U.S. patent application number 10/946969 was filed with the patent office on 2005-02-17 for composition and methods for treatment of sexual dysfunction.
This patent application is currently assigned to Palatin Technologies, Inc.. Invention is credited to Bernstein, Joanna K., Blood, Christine H., Herbert, Guy W., Shadiack, Annette M..
Application Number | 20050037951 10/946969 |
Document ID | / |
Family ID | 27385799 |
Filed Date | 2005-02-17 |
United States Patent
Application |
20050037951 |
Kind Code |
A1 |
Blood, Christine H. ; et
al. |
February 17, 2005 |
Composition and methods for treatment of sexual dysfunction
Abstract
Compositions and methods are provided for treatment of sexual
dysfunction in mammals, including male sexual dysfunction, such as
erectile dysfunction, and female sexual dysfunction, wherein
pharmaceutically sufficient quantities of a linear or cyclic
peptide with a carboxyl terminus --OH group or pharmaceutically
acceptable salt thereof and a core His-Phe-Arg-Trp (SEQ ID NO:1) or
His-D-Phe-Arg-Trp sequence are administered. Methods of
administration include injection, oral, nasal and mucosal
administration.
Inventors: |
Blood, Christine H.;
(Morristown, NJ) ; Shadiack, Annette M.;
(Somerset, NJ) ; Herbert, Guy W.; (Fords, NJ)
; Bernstein, Joanna K.; (North Brunswick, NJ) |
Correspondence
Address: |
PEACOCK MYERS AND ADAMS P C
P O BOX 26927
ALBUQUERQUE
NM
871256927
|
Assignee: |
Palatin Technologies, Inc.
Cranbury
NJ
|
Family ID: |
27385799 |
Appl. No.: |
10/946969 |
Filed: |
September 20, 2004 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10946969 |
Sep 20, 2004 |
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10040547 |
Jan 4, 2002 |
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6794489 |
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10040547 |
Jan 4, 2002 |
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09606501 |
Jun 28, 2000 |
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6579968 |
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60142346 |
Jun 29, 1999 |
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60194987 |
Apr 5, 2000 |
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Current U.S.
Class: |
514/10.7 ;
514/21.1 |
Current CPC
Class: |
Y10S 436/811 20130101;
C07K 14/685 20130101; A61K 38/00 20130101; A61P 15/00 20180101;
A61P 15/10 20180101 |
Class at
Publication: |
514/009 ;
514/018 |
International
Class: |
A61K 038/12 |
Foreign Application Data
Date |
Code |
Application Number |
Jun 29, 2000 |
WO |
PCT/US00/18217 |
Claims
What is claimed is:
1. A method for stimulating sexual response in a mammal, comprising
administering a pharmaceutically sufficient amount of a composition
comprising a peptide with a carboxyl terminus --OH group or
pharmaceutically acceptable salt thereof and a core His-Phe-Arg-Trp
(SEQ ID NO:1) or His-D-Phe-Arg-Trp sequence.
2. The method of claim 1, wherein the peptide is a linear
peptide.
3. The method of claim 1, wherein the peptide is a cyclic
peptide.
4. The method of claim 3, wherein the peptide is cyclicized through
a covalent bond comprising two reactive amino acid side chains.
5. The method of claim 1, wherein the mammal is a male.
6. The method of claim 1, wherein the mammal is a female.
7. The method of claim 1, wherein the composition further comprises
a pharmaceutically acceptable carrier.
8. The method of claim 1, wherein administering comprises
administering by a method of administration selected from the group
consisting of administration by injection, administration through
mucous membranes, buccal administration, oral administration,
dermal administration, inhalation administration and nasal
administration.
9. The method of claim 1, wherein administering comprises nasal
administration of a metered amount of a formulation comprising an
aqueous buffer.
10. The method of claim 9, wherein the aqueous buffer is a member
selected from the group consisting of saline and citrate
buffer.
11. A method for stimulating sexual response in a mammal,
comprising administering a pharmaceutically sufficient amount of a
composition comprising a linear peptide with a carboxyl terminus
--OH group or pharmaceutically acceptable salt thereof and
including the sequence His-Phe-Arg-Trp (SEQ ID NO:1) or
His-D-Phe-Arg-Trp.
12. The method of claim 11, wherein the mammal is a male.
13. The method of claim 11, wherein the mammal is a female.
14. The method of claim 11, wherein the composition further
comprises a pharmaceutically acceptable carrier.
15. The method of claim 11, wherein administering comprises
administering by a method of administration selected from the group
consisting of administration by injection, administration through
mucous membranes, buccal administration, oral administration,
dermal administration, inhalation administration and nasal
administration.
16. The method of claim 11, wherein administering comprises nasal
administration of a metered amount of a formulation comprising an
aqueous buffer.
17. The method of claim 16, wherein the aqueous buffer is a member
selected from the group consisting of saline and citrate
buffer.
18. A method for stimulating sexual response in a mammal,
comprising administering a pharmaceutically sufficient amount of a
composition comprising a cyclic peptide with a carboxyl terminus
--OH group or pharmaceutically acceptable salt thereof and
including the sequence His-Phe-Arg-Trp (SEQ ID NO:1) or
His-D-Phe-Arg-Trp sequence.
19. The method of claim 18, wherein the cyclic peptide is
cyclicized through a covalent bond comprising two reactive amino
acid side chains.
20. The method of claim 18, wherein the mammal is a male.
21. The method of claim 18, wherein the mammal is a female.
22. The method of claim 18, wherein the composition further
comprises a pharmaceutically acceptable carrier.
23. The method of claim 18, wherein administering comprises
administering by a method of administration selected from the group
consisting of administration by injection, administration through
mucous membranes, buccal administration, oral administration,
dermal administration, inhalation administration and nasal
administration.
24. The method of claim 18, wherein administering comprises nasal
administration of a metered amount of a formulation comprising an
aqueous buffer.
25. The method of claim 24, wherein the aqueous buffer is a member
selected from the group consisting of saline and citrate buffer.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application is a continuation application of U.S.
patent application Ser. No. 10/040,547, now U.S. Pat. No.
6,794,489, entitled Compositions and Methods for Treatment of
Sexual Dysfunction, which application was filed on Jan. 4, 2002 and
issued as a patent on Sep. 21, 2004, which is a
continuation-in-part application of U.S. patent application Ser.
No. 09/606,501, now U.S. Pat. No. 6,579,968, entitled Composition
and Methods for Treatment of Sexual Dysfunction, which application
was filed on Jun. 28, 2000 and issued as a patent on Jun. 17, 2003,
and the specification thereof of each is incorporated herein by
reference.
[0002] This application claims the benefit of the filing of U.S.
Provisional Patent Application Ser. 60/142,346, entitled
Compositions And Methods For Treatment Of Sexual Dysfunction, filed
on Jun. 29, 1999, and U.S. Provisional Patent Application Ser. No.
60/194,987, entitled Compositions And Nasal Delivery Methods For
Treatment Of Sexual Dysfunction, filed on Apr. 5, 2000, and the
specification thereof of each is incorporated herein by
reference.
BACKGROUND OF THE INVENTION
[0003] 1. Field of the Invention (Technical Field)
[0004] The present invention relates to peptide constructs for the
treatment of sexual dysfunction in animals, including both male
erectile dysfunction and female sexual dysfunction, including
methods and formulations for the use and administration of the
same.
