U.S. patent application number 10/405701 was filed with the patent office on 2003-10-30 for preventatives/remedies for skin aging.
This patent application is currently assigned to Fujisawa Pharmaceutical Co. Ltd.. Invention is credited to Furukawa, Yusuke, Murakami, Manabu, Nakahara, Kunio, Nomura, Kazuhiko, Yabuta, Tsuguo, Yasumura, Mitsuru.
Application Number | 20030203853 10/405701 |
Document ID | / |
Family ID | 12609497 |
Filed Date | 2003-10-30 |
United States Patent
Application |
20030203853 |
Kind Code |
A1 |
Yabuta, Tsuguo ; et
al. |
October 30, 2003 |
Preventatives/remedies for skin aging
Abstract
A composition for prophylaxis and therapy of dermal aging which
comprises a substance having human leukocyte elastase inhibitory
activity as an active ingredient.
Inventors: |
Yabuta, Tsuguo; (Osaka,
JP) ; Yasumura, Mitsuru; (Hyogo, JP) ;
Nakahara, Kunio; (Hyogo, JP) ; Furukawa, Yusuke;
(Nara, JP) ; Nomura, Kazuhiko; (Osaka, JP)
; Murakami, Manabu; (Osaka, JP) |
Correspondence
Address: |
OBLON, SPIVAK, MCCLELLAND, MAIER & NEUSTADT, P.C.
1940 DUKE STREET
ALEXANDRIA
VA
22314
US
|
Assignee: |
Fujisawa Pharmaceutical Co.
Ltd.
Osaka
JP
|
Family ID: |
12609497 |
Appl. No.: |
10/405701 |
Filed: |
April 3, 2003 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10405701 |
Apr 3, 2003 |
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09622053 |
Dec 15, 2000 |
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6610748 |
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09622053 |
Dec 15, 2000 |
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PCT/JP99/00761 |
Feb 19, 1999 |
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Current U.S.
Class: |
514/621 ;
424/401; 514/18.8; 514/20.4 |
Current CPC
Class: |
A61P 17/00 20180101;
A61K 38/005 20130101; A61K 38/03 20130101; A61K 38/04 20130101 |
Class at
Publication: |
514/19 ;
424/401 |
International
Class: |
A61K 038/04; A61K
007/00 |
Foreign Application Data
Date |
Code |
Application Number |
Feb 24, 1998 |
JP |
10/41479 |
Claims
1. A composition for prophylaxis and therapy of dermal aging which
comprises a substance having human leukocyte elastase inhibitory
activity as an active ingredient.
2. A composition for prophylaxis and therapy of wrinkles which
comprises WS7622A mono- or disulfate ester or a pharmaceutically
acceptable salt thereof as an active ingredient.
3. A composition for prophylaxis and therapy of dermal aging which
comprises a trifluoromethylketone derivative of the following
formula or a pharmaceutically acceptable salt thereof as an active
ingredient. 7[wherein R.sup.1 represents a lower alkyl group having
1 or 2 substituents selected from among carboxy, esterified carboxy
and di-lower alkylcarbamoyl; a phenyl (lower) alkyl group which may
optionally have halogen, amino or nitro on the phenyl moiety
thereof and carboxy or esterified carboxy on the alkyl moiety
thereof; a halophenyl group; a morpholino group; or a morpholino
(lower) alkyl group; R.sup.2 and R.sup.3 each represents a lower
alkyl group; X represents-- or --NH--; 8
4. A composition for prophylaxis and therapy of wrinkles which
comprises a trifluoromethylketone derivative of the following
formula or a pharmaceutically acceptable salt thereof as an active
ingredient. 9[wherein R.sup.1 represents a lower alkyl group having
1 or 2 substituents selected from among carboxy, esterified carboxy
and di-lower alkylcarbamoyl; a phenyl (lower) alkyl group which may
optionally have halogen, amino or nitro on the phenyl moiety
thereof and carboxy or esterified carboxy on the alkyl moiety
thereof; a halophenyl group; a morpholino group; or a morpholino
(lower) alkyl group; R.sup.2 and R.sup.3 each represents a lower
alkyl group; X represents-- or --NH--; 10
5. The composition for prophylaxis and therapy of dermal aging as
claimed in claim 1 wherein the substance having human leukocyte
elastase inhibitory activity is Substance FR134043.
