found as a single or repeated domain from a larger biomolecule.
Examples of domain derived antibody mimetics included AdNectins
that utilize the 10th fibronectin III domain (Lipov{hacek over
(s)}ek D. Protein Eng Des Sel, 2010, 24:3-9); Affibodies that
utilize the Z domain of staphylococcal protein A (Nord K et al. Nat
Biotechnol. 1997, 15: 772-777) and DARPins that utilize the
consensus ankyrin repeat domain (Amstutz P. Protein Eng Des Sel.
2006, 19:219-229. Alternatively, antibody mimetics can also be
based on the entire structure of a smaller biomolecule, such as
Anticalins that utilize the lipocalin structure (Beste G et al.
Proc Natl Acad Sci USA. 1999, 5:1898-1903)
[0052] As referred to herein, the phrase "an antibody that
specifically binds to CTGF" includes any antibody that binds to
CTGF with high affinity. Affinity can be calculated from the
following equation:
Affinity = K a = [ Ab Ag ] [ Ab ] [ Ag ] = 1 K d ##EQU00001##
where [Ab] is the concentration of the free antigen binding site on
the antibody, [Ag] is the concentration of the free antigen, [AbAg]
is the concentration of occupied antigen binding sites, K.sub.a is
the association constant of the complex of antigen with antigen
binding site, and K.sub.d is the dissociation constant of the
complex.
[0053] A high-affinity antibody typically has an affinity at least
on the order of 10.sup.8M.sup.-1, 10.sup.9 M.sup.-1 or 10.sup.10 In
particular embodiments, an antibody for use in the present methods
will have a binding affinity for CTGF between of 10.sup.8 M.sup.-1
and 10.sup.10 M.sup.-1, between 10.sup.8 M.sup.-1 and 10.sup.9
M.sup.-1 or between 10.sup.9M.sup.-1 and 10.sup.10 M.sup.-1. In
some embodiments the high-affinity antibody has an affinity of
about 10.sup.8M.sup.-1, 10.sup.9 M.sup.-1 or 10.sup.10
M.sup.-1.
[0054] In some embodiments, polynucleotide inhibitors of CTGF,
including small interfering ribonucleic acids (siRNAs), micro-RNAs
(miRNAs), and antisense sequences may be used in the present
methods to inhibit expression and/or production of CTGF. (See,
e.g., Kondo et al. (2000) Biochem Biophys Res Commun 278:119-124.)
Such techniques are well-known to those of skill in the relevant
art. Polynucleotide inhibitors that target CTGF expression have
been described and utilized to reduce CTGF expression in various
cell types. (See, e.g., International Publication No. WO 96/38172;
International Publication No. WO 00/27868; International
Publication No. WO 00/35936; International Publication No. WO
03/053340; Kothapalli et al. (1997) Cell Growth Differ 8:61-68;
Shimo et al. (1998) J Biochem (Tokyo) 124:130-140; Uchio et al.
(2004) Wound Repair Regen 12:60-66; Guha et al. (2007) FASEB J
21:3355-3368; U.S. Pat. No. 6,358,741; U.S. Pat. No. 6,965,025;
U.S. Pat. No. 7,462,602; U.S. Patent Application Publication No.
2008/0070856; U.S. Patent Application Publication No.
2008/0176964.) CTGF antisense constructs and other types of
polynucleotide inhibitors of CTGF can be used to reduce expression
of CTGF and thereby treat lung remodeling disease. Such constructs
can be designed using appropriate vectors and expressional
regulators for cell- or tissue-specific expression and constitutive
or inducible expression. Such genetic constructs can be formulated
and administered according to established procedures within the
art. The polynucleotide inhibitors used in the present methods and
medicaments may be made using solid phase synthesis techniques
known to those of skill in the art and available through various
vendors including GE Healthcare Biosciences (Piscataway, N.J.). Any
other means for such synthesis known in the art may additionally or
alternatively be employed.
Pharmaceutical Formulations and Routes of Administration
[0055] The anti-CTGF agents used in the methods of the present
invention can be delivered directly or in pharmaceutical
compositions containing excipients, as is well known in the art. An
effective amount of anti-CTGF agent can readily be determined by
routine experimentation, as can an effective and convenient route
of administration and an appropriate formulation. Various
formulations and drug delivery systems are available in the art.
(See, e.g., Gennaro, ed. (2000) Remington's Pharmaceutical
Sciences, supra; and Goodman and Gilman's The Pharmacological Basis
of Therapeutics, 10.sup.th Ed. (2001), Hardman, Limbird, and
Gilman, eds. MacGraw Hill Intl.) Typically, the anti-CTGF agent is
administered in an amount sufficient to provide therapeutic
efficacy over the treatment time course. Therapeutic efficacy can
be measured using any parameter provided herein, including
improvement in any pathological feature of lung remodeling disease
and/or improvement in lung function.
[0056] Therefore, in one embodiment, the present invention provides
methods for treatment of lung remodeling disease in a subject,
wherein the method comprises administering to the subject an
anti-CTGF agent. In another embodiment, the present invention
provides methods for pre-treating a subject at increased
probability of being afflicted with lung remodeling disease,
wherein the method comprises administering to the subject an
anti-CTGF agent. The methods of the present invention prevent,
reduce, stabilize, normalize or reverse various pathological
features of lung remodeling disease. Such features include, but are
not limited to, decreased lung volume, increased lung density,
remodeled lung tissue, and increased mortality. Thus, in one
embodiment, the present methods provide a method of reducing,
stabilizing or normalizing a pathological feature associated with
lung remodeling disease in a subject, the method comprising
administering to the subject an anti-CTGF agent, thereby reducing,
stabilizing or normalizing the pathological feature of lung
remodeling disease. Stabilizing of pathological features is defined
as reducing, slowing or freezing the rate of decline of a
pathological change in a lung tissue or in a lung parameter. In
some embodiments, stabilization is the reduction in the rate at
which a particular lung function is loss. In certain embodiments,
stabilization reduces the rate of a pathological change in a
particular lung function by at least 5%, 10%, 15%, 20%, 25%, 30%,
35%, 40%, 45%, 50%, 75% or 100% compared to the expected rate of
change based on a control population or a historical population. In
other embodiments, stabilization results in a halting of the rate
of change in a particular lung function by the pathologic process
thereby halting the deterioration in the patient's lung function.
In other embodiments, normalization is the return to a normal range
of a characteristic of a lung tissue or lung function. In some
embodiments, normalization results in the characteristic of the
lung tissue or lung parameter being within the normal range or less
than 5%, 10%, 15% or 20% away from the normal range. For instance,
a patient with a lung remodeling disease that had an initial lung
density of -300 HU that increased to a lung density within the
normal range of -800 to -900 HU after therapy with an anti-CTGF
agent would be classified as having normalized lung density.
Similarly, a patient with a lung remodeling disease that had an
initial PaO.sub.2 of 68% that increased to a PaO.sub.2 of 81% after
treatment with an anti-CTGF agent would be classified as having a
normalized PaO.sub.2 as a value of 80% or above is within the
normal range, see infra.
[0057] In another embodiment, the present invention provides a
method for pre-treating a subject at increased probability of being
afflicted with lung remodeling disease to prevent or reduce a
resulting pathological feature of lung remodeling disease, the
method comprising administering to the subject an anti-CTGF agent,
thereby preventing or reducing a resulting pathological feature of
lung remodeling disease. In various embodiments, the pathological
feature of lung remodeling disease is selected from the group
consisting of decreased lung volume, increased lung density,
remodeled lung tissue, and increased mortality.