[0005] 2. Background Art
[0006] Note that the following discussion refers to a number of
publications by author(s) and year of publication, and that due to
recent publication dates certain publications are not to be
considered as prior art vis-a-vis the present invention. Discussion
of such publications herein is given for more complete background
and is not to be construed as an admission that such publications
are prior art for patentability determination purposes.
[0007] Sexual dysfunction, including both penile erectile
dysfunction or impotence and female sexual dysfunction, are common
medical problems. Significant effort has been devoted over the last
twenty or more years to develop methods, devices and compounds for
treatment of sexual dysfunction. While more effort has been
undertaken for treatment of penile erectile dysfunction, female
sexual dysfunction is also an area to which significant research
and effort has been devoted.
[0008] At present, one commonly used orally administered drug for
treatment of sexual dysfunction in the male is Viagra.RTM., a brand
of sildenafil, which is a phosphodiesterase 5 inhibitor, increasing
the persistence of cyclic guanosine monophosphate and thereby
enhancing erectile response. There are several other medical
treatment alternatives currently available depending on the nature
and cause of the impotence problem. Some men have abnormally low
levels of the male hormone testosterone, and treatment with
testosterone injections or pills may be beneficial. However,
comparatively few impotent men have low testosterone levels. For
many forms of erectile dysfunction, treatment may be undertaken
with drugs injected directly into the penis, including drugs such
as papaverin, prostaglandin E.sub.1, phenoxybenzamine or
phentolamine. These all work primarily by dilating the arterial
blood vessels and decreasing the venous drainage. Urethral inserts,
such as with suppositories containing prostaglandin, may also be
employed. In addition, a variety of mechanical aids are employed,
including constriction devices and penile implants.
[0009] A variety of treatments have also been explored for female
sexual dysfunction, including use of sildenafil, although the Food
and Drug Administration has not specifically approved such use.
Testosterone propionate has also been employed to increase or
augment female libido.
[0010] Melanocortin receptor-specific compounds have been explored
for use of treatment of sexual dysfunction. In one report, a cyclic
.alpha.-melanocyte-stimulating hormone (".alpha.-MSH") analog,
called Melanotan-II, was evaluated for erectogenic properties for
treatment of men with psychogenic erectile dysfunction. Wessells H.
et al., J Urology 160:389-393 (1998); see also U.S. Pat. No.
5,576,290, issued Nov. 19, 1996 to M. E. Hadley, entitled
Compositions and Methods for the Diagnosis and Treatment of
Psychogenic Erectile Dysfunction and U.S. Pat. No. 6,051,555,
issued Apr. 18, 2000, also to M. E. Hadley, entitled Stimulating
Sexual Response in Females. The peptides used in U.S. Pat. Nos.
5,576,290 and 6,051,555 are also described in U.S. Pat. No.
5,674,839, issued Oct. 7, 1997, to V. J. Hruby, M. E. Hadley and F.
Al-Obeidi, entitled Cyclic Analogs of Alpha-MSH Fragments, and in
U.S. Pat. No. 5,714,576, issued Feb. 3, 1998, to V. J. Hruby, M. E.
Hadley and F. Al-Obeidi, entitled Linear Analogs of Alpha-MSH
Fragments. Melanotan-II is a peptide of the following formula:
1
[0011] Additional related peptides are disclosed in U.S. Pat. Nos.
5,576,290, 5,674,839, 5,714,576 and 6,051,555. These peptides are
described as being useful for both the diagnosis and treatment of
psychogenic sexual dysfunction in males and females. These peptides
are related to the structure of melanocortins.
[0012] In use of Melanotan-II, significant erectile responses were
observed, with 8 of 10 treated men developing clinically apparent
erections, and with a mean duration of tip rigidity greater than
80% for 38 minutes with Melanotan-II compared to 3.0 minutes with a
placebo (p=0.0045). The drug was administered by subcutaneous
abdominal wall injection, at doses ranging from 0.025 to 0.157
mg/kg body weight. Transient side effects were observed, including
nausea, stretching and yawning, and decreased appetite.
[0013] The minimum peptide fragment of native .alpha.-MSH needed
for erectile response is the central tetrapeptide sequence,
His.sup.6-Phe.sup.7-Arg.sup.8-Trp.sup.9 (SEQ ID NO:1). In general,
all melanocortin peptides share the same active core sequence,
His-Phe-Arg-Trp (SEQ ID NO:1), including melanotropin neuropeptides
and adrenocorticotropin. Five distinct melanocortin receptor
subtypes have been identified, called MC1-R through MC5-R, and of
these MC3-R and MC4-R are believed to be expressed in the human
brain. MC3-R has the highest expression in the arcuate nucleus of
the hypothalamus, while MC4-R is more widely expressed in the
thalamus, hypothalamus and hippocampus. A central nervous system
mechanism for melanocortins in the induction of penile erection has
been suggested by experiments demonstrating penile erection
resulting from central intracerebroventricular administration of
melanocortins in rats. While the mechanism of His-Phe-Arg-Trp (SEQ
ID NO:1) induction of erectile response has not been fully
elucidated, it has been hypothesized that it involves the central
nervous system, and probably binding to MC3-R and/or MC4-R.
[0014] Other peptides and constructs have been proposed which are
ligands that alter or regulate the activity of one or more
melanocortin receptors. For example, International Patent
Application No. PCT/US99/09216, entitled Isoquinoline Compound
Melanocortin Receptor Ligands and Methods of Using Same, discloses
two compounds that induce penile erections in rats. However, these
compounds were administered by injection at doses of 1.8 mg/kg and
3.6 mg/kg, respectively, and at least one compound resulted in
observable side effects, including yawning and stretching. Other
melanocortin receptor-specific compounds with claimed application
for treatment of sexual dysfunction are disclosed in International
Patent Application No. PCT/US99/13252, entitled Spiropiperidine
Derivatives as Melanocortin Receptor Agonists.
[0015] Both cyclic and linear .alpha.-MSH peptides have been
studied; however, the peptides heretofore evaluated have had an
amide or --NH.sub.2 group at the carboxyl terminus. See, for
example, Wessells H. et al., J Urology, cited above;
Haskell-Luevano C. et al., J Med Chem 40:2133-39 (1997); Schioth H.
B. et al., Brit J Pharmacol 124:75-82 (1998); Schioth H. B. et al.,
Eur J Pharmacol 349:359-66 (1998); Hadley M. E. et al., Pigment
Cell Res 9:213-34 (1996); Bednarek M. A. et al., Peptides 20:401-09
(1999); U.S. Pat. Nos. 6,054,556, 6,051,555 and 5,576,290; and,
International Patent Applications PCT/US99/04111 and
PCT/US98/03298. While significant research has been conducted in an
effort to determine the optimal structure of .alpha.-MSH peptides,
including a variety of structure-function, agonist-antagonist,
molecular modeling and pharmacophore studies, such studies have
relied upon peptides with an art conventional --NH.sub.2 group at
the carboxyl terminus. Further, it has long been believed that
biologically active neuropeptides, including .alpha.-MSH peptides,
are amidated, with an --NH.sub.2 group at the carboxyl terminus,
and that such amidation is required both for biological activity
and stability. See, for example, Metabolism of Brain Peptides, Ed.