6. The composition for prophylaxis and therapy of wrinkles as
claimed in claim 2 wherein the substance having human leukocyte
elastase inhibitory activity is Substance FR134043.
7. The composition for prophylaxis and therapy of dermal aging as
claimed in claim 1 wherein the substance having human leukocyte
elastase inhibitory activity is Substance FK706.
8. The composition for prophylaxis and therapy of wrinkles as
claimed in claim 4 wherein the substance having human leukocyte
elastase inhibitory activity is Substance FK706.
9. A method for prophylaxis and therapy of dermal aging which
comprises administering a substance having human leukocyte elastase
inhibitory activity to a patient for the prevention or treatment of
dermal aging.
10. Use of a substance having human leukocyte elastase inhibitory
activity for the production of a pharmaceutical composition for the
prophylaxis and therapy of dermal aging.
Description
TECHNICAL FIELD
[0001] This invention relates to a composition for prophylaxis and
therapy of dermal aging which comprises a substance having human
leukocyte elastase inhibitory activity as an active ingredient.
BACKGROUND ART
[0002] Aging of the skin is said to begin after the third decade of
life. Obvious signs of aging include decreases in the moistness,
gloss, smoothness, tonus, etc. of the skin and an increased number
of wrinkles. These are suspected to result from the morphological
and functional changes of the organs and tissues making up the
skin. Thus, aging is accompanied by thinning of the epidermis and
loss of oxytalan fiber in the papillary layer of the dermis.
[0003] Dermal aging, epitomized by wrinkling of the skin, is a
serious beauty problem for women but no satisfactory remedy has
been available to this day.
[0004] The present inventors discovered that a substance having
human leukocyte elastase inhibitory activity is effective in the
prevention and treatment of dermal aging and has perfected this
invention.
DISCLOSURE OF INVENTION
[0005] This invention is directed to a composition for prophylaxis
and therapy of dermal aging which comprises a substance having
human leukocyte elastase inhibitory activity as an active
ingredient.
[0006] The substance having human leukocyte elastase inhibitory
activity which can be used as the active ingredient of this
composition for prophylaxis and therapy of dermal aging may be any
substance that has human leukocyte elastase inhibitory activity.
Furthermore, the substance having human leukocyte elastase activity
which can be used in this invention includes not only substances
which are direct inhibitors of that activity but also substances
which inhibit leukocyte elastase activity indirectly through
suppression of leukocyte infiltration and inhibition of elastase
production. Thus, while many substances having such activity are
known, any novel substance that may have human leukocyte elastase
inhibitory activity can also be employed. The following is a
partial listing of the preferred compounds in this category. (1)
WS7622A mono- or disulfate and pharmaceutically acceptable salts
thereof; among these, WS7622A disulfate ester disodium salt and
WS7622A disulfate ester dipotassium salt are known substances which
have the following physicochemical properties (JP Kokai
H4-279600).
[0007] WS7622A disulfate ester disodium salt (hereinafter sometimes
referred to briefly as FR134043):
