U.S. patent application number 14/982139 was filed with the patent office on 2016-04-28 for system and method for diagnosis and treatment.
The applicant listed for this patent is Pop Test Oncology Limited Liability Company. Invention is credited to Randice Lisa Altschul, Rebecca O'Brien, Myron Rapkin, Neil David Theise.
Application Number | 20160113948 14/982139 |
Document ID | / |
Family ID | 46600751 |
Filed Date | 2016-04-28 |
United States Patent
Application |
20160113948 |
Kind Code |
A1 |
Altschul; Randice Lisa ; et
al. |
April 28, 2016 |
System and Method for Diagnosis and Treatment
Abstract
This invention relates to a low cost rapid response diagnostic
system to determine cortisol levels in patients selected as
potential candidates for GCR (glucocorticoid receptor) antagonist
therapy utilizing a GCR antagonist, such as ORG 34517. The rapid,
sensitive, and inexpensive test can be used to determine patients
who have non-normal cortisol production or disordered circadian
rhythms as a method for selecting subjects for GCR antagonist
therapy for whom it is likely to have beneficial and/or therapeutic
effects, and can also be used to monitor changes in cortisol levels
in response to treatment.
Inventors: |
Altschul; Randice Lisa;
(Cliffside Park, NJ) ; Theise; Neil David; (New
York, NY) ; Rapkin; Myron; (Indianapolis, IN)
; O'Brien; Rebecca; (Shell Knob, MO) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Pop Test Oncology Limited Liability Company |
Cliffside Park |
NJ |
US |
|
|
Family ID: |
46600751 |
Appl. No.: |
14/982139 |
Filed: |
December 29, 2015 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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14100714 |
Dec 9, 2013 |
9114147 |
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14982139 |
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13364651 |
Feb 2, 2012 |
8658128 |
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14100714 |
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61462492 |
Feb 3, 2011 |
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61463212 |
Feb 14, 2011 |
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61465703 |
Mar 23, 2011 |
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61518248 |
May 3, 2011 |
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61519323 |
May 20, 2011 |
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Current U.S.
Class: |
514/171 |
Current CPC
Class: |
A61K 2300/00 20130101;
A61K 9/08 20130101; A61K 47/44 20130101; A61P 13/08 20180101; A61P
43/00 20180101; A61K 47/34 20130101; G01N 2800/04 20130101; G01N
33/743 20130101; A61K 31/00 20130101; A61K 47/26 20130101; G01N
2800/304 20130101; B65D 75/367 20130101; A61P 15/00 20180101; A61K
2300/00 20130101; G01N 2800/7004 20130101; G01N 2800/52 20130101;
A61K 45/06 20130101; A61K 31/337 20130101; A61K 9/107 20130101;
A61K 31/58 20130101; A61P 25/20 20180101; A61P 3/08 20180101; A61N
5/1001 20130101; C07J 17/00 20130101; A61P 25/00 20180101; A61P
25/22 20180101; A61K 31/337 20130101; A61P 35/02 20180101; A61P
25/24 20180101; A61K 9/10 20130101; A61K 31/58 20130101; A61P 1/16
20180101; A61K 9/0019 20130101; A61P 5/00 20180101; A61K 47/10
20130101; A61P 35/00 20180101; A61P 11/00 20180101; A61P 25/18
20180101; A61K 47/42 20130101 |
International
Class: |
A61K 31/58 20060101
A61K031/58; A61K 31/337 20060101 A61K031/337 |
Claims
1. A method for treating neoplasia characterized by expression of a
glucocorticoid receptor, in a patient in need of such treatment,
comprising: administering to said patient therapeutically effective
amounts of each of: at least one neoplasia-treating agent selected
from the group consisting of taxanes, and combinations thereof; and
a GCR (glucocorticoid receptor) antagonist selected from the group
consisting of ORG 34517, 11-(substituted phenyl)-estra-4,9-diene
derivatives, and 11-(substituted phenyl)-estra-4,9-diene
derivatives of formula I ##STR00015## wherein A is a residue of a
5- or 6-membered ring containing 2 heteroatoms which are not
connected to each other and independently selected from O and S,
the ring being optionally substituted with one or more halogen
atoms, or A is a residue of a 5- or 6-membered ring wherein no
double C--C bonds are present, containing 1 heteroatom selected
from O and S, which heteroatom is connected to the phenyl group at
the position indicated with an asterisk, the ring being optionally
substituted with one or more halogen atoms; R1 is H or
1-oxo(1-4C)alkyl; R2 is H, (1-8C)alkyl, halogen or CF3; X is
selected from (H,OH), O, and NOH; and the interrupted line
represents an optional bond, wherein the neoplasia is selected from
the group consisting of hepatocellular carcinoma, breast cancer,
prostate cancer, lung cancer, and ovarian cancer.
2. The method of claim 1, wherein ORG34517 is administered as an
agent which directly effects tumor growth, independent of other
administered treatment modalities, for palliation, remission, or
cure.
3. The method of claim 1, wherein the neoplasia is chemo-resistant
ER/GR+ breast cancer.
4. The method of claim 1, wherein the ORG 34517 given systemically
through oral or intravenous routes.
5. The method of claim 1, wherein the ORG 34517 is given to
accomplish cure or remission of tumor.
6. The method of claim 1, wherein the ORG 34517 is given to
accomplish reduction of tumor burden to enhance effectiveness of
subsequent surgical resection.
7. The method of claim 1, wherein the ORG 34517 is given to
accomplish reduction of tumor burden to make an unresectable tumor
resectable.
8. The method of claim 1, wherein the taxane is selected from the
group consisting of paclitaxel, docetaxel, and combinations
thereof.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application is a Divisional application of U.S. Ser.
No. 14/100,714 filed Dec. 9, 2013 which is a Divisional application
of U.S. Ser. No. 13/364,651, filed Feb. 2, 2012, now U.S. Pat. No.
8,658,128, issued Feb. 25, 2014, which claims benefit under 35
U.S.C. .sctn.119(e) to U.S. Provisional Patent Application
61/462,492 filed Feb. 3, 2011; U.S. Provisional Patent Application
61/463,212 filed Feb. 14, 2011; U.S. Provisional Patent Application
61/465,703 filed Mar. 23, 2011; U.S. Provisional Patent Application
61/518,248 filed May 3, 2011; and U.S. Provisional Patent
Application 61/519,323 filed May 20, 2011, the disclosures of which
are incorporated herein in their entirety.
BACKGROUND OF THE INVENTION
[0002] 1. Field of Invention
[0003] This invention relates to a low cost rapid response
diagnostic system to determine cortisol levels in patients selected
as potential candidates for GCR (glucocorticoid receptor)
antagonist therapy utilizing a GCR antagonist, such as ORG 34517.
The rapid, sensitive, and inexpensive test can be used to determine
patients who have non-normal cortisol production or disordered
circadian rhythms as a method for selecting subjects for GCR
antagonist therapy for whom it is likely to have beneficial and/or
therapeutic effects, and can also be used to monitor changes in
cortisol levels in response to treatment.
[0004] 2. Description of Related Art
[0005] ORG 34517 is one of a class of therapeutic agents designed
to block the glucorticoid receptor (GR), acting as an antagonist
for endogenous cortisol. Its primary developmental pathway has been
as a treatment for neuropsychiatric diseases that are characterized
by dysregulated signaling in the hypothalamic-pituitary-adrenal
axis, often with higher than normal circulating levels of
endogenous cortisol. Of particular note are the phase 2 clinical
trials that have been completed for the treatment of psychotic
depression. Other possible uses in this disease category which are
under investigation include: post-traumatic stress disorder, weight
gain in patients requiring long term anti-psychotic medication,
hospital delirium of the elderly, etc. In addition, the diverse
data indicate a possible role for GR-blockade as a means of
promoting chemo-sensitization of target tumors. Pre-clinical trials
demonstrate significant outcomes--breast cancer growth slowed and
reversed. These are pre-clinical trials in which the company has
successfully demonstrated the efficacy of a chemotherapy sensitizer
for "triple negative" breast cancer.
[0006] The "triple negative" breast cancer is the most difficult to
treat type of breast cancer, and is indicated by the patient
testing negative for estrogen-receptor, progesterone-receptor and
her-2/neu. The triple negative breast cancer is resistant to
chemotherapy. Primary drug resistance and early onset of resistance
are seen in other tumor types, as well, for example in liver and
ovarian cancers, where there is a significant unmet medical need
for effective therapy. Chemotherapy is still a key approach to
cancer treatment. Chemosensitizers would contribute to improve the
efficacy of current therapeutic drugs and potentially improve their
side effect profile. The world cancer market was estimated at $23
billion in 2004 and is expected to grow to at least $61 billion by
2013 with a CAGR of 14.7%. U.S. Patent Application Publication no.
2011/0269728 (Pan et al.), incorporated by reference herein in its
entirety, discloses a method of killing breast cancer cells
comprising administering to a breast cancer patient an effective
amount of a combination of anti-cancer compounds, wherein the
anticancer compounds comprise a glucocorticoid receptor antagonist
and a chemotherapeutic.
[0007] The present invention provides a low cost rapid response
diagnostic system to determine salivary cortisol levels in patients
selected as potential candidates for GCR (glucocorticoid receptor)
antagonist therapy utilizing a GCR antagonist such as ORG 34517.
The inventors have developed a saliva based diagnostic device for
cortisol detection to accompany the development of ORG34517 as a
therapeutic agent for multiple indications.
[0008] Clinical testing of cortisol levels in patients is a high
cost, laborious test that can be salivary or serum, with samples
taken from a patient and sent to a lab to await results. The cost
and time factor for such tests has, to date, been prohibitive,
preventing the rapid quantitative determination necessary to assign
treatment with a glucocorticoid receptor (GCR) antagonist due to
the inability to make the determinations of cortisol levels at
point of need or to monitor changes in cortisol as a measure of
treatment response. By allowing the physician to determine the
elevated cortisol level of a patient and in turn provide a
therapeutic for such elevation at point of measurement, the
physician can qualify the best candidates suited for this type of
therapeutic. The system also enables continual monitoring of the
patient during treatment for assessment of responsiveness to
treatment.
[0009] The present invention provides a system in which an
apparatus uses a high void volume carrier to absorb sufficient
amounts of saliva to then be placed into a reaction vessel with a
reagent. The reagent is mixed with the sample and then is combined
with, for example, a fluorescent ligand or pigment-labeled ligand
and placed into a device to determine salivary cortisol levels of
the patient in less than 5 minutes, in either a portable,
miniaturized fluorescence polarization reader (in the former case)
or into a lateral flow device (in the latter) for measuring amounts
of substrate in a small amount of fluid by direct or indirect
methods.
[0010] The reader apparatus, for example, provides temperature
control and on-board mixing as an aid in viscosity control of the
reaction to ensure better accuracy and precision.
[0011] The invention and method for non-invasive sampling and
detecting the presence of a biological substance of interest in a
test sample of, for example, saliva, or a bodily fluid, combining
said test sample with a buffering system (Reagent 1) containing
viscosity controllers and stabilizers in a reaction vessel, mix
solution well, combining said test sample and buffering system
mixture with a fluorescence-labeled ligand (Reagent 2) to said
biological substance (assay solution) in a reaction vessel, mix
solution well, and detecting a change of the assay solution in the
fluorescence polarization reader, or a pigment labeled ligand.
[0012] The present invention relates to the use of glucocorticoid
receptor (GCR) antagonists (e.g. ORG 34517) enabled by a device for
rapidly, sensitively, specifically quantifying salivary cortisol
levels as a surrogate for serum cortisol levels in a low cost
manner. One purpose of this combination of inventions is to
determine patients who have non-normal cortisol produced by the
adrenal cortex or disordered circadian rhythms as a method for
selecting subjects for GCR antagonist therapy for whom it is likely
to have beneficial and/or therapeutic effects, i.e., those with
abnormal high levels (but maintained circadian rhythm), over
responsiveness to normal levels, high night-time cortisol levels as
a feature of disrupted circadian rhythm. The rapid, sensitive, and
inexpensive test can also be used to monitor changes in cortisol
levels in response to treatment, in patients who have non-normal
cortisol produced by the adrenal cortex or disordered circadian
rhythms as a method for selecting subjects for GCR antagonist
therapy for whom it is likely to have beneficial and/or therapeutic
effects, but also in patients having normal baseline cortisol at
the start of treatment, but for whom changing cortisol levels
during treatment will indicate responsiveness to the GCR
antagonist.
[0013] The endogenous glucocorticoids are steroids predominantly
produced in the adrenal cortex. Glucocorticoids are important
steroids for intermediary metabolism, immune, musculosceletal,
connective tissue and brain function. The main glucocorticoid in
the body is cortisol. The production and secretion of cortisol is
governed by a complex and highly efficient system that includes the
hypothalamus, pituitary and the adrenal glands i.e.,
hypothalamic-pituitary-adrenal axis (HPA). Cortisol secretion has a
circadian release rhythm with peak values in early morning and
trough values at midnight.
[0014] The production and secretion of the most important
glucocorticoid, cortisol, is governed by a complex and highly
efficient system that includes the hypothalamus, pituitary and the
adrenal glands i.e., hypothalamic-pituitary-adrenal axis. Cortisol
secretion is regulated by the suprachiasmatic nucleus of the
hypothalamus into a circadian release rhythm. The timing is
synchronized with the solar day by dark-light shifts, which
normally reflect the habitual sleep-wake pattern. Therefore in
healthy persons, the cortisol secretion has a 24-hour circadian
pattern with peak serum levels in the early morning, 3-6 hours
after onset of sleep, and nadir levels around midnight. Physical
and psychological stressors also activate cortisol secretion.
Changed patterns of serum cortisol levels have been observed in
connection with abnormal adrenocorticotropic hormone (ACTH),
levels, clinical depression, psychological stress, and
physiological stressors such as hypoglycemia, illness, fever,
trauma, surgery, fear, pain, physical exertion, or temperature
extremes. Cortisol levels and responsiveness may also differ from
normal for elderly individuals and in individuals with autism or
Asperger's syndrome.
[0015] Glucocorticoids (GCs) such as, in humans, cortisol, perform
several important functions. These include participating in the
regulation of carbohydrate, protein and fat metabolism by signaling
the liver to make glucose and glycogen, the adipose tissues to
release lipids and fatty acids into the bloodstream, and the
skeletal muscles to release proteins or amino acids into the
bloodstream. GCs also decrease bone formation.
[0016] GCs also regulate the body's inflammatory response as well.
GCs are part of the feedback mechanism in the immune system that
inhibits immune activity (i.e., inflammation). GCs cause their
effects by binding to the GCR. The activated GCR complex in turn
up-regulates the expression of anti-inflammatory proteins in the
nucleus (a process known as transactivation) and represses the
expression of pro-inflammatory proteins in the cytosol by
preventing the translocation of other transcription factors from
the cytosol into the nucleus (transrepression) (Rhen T and
Cidlowski J A. NEJM 2005; 353: 1711-23).
[0017] GCR antagonist therapy is helpful in patients with
abnormally high levels of cortisol (but maintained circadian
rhythm), over responsiveness to normal levels, or high night time
cortisol levels as a feature of disrupted circadian rhythm.
Successful therapeutic use of such agents is thus dependent on
determining circadian cortisol levels (either peak levels during
the day, e.g., at noon, or measurements taken every 4 hours or 6
hours over a 24 hour period). This combined system of salivary
cortisol quantification as an enabling device for its paired GCR
antagonist will identify individuals for whom GCR antagonist
therapy has a benefit.
[0018] The glucocorticoid receptor (GR) is expressed at high levels
in some normal tissues, but not in others. Likewise, malignant
tumors of diverse types and sites have variable GR expression. When
present in normal or tumor (benign or malignant) tissues, this GR
expression may be variously located in some or all of their
cellular sub-compartments: 1. stem cells; 2. progenitor (so called
"transit amplifying") cell descendents of activated stem cells; and
3. differentiated progeny of activated stem or progenitor
cells.
[0019] As an example, in the gastrointestinal tract, GR are highly
expressed in esophageal squamous epithelia, hepatocytes, and
pancreatic islet cells, but are not highly expressed in other
gastrointestinal epithelia (stomach, small and large intestines,
pancreatic and biliary ducts). In corresponding malignancies
arising in these epithelia, hepatocellular carcinoma (HCC) and
squamous cell carcinomas (SCC) of the esophagus have consistently
high GR expression. Gastric and colorectal adenocarcinomas have
little to no GR expression.
[0020] Dexamethasone (DEX), a binding activator of GR, has been
found to confer chemoresistance in oesophageal SCC and HCC cells,
suggesting that GR expression may be biologically important in some
GR-expressing carcinomas. This not only suggests why DEX or other
glucocorticoids are not useful in treatment of these malignancies,
but it implies that endogenous, circulating cortisol itself may
actually promote chemoresistance, even in the absence of iatrogenic
glucocorticoid administration. Therefore, these findings suggest
that blockade of GR within such malignant tumors, by preventing
activation by endogenous, circulating cortisol, can play a role in
maintaining or promoting chemosensitivity and/or treating
neoplasia.
[0021] The present invention therefore relates to the use of GR
antagonists (e.g., ORG 34517, RU486, and others) for the treatment
of, for example, esophageal SCC and HCC or other tumors with high
GR expression as a means of inhibiting promotion of chemoresistance
by endogenous cortisol. These effects may be present in all tumor
cells or, when tumors have stem or progenitor cell compartments,
these, specifically, as well. Thus, the present invention relates
to the inhibition of chemoprevention in the bulk of cells making up
a given tumor and/or in the rare stem/progenitor cells within the
tumor that are often responsible for tumor resistance to therapy
and re-occurrence, i.e., as a novel, targeted "cancer stem cell"
treatment.
[0022] To avoid possible negative side effects of systemic blockade
of GR, the present invention further relates to localized tumor
treatment with GR antagonists through direct vascular infusion of
tumor feeding vessels or by direct, intratumoral injection.
[0023] The present invention relates to the use of GR antagonists
for the treatment of, for example, breast and other cancers. The
invention is based on the observation that GR inhibition will
increase tumor cell susceptibility. GR antagonists will block
anti-apoptotic GR signaling in GR-overexpressing breast cancer
cells and subsequently render breast cancer cells more susceptible
to conventional and novel cytotoxic therapies (via blocking GR's
pro-cell survival signaling pathway).
[0024] All references cited herein are incorporated herein by
reference in their entireties.
BRIEF SUMMARY OF THE INVENTION
[0025] The present invention relates to a method for detecting the
presence of a biological substance in a test sample, comprising the
steps of: providing a test sample consisting of, for example,
saliva, or a bodily fluid sample from a subject with, for example,
a lollipop-like apparatus including a stem integrated with the base
and a head integrated with the stem. The stem head includes a
receptor of a sponge like carrier to ensure a high void volume to
absorb sufficient saliva, oral fluid or a bodily fluid sample.
[0026] Combining the test sample with a buffering system (Reagent
1) containing viscosity controllers and stabilizers into a reaction
vessel, mixing the solution well, and expressing all the liquid
from the sample carrier apparatus into Reagent 1 in the reaction
vessel and discarding. Reading the reaction vessel with sample and
buffer for a fluorescence polarization blank and then combining the
test sample and buffer mixture with a fluorescence-labeled ligand
(Reagent 2) to said biological substance in the reaction vessel,
mix solution well, to produce an assay solution. Furthermore,
Reagent 2 may be delivered to the reaction vessel without further
dilution volume of the assay solution.
[0027] The invention provides a method for screening a patient for
a disease state suitable for GCR (glucocorticoid receptor)
antagonist therapy, comprising the steps of: a) obtaining a test
sample from the patient, optionally at a predetermined time, using
a test sample collection unit; b) combining said test sample with a
buffering system to form a mixture in a reaction unit; c) measuring
a parameter of the mixture to determine a blank measurement; d)
combining said test sample and buffer mixture with a labeled ligand
which binds cortisol, wherein the labeled ligand is provided in a
label unit, in the reaction unit to produce an assay solution; or
combining said test sample and buffer mixture and delivering it to
a carrier containing a labeled ligand which binds cortisol, wherein
the labeled ligand is provided in a label unit, in the reaction
unit to produce an assay immobilized complex; e) measuring a
parameter of said assay solution or complex; f) comparing the
measurement of the assay solution relative to the blank
measurement; g) determining the patient's circulating cortisol
levels based on the change of the measurement; and h) comparing the
patient's measured cortisol levels to a predetermined reference
range cortisol levels, wherein when the level of cortisol is
elevated relative to the predetermined reference range, then the
patient has a disease state which involves elevated cortisol, and
thus has a disease state which is a potential candidate for GCR
antagonist therapy. The invention further provides a method wherein
the patient's test sample is selected from the group consisting of
saliva, blood, plasma, serum, urine, other bodily fluids, and
combinations thereof. The invention further provides a method
wherein the sample is obtained from the patient over more than one
time, and the predetermined time is selected from the group
consisting of morning, noon, and evening. The invention further
provides a method wherein the sample is obtained from the patient
over consecutive days. The invention further provides a method
wherein the method is to determine the circadian cycle of the
cortisol levels in the patient, and the predetermined time is
selected from the group consisting of about hourly, every 4 hours,
every 6 hours, every 8 hours, and every 12 hours. The invention
further provides a method wherein the sample is obtained from the
patient over consecutive days. The invention further provides a
method wherein the predetermined reference range is a medically
standard reference range. The invention further provides a method
wherein the predetermined reference range is the patient's
previously measured level. The invention further provides a method
wherein the ligand is detectably labeled with a moiety selected
from the group consisting of a radioisotope, a fluorophore, a
quencher of fluorescence, an enzyme, an affinity tag, and an
epitope tag. The invention further provides a method wherein said
measuring of said parameter of said mixture and said assay solution
is performed using a method selected from spectroscopic,
photochemical, radiochemical, biochemical, enzymatic,
immunochemical, chemical label displacement, surface plasmon
resonance, fluorescence resonance energy transfer, fluorescence
quenching, lateral flow, and fluorescence polarization means. The
invention further provides a method further comprising the steps
of: i) determining a patient's elevated cortisol level; and j)
providing a therapeutic for such elevated cortisol level, wherein
the therapeutic comprises GCR antagonist therapy. The invention
further provides a method wherein the patient has changed patterns
of cortisol levels that have been observed in connection with
abnormal Adrenocorticotropic hormone (ACTH) levels. The invention
further provides a method wherein the patient has non-normal
cortisol levels produced by the adrenal cortex or disordered
circadian rhythms, as a method for selecting subjects for GCR
antagonist therapy wherein the patient has cortisol levels selected
from the group consisting abnormally high cortisol levels but
maintained circadian rhythm, over responsiveness to normal levels,
and high night time cortisol levels as a feature of disrupted
circadian rhythm. The invention further provides a method wherein
the disease state is selected from the group consisting of cancer,
clinical depression, psychological stress, and physiological
stressors such as hypoglycemia, illness, fever, trauma, surgery,
fear, pain, physical exertion, or temperature extremes.
[0028] The invention provides a method for monitoring changes in
cortisol levels in response to treatment, in patients who have
non-normal cortisol levels produced by the adrenal cortex,
comprising: a) obtaining a test sample from the patient, optionally
at a predetermined time, using a test sample collection unit; b)
combining said test sample with a buffering system to form a
mixture in a reaction unit; c) measuring a parameter of the mixture
to determine a blank measurement; d) combining said test sample and
buffer mixture with a labeled ligand which binds cortisol, wherein
the labeled ligand is provided in a label unit, in the reaction
unit to produce an assay solution; or combining said test sample
and buffer mixture and delivering it to a carrier containing a
labeled ligand which binds cortisol, wherein the labeled ligand is
provided in a label unit, in the reaction unit to produce an assay
immobilized complex; e) measuring a parameter of said assay
solution or complex; f) comparing the measurement of the assay
solution relative to the blank measurement; g) determining the
patient's circulating cortisol levels based on the change of the
measurement; h) administering a GCR antagonist; i) repeating steps
a) to 1) after the therapy has been administered; and j)
determining the patient's circulating cortisol levels post-therapy,
wherein when the cortisol levels change in response to treatment to
indicate responsiveness to the GCR antagonist. The invention
further provides a method wherein the patient's test sample is
selected from the group consisting of saliva, blood, plasma, serum,
urine, other bodily fluids, and combinations thereof. The invention
further provides a method wherein the sample is obtained from the
patient over more than one day, and the predetermined time is
selected from the group consisting of morning, noon, and evening.
The invention further provides a method wherein the sample is
obtained from the patient over consecutive days. The invention
further provides a method wherein the method is to determine the
circadian cycle of the cortisol levels in the patient, and the
predetermined time is selected from the group consisting of hourly,
every 4 hours, every 6 hours, every 8 hours, and every 12 hours.
The invention further provides a method wherein the sample is
obtained from the patient over consecutive days. The invention
further provides a method wherein the predetermined reference range
is a medically standard reference range. The invention further
provides a method wherein the predetermined reference range is the
patient's previously measured level. The invention further provides
a method wherein the ligand is detectably labeled with a moiety
selected from the group consisting of a radioisotope, a
fluorophore, a quencher of fluorescence, an enzyme, an affinity
tag, and an epitope tag. The invention further provides a method
wherein said measuring of said parameter of said mixture and said
assay solution is performed using a method selected from
spectroscopic, photochemical, radiochemical, biochemical,
enzymatic, immunochemical, chemical label displacement, surface
plasmon resonance, fluorescence resonance energy transfer,
fluorescence quenching, lateral flow, and fluorescence polarization
means.