G. O'Cuinn, CRC Press, New York, 1995, pp.1-9 and 99-101.
SUMMARY OF THE INVENTION (DISCLOSURE OF THE INVENTION)
[0016] The invention relates to a peptide that is a free acid or
pharmaceutically acceptable salt thereof that includes the sequence
His-Phe-Arg-Trp (SEQ ID NO:1), His-D-Phe-Arg-Trp, homologs of
His-Phe-Arg-Trp (SEQ ID NO:1) or homologs of His-D-Phe-Arg-Trp. The
peptide is preferably a cyclic peptide, and preferable has a
terminal --OH at the carboxyl terminus. In a preferred embodiment,
the peptide is Ac-Nle-cyclo(-Asp-His-D-Phe-Arg-Trp-Lys)-OH.
[0017] The invention also includes pharmaceutical compositions of
matter, including a peptide of this invention and a
pharmaceutically acceptable carrier. The pharmaceutically
acceptable carrier may be a buffered aqueous carrier, and
preferably a saline or citrate buffered carrier.
[0018] The peptide of this invention, and pharmaceutical
compositions of this invention, may be used for stimulating sexual
response in a mammal. The invention thus also includes a method for
stimulating sexual response in a mammal, in which a
pharmaceutically sufficient amount of a composition including
His-Phe-Arg-Trp (SEQ ID NO:1), His-D-Phe-Arg-Trp, homologs of
His-Phe-Arg-Trp (SEQ ID NO:1) or homologs of His-D-Phe-Arg-Trp is
administered. In a preferred embodiment, the composition includes a
peptide or pharmaceutically acceptable salt thereof of the formula
Ac-Nle-cyclo(-Asp-His-D-Phe-Arg-Trp-Lys)-OH. The mammal may be male
or female. In this method, the composition can also include a
pharmaceutically acceptable carrier. The peptide or pharmaceutical
composition may be administered by any means known in the art,
including administration by injection, administration through
mucous membranes, buccal administration, oral administration,
dermal administration, inhalation administration and nasal
administration. In a preferred embodiment, administration is by
nasal administration of a metered amount of a formulation including
an aqueous buffer, which buffer may be a saline or citrate
buffer.
[0019] A primary object of the present invention is a melanocortin
receptor-specific pharmaceutical for use in treatment of sexual
dysfunction.
[0020] A second object is to provide a peptide-based melanocortin
receptor-specific pharmaceutical for use in treatment of male
sexual dysfunction, including erectile dysfunction.
[0021] Yet another object is to provide a peptide-based
melanocortin receptor-specific pharmaceutical for use in treatment
of female sexual dysfunction.
[0022] Yet another object is to provide a peptide-based
melanocortin receptor-specific pharmaceutical for use in treatment
of sexual dysfunction with substantially reduced incidence of
undesirable side effects.
[0023] A primary advantage of the present invention is that it is
efficacious at doses that do not cause deleterious side effects,
such side effects including nausea, yawning, stretching, decreased
appetite and other effects observed with Melanotan-II, or that
causes decreased deleterious side effects as compared to
Melanotan-II.
[0024] A second advantage of the present invention is that it
provides compositions with a larger therapeutic window between
desired therapeutic effects and the onset of undesired side effects
than other melanocortin receptor-specific agents for the intended
purpose.
[0025] Yet another advantage of the present invention is that it
provides compositions with a greater safety margin between desired
therapeutic effects and the onset of undesired side effects than
other melanocortin receptor-specific agents for the intended
purpose.
[0026] Yet another advantage of the present invention is that it
provides a peptide-based melanocortin receptor-specific
pharmaceutical that is effective over a greater dose range, without
deleterious side effects, than Melnotan-II or other melanocortin
receptor-specific agents.
[0027] Yet another advantage of the present invention is that it
provides a peptide-based melanocortin receptor-specific
pharmaceutical for use in treatment of sexual dysfunction which is
pharmaceutically active more rapidly following administration than
Melanotan-II or other peptide-based melanocortin receptor-specific
agents.
[0028] Yet another advantage of the present invention is that it
provides a peptide-based melanocortin receptor-specific
pharmaceutical for use in treatment of sexual dysfunction which,
because of increased efficacy at low doses, may be administered by
delivery systems other than art conventional intravenous,
subcutaneous or intramuscular injection, including but not limited
to nasal delivery systems and mucous membrane delivery systems.
[0029] Other objects, advantages and novel features, and further
scope of applicability of the present invention will be set forth
in part in the detailed description to follow, taken in conjunction
with the accompanying drawings, and in part will become apparent to
those skilled in the art upon examination of the following, or may
be learned by practice of the invention. The objects and advantages
of the invention may be realized and attained by means of the
instrumentalities and combinations particularly pointed out in the
appended claims.
BRIEF DESCRIPTION OF THE DRAWINGS
[0030] The accompanying drawings, which are incorporated into and
form a part of the specification, illustrate several embodiments of
the present invention and, together with the description, serve to
explain the principles of the invention. The drawings are only for
the purpose of illustrating a preferred embodiment of the invention
and are not to be construed as limiting the invention. In the
drawings:
[0031] FIG. 1 is a plot of the pharmacokinetic profile in rats
intravenously administered 100 .mu.g/kg body weight of Compound
1;
[0032] FIG. 2 is a plot of the pharmacokinetic profile in monkeys
intravenously administered 50 .mu.g/kg body weight of Compound
1;
[0033] FIG. 3 is a plot of the pharmacokinetic profile in a monkey
intranasally administered 50 .mu.g/kg body weight of Compound
1;
[0034] FIG. 4 is a plot of the intranasal efficacy in rats of
different amounts of Compound 1;
[0035] FIG. 5 is a plot of pharmacokinetic profile in Beagle dogs
intranasally administered 30, 150 and 650 .mu.g/kg body weight of
Compound 1;
[0036] FIG. 6 is a graph of solicitations in fully primed (estrogen
and progesterone) female rats following administration of 0, 50,
100 and 200 .mu.g/kg body weight of Compound 1;
[0037] FIG. 7 is a graph of solicitations in partially primed
(estrogen alone) female rats following administration of 0and 200
.mu.g/kg body weight of Compound 1;
[0038] FIG. 8A is a graph of solicitations in fully primed female
rats following administration of 0, 50, 100 and 200 .mu.g/kg body
weight of Compound 1, where ANOVA P<0.001 for the
experiment;
[0039] FIG. 8B is a graph of lordosis magnitude in fully primed
female rats following administration of 0, 50, 100 and 200 .mu.g/kg
body weight of Compound 1;
[0040] FIG. 8C is a graph of hops and darts in fully primed female
rats following administration of 0, 50, 100 and 200 .mu.g/kg body
weight of Compound 1, where ANOVA P<0.02 for the experiment;
and
[0041] FIG. 8D is a graph of return latency, in seconds, after
ejaculation in fully primed female rats following administration of
0, 50, 100 and 200 .mu.g/kg body weight of Compound 1, where ANOVA
P<0.02 for the experiment.