[0058] In one embodiment, the present invention provides a method
of normalizing or stabilizing lung density in a subject having lung
remodeling disease, the method comprising administering to the
subject an anti-CTGF agent, thereby normalizing or stabilizing lung
density in the subject. In another embodiment, the present
invention provides methods of preventing or reducing a pathological
change in lung density in a subject at increased probability of
being afflicted with lung remodeling disease, the method comprising
administering to the subject an anti-CTGF agent, thereby preventing
or reducing the pathological change in lung density in the subject.
Lung density may be measured by any method known to one of skill in
the art. In one particular aspect, the lung density is measured by
computed tomography (CT) scan; more particularly, lung density is
measured by high resolution CT (HRCT). In particular embodiments,
lung density is measured in Hounsfield Units (HU), and improvement
in lung density as a result of the present methods is measured as a
decrease in measured HU. On the Hounsfield scale, lung zones with a
density between -900 and -800 HU are typically considered within a
normal aerated range, while those between -500 and -100 are poorly
aerated and those between -100 and +100 are nonaerated. In some
embodiments, the subject having increased lung density has a lung
density of greater than -800 HU, greater than -700 HU, greater than
-600 HU, greater than -500 HU, greater than -400 HU, or greater
than -300 HU. In some embodiments, the subject having increased
lung density has a lung density between -700 and +100 HU, between
-600 and +100 HU, between -500 and +100 HU, between -500 and -100
HU, between -700 and +100 HU or between -500 and -300 HU. In some
embodiments, the subject having decreased lung density has a lung
density of less than -800 HU, less than -900 HU, less than -910 HU,
less than -950 HU. In some embodiments, treatment with an anti-CTGF
agent changes the lung density of the patient by at least 100 HU,
at least 150 HU, at least 200 HU, at least 250 HU or at least 300
HU towards the normal range HU range of -800 to -900 HU. In other
embodiments, treatment with an anti-CTGF agent normalizes the
patient's lung density, i.e., changes the lung density so that
after treatment the lung density is within the normal range of -800
to -900 HU. Improvement in lung density may additionally be
measured by or associated with improved lung function as described
infra.
[0059] In another embodiment, the present invention provides a
method of reducing or stabilizing lung remodeling in a subject
having lung remodeling disease, the method comprising administering
to the subject an anti-CTGF agent, thereby reducing or stabilizing
lung remodeling in the subject. In another embodiment, the present
invention provides a method of preventing or reducing lung
remodeling in a subject at increased probability of being afflicted
with lung remodeling disease, the method comprising administering
to the subject an anti-CTGF agent, thereby preventing or reducing
lung remodeling in the subject. Lung remodeling may be measured by
any method known to one of skill in the art. In some embodiments,
lung remodeling is measured using lung images from CT scan; more
particularly, by HRCT. In other embodiments, lung remodeling is
measured by lung biopsy and histology. In subjects having lung
remodeling disease, portions of normal lung may be replaced by
fibrotic septae between dilated airspaces, the gross appearance
being referred to as `honeycomb changes.` Lung remodeling in lung
remodeling disease generally shows patchy, heterogeneous regions of
dense fibrosis and mild or moderate interstitial lymphoplasmacytic
infiltrates, architectural remodeling, honeycomb change, and
fibroblastic foci. Fibroblastic foci represent zones of disease
activity whose extensiveness has been linked to survival. In some
embodiments, lung remodeling is measured as a change in potential
airspace, i.e., the fraction of lung not occupied by tissue as
assessed, e.g., by histology of biopsy material. In particular
embodiments, the present methods are used to treat a subject having
decreased potential airspace relative to normal. In other
embodiments, lung remodeling is measured by percentage of lung
showing honeycomb changes or fibroblastic foci. In particular
embodiments, the present methods are used to treat a subject having
increased percentage of honeycomb change or increased number of
fibroblastic foci. Lung remodeling diseases are frequently
associated with airway smooth muscle hypertrophy allowing for
measurement of lung remodeling based on the extent of hypertrophy.
Lung remodeling may additionally be measured by or associated with
decreased lung function as described infra.
[0060] Subjects having lung remodeling disease are at high risk of
acute or chronic respiratory failure and cardiovascular
complications which lead to increased mortality. Thus, in yet
another embodiment, the present invention provides a method of
increasing the likelihood of survival in a subject having lung
remodeling disease, the method comprising administering to the
subject an anti-CTGF agent, thereby increasing the likelihood of
survival in the subject. Increased likelihood of survival may also
be associated with improved lung function as described infra. In
further embodiments, the methods of the invention improve the
survival of subjects treated with an anti-CTGF agent by at least
5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 50%, 60%, 65%, 70%, 75%,
80%, 90%, 100%, 125%, 150% or 200% compared to a control group that
is not treated with an anti-CTGF agent.
[0061] In various embodiments, the methods of the invention improve
lung function. Improved lung function may be determined by any
measure known to those of skill in the art. In some embodiments,
lung function is determined by measuring blood gas parameters,
e.g., partial arterial pressure of oxygen (PaO.sub.2) or percent
oxygen saturation of blood. For instance, improvement in lung
function can be demonstrated in subjects having low PaO.sub.2 by
measuring the ability of the present methods to increase PaO.sub.2.
Typically, values for PaO.sub.2 greater than 80 mmHg are considered
normal, whereas values of 80 mmHg or less indicate a state of
hypoxia or hypoxemia. Although oxygen saturation (O.sub.2 Sat)
usually correlates with PaO.sub.2, the relationship is not linear.
For non-shifted O.sub.2 Sat, a value of 100% corresponds to 90 mmHg
PaO.sub.2, a value of 90% corresponds to 60 mmHg, and a value of
60% corresponds to 30 mmHg Factors that can cause a shift in the
correlative values include temperature and pH. In various
embodiments, the present methods are used to treat a subject having
a PaO.sub.2 of below 80 mmHg, particularly below 75 mmHg, and more
particularly below 70 mmHg. In particular embodiments, blood gas
parameters are normalized over the treatment time course. In some
embodiments, treatment with an anti-CTGF agent results in an
increase in a subject's PaO.sub.2 value of at least 1%, 2%, 3%, 4%,
5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25% or 30% above an initial
baseline value obtained prior to the initiation of treatment.
[0062] In some embodiments, lung function is determined by
measuring lung volume parameters or lung function parameters, e.g.,
vital capacity, residual volume (RV), forced expiratory volume
(FEV), forced vital capacity (FVC), forced expiratory flow (FEF),
tidal volume (TV), maximum flow (Vmax), peak expiratory flow rate
(PEFR), inspiratory reserve volume (IRV), functional residual
capacity (FRC), total lung capacity (TLC), expiratory reserve
volume (ERV) and/or maximum voluntary ventilation (MVV). Total lung
volume or total lung capacity refers to the volume in the lungs
upon maximal inspiration, and in a normal adult is 4-6 L. Tables 1
and 2 provide various measured and derived lung volumes. The vital
capacity and force vital capacity is the maximal volume expelled
from the lungs after maximal inspiration and is typically about 4.8
L for men and about 3.2 L for women. Residual volume refers to the
volume remaining in the lungs after maximal expiration, and in a
normal adult is about 1.0-1.2 L. FEV measures the volume of air
exhaled over a pre-determined period of time by a forced expiration
immediately after a full inspiration, typically over 1 second
(FEV.sub.1) or six seconds (FEV.sub.6). Vmax is the maximum flow
measured during FVC. PEFR measures the maximum flow rate during a
forced exhale starting from full inspiration.