[0029] The invention provides a method for monitoring changes in
cortisol levels in response to treatment and adjusting the
treatment in response to these changes in a patient who has
non-normal cortisol levels produced by the adrenal cortex,
comprising: a) obtaining a test sample from the patient, optionally
at a predetermined time, using a test sample collection unit; b)
combining said test sample with a buffering system to form a
mixture in a reaction unit; c) measuring a parameter of the mixture
to determine a blank measurement; d) combining said test sample and
buffer mixture with a labeled ligand which binds cortisol, wherein
the labeled ligand is provided in a label unit, in the reaction
unit to produce an assay solution; or combining said test sample
and buffer mixture and delivering it to a carrier containing a
labeled ligand which binds cortisol, wherein the labeled ligand is
provided in a label unit, in the reaction unit to produce an assay
immobilized complex; e) measuring a parameter of said assay
solution or complex; 0 comparing the measurement of the assay
solution relative to the blank measurement; g) determining the
patient's circulating cortisol levels based on the change of the
measurement; h) administering a GCR antagonist; i) repeating steps
a) to f) after the therapy has been administered; j) determining
the patient's circulating cortisol levels post-therapy; and k)
adjusting the GCR antagonist therapy in response to changes in the
patient's cortisol levels post-therapy, wherein the adjustment in
GCR antagonist therapy is to enhance therapeutic efficacy.
[0030] The invention further provides a method wherein the
patient's test sample is selected from the group consisting of
saliva, blood, plasma, serum, urine, other bodily fluids, and
combinations thereof. The invention further provides a method
wherein the sample is obtained from the patient over more than one
time, and the predetermined time is selected from the group
consisting of morning, noon, and evening. The invention further
provides a method wherein the sample is obtained from the patient
over consecutive days. The invention further provides a method
wherein the method is to determine the circadian cycle of the
cortisol levels in the patient, and the predetermined time is
selected from the group consisting of hourly, every 4 hours, every
6 hours, every 8 hours, and every 12 hours. The invention further
provides a method wherein the sample is obtained from the patient
over consecutive days. The invention further provides a method
wherein the predetermined reference range is a medically standard
reference range. The invention further provides a method wherein
the predetermined reference range is the patient's previously
measured level. The invention further provides a method wherein the
ligand is detectably labeled with a moiety selected from the group
consisting of a radioisotope, a fluorophore, a quencher of
fluorescence, an enzyme, an affinity tag, and an epitope tag. The
invention further provides a method wherein said measuring of said
parameter of said mixture and said assay solution is performed
using a method selected from spectroscopic, photochemical,
radiochemical, biochemical, enzymatic, immunochemical, chemical
label displacement, surface plasmon resonance, fluorescence
resonance energy transfer, fluorescence quenching, lateral flow,
and fluorescence polarization means. The invention further provides
a method wherein the GCR antagonist is selected from the group
consisting of compounds which are selective for GCR, compounds
which non-specifically bind steroid hormone receptors, and
compounds which cross-react to both GCR and other steroid hormone
receptors. The invention further provides a method wherein a
decision to adjust the GCR antagonist therapy in response to
changes in the cortisol levels, post-therapy, is made by a medical
professional. The invention further provides a method further
comprising the step of monitoring changes in biomarker expression
using a nucleic acid microarray. The invention further provides a
method wherein in the patients having normal baseline cortisol at
the start of treatment, and changing cortisol levels during
treatment indicate responsiveness to the GCR antagonist. The
invention further provides a method wherein the combined system of
salivary cortisol quantification as an enabling device for its
paired GCR antagonist will identify patients for whom GCR
antagonism has a likely benefit. The invention further provides a
method wherein the GCR antagonist is selected from the group
consisting of ORG 34517, 11-(substituted phenyl)-estra-4,9-diene
derivatives, and 11-(substituted phenyl)-estra-4,9-diene
derivatives of formula I
##STR00001##
wherein A is a residue of a 5- or 6-membered ring containing 2
heteroatoms which are not connected to each other and independently
selected from O and S, the ring being optionally substituted with
one or more halogen atoms, or A is a residue of a 5- or 6-membered
ring wherein no double C--C bonds are present, containing 1
heteroatom selected from O and S, which heteroatom is connected to
the phenyl group at the position indicated with an asterisk, the
ring being optionally substituted with one or more halogen atoms;
R1 is H or 1-oxo(1-4C)alkyl; R2 is H, (1-8C)alkyl, halogen or CF3;
X is selected from (H,OH), O, and NOH; and the interrupted line
represents an optional bond. The invention further provides a
method wherein the buffering system comprises additional components
selected form the group consisting of viscosity controllers,
stabilizers, and combinations thereof. The invention further
provides a method wherein the fluorescence-labeled ligand which
binds cortisol is selected from the group consisting of an aptamer,
an antibody, an antibody fragment, a receptor, a receptor fragment,
a binding polypeptide, a binding peptide, and combinations thereof.
The invention further provides a method wherein the test sample is
collected from the patient with a lollipop-like apparatus,
including a stem integrated with the base and a head integrated
with the stem, and further wherein the stem head including a
receptor of a sponge like carrier to ensure a high void volume to
absorb sufficient sample.
[0031] The invention provides a method of treating major depressive
disorder in a patient in need thereof by determining whether the
patient has major depressive disorder suitable for GCR
(glucocorticoid receptor) antagonist therapy, comprising the steps
of a) obtaining a test sample from the patient, optionally at a
predetermined time, using a test sample collection unit; b)
combining said test sample with a buffering system to form a
mixture in a reaction unit; c) measuring a parameter of the mixture
to determine a blank measurement; d) combining said test sample and
buffer mixture with a labeled ligand which binds cortisol, wherein
the labeled ligand is provided in a label unit, in the reaction
unit to produce an assay solution; or combining said test sample
and buffer mixture and delivering it to a carrier containing a
labeled ligand which binds cortisol, wherein the labeled ligand is
provided in a label unit, in the reaction unit to produce an assay
immobilized complex; e) measuring a parameter of said assay
solution or complex; f) comparing the measurement of the assay
solution relative to the blank measurement; g) determining the
patient's circulating cortisol levels based on the change of the
measurement; h) comparing the measured cortisol levels to a
predetermined reference range cortisol levels, wherein when the
level of cortisol is elevated relative to the a predetermined
reference range, then the patient has major depressive disorder
which involves elevated cortisol, and thus has major depressive
disorder suitable for GCR (glucocorticoid receptor) antagonist
therapy; and i) when the patient has major depressive disorder
suitable for GCR antagonist therapy, a administering at least one
GCR antagonist. The invention further provides a method wherein the
patient's test sample is selected from the group consisting of
saliva, blood, plasma, serum, urine, other bodily fluids, and
combinations thereof. The invention further provides a method
wherein the sample is obtained from the patient over more than one
time, and the predetermined time is selected from the group
consisting of morning, noon, and evening. The invention further
provides a method wherein the sample is obtained from the patient
over consecutive days. The invention further provides a method
wherein the method is to determine the circadian cycle of the
cortisol levels in the patient, and the predetermined time is
selected from the group consisting of hourly, every 4 hours, every
6 hours, every 8 hours, and every 12 hours. The invention further
provides a method wherein the sample is obtained from the patient
over consecutive days. The invention further provides a method
wherein the predetermined reference range is a medically standard
reference range. The invention further provides a method wherein
the predetermined reference range is the patient's previously
measured level. The invention further provides a method wherein the
ligand is detectably labeled with a moiety selected from the group
consisting of a radioisotope, a fluorophore, a quencher of
fluorescence, an enzyme, an affinity tag, and an epitope tag. The
invention further provides a method wherein said measuring of said
parameter of said mixture and said assay solution is performed
using a method selected from spectroscopic, photochemical,
radiochemical, biochemical, enzymatic, immunochemical, chemical
label displacement, surface plasmon resonance, fluorescence
resonance energy transfer, fluorescence quenching, lateral flow,
and fluorescence polarization means. The invention further provides
a method wherein the GCR (glucocorticoid receptor) antagonist is
selected from the group consisting of ORG 34517, 11-(substituted
phenyl)-estra-4,9-diene derivatives, and 11-(substituted
phenyl)-estra-4,9-diene derivatives of formula I
##STR00002##
wherein A is a residue of a 5- or 6-membered ring containing 2
heteroatoms which are not connected to each other and independently
selected from O and S, the ring being optionally substituted with
one or more halogen atoms, or A is a residue of a 5- or 6-membered
ring wherein no double C--C bonds are present, containing 1
heteroatom selected from O and S, which heteroatom is connected to
the phenyl group at the position indicated with an asterisk, the
ring being optionally substituted with one or more halogen atoms;
R1 is H or 1-oxo(1-4C)alkyl; R2 is H, (1-8C)alkyl, halogen or CF3;
X is selected from (H,OH), O, and NOH; and the interrupted line
represents an optional bond.
[0032] The invention provides a method of treating psychotic
depression in a patient in need thereof by determining whether the
patient has psychotic depression suitable for GCR (glucocorticoid
receptor) antagonist therapy, comprising the steps of a) obtaining
a test sample from the patient, optionally at a predetermined time
using a test sample collection unit; b) combining said test sample
with a buffering system to form a mixture in a reaction unit; c)
measuring a parameter of the mixture to determine a blank
measurement; d) combining said test sample and buffer mixture with
a labeled ligand which binds cortisol, wherein the labeled ligand
is provided in a label unit, in the reaction unit to produce an
assay solution; or combining said test sample and buffer mixture
and delivering it to a carrier containing a labeled ligand which
binds cortisol, wherein the labeled ligand is provided in a label
unit, in the reaction unit to produce an assay immobilized complex;
e) measuring a parameter of said assay solution or complex; 1)
comparing the measurement of the assay solution relative to the
blank measurement; g) determining the patient's circulating
cortisol levels based on the change of the measurement; h)
comparing the measured cortisol levels to a predetermined reference
range cortisol levels, wherein when the level of cortisol is
elevated relative to the predetermined reference range, then the
patient has psychotic depression which involves elevated cortisol,
and thus has psychotic depression suitable for GCR antagonist
therapy; and i) when the patient has psychotic depression suitable
for GCR antagonist therapy, administering at least one GCR
antagonist. The invention further provides a method wherein the
patient's test sample is selected from the group consisting of
saliva, blood, plasma, serum, urine, other bodily fluids, and
combinations thereof. The invention further provides a method
wherein the sample is obtained from the patient over more than one
time, and the predetermined time is selected from the group
consisting of morning, noon, and evening. The invention further
provides a method wherein the sample is obtained from the patient
over consecutive days. The invention further provides a method
wherein the method is to determine the circadian cycle of the
cortisol levels in the patient, and the predetermined time is
selected from the group consisting of hourly, every 4 hours, every
6 hours, every 8 hours, and every 12 hours. The invention further
provides a method wherein the sample is obtained from the patient
over consecutive days. The invention further provides a method
wherein the predetermined reference range is a medically standard
reference range. The invention further provides a method wherein
the predetermined reference range is the patient's previously
measured level. The invention further provides a method wherein the
ligand is detectably labeled with a moiety selected from the group
consisting of a radioisotope, a fluorophore, a quencher of
fluorescence, an enzyme, an affinity tag, and an epitope tag. The
invention further provides a method wherein said measuring of said
parameter of said mixture and said assay solution is performed
using a method selected from spectroscopic, photochemical,
radiochemical, biochemical, enzymatic, immunochemical, chemical
label displacement, surface plasmon resonance, fluorescence
resonance energy transfer, fluorescence quenching, lateral flow,
and fluorescence polarization means. The invention further provides
a method wherein the GCR antagonist is selected from the group
consisting of ORG 34517, 11-(substituted phenyl)-estra-4,9-diene
derivatives, and 11-(substituted phenyl)-estra-4,9-diene
derivatives of formula I
##STR00003##
wherein A is a residue of a 5- or 6-membered ring containing 2
heteroatoms which are not connected to each other and independently
selected from O and S, the ring being optionally substituted with
one or more halogen atoms, or A is a residue of a 5- or 6-membered
ring wherein no double C--C bonds are present, containing 1
heteroatom selected from O and S, which heteroatom is connected to
the phenyl group at the position indicated with an asterisk, the
ring being optionally substituted with one or more halogen atoms;
R1 is H or 1-oxo(1-4C)alkyl; R2 is H, (1-8C)alkyl, halogen or CF3;
X is selected from (H,OH), O, and NOH; and the interrupted line
represents an optional bond.
[0033] The invention provides a method of treating stress-induced
cortisol elevation in a patient in need thereof by determining
whether the patient has stress-induced cortisol elevation suitable
for GCR (glucocorticoid receptor) antagonist therapy, comprising
the steps of: a) obtaining a test sample from the patient,
optionally at a predetermined time, using a test sample collection
unit; b) combining said test sample with a buffering system to form
a mixture in a reaction unit; c) measuring a parameter of the
mixture to determine a blank measurement; d) combining said test
sample and buffer mixture with a labeled ligand which binds
cortisol, wherein the labeled ligand is provided in a label unit,
in the reaction unit to produce an assay solution; or combining
said test sample and buffer mixture and delivering it to a carrier
containing a labeled ligand which binds cortisol, wherein the
labeled ligand is provided in a label unit, in the reaction unit to
produce an assay immobilized complex; e) measuring a parameter of
said assay solution or complex;
f) comparing the measurement of the assay solution relative to the
blank measurement; g) determining the patient's circulating
cortisol levels based on the change of the measurement; h)
comparing the measured cortisol levels to a predetermined reference
range cortisol levels, wherein when the level of cortisol is
elevated relative to the predetermined reference range, then the
patient has stress-induced cortisol elevation which involves
elevated cortisol, and thus has stress-induced cortisol elevation
suitable for GCR antagonist therapy; and i) when the patient has
stress-induced cortisol elevation suitable for GCR antagonist
therapy, administering at least one GCR antagonist. The invention
further provides a method wherein the patient's test sample is
selected from the group consisting of saliva, blood, plasma, serum,
urine, other bodily fluids, and combinations thereof. The invention
further provides a method wherein the stress-related cortisol
elevation is related to a hospital stay, medical treatment, an
institutional stay, clinical depression, psychological stress,
physiological stressors, hypoglycemia, illness, fever, trauma,
surgery, fear, pain, physical exertion, or temperature extremes.
The invention further provides a method wherein the patient is an
elderly individual. The invention further provides a method wherein
the patient has autism or Asperger's syndrome. The invention
further provides a method wherein the sample is obtained from the
patient over more than one time, and the predetermined time is
selected from the group consisting of morning, noon, and evening.
The invention further provides a method wherein the sample is
obtained from the patient over consecutive days. The invention
further provides a method wherein the method is to determine the
circadian cycle of the cortisol levels in the patient, and the
predetermined time is selected from the group consisting of hourly,
every 4 hours, every 6 hours, every 8 hours, and every 12 hours.
The invention further provides a method wherein the sample is
obtained from the patient over consecutive days. The invention
further provides a method wherein the predetermined reference range
is a medically standard reference range. The invention further
provides a method wherein the predetermined reference range is the
patient's previously measured level. The invention further provides
a method wherein the ligand is detectably labeled with a moiety
selected from the group consisting of a radioisotope, a
fluorophore, a quencher of fluorescence, an enzyme, an affinity
tag, and an epitope tag. The invention further provides a method
wherein said measuring of said parameter of said mixture and said
assay solution is performed using a method selected from
spectroscopic, photochemical, radiochemical, biochemical,
enzymatic, immunochemical, chemical label displacement, surface
plasmon resonance, fluorescence resonance energy transfer,
fluorescence quenching, lateral flow, and fluorescence polarization
means. The invention further provides a method wherein the GCR
antagonist is selected from the group consisting of ORG 34517,
11-(substituted phenyl)-estra-4,9-diene derivatives, and
11-(substituted phenyl)-estra-4,9-diene derivatives of formula
I
##STR00004##
wherein A is a residue of a 5- or 6-membered ring containing 2
heteroatoms which are not connected to each other and independently
selected from O and S, the ring being optionally substituted with
one or more halogen atoms, or A is a residue of a 5- or 6-membered
ring wherein no double C--C bonds are present, containing 1
heteroatom selected from O and S, which heteroatom is connected to
the phenyl group at the position indicated with an asterisk, the
ring being optionally substituted with one or more halogen atoms;
R1 is H or 1-oxo(1-4C)alkyl; R2 is H, (1-8C)alkyl, halogen or CF3;
X is selected from (H,OH), O, and NOH; and the interrupted line
represents an optional bond.
[0034] The invention provides a method of treating post-traumatic
stress disorder in a patient in need thereof by determining whether
the patient has post-traumatic stress disorder suitable for GCR
(glucocorticoid receptor) antagonist therapy, comprising the steps
of: a) obtaining a test sample from the patient, optionally at a
predetermined time, using a test sample collection unit; b)
combining said test sample with a buffering system to form a
mixture in a reaction unit; c) measuring a parameter of the mixture
to determine a blank measurement; d) combining said test sample and
buffer mixture with a labeled ligand which binds cortisol, wherein
the labeled ligand is provided in a label unit, in the reaction
unit to produce an assay solution; or combining said test sample
and buffer mixture and delivering it to a carrier containing a
labeled ligand which binds cortisol, wherein the labeled ligand is
provided in a label unit, in the reaction unit to produce an assay
immobilized complex; e) measuring a parameter of said assay
solution or complex; comparing the measurement of the assay
solution relative to the blank measurement; g) determining the
patient's circulating cortisol levels based on the change of the
measurement; h) comparing the measured cortisol levels to a
predetermined reference range cortisol levels, wherein when the
level of cortisol is elevated relative to the predetermined
reference range, then the patient has post-traumatic stress
disorder which involves elevated cortisol, and thus has
post-traumatic stress disorder suitable for GCR (glucocorticoid
receptor) antagonist therapy; and i) when the patient has
post-traumatic stress disorder suitable for GCR antagonist therapy,
administering at least one GCR antagonist. The invention further
provides a method wherein the patient's test sample is selected
from the group consisting of saliva, blood, plasma, serum, urine,
other bodily fluids, and combinations thereof. The invention
further provides a method wherein the sample is obtained from the
patient over more than one time, and the predetermined time is
selected from the group consisting of morning, noon, and evening.
The invention further provides a method wherein the sample is
obtained from the patient over consecutive days. The invention
further provides a method wherein the method is to determine the
circadian cycle of the cortisol levels in the patient, and the
predetermined time is selected from the group consisting of hourly,
every 4 hours, every 6 hours, every 8 hours, and every 12 hours.
The invention further provides a method wherein the sample is
obtained from the patient over consecutive days. The invention
further provides a method wherein the predetermined reference range
is a medically standard reference range. The invention further
provides a method wherein the predetermined reference range is the
patient's previously measured level. The invention further provides
a method wherein the ligand is detectably labeled with a moiety
selected from the group consisting of a radioisotope, a
fluorophore, a quencher of fluorescence, an enzyme, an affinity
tag, and an epitope tag. The invention further provides a method
wherein said measuring of said parameter of said mixture and said
assay solution is performed using a method selected from
spectroscopic, photochemical, radiochemical, biochemical,
enzymatic, immunochemical, chemical label displacement, surface
plasmon resonance, fluorescence resonance energy transfer,
fluorescence quenching, lateral flow, and fluorescence polarization
means. The invention further provides a method wherein the GCR
antagonist is selected from the group consisting of ORG 34517,
11-(substituted phenyl)-estra-4,9-diene derivatives, and
11-(substituted phenyl)-estra-4,9-diene derivatives of formula
I
##STR00005##
wherein A is a residue of a 5- or 6-membered ring containing 2
heteroatoms which are not connected to each other and independently
selected from O and S, the ring being optionally substituted with
one or more halogen atoms, or A is a residue of a 5- or 6-membered
ring wherein no double C--C bonds are present, containing 1
heteroatom selected from O and S, which heteroatom is connected to
the phenyl group at the position indicated with an asterisk, the
ring being optionally substituted with one or more halogen atoms;
R1 is H or 1-oxo(1-4C)alkyl; R2 is H, (1-8C)alkyl, halogen or CF3;
X is selected from (H,OH), O, and NOH; and the interrupted line
represents an optional bond.
[0035] The invention provides a method of prevention of weight gain
in patients using anti-psychotic or anti-depressant medications in
a patient in need thereof, wherein the weight gain is suitable for
GCR (glucocorticoid receptor) antagonist therapy, comprising the
steps of a) obtaining a test sample from the patient, optionally at
a predetermined time using a test sample collection unit; b)
combining said test sample with a buffering system to form a
mixture in a reaction unit; c) measuring a parameter of the mixture
to determine a blank measurement; d) combining said test sample and
buffer mixture with a labeled ligand which binds cortisol, wherein
the labeled ligand is provided in a label unit, in the reaction
unit to produce an assay solution; or combining said test sample
and buffer mixture and delivering it to a carrier containing a
labeled ligand which binds cortisol, wherein the labeled ligand is
provided in a label unit, in the reaction unit to produce an assay
immobilized complex; e) measuring a parameter of said assay
solution or complex;
f) comparing the measurement of the assay solution relative to the
blank measurement; g) determining the patient's circulating
cortisol levels based on the change of the measurement; h)
comparing the measured cortisol levels to a predetermined reference
range cortisol levels, wherein when the level of cortisol is
elevated relative to the predetermined reference range, then the
patient has weight gain which involves elevated cortisol, which is
suitable for GCR (glucocorticoid receptor) antagonist therapy; and
i) when the patient has weight gain suitable for GCR antagonist
therapy, administering at least one GCR antagonist. The invention
further provides a method wherein the patient's test sample is
selected from the group consisting of saliva, blood, plasma, serum,
urine, other bodily fluids, and combinations thereof. The invention
further provides a method wherein the sample is obtained from the
patient over more than one time, and the predetermined time is
selected from the group consisting of morning, noon, and evening.
The invention further provides a method wherein the sample is
obtained from the patient over consecutive days. The invention
further provides a method wherein the method is to determine the
circadian cycle of the cortisol levels in the patient, and the
predetermined time is selected from the group consisting of hourly,
every 4 hours, every 6 hours, every 8 hours, and every 12 hours.
The invention further provides a method wherein the sample is
obtained from the patient over consecutive days. The invention
further provides a method wherein the predetermined reference range
is a medically standard reference range. The invention further
provides a method wherein the predetermined reference range is the
patient's previously measured level. The invention further provides
a method wherein the ligand is detectably labeled with a moiety
selected from the group consisting of a radioisotope, a
fluorophore, a quencher of fluorescence, an enzyme, an affinity
tag, and an epitope tag. The invention further provides a method
wherein said measuring of said parameter of said mixture and said
assay solution is performed using a method selected from
spectroscopic, photochemical, radiochemical, biochemical,
enzymatic, immunochemical, chemical label displacement, surface
plasmon resonance, fluorescence resonance energy transfer,
fluorescence quenching, lateral flow, and fluorescence polarization
means. The invention further provides a method wherein the GCR
antagonist is selected from the group consisting of ORG 34517,
11-(substituted phenyl)-estra-4,9-diene derivatives, and
11-(substituted phenyl)-estra-4,9-diene derivatives of formula
I
##STR00006##
wherein A is a residue of a 5- or 6-membered ring containing 2
heteroatoms which are not connected to each other and independently
selected from O and S, the ring being optionally substituted with
one or more halogen atoms, or A is a residue of a 5- or 6-membered
ring wherein no double C--C bonds are present, containing 1
heteroatom selected from O and S, which heteroatom is connected to
the phenyl group at the position indicated with an asterisk, the
ring being optionally substituted with one or more halogen atoms;
R1 is H or 1-oxo(1-4C)alkyl; R2 is H, (1-8C)alkyl, halogen or CF3;
X is selected from (H,OH), O, and NOH; and the interrupted line
represents an optional bond.
[0036] The invention provides a method of treating Cushing's
syndrome in a patient in need thereof by determining whether the
patient has Cushing's syndrome suitable for GCR (glucocorticoid
receptor) antagonist therapy, comprising the steps of a) obtaining
a test sample from the patient, optionally at a predetermined time,
using a test sample collection unit; b) combining said test sample
with a buffering system to form a mixture in a reaction unit; c)
measuring a parameter of the mixture to determine a blank
measurement; d) combining said test sample and buffer mixture with
a labeled ligand which binds cortisol, wherein the labeled ligand
is provided in a label unit, in the reaction unit to produce an
assay solution; or combining said test sample and buffer mixture
and delivering it to a carrier containing a labeled ligand which
binds cortisol, wherein the labeled ligand is provided in a label
unit, in the reaction unit to produce an assay immobilized complex;
e) measuring a parameter of said assay solution or complex; f)
comparing the measurement of the assay solution relative to the
blank measurement; g) determining the patient's circulating
cortisol levels based on the change of the measurement; h)
comparing the measured cortisol levels to a predetermined reference
range cortisol levels, wherein when the level of cortisol is
elevated relative to the predetermined reference range, then the
patient has Cushing's syndrome which involves elevated cortisol,
which is suitable for GCR (glucocorticoid receptor) antagonist
therapy; and i) when the patient has Cushing's syndrome suitable
for GCR antagonist therapy, administering at least one GCR
antagonist. The invention further provides a method wherein the
patient's test sample is selected from the group consisting of
saliva, blood, plasma, serum, urine, other bodily fluids, and
combinations thereof. The invention further provides a method
wherein the sample is obtained from the patient over more than one
time, and the predetermined time is selected from the group
consisting of morning, noon, and evening. The invention further
provides a method wherein the sample is obtained from the patient
over consecutive days. The invention further provides a method
wherein the method is to determine the circadian cycle of the
cortisol levels in the patient, and the predetermined time is
selected from the group consisting of hourly, every 4 hours, every
6 hours, every 8 hours, and every 12 hours. The invention further
provides a method wherein the sample is obtained from the patient
over consecutive days. The invention further provides a method
wherein the predetermined reference range is a medically standard
reference range. The invention further provides a method wherein
the predetermined reference range is the patient's previously
measured level. The invention further provides a method wherein the
ligand is detectably labeled with a moiety selected from the group
consisting of a radioisotope, a fluorophore, a quencher of
fluorescence, an enzyme, an affinity tag, and an epitope tag. The
invention further provides a method wherein said measuring of said
parameter of said mixture and said assay solution is performed
using a method selected from spectroscopic, photochemical,
radiochemical, biochemical, enzymatic, immunochemical, chemical
label displacement, surface plasmon resonance, fluorescence
resonance energy transfer, fluorescence quenching, lateral flow,
and fluorescence polarization means. The invention further provides
a method wherein the GCR antagonist is selected from the group
consisting of ORG 34517, 11-(substituted phenyl)-estra-4,9-diene
derivatives, and 11-(substituted phenyl)-estra-4,9-diene
derivatives of formula I
##STR00007##
wherein A is a residue of a 5- or 6-membered ring containing 2
heteroatoms which are not connected to each other and independently
selected from O and S, the ring being optionally substituted with
one or more halogen atoms, or A is a residue of a 5- or 6-membered
ring wherein no double C--C bonds are present, containing 1
heteroatom selected from O and S, which heteroatom is connected to
the phenyl group at the position indicated with an asterisk, the
ring being optionally substituted with one or more halogen atoms;
R1 is H or 1-oxo(1-4C)alkyl; R2 is H, (1-8C)alkyl, halogen or CF3;
X is selected from (H,OH), O, and NOH; and the interrupted line
represents an optional bond.