DESCRIPTION OF THE PREFERRED EMBODIMENTS
(BEST MODES FOR CARRYING OUT THE INVENTION)
[0042] In the listing of compounds according to the present
invention, the amino acid residues have their conventional meaning
as given in Chapter 2400 of the Manual of Patent Examining
Procedure, 7.sup.th Ed. Thus, "Nle" is norleucine; "Asp" is
aspartic acid; "His" is histidine; "D-Phe" is D-phenylalanine;
"Arg" is arginine; "Trp" is tryptophan; "Lys" is lysine; "Gly" is
glycine; "Pro" is proline; "Tyr" is tyrosine, and "Ser" is serine.
"Ac" refers to a peptide or amino acid sequence that is
acetylated.
[0043] The invention provides deamidated .alpha.-MSH peptides,
which are peptides that include the core .alpha.-MSH sequence
His-Phe-Arg-Trp (SEQ ID NO:1), His-D-Phe-Arg-Trp, or homologs or
analogs of either of the foregoing, in which the peptide is
deamidated, which is to say that it does not include an --NH.sub.2
group at the carboxyl terminus. In a preferred embodiment, the
deamidated .alpha.-MSH peptides of this invention have an --OH
group at the carboxyl terminus, and are thus a free acid form of
peptide.
[0044] Peptides with a His-D-Phe-Arg-Trp core .alpha.-MSH sequence
are described in U.S. Pat. Nos. 5,576,290, 5,674,839, 5,714,576 and
6,051,555, disclosed above, and include the following
representative linear peptides:
1 Ac-Ser-Tyr-Ser-Nle-Glu-His-D-Phe-Arg-Trp-
Lys-Gly-Pro-Val-NH.sub.2 Ac-Ser-Tyr-Ser-Nle-Asp-His-D-Phe-
-Arg-Trp- Lys-Gly-Pro-Val-NH.sub.2
Ac-Nle-Glu-His-D-Phe-Arg-Trp-Lys-Gly-Pro-Val-NH.sub.2
Ac-Nle-Asp-His-D-Phe-Arg-Trp-Lys-Gly-Pro-Val-NH.sub.2
Ac-Nle-Asp-His-D-Phe-Arg-Trp-Gly-NH.sub.2
Ac-Nle-Glu-His-D-Phe-Arg-Trp-Lys-NH.sub.2
Ac-Nle-Asp-His-D-Phe-Arg-Trp-Lys-NH.sub.2
Ac-Nle-Glu-His-D-Phe-Arg-Trp-Orn-NH.sub.2
Ac-Nle-Asp-His-D-Phe-Arg-Trp-Orn-NH.sub.2
Ac-Nle-Glu-His-D-Phe-Arg-Trp-Dab-NH.sub.2
Ac-Nle-Asp-His-D-Phe-Arg-Trp-Dab-NH.sub.2
Ac-Nle-Asp-His-D-Phe-Arg-Trp-Dpr-NH.sub.2
[0045] Cyclic peptides with a His-D-Phe-Arg-Trp core .alpha.-MSH
sequence are similarly described in the U.S. Pat. Nos. 5,576,290,
5,674,839, 5,714,576 and 6,051,555, disclosed above, and include
the following representative cyclic peptides:
2 Ac-Nle-cyclo(-Glu-His-D-Phe-Arg-Trp-Lys)- Gly-Pro-Val-NH.sub.2
Ac-Nle-cyclo(-Glu-His-D-Phe-Arg-Tr- p-Lys)-NH.sub.2
Ac-Nle-cyclo(-Asp-His-D-Phe-Arg-Trp-Lys)- -NH.sub.2
Ac-Nle-cyclo(-Asp-His-D-Phe-Arg-Trp-Orn)-NH.su- b.2
Ac-Nle-cyclo(-Asp-His-D-Phe-Arg-Trp-Dab)-NH.sub.2
Ac-Nle-cyclo(-Asp-His-D-Phe-Arg-Trp-Dpr)-NH.sub.2
Ac-Ser-Tyr-Ser-Nle-cyclo(-Asp-His-D-Phe-
Arg-Trp-Lys)-Gly-Pro-Val-NH.sub.2
Ac-Ser-Tyr-Ser-Nle-cyclo(-Asp-His-D-Phe- Arg-Trp-Lys)-NH.sub.2
Ac-Tyr-Ser-Nle-cyclo(-Asp-His-D-P- he-Arg- Trp-Lys)-NH.sub.2
Ac-Ser-Nle-cyclo(-Asp-His-D-Phe-Arg-Trp- Lys)-NH.sub.2
Ac-Nle-cyclo(-Asp-His-D-Phe-Arg-Trp-Lys)-NH.sub.2
Ac-Nle-cyclo(-Asp-His-D-Phe-Arg-Trp-Lys)- Gly-NH.sub.2
Ac-Nle-cyclo(-Asp-His-D-Phe-Arg-Trp-Lys)- Gly-Pro-NH.sub.2
Ac-Nle-cyclo(-Asp-His-D-Phe-Arg-Trp-Lys)- Gly-Pro-Val-NH.sub.2
Ac-Ser-Nle-cyclo(-Asp-His-D-Phe-Ar- g-Trp-
Lys)-Gly-Pro-Val-NH.sub.2 Ac-Thr-Ser-Nle-cyclo(-Asp-His-D-Phe-Arg-
Trp-Lys)-NH.sub.2 Ac-Ser-Thr-Ser-Nle-cyclo(-Asp-His-D-P- he-
Arg-Trp-Lys)-NH.sub.2
[0046] In the forgoing list, Orn is ornithine, Dab is 2,4
diaminobutyric acid, and Dpr is 2,3 diaminopropionic acid.
[0047] Peptides with a His-D-Phe-Arg-Trp core .alpha.-MSH sequence
are disclosed in International Patent Application PCT/US98/03298,
disclosed above, and include the following representative linear
peptides:
3 Ac-Nle-Gln-His-(p(l)-D-Phe)-Arg-(D-Trp)-Gly-NH.sub.2
Ac-Nle-Gln-His-(p(.sup.125l)-D-Phe)-Arg-(D-Trp)-Gly-NH.sub.2
[0048] In a preferred embodiment, the invention provides the
peptide
4 Ac-Nle-cyclo(-Asp-His-D-Phe-Arg-Trp- Compound 1 Lys)-OH
[0049] The peptide of Compound 1 has a formula of
C.sub.50H.sub.68N.sub.14- O.sub.10, and a net molecular weight of
1025.18. This peptide may be synthesized by solid-phase means and
purified to greater than 96% purity by HPLC, yielding a white
powder that is a clear, colorless solution in water. The structure
of Compound 1 is: 2
[0050] In general, the peptide compounds of this invention may be
synthesized by solid-phase synthesis and purified according to
methods known in the art. Any of a number of well-known procedures
utilizing a variety of resins and reagents may be used to prepare
the compounds of this invention.
[0051] The peptides of this invention may be in the form of any
pharmaceutically acceptable salt. Acid addition salts of the
compounds of this invention are prepared in a suitable solvent from
the peptide and an excess of an acid, such as hydrochloric,
hydrobromic, sulfuric, phosphoric, acetic, trifluoroacetic, maleic,
succinic or methanesulfonic. The acetate salt form is especially
useful. Where the compounds of this invention include an acidic
moiety, suitable pharmaceutically acceptable salts may include
alkali metal salts, such as sodium or potassium salts, or alkaline
earth metal salts, such as calcium or magnesium salts.