[0063] The various lung volume measurements and other lung function
parameters can be readily measured through the use of a spirometer
as is well known in the art. Lung volume measurements and other
lung function parameters depend on the gender, age, ethnicity and
height of the subject, but spirometric reference values are widely
known and available in the art. (See Hankinson et al. Am J Respir
Crit Care Med. 1999; 159(1):179-187) Further, computer programs are
readily available for the calculation of spirometry reference
values based on a patient's specific gender, ethnicity, age and
height. (See Centers for Disease Control and Prevention
website)
TABLE-US-00001 TABLE 1 Average lung volumes in healthy adults Value
(liters) Volume In men In women Inspiratory reserve volume 3.3 1.9
Tidal volume 0.5 0.5 Expiratory reserve volume 1.0 0.7 Residual
volume 1.2 1.1 Ganong, W. Review of Medical Physiology (21st
ed.)
TABLE-US-00002 TABLE 2 Lung capacities in healthy adults Average
value (liters) Volume In men In women Derivation Vital capacity 4.8
3.1 IRV plus TV plus ERV Inspiratory capacity 3.8 2.4 IRV plus TV
Functional residual capacity 2.2 1.8 ERV plus RV Total lung
capacity 6.0 4.2 IRV plus TV plus ERV plus RV Ganong, W. Review of
Medical Physiology (21st ed.)
[0064] In some embodiments, the present methods improve lung
function in a subject having reduced lung function associated with
a lung remodeling disease as determined by measuring one or more
lung volume parameters. In certain embodiments, the subject has
below inspiratory reserve volume, expiratory reserve volume, vital
capacity and/or total lung capacity and the methods of the
invention increase one or more of these lung volume parameters. In
particular embodiments, one or more lung volume parameters are
normalized over the treatment time course.
[0065] In some embodiments, the methods of the invention will
increase at least one lung volume parameter of a patient suffering
for a lung remodeling disease associated with in a decrease in a
lung volume parameter or a restriction in lung expansion. In
certain embodiments, the increase in the at least one lung volume
parameter is an at least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40% or
50% increase compared to a baseline measurement taken before
treatment.
[0066] In some embodiments, lung function is determined by
measuring the cell number and cellular make-up of bronchoalveolar
lavage (BAL) fluid and/or sputum. Table 3 provides normal sputum
cell count values. (See Brightling C E. Chest. 2006:
129(5):1344-1348) The total cell count gives a measure of the
intensity of any inflammation and the differential of the type. At
early stages of lung remodeling disease, the alveolar and adjacent
capillary endothelial cells become leaky, leading to alveolar and
interstitial edema that is accompanied by an increase in the number
of immune cells found in BAL fluid and/or sputum. In particular,
the number of polymorphonuclear leukocytes (PMNs), which normally
comprise about 1-3% of the cellular component of BAL or sputum, can
increase to 20% or more. Therefore, in various embodiments, the
present methods are used to treat a subject having an elevated
percentage of PMNs in BAL fluid and/or sputum and the methods of
the invention reduce the percentage of PMNs in BAL fluid and/or
sputum. In particular embodiments, the present methods are used to
treat a subject having greater than 5% PMNs in BAL fluid and/or
sputum, particularly greater than 10%, and more particularly
greater than 15%. In particular embodiments, the cellular make-up
of BAL fluid and/or sputum is normalized over the treatment time
course. Sputum sample can be spontaneously obtained or induced with
an aerosol of normal or hypertonic saline. Treatment of a lung
remodeling disease with an anti-CTGF agent can be monitored through
analysis of BAL fluid and/or sputum and the dose adjusted to reduce
the total cell count and also to change the percent composition of
the BAL fluid and/or sputum.
TABLE-US-00003 TABLE 3 Normal Sputum Cell Count Values Sputum Cell
Count Mean Mean + 2 SD Median 90th Percentile Total cells,
10.sup.6/g 4.1 13.8 2.4 9.7 Eosinophils, % 0.4 2.2 0.0 1.1
Neutrophils, % 37.5 77.7 36.7 64.4 Macrophages, % 58.8 100 60.8
86.1 Lymphocytes, % 1.0 3.2 0.5 2.6 Metachromatic, % 0.0 0.1 0.0
0.04
[0067] In various embodiments, the method may be applied at
appropriate intervals to achieve the claimed result, such as
improved lung function as measured by any of the parameters
provided herein. In particular embodiments, the methods may be
applied 1, 2, 3, 4, or 5 time per month. In some embodiments, the
methods are applied 1 time per week or 1 time every other week. In
various embodiments, the methods are continued until the
pathological feature or functional parameter is essentially
normalized or the subject is no longer considered at risk. The
anti-CTGF agent can be administered by any conventional route with
systemic and pulmonary administration preferred.
[0068] In various embodiments, the anti-CTGF agent may be
administered at appropriate levels to achieve the desired
pharmacological effect. In particular embodiments, the anti-CTGF
agent is an antibody that binds specifically to CTGF. In various
embodiments, the antibody may be administered at a dose of from
0.01 to 100 mg of antibody/kg of patient weight, more particularly
from 0.1 to 50 mg/kg, and even more particularly from 1-30 mg/kg.
Doses particularly contemplated for use in the present methods
include, but are not limited to, at least 3 mg/kg, 5 mg/kg, 10
mg/kg, 15 mg/kg 20 mg/kg, 25 mg/kg, 30 mg/kg, 35 mg/kg, 40 mg/kg,
45 mg/kg, 50 mg/kg, 60 mg/kg, 70 mg/kg, 80 mg/kg, 90 mg/kg or 100
mg/kg. Depending on the type and severity of the disease, about
0.01 to 100 mg/kg is an initial candidate dosage for administration
to the patient, whether, for example, by one or more separate
administrations, or by continuous infusion. For repeated
administrations over several days or longer, depending on the
condition, the treatment is repeated until a desired suppression or
reversal of disease symptoms occurs. In some embodiments, an
initial higher antibody induction or loading dose is first used to
suppress or reverse a disease symptom and then the patient is
treated with a lower antibody maintenance dose. Other dosage
regimens may be useful and are not excluded from the present
invention.
[0069] In various embodiments, the anti-CTGF agent may be
administered by any route that provides a suitable pharmacokinetic
profile. In particular embodiments, the anti-CTGF agent is
administered intravenously. In further embodiments, the anti-CTGF
agent is administered intravenously in a single bolus injection. In
other embodiments, the anti-CTGF agent is administered
intravenously by infusion. In additional embodiments, the anti-CTGF
agent is administered as particulate or aerosol directly to the
lungs. In other embodiments, the anti-CTGF agent may be
administered subcutaneously, intramuscularly, or
intraperitoneally.
[0070] The methods of the present invention, as exemplified herein,
clearly demonstrate that the administration of an anti-CTGF agent
significantly improves lung density, lung function, and survival at
all treatment periods. Thus, administration of an anti-CTGF agent
provides significant benefit to a subject whether the anti-CTGF
agent is administered prior to an initial insult or at any time
subsequent to the initial insult. The data also clearly
demonstrates that administration of an anti-CTGF agent does benefit
a patient already in a chronic progressive phase of a lung
remodeling disease. Therefore, in some embodiments, the anti-CTGF
agent is administered at or near the time of initial diagnosis of a
lung remodeling disease. In other embodiments, the anti-CTGF agent
may be initiated prior to an event associated with increased
probability or likelihood of being afflicted with lung remodeling
disease. Such an event may be an occupational event, e.g.,
potential exposure to an agent known or suspected of causing lung
remodeling disease. For example, the anti-CTGF agent may be used to
pre-treat a worker that may be exposed to asbestos as part of an
asbestos abatement project. Such an event may be a medical event,
e.g., exposure to a medical therapy or procedure known or suspected
of causing a lung remodeling disease. For example, the anti-CTGF
agent may be used to pre-treat a patient that is going to be
exposed to ionizing radiation as part of radiation therapy.