[0037] The invention provides a pharmaceutical composition
comprising: (a) GCR (glucocorticoid receptor) antagonist; (b) the
pharmaceutical composition of (a), further comprising at least one
pharmaceutically acceptable excipient; (c) the pharmaceutical
composition of (a) or (b), wherein the pharmaceutical composition
is formulated or manufactured as a liquid, an elixir, an aerosol, a
spray, a powder, a tablet, a pill, a capsule, a gel, a geltab, a
nanosuspension, a nanoparticle, an extended release dosage form, or
a topical formulation. The invention provides a method for treating
a condition selected from the group consisting of major depressive
disorder, psychotic depression, stress-induced cortisol elevation,
post-traumatic stress disorder, preventing weight gain in patients
using anti-psychotic and anti-depressant medications, or having
Cushing's syndrome, in a patient in need of such treatment
comprising administering the pharmaceutical composition to the
patient.
[0038] The invention provides a kit for the treatment, amelioration
or prevention of a condition selected from the group consisting of
major depressive disorder, psychotic depression, stress-induced
cortisol elevation, post-traumatic stress disorder, preventing
weight gain in patients using anti-psychotic and anti-depressant
medications, or having Cushing's syndrome, in a patient in need of
such treatment comprising: (a) the pharmaceutical composition; and
(b) at least one blister package; a lidded blister; a blister card
or packet; a clamshell; an intravenous (IV) package, IV packette or
IV container; a tray or a shrink wrap comprising the pharmaceutical
composition of (a) and instructions for use of the pharmaceutical
composition.
[0039] The invention provides a product of manufacture comprising a
blister package; a lidded blister; a blister card or packet; a
clamshell; an intravenous (N) package, N packette or IV container;
a tray or a shrink wrap comprising the pharmaceutical composition
and instructions for use of the pharmaceutical composition. The
invention further provides a method wherein the GCR (glucocorticoid
receptor) antagonist selected from the group consisting of ORG
34517, 11-(substituted phenyl)-estra-4,9-diene derivatives, and
11-(substituted phenyl)-estra-4,9-diene derivatives of formula
I
##STR00008##
wherein A is a residue of a 5- or 6-membered ring containing 2
heteroatoms which are not connected to each other and independently
selected from O and S, the ring being optionally substituted with
one or more halogen atoms, or A is a residue of a 5- or 6-membered
ring wherein no double C--C bonds are present, containing 1
heteroatom selected from O and S, which heteroatom is connected to
the phenyl group at the position indicated with an asterisk, the
ring being optionally substituted with one or more halogen atoms;
R1 is H or 1-oxo(1-4C)alkyl; R2 is H, (1-8C)alkyl, halogen or CF3;
X is selected from (H,OH), O, and NOH; and the interrupted line
represents an optional bond.
[0040] The invention provides a method for treating neoplasia
characterized by expression of a glucocorticoid receptor, in a
patient in need of such treatment, comprising: administering to
said animal or human therapeutically effective amounts of each of
at least one neoplasia-treating agent and a GCR (glucocorticoid
receptor) antagonist selected from the group consisting of ORG
34517, 11-(substituted phenyl)-estra-4,9-diene derivatives, and
11-(substituted phenyl)-estra-4,9-diene derivatives of formula
I
##STR00009##
wherein A is a residue of a 5- or 6-membered ring containing 2
heteroatoms which are not connected to each other and independently
selected from O and S, the ring being optionally substituted with
one or more halogen atoms, or A is a residue of a 5- or 6-membered
ring wherein no double C--C bonds are present, containing 1
heteroatom selected from O and S, which heteroatom is connected to
the phenyl group at the position indicated with an asterisk, the
ring being optionally substituted with one or more halogen atoms;
R1 is H or 1-oxo(1-4C)alkyl; R2 is H, (1-8C)alkyl, halogen or CF3;
X is selected from (H,OH), O, and NOH; and the interrupted line
represents an optional bond. The invention further provides a
method wherein ORG34517 is administered as an agent which directly
effects tumor growth, independent of other administered treatment
modalities, for palliation, remission, or cure. The invention
further provides a method wherein said neoplasia treating agent is
radiation. The invention further provides a method wherein said
neoplasia treating agent is a biotherapy agent. The invention
further provides a method wherein said neoplasia treating agent is
a chemotherapy agent. The invention further provides a method
wherein said neoplasia treating agent is a radionuclide. The
invention further provides a method wherein the neoplasia is
selected for the group consisting of hepatocellular carcinoma,
esophageal squamous cell carcinoma, breast cancer, pancreatic
cancer, squamous cell cancer or adenocarcinoma of the head and
neck, colorectal cancer, renal cancer, brain cancer, prostate
cancer, small and non-small cell lung cancer, bladder cancer, bone
or joint cancer, uterine cancer, cervical cancer, multiple myeloma,
hematopoietic malignancies, lymphoma, Hodgkin's disease,
non-Hodgkin's lymphoma, skin cancer, melanoma, squamous cell
carcinoma, leukemia, lung cancer, ovarian cancer, stomach cancer,
Kaposi's sarcoma, laryngeal cancer, endocrine carcinomas, cancer of
the thyroid gland, cancer of the parathyroid gland, cancer of the
pituitary gland, cancer of the adrenal gland, and combinations
thereof. The invention further provides a method wherein the
neoplasia expresses multidrug resistance genes when GR is activated
through binding by endogenous cortisol. The invention further
provides a method wherein the neoplasia expresses proteins of cell
survival pathway (including inhibition of apoptosis) genes when GR
is activated through binding by endogenous cortisol. The invention
further provides a method wherein the neoplasia expresses genes
responsible for epithelial-mesenchymal transition and cell shape
maintenance are repressed when GR is activated through binding by
endogenous cortisol. The invention further provides a method
wherein the neoplasia expresses genes involved in signal
transduction pathways, lipid/fatty acid metabolism, inflammation
and macrophage regulation, transcriptional regulation and chromatin
remodeling, and cell metabolic pathways. The invention further
provides a method wherein tumor stem cells (TSC) express GR,
blockade of which by ORG34517 results in anti-TSC therapy. The
invention further provides a method wherein TSC express multidrug
resistance genes when GR is activated through binding by endogenous
cortisol. The invention further provides a method wherein TSC
express proteins of cell survival pathways (including inhibition of
apoptosis) genes when GR is activated through binding by endogenous
cortisol. The invention further provides a method wherein TSC
express genes responsible for epithelial-mesenchymal transition and
cell shape maintenance are repressed when GR is activated through
binding by endogenous cortisol. The invention further provides a
method wherein TSC express genes involved in signal transduction
pathways, lipid/fatty acid metabolism, inflammation and macrophage
regulation, transcriptional regulation and chromatin remodeling,
and cell metabolic pathways. The invention further provides a
method wherein the neoplasia is chemo-resistant ER/GR+ breast
cancer. The invention further provides a method wherein the
administration of ORG 34517 for GR-blockade, reduces toxicities and
side effects when given systemically. The invention further
provides a method wherein the ORG 34517 given systemically through
oral or intravenous routes. The invention further provides a method
wherein the ORG 34517 is targeted to tumor by intra-arterial
infusion to reduce systemic side effects of GR blockade. The
invention further provides a method wherein the ORG 34517 is given
to accomplish cure or remission of tumor. The invention further
provides a method wherein the ORG 34517 is given to accomplish
reduction of tumor burden to enhance effectiveness of subsequent
surgical resection. The invention further provides a method wherein
the ORG 34517 is given to accomplish reduction of tumor burden to
make an unresectable tumor resectable.
[0041] The invention provides a method for treatment of neoplasia
in a patient comprising targeted delivery of ORG 34517, wherein the
neoplasia expresses multidrug resistance genes when GR is activated
through binding by endogenous cortisol. The invention further
provides a method wherein the HCC patient is not a candidate to
undergo surgical intervention because tumor is too large or
encroaches on liver anatomy in a manner that prevents resection,
delivery of ORG34517 prior to ablative or chemotherapy to shrink
the tumor and make it resectable. The invention further provides a
method wherein the HCC is present in cirrhosis and the patient is
not a candidate for transplantation because of large tumor size,
administration of ORG34517 making the tumor amenable to ablative or
chemotherapy to shrink the tumor and allow the patient to be
eligible for liver transplant. The invention further provides a
method wherein the HCC is present in cirrhosis and the patient is a
candidate for transplantation, administration of ORG34517 making
the tumor amenable to ablative therapy to manage tumor while
patient remains on the liver transplant waitlist.
[0042] The invention provides a method for low toxicity
chemoprevention by targeted liver infusion in patients with forms
of established cirrhosis that are high risk for emergence of HCC,
including those with premalignant lesions diagnosed on biopsy or by
radiology, comprising targeted delivery of Org34517 to intrahepatic
lesions, wherein the targeted delivery of ORG g34517 to
intrahepatic lesions prevents emergence of HCC. The invention
further provides a method wherein the patient with HCC is not a
candidate for undergo surgical intervention. The invention further
provides a method wherein the HCC resides in locations where
surgical or ablative interventions are not available. The invention
further provides a method wherein the patient with HCC has
cirrhosis that is too advanced to make partial hepatectomy safe.
The invention further provides a method wherein the patient with
HCC is too early in their chronic liver disease to qualify for
transplantation. The invention further provides a method wherein
the HCC is too advanced for localized treatments.
[0043] The invention provides a method for treatment of HCC
comprising: a) targeted delivery of ORG 34517 to intrahepatic
lesions, wherein the targeted delivery of ORG 34517 to intrahepatic
lesions improves outcomes of localized chemo-ablative therapies.
The invention further provides a method wherein the treatment is to
help patients qualify for liver transplantation.
[0044] The invention provides a method for low toxicity
chemoprevention by targeted liver infusion in patients with forms
of established cirrhosis that are high risk for emergence of HCC,
including those with premalignant lesions diagnosed on biopsy or by
radiology comprising targeted delivery of ORG 34517 to intrahepatic
lesions, wherein the targeted delivery of ORG 34517 to intrahepatic
lesions prevents emergence of HCC. The invention further provides a
method wherein the neoplasia is eSCC.
[0045] The invention provides a method for treatment of eSCC in a
patient with unresectable eSCC where systemic or targeted
administration of ORG 34517 makes tumor responsive to ablative or
chemotherapies as palliative or curative treatment.
[0046] The invention provides method for treatment of eSCC in a
patient with unresectable eSCC where systemic or targeted
administration of ORG 34517 makes tumor responsive to ablative or
chemotherapies to shrink the tumor and enhance resectability. The
invention further provides a method wherein the neoplasia-treating
agent is a chemotherapeutic agent including but not limited to
gemcitabine, paclitaxel, carboplatin, cisplatin, and
5-fluorouracil. The invention further provides a method wherein the
therapeutic effective amount of glucocorticoid administered is
about 100 to 400 microg/kg body weight per day when administered
intravenously.
[0047] The invention provides a method for treating neoplasia, in
an animal or human in need of such treatment, wherein said
neoplasia comprises neoplastic stem cells characterized by
expression of a glucocorticoid receptor, and further characterized
by expression of multidrug resistance genes or other stem cell
related means of survival when GR is activated through binding by
endogenous cortisol, the method comprising: a) administering to
said animal or human a therapeutically effective amount of ORG
34517; and b) administering to said animal or human a
therapeutically effective amount of the at least one
neoplasia-treating agent, wherein said therapeutically effective
amount of ORG 34517 is an amount sufficient to promote
susceptibility of the neoplastic stem cells to at least one
neoplasia-treating agent. The invention further provides a method
wherein said neoplasia-treating agent is radiation selected from
the group consisting of external beam or radionuclide therapy. The
invention further provides a method wherein said neoplasia-treating
agent is a biotherapy agent. The invention further provides a
method wherein said neoplasia-treating agent is a chemotherapy
agent. The invention further provides a method wherein said
neoplasia-treating agent is a radionuclide. The invention further
provides a method wherein the neoplasia is selected from the group
consisting of hepatocellular carcinoma, esophageal squamous cell
carcinoma, breast cancer, pancreatic cancer, squamous cell cancer
or adenocarcinoma of the head and neck, colorectal cancer, renal
cancer, brain cancer, prostate cancer, small and non-small cell
lung cancer, bladder cancer, bone or joint cancer, uterine cancer,
cervical cancer, multiple myeloma, hematopoietic malignancies,
lymphoma, Hodgkin's disease, non-Hodgkin's lymphoma, skin cancer,
melanoma, squamous cell carcinoma, leukemia, lung cancer, ovarian
cancer, stomach cancer, Kaposi's sarcoma, laryngeal cancer,
endocrine carcinomas, cancer of the thyroid gland, cancer of the
parathyroid gland, cancer of the pituitary gland, cancer of the
adrenal gland, and combinations thereof. The invention further
provides a method wherein the neoplasia expresses multidrug
resistance genes when GR is activated through binding by endogenous
cortisol.
[0048] The invention provides a pharmaceutical composition for
treating neoplasia in a patient which is characterized by
expression of a glucocorticoid receptor, comprising: a)
therapeutically effective amounts of at least one
neoplasia-treating agent; b) a GCR (glucocorticoid receptor)
antagonist selected from the group consisting of ORG 34517,
11-(substituted phenyl)-estra-4,9-diene derivatives, and
11-(substituted phenyl)-estra-4,9-diene derivatives of formula
I
##STR00010##
wherein A is a residue of a 5- or 6-membered ring containing 2
heteroatoms which are not connected to each other and independently
selected from O and S, the ring being optionally substituted with
one or more halogen atoms, or A is a residue of a 5- or 6-membered
ring wherein no double C--C bonds are present, containing 1
heteroatom selected from O and S, which heteroatom is connected to
the phenyl group at the position indicated with an asterisk, the
ring being optionally substituted with one or more halogen atoms;
R1 is H or 1-oxo(1-4C)alkyl; R2 is H, (1-8C)alkyl, halogen or CF3;
X is selected from (H,OH), O, and NOH; and the interrupted line
represents an optional bond; and c) optionally, at least one
pharmaceutically acceptable carrier. The invention further provides
a method wherein said neoplasia-treating agent is selected from the
group consisting of a chemotherapeutic agent, a biotherapeutic
agent, a radionuclide agent, and combinations thereof. The
invention further provides a method wherein the neoplasia is
selected from the group consisting of hepatocellular carcinoma,
esophageal squamous cell carcinoma, breast cancer, pancreatic
cancer, squamous cell cancer or adenocarcinoma of the head and
neck, colorectal cancer, renal cancer, brain cancer, prostate
cancer, small and non-small cell lung cancer, bladder cancer, bone
or joint cancer, uterine cancer, cervical cancer, multiple myeloma,
hematopoietic malignancies, lymphoma, Hodgkin's disease,
non-Hodgkin's lymphoma, skin cancer, melanoma, squamous cell
carcinoma, leukemia, lung cancer, ovarian cancer, stomach cancer,
Kaposi's sarcoma, laryngeal cancer, endocrine carcinomas, cancer of
the thyroid gland, cancer of the parathyroid gland, cancer of the
pituitary gland, cancer of the adrenal gland, and combinations
thereof. The invention further provides a method, wherein the
neoplasia is chemo-resistant ER-GR+ breast cancer. The invention
further provides a method wherein the neoplasia expresses multidrug
resistance genes when GR is activated through binding by endogenous
cortisol. The invention further provides a method wherein the
chemotherapeutic agent is selected from the group comprising:
busulfan, cisplatin, carboplatin, chlorambucil, cyclophosphamide,
ifosfamide, dacarbazine (DTIC), mechlorethamine (nitrogen mustard),
melphalan carmustine (BCNU) lomustine (CCNU), 5-FU, capecitabine,
methotrexate, gemcitabine, cytarabine (ara-C), fludarabine
dactinomycin, daunorubicin, doxorubicin (Adriamycin), idarubicin,
mitoxantrone, paclitaxel, docetaxel, etoposide (VP-16),
vinblastine, vincristine, vinorelbine prednisone, dexamethasone,
tamoxifen, fulvestrant, anastrozole, letrozole, megestrol acetate,
bicalutamide, flutamide, leuprolide, goserelin, L-asparaginase, and
tretinoin, gemcitabine, paclitaxel, carboplatin, 5-FU, and
combinations thereof.
[0049] The invention provides a diagnostic kit comprises the
following components: a) test sample collection unit; b) the buffer
system unit; c) the reaction unit; and d) the label unit, wherein
the components are in a blister package; a lidded blister; a
blister card or packet; a clamshell; a tray, or a shrink wrap, and
instructions for use of the kit. The invention further provides
test sample collection unit wherein the test sample collection unit
comprises a stem integrated with a base, and a head integrated with
the stem. The invention further provides a test sample collection
unit wherein the stem head comprises a receptor of a sponge-like
carrier sufficient to ensure a high void volume to absorb
sufficient saliva, oral fluid or a bodily fluid sample. The
invention further provides a buffer system unit wherein the buffer
system unit comprises additional components selected from the group
consisting of viscosity controllers, stabilizers, and combinations
thereof. The invention further provides a reaction unit wherein the
reaction unit is adapted to fit in a fluorescent polarization
reader. The invention further provides a label unit wherein the
label unit comprises a fluorescence-labeled ligand which binds
cortisol, wherein the fluorescence-labeled ligand which binds
cortisol is selected from the group consisting of an aptamer, an
antibody, an antibody fragment, a receptor, a receptor fragment, a
binding polypeptide, a binding peptide, and combinations thereof.
The invention further provides a diagnostic kit wherein the reader
apparatus provides temperature control and on-board mixing as an
aid in viscosity control of the reaction to ensure better accuracy
and precision. The invention further provides a diagnostic kit
wherein the reader is a miniaturized, portable apparatus for
measuring fluorescence polarization of a liquid sample by direct or
indirect methods. The invention further provides a diagnostic kit
wherein system also enables continual monitoring of the patient
during treatment for assessment of responsiveness to treatment.
BRIEF DESCRIPTION OF SEVERAL VIEWS OF THE DRAWINGS
[0050] The invention will be described in conjunction with the
following drawings in which like reference numerals designate like
elements and wherein:
[0051] FIG. 1 is an example of a sample carrier.
[0052] FIG. 2 is an example of glass reaction vessel with reagent
one dispensed inside.
[0053] FIG. 3 is an example of a fluorescent ligand reagent two,
dry formulation to be dissolved in the reaction vessel.
[0054] FIGS. 4A through 4D show examples of additional embodiments
of the chemistry process configuration. FIG. 4A shows a blister
pack of reagent cartridge units. FIG. 4B shows an exemplary
reaction vessel and cap. FIG. 4C shows an exemplary sample carrier.
FIG. 4D shows an exemplary reagent 2 (fluorescent ligand)
carrier.
[0055] FIGS. 5A through 5C show examples of additional embodiments
of the chemistry process configuration. FIG. 5A shows an exemplary
sample carrier. FIG. 5B shows an exemplary reaction vessel and cap.
FIG. 5C shows an exemplary reagent 2 (fluorescent ligand)
carrier.
[0056] FIGS. 6A and 6B show additional embodiments of chemistry
process configuration-plastic cartridge. FIG. 6A shows an exemplary
sample carrier. FIG. 6B shows an exemplary reagent 2 (fluorescent
ligand) carrier.
[0057] FIG. 7 is an example of a Fluorescence Polarization Reader
(DC and battery operated wt.<3 lbs).
[0058] FIG. 8 is Effect of Vehicle (Ethanol/Castor Oil)/Vehicle
(Ethanol/Sesame Oil), Vehicle (Ethanol/Castor Oil)/Taxol (10
mg/kg/day) and ORG 34517 (20.5 mg/kg/day)/Taxol (10 mg/kg/day) on
relative tumor growth. Compounds are administered day 1-5.
[0059] FIG. 9 shows the Effect of Vehicle (Ethanol/Castor
Oil)/Taxol (10 mg/kg/day) and ORG 34517 (20.5 mg/kg/day)/Taxol (10
mg/kg/day) on relative tumor growth. The average curve for Veh/Taxo
is shown. 517/Taxocurves are show for each individual animal.
Compounds are administered day 1-5.
[0060] FIG. 10 shows the results of an experiment in which mice
were implanted with cultured ER-GR+ human breast cancer cells. As
tumor volume in each mouse approached the test threshold of 200
mm3, the mice were randomized to receive intraperitoneal injections
of vehicle alone, chemotherapy (Paclitaxel) alone, and chemotherapy
and ORG 34517. Each group contained 3 mice. Results show
significant differences in attained tumor volume
[0061] FIG. 11 shows the AUC.sub.0-24. Two dogs were included in
the test. The nanosuspension increases the exposure of
(11b,17b)-11-(1,3-benzodioxol-5-yl)-17-hydroxy-17-(1-propynyl)estra-4,9-d-
ien-3-one.
DETAILED DESCRIPTION OF THE INVENTION
[0062] This invention relates to a low cost rapid response
diagnostic system to determine salivary cortisol levels in patients
selected as potential candidates for GCR (glucocorticoid receptor)
antagonist therapy utilizing a GCR antagonist, such as ORG 34517.
The rapid, sensitive, and inexpensive test can be used to determine
patients who have non-normal cortisol production or disordered
circadian rhythms as a method for selecting subjects for GCR
antagonist therapy for whom it is likely to have beneficial and/or
therapeutic effects, and can also be used to monitor changes in
cortisol levels in response to treatment.
[0063] As used herein, the term "agent" refers to any molecule,
compound, methodology and/or substance for use in the prevention,
treatment, management and/or diagnosis of cancer. Non-limiting
examples of cancer therapies include chemotherapies, radiation
therapies, hormonal therapies, anti-angiogenesis therapies,
targeted therapies, and/or biological therapies including
immunotherapies and surgery
[0064] As used herein, the term "cancer cells" refer to cells that
acquire a characteristic set of functional capabilities during
their development, including the ability to evade apoptosis,
self-sufficiency in growth signals, insensitivity to anti-growth
signals, tissue invasion/metastasis, significant growth potential,
and/or sustained angiogenesis. The term "cancer cell" is meant to
encompass both pre-malignant and malignant cancer cells.
[0065] As used herein, the term "cancer stem cell(s)" refers to a
cell that can be a progenitor of a highly proliferative cancer
cell. A cancer stem cell has the ability to re-grow a tumor as
demonstrated by its ability to form tumors in immunocompromised
mice, and typically to form tumors upon subsequent serial
transplantation in immunocompromised mice. Cancer stem cells are
also typically slow-growing relative to the bulk of a tumor; that
is, cancer stem cells are generally quiescent. In certain
embodiments, but not all, the cancer stem cell may represent
approximately 0.1 to 10% of a tumor.
[0066] As used herein, the phrase "diagnostic agent" refers to any
molecule, compound, and/or substance that is used for the purpose
of diagnosing a disease or condition. Non-limiting examples of
diagnostic agents include antibodies, antibody fragments, or other
proteins, including those conjugated to a detectable agent. As used
herein, the term "detectable agents" refer to any molecule,
compound and/or substance that is detectable by any methodology
available to one of skill in the art. Non-limiting examples of
detectable agents include dyes, gases, metals, or radioisotopes. As
used herein, diagnostic agent and "imaging agent" are equivalent
terms.
[0067] As used herein, the term "effective amount" refers to the
amount of a therapy that is sufficient to result in the prevention
of the development, recurrence, or onset of a disease or condition,
such as cancer, and one or more symptoms thereof, to enhance or
improve the prophylactic effect(s) of another therapy, reduce the
severity, the duration of a disease or condition, such as cancer,
ameliorate one or more symptoms of a disease or condition, such as
cancer, prevent the advancement of a disease or condition, such as
cancer, cause regression of a disease or condition, such as cancer,
and/or enhance or improve the therapeutic effect(s) of another
therapy.