[0008] Description: Colorless crystals
[0009] Solubility: Soluble: water, methanol
[0010] Insoluble: chloroform, n-hexane
[0011] Melting point: 257.about.263.degree. C. (decomp.) Optical
rotation: [.alpha.].sup.23.sub.D+37.5.degree. (c=1, methanol)
Molecular formula: C.sub.47H.sub.61N.sub.9O.sub.19S.sub.2Na.sub.2
Elemental analysis: Calcd.: (for
C.sub.47H.sub.61N.sub.9O.sub.19S.sub.2Na.sub.2.6H.sub.2O): C,
44.30; H, 5.77; N, 9.89; S, 5.03; Na, 3.61% Found: C, 44.98; H,
5.90; N, 10.06; S, 5.00; Na, 3.98% Molecular weight: FAB-MS m/z
1188 (M+Na).sup.+
[0012] Thin-Layer Chromatography:
1 TABLE 1 Stationary Developer phase solvent Rf Silica gel
CHCl.sub.3--CH.sub.3OH--H.sub.2O 0.11 (Merck Art. 5715) (65:25:4)
n-butanol-acetic 0.29 acid-water (4:2:1) Infrared absorption
spectrum: .upsilon..sup.KBr.sub.max: 3360, 2960, 1735, 1660, 1640,
1530, 1500, 1380, 1250, 1200, 1060, 1030, 940, 890 cm.sup.-1
.sup.1H Nuclear magnetic resonance spectrum: (400 MHz, D.sub.2O)
.delta. 7.50 (1H, s) 7.27 (1H, s) 7.33-7.24 (3H, m) 6.94 (1H, q,
J=7 Hz) 6.85 (2H, br d, J=8 Hz) 5.53 (1H, m) 5.37 (1H, m) 4.80 (1H,
br s) 4.63-4.57 (2H, m) 4.53 (1H, m) 4.06 (1H, m) 3.99 (1H, d, J=10
Hz) 3.56 (1H, br d, J=14 Hz) 3.46 (1H, m) 2.97 (3H, s) 2.97-2.88
(2H, m) 2.72 (1H, m) 2.59 (1H, m) 2.51-2.38 (2H, m) 2.09-1.91 (4H,
m) 1.82-1.60 (3H, m) 1.77 (3H, d, J=7 Hz) 1.50 (3H, d, J=6.5 Hz)
1.40 (1H, m) 1.11 (6H, d, J=7 Hz) 0.99 (3H, d, J=6.5 Hz) 0.97 (3H,
d, J=6.5 Hz) .sup.13C Nuclear magnetic resonance spectrum: (100
MHz, D.sub.2O) .delta. 183.6 (s) 177.9 (s) 177.7 (s) 174.8 (s)
173.8 (s) 173.3 (s) 172.4 (s) 167.8 (s) 161.5 (s) 145.5 (s) 144.9
(s) 139.6 (d) 139.0 (s) 137.0 (s) 136.0 (s) 132.3 (d).times.2 131.0
(d).times.2 129.6 (d) 127.4 (d) 125.9 (d) 77.4 (d) 75.1 (d) 63.8
(d) 62.7 (d) 59.1 (d) 55.9 (d) 54.9 (d) 51.9 (d) 41.9 (t) 37.2 (d)
36.9 (t) 34.1 (q) 32.3 (d) 31.9 (t) 31.8 (t) 31.2 (t) 27.5 (t) 23.7
(t) 21.7 (q) 21.4 (q).times.2 21.3 (q) 21.1 (q) 15.5 (q)
[0013] Amino Acid Analysis:
[0014] WS7622A disulfate ester disodium salt (1 mg) was hydrolyzed
with 6 N-hydrochloric acid at 110.degree. C. for 20 hours and,
then, concentrated to dryness under reduced pressure, and the
residue was analyzed with Hitachi 835 Automatic Amino Acid
Analyzer. As the amino acids standard solution, Wako Pure
Chemical's Type H (Wako Code 013-08391) and Type B (016-08641) were
used.
[0015] As a result, threonine, valine, phenylalanine, ornithine,
ammonia and several unknown ninhydrin-positive components were
detected.
[0016] As a partial chemical structure of WS7622A disulfate ester
disodium salt, the following formula is proposed. 1
[0017] WS7622A disulfate ester dipotassium salt:
[0018] Description: Colorless amorphous powders
[0019] Solubility: Soluble: water, methanol
[0020] Insoluble: chloroform, n-hexane
[0021] Melting point: 230.about.237.degree. C. (decomp.) Optical
rotation: [.alpha.].sup.23.sub.D+34.degree. (c=1, methanol)
Molecular formula: C.sub.47H.sub.61N.sub.9O.sub.19S.sub.2K.sub.2
Elemental analysis: Calcd.: (for
C.sub.47H.sub.61N.sub.9O.sub.19S.sub.2K.sub.2.6H.sub.2O ): C,
43.21; H, 5.63; N, 9.65; S, 7.91; Na, 5.99% Found: C, 43.96; H,
5.44; N, 9.97; S, 5.09; Na, 4.49% Molecular weight: FAB-MS m/z 1236
(M+K).sup.+
[0022] Thin-Layer Chromatography:
2 TABLE 2 Stationary Developer phase solvent Rf Silica gel
CHCl.sub.3--CH.sub.3OH--H.sub.2O 0.13 (Merck Art. 5715) Infrared