[0071] In another aspect, the present invention provides
medicaments for treatment of lung remodeling disease. In one
embodiment, the present invention provides the use of an anti-CTGF
agent in preparing a medicament for treating lung remodeling
disease. The medicament may be used to prevent, reduce, reverse,
normalize, and/or stabilize various pathological features of lung
remodeling disease. Such features include, but are not limited to,
decreased lung volume, increased lung density, the presence of
remodeled lung tissue, and increased probability of mortality. In
another embodiment, the present invention provides the use of an
anti-CTGF agent in preparing a medicament for preventing or
reducing a pulmonary disorder in a subject at increased probability
of being afflicted with a lung remodeling disease. In various
embodiments, the medicament also improves lung function in a
subject having lung remodeling disease.
[0072] Any anti-CTGF agent that directly inhibits the expression or
activity of CTGF may be used in formulating the present
medicaments. In particular embodiments, the anti-CTGF agent is an
antibody that binds specifically to CTGF, or a polynucleotide
inhibitor of CTGF expression (for example, an antisense
oligonucleotide, siRNA, shRNA, or miRNA). In a preferred
embodiment, the anti-CTGF agent is an antibody that binds
specifically to CTGF. Any antibody that specifically binds to CTGF
may be used in formulating the present medicaments. Antibodies for
use in the present medicaments are described supra. In various
embodiments, the antibody for use in the present medicaments is an
antibody described and claimed in U.S. Pat. No. 7,871,617. In
particular embodiments, the antibody has the amino acid sequence of
the antibody produced by the cell line identified by ATCC Accession
No. PTA-6006. In other embodiments, the antibody competitively
binds to CTGF with an antibody produced by ATCC Accession No.
PTA-6006. A particular antibody for use in the present medicaments
is CLN1 or mAb1 as described in U.S. Pat. No. 7,405,274, or an
antibody substantially equivalent thereto or derived therefrom.
[0073] The medicament may be formulated for the intended route of
administration. Such formulations may encompass pharmaceutically
acceptable carriers that are inherently nontoxic and
nontherapeutic. Polynucleotide inhibitors may be admixed,
encapsulated, conjugated or otherwise associated with other
molecules, molecule structures or mixtures of compounds, e.g.,
liposomes, receptor targeted molecules, oral, rectal, topical,
systemic, aerosolized, nebulized or other formulations, for
assisting in uptake, distribution and/or absorption. (See, e.g.,
U.S. Pat. No. 5,108,921; U.S. Pat. No. 5,354,844; U.S. Pat. No.
5,416,016; U.S. Pat. No. 5,459,127; U.S. Pat. No. 5,521,291; U.S.
Pat. No. 5,543,158; U.S. Pat. No. 5,547,932; U.S. Pat. No.
5,583,020; U.S. Pat. No. 5,591,721; U.S. Pat. No. 4,426,330; U.S.
Pat. No. 4,534,899; U.S. Pat. No. 5,013,556; U.S. Pat. No.
5,108,921; U.S. Pat. No. 5,213,804; U.S. Pat. No. 5,227,170; U.S.
Pat. No. 5,264,221; U.S. Pat. No. 5,356,633; U.S. Pat. No.
5,395,619; U.S. Pat. No. 5,416,016; U.S. Pat. No. 5,417,978; U.S.
Pat. No. 5,462,854; U.S. Pat. No. 5,469,854; U.S. Pat. No.
5,512,295; U.S. Pat. No. 5,527,528; U.S. Pat. No. 5,534,259; U.S.
Pat. No. 5,543,152; U.S. Pat. No. 5,556,948; U.S. Pat. No.
5,580,575; and U.S. Pat. No. 5,595,756.)
[0074] Examples of carriers for use with antibodies include ion
exchangers, alumina, aluminum stearate, lecithin; serum proteins
such as human serum albumin; buffers such as phosphate, histidine,
or glycine; sorbic acid, potassium sorbate, partial glyceride
mixtures of saturated vegetable fatty acids, water, salts; or
electrolytes such as protamine sulfate, sodium chloride, metal
salts, colloidal silica, magnesium trisilicate, polyvinyl
pyrrolidone, cellulosic polymers, and polyethylene glycol. Carriers
for topical or gel-based forms of antibody include polysaccharides
such as sodium carboxymethylcellulose or methylcellulose,
polyvinylpyrrolidone, polyacrylates,
polyoxyethylene-polyoxypropylene-block polymers, polyethylene
glycol, and wood wax alcohols. Conventional depot forms include,
for example, microcapsules, nano-capsules, liposomes, plasters,
sublingual tablets, and polymer matrices such as
polylactide:polyglycolide copolymers. When present in an aqueous
dosage form, rather than being lyophilized, the antibody typically
will be formulated at a concentration of about 0.1 mg/ml to 100
mg/ml, although wide variation outside of these ranges is
permitted.
[0075] The present invention contemplates the use of the present
methods in combination with other therapies. In one embodiment, the
method is used in combination with another therapy, e.g., to
further augment therapeutic effect on certain pathological events,
etc. The two treatments may be administered at the same time or
consecutively, e.g., during a treatment time course or following
disease progression and remission. In another embodiment, the
method is used in combination with another therapeutic method
having a similar or different mode of action, e.g., a
corticosteroid. Current therapeutic approaches to treat lung
remodeling are known by one of skill in the art, and include, for
example, corticosteroids, bronchodilators, anticholinergics,
vasodilators, diuretics, anti-hypertensive agents, acetazolamide,
antibiotics, immunosuppressive drugs, surfactants, supplemental
oxygen and mechanical ventilation. Use of any of these therapeutic
agents in combination with the use of methods of the present
invention is specifically contemplated.
[0076] The medicaments may, if desired, be presented in a pack or
dispenser device containing one or more unit dosage forms
containing the active ingredient. Such a pack or device may, for
example, comprise metal or plastic foil, such as a blister pack; or
glass and rubber stoppers such as in vials. The pack or dispenser
device may be accompanied by instructions for administration.
Compositions comprising an agent of the invention formulated in a
compatible pharmaceutical carrier may also be prepared, placed in
an appropriate container, and labeled for treatment of an indicated
condition.
[0077] These and other embodiments of the present invention will
readily occur to those of ordinary skill in the art in view of the
disclosure herein.
EXAMPLES
[0078] The invention will be further understood by reference to the
following examples, which are intended to be purely exemplary of
the invention. The present invention is not limited in scope by the
exemplified embodiments, which are intended as illustrations of
single aspects of the invention only. Any methods that are
functionally equivalent are within the scope of the invention.
Various modifications of the invention in addition to those
described herein will become apparent to those skilled in the art
from the foregoing description and accompanying figures. Such
modifications are intended to fall within the scope of the appended
claims.
[0079] The invention will be further understood by reference to the
following examples, which are intended to be purely exemplary of
the invention. The present invention is not limited in scope by the
exemplified embodiments, which are intended as illustrations of
single aspects of the invention only. Any methods that are
functionally equivalent are within the scope of the invention.
Various modifications of the invention in addition to those
described herein will become apparent to those skilled in the art
from the foregoing description and accompanying figures. Such
modifications are intended to fall within the scope of the appended
claims.
Example 1
Radiation Induced Lung Remodeling
Materials and Methods
Experimental Protocol and Animal Model
[0080] Female C57BL/6 mice (8-wk-old; approximate body weight: 20
g; Charles River Laboratories) were randomized into 8 groups of 25
animals and were supplied with food and water ad libitum. Six
groups (150 mice) were irradiated (IR) with a single dose of 20 Gy
at day 0. Mice were anesthetized by intraperitoneal application of
0.2 mg/kg Rompun (Bayer) and 100 mg/kg ketamin 10% (Parke-Davis).