[0068] In an embodiment of the invention, the amount of a therapy
is effective to achieve one, two, three, or more results following
the administration of one, two, three or more therapies: (1) a
stabilization, reduction or elimination of the cancer stem cell
population; (2) a stabilization, reduction or elimination in the
cancer cell population; (3) a stabilization or reduction in the
growth of a tumor or neoplasm; (4) an impairment in the formation
of a tumor; (5) eradication, removal, or control of primary,
regional and/or metastatic cancer; (6) a reduction in mortality;
(7) an increase in disease-free, relapse-free, progression-free,
and/or overall survival, duration, or rate; (8) an increase in the
response rate, the durability of response, or number of patients
who respond or are in remission; (9) a decrease in hospitalization
rate; (10) a decrease in hospitalization lengths; (11) the size of
the tumor is maintained and does not increase or increases by less
than 10%, preferably less than 5%, preferably less than 4%,
preferably less than 2%; (12) an increase in the number of patients
in remission; (13) an increase in the length or duration of
remission; (14) a decrease in the recurrence rate of cancer; (15)
an increase in the time to recurrence of cancer; and (16) an
amelioration of cancer-related symptoms and/or quality of life.
[0069] As used herein, the phrase "pharmaceutically acceptable"
means approved by a regulatory agency of the federal or a state
government, or listed in the U.S. Pharmacopeia, European
Pharmacopeia, or other generally recognized pharmacopeia for use in
animals, and more particularly, in humans.
[0070] As used herein, the term "predetermined reference range"
refers to a reference range for the particular biological entity,
e.g., cortisol, for a subject or a population of subjects. Each
laboratory may establish its own reference range for each
particular assay, or a standard reference range for each assay may
be made available and used locally, regionally, nationally, or
worldwide or may be patient-specific. In one specific embodiment,
the term refers to a reference range for the amount of cortisol in
a patient or a specimen from a patient. In another specific
embodiment, the term refers to a reference range for the amount of
cortisol in a patient or a specimen from a patient.
[0071] As used herein, the terms "prevent," "preventing" and
"prevention" in the context of the administration of a therapy to a
subject refer to the prevention or inhibition of the recurrence,
onset, and/or development of a a disease or condition, such as
cancer or a symptom thereof in a subject resulting from the
administration of a therapy (e.g., a prophylactic or therapeutic
agent), or a combination of therapies (e.g., a combination of
prophylactic or therapeutic agents). In some embodiments, such
terms refer to one, two, three, or more results following the
administration of one or more therapies: (1) a stabilization,
reduction or elimination of the cancer stem cell population; (2) a
stabilization, reduction or elimination in the cancer cell
population; (3) an increase in response rate; (4) an increase in
the length or duration of remission; (5) a decrease in the
recurrence rate of cancer; (6) an increase in the time to
recurrence of cancer; (7) an increase in the disease-free,
relapse-free, progression-free, and/or overall survival of the
patient; and (8) an amelioration of cancer-related symptoms and/or
quality of life. In specific embodiments, such terms refer to a
stabilization, reduction or elimination of the cancer stem cell
population.
[0072] As used herein, the terms "subject" and "patient" are used
interchangeably. As used herein, the term "patient" refers to an
animal, preferably a mammal such as a non-primate (e.g., cows,
pigs, horses, cats, dogs, rats etc.) and a primate (e.g., monkey
and human), and most preferably a human. In some embodiments, the
subject is a non-human animal such as a farm animal (e.g., a horse,
pig, or cow) or a pet (e.g., a dog or cat). In a specific
embodiment, the subject is an elderly human. In another embodiment,
the subject is a human adult. In another embodiment, the subject is
a human child. In yet another embodiment, the subject is a human
infant.
[0073] As used herein, the term "therapeutic agent" refers to any
molecule, compound, and/or substance that is used for the purpose
of treating and/or managing a disease or disorder. Examples of
therapeutic agents include, but are not limited to, proteins,
immunoglobulins (e.g., multi-specific Igs, single chain Igs, Ig
fragments, polyclonal antibodies and their fragments, monoclonal
antibodies and their fragments), peptides (e.g., peptide receptors,
selectins), binding proteins, biologics, chemospecific agents,
chemotoxic agents (e.g., anti-cancer agents), proliferation-based
therapy, radiation, chemotherapy, anti-angiogenic agents, and small
molecule drugs.
[0074] As used herein, the terms "therapies" and "therapy" can
refer to any method(s), composition(s), and/or agent(s) that can be
used in the prevention, treatment and/or management of a disease or
condition, such as cancer, or one or more symptoms thereof. In
certain embodiments, the terms "therapy" and "therapies" refer to
chemotherapy, small molecule therapy, radioimmunotherapy, toxin
therapy, prodrug-activating enzyme therapy, biologic therapy,
antibody therapy, surgical therapy, hormone therapy, immunotherapy,
anti-angiogenic therapy, targeted therapy, epigenetic therapy,
demethylation therapy, histone deacetylase inhibitor therapy,
differentiation therapy, radiation therapy, or a combination of the
foregoing and/or other therapies useful in the prevention,
management and/or treatment of a cancer or one or more symptoms
thereof.
[0075] As used herein, the terms "treat," "treatment," and
"treating" in the context of the administration of a therapy to a
subject refer to the reduction or inhibition of the progression
and/or duration of a disease or condition, such as cancer, the
reduction or amelioration of the severity of a disease or
condition, such as cancer, and/or the amelioration of one or more
symptoms thereof resulting from the administration of one or more
therapies. In specific embodiments, such terms refer to one, two or
three or more results following the administration of one, two,
three or more therapies: (1) a stabilization, reduction or
elimination of the cancer stem cell population; (2) a
stabilization, reduction or elimination in the cancer cell
population; (3) a stabilization or reduction in the growth of a
tumor or neoplasm; (4) an impairment in the formation of a tumor;
(5) eradication, removal, or control of primary, regional and/or
metastatic cancer; (6) a reduction in mortality; (7) an increase in
disease-free, relapse-free, progression-free, and/or overall
survival, duration, or rate; (8) an increase in the response rate,
the durability of response, or number of patients who respond or
are in remission; (9) a decrease in hospitalization rate; (10) a
decrease in hospitalization lengths; (11) the size of the tumor is
maintained and does not increase or increases by less than 10%,
preferably less than 5%, preferably less than 4%, preferably less
than 2%; and (12) an increase in the number of patients in
remission. In certain embodiments, such terms refer to a
stabilization or reduction in the cancer stem cell population. In
some embodiments, such terms refer to a stabilization or reduction
in the growth of cancer cells. In some embodiments, such terms
refer to a stabilization or reduction in the cancer stem cell
population and a reduction in the cancer cell population. In some
embodiments, such terms refer to a stabilization or reduction in
the growth and/or formation of a tumor. In some embodiments, such
terms refer to the eradication, removal, or control of primary,
regional, or metastatic cancer (e.g., the minimization or delay of
the spread of cancer). In some embodiments, such terms refer to a
reduction in mortality and/or an increase in survival rate of a
patient population. In further embodiments, such terms refer to an
increase in the response rate, the durability of response, or
number of patients who respond or are in remission. In some
embodiments, such terms refer to a decrease in hospitalization rate
of a patient population and/or a decrease in hospitalization length
for a patient population.
Cortisol
[0076] The estimated daily cortisol production rate in normal
subjects varies between 4-15 mg/m.sup.2 per day or, according to
more recent studies between 9 and 11 mg/m.sup.2 per day. In order
to describe the 24-hour variation in serum cortisol levels
adequately, the day may be divided into, for example, four phases.
Phase 1 is a 6-hours period of minimal secretory activity 4 h
before and 2 h after onset of sleep. Phase 2 refers to the 3rd to
5th hours of sleep when there is a preliminary nocturnal secretory
episode. Phase 3 is a 4-hour main secretory phase during the last 3
h of sleep and the first hour after wakening. Phase 4 is an 11-hour
phase of intermittent secretory activity when there is a slow
decline in serum levels of cortisol.
[0077] In a study by Mah et al. (Clinical Endocrinology (2004) 61,
367-375) the circadian rhythm of serum cortisol of normal subjects
is described. Peak levels of about 400-800 mmol/l, about 150-300
mmol/l and about 150 mmol/1 are observed at about 6 am, 2 pm and 9
pm, respectively, and the lowest level is about midnight. In this
study it is observed that the endogenous cortisol levels reach
their highest levels within 30 minutes after wake-up. In order to
mimic the circadian rhythm, Mah et al. recommend a thrice-daily
treatment regimen of hydrocortisone, the first dose taken in the
fasted state and delaying the breakfast 1-3 hours and the other two
doses taken 15-60 min before food. A trice-daily regimen is also
recommended in a recent review by Czock et al. (Clin. Pharmacokinet
(2005) 44, 61-98) due to the short half-life of hydrocortisone, and
for prednisolone a twice-daily regimen is preferred over a
once-daily regimen.
Cortisol Test
[0078] The absence of rapid response and inexpensive testing for
cortisol has, heretofore, prevented the linking of GCR antagonists
(e.g., ORG 34517) to a cortisol pre-test for entry into clinical
trials for GCR antagonists and will inhibit the ability to select
the patients most likely to receive the benefit of treatment with
the compounds when available for clinical use. The invention
provides the pairing of an affordable, real-time cortisol test
(e.g., PopTest Cortisol) which will enable the successful
completion of clinical trials for this class of drugs as well as
form the basis for their future, anticipated therapeutic
use(s).
[0079] Conditions that may be treated using, for example, a linked
salivary cortisol quantification test and GCR antagonist (e.g., ORG
34517) system include, but are not limited to the following:
[0080] Major Depressive Disorder (MDD).
[0081] MDD is a psychiatric disorder which has a lifetime
prevalence of around 8%. One of the most consistent findings in
psychiatry is that patients with major depression present with
alterations in the hypothalamic-pituitary-adrenal (HPA) axis. A
significant percentage of depressed patients exhibit hypersecretion
cortisol, as manifested by elevated plasma, cerebrospinal fluid,
and salivary concentrations of cortisol and increased urinary free
cortisol. In addition, many depressed patients exhibit a clear
inability to switch off endogenous cortisol release following
exogenous challenge with the potent synthetic glucocorticoid
dexamethasone (the so-called dexamethasone non-suppressors) (Gold
P. W., et al., Clinical and biochemical manifestations of
depression: relation to neurobiology of stress. New England J. Med.
319, 413-420, 1988). This `sub-group` of severely compromised
patients are most often the ones in whom depression becomes a
life-threatening illness that warrants hospitalization.
[0082] Other abnormalities of the HPA axis found in depressed
patients are increased cortisol response to corticotrophin, a
blunted corticotrophin response to CRH (corticotrophin releasing
hormone), and adrenal and pituitary enlargement (for a review see
Holsboer, F. and Barden, N.: Endocrine Reviews 1996, 17, 187-205).
These observations have been interpreted to suggest a causal
relationship between disturbed functioning of the HPA axis and the
pathology of depression (Murphy, B. E. P. J. of Steroid Biochem.
and Mol. Biol. 1991, 38, 537-559). Therapeutic efficacy of
classical antidepressants has been shown to be preceded by or to
coincide with restoration of the disturbed HPA axis in depression
(Holsboer and Barden, 1996, supra). It has been postulated that any
intervention which can restore this HPA dysfunction may have
antidepressant potential.
[0083] One type of such intervention, studies of which support the
impression that HPA-axis functioning and high circulating cortisol
is a major instigator of major depression is the administration of
glucocorticoid synthesis inhibitors, as has been shown in patients
suffering from Cushing's Syndrome, which is a condition in which
high cortisol levels are reported as a result of adrenal gland
malfunction (due to a pituitary tumour or a secondary tumour, both
producing the cortisol secretagogue ACTH). The depressive symptoms
associated with Cushing's disappear relatively quickly with the
return of cortisol levels to normal. Such treatment may involve
removal of the offending tumour or treatment with cortisol
synthesis inhibitors such as metyrapone, ketoconozole, or
aminoglutethimide (Murphy, B. E. P., Steroids and Depression. J.
Steroid Biochem & Mol. Biol. 38, 537-558, 1991). Similarly,
relatively recent clinical trials have demonstrated that cortisol
synthesis inhibitors can be used to ameliorate depressive symptoms
in severe, treatment-resistant non-Cushing depressives (Murphy, B.
E. P., Can. J. Psych. 43, 279-286, 1998; see also U.S. Pat. No.
4,814,333 (Ravaris, C. L.)).
[0084] Another type of intervention is the use of direct GCR
antagonists, which have much more specific pharmacological effects
as compared to synthesis inhibitors and which may help restore HPA
activity. Small scale pilot clinical studies have been conducted in
order to study the antidepressant activity of the non-selective
glucocorticoid receptor antagonist RU 486 (mifepristone; Murphy, B.
E. P. et al. J. Psychiat. Neurosc. 18, 209-213, 1993). More
recently (Nemeroff, C., Remeron Scientific Expert Meeting,
Budapest, Mar. 29-Apr. 1, 2001) it was demonstrated in a Phase IIB
continuation of this study, that both the number of responders as
well as the efficacy of the psychosis treatment increased with
increasing daily dose of mifepristone as measured by the change in
Brief Psychiatric Rating Scale (50 mg-33% change; 600 mg-40% change
and 1200 mg-52% change). These data indicate that a higher dose of
glucocorticoid receptor antagonist is correlated with a higher
clinical efficacy.
[0085] Non-response to standard treatments, however, reach levels
as high as 50%. (Connolly K R, Drugs. 2011; 71: 43-64.) Frequently,
extra interventions are necessary to get patients to achieve
remission. Various augmentation and combination strategies have
been described in the literature for difficult to treat major MMD
patients.
[0086] Use of an HPA-axis modulating drug in these patients has not
been studied in spite of the fact that there is clear evidence that
at least a sub-group of MDD patients have significant HPA-axis
dysfunction, as noted above. Biological symptoms, indicative of
excessive activity of the HPA-axis, have been reported with great
consistency. In parallel, there is a body of evidence suggesting
that there is an association between HPA-axis functioning and
treatment response, where high HPA-axis activation at baseline, or
post-treatment, is associated with a poorer response to SSRI
treatment or a higher relapse risk.
[0087] Preclinical studies indicate that HPA-axis dysfunction of
the type seen in affective disorders can attenuate the
neurochemical effects of a selective serotonin re-uptake inhibitor
(SSRI) antidepressant. Conversely, in animals with normal HPA axis
function, co-administration of GR antagonists augmented the
neurochemical effects of an SSRI. These data provide a mechanistic
underpinning of the GR antagonist augmentation strategy, and
moreover indicate that the strategy may prove efficacious in
patients both with and without HPA axis dysfunction.
[0088] Small scale pilot clinical studies were conducted in order
to study the antidepressant activity of the non-selective
glucocorticoid receptor antagonist RU 486 (mifepristone; Murphy, B.
E. P. et al. J. Psychiat. Neurosc. 18, 209-213, 1993). A double
blind, 4 week, paroxetine controlled study of ORG 34517 in
depressed patients was carried out. Paroxetine is a selective
serotonin re-uptake inhibitor which is recognized as an effective
antidepressant for major depression. Patients were selected which
had a primary depressive disorder fulfilling the diagnostic
criteria of a MDD as defined by the DSM-IV for recurrent (296.3)
episodes, and who had a severity of depression which resulted in a
total score of at least 22 on the HAMD-21 (Hamilton Rating Scale
for Depression; see Hamilton, M. "A rating scale for depression."
J. Neurol. Neurosurg. Psychiat. 1960, 23, 56-62) scale at baseline.
Patients had an episode of depression which had lasted at least 2
weeks before baseline.
[0089] In this study, patients were randomly allocated to one of
three treatment groups. Group I patients (50 patients) received 2
capsules with 75 mg of ORG 34517 and one placebo (total daily dose
150 mg) for the first 2 weeks and 2 capsules with 75 mg ORG 34517
and 1 capsule with 150 mg (total daily dose 300 mg) the next 2
weeks; Group II patients (46 patients) received 3 capsules with 150
mg ORG 34517 (total daily dose 450 mg) in the first 2 weeks and 4
capsules of ORG 34517 (total daily dose 600 mg) in the next 2
weeks; Group III patients (44 patients) received 2 capsules with 10
mg paroxetine and one placebo capsule (total daily dose 20 mg) for
the first 2 weeks, followed by 2 capsules of 10 mg and one capsule
of 20 mg paroxetine (total daily dose 40 mg) in the next 2 weeks.
Medication was administered orally in the morning. Efficacy
assessment was done on days 4, 7, 10, 14, 21, 28 and 35 by using
the 21-item HAMD scale.
[0090] Thus, GCR antagonist therapy could prove a useful mechanism
for treatment of selected individuals who fail to respond to
current anti-depressant therapies such as SSRIs, providing a way to
enhance responsiveness or as an alternate means of achieving a
maintained euthymia.
[0091] Psychotic Depression.
[0092] Psychotic major depression has long been recognized as a
distinct psychiatric illness, having both psychotic and depressive
components in a differential diagnosis. Psychotic major depression
is very common. It has been estimated that twenty five percent of
depressed patients admitted to the hospital have psychotic major
depression (Coryell (1984) J. Nerv. Ment. Dis. 172:521). Like major
depression, psychotic depression is often also a result of high
circulating cortisol levels. Various evidence supports this
concept. Psychosis has been associated with Cushing's syndrome
(Gerson (1985) Can. J. Psychiatry 30:223-224; Saad (1984) Am. J.
Med. 76:759-766). A GR antagonist has been used to treat acute
psychiatric disturbances secondary to Cushing's syndrome. One study
showed that a relatively high dose of such a GR antagonist (400 to
800 mg per day) was useful in rapidly reversing acute psychosis in
patients with severe Cushing's' Syndrome due to adrenal cancers and
ectopic secretion of ACTH from lung cancer (Van der Lely (1991)
Ann. Intern. Med. 114:143; Van der Lely (1993) Pharmacy World &
Science 15:89-90; Sartor (1996) supra). Relatively high dose
mifepristone, in the range of 8-12 mg/kg/day, over a relatively
short period of time (4 days), was also shown to be effective in
the treatment of psychosis associated with psychotic major
depression (International Patent Application WO 99/17779;
Schatzberg and Belanoff).
[0093] Surgery-Associated Immune Suppression in the Elderly.
[0094] In healthy, young to middle aged subjects suffering from
stress, there is a physiological balance between pro-inflammatory
and anti-inflammatory mediators. In the elderly, the immune
response is blunted as a result of the decline in several
components of the immune system (immune senescence) and a shifting
to a chronic pro-inflammatory status (the so-called "inflammaging"
effect (Butcher and Lord, (2004) Aging Cell, pp. 151-160). As
production of cortisol remains reasonably constant with age,
whereas summed levels of DHEA and DHEAS decrease gradually from the
third decade, reaching 10-20% of their maximum by the eighth
decade, Butcher and Lord (2004, supra) propose a model for age and
stress, in which the age-related increase in the ratio of cortisol
to DHEAS, combined with an elevated cortisol release during stress,
leads to a significant reduction of immunity in aging subjects.
This is proposed to explain that aging subjects are far more prone
to infections under conditions of stress. (Butcher and Lord (2004,
supra); Butcher et al. (2005, Aging Cell 5, pp. 319-324).
[0095] The present invention relates to the use of a GCR antagonist
for the prevention or treatment of infections or infectious
conditions, in an aging patient, such as a human subject. The
beneficial effects of said GCR antagonists may be explained on the
basis of their correcting influence on the cortisol/DHEA(S) ratio.
It is believed that the effect in selected subjects, found to have
high circulating cortisol by a saliva test as provided for by this
invention, can be explained by the unbalanced immunosuppressive
role of the increased cortisol/DHEAS ratio in the aged group in
comparison to the balanced influence of cortisol and DHEAS on the
immune system in normal subjects.
[0096] The meaning of the term `aging subject` or `aged subject`
will be well understood in the context of the use according to this
invention. Although it is not linked to an exact lower age limit
this general notion refers in the human situation usually to a
person of at least 55 years old, but it is more clear with a lowest
age limit set at 60, 65, 70 or 75 years.
[0097] In the context of the invention, the infection or infectious
condition can be caused by any of several agents, e.g., by
bacteria, by viruses or by fungi. Also in the context of the
present invention, the expression "infectious conditions" means
silent or subclinical infections as well as conditions not
resulting in a manifest infectious disease, but in which at least
one parameter associated with an infectious disease, such as the
white blood (e.g., neutrophil, basophil or eosinophil) cell counts
or the level of some antibodies or some cytokines is higher than
normal. Normal values are known to the expert and may be found in
standard medical manuals.
[0098] Particular uses according to the invention relate to aging
subjects suffering from an infection or an infectious condition
concomitant to stress resulting from a trauma. The invention
particularly relates to uses wherein the subject suffers from the
consequences of a bone fracture and/or bone surgery, either for
such injury or for joint replacement for osteoarthritis or
rheumatoid arthritis. The invention also relates to uses wherein
the subject suffers from an infection or an infectious condition
concomitant to psychological stress, particularly acute emotional
stress.
[0099] Post Traumatic Stress Disorder (PTSD).
[0100] PTSD is a severe anxiety disorder that can develop after
exposure to any event that results in psychological trauma. This
event may involve the threat of death to oneself or to someone
else, or to one's own or someone else's physical, sexual, or
psychological integrity, overwhelming the individual's ability to
cope. As an effect of psychological trauma, PTSD is less frequent
and more enduring than the more commonly seen acute stress
response. Diagnostic symptoms for PTSD include re-experiencing the
original trauma(s) through flashbacks or nightmares, avoidance of
stimuli associated with the trauma, and increased arousal, such as
difficulty falling or staying asleep, anger, and hyper-vigilance.
Formal diagnostic criteria (both DSM-IV-TR and ICD-9) require that
the symptoms last more than one month and cause significant
impairment in social, occupational, or other important areas of
functioning. (Diagnostic and statistical manual of mental
disorders: DSM-IV. American Psychiatric Association. 1994.
Washington, D.C.: American Psychiatric Association.)
[0101] PTSD displays biochemical changes in the brain and body that
differ from other psychiatric disorders such as major depression.
Abundant evidence suggests derangement of HPA-axis physiology in
individuals diagnosed with PTSD, though the nature of the
derangements is variable: some have low cortisol, some have normal
levels, others have high levels of cortisol and for some, levels
may be normal, but circadian rhythm is lost. It is postulated that
these reflect different baseline mechanisms, but that when cortisol
is high, either in a sustained way through the day or by loss of
circadian rhythm with elevated night time levels, it is likely to
be an important component of the clinical symptomatology (Lindley S
E, et al. Basal and dexamethasone suppressed salivary cortisol
concentrations in a community sample of patients with posttraumatic
stress disorder. Biol. Psychiatry 2004; 55: 940-5). In such
patients, determined by salivary cortisol testing, administration
of a GCR antagonist is expected to be therapeutic or beneficial for
the symptoms of PTSD.
[0102] Prevention of Weight Gain in Patients Using Anti-Psychotic
and Anti-Depressant Medications.
[0103] Anti-psychotic and some anti-depressant medications (e.g.,
SSRIs) are amongst the most important tools for treating
psychiatric conditions of all kinds. However, management of
patients on who take many of these medications for chronic, long
term disease is made difficult by their significant side effect
profiles. One of the most important of these is weight gain and the
attendant metabolic syndrome that follows. For example, it is
estimated that 40-80% of patients who are under chronic
anti-psychotic administration experience substantial weight gain,
often exceeding 20% or more over their ideal body weights (Umbricht
et al. J Clin. Psychiatry 1994; 55: 157-160; Khan A Y, et al. J
Psychiatr Pract. 2010; 16: 289-96; Pramyothin P, Khaodhiar L. Curr
Opin Endocrinol Diabetes Obes. 2010; 17: 460-6; Rummel-Kluge C et
al. Schizophr Res. 2010; 123: 225-33). Such weight gain is one of
the most common causes of poor compliance with anti-psychotic and
anti-depressant regimens and, therefore, of long term failure of
therapy. Furthermore, anti-psychotic medications specifically are
commonly associated with development of insulin resistance and
metabolic syndrome (with development of type 2 diabetes mellitus
and hyper/dyslipidemia states) and the potentially and
significantly increased risks for cardiovascular disease; these
conditions are of tremendous medical consequence for patients who
are thereby caught in a "can't live with them, can't live without
them" treatment scenario. While weight gain is potentially seen
with all anti-psychotic medications, they are particularly common
and tend to more severe with the newer or "atypical" AP drugs
(Allison et al. Am J Psychiatry 1999; 156:1686-1696; Rummel-Kluge C
et al. Schizophr Res. 2010; 123: 225-33).
[0104] Elevations in cortisol are associated with changes in body
fat and insulin resistance. Several years ago, in a proof of
principle clinical experiment, it was reported that one GCR
anatogonist (mifepristone) was a highly effective treatment for
multiple medical complications in a patient with Cushing's disease
whose illness had not responded to surgery and radiation, including
reversal of insulin dependent diabetes: the patient was able to
stop insulin within a month (Chu et al., J. Clin. Endocrinol.
Metab. 2001; 86, 3568-3573.). These data suggest that a GCR
antagonist could be useful for blocking and reversing the insulin
resistance and weight changes seen in some patients treated with
atypical antipsychotic agents. To this end, this compound was
tested in rats who had olanzapine-induced weight gain and increases
in abdominal fat; reversal of weight gain was seen and reduction of
abdominal fat was obtained (Beebe et al. Behav. Brain Res. 2006;
171, 225-229). A clinical trial with this compound then confirmed
this benefit in humans with a 2 week study of 600 mg/day of
mifepristone that reduced olanzapine-induced weight gain in 57
non-overweight healthy males with Body Mass Indices less than 25
(Gross et al., Adv Ther. 2009; 26: 959-69.). Thus, GCR antagonist
therapy could prove a useful mechanism to target in treating
psychotic patients with atypical antipsychotic agents.