absorption spectrum: .upsilon..sup.KBr.sub.max: 3360, 2960, 1735,
1660, 1640, 1530, 1500, 1405, 1380, 1250, 1200, 1050, 1030, 940,
890 cm.sup.-1 .sup.1H Nuclear magnetic resonance spectrum: (400
MHz, D.sub.2O) .delta.: 7.52 (1H, s) 7.28 (1H, s) 7.34-7.25 (3H, m)
6.96 (1H, q, J=7 Hz) 6.87 (2H, br d, J=8 Hz) 5.56 (1H, m) 5.40 (1H,
m) 4.84 (1H, br s) 4.70-4.55 (3H, m) 4.10 (1H, m) 4.03 (1H, m) 3.60
(1H, br d, J=14 Hz) 3.50 (1H, m) 3.00 (3H, s) 3.00-2.85 (2H, m)
2.76 (1H, m) 2.62 (1H, m) 2.55-2.40 (2H, m) 2.12-1.95 (4H, m)
1.90-1.65 (3H, m) 1.79 (3H, d, J=7 Hz) 1.53 (3H, d, J=6.5 Hz) 1.45
(1H, m) 1.14 (6H, d, J=7 Hz) 1.02 (3H, d, J=6.5 Hz) 1.00 (3H, d,
J=6.5 Hz)
[0023] Amino Acid Analysis:
[0024] WS7622A disulfate ester dipotassium salt (1 mg) was
hydrolyzed with 6 N-hydrochloric acid (1 ml) at 110.degree. C. for
20 hours and, then, concentrated to dryness under reduced pressure,
and the residue was analyzed with Hitachi 835 Automatic Amino Acid
Analyzer. As the amino acids standard solution, Wako Pure
Chemical's Type H (Wako Code 013-08391) and Type B (016-08641) were
used.
[0025] As a result, threonine, valine, phenylalanine, ornithine,
ammonia and several unknown ninhydrin-positive components were
detected.
[0026] As a partial chemical structure of WS7622A disulfate ester
dipotassium salt, the following formula is proposed. 2
[0027] As the pharmaceutically acceptable salt of WS7622A mono- or
disulfate ester, there can be mentioned the mono- or di-salts with
inorganic or organic bases, such as alkali metal salts (e.g. sodium
salt, potassium salt, etc.), alkaline earth metal salts (e.g.
calcium salt etc.), ammonium salt, ethanolamine salt, triethylamine
salt, dicyclohexylamine salt and pyridine salt, among others.
[0028] Substance WS7622A which is a starting compound for synthesis
of said WS7622A mono- or disulfate ester also has human leukocyte
elastase inhibitory activity and can, therefore, be used as a
prophylactic and therapeutic agent for wrinkles. This substance is
known to have the following physicochemical properties (JP Kokai
H3-218387, JP Kokai H4-279600).
[0029] Physicochemical properties of Substance WS7622A:
[0030] Description: colorless prisms
[0031] Acidity-alkalinity: acidic
[0032] Color reactions:
[0033] Positive: cerium sulfate reaction, iodine vapor reaction,
ferric chloride reaction
[0034] Negative: ninhydrin reaction, Molisch reaction, Dragendorff
reaction
[0035] Solubility:
[0036] Soluble: methanol, ethanol, n-butanol
[0037] Slightly soluble: chloroform, acetone, ethyl acetate
[0038] Insoluble: water, n-hexane
[0039] Thin-layer chromatography (TLC):
[0040] Chloroform-methanol (5:1, v/v)
[0041] Rf 0.51
[0042] Acetone-methanol (10:1)
[0043] Rf 0.62
[0044] (Kieselgel 60F.sub.254 silica gel plate, Merck)
[0045] Melting point: 250.about.252.degree. C. (decomp.) Optical
rotation: [.alpha.].sup.23.sub.D+36.degree. (c=1, methanol) UV
absorption spectrum: .lambda..sup.MeOH.sub.max 287 nm
(.epsilon.=3600) .lambda..sup.MeOH--HC1.- sub.max 287 nm
.lambda..sup.MeOH--NaOH.sub.max 298 nm Molecular formula:
C.sub.47H.sub.63N.sub.9O.sub.13 Elemental analysis: Calcd.: (for
C.sub.47H.sub.63N.sub.9O.sub.13.2H.sub.2O): C, 56.56; H, 6.77; N,
12.63% Found: C, 56.65; H, 6.62; N, 12.27% Molecular weight: FAB-MS
m/z 984 (M+Na).sup.+Infrared absorption spectrum:
.upsilon..sup.KBr.sub.max: 3400, 3300, 3060, 2980, 2940, 1735,
1710, 1690, 1670, 1660, 1640, 1540, 1520, 1470, 1380, 1330, 1300,