Photon irradiation was administered as a single 20 Gy dose to the
entire thorax (Siemens linear accelerator; source surface distance
of 1 m; irradiation field of 0.02.times.0.2 m). Other organs, above
and below the thorax, were shielded.
[0081] The anti-CTGF monoclonal antibody (CLN1) used in this
experiment is described and claimed in U.S. Pat. No. 7,405,274.
Irradiated mice were administered either no therapeutic (25 mice),
control IgG (25 mice), or CLN1 (100 mice). Animals receiving CLN1
were randomized into groups to receive antibody therapy beginning
either 2 days before (25 mice; d-2), 2 days after (25 mice; d+2),
20 days after (25 mice; d+20), or 112 days after (25 mice; d+112)
irradiation. In each CLN1 group, antibody was administered at a
dose of 10 mg/kg by intraperitoneal (i.p.) injection three times
per week for a dosing period of 8 weeks. The two unirradiated
groups received either control IgG or CLN1 for the first 8 weeks of
the experiment. A schematic of the different dosing schedules is
provided in FIG. 1A.
Lung Imaging
[0082] To obtain independent qualitative and quantitative measures
for lung remodeling in the mice, high resolution computed
tomography (HRCT) was performed in every surviving mouse from each
group on weeks 4, 8, 12, 16, 18, 20, 22, 24, 30 and 48 after
irradiation. CT images were captured on a SOMATOM PLUS 4 VOLUME
ZOOM multi-slice CT scanner (Siemens). 120 kV with 100 mAS were
applied. 0.5-mm thin slices with 0.5-mm inter-slice distance
spanned the complete mouse chest (a total acquisition time of 0.5
s). Multiplanar reconstructions were performed for semiquantitative
analysis. The Hounsfield units (HU) of section slices from the
upper, middle and lower lung regions were determined. Density on CT
is often described by Hounsefield Units, where pure water measures
0 HU, air measures -1,000 HU, and very dense structures such as
bone approach +1,000 HU. After an initial examination of the whole
lung, representative slides were chosen to undergo further
analysis. Three slides of the lung, representing the upper (5
slides below the apex), middle (divorce of the trachea) and lower
(about 5 slides above the diaphragmatic dome) region were selected
and measured quantitatively by Hounsfield Units. Six circles were
set in the selected fields of both sides of the lung, representing
the upper anterior and posterior, the middle anterior and posterior
and the lower anterior and posterior region, thus collecting twelve
sets of data per mouse. Circles were set as large as possible, but
avoiding big bronchi and vessels. All examinations were performed
with the same window and level settings (400/1000). Total
arithmetic means.+-.standard error of the mean (SEM) of the HU were
calculated.
Lung Histology
[0083] Histological analysis from mice tissues was performed as
described in Plathow et al. (2004) Invest. Radiol. 39:600-609. In
brief, lungs were fixed by intratracheal instillation of 4%
formalin followed by overnight fixation, embedded in paraffin,
sectioned at 5 .mu.m, and stained with hematoxylin and eosin
(H&E), Sirius red or Masson's trichrome. The total count of
leukocytes, and septal thickness were determined by morphometric
evaluation (Q 600 Quantimet; Leica).
Statistics
[0084] Mouse survival curves after thoracic irradiation and CLN1
treatments were calculated with the Kaplan-Meier method and
compared using the log-rank test. Other quantitative data are given
as mean values.+-.SEM or as indicated. For analysis of differences
between the groups, ANOVA followed by the appropriate post hoc test
for individual comparisons between the groups was performed. All
tests were two-tailed. P<0.05 was considered statistically
significant.
Results
Course of Radiation-Induced Lung Damage
[0085] The present model of radiation-induced pulmonary disorders
utilizes a single dose of ionizing radiation as an initial insult.
Exposure of normal lung tissue to irradiation produces an acute
pneumonitis and a progressive, long-term fibrosis (see, e.g.,
Movsas et al. (1997) Chest 111:1061-1076). Characteristic
histologic findings in the pneumonitis phase of the radiation
response include prominent inflammatory cell infiltrates in the
alveoli and lung interstitium with simultaneous interstitial edema.
Both parameters typically exhibit similar kinetics in the acute
phase, reaching their maximum about 72 h after radiation injury.
After the acute radiation response, both leukocyte count and septal
edema spontaneously subside within a few days. (See FIGS. 1B and
1C.)
[0086] The later fibrogenesis phase is accompanied by a strong
second onset of leukocyte infiltration that typically begins
several weeks after irradiation and reaches a peak at about 20
weeks after irradiation. Development of fibrosis by progressive
collagen deposition detectable by Masson's trichrome staining of
irradiated lungs is usually evident after week 12. This
fibrogenesis phase is characterized by development of typical
fibroblast foci, with abnormal wound healing/repair leading to
replication of mesenchymal cells, as characterized by
fibroblast/myofibroblast migration and proliferation and exuberant
deposition of extracellular matrix in irradiated lungs. At later
time points (>20 wk), the fibroblast foci evolve and coalesce
into more widespread fibrosis with remodeling of the lung
architecture. The second onset of progressive fibrogenesis-related
infiltration of leukocytes typically persists until the
morphologically described fibrosis process is completed (after week
26). (See FIGS. 1B and 1C.) Fibrosis can be quantitatively assessed
by measuring lung density (quantified in Hounsfield units, HU)
using HRCT. Lung density dramatically increases during weeks 12-24
after radiotherapy in irradiated animals. (See FIG. 1D.)
Lung Remodeling Shown by Histology
[0087] To monitor the pathogenesis of the radiation-induced lung
remodeling process and to evaluate the effects of CLN1, mice were
selected for analysis of leukocyte infiltration and collagen
deposition with associated thickening of the alveolar septum.
Histological examination of H&E stained lung sections taken
from mice at various time points after irradiation demonstrated
that significant lung remodeling occurred between 13 and 19 weeks
after irradiation in the irradiated but untreated mice, and
administration of CLN1 attenuated this remodeling in a
schedule-dependent manner in the mice treated with CLN1 (data not
shown).
Potential Airspace
[0088] Evaluation of potential airspace in the lung, as assessed by
image analysis of histological specimens, demonstrated that the
present methods attenuate lung remodeling. (FIG. 2) Image analysis
of the H&E stained lung sections from the various groups were
processed to quantify the amount of solid tissue vs. empty space.
The ratio of empty space to tissue was called the fraction
potential airspace. No significant change in potential airspace was
observed in the first 12-13 weeks after irradiation in any of the
groups. By week 19, however, the fraction potential airspace in the
irradiated but untreated mice significantly decreased and continued
to decrease until week 31. By week 31 after irradiation, all of the
groups that had been treated with CLN1 exhibited larger fraction
potential airspace than the irradiated untreated group. As shown in
FIG. 2A, potential airspace in irradiated lungs improved when CLN1
administration was initiated 2 days before or 2 days after
irradiation and continued for 8 weeks. As shown in FIGS. 2B and 2C,
potential airspace in irradiated lungs was essentially normalized
(that is, was very similar to the potential airspace seen in
unirradiated mice) when CLN1 administration was initiated 20 days
or even 112 days after irradiation, and treatment was continued for
8 weeks. Thus, the methods of the present invention reduce, reverse
and even normalize lung remodeling in a subject having a
radiation-induced pulmonary disorder.