[0105] Cushing's Syndrome.
[0106] Cushing's Syndrome is a set of conditions in which high
levels of circulating cortisol or other GCR agonists cause a set of
seriously debilitating and sometimes life threatening signs and
symptoms including, but not limited to, psychiatric disturbances
(e.g. anxiety, depression, psychosis), immunosuppression, insulin
resistance and metabolic syndrome, skin conditions, hypertension
and osteoporosis. Endogenous cortisol may be produced by
ACTH-secreting, benign or malignant tumors of the pituitary gland
("Cushing's Disease") or of the adrenal cortex. These are rare
conditions and therefore Cushing's Syndrome is considered an
"orphan disease."
[0107] A proof of concept trial using RU486 to treat patients with
tumor-related Cushing's Syndrome demonstrated efficacy in remitting
symptoms such as glucose metabolic abnormalities (i.e., glucose
intolerance; (group 1) and hypertension (group 2). Statistically
significant improvement was achieved for both groups: with 60%
responding in the glucose intolerant group and 43% in the
hypertensive group (Corcept Therapeutics Press Release Dec. 22,
2010). Thus, GCR antagonist therapy can be expected to provide
clinical benefits for patients with Cushing's Syndrome administered
prior to tumor surgery to improve surgical outcomes and/or
post-surgery to mitigate symptoms in patients for whom surgical
cure is not achievable.
[0108] In addition, GCR antagonist therapy can be expected to
provide clinical benefits for patients, for example, in hospitals,
nursing homes, nurseries, daycares, schools, work environments,
public transportation, healthcare settings, psychiatric
institutions, and long-term nursing facilities
Diagnostic Systems and Kits
[0109] A diagnostic kit may comprise some or all of the following
components: 1) one or more standards comprised of one or more of
the biomarker(s) of the invention, such as cortisol; 2) a ligand,
such as an antibody or a plurality of antibodies, that are specific
for the biomarker(s) that are to be assayed for using the kit; 3)
written instructions; 4) diluents for samples and the standards; 5)
a wash buffer; 6) color reagents; 7) stop solution; and 8) a ligand
carrier, such as an antibody carrier, for example, a lateral flow
device, or a microplate with bound antibody, or polystyrene
beads.
[0110] An example of such a kit is a quantitative ELISA
(enzyme-linked immunosorbent assay) that determines the
concentration or concentrations of the biomarker or biomarker(s) in
accordance with methods embodied by the invention. The principle of
the assay is to use the quantitative sandwich enzyme immunoassay
technique wherein a monoclonal or polyclonal antibody selective for
a biomarker is pre-coated onto a carrier such as a microplate into
its wells. The standards and sample are then pipetted into the
wells and any of the biomarker that is present is bound to this
immobilized antibody. Next, the wells are washed with washing
buffer, and an enzyme-linked monoclonal or polyclonal antibody that
is specific for the biomarker is added to the wells. Washing is
again performed, then a substrate solution is added to the wells.
Color subsequently develops in proportion to the amount of
polypeptide of the invention that is bound in the first step. The
color development is stopped using a stop solution, and the
intensity of the color is measured by a microplate reader.
[0111] The methods of the invention may be carried out using, for
example, a lateral flow assay. Such lateral flow assays have the
potential to be a cost-effective, fast, simple, and sensitive
method, for instance for on-site screening assays. The lateral flow
assay comprises a carrier that allows a lateral flow to occur
wherein either the sample or the detection reagent is displaced
form one location on the carrier to another. There are many formats
of lateral flow assays suitable for use in a method embodied by the
invention, and the skilled person will readily know how to select
and optimize a particular format. An example of a lateral flow test
strip of the invention comprises, for example, the following
components:
[0112] 1. Sample pad--an absorbent pad onto which the test sample
is applied.
[0113] 2. Conjugate or reagent pad--this contains antibodies
specific to the target analyte conjugated to colored particles
(usually colloidal gold particles, or latex microspheres).
[0114] 3. Reaction membrane--typically a hydrophobic nitrocellulose
or cellulose acetate membrane onto which anti-target analyte
antibodies are immobilized in a line across the membrane as a
capture zone or test line (a control zone may also be present,
containing antibodies specific for the conjugate antibodies).
[0115] 4. Wick or waste reservoir--a further absorbent pad designed
to draw the sample across the reaction membrane by capillary action
and collect it.
[0116] Double Antibody Sandwich Assays--
[0117] In this format the sample migrates from the sample pad
through the conjugate pad where any target analyte present will
bind to the conjugate. The sample then continues to migrate across
the membrane until it reaches the capture zone where the
target/conjugate complex will bind to the immobilized antibodies
producing a visible line on the membrane. The sample then migrates
further along the strip until it reaches the control zone, where
excess conjugate will bind and produce a second visible line on the
membrane. This control line indicates that the sample has migrated
across the membrane as intended. Two clear lines on the membrane is
a positive result. A single line in the control zone is a negative
result. Double antibody sandwich assays are most suitable for
larger analytes, such as bacterial pathogens and viruses, with
multiple antigenic sites. Competitive assays are primarily used for
testing small molecules and differ from the double antibody
sandwich format in that the conjugate pad contains antibodies that
are already bound to the target analyte, or to an analogue of it.
If the target analyte is present in the sample it will therefore
not bind with the conjugate and will remain unlabelled. As the
sample migrates along the membrane and reaches the capture zone, an
excess of unlabelled analyte will bind to the immobilized
antibodies and block the capture of the conjugate, so that no
visible line is produced. The unbound conjugate will then bind to
the antibodies in the control zone producing a visible control
line. A single control line on the membrane is a positive result.
Two visible lines in the capture and control zones is a negative
result. However, if an excess of unlabelled target analyte is not
present, a weak line may be produced in the capture zone,
indicating an inconclusive result. Competitive assays are most
suitable for testing for small molecules, such as mycotoxins,
unable to bind to more than one antibody simultaneously. There are
a number of variations on lateral flow technology. The capture zone
on the membrane may contain immobilized antigens or
enzymes--depending on the target analyte--rather than antibodies.
It is also possible to apply multiple capture zones to create a
multiplex test. For example, commercial test strips able to detect
both EHEC Shiga toxins ST1 and ST2 separately in the same sample
have been developed. Lateral flow immunoassays are simple to use by
untrained operators and generally produce a result within 15
minutes. They are very stable and robust, have a long shelf life
and do not usually require refrigeration. They are also relatively
inexpensive to produce. These features make them ideal for use at
the point-of-care and for testing samples in the field, as well as
in the laboratory. However, their sensitivity is limited without
additional concentration or culture procedures.
[0118] Quantitative Tests--
[0119] While most lateral flow immunoassays are only capable of
providing a qualitative result, it is possible to obtain some
degree of quantification by measuring the amount of conjugate bound
to the capture zone. This can be done using a dedicated reader to
measure the intensity of the colored test line. For example, the
Neogen Corporation has developed the Accuscan.TM. lateral flow
reader for use with its range of Reveal.RTM. assay kits and Charm
Sciences also supplies a reader for its Rosa.RTM. range of
mycotoxin test strips. More sophisticated techniques, such as
fluorescent dye labeled conjugates, have also been developed to
improve the quantitative potential of lateral flow assays.
Applications in the 20 years since the first lateral flow test was
launched have expanded to include a huge range of different tests
that have been developed based on the same technology. The first
commercially available kits were aimed at the clinical diagnostics
field, but there are now products with applications in almost every
branch of microbiology. Clinical microbiology--lateral flow tests
have been developed for bacterial pathogens, respiratory and
enteric viruses, intestinal parasites and bacterial toxins. Many of
the lateral flow immunoassay products designed for the clinical
sector were intended to be used at the point-of-care for direct
testing of fecal, blood and urine samples and nose and throat
swabs, where the simple operation and speed of the tests is key to
their use outside of the laboratory. However, the same test strips
may also be useful as a quick confirmatory test following
laboratory culture of clinical samples. Food and agricultural
microbiology--test strips are available for food borne bacterial
pathogens, bacterial and fungal toxins. In the food microbiology
sector, the main applications are more likely to be in the
laboratory, although there are field test kits for mycotoxins in
grain samples. Testing for food borne bacterial pathogens generally
involves at least one enrichment stage before the assay strip is
used to confirm the presence or absence of the pathogen. Some
manufacturers, such as Dupont.RTM., have developed enrichment media
and methods specifically designed for use with lateral flow test
strips. Test strips may also be useful for rapid confirmation of
the identity of bacterial isolates from conventional
microbiological testing.
[0120] A diagnostic system in kit form of the present invention
includes, for example, in an amount sufficient for at least one
assay, a polypeptide, antibody composition or monoclonal antibody
composition of the present invention, as a packaged reagent.
Instructions for use of the packaged reagent are also typically
included.
[0121] A diagnostic system in kit form of the present invention may
include, for example, a means for detecting the presence of a
biological substance in a test sample, comprising for example, a
lollipop-like apparatus including a stem integrated with the base
and a head integrated with the stem, for collecting a test sample
consisting of, for example, saliva, or a bodily fluid sample from a
subject. The stem head may include a receptor of a sponge like
carrier to ensure a high void volume to absorb sufficient saliva,
or bodily fluid sample. See U.S. Pat. No. 7,993,283, incorporated
by reference herein in its entirety.
[0122] A diagnostic system in kit form of the present invention may
include, for example, a means for combining the test sample with a
buffering system (Reagent 1) containing viscosity controllers and
stabilizers into a reaction vessel and mixing the solution. A
diagnostic system in kit form of the present invention may include,
for example, a means for reading the a parameter of the reaction
vessel with sample and buffer, and further means for combining the
test sample and buffer mixture with a fluorescence-labeled ligand
(Reagent 2) to said biological substance in the reaction vessel,
mixing the solution to produce an assay solution. Furthermore,
Reagent 2 may be delivered to the reaction vessel without further
dilution volume of the assay solution.
[0123] As used herein, the term "package" refers to a solid matrix
or material such as glass, plastic, paper, foil and the like
capable of holding within fixed limits a polypeptide, antibody
composition or monoclonal antibody composition of the present
invention. Thus, for example, a package can be a glass vial used to
contain milligram quantities of a contemplated polypeptide or it
can be a microliter plate well to which microgram quantities of a
contemplated polypeptide have been operatively affixed, i.e.,
linked so as to be capable of being immunologically bound by an
antibody.
[0124] "Instructions for use" typically include a tangible
expression describing the reagent concentration or at least one
assay method parameter such as the relative amounts of reagent and
sample to be admixed, maintenance time periods for reagent/sample
admixtures, temperature, buffer conditions and the like.
[0125] In preferred embodiments, a diagnostic system of the present
invention further includes a label or indicating means capable of
signaling the formation of a complex containing a polypeptide or
antibody molecule of the present invention.
[0126] The word "complex" as used herein refers to the product of a
specific binding reaction such as an antibody-antigen or
receptor-ligand reaction. Exemplary complexes are immunoreaction
products.
[0127] As used herein, the terms "label" and "indicating means" in
their various grammatical forms refer to single atoms and molecules
that are either directly or indirectly involved in the production
of a detectable signal to indicate the presence of a complex. Any
label or indicating means can be linked to or incorporated in an
expressed protein, polypeptide, or antibody molecule that is part
of an antibody or monoclonal antibody composition of the present
invention, or used separately, and those atoms or molecules can be
used alone or in conjunction with additional reagents such labels
are themselves well-known in clinical diagnostic chemistry and
constitute a part of this invention only insofar as they are
utilized with otherwise novel proteins methods and/or systems.
[0128] The labeling means can be a fluorescent labeling agent that
chemically binds to antibodies or antigens without denaturing them
to form a fluorochrome (dye) that is a useful immunofluorescent
tracer. Suitable fluorescent labeling agents are fluorochromes such
as fluorescein isocyanate (FIC), fluorescein isothiocyante (FITC),
5-dimethylamine-1-naphthalenesulfonyl chloride (DANSC),
tetramethylrhodamine isothiocyanate (TRITC), lissamine, rhodamine
8200 sulphonyl chloride (RB 200 SC) and the like. A description of
immunofluorescence analysis techniques is found in DeLuca,
"Immunofluorescence Analysis", in Antibody As a Tool, Marchalonis,
et al., eds., John Wiley & Sons, Ltd., pp. 189-231 (1982),
which is incorporated herein by reference.
[0129] In preferred embodiments, the indicating group is an enzyme,
such as horseradish peroxidase (HRP), glucose oxidase, or the like.
In such cases where the principal indicating group is an enzyme
such as HRP or glucose oxidase, additional reagents are required to
visualize the fact that a receptor-ligand complex (immunoreactant)
has formed. Such additional reagents for HRP include hydrogen
peroxide and an oxidation dye precursor such as diaminobenzidine.
An additional reagent useful with glucose oxidase is
2,2'-azino-di-(3-ethyl-benzthiazoline-G-sulfonic acid) (ABTS).
[0130] Radioactive elements are also useful labeling agents and are
used illustratively herein. An exemplary radiolabeling agent is a
radioactive element that produces gamma ray emissions. Elements
which themselves emit gamma rays, such as .sup.124I, .sup.125I,
.sup.128I, .sup.132I and .sup.51Cr represent one class of gamma ray
emission-producing radioactive element indicating groups.
Particularly preferred is .sup.125I. Another group of useful
labeling means are those elements such as .sup.11C, .sup.18F,
.sup.15O and .sup.13N which themselves emit positrons. The
positrons so emitted produce gamma rays upon encounters with
electrons present in the animal's body. Also useful is a beta
emitter, such .sup.111 indium or .sup.3H.
[0131] The linking of labels, i.e., labeling of, polypeptides and
proteins is well known in the art. For instance, antibody molecules
produced by a hybridoma can be labeled by metabolic incorporation
of radioisotope-containing amino acids provided as a component in
the culture medium. See, for example, Galfre et al., Meth.
Enzymol., 73:3-46 (1981). The techniques of protein conjugation or
coupling through activated functional groups are particularly
applicable. See, for example, Aurameas, et al., Scand. J. Immunol.,
Vol. 8 Suppl. 7:7-23 (1978), Rodwell et al., Biotech., 3:889-894
(1984), and U.S. Pat. No. 4,493,795, which are all incorporated
herein by reference.
[0132] The diagnostic systems can also include, preferably as a
separate package, a specific binding agent. A "specific binding
agent" is a molecular entity capable of selectively binding a
reagent species of the present invention or a complex containing
such a species, but is not itself a polypeptide or antibody
molecule composition of the present invention. Exemplary specific
binding agents are second antibody molecules, complement proteins
or fragments thereof, S. aureus protein A, and the like. Preferably
the specific binding agent binds the reagent species when that
species is present as part of a complex.
[0133] In preferred embodiments, the specific binding agent is
labeled. However, when the diagnostic system includes a specific
binding agent that is not labeled, the agent is typically used as
an amplifying means or reagent. In these embodiments, the labeled
specific binding agent is capable of specifically binding the
amplifying means when the amplifying means is bound to a reagent
species-containing complex.
[0134] The diagnostic kits of the present invention can be used in
an "ELISA" format to detect, for example, the presence or quantity
of cortisol in a body fluid sample such as serum, plasma, or urine,
etc. "ELISA" refers to an enzyme-linked immunosorbent assay that
employs an antibody or antigen bound to a solid phase and an
enzyme-antigen or enzyme-antibody conjugate to detect and quantify
the amount of an antigen or antibody present in a sample. A
description of the ELISA technique is found in Chapter 22 of the
4th Edition of Basic and Clinical Immunology by D. P. Sites et al.,
published by Lange Medical Publications of Los Altos, Calif. in
1982 and in U.S. Pat. No. 3,654,090; U.S. Pat. No. 3,850,752; and
U.S. Pat. No. 4,016,043, which are all incorporated herein by
reference.
[0135] Thus, for example, a polypeptide, antibody molecule
composition or monoclonal antibody molecule composition of the
present invention can be affixed to a solid matrix to form a solid
support that comprises a package in the subject diagnostic systems.
The reagent is typically affixed to the solid matrix by adsorption
from an aqueous medium although other modes of affixation, well
known to those skilled in the art, can be used.
[0136] Useful solid matrices are also well known in the art. Such
materials are water insoluble and include cross-linked dextran;
agarose; beads of polystyrene beads about 1 micron to about 5
millimeters in diameter; polyvinyl chloride, polystyrene,
cross-linked polyacrylamide, nitrocellulose- or nylon-based webs
such as sheets, strips or paddles; or tubes, plates or the wells of
a microtiter plate such as those made from polystyrene or
polyvinylchloride.
[0137] The reagent species, labeled specific binding agent or
amplifying reagent of any diagnostic system described herein can be
provided in solution, as a liquid dispersion or as a substantially
dry power, e.g., in lyophilized form. Where the indicating means is
an enzyme, the enzyme's substrate can also be provided in a
separate package of a system. A solid support such as the
before-described microtiter plate and one or more buffers can also
be included as separately packaged elements in this diagnostic
assay system.
[0138] The packaging materials discussed herein in relation to
diagnostic systems are those customarily utilized in diagnostic
systems. Such materials include glass and plastic (e.g.,
polyethylene, polypropylene and polycarbonate) bottles, vials,
plastic and plastic-foil laminated envelopes and the like. In one
embodiment a diagnostic system of the present invention is useful
for assaying for the presence of, for example, cortisol. Such a
system comprises, in kit form, a package containing an antibody to,
for example, cortisol.
[0139] "Sample" refers to, for example, essentially any source from
which materials of interest to be analyzed (e.g., ligands and
antiligands, such as antibodies and antigens, and nucleic acids and
their complements) can be obtained. A sample may be acquired from
essentially any organism, including animals and plants, as well as
cell cultures, recombinant cells and cell components. Samples can
be from a biological tissue, fluid or specimen and may be obtained
from a diseased or healthy organism. Samples may include, but are
not limited to, saliva, sputum, amniotic fluid, blood, blood cells
(e.g., white cells), urine, semen, peritoneal fluid, pleural fluid,
tissue or fine needle biopsy samples, and tissue homogenates.
Samples may also include sections of tissues such as frozen
sections taken for histological purposes. Typically, samples are
taken from a human. However, samples can be obtained from other
mammals'also, including by way of example and not limitation, dogs,
cats, sheep, cattle, and pigs. The sample may be pretreated as
necessary by dilution in an appropriate buffer solution or
concentrated, if desired. Any of a number of standard aqueous
buffer solutions, employing one of a variety of buffers, such as
phosphate, Tris, or the like, preferably at physiological pH can be
used.
[0140] Biological samples can be derived from patients using well
known techniques such as venipuncture, lumbar puncture, fluid
sample such as saliva or urine, or tissue biopsy and the like. When
the biological material is derived from non-humans, such as
commercially relevant livestock, blood and tissue samples are
conveniently obtained from livestock processing plants.
Alternatively, a biological sample may be obtained from a cell or
blood bank where tissue and/or blood are stored, or from an in
vitro source, such as a culture of cells. Techniques for
establishing a culture of cells for use as a source for biological
materials are well known to those of skill in the art.
[0141] In one embodiment the sample is selected from or is derived
from, for example, microbial products or biological products.
[0142] Although the above described example relates to the antigens
relating to disease, the immunoassay apparatus could be used, for
example, as an allergy test kit, as a test kit for drugs of abuse
or for analyzing non-human derived samples e.g. bovine, porcine,
and veterinary tests.
[0143] Specific reagents used in the assay device will be selected
so as to ensure that the particular target analyte is detected as
is well known in the art. The target analyte may be any analyte for
example a chemical reagent which may be organic or inorganic, and
which optionally comprises a hapten, a protein, a polypeptide, a
microorganism or a nucleic acid sequence.
[0144] In particular, the analyte is a hormone such as a fertility
hormone like progesterone or a stress hormone such as cortisol.
However, there is a wide range of applications of these types of
tests across the entire field of diagnostics and analysis.
Detection of marker proteins or hormones can be diagnostic of
certain disease conditions in humans or animals, and the presence
of drugs or drug residues may also be required to be detected, for
example, in animal husbandry, forensic medicine or in the testing
for banned or prohibited drug substances.
[0145] Alternatively, the analyte is a chemical reagent, for
instance a small molecule, which suitably comprises a hapten. Small
molecules will generally comprise a single recognizable binding
site. Typically they will have a molecular weight of less than 1
kDa.
[0146] Where the assay utilizes a labelled binding partner for the
analyte and the analyte is a chemical reagent, the binding partner
may comprise any other reagent which reacts with or otherwise
becomes associated with the chemical reagent, either because it
forms covalent or ionic bonds with the reagent, or by the formation
of other interactions, such as hydrogen bonding or Van der Waals
interactions. For example, where the chemical reagent is an acid,
the binding partner may comprise an alcohol or an amine that forms
an ester or amide with the acid under the sorts of conditions found
in the test. Alternatively the binding partner may comprise a base
that forms a salt with the acid. Conversely, where the binding
partner may comprise the acid part of the reactive pair.
[0147] Where the analyte is or comprises a hapten or a protein
antigen, the binding partner may comprise an antibody or a binding
fragment thereof, which may be monoclonal, polyclonal or
recombinant, but preferably is monoclonal. Where the analyte is a
hormone or enzyme, the labelled binding partner may comprise a
labelled receptor for the analyte. However, where the analyte is
itself an immunoglobulin, and in particular, an antibody, the
labelled binding partner may also comprise for instance, an antigen
or recombinant antigen, as well as anti-antibody immunoglobulin
such as anti-sera.
[0148] Antibodies or binding fragments to small molecules such as
haptens, are generated by attaching the molecule to an immunogenic
reagent and administering this to an animal such as a mouse or
rabbit. Antibodies are then harvested from the animal in the usual
way. Monoclonal antibodies are obtained by fusing spleen cells to
hybridoma cells, and selecting those which bind the hapten, using
routine procedures.
[0149] For example, where the analyte is a biologically active
material such as an active agrochemical as discussed above,
specific reagents used in the assay device will be selected so as
to ensure that the particular target biologically active material
is detected as is well known in the art. The biologically active
material may be any active chemical such as an agrochemical, for
example a chemical reagent which may be organic or inorganic, and
which optionally comprises a hapten, a protein, a polypeptide, a
microorganism or a nucleic acid sequence. Most preferably the
biologically active material is a chemical reagent, for instance a
small molecule, which suitably comprises a hapten. Small molecules
will generally have a single antibody binding site. Typically they
will have a molecular weight of less that 1 kDa.
[0150] Antibodies or binding fragments to small molecules such as
haptens, are generated by attaching the molecule to an immunogenic
reagent and administering this to an animal such as a mouse or
rabbit. Antibodies are then harvested from the animal in the usual
way. Monoclonal antibodies are obtained by fusing spleen cells to
hybridoma cells, and selecting those which bind the hapten, using
routine procedures.
Microarrays
[0151] The method of the invention is particularly useful in
combination with the analysis of gene expression profiles. In some
embodiments, a gene expression profile, such as a collection of
transcription rates of a number of genes, is converted to a
projected gene expression profile. The projected gene expression
profile is a collection of expression values. The conversion is
achieved, in some embodiments, by averaging the transcription rate
of the genes. In some other embodiments, other linear projection
processes may be used.
[0152] Microarrays may be prepared and analyzed using methods known
in the art. Oligonucleotides may be used as either probes or
targets in a microarray. The microarray can be used to monitor the
expression level of large numbers of genes simultaneously and to
identify genetic variants, mutations, and single nucleotide
polymorphisms. Such information may be used to determine gene
function; to understand the genetic basis of a condition, disease,
or disorder; to diagnose a condition, disease, or disorder; and to
develop and monitor the activities of therapeutic agents. (See,
e.g., Brennan et al. (1995) U.S. Pat. No. 5,474,796; Schena et al.
(1996) Proc. Natd. Acad. Sci. 93:10614-10619; Baldeschweiler et al.
(1995) PCT application WO95/251116; Shalon et al. (1995) PCT
application WO95/35505; Heller et al. (1997) Proc. Natl. Acad. Sci.
94:2150-2155; and Heller et al. (1997); U. S. Pat. No. 5,605,662.)
Hybridization probes are also useful in mapping the naturally
occurring genomic sequence. The sequences may be mapped to a
particular chromosome, to a specific region of a chromosome, or to
artificial chromosome constructions, e.g., human artificial
chromosomes (HACs), yeast artificial chromosomes (YACs), bacterial
artificial chromosomes (BACs), bacterial P1 constructions, or
single chromosome DNA libraries.
[0153] Using the methods of the invention a skilled artisan can
readily select and prepare probes for a microarray wherein the
microarray contains specific individual probes for less than all
the genes in the genome and less than all the genes in the genome.
In such embodiments, the microarray contains one or two or more
individual probes, each of which hybridizes to an expression
product (e.g., mRNA, or cDNA or cRNA derived therefrom) for a
desired number of genes. Thus, for example, changes in the
expression of all or most of the genes in the entire genome of a
cell or organism can thereby be monitored by use of a surrogate and
on a single microarray by measuring expression of the group of
genes that are representative of all or most of the genes of the
genome. Such microarrays can be prepared using the selected probes
and are therefore part of the present invention.