1260, 1220, 1200, 1160, 1130, 1090, 1000, 980, 940, 920
cm.sup.-1
3 .sup.1H Nuclear magnetic resonance spectrum: (400 MHz,
CD.sub.3OD) .delta.: 7.22-7.09 (3H, m) 6.88-6.77 (3H, m) 6.74 (1H,
s) 6.46 (1H, s) 5.46 (1H, m) 5.18 (1H, s) 4.85 (1H, s) 4.77 (1H, m)
4.65 (1H, m) 4.50 (1H, m) 3.96 (1H, m) 3.91 (1H, d, J=9 Hz)
3.60-3.47 (2H, m) 3.03 (1H, m) 2.90 (3H, s) 2.86 (1H, m) 2.59-2.49
(2H, m) 2.39 (1H, m) 2.29-2.16 (2H, m) 2.00 (1H, m) 1.84 (1H, m)
1.74 (3H, d, J=6 Hz) 1.72-1.53 (4H, m) 1.44 (3H, d, J=6 Hz) 1.12
(1H, m) 1.10 (6H, d, J=6 Hz) 0.99 (3H, d, J=6 Hz) 0.94 (3H, d, J=6
Hz) .sup.13C Nuclear magnetic resonance spectrum: (100 MHz,
CD.sub.3OD) .delta. 179.7 (s) 176.3 (s) 174.7 (s) 173.3 (s) 172.4
(s) 171.4 (s) 170.3 (s) 165.8 (s) 160.2 (s) 145.7 (s) 145.6 (s)
137.5 (s) 134.0 (d) 131.4 (s) 130.6 (d).times.2 129.8 (s) 129.1
(d).times.2 129.1 (s) 127.6 (d) 119.1 (d) 118.0 (d) 76.0 (d) 73.4
(d) 63.1 (d) 61.4 (d) 57.1 (d) 53.6 (d) 52.7 (d) 50.5 (d) 39.9 (t)
36.1 (t) 35.8 (d) 31.8 (q) 31.0 (t) 30.8 (d) 29.9 (t) 29.7 (t) 25.2
(t) 22.3 (t) 20.2 (q) 20.0 (q).times.2 19.7 (q) 19.5 (q) 13.3
(q)
[0046] Amino Acid Analysis:
[0047] WS7622A (1 mg) was hydrolyzed with 6 N-hydrochloric acid (1
m) at 110.degree. C. for 20 hours and concentrated to dryness under
reduced pressure, and the residue was analyzed with Hitachi 835
Automatic Amino Acid Analyzer. As the amino acids standard
solution, Wako Pure Chemical's Type H (Wako Code 013-08391) and
Type B (016-08641) were used.
[0048] As a result, threonine, valine, phenylalanine, ornithine,
ammonia and several unknown ninhydrin-positive components were
detected.
[0049] As a partial chemical structure of WS7622A, the following
formula is proposed. 3
[0050] The salt of Substance WS7622A includes salts with inorganic
or organic bases, such as alkali metal salts (e.g. sodium salt,
potassium salt, etc.), alkaline earth metal salts (e.g. calcium
salt etc. ), ammonium salt, ethanolamine salt, triethylamine salt
and dicyclohexylamine salt, among others.
[0051] Substances WS7622B, C and D and derivatives thereof (JP
Kokai H3-218387) , all of which have human leukocyte elastase
inhibitory activity as well can also be used as prophylactic and
therapeutic agents for wrinkles.
[0052] The above-mentioned Substance WS7622A (as well as Substances
WS7622B, C and D) can be produced by growing Streptomyces
resistomycificus No. 7622, which strain has been deposited with
National Institute of Bioscience and Human Technology, one of the
international culture collections under Budapest Treaty, with the
accession number of FERM BP-2306 assigned.
[0053] (2) A trifluoromethylketone derivative of the following
formula and its pharmaceutically acceptable salt: 4
[0054] [wherein R.sup.1 represents a lower alkyl group having 1 or
2 substituents selected from among carboxy, esterified carboxy and
di-lower alkylcarbamoyl; a phenyl (lower) alkyl group which may
optionally have halogen, amino or nitro on the phenyl moiety
thereof and carboxy or esterified carboxy on the alkyl moiety
thereof; a halophenyl group; a morpholino group; or a morpholino
(lower) alkyl group; R.sup.2 and R.sup.3 each represents a lower
alkyl group; X represents-- or --NH--; 5
[0055] (3) a trifluoromethylketone derivative of the following
formula and its pharmaceutically acceptable salt: 6
[0056] (wherein R.sup.1.about.R.sup.3 have the same meanings as
defined above (2)).