Lung Density
[0089] To monitor changes in lung density following irradiation and
to evaluate the effects of CLN1 on the changes in lung density, the
density of the lungs of all mice were measured by micro-CT at
various times after irradiation. The density of the lungs of
unirradiated mice (treated with either IgG or CLN1) was unchanged
over the course of the experiment. In contrast, the lung densities
in irradiated but untreated mice (either no treatment or treatment
with IgG) progressively increased until about 30 weeks after
irradiation, after which, the lung densities of the few surviving
mice did not increase further. The changes in lung density observed
in the irradiated but untreated mice were attenuated by the present
methods when CLN 1 was administered for 8 weeks beginning
immediately before (d-2) or after (d+2) irradiation (FIG. 3A.)
Administration of CLN1 beginning 20 days (d+20) or 112 days (d+112)
after irradiation had a more dramatic effect on lung densities,
such that the lung densities in the irradiated groups treated using
the methods of the present invention were indistinguishable from
those of unirradiated mice (FIGS. 3B and 3C) demonstrating the
ability of an anti-CTGF agent to reverse and normalize a pathology
associated with a lung remodeling disorder. Further, administration
of CLN1 112 days after irradiation showed a reversal of lung
density increases that occurred prior to treatment initiation (FIG.
3C.) Together, these data demonstrate that the methods of the
invention are capable of reducing, reversing and normalizing the
increased lung density that occurs in a subject having a
radiation-induced pulmonary disorder.
Partial Pressure O.sub.2 and Oxygen Saturation Percent
[0090] To monitor changes in lung function and to evaluate the
effects of CLN1, blood was sampled from the tail capillary of mice
in various groups and oxygen partial pressure was measured. FIG. 4
sets forth blood partial oxygen pressure (PaO.sub.2) 30 weeks after
irradiation for the treatment groups. The PaO.sub.2 for mice in the
unirradiated groups (IgG only or CLN1 only) was in the normal range
(normal range is >80 mm Hg shown as the striped area). As can be
seen in FIG. 4A, the irradiated but untreated mice (either no
treatment or IgG treated) exhibited PaO.sub.2 well below normal.
However, irradiated mice that were treated with CLN1 beginning
either at 20 days (d+20) or 112 days (d+112) after irradiation had
PaO.sub.2 in the normal range (FIG. 4A.) The oxygen partial
pressure of the blood was converted to a percent saturation and
compared to the lung densities measured at week 30 (FIG. 4B). As
can be seen in FIG. 4B, the correlation of the lung density and the
oxygen saturation is quite striking and suggested that reduction in
lung density is a good surrogate for improvement in lung function,
and both parameters are normalized in animals treated with the
CLN1. Additional blood samples were collected at week 48 and
measured for PaO.sub.2 (data not shown). No mice remained alive in
the irradiated untreated group at this time point but samples from
1-3 mice from the other groups were examined including remaining
mice from the irradiated IgG treated group. Oxygen saturation was
below normal in the irradiated IgG treated mice, but oxygen
saturation was now in the normal range for all of the CLN1 treated
groups demonstrating the ability of an anti-CTGF agent to increase
oxygen saturation in a subject with a lung remodeling disease
including normalizing oxygen saturation.
Septal Thickness
[0091] Another way to assess remodeling of the lungs is to measure
the thickness of the septa between the alveoli. The thickness of
the alveolar septa were measured manually from photographs of lung
sections from mice in all groups. The mean of 100 measurements per
lung section were plotted as a function of time (data not shown).
Little change in the alveolar septa thickness was observed in the
unirradiated mice. In contrast, the septa of irradiated and
untreated (either no treatment or IgG treated) mice exhibited
progressive thickening between 12 and 30 weeks after irradiation.
Septa of irradiated mice that were treated with CLN1 beginning 2
days before or 2 days after irradiation also exhibited progressive
thickening albeit with different time courses than the irradiated
untreated controls. However, septa of irradiated mice that were
treated with CLN1 beginning at 20 days or 112 days after
irradiation exhibited little change in alveolar septa thickness
(although there appeared to be a slight thickening at 12 weeks that
subsequently resolved) and at 30 weeks after irradiation were
indistinguishable from the unirradiated controls demonstrating that
treatment with an anti-CTGF agent can prevent, reverse and
normalize septal thickening associated with a lung remodeling
disease.
Survival
[0092] FIG. 5 shows the percent survival of mice from treatment
groups over the 48 weeks of the experiment. Over the course of the
48 week experiment, no mice in the unirradiated +CLN1 control group
died, while one mouse in the unirradiated +IgG control group died
(unirradiated +IgG group is not shown on FIG. 5). The survival of
the irradiated and untreated mice (IR) and those that were
irradiated and exposed to placebo (IR+IgG) were similar, with a
median survival of 167 and 161 days, respectively (irradiated +IgG
group not shown on FIG. 5). The group that began receiving CLN1 two
days before irradiation (d-2) exhibited a statistically significant
improvement in survival (p=0.041) with a median survival >336
days (FIG. 5A) demonstrating that treatment with an anti-CTGF agent
can extend survival by at least 200% compared to animals that do
not receive an anti-CTGF agent. The group that began receiving CLN1
20 days after irradiation (d+20) also exhibited a statistically
significant improvement in survival relative to the irradiated
placebo-treated group (p=0.021) with a median survival >336 days
FIG. 5B) demonstrating that treatment with an anti-CTGF agent can
extend survival by at least 200% compared to animals that do not
receive an anti-CTGF agent. The group that began receiving CLN1 at
16 weeks after irradiation (d+112) exhibited an increase in median
survival (246 days) over the irradiated untreated groups (FIG. 5C)
demonstrating that treatment with an anti-CTGF agent can extend
survival by at least 140% compared to animals that do not receive
an anti-CTGF agent. The group that began receiving CLN1 2 days
after irradiation (d+2) exhibited a median survival of 202 days
(FIG. 5A) demonstrating that treatment with an anti-CTGF agent can
extend survival by at least 120% compared to animals that do not
receive an anti-CTGF agent. Thus all of the irradiated groups that
received CLN1 treatment exhibited greater median survival rates
after receiving a lethal ionizing radiation dose than the
irradiated and untreated (or placebo treated) control groups.
Together, these data demonstrate that the methods of the invention
improve survivability and survival rate in a subject having a
radiation-induced pulmonary disorder. In further embodiments, the
methods of the invention improve the survival of subjects afflicted
with a lung remodeling disease when treated with an anti-CTGF
agent, thereby increasing the survival of the treated group by at
least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 50%, 60%, 65%, 70%,
75%, 80%, 90%, 100%, 125%, 150% or 200% compared to a control group
not treated with the anti-CTGF agent.
Leukocyte Infiltration
[0093] The difference in survival between the group that began
receiving CLN1 2 days before or 2 days after irradiation was
unexpected, since their treatment periods almost completely
overlapped, with less than 1 week difference between them. The
acute response to irradiation is characterized by edema and
leukocyte infiltration, and CTGF has the potential to alter either
or both of these events. CTGF has been reported to modulate the
motility of immune cells such as macrophages, suggesting that CLN1
could directly affect leukocyte infiltration. In addition, CTGF may
alter secretion of chemokines and cytokines that recruit and
maintain leukocytes. Therefore, CTGF may have direct and indirect
effects on leukocyte infiltration that could be altered by the
presence of CLN1 before irradiation. To test this hypothesis, the
number of leukocytes infiltrating the lungs at 2 days after
irradiation (see Plathow et al. (2004) Invest. Radiol. 39:600-609)
was compared for the irradiated and untreated group (IR) and the
group that was pretreated with CLN1 for 2 days prior to irradiation
(d-2). A slightly smaller number of leukocytes per high power field
were observed in lung slices from the CLN1-pretreated mice than
from the irradiated, untreated mice (data not shown).