Glucocorticoid Receptor
[0154] The glucocorticoid receptor is widely distributed and
expressed in many cultured cell lines, and the control of gene
expression by glucocorticoids, therefore, has been widely studied
as a model for transcriptional regulation. A number of
glucocorticoid-responsive transcription units, including mouse
mammary tumor virus (MMTV) (Ringold, et al., 1975; Parks, et al.,
1974), mouse and human metallothionein (Eager, et al., 1981; Karin,
et al., 1980), rat alpha.sub.2M-globulin (Kurtz, et al., 1977) and
rat and human growth hormone (Spindler, et al., 1982; Evans, et
al., 1982; Robins, et al., 1982) genes have been identified. DNA
sequences mediating transcriptional stimulation of several of these
genes have been localized. For MMTV, these sequences are discrete
genomic regions upstream of the transcriptional start site which
appear to exert their actions independently of orientation and
position (Chandler, et al., 1983; Ostrowski, et al., 1984). The
steroid/receptor complex appears to bind to these regulatory
sequences and purified receptor has been used to define the
specific binding sites (Govinda, et al., 1982; Scheidereit, et al.,
1983; Pfahl, 1982; Payvar, et al., 1983). The ability of the
glucocorticoid-responsive element (GRE) to alter its position and
orientation yet still maintain promoter inducibility suggests that
it resembles the class of cis-acting regulatory sequences termed
enhancers (Chandler, et al., 1983). First discovered in viruses and
subsequently in cellular genes, these sequences are necessary for
efficient transcription in vivo (Laimonis, et al., 1982; Benoist,
et al., 1981; Baerji, et al., 1983). It has been suggested that
enhancers are recognized by trans-acting factors that mediate
regulatory effects by tissue-specific transcriptional control.
Although the enhancer factors have not been well characterized, the
glucocorticoid receptor may serve as a paradigm for these putative
gene activator proteins.
[0155] It is generally accepted that the unliganded glucocorticoid
receptor (GR) resides in the cytoplasm, and that hormone activation
leads both to nuclear accumulation and gene activation. (Gasc,
J.-M. & Baulieu, E. E. (1987) in Steroid Hormone Receptors:
Their Intracellular Localisation, ed. Clark, C. R. (Ellis Horwood
Ltd., Chichester, England), pp. 233-250; Beato, M. (1989) Cell 56,
335-344; Carson-Jurica, M. A., Schrader, W. T. & O'Malley, B.
W. (1990) Endocr. Rev. 11, 201-220; Gronemeyer, H. (1993) in
Steroid Hormone Action, ed. Parker, M. G. (Oxford University Press,
New York), pp. 94-117; Tsai, M. J. & O'Malley, B. W. (1994)
Annu. Rev. Biochem. 63, 451-486; Akner, G., Wikstrom, A. C. &
Gustafsson, J. A. (1995) J. Steroid Biochem. Mol. Biol. 52, 1-16),
and references therein. However, the mechanisms involved in nuclear
translocation and targeting of steroid receptors to regulatory
sites in chromatin have been poorly understood. It has previously
been difficult to discriminate between the ability of a given
receptor mutant, or a given receptor/ligand combination, to
participate in the separate processes of receptor activation,
nuclear translocation, sequence-specific binding, and promoter
activation.
[0156] The glucocorticoid receptor (GR) is expressed in a subset of
both ERalpha-positive and -negative human breast cancers as well as
in some ovarian cancers. In vitro and in vivo experiments suggest
that activation of the GR in ER-negative pre-malignant breast
epithelial and cancer cells initiates cell survival pathways under
stress conditions that normally induce significant cell death (e.g.
chemotherapy, radiation, growth factor deprivation).
[0157] The glucocorticoid receptor (GR) is present in
glucocorticoid responsive cells where it resides in the cytosol in
an inactive state until it is stimulated by an agonist. Upon
stimulation the glucocorticoid receptor translocates to the cell
nucleus where it specifically interacts with DNA and/or protein(s)
and regulates transcription in a glucocorticoid responsive manner.
Two examples of proteins that interact with the glucocorticoid
receptor are the transcription factors, API and NF.kappa.-B. Such
interactions result in inhibition of API- and NF.kappa.-B-mediated
transcription and are believed to be responsible for some of the
anti-inflammatory activity of endogenously administered
glucocorticoids. In addition, glucocorticoids may also exert
physiologic effects independent of nuclear transcription.
Biologically relevant glucocorticoid receptor agonists include
cortisol and corticosterone. Many synthetic glucocorticoid receptor
agonists exist including dexamethasone, prednisone and
prednisolone.
Glucocorticoid Receptor Antagonists
[0158] Glucocorticoid receptor antagonists bind to the receptor and
prevent glucocorticoid receptor agonists from binding and eliciting
GR mediated events, including transcription. RU486 is an example of
a non-selective glucocorticoid receptor antagonist.
[0159] Compounds having high glucocorticoid receptor binding
affinity and, in addition, high in vivo anti-glucocorticoid
activity, while having, for example, low androgenic and
progestagenic activities are disclosed in U.S. Pat. No. 6,011,025,
incorporated herein by reference in its entirety. ORG 34517 is an
example of a compound with high glucocorticoid receptor binding
affinity while having low androgenic and progestagenic
activities.
[0160] It has been found that 11-(substituted
phenyl)-estra-4,9-diene derivatives of formula I
##STR00011##
wherein A is a residue of a 5- or 6-membered ring containing 2
heteroatoms which are not connected to each other and independently
selected from O and S, the ring being optionally substituted with
one or more halogen atoms, or A is a residue of a 5- or 6-membered
ring wherein no double C--C bonds are present, containing 1
heteroatom selected from O and S, which heteroatom is connected to
the phenyl group at the position indicated with an asterisk, the
ring being optionally substituted with one or more halogen atoms;
R1 is H or 1-oxo(1-4C)alkyl; R2 is H, (1-8C)alkyl, halogen or CF3;
X is selected from (H,OH), O, and NOH; and the interrupted line
represents an optional bond, show specific and high glucocorticoid
receptor binding affinity and are highly active in vivo showing
predominant anti-glucocorticoid activity.
[0161] The compounds lack appreciable affinity for
mineralocorticoid, progesterone, estrogen and androgen receptors,
indicating a clean side effect profile.
[0162] The 11-(substituted phenyl)-estra-4,9-diene derivatives of
the invention can be used in the prevention and treatment of
glucocorticoid dependent diseases or symptoms, like Cushing
syndrome, diabetes, glaucoma, sleep disturbances, depression,
anxiety, atherosclerosis, hypertension, adiposity, osteoporosis and
withdrawal symptoms from narcotics and their mixtures.
[0163] Preferred compounds according to this invention are
11-(substituted phenyl)estra-4,9-diene derivatives, wherein the
heteroatom(s) are (is) O, the 5- or 6-membered ring being
optionally substituted with one or more fluorine atoms; R1 is H;
and X is O or NOH. More preferred compounds are 11-(substituted
phenyl)estra-4,9-diene derivatives wherein A is a residue of a
5-membered ring. Particularly preferred are 11-(substituted
phenyl)estra-4,9-diene derivatives wherein A contains 2 heteroatoms
being O.
[0164] Especially preferred are 11-(substituted
phenyl)estra-4,9-diene derivatives wherein R2 is methyl and the
interrupted line represents a bond.
[0165] The most preferred compound is
(11.beta.,17.beta.)-11-(1,3-benzodioxol-5-yl)-17-hydroxy-17-(1-propynyl)e-
stra-4,9-dien-3-one (ORG 34517).
##STR00012##
The term halogen means a fluorine, chlorine, bromine or iodine
atom. Fluorine is the preferred halogen in ring A and when R2 is
halogen, chlorine is preferred. The terms (1-4C)alkyl and
(1-8C)alkyl, as used in the definitions of R1 and R2, respectively,
mean alkyl groups having 1-4 and 1-8 carbon atoms, respectively,
for example methyl, ethyl, propyl, isopropyl, butyl, sec-butyl,
tert-butyl, pentyl, neopentyl, hexyl, octyl.
[0166] The 11-(substituted phenyl)-estra-4,9-diene derivatives
according to the present invention can be prepared by a process
wherein a compound of formula II
##STR00013##
wherein A, R2 and the interrupted line have the meanings as
previously defined, R1 is H, and P is a protected keto-group, is
dehydrated and deprotected, after which the 17.beta.-OH is
optionally esterified by reaction with an appropriate carboxylic
acid to give a derivative wherein R1 is 1-oxo(1-4C)alkyl, and
optionally the 3-oxo group is converted into the corresponding
3-hydroxy--or 3-oxime derivative. The 3-oxo group can be reduced to
form the 3-hydroxy-derivative by using a suitable reducing agent,
such as sodium borohydride. The 3-oxime derivatives can be prepared
by hydroxylamine treatment in a suitable solvent, like
pyridine.
[0167] The derivatives of formula II may be prepared according to
well known methods described and used for the preparation of
steroids.
[0168] A suitable process for the preparation of derivatives of
formula II starts from estra-4,9-diene-3,17-dione. Selective
reduction of the 17-keto group to 17.beta.-OH, 17.alpha.-H, e.g.
with sodium borohydride, followed by protection of the 3-keto
group, e.g., by ketalisation with ethyleneglycol,
triethylorthoformate and p-toluenesulfonic acid, and oxidation of
the 17-hydroxy group, e.g. with pyridinium chlorochromate, provides
the 3-ketoprotected estra-5(10),9(11)-diene-3,17-dione.
Alkynylation at the 17-position (yielding a
17.alpha.-alkynyl,17.beta.-OH derivative), followed by epoxidation
of the 5(10) double bond, e.g. with hydrogen peroxide,
trifluoroacetophenone, and pyridine in dichloromethane according to
the method as disclosed in European patent application EP 0 298
020, provides the 3-ketoprotected
5.alpha.,10.alpha.-epoxy-17.alpha.-alkynyl-17.beta.-hydroxy-estr-9(11)-en-
e-3-one.
[0169] Subsequently, compounds of formula II are formed from this
epoxide derivative, for example by reaction with an organometallic
compound of the formula
##STR00014##
wherein X is a (alkali)metal, like lithium, or a magnesiumhalide,
preferably magnesium bromide.
[0170] Suitable protective groups and methods to remove these
groups are known in the art, for example from T. W. Green:
Protective Groups in Organic Synthesis (Wiley, N Y, 1981).
Particularly suitable protective groups for the protection of keto
groups are acetals, e.g. 1,2-ethylene ketal.
[0171] The specificity of ORG 34517 for GR blockade, without
significant cross-binding to other related steroidal hormone
receptors (such as those for estrogen and progesterone), eliminates
the likelihood of significant toxicities and side effects. Indeed,
none were identified in all the substantial phase I and phase II
clinical trials that already have been performed with the compound.
Because the drug is envisioned as being used in limited dosing over
time, coordinated with the intermittent dosing strategies typical
for chemotherapeutic agents, the GR blockade also would not lead to
significant alteration of HPA-axis functioning, with rapid
restitution of the HPA-axis to baseline following dosing.
Formulations
[0172] The compounds of the invention may be administered enterally
or parenterally. Mixed with pharmaceutically suitable auxiliaries,
e.g., as described in the standard reference, Gennaro et al.,
Remington's Pharmaceutical Sciences. The compounds may be
compressed into solid dosage units, such as pills, tablets, or be
processed into capsules or suppositories. By means of
pharmaceutically suitable liquids the compounds can also be applied
in the form of a solution, suspension, emulsion, e.g. for use as an
injection preparation or eye drops, or as a spray, e.g. for use as
a nasal spray.
[0173] For making dosage units, e.g., tablets, the use of
conventional additives such as fillers, colorants, polymeric
binders and the like is contemplated. In general, any
pharmaceutically acceptable additive which does not interfere with
the function of the active compounds can be used. Suitable carriers
with which the compositions can be administered include lactose,
starch, cellulose derivatives and the like, or mixtures thereof,
used in suitable amounts.
Dosage Forms
[0174] The compositions of the present invention can be processed
by agglomeration, air suspension chilling, air suspension drying,
balling, coacervation, coating, comminution, compression,
cryopelletization, encapsulation, extrusion, wet granulation, dry
granulation, homogenization, inclusion complexation,
lyophilization, melting, microencapsulation, mixing, molding, pan
coating, solvent dehydration, sonication, spheronization, spray
chilling, spray congealing, spray drying, or other processes known
in the art. The compositions can be provided in the form of a
minicapsule, a capsule, a tablet, an implant, a troche, a lozenge
(minitablet), a temporary or permanent suspension, an ovule, a
suppository, a wafer, a chewable tablet, a quick or fast dissolving
tablet, an effervescent tablet, a buccal or sublingual solid, a
granule, a film, a sprinkle, a pellet, a bead, a pill, a powder, a
triturate, a platelet, a strip or a sachet. Compositions can also
be administered as a "dry syrup", where the finished dosage form is
placed directly on the tongue and swallowed or followed with a
drink or beverage. These forms are well known in the art and are
packaged appropriately. The compositions can be formulated for
oral, nasal, buccal, ocular, urethral, transmucosal, vaginal,
topical or rectal delivery.
[0175] The pharmaceutical composition can be coated with one or
more enteric coatings, seal coatings, film coatings, barrier
coatings, compress coatings, fast disintegrating coatings, or
enzyme degradable coatings. Multiple coatings can be applied for
desired performance. Further, the dosage form can be designed for
immediate release, pulsatile release, controlled release, extended
release, delayed release, targeted release, synchronized release,
or targeted delayed release. For release/absorption control, solid
carriers can be made of various component types and levels or
thicknesses of coats, with or without an active ingredient. Such
diverse solid carriers can be blended in a dosage form to achieve a
desired performance. The definitions of these terms are known to
those skilled in the art. In addition, the dosage form release
profile can be affected by a polymeric matrix composition, a coated
matrix composition, a multiparticulate composition, a coated
multiparticulate composition, an ion-exchange resin-based
composition, an osmosis-based composition, or a biodegradable
polymeric composition. Without wishing to be bound by theory, it is
believed that the release may be effected through favorable
diffusion, dissolution, erosion, ion-exchange, osmosis or
combinations thereof.
[0176] When formulated as a capsule, the capsule can be a hard or
soft gelatin capsule, a starch capsule, or a cellulosic capsule.
Although not limited to capsules, such dosage forms can further be
coated with, for example, a seal coating, an enteric coating, an
extended release coating, or a targeted delayed release coating.
These various coatings are known in the art, but for clarity, the
following brief descriptions are provided: seal coating, or coating
with isolation layers: Thin layers of up to 20 microns in thickness
can be applied for variety of reasons, including for particle
porosity reduction, to reduce dust, for chemical protection, to
mask taste, to reduce odor, to minimize gastrointestinal
irritation, etc. The isolating effect is proportional to the
thickness of the coating. Water soluble cellulose ethers are
preferred for this application. HPMC and ethyl cellulose in
combination, or Eudragit E 100, may be particularly suitable for
taste masking applications. Traditional enteric coating materials
listed elsewhere can also be applied to form an isolating
layer.
[0177] Extended release coatings are designed to effect delivery
over an extended period of time. The extended release coating is a
pH-independent coating formed of, for example, ethyl cellulose,
hydroxypropyl cellulose, methylcellulose, hydroxymethyl cellulose,
hydroxyethyl cellulose, acrylic esters, or sodium carboxymethyl
cellulose. Various extended release dosage forms can be readily
designed by one skilled in art to achieve delivery to both the
small and large intestines, to only the small intestine, or to only
the large intestine, depending upon the choice of coating materials
and/or coating thickness.
[0178] Enteric coatings are mixtures of pharmaceutically acceptable
excipients which are applied to, combined with, mixed with or
otherwise added to the carrier or composition. The coating may be
applied to a compressed or molded or extruded tablet, a gelatin
capsule, and/or pellets, beads, granules or particles of the
carrier or composition. The coating may be applied through an
aqueous dispersion or after dissolving in appropriate solvent.
Additional additives and their levels, and selection of a primary
coating material or materials will depend on the following
properties: 1. resistance to dissolution and disintegration in the
stomach; 2. impermeability to gastric fluids and
drug/carrier/enzyme while in the stomach; 3. ability to dissolve or
disintegrate rapidly at the target intestine site; 4. physical and
chemical stability during storage; 5. non-toxicity; 6. easy
application as a coating (substrate friendly); and 7. economical
practicality.
[0179] Dosage forms of the compositions of the present invention
can also be formulated as enteric coated delayed release oral
dosage forms, i.e., as an oral dosage form of a pharmaceutical
composition as described herein which utilizes an enteric coating
to affect release in the lower gastrointestinal tract. The enteric
coated dosage form may be a compressed or molded or extruded
tablet/mold (coated or uncoated) containing granules, pellets,
beads or particles of the active ingredient and/or other
composition components, which are themselves coated or uncoated.
The enteric coated oral dosage form may also be a capsule (coated
or uncoated) containing pellets, beads or granules of the solid
carrier or the composition, which are themselves coated or
uncoated.
[0180] Delayed release generally refers to the delivery so that the
release can be accomplished at some generally predictable location
in the lower intestinal tract more distal to that which would have
been accomplished if there had been no delayed release alterations.
The preferred method for delay of release is coating. Any coatings
should be applied to a sufficient thickness such that the entire
coating does not dissolve in the gastrointestinal fluids at pH
below about 5, but does dissolve at pH about 5 and above. It is
expected that any anionic polymer exhibiting a pH-dependent
solubility profile can be used as an enteric coating in the
practice of the present invention to achieve delivery to the lower
gastrointestinal tract. Polymers for use in the present invention
are anionic carboxylic polymers.
[0181] Shellac, also called purified 1ac, a refined product
obtained from the, resinous secretion of an insect. This coating
dissolves in media of pH>7.
[0182] Colorants, detackifiers, surfactants, antifoaming agents,
lubricants, stabilizers such as hydroxy propyl cellulose, acid/base
may be added to the coatings besides plasticizers to solubilize or
disperse the coating material, and to improve coating performance
and the coated product.
[0183] In carrying out the method of the present invention, the
combination of the invention may be administered to mammalian
species, such as dogs, cats, humans, etc. and as such may be
incorporated in a conventional systemic dosage form, such as a
tablet, capsule, elixir or injectable. The above dosage forms will
also include the necessary carrier material, excipient, lubricant,
buffer, antibacterial, bulking agent (such as mannitol),
anti-oxidants (ascorbic acid of sodium bisulfate) or the like.
[0184] The dose administered must be carefully adjusted according
to age, weight and condition of the patient, as well as the route
of administration, dosage form and regimen and the desired
result.
[0185] The pharmaceutical compositions of the invention may be
administered in the dosage forms in single or divided doses of one
to four times daily. It may be advisable to start a patient on a
low dose combination and work up gradually to a high dose
combination.
[0186] Tablets of various sizes can be prepared, e.g., of about 1
to 2000 mg in total weight, containing one or both of the active
pharmaceutical ingredients, with the remainder being a
physiologically acceptable carrier of other materials according to
accepted pharmaceutical practice. These tablets can be scored to
provide for fractional doses. Gelatin capsules can be similarly
formulated.
[0187] Liquid formulations can also be prepared by dissolving or
suspending one or the combination of active substances in a
conventional liquid vehicle acceptable for pharmaceutical
administration so as to provide the desired dosage in one to four
teaspoonfuls.
[0188] Dosage forms can be administered to the patient on a regimen
of, for example, one, two, three, four, five, six, or other doses
per day
[0189] In order to more finely regulate the dosage schedule, the
active substances may be administered separately in individual
dosage units at the same time or carefully coordinated times. Since
blood levels are built up and maintained by a regulated schedule of
administration, the same result is achieved by the simultaneous
presence of the two substances. The respective substances can be
individually formulated in separate unit dosage forms in a manner
similar to that described above.
[0190] In formulating the compositions, the active substances, in
the amounts described above, may be compounded according to
accepted pharmaceutical practice with a physiologically acceptable
vehicle, carrier, excipient, binder, preservative, stabilizer,
flavor, etc., in the particular type of unit dosage form.
[0191] Illustrative of the adjuvants which may be incorporated in
tablets are the following: a binder such as gum tragacanth, acacia,
corn starch or gelatin; an excipient such as dicalcium phosphate or
cellulose; a disintegrating agent such as corn starch, potato
starch, alginic acid or the like; a lubricant such as stearic acid
or magnesium stearate; a sweetening agent such as sucrose,
aspartame, lactose or saccharin; a flavoring agent such as orange,
peppermint, oil of wintergreen or cherry. When the dosage unit form
is a capsule, it may contain in addition to materials of the above
type a liquid carrier such as a fatty oil. Various other materials
may be present as coatings or to otherwise modify the physical form
of the dosage unit. For instance, tablets or capsules may be coated
with shellac, sugar or both. A syrup of elixir may contain the
active compound, water, alcohol or the like as the carrier,
glycerol as solubilizer, sucrose as sweetening agent, methyl and
propyl parabens as preservatives, a dye and a flavoring such as
cherry or orange.
[0192] One embodiment of this invention includes methods of
treating, preventing, or diagnosing a particular disease or
condition by administering the disclosed nanoparticles, composite
nanoparticles, nanosuspension, or nanocapsules to a subject. In
many instances, the nanoparticles, composite nanoparticles, or
nanocapsules are administered alone or can be included within a
pharmaceutical composition. An effective amount of a pharmaceutical
composition, generally, is defined as that amount sufficient to
ameliorate, reduce, minimize, or limit the extent of the disease or
condition. More rigorous definitions may apply, including
elimination, eradication, or cure of the disease or condition.
[0193] "Nanoparticles" are solid particles of an average particle
diameter of, for example, less than about 1 micron (micrometer).
One micron is 1,000 nanometers (nm).
[0194] "Stabilized" nanoparticles are nanoparticles coated with a
stabilizing material and having a reduced tendency for aggregation
and loss of dispersion with respect to nanoparticles of the
compound of the invention without a stabilizing coating.
[0195] A nano-spray is a spray containing nanoparticles or a spray
that produces nanoparticles. A nanodispersion is a dispersion
containing nanoparticles. A nanosuspension is a suspension
containing nanoparticles.
[0196] The liquid formulations useful herein may comprise a
solvent, solution, suspension, microsuspension, nanosuspension,
emulsion, microemulsion, gel or even a melt containing the active
component or components. In some embodiments the nanoparticles,
nanofibers, or nanofibrils may be in the form of, or within or on,
granules, powders, suspensions, solutions, dissolvable films, mats,
webs, tablets, or releasable forms particularly releasable dosage
forms. Other particular useful forms are concentrates to which a
diluting liquid is added prior to use. The product may also be
sprayed onto the inner surface of a container to which a liquid is
added later prior to use and the nanoparticles, nanofibers, or
nanofibrils, are released into the liquid.
[0197] Pharmaceutical compositions of the present invention can
include nanoparticles, composite nanoparticles, nanosuspension, or
nanocapsules of the present invention.
[0198] In certain non-limiting embodiments, pharmaceutical
compositions may comprise, for example, at least about 0.1% of an
active ingredient or nanoparticles, composite nanoparticles, or
nanocapsules, for example. In other embodiments, the an active
ingredient or nanoparticles, composite nanoparticles, or
nanocapsules may comprise between about 2% to about 75% of the
weight of the unit, or between about 25% to about 60%, for example,
and any range derivable therein. In non-limiting examples of a
derivable range from the numbers listed herein, a range of about 5
mg/kg/body weight to about 100 mg/kg/body weight, about 5
microgram/kg/body weight to about 500 milligram/kg/body weight,
etc., can be administered.
[0199] The composition may also include various antioxidants to
retard oxidation of one or more active ingredient or nanoparticles,
composite nanoparticles, nanosuspension, or nanocapsules. The
prevention of the action of microorganisms can be brought about by
preservatives such as various antibacterial and antifungal agents,
including but not limited to parabens (e.g., methylparabens,
propylparabens), chlorobutanol, phenol, sorbic acid, thimerosal or
combinations thereof.
[0200] In order to increase the effectiveness of a treatment with
the nanoparticles, nanogels, composite nanoparticles,
nanosuspension, or nanocapsules of the present invention, it may be
desirable to combine these nanoparticles, composite nanoparticles,
or nanocapsules with other therapies effective in the treatment of
a particular disease or condition.
[0201] The formulations as described above may be administered for
a prolonged period, that is, for as long as the potential for a
disease or condition remains or the symptoms continue.
Packaging/Treatment Kits
[0202] The present invention relates to a kit for conveniently and
effectively carrying out the methods in accordance with the present
invention. Such kits may be suited for the delivery of solid oral
forms such as tablets or capsules. Such a kit may include a number
of unit dosages. Such kits can include a means for containing the
dosages oriented in the order of their intended use. An example of
a means for containing the dosages in the order of their intended
uses is a card. An example of such a kit is a "blister pack".
Blister packs are well known in the packaging industry and are
widely used for packaging pharmaceutical unit dosage forms. If
desired, the blister can be in the form of a childproof blister,
i.e., a blister that is difficult for a child to open, yet can be
readily opened by an adult. If desired, a memory aid can be
provided, for example in the form of numbers, letters, or other
markings or with a calendar feature and/or calendar insert,
designating the days and the sections of a day in the treatment
schedule in which the dosages can be administered, such as an AM
dose is packaged with a "mid day" and a PM dose; or an AM dose is
packaged with a PM dose. Alternatively, placebo dosages, or vitamin
or dietary supplements, either in a form similar to or distinct
from the pharmaceutical active dosages, can be included.
[0203] In one aspect, the package, kit or container comprises a
"blister package" (also called a blister pack, or bubble pack). In
one aspect, the blister package consists of two or more separate
compartments: Am dosage of this invention, and PM dosage of this
invention, or mid-day dosage of this invention. This blister
package is made up of two separate material elements: a transparent
plastic cavity shaped to the product and its blister board backing.