[0057] (4) 3 (RS)-[[4-(carboxymethylaminocarbonyl)
phenyl-carbonyl]-L-valy-
l-L-prolyl]amino-1,1,1-trifluoro-4-methyl-2-oxopentane or its
sodium salt (this sodium salt will sometimes be referred to briefly
as FK706).
[0058] The compounds mentioned in the above paragraphs
(2).about.(4) are known compounds as described in JP Kokai
H4-297446, for instance. As the pharmaceutically acceptable salts
of said compounds (2).about.(3) , there can be mentioned the
respective salts with organic or inorganic bases, such as alkali
metal salts (e.g. sodium salts, potassium salts, etc.), alkaline
earth metal salts (e.g. calcium salts etc.), ammonium salts,
ethanolamine salts, triethylamine salts, dicyclo-hexylamine salts,
etc. , methanesulfonates, and organic or inorganic acid addition
salts such as hydrochlorides, sulfates, nitrates, phosphates and so
forth.
[0059] The preferred examples relevant to the various definitions
given hereinbefore are now set forth in detail. The term "lower"
means 1.about.6 carbon atoms unless otherwise specified. As the
preferred examples of "halogen", there can be mentioned fluoro,
chloro, bromo and iodo. The preferred "lower alkyl group" includes
residues of straight-chain or branched-chain alkanes containing
1.about.6 carbon atoms, such as methyl, ethyl, propyl, isopropyl,
butyl, isobutyl, t-butyl, pentyl, neopentyl and hexyl, preferably
those having 1.about.4 carbon atoms. The preferred "esterified
carboxy" includes, among others, alkyl esters, i.e. alkoxycarbonyl
groups such as lower alkoxycarbonyl (e.g. methoxycarbonyl,
ethoxycarbonyl, propoxycarbonyl, butoxycarbonyl,
tert-butoxycarbonyl, etc.), and phenyl (lower) alkyl esters, i.e.
phenyl (lower) alkoxycarbonyl groups such as benzyloxycarbonyl
etc., and benzoyl (lower) alkyl esters, i.e. benzoyl (lower)
alkoxycarbonyl groups such as benzoylmethoxycarbonyl and so
forth.
[0060] The preferred "lower alkylene group" includes but is not
limited to methylene, ethylene, propylene and isopropylene. The
preferred "di-lower-alkylcarbamoyl group" includes
N,N-dimethylcarbamoyl and N,N-diethylcarbamoyl, among others.
[0061] (5) Substance FR901451 having the following physicochemical
properties and its pharmaceutically acceptable salt:
[0062] Description: white powders
[0063] Color reactions:
[0064] Positive: cerium sulfate, iodine vapor, Ehrlich, and
ninhydrin reactions
[0065] Negative: Molisch reaction
[0066] Solubility:
[0067] Soluble: water, methanol, dimethyl sulfoxide
[0068] Hardly soluble: acetate
[0069] Insoluble: ethyl acetate
[0070] Melting point: 243.about.245.degree. C. (decomp.) Optical
rotation: [.alpha.].sup.23.sub.D-15.degree. (c=0.65, H.sub.2O)
Ultraviolet absorption spectrum: .lambda..sup.MeOH.sub.max
nm(.epsilon.): 275 (4300), 281 (4500), 290 (3900) Molecular
formula: C.sub.60H.sub.79N.sub.13O.sub.1- 8 Elemental analysis:
Calcd.: (for C.sub.60H.sub.79N.sub.13 O.sub.18.10H.sub.2O): C,
49.68; H, 6.88; N, 12.55 Found: C, 49.95; H, 6.28; N, 12.42
Molecular weight: FAB-MS m/z 1270 (M+H).sup.+
[0071] Thin-Layer Chromatography:
4 TABLE 3 Stationary Developer phase solvent Rf Silica gel
CHCl.sub.3:MeOH:NH.sub.4OH 0.60 (Merck) (15:11:5) RP-18 70% aqueous