[0052] The invention provides a pharmaceutical composition that
includes a peptide of this invention and a pharmaceutically
acceptable carrier. The carrier may be a liquid formulation, and is
preferably a buffered, isotonic, aqueous solution. Pharmaceutically
acceptable carriers also include excipients, such as diluents,
carriers and the like, and additives, such as stabilizing agents,
preservatives, solubilizing agents, buffers and the like, as
hereafter described.
[0053] Routes of Administration. Because, in part, of the increased
potency of Compound 1 and the other peptides of this invention,
these peptides may be administered by means other than by
injection. If it is administered by injection, the injection may be
intravenous, subcutaneous, intramuscular, intraperitoneal or other
means known in the art. The peptides of this invention may be
formulated by any means known in the art, including but not limited
to formulation as tablets, capsules, caplets, suspensions, powders,
lyophilized preparations, suppositories, ocular drops, skin
patches, oral soluble formulations, sprays, aerosols and the like,
and may be mixed and formulated with buffers, binders, excipients,
stabilizers, anti-oxidants and other agents known in the art. In
general, any route of administration by which the peptides of
invention are introduced across an epidermal layer of cells may be
employed. Administration means may include administration through
mucous membranes, buccal administration, oral administration,
dermal administration, inhalation administration, nasal
administration and the like. The dosage for treatment of male
erectile dysfunction is administration, by any of the foregoing
means or any other means known in the art, of an amount sufficient
to bring about an erection of the penis in a male. The dosage for
treatment of female sexual dysfunction is administration, by any of
the foregoing means or any other means known in the art, of an
amount sufficient to bring about the desired response.
[0054] The peptides of this invention may be formulated or
compounded into pharmaceutical compositions that include at least
one peptide of this invention together with one or more
pharmaceutically acceptable carriers, including excipients, such as
diluents, carriers and the like, and additives, such as stabilizing
agents, preservatives, solubilizing agents, buffers and the like,
as may be desired. Formulation excipients may include
polyvinylpyrrolidone, gelatin, hydroxy cellulose, acacia,
polyethylene glycol, manniton, sodium chloride or sodium citrate.
For injection or other liquid administration formulations, water
containing at least one or more buffering constituents is
preferred, and stabilizing agents, preservatives and solubilizing
agents may also be employed. For solid administration formulations,
any of a variety of thickening, filler, bulking and carrier
additives may be employed, such as starches, sugars, fatty acids
and the like. For topical administration formulations, any of a
variety of creams, ointments, gels, lotions and the like may be
employed. For most pharmaceutical formulations, non-active
ingredients will constitute the greater part, by weight or volume,
of the preparation. For pharmaceutical formulations, it is also
contemplated that any of a variety of measured-release,
slow-release or time-release formulations and additives may be
employed, so that the dosage may be formulated so as to effect
delivery of a peptide of this invention over a period of time.
[0055] In general, the actual quantity of peptides of this
invention administered to a patient will vary between fairly wide
ranges depending upon the mode of administration, the formulation
used, and the response desired.
[0056] Nasal or Intrapulmonary Administration. By "nasal
administration" is meant any form of intranasal administration of
any of the peptides of this invention. The peptides may be in an
aqueous solution, such as a solution including saline, citrate or
other common excipients or preservatives. The peptides may also be
in a dry or powder formulation.
[0057] In an alternative embodiment, peptides of this invention may
be administered directly into the lung. Intrapulmonary
administration may be performed by means of a metered dose inhaler,
a device allowing self-administration of a metered bolus of a
peptide of this invention when actuated by a patient during
inspiration.
[0058] The peptides of this invention may be formulated with any of
a variety of agents that increase effective nasal absorption of
drugs, including peptide drugs. These agents should increase nasal
absorption without unacceptable damage to the mucosal membrane.
U.S. Pat. Nos. 5,693,608, 5,977,070 and 5,908,825, among others,
teach a number of pharmaceutical compositions that may be employed,
including absorption enhancers, and the teachings of each of the
foregoing, and all references and patents cited therein, are
incorporated by reference.
[0059] If in an aqueous solution, the peptide may be appropriately
buffered by means of saline, acetate, phosphate, citrate, acetate
or other buffering agents, which may be at any physiologically
acceptable pH, generally from about pH 4 to about pH 7. A
combination of buffering agents may also be employed, such as
phosphate buffered saline, a saline and acetate buffer, and the
like. In the case of saline, a 0.9% saline solution may be
employed. In the case of acetate, phosphate, citrate, acetate and
the like, a 50 mM solution may be employed. In addition to
buffering agents, a suitable preservative may be employed, to
prevent or limit bacteria and other microbial growth. One such
preservative that may be employed is 0.05% benzalkonium
chloride.
[0060] It is also possible and contemplated that the peptide may be
in a dried and particulate form. In a preferred embodiment, the
particles are between about 0.5 and 6.0 .mu.m, such that the
particles have sufficient mass to settle on the lung surface, and
not be exhaled, but are small enough that they are not deposited on
surfaces of the air passages prior to reaching the lung. Any of a
variety of different techniques may be used to make dry powder
microparticles, including but not limited to micro-milling, spray
drying and a quick freeze aerosol followed by lyophilization. With
micro-particles, the peptides may be deposited to the deep lung,
thereby providing quick and efficient absorption into the
bloodstream. Further, with such approach penetration enhancers are
not required, as is sometimes the case in transdermal, nasal or
oral mucosal delivery routes. Any of a variety of inhalers can be
employed, including propellant-based aerosols, nebulizers, single
dose dry powder inhalers and multidose dry powder inhalers. Common
devices in current use include metered dose inhalers, which are
used to deliver medications for the treatment of asthma, chronic
obstructive pulmonary disease and the like. Preferred devices
include dry powder inhalers, designed to form a cloud or aerosol of
fine powder with a particle size that is always less than about 6.0
.mu.m. One type of dry powder inhaler in current use is Glaxo's
Rotahaler.TM., which dispenses a unit dose of powder into a tube,
and employs patient suction for inhalation of the powder. Other,
more advanced and preferred dry powder inhalers have been or are in
development, which include propellants and the like.
[0061] Microparticle size, including mean size distribution, may be
controlled by means of the method of making. For micro-milling, the
size of the milling head, speed of the rotor, time of processing
and the like control the microparticle size. For spray drying, the
nozzle size, flow rate, dryer heat and the like control the
microparticle size. For making by means of quick freeze aerosol
followed by lyophilization, the nozzle size, flow rate,
concentration of aerosoled solution and the like control the
microparticle size. These parameters and others may be employed to
control the microparticle size.
[0062] In one preferred embodiment, a dry powder inhaler is
employed which includes a piezoelectric crystal that deaggregates a
dry powder dose, creating a small powder "cloud." Once the powder
cloud is generated, an electricostatically charged plated above the
powder cloud lifts the drug into the air stream. The user with one
relatively easy breath can then inhale the powder. The device may
be breath activated, utilizing a flow sensor that activates the
electronic components upon the start of inhalation, and thereby
eliminating the need for coordination of activation and breathing
rhythms by the user.