These two elements are then joined together with a heat sealing
process which allows the product to be hung or displayed. Exemplary
types of "blister packages" include: Face seal blister packages,
gang run blister packages, mock blister packages, interactive
blister packages, slide blister packages.
[0204] Blister packs, clamshells or trays are forms of packaging
used for goods; thus, the invention provides for blister packs,
clamshells or trays comprising a composition (e.g., a (the
multi-ingredient combination of drugs of the invention) combination
of active ingredients) of the invention. Blister packs, clamshells
or trays can be designed to be non-reclosable, so consumers can
tell if a package has already opened. They are used to package for
sale goods where product tampering is a consideration, such as the
pharmaceuticals of the invention. In one aspect, a blister pack of
the invention comprises a moulded PVC base, with raised areas (the
"blisters") to contain the tablets, pills, etc. comprising the
combinations of the invention, covered by a foil laminate. Tablets,
pills, etc. are removed from the pack either by peeling the foil
back or by pushing the blister to force the tablet to break the
foil. In one aspect, a specialized form of a blister pack is a
strip pack.
[0205] In one aspect, a blister pack also comprises a method of
packaging where the compositions comprising combinations of
ingredients of the invention are contained in-between a card and a
clear PVC. The PVC can be transparent so the item (pill, tablet,
geltab, etc.) can be seen and examined easily; and in one aspect,
can be vacuum-formed around a mould so it can contain the item
snugly and have room to be opened upon purchase. In one aspect, the
card is brightly colored and designed depending on the item (pill,
tablet, geltab, etc.) inside, and the PVC is affixed to the card
using pre-formed tabs where the adhesive is placed. The adhesive
can be strong enough so that the pack may hang on a peg, but weak
enough so that this way one can tear open the join and access the
item. Sometimes with large items or multiple enclosed pills,
tablets, geltabs, etc., the card has a perforated window for
access. In one aspect, more secure blister packs, e.g., for items
such as pills, tablets, geltabs, etc. of the invention are used,
and they can comprise of two vacuum-formed PVC sheets meshed
together at the edges, with the informative card inside.
[0206] In one aspect, blister packaging comprises at least two
components (e.g., is a multi-ingredient combination of drugs of the
invention): a thermoformed "blister" which houses the product
(e.g., a pharmaceutical combination of the invention), and then a
"blister card" that is a printed card with an adhesive coating on
the front surface. During the assembly process, the blister
component, which is most commonly made out of PVC, is attached to
the blister card using a blister machine. This machine introduces
heat to the flange area of the blister which activates the glue on
the card in that specific area and ultimately secures the PVG
blister to the printed blister card. The thermoformed PVG blister
and the printed blister card can be as small or large. Conventional
blister packs can also be sealed (e.g., using an AERGO 8 DUO.RTM.,
SCA Consumer Packaging, Inc., DeKalb, Ill.) using regular heat seal
tooling. This alternative aspect, using heat seal tooling, can seal
common types of thermoformed packaging.
[0207] As discussed herein, the products of manufacture of the
invention can comprise the packaging of the therapeutic drug
combinations of the invention, alone or in combination, as "blister
packages" or as a plurality of packettes, including as lidded
blister packages, lidded blister or blister card or packets, or a
shrink wrap.
[0208] In one aspect, laminated aluminum foil blister packs are
used, e.g., for the preparation of drugs designed to dissolve
immediately in the mouth of a patient. This exemplary process
comprises having the drug combinations of the invention prepared as
an aqueous solution(s) which are dispensed (e.g., by measured dose)
into an aluminum (e.g., alufoil) laminated tray portion of a
blister pack. This tray is then freeze-dried to form tablets which
take the shape of the blister pockets. The alufoil laminate of both
the tray and lid fully protects any highly hygroscopic and/or
sensitive individual doses. In one aspect, the pack incorporates a
child-proof peel open security laminate. In one aspect, the system
give tablets an identification mark by embossing a design into the
alufoil pocket that is taken up by the tablets when they change
from aqueous to solid state. In one aspect, individual
`push-through` blister packs/packettes are used, e.g., using hard
temper aluminum (e.g., alufoil) lidding material. In one aspect,
hermetically-sealed high barrier aluminum (e.g., alufoil) laminates
are used. In one aspect, any of the invention's products of
manufacture, including kits or blister packs, use foil laminations
and strip packs, stick packs, sachets and pouches, peelable and
non-peelable laminations combining foil, paper, and film for high
barrier packaging.
[0209] Other means for containing said unit dosages can include
bottles and vials, wherein the bottle or vial comprises a memory
aid, such as a printed label for administering said unit dosage or
dosages. The label can also contain removable reminder stickers for
placement on a calendar or dayminder to further help the patient to
remember when to take a dosage or when a dosage has been taken.
ORG 34517 Promotes Sensitivity to Chemotherapy
[0210] The glucocorticoid receptor (GR) is expressed in a subset of
both ERalpha-positive and -negative human breast cancers as well as
in some ovarian cancers. In vitro and in vivo experiments suggest
that activation of the GR in ER-negative pre-malignant breast
epithelial and cancer cells iniates cell survival pathways under
stress conditions that normally induce significant cell death (e.g.
chemotherapy, radiation, growth factor deprivation). Thus, GR
antagonism is predicted to enhance chemotherapy sensitivity of
GR+/ER- breast cancer cells by blocking stress-mediated cell
survival pathways that would otherwise counteract
chemotherapy-induced apoptosis.
[0211] In support of the present invention that GR activation
mediates chemo-resistance (and associated increase risk of relapse
for early stage ER- breast cancers), recently the association was
examined between GR (NR3C1) gene expression and GR target gene
expression in human ER- breast cancers and cell lines. It was found
that in ER- breast cancers, high GR expression is associated with a
significantly increased probability of relapse in early stage
patients. This analysis and previous data in breast and ovarian
cancer have led to the present invention wherein "GR high" breast
tumors may respond better to cytotoxic therapies if glucocorticoid
receptor (GR) antagonist therapy is administered with chemotherapy.
Furthermore, GR antagonist therapy should not worsen the
lymphopenia sometimes associated with taxane therapy because GR
signaling is pro-apoptotic to lymphocytes.
Cancer/Tumor Stem Cells
[0212] Cancer stem cells comprise a unique subpopulation (often
0.1-10% or so) of a tumor that, relative to the remaining 90% or so
of the tumor (i.e., the tumor bulk), are more tumorigenic,
relatively more slow-growing or quiescent, and often relatively
more chemoresistant than the tumor bulk. Given that conventional
therapies and regimens have, in large part, been designed to attack
rapidly proliferating cells (i.e., those cancer cells that comprise
the tumor bulk), cancer stem cells which are often slow-growing may
be relatively more resistant than faster growing tumor bulk to
conventional therapies and regimens. Cancer stem cells can express
other features which make them relatively chemoresistant such as
multi-drug resistance and anti-apoptotic pathways. The
aforementioned would constitute a key reason for the failure of
standard oncology treatment regimens to ensure long-term benefit in
most patients with advanced stage cancers--i.e., the failure to
adequately target and eradicate cancer stem cells. In some
instances, a cancer stem cell(s) is the founder cell of a tumor
(i.e., it is the progenitor of the cancer cells that comprise the
tumor bulk).
[0213] Cancer stem cells have been identified in a large variety of
cancer types. For instance, Bonnet et al., using flow cytometry
were able to isolate the leukemia cells bearing the specific
phenotype CD34+ CD38-, and subsequently demonstrate that it is
these cells (comprising <1% of a given leukemia), unlike the
remaining 99+% of the leukemia bulk, that are able to recapitulate
the leukemia from which it was derived when transferred into
immunodeficient mice. See, e.g., Nat Med 3:730-737 (1997). That is,
these cancer stem cells were found as <1 in 10,000 leukemia
cells yet this low frequency population was able to initiate and
serially transfer a human leukemia into severe combined
immunodeficiency/non-obese diabetic (NOD/SCID) mice with the same
histologic phenotype as in the original tumor.
[0214] Cox et al. identified small subfractions of human acute
lymphoblastic leukemia (ALL) cells which had the phenotypes
CD34.sup.+/CD10.sup.- and CD34.sup.+/CD19.sup.-, and were capable
of engrafting ALL tumors in immunocompromised mice--i.e. the cancer
stem cells. In contrast, no engraftment of the mice was observed
using the ALL bulk, despite, in some cases, injecting 10-fold more
cells. See Cox et al., Blood 104(19): 2919-2925 (2004).
[0215] Multiple myeloma was found to contain small subpopulations
of cells that were CD138- and, relative to the large bulk
population of CD138+ myeloma cells, had greater clonogenic and
tumorigenic potential. See Matsui et al., "Characterization of
clonogenic multiple myeloma cells," Blood 103(6): 2332. The authors
concluded that the CD138-subpopulation of multiple myeloma was the
cancer stem cell population.
[0216] Kondo et al. isolated a small population of cells from a
C6-glioma cell line, which was identified as the cancer stem cell
population by virtue of its ability to self-renew and recapitulate
gliomas in immunocompromised mice. See Kondo et al., Proc. Natl.
Acad. Sci. USA 101:781-786 (2004). In this study, Kondo et al.
determined that cancer cell lines contain a population of cancer
stem cells that confer the ability of the line to engraft
immunodeficient mice.
[0217] Breast cancers were shown to contain a small population of
cells with stem cell characteristics (bearing surface markers
CD44+CD24low lin-). See Al-Hajj et al., Proc. Natl. Acad. Sci. USA
100:3983-3988 (2003). As few as 200 of these cells, corresponding
to 1-10% of the total tumor cell population, are able to form
tumors in NOD/SCID mice. In contrast, implantation of 20,000 cells
that lacked this phenotype (i.e. the tumor bulk) was unable to
re-grow the tumor.
[0218] A subpopulation of cells derived from human prostate tumors
was found to self-renew and to recapitulate the phenotype of the
prostate tumor from which they were derived thereby constituting
the prostate cancer stem cell population. See Collins et al.,
"Prospective Identification of Tumorigenic Prostate Cancer Stem
Cells," Cancer Res 65(23):10946-10951 (2005).
[0219] Fang et al. isolated a subpopulation of cells from melanoma
with cancer stem cell properties. In particular, this subpopulation
of cells could differentiate and self-renew. In culture, the
subpopulation formed spheres whereas the more differentiated cell
fraction from the lesions were more adherent. Moreover, the
subpopulation containing sphere-like cells were more tumorigenic
than the adherent cells when grafted into mice. See Fang et al., "A
Tumorigenic Subpopulation with Stem Cell Properties in Melanomas,"
Cancer Res 65(20): 9328-9337 (2005).
[0220] Singh et al. identified brain tumor stem cells. When
isolated and transplanted into nude mice, the CD133+ cancer stem
cells, unlike the CD133-tumor bulk cells, form tumors that can then
be serially transplanted. See Singh et al., "Identification of
human brain tumor initiating cells," Nature 432:396-401 (2004);
Singh et al., "Cancer stem cells in nervous system tumors,"
Oncogene 23:7267-7273 (2004); Singh et al., "Identification of a
cancer stem cell in human brain tumors," Cancer Res. 63:5821-5828
(2003).
[0221] Since conventional cancer therapies target rapidly
proliferating cells (i.e., cells that form the tumor bulk) these
treatments are believed to be relatively ineffective at targeting
and impairing cancer stem cells. In fact, cancer stem cells,
including leukemia stem cells, have indeed been shown to be
relatively resistant to conventional chemotherapeutic therapies
(e.g. Ara-C, daunorubicin) as well as other targeted therapies
(e.g. Gleevec.RTM., Velcade.RTM.).
Cancer Therapies
[0222] Any therapy (e.g., therapeutic or prophylactic agent) which
is useful, has been used, is currently being used, or may be used
for the prevention, treatment and/or management of cancer can be
used to prevent, treat, and/or manage the patient whose neoplasia
and/or cancer stem cells are monitored in accordance with the
methods of the invention. Also, such neoplasia and/or cancer stem
cell monitoring can be employed in conjunction with any therapy for
cancer according to the instant invention. Therapies (e.g.,
therapeutic or prophylactic agents) include, but are not limited
to, peptides, polypeptides, fusion proteins, nucleic acid
molecules, small molecules, mimetic agents, synthetic drugs,
inorganic molecules, and organic molecules. Non-limiting examples
of cancer therapies include chemotherapies, radiation therapies,
hormonal therapies, anti-angiogenesis therapies, targeted
therapies, and/or biological therapies including immunotherapies
and surgery. In certain embodiments, a prophylactically and/or
therapeutically effective regimen comprises the administration of a
combination of therapies. In certain embodiments, ORG 34517 can be
administered as an agent to treat or prevent neoplasia. In certain
embodiments, RU486 (mifepristone) can be administered as an agent
to treat or prevent neoplasia.
[0223] Examples of cancer therapies include, but are not limited
to: acivicin; aclarubicin; acodazole hydrochloride; acronine;
adozelesin; aldesleukin; altretamine; ambomycin; ametantrone
acetate; aminoglutethimide; amsacrine; anastrozole; anthracyclin;
anthramycin; asparaginase; asperlin; azacitidine (Vidaza); azetepa;
azotomycin; batimastat; benzodepa; bicalutamide; bisantrene
hydrochloride; bisnafide dimesylate; bisphosphonates (e.g.,
pamidronate (Aredria), sodium clondronate (Bonefos), zoledronic
acid (Zometa), alendronate (Fosamax), etidronate, ibandornate,
cimadronate, risedromate, and tiludromate); bizelesin; bleomycin
sulfate; brequinar sodium; bropirimine; busulfan; cactinomycin;
calusterone; caracemide; carbetimer; carboplatin; carmustine;
carubicin hydrochloride; carzelesin; cedefingol; chlorambucil;
cirolemycin; cisplatin; cladribine; crisnatol mesylate;
cyclophosphamide; cytarabine (Ara-C); dacarbazine; dactinomycin;
daunorubicin hydrochloride; decitabine (Dacogen); demethylation
agents, dexormaplatin; dezaguanine; dezaguanine mesylate;
diaziquone; docetaxel; doxorubicin; doxorubicin hydrochloride;
droloxifene; droloxifene citrate; dromostanolone propionate;
duazomycin; edatrexate; eflornithine hydrochloride; EphA2
inhibitors; elsamitrucin; enloplatin; enpromate; epipropidine;
epirubicin hydrochloride; erbulozole; esorubicin hydrochloride;
estramustine; estramustine phosphate sodium; etanidazole;
etoposide; etoposide phosphate; etoprine; fadrozole hydrochloride;
fazarabine; fenretinide; floxuridine; fludarabine phosphate;
fluorouracil; fluorocitabine; fosquidone; fostriecin sodium;
gemcitabine; gemcitabine hydrochloride; histone deacetylase
inhibitors (HDAC-Is) hydroxyurea; idarubicin hydrochloride;
ifosfamide; ilmofosine; imatinib mesylate (Gleevec, Glivec);
interleukin II (including recombinant interleukin II, or rIL2),
interferon alpha-2a; interferon alpha-2b; interferon alpha-n1;
interferon alpha-n3; interferon beta-I a; interferon gamma-I b;
iproplatin; irinotecan hydrochloride; lanreotide acetate;
lenalidomide (Revlimid); letrozole; leuprolide acetate; liarozole
hydrochloride; lometrexol sodium; lomustine; losoxantrone
hydrochloride; masoprocol; maytansine; mechlorethamine
hydrochloride; anti-CD2 antibodies (e.g., siplizumab (MedImmune
Inc.; International Publication No. WO 02/098370, which is
incorporated herein by reference in its entirety)); megestrol
acetate; melengestrol acetate; melphalan; menogaril;
mercaptopurine; methotrexate; methotrexate sodium; metoprine;
meturedepa; mifepristone; mitindomide; mitocarcin; mitocromin;
mitogillin; mitomalcin; mitomycin; mitosper; mitotane; mitoxantrone
hydrochloride; mycophenolic acid; nocodazole; nogalamycin; ORG
34517; ormaplatin; oxaliplatin; oxisuran; paclitaxel; pegaspargase;
peliomycin; pentamustine; peplomycin sulfate; perfosfamide;
pipobroman; piposulfan; piroxantrone hydrochloride; plicamycin;
plomestane; porfimer sodium; porfiromycin; prednimustine;
procarbazine hydrochloride; puromycin; puromycin hydrochloride;
pyrazofurin; riboprine; rogletimide; RU486; safingol; safingol
hydrochloride; semustine; simtrazene; sparfosate sodium;
sparsomycin; spirogermanium hydrochloride; spiromustine;
spiroplatin; streptonigrin; streptozocin; sulofenur; talisomycin;
tecogalan sodium; tegafur; teloxantrone hydrochloride; temoporfin;
teniposide; teroxirone; testolactone; thiamiprine; thioguanine;
thiotepa; tiazofurin; tirapazamine; toremifene citrate; trestolone
acetate; triciribine phosphate; trimetrexate; trimetrexate
glucuronate; triptorelin; tubulozole hydrochloride; uracil mustard;
uredepa; vapreotide; verteporfin; vinblastine sulfate; vincristine
sulfate; vindesine; vindesine sulfate; vinepidine sulfate;
vinglycinate sulfate; vinleurosine sulfate; vinorelbine tartrate;
vinrosidine sulfate; vinzolidine sulfate; vorozole; zeniplatin;
zinostatin; zorubicin hydrochloride.
[0224] Other examples of cancer therapies include, but are not
limited to: 20-epi-1,25 dihydroxyvitamin D3; 5-ethynyluracil;
abiraterone; aclarubicin; acylfulvene; adecypenol; adozelesin;
aldesleukin; ALL-TK antagonists; altretamine; ambamustine; amidox;
amifostine; aminolevulinic acid; amrubicin; amsacrine; anagrelide;
anastrozole; andrographolide; angiogenesis inhibitors; antagonist
D; antagonist G; antarelix; anti-dorsalizing morphogenetic
protein-1; antiandrogen, prostatic carcinoma; antiestrogen;
antineoplaston; antisense oligonucleotides; aphidicolin glycinate;
apoptosis gene modulators; apoptosis regulators; apurinic acid;
ara-CDP-DL-PTBA; arginine deaminase; asulacrine; atamestane;
atrimustine; axinastatin 1; axinastatin 2; axinastatin 3;
azasetron; azatoxin; azatyrosine; baccatin III derivatives;
balanol; batimastat; BCR/ABL antagonists; benzochlorins;
benzoylstaurosporine; beta lactam derivatives; beta-alethine;
betaclamycin B; betulinic acid; bFGF inhibitor; bicalutamide;
bisantrene; bisaziridinylspermine; bisnafide; bistratene A;
bizelesin; breflate; bropirimine; budotitane; buthionine
sulfoximine; calcipotriol; calphostin C; camptothecin derivatives;
canarypox IL-2; capecitabine; carboxamide-amino-triazole;
carboxyamidotriazole; CaRest M3; CARN 700; cartilage derived
inhibitor; carzelesin; casein kinase inhibitors (ICOS);
castanospermine; cecropin B; cetrorelix; chlorins;
chloroquinoxaline sulfonamide; cicaprost; cis-porphyrin;
cladribine; clomifene analogues; clotrimazole; collismycin A;
collismycin B; combretastatin A4; combretastatin analogue;
conagenin; crambescidin 816; crisnatol; cryptophycin 8;
cryptophycin A derivatives; curacin A; cyclopentanthraquinones;
cycloplatam; cypemycin; cytarabine ocfosfate; cytolytic factor;
cytostatin; dacliximab; decitabine; dehydrodidemnin B; deslorelin;
dexamethasone; dexifosfamide; dexrazoxane; dexverapamil;
diaziquone; didemnin B; didox; diethylnorspermine;
dihydro-5-azacytidine; dihydrotaxol, dioxamycin; diphenyl
spiromustine; docetaxel; docosanol; dolasetron; doxifluridine;
droloxifene; dronabinol; duocarmycin SA; ebselen; ecomustine;
edelfosine; edrecolomab; eflomithine; elemene; emitefur;
epirubicin; epristeride; estramustine analogue; estrogen agonists;
estrogen antagonists; etanidazole; etoposide phosphate; exemestane;
fadrozole; fazarabine; fenretinide; filgrastim; finasteride;
flavopiridol; flezelastine; fluasterone; fludarabine;
fluorodaunorunicin hydrochloride; forfenimex; formestane;
fostriecin; fotemustine; gadolinium texaphyrin; gallium nitrate;
galocitabine; ganirelix; gelatinase inhibitors; gemcitabine;
glutathione inhibitors; HMG CoA reductase inhibitors (e.g.,
atorvastatin, cerivastatin, fluvastatin, lescol, lupitor,
lovastatin, rosuvastatin, and simvastatin); hepsulfam; heregulin;
hexamethylene bisacetamide; hypericin; ibandronic acid; idarubicin;
idoxifene; idramantone; ilmofosine; ilomastat; imidazoacridones;
imiquimod; immunostimulant peptides; insulin-like growth factor-1
receptor inhibitor; interferon agonists; interferons; interleukins;
iobenguane; iododoxorubicin; ipomeanol, 4-iroplact; irsogladine;
isobengazole; isohomohalicondrin B; itasetron; jasplakinolide;
kahalalide F; lamellarin-N triacetate; lanreotide; leinamycin;
lenograstim; lentinan sulfate; leptolstatin; letrozole; leukemia
inhibiting factor; leukocyte alpha interferon;
leuprolide+estrogen+progesterone; leuprorelin; levamisole; LFA-3TIP
(Biogen, Cambridge, Mass.; International Publication No. WO 93/0686
and U.S. Pat. No. 6,162,432); liarozole; linear polyamine analogue;
lipophilic disaccharide peptide; lipophilic platinum compounds;
lissoclinamide 7; lobaplatin; lombricine; lometrexol; lonidamine;
losoxantrone; lovastatin; loxoribine; lurtotecan; lutetium
texaphyrin; lysofylline; lytic peptides; maitansine; mannostatin A;
marimastat; masoprocol; maspin; matrilysin inhibitors; matrix
metalloproteinase inhibitors; menogaril; merbarone; meterelin;
methioninase; metoclopramide; MIF inhibitor; mifepristone;
miltefosine; mirimostim; mismatched double stranded RNA;
mitoguazone; mitolactol; mitomycin analogues; mitonafide; mitotoxin
fibroblast growth factor-saporin; mitoxantrone; mofarotene;
molgramostim; monoclonal antibody, human chorionic gonadotrophin;
monophosphoryl lipid A+myobacterium cell wall sk; mopidamol;
multiple drug resistance gene inhibitor; multiple tumor suppressor
1-based therapy; mustard anticancer agent; mycaperoxide B;
mycobacterial cell wall extract; myriaporone; N-acetyldinaline;
N-substituted benzamides; nafarelin; nagrestip;
naloxone+pentazocine; napavin; naphterpin; nartograstim;
nedaplatin; nemorubicin; neridronic acid; neutral endopeptidase;
nilutamide; nisamycin; nitric oxide modulators; nitroxide
antioxidant; nitrullyn; 06-benzylguanine; octreotide; okicenone;
oligonucleotides; onapristone; oracin; oral cytokine inducer;
ormaplatin; osaterone; oxaliplatin; oxaunomycin; paclitaxel;
paclitaxel analogues; paclitaxel derivatives; palauamine;
palmitoylrhizoxin; pamidronic acid; panaxytriol; panomifene;
parabactin; pazelliptine; pegaspargase; peldesine; pentosan
polysulfate sodium; pentostatin; pentrozole; perflubron;
perfosfamide; perillyl alcohol; phenazinomycin; phenylacetate;
phosphatase inhibitors; picibanil; pilocarpine hydrochloride;
pirarubicin; piritrexim; placetin A; placetin B; plasminogen
activator inhibitor; platinum complex; platinum compounds;
platinum-triamine complex; porfimer sodium; porfiromycin;
prednisone; propyl bis-acridone; prostaglandin J2; proteasome
inhibitors; protein A-based immune modulator; protein kinase C
inhibitor; protein kinase C inhibitors, microalgal; protein
tyrosine phosphatase inhibitors; purine nucleoside phosphorylase
inhibitors; purpurins; pyrazoloacridine; pyridoxylated hemoglobin
polyoxyethylene; raf antagonists; raltitrexed; ramosetron; ras
farnesyl protein transferase inhibitors; ras inhibitors; ras-GAP
inhibitor; retelliptine demethylated; rhenium Re 186 etidronate;
rhizoxin; ribozymes; RII retinamide; rogletimide; rohitukine;
romurtide; roquinimex; rubiginone B1; ruboxyl; safingol; saintopin;
SarCNU; sarcophytol A; sargramostim; Sdi 1 mimetics; semustine;
senescence derived inhibitor 1; sense oligonucleotides; signal
transduction inhibitors; signal transduction modulators; gamma
secretase inhibitors, single chain antigen binding protein;
sizofuran; sobuzoxane; sodium borocaptate; sodium phenylacetate;
solverol; somatomedin binding protein; sonermin; sparfosic acid;
spicamycin D; spiromustine; splenopentin; spongistatin 1;
squalamine; stem cell inhibitor; stem-cell division inhibitors;
stipiamide; stromelysin inhibitors; sulfinosine; superactive
vasoactive intestinal peptide antagonist; suradista; suramin;
swainsonine; synthetic glycosaminoglycans; tallimustine;
5-fluorouracil; leucovorin; tamoxifen methiodide; tauromustine;
tazarotene; tecogalan sodium; tegafur; tellurapyrylium; telomerase
inhibitors; temoporfin; temozolomide; teniposide;
tetrachlorodecaoxide; tetrazomine; thaliblastine; thiocoraline;
thrombopoietin; thrombopoietin mimetic; thymalfasin; thymopoietin
receptor agonist; thymotrinan; thyroid stimulating hormone; tin
ethyl etiopurpurin; tirapazamine; titanocene bichloride; topsentin;
toremifene; totipotent stem cell factor; translation inhibitors;
tretinoin; triacetyluridine; triciribine; trimetrexate;
triptorelin; tropisetron; turosteride; tyrosine kinase inhibitors;
tyrphostins; UBC inhibitors; ubenimex; urogenital sinus-derived
growth inhibitory factor; urokinase receptor antagonists;
vapreotide; variolin B; vector system, erythrocyte gene therapy;
thalidomide; velaresol; veramine; verdins; verteporfin;
vinorelbine; vinxaltine; anti-integrin antibodies (e.g.,
anti-integrin a.sub.vb.sub.3 antibodies); vorozole; zanoterone;
zeniplatin; zilascorb; and zinostatin stimalamer.