[0094] Leukocyte infiltration into the lungs of mice from all
groups was also examined at 18 weeks after irradiation which
corresponds to about the mid-point of the chronic response to
irradiation. By 18 weeks, about 5 times the number of leukocytes
was observed per field in the lungs of irradiated mice compared to
unirradiated mice. Relative to irradiated and placebo-treated mice
(IR+IgG), pretreatment of mice with CLN1 (d-2) decreased leukocyte
infiltration by a small but statistically significant amount, while
treatment with CLN1 beginning 2 days after irradiation (d+2) did
not alter the number of leukocytes in the lungs at 18 weeks after
administration. Administration of CLN1 beginning 20 days or 16
weeks (112 days) after irradiation demonstrated a greater
inhibition of leukocyte infiltration into the lungs at 18 weeks
after irradiation, such that it was indistinguishable from that of
unirradiated mice demonstrating the ability of treatment with an
anti-CTGF agent to prevent, reverse or normalize leukocyte
infiltration associated with a lung remodeling disease.
[0095] Although the present examples were limited to 8 week
treatment windows due to potential immune reaction in mice being
treated with a human monoclonal antibody, the results clearly
demonstrate that the methods provide significant improvement in all
treatment periods. Thus, benefit can be seen in a subject whether
the present methods are initiated prior to thoracic exposure to
ionizing radiation or at times subsequent to exposure and even at
times subsequent to the manifestation of acute or chronic effects
of the radiation. The data clearly demonstrate that, to the extent
possible, the methods should be initiated as early as possible and
maintained throughout the period that the subject remains at risk
for diminished organ function and compromised survivability. The
data also clearly demonstrate that patients in the chronic
progressive phase of the disorder can still benefit from the
methods and medicaments of the present invention, improving lung
function, reversing lung remodeling and improving survival.
Example 2
Human Lung Specimens
[0096] Paraffin embedded lung tissue sections prepared from autopsy
specimens were obtained from 6 patients with BPD and 3 newborn
patients without lung disease. The BPD patients were born between
23 to 28 weeks of gestational age and died between 2 to 11 months
of chronological age. Diagnosis of BPD was made based on
standardized criteria including oxygen requirement for more than 28
days at 36 weeks postmenstrual age (Jobe A H et al. Am J Respir
Crit Care Med. 2001; 163: 1723-1729). The non-BPD patients born
between 34 to 36 weeks of gestational age were diagnosed with
trisomy 18, complex congenital heart disease and anencephaly and
died within 2 to 6 h.
[0097] Examination of CTGF expression in lung tissue sections
revealed that CTGF expression was dramatically increased in the
thickening alveolar septa in BPD lungs as compared with the control
lungs where CTGF expression was low. These data support a role of
CTGF in the pathogenesis of BPD.
Example 3
Hyperoxia-Induced Lung Remodeling
[0098] The lungs of neonates and premature infants are often not
sufficiently developed to oxygenate blood effectively.
Consequently, these infants are treated with oxygen, in hyperbaric
chambers, to keep the infants alive until they can survive in room
air. However, even short term exposure to pure oxygen (or high
concentration of oxygen) can be toxic or result in serious
long-term effects, for example, pulmonary and cardiovascular
remodeling. Exposure of the underdeveloped lungs to high oxygen
concentrations may trigger pulmonary hypertension or cancers, and
may affect organs beyond the lungs including kidney, heart, brain,
eye, and vasculature.
[0099] CTGF was known to be elevated during neonatal hyperoxia
(Chen et al. (2007) Pediatric Res. 62:128-133) and confirmed by
Example 2, supra, but the functional significance of this is not
completely understood.
[0100] Objective: To determine the effect of anti-CTGF antibody on
prevention and treatment of hyperoxia-induced bronchopulmonary
dysplasia in neonatal rats. The anti-CTGF antibody used in these
experiments, Ab3149, is a rodent-specific chimeric antibody derived
from the CLN1 antibody described herein, that was designed to
eliminate the problem of mouse anti-human antibodies that can
result when a human anti-CTGF antibody is used for experiments in
mice or rats. Ab3149 contains the variable regions of the human
CLN1 and the constant region of a mouse IgG2a, and is
indistinguishable from CLN1 in binding to CTGF, but importantly, no
rat anti-mouse antibodies were seen in rats after 15 weeks of
dosing with Ab3149.
[0101] Study Design: Hyperoxia exposure and anti-CTGF antibody
treatment: Within 24 h after birth Sprague Dawley rat pups were
randomly allocated to receive room air (21% O2) or 90% O2.
Continuous 90% O2 exposure was achieved in a Plexiglas chamber
(77.times.64.times.37 cm) by a flow-through system. The oxygen
level inside the Plexiglas chamber was monitored continuously with
a Ceramatec (MAXO2) oxygen analyzer. Nursing dams were rotated
between room air and hyperoxia every 24 h to prevent oxygen
toxicity in the dams. The room air and hyperoxia groups were
subdivided to receive Ab3149 (10 mg/kg) or placebo (control IgG)
(20 mg/kg) i.p. injection. To prevent BPD, rat pups received Ab3149
or placebo on day 1 and then every other day during hyperoxia or
room air exposure until sacrificed at day 14.
[0102] Lung histology and morphometry: On day 14, rat pups were
sacrificed and lungs were pressure fixed for parafilm embedding and
section. Lung tissue sections were stained by H&E for histology
and morphometry. Lung morphometry was performed as previously
described with some modification (Chen S et al. Am J Physiol Lung
Cell Mol Physiol. 2011; 300(3):L330-40.). Briefly, 10 random
non-overlapping images are taken under 20.times. magnification on
three H&E stained tissue sections of every 100 .mu.m distance
from each lung by a research staff blind to experimental
conditions. The large airways and blood vessels are excluded from
the analysis. Quantification of mean linear intercept (MLI), radio
alveolar count (RAC) and secondary septa were performed using
MetaMorph Imaging System by an Analytical Imaging Core Facility
member blind to experimental conditions. At least 8 animals were
evaluated in each group (total 48).
[0103] Mean linear intercept (MLI), vascular density (VD) and
medial wall thickness (MWT) of pulmonary arterioles were determined
as a means of assessing the extent of pulmonary vascular
remodeling. PH was determined by RVSP and RVH. .beta.-catenin
signaling was assessed by .beta.-catenin nuclear translocation and
cyclin D1 expression. Western blot analysis for CTGF expression was
performed on lung homogenates using the method of Wu S et al. (Wu S
et al. Am J Respir Cell Mol Biol 2010; 42:552-63). Lung
gelatinolytic activity of matrix metalloproteinase-2 (MMP-2) was
evaluated with zymography using the method of Fan W H et al. (Fan W
H et al. J Biol Chem 2002; 277:9800-9805). Data were expressed as
means.+-.SD. Comparison among the groups was performed by one-way
ANOVA followed by Student-Newman Keuls test. A P<0.05 was
considered significant.
Results:
[0104] Treatment with an anti-CTGF agent improved alveolarization
in the hyperoxia treated animals. The lungs from the hyperoxia +IgG
treated animals had an increased MLI compared to the control (room
air +IgG) animals. Treatment with Ab3149 reduced the increase in
MLI of the hyperoxia treated animals. Stained lung sections from
the hyperoxia+Ab3149 animals showed increased alveolarization
compared to the lung sections from the hyperoxia +IgG animals.