[0063] Induction of Penile Response. Both Compound 1 and
Melanotan-II induce penile erections in experimental rat models
when administered by intravenous routes and by other routes,
including nasal administration. However, Compound 1 is
approximately 100-fold more potent than Melanotan-II for inducing
penile erection in the rat model. In intravenous dosing studies,
penile erection was induced by Compound 1 in a broad range of
concentrations, from 0.5 .mu.g/kg of body weight to 25 .mu.g/kg of
body weight. At intravenous doses of 2 .mu.g/kg of body weight,
Compound 1 resulted in penile erection in 100% of rats tested, with
approximately 50% of the rats having multiple erections during a 30
minutes observation period. By comparison, the optimal efficacious
dose intravenously of Melanotan-II in the same rat model was 100
.mu.g/kg of body weight.
[0064] The therapeutic window (the range from the desired
therapeutic effect to observed adverse effects) for Compound 1 is
on the order of >1,000-fold, compared to a 3- to 4-fold
therapeutic window for Melanotan-II. That is, the optimal
efficacious dose of Melanotan-II is approximately one-third to
one-fourth the dose that causes observed adverse effects. For
Compound 1, the optimal efficacious dose is less than one-one
thousandth the dose that causes observed adverse effects. The
significantly greater therapeutic window for Compound 1
demonstrates that it is better tolerated than Melanontan-II.
[0065] The potency of Compound 1 is significantly higher than that
of Melanotan-II, meaning that significantly less Compound 1 is
required for a desired effect, as compared to Melanotan-II. For
example, 2 .mu.g/kg of body weight of Compound 1 resulted in penile
response by intravenous injection in rats, while 100 .mu.g/kg of
body weight of Melanotan-II was required for an equivalent penile
response. The significantly higher potency results in less product
being required. It further permits utilization of alternative
delivery routes, including dermal, nasal and similar delivery
routes, wherein higher quantities of drug may be required in order
to achieve the desired effect. For example, intranasal routes of
administration typically have a bioavailability substantially less
than that achieved with intravenous dosing, and thus more drug must
be administered by intranasal routes in order to achieve the
equivalent pharmacological response.
[0066] Significant commercial advantage for Compound 1 is
demonstrated by the reduced effective dose, compared to
Melanotan-II, as a treatment or diagnostic for erectile
dysfunction. Among the potential commercial advantages of Compound
1 are improved effectiveness; lower costs of the product; improved
convenience, particularly in that the lower dose permits
non-conventional delivery methods that are not feasible with
Melanotan-II doses; and improved safety with a decreased risk of
side-effects.
[0067] Industrial Applicability:
[0068] The invention is further illustrated by the following
non-limiting examples.
EXAMPLE 1
Peptide Synthesis
[0069] The peptide Ac-Nle-cyclo(-Asp-His-D-Phe-Arg-Trp-Lys)-OH was
synthesized by standard solid phase peptide synthesis methods, and
is a cyclic heptapeptide melanocortin peptide analog with a free
acid at the carboxyl terminus and an acetylated amino group at the
amino terminus, with the structure: 3
[0070] The peptide has a net molecular weight of 1025.18, and is
supplied in an acetate salt form. The peptide is a white, odorless
amorphous hygroscopic powder, soluble in 0.9% saline, composed of
C.sub.50H.sub.68N.sub.14O.sub.10. For synthesis, an
Fmoc-Lys(R.sup.3)-p-alkoxybenzyl alcohol resin was transferred to a
solid phase peptide synthesizer reactor with a mechanical stirrer,
where the R.sup.3 group was a group such as
1-(1'-adamantyl)-1-methyl-ethoxycarbony- l (Adpoc),
allyloxycarbonyl (Aloc) or 4-methyltrityl (Mtt). The Fmoc group was
removed and the next Fmoc-protected amino acid (Fmoc-Trp(Boc)-OH)
was added to the resin through standard coupling procedures. The
Fmoc protective group was removed and the remaining amino acids
added individually in the correct sequence, by repeating coupling
and deprotection procedures until the amino acid sequence was
completed. After completion of coupling with the last Fmoc-amino
acid derivative, Fmoc-Nle-OH, and cleavage of the Fmoc protective
group, the exposed terminal amino group was acetylated with acetic
anhydride and pyridine in N,N-dimethylformamide (DMF). The Lys and
Asp protective groups were then cleaved. The Lys and Asp
deprotected peptide resin was suspended in a suitable solvent, such
as DMF, dichloromethane (DCM) or 1-methyl-2-pyrrolidone (NMP), a
suitable cyclic coupling reagent, such as
2-(1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluronium
tetrafluoroborate (TBTU),
2-(7-aza-1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluronium
tetrafluoroborate (TATU),
2-(2-oxo-1(2H)-pyridyl)-1,1,3,3-tetramethyluron- ium
tetrafluoroborate (TBTU) or
N,N'-dicyclohexylcarbodiimide/1-hydroxyben- zotriazole (DCCI/HOBt)
was added, and coupling initiated by use of a suitable base, such
as N,N-diispropylethylamine (DIPEA), sym-collidine or
N-methylmorpholine (NM M). After cyclization, the peptide-resin was
washed and the peptide cleaved from the resin and any remaining
protective groups using trifluoroacetic acid (TFA) in the presence
of water and 1,2-ethanedithiol (EDT). The final product was
precipitated by adding cold ether and collected by filtration.
Final purification was by reversed phase HPLC using a C.sub.18
column. The purified peptide was converted to acetate salt by
passage through an ion-exchange column.
EXAMPLE 2
Binding of Compound 1 and Melanotan-II to Melanocortin
Receptors
[0071] The binding of Compound 1, made as in Example 1 above, to
selected melanocortin receptors was compared to binding of
Melanotan-II. Melanotan-II was also made by conventional peptide
synthesis, as generally described in Example 1 above.
[0072] Relative binding was determined by competitive inhibition
using .alpha.-MSH. B16-F1 mouse melanoma cells were used as the
source of MC1 receptors; HEK 293 cells, transfected with human
melanocortin receptor sequences, were used as the source of MC3,
MC4 and MC5 receptors; and Y1 cells were used as the source of MC2
receptors. A standard competitive binding assay protocol was
following, using .sup.125I-NDP-MSH as the radioligand.
[0073] Utilizing the assay procedure, the following relative
binding was determined:
5TABLE 1 RELATIVE BINDING.sub.(KI) MC1-R MC3-R MC4-R MC5-R COMPOUND
(nM) MC2-R (nM) (nM) (nM) Compound 1 0.68 >10.sup.-6 M 72.07
19.25 166.8 Melanotan-II 0.12 >10.sup.-6 M 23.7 1.03 69.48
EXAMPLE 3
Agonist Activity of Compound 1
[0074] The agonist activity of Compound 1, made as in Example 1
above, was evaluated by cyclic adenosine 3':5'-cyclic monophosphate
(cAMP) detection using a commercially available cAMP kit (R&D
Systems, DE0350, low pH). HEK 293 cells, stably transfected with
human MC4-R, were used. Compound 1 was evaluated by addition of
each to cells both with and without concomitant addition of
.alpha.-MSH, with .alpha.-MSH also used as a positive control.
Following incubation, the medium was aspirated and cell layers
extracted. The total cAMP accumulation in fixed quantities of cell
extract was determined by competitive immunoassay using the CAMP
kit. These results are shown on Table 2.
6TABLE 2 cAMP CONCENTRATION (pMol/mL) Concentration Compound 1 + of
Test 100 nM Blank Compound Compound 1 .alpha.-MSH .alpha.-MSH
(DMEM) .sup. 1 .times. 10.sup.-7 M 270.2 180.9 107.7 8.24 .sup. 2.5
.times. 10.sup.-8 M 85.1 218.5 146.3 1.9 .sup. 6.25 .times.