[0225] A non-limiting list of compounds that could be used to
target cancer stem cells includes: inhibitors of interleukin-3
receptor (IL-3R) and CD123 (including peptides, peptide-conjugates,
antibodies, antibody-conjugates, antibody fragments, and antibody
fragment-conjugates that target IL-3R or CD123); cantharidin;
norcantharidin and analogs and derivatives thereof; Notch pathway
inhibitors including gamma secretase inhibitors; sonic
hedgehog/smoothened pathway inhibitors including cyclopamine and
analogs thereof; antibodies to CD96; certain NF-kB/proteasome
inhibitors including parthenolide and analogs thereof; certain
triterpenes including celastrol; certain mTOR inhibitors; compounds
and antibodies that target the urokinase receptor; sinefungin;
certain inosine monophosphate dehydrogenase (IMPDH) inhibitors;
PPAR-alpha and PPAR-gamma agonists and antagonists (including
pioglitazone, tesaslitazar, muraglitazar, peliglitazar,
lobeglitazone, balaglitazone, ragaglitazar, rosiglitazone,
farglitazar, sodeiglitazar, reglitazar, naveglitazar, oxeglitazar,
metaglidasen, netoglitazone, darglitazone, englitazone,
thiazolidinediones, aleglitazar, edaglitazone, rivoglitazone,
troglitazone, imiglitazar, and sipoglitazar); telomerase
inhibitors; antibodies to EpCAM (ESA); GSK-3 beta agonists and
antagonists (including Lithium, 6-bromoinirubin-3'-oxime (BIO),
TDZD8); Wnt pathway inhibitors including antibodies to frizzled or
small molecules that inhibit disheveled/frizzled or beta catenin;
anti-CD20 antibodies and conjugates (e.g. Rituxan, Bexxar, Zevalin)
for novel use in multiple myeloma or melanoma; anti-CD133 antibody;
anti-CD44 antibody; antibodies to IL-4; certain differentiation
agents such as versnarinone; compounds that target CD33 such as an
antibody or betulinic acid; compounds that target lactadherin such
as an antibody; small molecules or antibodies that target CXCR4 or
SDF-1; small molecules or antibodies that target multi-drug
resistance pumps; inhibitors of survivin; inhibitors of XIAP; small
molecules that target Bcl-2; antibodies to CLL-1; and furin
inhibitors (such as cucurbitacins).
[0226] An additional non-limiting list of compounds that could also
be used to target cancer and/or cancer stem cells includes: i)
antibodies, antibody fragments, and proteins that are either naked
or conjugated to a therapeutic moiety that target certain cell
surface targets on cancer stem cells, or ii) small molecules known
in the art including ones that can be further optimized (e.g., via
chemistry) or identified via a cancer stem cell-based screen (e.g.,
such as one that would determine whether a compound impairs
proliferation or viability of a cancer stem cell through standard
methods, the cell surface and intracellular targets including (not
meant to be exhaustive) are: Rex1 (Zfp42), CTGF, Activin A, Wnt,
FGF-2, HIF-1, AP-2gamma, Bmi-1, nucleostemin, hiwi, Moz-TIF2,
Nanog, beta-arrestin-2, Oct-4, Sox2, stella, GDF3, RUNX3, EBAF,
TDGF-1, nodal, ZFPY, PTNE, Evi-1, Pax3, Mcl-1, c-kit, Lex-1, Zfx,
lactadherin, aldehyde dehydrogenase, BCRP, telomerase, CD133,
Bcl-2, CD26, Gremlin, and FoxC2.
[0227] In some embodiments, the therapy(ies) is an immunomodulatory
agent. Non-limiting examples of immunomodulatory agents include
proteinaceous agents such as cytokines, peptide mimetics, and
antibodies (e.g., human, humanized, chimeric, monoclonal,
polyclonal, Fvs, ScFvs, Fab or F(ab)2 fragments or epitope binding
fragments), nucleic acid molecules (e.g., antisense nucleic acid
molecules and triple helices), small molecules, organic compounds,
and inorganic compounds. In particular, immunomodulatory agents
include, but are not limited to, methotrexate, leflunomide,
cyclophosphamide, cytoxan, Immuran, cyclosporine A, minocycline,
azathioprine, antibiotics (e.g., FK506 (tacrolimus)),
methylprednisolone (MP), corticosteroids, steroids, mycophenolate
mofetil, rapamycin (sirolimus), mizoribine, deoxyspergualin,
brequinar, malononitriloamides (e.g., leflunamide), T cell receptor
modulators, cytokine receptor modulators, and modulators mast cell
modulators. Other examples of immunomodulatory agents can be found,
e.g., in U.S. Publication No. 2005/0002934 A1 at paragraphs 259-275
which is incorporated herein by reference in its entirety. In one
embodiment, the immunomodulatory agent is a chemotherapeutic agent.
In an alternative embodiment, the immunomodulatory agent is an
immunomodulatory agent other than a chemotherapeutic agent. In some
embodiments, the therapy(ies) used in accordance with the invention
is not an immunomodulatory agent.
[0228] In some embodiments, the therapy(ies) is an anti-angiogenic
agent. Non-limiting examples of anti-angiogenic agents include
proteins, polypeptides, peptides, fusion proteins, antibodies
(e.g., human, humanized, chimeric, monoclonal, polyclonal, Fvs,
ScFvs, Fab fragments, F(ab).sub.2 fragments, and antigen-binding
fragments thereof) such as antibodies that specifically bind to
TNF-alpha, nucleic acid molecules (e.g., antisense molecules or
triple helices), organic molecules, inorganic molecules, and small
molecules that reduce or inhibit angiogenesis. Other examples of
anti-angiogenic agents can be found, e.g., in U.S. Publication No.
2005/0002934 A1 at paragraphs 277-282, which is incorporated by
reference in its entirety. In other embodiments, the therapy(ies)
is not an anti-angiogenic agent.
[0229] In certain embodiments, the therapy(ies) is an alkylating
agent, a nitrosourea, an antimetabolite, and anthracyclin, a
topoisomerase II inhibitor, or a mitotic inhibitor. Alkylating
agents include, but are not limited to, busulfan, cisplatin,
carboplatin, chlorambucil, cyclophosphamide, ifosfamide,
decarbazine, mechlorethamine, mephalen, and themozolomide.
Nitrosoureas include, but are not limited to carmustine (BCNU) and
lomustine (CCNU). Antimetabolites include but are not limited to
5-fluorouracil, capecitabine, methotrexate, gemcitabine,
cytarabine, and fludarabine. Anthracyclins include but are not
limited to daunorubicin, doxorubicin, epirubicin, idarubicin, and
mitoxantrone. Topoisomerase II inhibitors include, but are not
limited to, topotecan, irinotecan, etopiside (VP-16), and
teniposide. Mitotic inhibitors include, but are not limited to
taxanes (paclitaxel, docetaxel), and the vinca alkaloids
(vinblastine, vincristine, and vinorelbine). In some embodiments of
the invention, the therapy(ies) includes the administration
cantharidin or an analog thereof. The invention includes the use of
agents that target cancer stem cells. In certain embodiments, the
agent acts alone. In other embodiments, the agent is attached
directly or indirectly to another therapeutic moiety. Non-limiting
examples of therapeutic moieties include, but are not limited to
alkylating agents, anti-metabolites, plant alkaloids, cytotoxic
agents, chemotherapeutic agents (e.g., a steroid, cytosine
arabinoside, fluoruracil, methotrexate, aminopterin, mitomycin C,
demecolcine, etoposide, mithramycin, calicheamicin, CC-1065,
chlorambucil or melphalan), radionuclides, therapeutic enzymes,
cytokines, toxins including plant-derived toxins, fungus-derived
toxins, bacteria-derived toxin (e.g., deglycosylated ricin A chain,
a ribosome inactivating protein, alpha-sarcin, aspergillin,
restirictocin, a ribonuclease, a diphtheria toxin, Pseudomonas
exotoxin, a bacterial endotoxin or the lipid A moiety of a
bacterial endotoxin), growth modulators and RNase. In some
embodiments, the agent used is an agent that binds to a marker,
e.g., an antigen on a cancer stem cell. In a specific embodiment,
the agent binds to an antigen that is expressed at a greater level
on cancer stem cells than on normal stem cells. In a specific
embodiment, the agent binds specifically to a cancer stem cell
antigen that is not a normal stem cell. In other embodiments, the
therapy(ies) is an agent that binds to a marker on cancer stem
cells. In one embodiment, the agent that binds to a marker on
cancer stem cells is an antibody or an antibody conjugated to a
therapeutic moiety or an antibody fragment conjugated to a
therapeutic moiety.
[0230] For example, in a specific embodiment, the agent binds
specifically to the IL-3 Receptor (IL-3R). In some embodiments, the
agent that binds to the IL-3R is an antibody or an antibody
fragment that is specific for IL-3R. In some embodiments, the
antibody or antibody fragment is conjugated either chemically or
via recombinant technology to a therapeutic moiety (e.g., a
chemotherapeutic agent, a plant-, fungus- or bacteria-derived
toxin, a radionuclide) using a linking agent to effect a cell
killing response. In certain embodiments, the antibody,
antibody-conjugate, antibody fragment, or antibody
fragment-conjugate binds to the .alpha.-subunit of IL-3R (i.e., the
CD123 antigen). In other embodiments, the antibody,
antibody-conjugate, antibody fragment, or antibody
fragment-conjugate binds to the IL-3R, containing both the .alpha.
and .beta. subunits. Methods for preparing antibodies to IL-3R and
mimetics of antibodies to IL-3R are described in U.S. Pat. No.
6,733,743 B2, which is incorporated herein by reference in its
entirety.
[0231] In other embodiments, the agent that binds to a marker on
cancer stem cells is a ligand. In some embodiments, the ligand is a
cytokine that binds to a cytokine receptor on cancer stem cells. In
a particular embodiment, the ligand is interleukin-3 (IL-3) which
can be conjugated to a therapeutic moiety that includes a
chemotherapeutic agent, a plant-, fungus-, or bacteria-derived
toxin, or a radionuclide. The IL-3-conjugate prophylactic and/or
therapeutic therapy or regimen can be in the form of a recombinant
fusion protein in embodiments where the conjugate is a toxin and
the toxin is a protein, such as diphtheria toxin. Methods for
preparing and isolating an IL-3-diphtheria toxin fusion protein
(IL3DT) are described in Frankel et al., "Diphtheria toxin fused to
human interleukin-3 is toxic to blasts from patients with myeloid
leukemias," Leukemia 14:576 (2000) and Urieto et al., Protein
Expression and Purification 33: 123-133 (2004), the disclosures of
which are incorporated by reference in their entireties.
[0232] In certain embodiments, antibodies or fragments thereof that
bind to a marker on cancer stem cells are substantially
non-immunogenic in the treated subject. Methods for obtaining
non-immunogenic antibodies include, but are not limited to,
chimerizing the antibody, humanizing the antibody, and isolating
antibodies from the same species as the subject receiving the
therapy. Antibodies or fragments thereof that bind to markers in
cancer stem cells can be produced using techniques known in the
art. See, for example, paragraphs 539-573 of U.S. Publication No.
2005/0002934 A1, which is incorporated by reference in its
entirety.
[0233] In some embodiments, the therapy comprises the use of
X-rays, gamma rays and other sources of radiation to destroy cancer
stem cells and/or cancer cells. In specific embodiments, the
radiation therapy is administered as external beam radiation or
teletherapy, wherein the radiation is directed from a remote
source. In other embodiments, the radiation therapy is administered
as internal therapy or brachytherapy wherein a radioactive source
is placed inside the body close to cancer stem cells, cancer cells
and/or a tumor mass.
[0234] In some embodiments, the therapy used is a proliferation
based therapy. Non-limiting examples of such therapies include a
chemotherapy and radiation therapy as described supra.
[0235] Currently available therapies and their dosages, routes of
administration and recommended usage are known in the art and have
been described in such literature as the Physician's Desk Reference
(60th ed., 2006).
[0236] In a specific embodiment, cycling therapy involves the
administration of a first cancer therapeutic for a period of time,
followed by the administration of a second cancer therapeutic for a
period of time, optionally, followed by the administration of a
third cancer therapeutic for a period of time and so forth, and
repeating this sequential administration, i.e., the cycle in order
to reduce the development of resistance to one of the cancer
therapeutics, to avoid or reduce the side effects of one of the
cancer therapeutics, and/or to improve the efficacy of the cancer
therapeutics.
[0237] When two prophylactically and/or therapeutically effective
regimens are administered to a subject concurrently, the term
"concurrently" is not limited to the administration of the cancer
therapeutics at exactly the same time, but rather, it is meant that
they are administered to a subject in a sequence and within a time
interval such that they can act together (e.g., synergistically to
provide an increased benefit than if they were administered
otherwise). For example, the cancer therapeutics may be
administered at the same time or sequentially in any order at
different points in time; however, if not administered at the same
time, they should be administered sufficiently close in time so as
to provide the desired therapeutic effect, preferably in a
synergistic fashion. The combination cancer therapeutics can be
administered separately, in any appropriate form and by any
suitable route. When the components of the combination cancer
therapeutics are not administered in the same pharmaceutical
composition, it is understood that they can be administered in any
order to a subject in need thereof. For example, a first
prophylactically and/or therapeutically effective regimen can be
administered prior to (e.g., 5 minutes, 15 minutes, 30 minutes, 45
minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48
hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5
weeks, 6 weeks, 8 weeks, or 12 weeks before), concomitantly with,
or subsequent to (e.g., 5 minutes, 15 minutes, 30 minutes, 45
minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48
hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5
weeks, 6 weeks, 8 weeks, or 12 weeks after) the administration of
the second cancer therapeutic, to a subject in need thereof. In
various embodiments, the cancer therapeutics are administered 1
minute apart, 10 minutes apart, 30 minutes apart, less than 1 hour
apart, 1 hour apart, 1 hour to 2 hours apart, 2 hours to 3 hours
apart, 3 hours to 4 hours apart, 4 hours to 5 hours apart, 5 hours
to 6 hours apart, 6 hours to 7 hours apart, 7 hours to 8 hours
apart, 8 hours to 9 hours apart, 9 hours to 10 hours apart, 10
hours to 11 hours apart, 11 hours to 12 hours apart, no more than
24 hours apart or no more than 48 hours apart. In one embodiment,
the cancer therapeutics are administered within the same office
visit. In another embodiment, the combination cancer therapeutics
are administered at 1 minute to 24 hours apart.
[0238] In a specific embodiment, the combination therapies have the
same mechanism of action. In another specific embodiment, the
combination therapies each have a different mechanism of
action.
Diagnostic System
[0239] The following are sample embodiments of the chemistry
process configuration. They each consist of a plastic cartridge or
glass reaction vessel with reagent dispensed inside. The user will
remove the seal from the cap on the reaction vessel, insert the
high void volume swab with saliva sample into cartridge or glass
reaction vessel and place the assay solution into the portable
fluorescent polarization apparatus for mixing with reagent one and
two for rapid response to determine the level of the constituent
e.g., salivary cortisol.
[0240] The invention will be illustrated in more detail with
reference to the following Examples, but it should be understood
that the present invention is not deemed to be limited thereto.
EXAMPLES
Example 1
[0241] This study is performed to obtain evidence for enhancement
of chemotherapy sensitivity of GR+/ER-/PR-/Her2-neu-breast cancer
cells by blocking stress-mediated cell survival pathways with a GR
antagonist. In the present pilot study, the GR antagonist ORG
34517, is investigated, in one dose, in combination with the
chemotherapeutic agent Taxol. Both the Taxol and ORG 34517 doses
are selected based on previous experiments.
[0242] Materials and Methods--1.times.107 TNBC (Triple Negative
Breast Cancer; MDA-MB-231) cells in 50 ul of PBS (phosphate
buffered saline) are injected into the mammary fat pad of 4-5 week
old CB-17 SCID mice (Taconic Labs). The mice are allowed to form
xenografted breast tumors of approximately 200 mm3 volume. When
tumors reach an average of 200 mm3 (Week "0"), 15 mice are
separated into the 3 treatment groups 1) vehicle, 2) paclitaxel
(Taxol), 3) ORG 34517+paclitaxel. The used paclitaxel dose is 10
mg/kg/day dissolved in Ethanol/Sesame Oil; ORG 34517 is
administered in the maximum soluble dose of 20.5 mg/kg/day
dissolved in Ethanol/Castor Oil. The mice receive intraperitoneal
injections (200 ul) of drugs for 5 consecutive days (day 1-5).
Tumors are measured three times per week. Tumor size is both
expressed as absolute volume and as relative growth in comparison
to baseline (day 0).
[0243] Results--Treatment of the animals is started when the tumor
has a volume of at least 100 mm3. The total experimental group
consisted of 15 animals. In 6 animals the tumor growth was
considered not sufficient. These animals were not treated which
resulted in individual group sizes of N=3. In the control situation
the tumor grows within 13 days with about 800 mm3 (see raw data).
To correct for baseline differences, for each animal the relative
tumor growth is calculated. Relative growth is defined as (tumor
size day x-tumor size day 0)/tumor size day 0. Group averages for
the relative tumor growth are depicted in FIG. 1. In the control
condition (VehNeh) the relative growth is 379%. This growth is
reduced to 183% by taxol treatment (Veh/Taxo). A further reduction
of the growth is induced by the addition of ORG 34517 (517/Taxo).
The relative growth of this latter group is 100%.
[0244] A repeated measures ANOVA did reveal a significant
time-effect (F5,36)=2,73; p=0,035), a significant treatment effect
(F(2,36)=26,86; p=0,000) and a significant interaction
(F(10,36)=2,36; p=0,029). It should be remembered that the data are
not normally distributed and therefore only for descriptive
purposes one-sided Student-t tests were performed and revealed a
significant difference (p<0,05) on day 4 and 11 when comparing
Veh/Veh and Veh/Taxo. Comparing Veh/Taxo and 517/Taxo showed
significances on day 4, 7, 9 and 13. This statistical analysis
should be considered explorative because of the pilot nature of the
study.
[0245] The individual scores for the 517/Taxo treated animals are
depicted in FIG. 2. All three animals show a decreased tumor growth
in comparison to Veh/Taxo treatment (mean scores). In 2 out of 3
animals a tumor shrinkage is observed, during treatment.
[0246] In this pilot study, the glucocorticoid receptor antagonist
ORG 34517 (when administered intraperitoneally in a dose of 20.5
mg/kg/day for 5 days) potentiated the chemotherapeutic effect of
Taxol. While Taxol injections reduced the tumor growth; Taxol+ORG
34517 induced a tumor shrinkage in 2 out of 3 animals, during
treatment.
Example 2
[0247] Mice were implanted with cultured ER-GR+ human breast cancer
cells. As tumor volume in each mouse approached the test threshold
of 200 mm3, the mice were randomized to receive intraperitoneal
injections of vehicle alone, chemotherapy (Paclitaxel) alone, and
chemotherapy and ORG 34517. Each group contained 3 mice. Results
show significant differences in attained tumor volume (see FIG.
10).
Example 3
[0248] In conventional dosage forms, like a capsule or a tablet
(11.beta.,17.beta.)-11-(1,3-benzodioxol-5-yl)-17-hydroxy-17-(1-propynyl)e-
stra-4,9-dien-3-one suffers from a low exposure, high dose
variability, large food effect and non-dose linearity. Another
dosage form than a conventional tablet or capsule may be able to
circumvent these issues.
[0249] The solubility of
(11.beta.,17.beta.)-11-(1,3-benzodioxol-5-yl)-17-hydroxy-17-(1-propynyl)e-
stra-4,9-dien-3-one is determined in different solvents. These
solutions can be of use for clinical treatment with the advantages
as mentioned above.
TABLE-US-00001 TABLE 1 Compound Solubility Acetone >10 mg/mL
Acetonitrile >10 mg/mL Arachis oil 2.46 mg/g Benzylalcohol 390
mg/g Castor oil 7.38 mg/g Castor oil:benzylbenzoate (90:10) 15.0
mg/g Castor oil:propylene glycol laurate (0.55:0.45; v:v) 9.4 mg/g
Corn oil 2.58 mg/g Cremophor EL 23 mg/g Dimethylsulfoxide >100
mg/mL Dog bile:phosphate buffer (pH = 6.5) (1:1) ~0.08 mg/mL
Ethanol >10 mg/mL Ethanol:PEG 400:Propylene glycol (10:50:40)
28.6 mg/mL Ethanol:PEG 400:Propylene glycol:water 14.0 mg/mL
(10:40:40:10) Gelicure 44/14 100 mg/mL Glucofurol:water (50:50) 0.7
mg/mL Glycoferol 118 mg/ml HP.beta.CD, 10% in water 0.207 mg/mL
HP.beta.CD, 20% in water 0.395 mg/mL HP.beta.CD, 40% in water 0.8
mg/mL Medium Chain monoglyceride/glycerol monocaprylate 28 mg/g
Methanol >10 mg/mL Miglyol 812 4.5 mg/g Mulgofen:water (5:95) 2
mg/mL Olive oil 2.26 mg/g PEG 1500 37 mg/mL PEG 400 34.4 mg/mL PEG
400:Water (60:40) 0.0006 mg/mL Polysorbate in water (0.5%) <LOD
Propylene glycol 8.9 mg/mL Propylene glycol 8.9 mg/mL Soy oil 2.70
mg/g Span 80 7 mg/g TPGS:PEG 400:propyleneglycol (20:60:5) 0.9
mg/mL TPGS:PEG 400:propyleneglycol (12:17:55) 0.7 mg/mL
TPGS:d-Alpha Tocopheryl Polyethylene Glycol 1000 1.1 mg/mL
Succinate (20%, as delivered) Water 0.003 mg/mL Water - Sodium
lauryl sulphate (0.5% w/v) 0.240 mg/mL Water - Sodium lauryl
sulphate (1.0% w/v) 0.377 mg/mL
Example 4
[0250] During toxicology studies also attempts were made to
increase the exposure to Org 34517 (6). A rat study showed
interesting results. Wistar rats 5 male and 5 female) were dosed
orally with three different formulations: [0251] a gelatin/mannitol
suspension (173.6 mg/kg) [0252] a solution of Org 34517 in arachis
oil/10% benzyl alcohol (38.4 mg/kg [0253] a dispersion of Org 34517
in Gelucire 44/14 tablets (a lipid based Self Emulsifying Drug
Delivery System, 7.4 mg/tablet, 37 mg/kg). Table 2 shows the dose
normalized results for Cm and AUC for the male and female rats.
Although the data show a large difference between male and female
rats, both the Gelucire and the Arachis oil show a much higher
bioavailability than the gelatin mannitol suspension. It must be
said that the data of arachis oil showed a much larger variation
than the other two formulations (50% versus 10%, not shown
here).
TABLE-US-00002 [0253] TABLE 2 Dose normalized toxicokinetic data of
3 formulations of Org 34517 in rats (M: Male, F: Female). Gelucire
Arachis oil/10% Gelatin 44/14 benzylalcohol Mannitol C.sub.max M
3.1 8.2 0.4 [ng/ml mg Org 34517] C.sub.max F 42.6 16.8 3.5 [ng/ml
mg Org 34517] AUC.sub.0-24 M 30.8 99.3 4.9 [ng hr/ml mg Org 34517]
AUC.sub.0-24 F 427.4 231.8 51.7 [ng hr/ml mg Org 34517]
[0254] The formulations which show a better bioavailability than
gelatin mannitol are thought to increase the solubility of Org
34517 in the GI tract.
[0255] Table 2 shows a finely milled suspension ("nanosuspension")
Study in DOG, doses 5 mg/kg; 15 mg/kg; 45 mg/kg comparison of SD
results. Relative bioavailability compares AUC.sub.0-24. Two dogs
were included in the test. The nanosuspension increases the
exposure of
(11.beta.,17.beta.)-11-(1,3-benzodioxol-5-yl)-17-hydroxy-17-(1-propynyl)e-
stra-4,9-dien-3-one (see FIG. 11).
TABLE-US-00003 TABLE 2 Relative bioavailability in dogs: comparison
of conventional suspension with finely milled suspension.
AUC.sub.0-24 (conventional AUC.sub.0-24 (finely milled Dose
suspension) ng*hr/mL suspension) ng*hr/mL F.sub.rel 5 mg/kg 10105
13288 1.31 15 mg/kg 21687 23095 1.06 45 mg/kg 118493 90536 0.76
[0256] While the invention has been described in detail and with
reference to specific examples thereof, it will be apparent to one
skilled in the art that various changes and modifications can be
made therein without departing from the spirit and scope
thereof.
* * * * *