Treatment with Ab3149 was also associated with increased numbers of
secondary septa (seen in the stained sections) in the hyperoxia
treated animals as compared to the IgG treated hyperoxia animals,
FIG. 6. This demonstrates that treatment of animals exposed to
hyperoxia with an anti-CTGF agent can increase alveolarization. The
hyperoxemic animals treated with Ab3149 also showed increased
vascular density, had more normalized arterial wall thickness,
showed improved pulmonary arterial hypertension, and showed reduced
macrophage infiltration, compared to the IgG treated hyperoxemic
animals. Nuclear translocation of .beta.-catenin was inhibited in
the Ab3149 treated hyperoxemic animals compared to the IgG treated
hyperoxemics and Ab3149 prevented the increase in cyclin D1 that
was seen in the IgG treated hyperoxemic animals. The data is
summarized in Table 4.
TABLE-US-00004 TABLE 4 RA-IgG O.sub.2-IgG RA-Ab3149 O.sub.2-Ab3149
MLI 42.2 .+-. 2.7 65.9 .+-. 8.4* 36.8 .+-. 1.5 48.5 .+-. 4.3** *p
< 0.001, **p < 0.001 VD 8.8 .+-. 1.9 2.6 .+-. 0.2* 8.6 .+-. 1
5.7 .+-. 0.7** *p < 0.001, **p < 0.001 MWT 0.1 .+-. 0.02 0.2
.+-. 0.03* 0.09 .+-. 0.01 0.18 .+-. 0.05** *p < 0.001, **p =
0.002 RVSP 15 .+-. 3 28.9 .+-. 5* 15 .+-. 1 22.8 .+-. 2** *p <
0.001, **p = 0.001 RVH 0.3 .+-. 0.04 0.4 .+-. 0.1* 0.3 .+-. 0.01
0.3 .+-. 0.04** *p = 0.03, **p < 0.001 .beta.-catenin 0.01 .+-.
0.01 0.35 .+-. 0.1* 0.01 .+-. 0.01 0.07 .+-. 0.02** *p < 0.001,
**p < 0.001 CD1 0.07 .+-. 0.02 2.04 .+-. 0.6* 0.7 .+-. 0.2 0.6
.+-. 0.1** *p = 0.001, **p = 0.002 MLI = Mean Linear Intercept VD =
Vascular Density MWT = Medial Wall Thickness RVSP = Right
Ventricular Systolic Pressure RVH = Right Ventricular Hypertrophy N
= 5-10/group, Mean .+-. SD, *compare to RA-IgG, **compare to
O.sub.2-IgG
[0105] These experiments demonstrate that hyperoxia alters
development of neonatal rat lungs. Hyperoxia decreased
alveolarization and vascularization resulting in fewer, larger
alveoli. Hyperoxia also induced cardiovascular remodeling,
pulmonary artery wall thickening and RV hypertrophy, leading to
pulmonary arterial hypertension. Hyperoxia also increased
epithelial to mesenchymal transition (EMT) in the lung and
hyperplasia.
[0106] Treatment with the anti-CTGF agent, Ab3149, improved the
lung development in hyperoxemic animals for all of the parameters
measured. MLI was significantly reduced in hyperoxemic animals
treated with Ab3149 being about 74% the size of MLI of hyperoxemic
animals treated with IgG. Treatment of hyperoxemic animals with an
anti-CTGF agent significantly reduced right ventricular systolic
pressure to about 79% and right ventricular hypertrophy to about
75% of the values seen in hyperoxemic animals treated with IgG
demonstrating the ability of an anti-CTGF agent to reduce hyperoxia
induced pulmonary vascular remodeling and thus attenuate pulmonary
hypertension.
[0107] Further, hyperoxemic animals treated with Ab3149 had
increased secondary septa formation compared to hyperoxemic animals
treated with IgG, FIG. 6. These facts demonstrate that treatment
with an anti-CTGF agent improves alveolarization in neonatal rats
exposed to hyperoxia.
[0108] Lung vascularization was increased by over 2-fold in
hyperoxemic animals treated with an anti-CTGF agent compared to
hyperoxemic animals treated with IgG demonstrating the ability of
an anti-CTGF agent to increase vascular density.
[0109] Additionally, treatment with an anti-CTGF agent
significantly decreased macrophage infiltration into the alveoli of
hyperoxemic animals compared to hyperoxemic animals treated with
IgG, FIG. 7, indicating the ability of an anti-CTGF agent to
decrease pulmonary inflammation.
[0110] MMP-2 is one of the major ECM-degrading protease that
specifically degrades basement membranes and is known to play a
role in systemic vascular remodeling. (See Galis Z S et al. Circ
Res 2002; 90:251-262) Hyperoxia exposure was found to increase
MMP-2 activity as evidenced by gelatin gel zymography. This
activity was completely blocked by the anti-CTGF agent
demonstrating the ability of an anti-CTGF agent to prevent
increased expression of an ECM-degrading protease associated with
lung tissue remodeling.
Example 4
Improvement in Lung Function of an Asthmatic
[0111] A 30-year-old patient is sent for reassessment of her asthma
because her daily symptoms of cough and wheeze are not controlled
on inhaled budesonide 400 .mu.g bid. For control she requires
treatment with salbutamol, two to three puffs daily. Her FEV.sub.1
and slow vital capacity (SVC) are 2.0 L and 3.4 L (68% and 80% of
predicted, respectively). Her FEV.sub.1 improves to 2.5 L after
inhaling 200 .mu.g of salbutamol, confirming the presence of
asthma. A sputum sample is taken for assessment of airway
inflammation based on the measurement of sputum cell counts. The
total cell count is 22 million cells/g (normal<9.7 million
cells/g) of which 88% are neutrophils (normal<64.4%) and 5% are
eosinophils (normal<2%). The sputum cell count indicates a
combined infective and eosinophilic bronchitis. She is treated with
an antibiotic and an anti-CTGF agent. Two months later, she no
longer requires daily salbutamol, her FEV.sub.1 is improved to 2.6
L presalbutamol representing a 30% improvement over baseline, and
sputum total cell count and composition return to nominal.
Example 5
Improvement in Lung Function of an Asthmatic Smoker
[0112] A 175-cm tall man that smokes and has asthma has an
FEV.sub.1 of 1.99 L. The average FEV.sub.1 for a man of his age
that does not smoke or have asthma is 3.05 L. Treatment with an
anti-CTGF agent is initiated. At a later period, the man's lung
function is tested again and the FEV.sub.1 is found to improve to
2.4 L, demonstrating over a 20% increase from his baseline
measurement.
Example 6
Improvement in Lung Function of COPD Patient
[0113] An elderly man suspected of having the emphysema phenotype
of COPD is tested for baseline inspiratory capacity and arterial
oxygen tension (PaO.sub.2) values. His FEV.sub.1 value is 1.17 L
that is 48.3% of the predicted value for his age and height. His
FEV.sub.1/FVC is 45.1%, FRC is 120.9% of predicted, his RV is 191%
of predicted, TLC is 120% of predicted and his PaO.sub.2 is 65.1.
These results confirm the suspected diagnosis of the emphysema
phenotype of COPD.
[0114] The patient is treated with an anti-CTGF agent. At a later
time period, the patient is retested for inspiratory capacity and
PaO.sub.2. His FEV.sub.1 value is now 1.45 L, representing an over
20% increase compared to the baseline measurement. His PaO.sub.2
value has risen to 69.3 representing an increase of over 6% from
his initial baseline value. Further, his RV is reduced by over 10%
demonstrating the ability of an anti-CTGF agent to improve the lung
function and blood oxygenation of a patient with COPD.
[0115] Various modifications of the invention, in addition to those
shown and described herein, will become apparent to those skilled
in the art from the foregoing description. Such modifications are
intended to fall within the scope of the appended claims.
[0116] All references cited herein are hereby incorporated by
reference herein in their entirety.
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