10.sup.-9 M 115.6 62.6 45.0 6.5 .sup. 1.56 .times. 10.sup.-9 M 17.9
245.8 107.7 0.7 3.91 .times. 10.sup.-10 M 22.7 127.0 20.2 2.6 0.98
.times. 10.sup.-10 M 5.8 251.8 40.0 0.5
[0075] These results demonstrate that Compound 1 exhibits agonist
activity to MC4-R.
EXAMPLE 4
Induction of Penile Erections Using Compound 1 and Melanotan-II
[0076] The efficacious dose of Compound 1, made as in Example 1
above, to induce penile erection in Sprague Dawley rats by
intravenous dosing was determined. In the rat model of penile
erection, the optimal efficacious dose by intravenous
administration is 100 .mu.g/kg body weight. Melanotan-II was also
made by conventional peptide synthesis, as generally described in
Example 1 above. Male Sprague Dawley rats were administered
Compound 1, in the indicated dose, Melanotan-II, or a saline
negative control, and were observed for erections and side effects,
including excessive grooming, yawning, vacuous chewing,
hypoactivity, and heaving. It is known that Melanotan-II induces
these side effects as doses as low as 300 .mu.g/kg body weight.
Table 3 sets forth the results obtained:
7TABLE 3 INDUCED PENILE ERECTION IN SPRAGUE DAWLEY RATS BY
INTRAVENOUS INJECTION Number of % Rats with Penile Erections/Rat/
Drug Administered Dose (.mu.g/kg) Erection 30 Minutes Compound 1
0.5 50% 0.5 Compound 1 0.75 50% 0.5 Compound 1 1.0 100% 1.0
Compound 1 2.0 100% 2.0 Compound 1 5.0 100% 1.8 Compound 1 10.0
100% 2.5 Compound 1 25.0 50% 0.5 Melanotan-II 10.0 17% 0.3
Melanotan-II 100.0 75% 1.0 Saline (Negative N/A 0% 0 Control)
[0077] Two to four rats were observed in each group, with
observation over a one-half hour period following administration.
No adverse side effects were observed in any rat administered
Compound 1. In this and comparable experiments, it was observed
that Compound 1 was efficacious at doses as low as 1 .mu.g/kg of
body weight when administered by intravenous injection, with
efficacy determined by induction of penile erection in 100% of
animals administered Compound 1. The comparable efficacious dose of
Melanotan-II, with efficacy also determined by induction of penile
erection in 100% of animals, was at least approximately 100
.mu.g/kg body weight. This resulted in the unexpected conclusion
that Compound 1 was at least approximately 50-fold more potent for
inducing erection in a rat model when administered by intravenous
injection than is Melanotan-II. In other studies, no adverse side
effects, including excessive grooming, yawning, vacuous chewing,
hypoactivity, or heaving, were observed on administration of
intravenous doses of Compound 1 as high as 3,000 .mu.g/kg body
weight.
EXAMPLE 5
Pharmacokinetics in Rats Following IV Administration of Compound
1
[0078] A sensitive bioassay, based on high affinity binding to
MCRs, was used to monitor plasma levels of Compound 1 in dosed
rats. Rats were dosed intravenously with 100 .mu.g/kg of Compound
1. A bi-phasic pharmacokinetic profile resulted, as shown in FIG.
1.
EXAMPLE 6
Pre-Formulation Studies for Nasal Delivery
[0079] A series of pre-formulation studies for nasal delivery were
conducted. This study shows the stability of the peptide
Ac-Nle-cyclo(-Asp-His-D-Phe-Arg-Trp-Lys)-OH, at a concentration of
0.825 mg/mL, over a period of twelve weeks, with storage at
40.degree. C.
EXAMPLE 7
Formulations for Nasal Delivery
[0080] Based on the pre-formulation study of Example 6, two
formulations were prepared. For the first, the peptide
Ac-Nle-cyclo(-Asp-His-D-Phe-Arg- -Trp-Lys)-OH was dissolved in a
0.9% saline solution, pH approximately 6.0, at a concentration of
0.825 mg per mL of solution (Nasal Formulation 1).
EXAMPLE 8
Inhibition of Compound 1 Using Melanocortin Antagonist
[0081] The effect of a nonselective melanocortin antagonist,
SHU9119 (Hruby V J, Lu D, Sharma S D, et al. J Med Chem
38:3454-3461 (1995)),
Ac-Nle-cyclo(-Asp-His-D-Nal(2')-Arg-Trp-Lys)-NH.sub.2, to inhibit
induction of penile erection with Compound 1 was studied. A group
of four Sprague Dawley rats were administered 5 .mu.g/kg of SHU9119
by intravenous administration, and five minutes later 25 or 50
.mu.g/kg of Compound 1 in Nasal Formulation 1 was administered by
nasal administration. No erections were observed in the rats over
the one-half hour observation period. No erections were observed in
rats administered a saline control by nasal administration. In
control groups, 100% of the rats administered the same dose of
Compound 1 by the same route of administration, but without
pre-administration of SHU9119, had observed erections. FIG. 4 shows
the number of penile erections per rat during the initial 30 minute
period post-administration; as shown for the 25 .mu.g/kg level, no
penile erections were observed with pre-administration of
SHU9119.
EXAMPLE 9
Nasal Administration to Cynomolgus Monkeys
[0082] Nasal Formulation 1 of Example 6 was administered to
cynomolgus monkeys by nasal spray, at a dose of approximately 50
.mu.g per kg of body weight. Compared to intravenous injection of
the same dose of Ac-Nle-cyclo(-Asp-His-D-Phe-Arg-Trp-Lys)-OH, the
mean bioavailability of the peptide using Nasal Formulation 1 was
approximately 0.73% .+-.0.05. In a related cardiovascular safety
study using cynomolgus monkeys with intranasal administration of
Nasal Formulation 1, an erection lasting approximately one hour was
observed in one of four monkeys.
EXAMPLE 10
Pharmacokinetic Profiles Following Administration to Cynomolgus
Monkeys
[0083] A sensitive radioimmunoassay was used to monitor plasma
levels of Compound 1 in cynomolgus monkeys. Monkeys were dosed
intravenously with 50 .mu.g/kg of Compound 1. A bi-phasic
pharmacokinetic profile resulted, as shown in FIG. 2. Selected
monkeys were intranasally administered 50 .mu.g/kg of Compound 1 in
Nasal Formulation 1, and the resulting pharmacokinetic profiles are
shown in FIG. 3.
EXAMPLE 11
Nasal Administration Dosing Study in Rats
[0084] In another series of experiments, a dosing study was
conducted in rats by nasal administration of from 15 .mu.g per kg
of body weight to 1500 .mu.g per kg of body weight, using Nasal
Formulation 1. Based on penile erectile response behavior, the
peptide was efficacious at all dose amounts administered, and
particularly from 25 .mu.g/kg to 75 .mu.g/kg, with pharacologic
effect observed over the range from 15 .mu.g/kg to 1500 .mu.g/kg.