0.32 (Merck) methanol FT-IR spectrum: .upsilon..sup.KBr.sub.max:
3390, 3070, 2970, 2880, 1740, 1660, 1530, 1450, 1410, 1380, 1350,
1250, 1190, 1110, 1080, 1010, 750, 700, 670, 660, 620, 600
cm.sup.-1 .sup.1H Nuclear magnetic resonance spectrum: (400 MHz,
D.sub.2O) .delta.: 7.70 (1H, d, J=7 Hz) 7.52 (1H, d, J=7.5 Hz)
7.44-7.23 (7H, m) 7.22 (1H, s) 5.59 (1H, q, J=7 Hz) 4.94 (1H, t,
J=4.5 Hz) 4.85-4.74 (3H, m) 4.58 (1H, dd, J=6 Hz, 10 Hz) 4.45-4.35
(3H, m) 4.30 (1H, dd, J=4 Hz, 7 Hz) 4.07 (1H, m) 3.99 (1H, dd, J=10
Hz, 4.5 Hz) 3.66-3.50 (3H, m) 3.44-3.25 (4H, m) 3.16-2.93 (4H, m)
2.87 (1H, d, J=18 Hz) 2.80-2.68 (2H, m) 2.56-2.48 (2H, m) 2.08 (1H,
dd, J=16 Hz, 4 Hz) 1.87-1.53 (9H, m) 1.43 (3H, d, J=7 Hz) 1.30 (3H,
d, J=6.5 Hz) 1.45-1.17 (4H, m) 0.95 (3H, d, J=6 Hz) 0.84 (3H, d,
J=6 Hz) .sup.13C Nuclear magnetic resonance spectrum: (100 MHz,
D.sub.2O) .delta. 177.2 (s) 130.0 (d).times.2 56.0 (d) 31.4 (t)
176.5 (s) 129.8 (d).times.2 54.1 (d) 28.8 (t) 174.6 (s) 128.5 (d)
53.8 (d) 26.6 (t) 174.2 (s) 127.8 (s) 53.2 (d) 25.1 (d) 174.0 (s)
125.5 (d) 53.1 (d) 23.2 (q) 173.2 (s) 123.2 (d) 52.9 (d) 23.2 (t)
173.0 (s) 120.9 (d) 52.8 (d) 23.1 (t) 172.8 (s) 118.7 (d) 49.5 (d)
20.8 (q) 172.6 (s) 113.1 (d) 48.6 (t) 19.4 (q) 172.5 (s) 108.8 (s)
40.1 (t) 18.3 (q) 172.1 (s) 73.3 (d) 39.6 (t) 171.7 (s) 69.7 (d)
39.4 (t) 171.4 (s) 64.3 (t) 38.9 (t) 170.3 (s) 62.1 (d) 35.3 (t)
137.2 (s) 60.9 (d) 34.8 (t) 136.0 (s) 57.1 (d) 31.7 (t)
[0072] The above Substance FR901451 is known as the substance
elaborated by Substance FR901451-producing strains of organisms
belonging to the genus Flexibacter (e.g. WO93/02203). Flexibacter
sp. No. 758, which is among such producer strains, has been
deposited with National Institute of Bioscience and Human
Technology, an international culture collection under Budapest
Treaty, with the accession number of FERM BP-3420 assigned.
[0073] The pharmaceutically acceptable salt of the above Substance
FR901451 includes the same kinds of salts as mentioned for the
pharmaceutically acceptable salts of the compounds
(2).about.(3).
[0074] In addition to the foregoing substances, the following can
be mentioned as examples of the substance having elastase
inhibitory activity: .alpha.1-antitrypsin, SLP1 (secretory
leukocyte protease inhibitor) [American Review of Respiratory
Diease Vol. 147, 1993, P442-446], urinastatin, colchicine,
erythromycin, clarithromycin, ICI200, 880, ONO-8046 [American
Journal of Respiratory and Critical Care Medicine Vol. 153,
P391-397], anti-elastase antibodies, and so forth.
[0075] For the purposes of this invention, a substance having human
leukocyte elastase inhibitory activity is efficacious and can be
administered for the prevention and treatment of dermal aging in
general. More particularly, it can be indicated for the prevention
and treatment of decreases in moistfulness, sheen, smoothness, and
tonus of the skin and even increased wrinkling, or the prevention
and treatment of skin flaccidity. These signs of dermal aging are
suspected to arise from morphological and functional changes of the
organs and tissues making up the skin, and actually, thinning of
the horny layer of the epidermis and loss of oxytalan fiber in the
papillary layer of the corium are noted.