In addition, the response rate and side effects differed
significantly from those demonstrated with nasal administration of
Melanotan-II. With nasal administration of the peptide of
Melanotan-II, the penile erection response was primarily in the
2.sup.nd and 3.sup.rd ten-minute period, while with Nasal
Formulation 1 the penile erection response was primarily in the
1.sup.st and 2.sup.nd ten-minute period. Additionally, no adverse
side effects were detected at any dosage level with Nasal
Formulation 1, while adverse side effects were seen with
administration of higher doses of the peptide of Melanotan-II.
Table 4 sets forth the comparative data:
8TABLE 4 INDUCED PENILE ERECTION IN SPRAGUE DAWLEY RATS BY
INTRANASAL ADMINISTRATION Drug % Rats with Number of Erections/
Administered Dose (.mu.g/kg) Penile Erection Rat/30 Minutes
Compound 1 15 50% 0.5 Compound 1 25 100% 2.0 Compound 1 37 100% 2.0
Compound 1 50 100% 2.0 Compound 1 75 100% 2.5 Compound 1 500 100%
1.5 Compound 1 1000 100% 1.5 Compound 1 1500 50% 0.5 Melanotan-II
100 25% 0.25 Melanotan-II 500 100% 1.0 Saline (Negative N/A 0% 0
Control)
EXAMPLE 12
Pharmacokinetic Profiles Following Administration to Beagle
Dogs
[0085] A sensitive bioassay, based on high affinity binding to
melanocortin receptors, was used to monitor plasma levels of
Compound 1 in Beagle dogs. The dogs were administered 0 (control),
30, 150 or 650 .mu.g/kg of Compound 1 in Nasal Formulation 1 by
nasal administration. The pharmacokinetic profile through four
hours post-administration is shown in FIG. 5.
EXAMPLE 13
Female Sexual Response in Rats
[0086] Female Long-Evans rats (200-250 g) were house in groups of 2
to 4 depending on size in plastic cages in a colony room maintained
on a reversed 12:12 hour light/dark cycle at approximately
21.degree. C. with food and water ad libitum. The rats were
ovariectomized bilaterally via lumbar incisions one week prior to
preliminary tests of sexual behavior. Intact Long-Evans male rats
served as stimulus males, and were sexual vigorous, initiating
copulatory activity with females within 15 seconds of being placed
into bilevel chambers containing female rats. Following preliminary
tests of sexual behavior, females were administered subcutaneous
injections of estradiol benzoate, 10 .mu.g in 0.1 mL of sesame oil,
forty-eight hours prior to testing, and progesterone, 500 .mu.g in
0.1 mL of sesame oil, four hours prior to testing. Administration
of estradiol benzoate and progesterone induced full sexual
receptivity in the female rats. Female rats were placed in bilevel
chambers five minutes prior to the introduction of a sexual
vigorous male. After females acquired baseline rates of sexual
activity, a dose response analysis of Compound 1 was conducted,
with 0, 50, 100 or 200 .mu.g/kg, injected subcutaneously, one
minute before each test. Tests were videotaped and score
subsequently for appetitive level changing during the five minute
period prior to the introduction of the male, lordosis quotients
and reflex magnitudes, incidents of solicitation (head-wise
orientation and runaway), hops and darts, pacing (level changes per
mount), female mounts and defensive responses. FIG. 6 depicts
results, with a between-subjects analysis of variance (ANOVA) of
P<0.004. In a second experiment, female rats were administered
only estradiol benzoate prior to administration of 0 or 200
.mu.g/kg of Compound 1 injected subcutaneously. The results are
shown in FIG. 7. In general, in both experiments the 200 .mu.g/kg
dose of Compound 1 increased the number of solicitations
significantly without affecting rates of appetitive level changing,
pacing, lordosis, female mounts or rejection responses.
EXAMPLE 14
Female Sexual Response in Rats
[0087] Female Long-Evans rats as in Example 13 were tested in a
unilevel pacing chambers bisected by a four-holed divider, as
generally described in Yang L. V. and Clemens L. G., Physiol Behav
61:889-894 (1997), for five minutes prior to the introduction of a
sexually vigorous male for a thirty-minute test of copulation. As
in Example 13, following priming with estradiol benzoate and
progesterone, 0, 50,100 or 200 .mu.g/kg of Compound 1 was
administered subcutaneously one minute before each test, with
female animals placed in the unilevel pacing chambers bisected by
the four-hole divider through which only the females could pass.
Tests were videotaped and scored as in Example 13, with pacing
defined as number of exits from and returns to the side with the
male; and exist and return latencies following mounts,
intromissions or ejaculations. FIGS. 8A, 8B, 8C and 8D show the
results. The 200 .mu.g/kg dose of Compound 1 increased the number
of solicitations, hops and darts, and female mounts significantly
compared to controls, but had no effect on lordosis or defensive
responding. This dose also increase the ejaculation return
latencies, but not the mount or intromission return latencies. This
dose had no significant effect on the number of exits or returns
made by the females.
EXAMPLE 15
Ferret Emesis Study
[0088] Groups of six ferrets were studied for emetic effect of
Compound 1 and Melanotan-II at various dose ranges with delivery
both by intravenous and intranasal administration. Ferrets are a
recognized model for emetic studies. The vehicle for both
intravenous and intranasal administration was saline. The positive
control was cisplatin, known to induce vomiting in ferrets. The
data is described as the number of animals out of a group of six
showing signs of vomiting.
9TABLE 5 EMETIC EFFECT COMPARISON Dosages in .mu.g/kg Signs of
Intravenous (IV) vomiting (proportions) ZZIntranasal (IN) Compound
1 Melanotan-II Vehicle (saline) 0/6 0/6 10 (IV) 0/6 1/6 30 (IV) 6/6
4/6 100 (IV) 6/6 6/6 100 (IN) 0/6 0/6 300 (IN) 1/6 5/6 500 (IN) 1/6
6/6
EXAMPLE 16
Administration For Treatment of Male Erectile Dysfunction
[0089] For treatment of male erectile dysfunction in human males
with psychogenic erectile dysfunction, a dose of Compound 1 in a
nasal formulation is administered by means of a nasal
administration metered dose device. The device dispenses a metered
volume, such as a 100 .mu.L spray volume, and is optionally
administered by the patient into a single nostril. If desired,
erectile response is measured using an appropriate device, such as
a Rigiscan (Dacomed).
EXAMPLE 17
Administration For Treatment of Female Sexual Dysfunction
[0090] For treatment of female sexual dysfunction in human females
with psychogenic sexual dysfunction, a dose of Compound 1 in a
nasal formulation is administered by means of a nasal
administration metered dose device. The device dispenses a metered
volume, such as a 100 .mu.L spray volume, and is optionally
administered by the patient into a single nostril.
[0091] The preceding examples can be repeated with similar success
by substituting the generically or specifically described reactants
and/or operating conditions of this invention for those used in the
preceding examples.
[0092] Although the invention has been described in detail with
particular reference to these preferred embodiments, other
embodiments can achieve the same results. Variations and
modifications of the present invention will be obvious to those
skilled in the art and it is intended to cover in the appended
claims all such modifications and equivalents. The entire
disclosures of all references, applications, patents, and
publications cited above are hereby incorporated by reference.
Sequence CWU 1
1
1 1 4 PRT Artificial Alpha-melanocyte-stimulation hormone
tetrapeptide core sequence 1 His Phe Arg Trp 1
* * * * *