[0076] The signs of dermal aging being as mentioned above, the
efficacy of a substance having human leukocyte elastase inhibitory
activity is particularly pronounced for the elimination or
diminution of wrinkles or prevention of increased wrinkling,
improvement of skin texture (fineness, handle) and amelioration of
the shade of the skin (shadowy complexion).
[0077] Furthermore, said substances are efficacious for promoting
neogenesis of oxytalan fiber in the region of dermal papillae and
neogenesis of collagen fibrils in the dermis immediately beneath
the epidermis, and for increasing the thickness of the epidermis,
among others.
[0078] In addition, substances having human leukocyte elastase
inhibitory activity are efficacious in various skin diseases such
as scleroderma, elephantiasis, scars, steroid-induced thinning of
the skin, keloids, pressure sores, wounds, refractory ulcers,
psoriasis, spots, freckles, senile plaques, rough skin and
pityriasis.
[0079] The following is an example of experimentation relevance to
this invention.
[0080] Object of Experiment:
[0081] The therapeutic efficacy of neutrophil (leukocyte) elastase
inhibitors in "dermal aging" was evaluated in hairless dogs.
[0082] Experimental Animals:
[0083] Two adult (old) experimental hairless dogs constructed by
cross-breeding of a Mexican hairless dog and a beagle dog were
used. Old dogs presenting with age-associated fine "wrinkles", not
observed in young dogs, on the body surface were used as subject
animals.
[0084] No. 8807 (10 yr old, male)
[0085] No. 8808 (10 yr old, female)
[0086] Investigational Drugs:
[0087] 1) FK706.multidot.0.2%
[0088] 2) FK706.multidot.0.02%
[0089] 3) FR134043.multidot.0.2%
[0090] 4) FR134043.multidot.0.02%
[0091] 5) Polyethylene glycol (PEG) (solvent control) (PEG was used
as the solvent for drugs 1.about.4)
[0092] Administration:
[0093] Each drug was applied in one location (a total of 5
locations) on the back (5.times.5 cm) of each dog. The drug was
applied in a dosing volume of about 4 .mu.L/cm.sup.2 once daily
(except on Saturdays, Sundays and holidays) for 3 months.
[0094] Evaluation Items:
[0095] 1) Skin condition: Gross observation and observation with a
video macroscope
[0096] 2) Histology: Observation of histological changes in biopsy
samples by H-E (hematoxylin-eosin) stain (general staining), van
Gieson's stain (staining of collagen fiber) and Weigert's stain
(staining of elastin fiber)
[0097] 3) Skin thickness: Using H-E stained tissue samples, changes
in thickness of the epidermis (excluding the horny layer) were
studied (only the location treated with FK706 0.2%, which showed a
marked improvement in skin condition, was evaluated)
[0098] Results:
[0099] 1) Skin Condition (Remission of Wrinkles)
[0100] (Gross Observation)
[0101] No. 8807: In the elimination or remission of wrinkles, the
order of efficacy was
FK706.multidot.0.1%>FK706.multidot.0.02%>FR134043.mult-
idot.0.2%=FR134043.multidot.0.02%. This finding of remission was
accompanied by improvements in skin texture (fineness, handle) and
paralleled depigmentation of the skin (change to fair.about.rosy
skin).
[0102] No. 8808: The order of efficacy was
FK706.multidot.0.2%>FK706.mu-
ltidot.0.02%>FR134043.multidot.0.2%=FR134043.multidot.0.02%.
[0103] In neither dog was found a side effect.
[0104] (Observation with a Video Macroscope)
[0105] The findings were substantially identical to the results of
gross observation.
[0106] No. 8807: The order of efficacy was
FK706.multidot.0.2%>FK706.mu-
ltidot.0.02%>FR134043.multidot.0.2%>FR134043.multidot.0.02%
[0107] No. 8808: The order of efficacy was
FK706.multidot.0.2%=FK706.multi-
dot.0.02%>FR134043.multidot.0.2%=FR134043.multidot.0.02%.
[0108] 2) Histological Findings
[0109] Van Gieson's Stain:
[0110] Neogenesis of a thin layer of collagen fiber was found
across the boundary between the epidermis and dermis in the case of
FK706.multidot.0.2% (Dog No. 8807)
[0111] Weigert's Strain:
[0112] Growth of elastin fibrils (suspected to be oxytalan fiber
which is said to disappear as increasing age) was found across the
boundary between the epidermis and dermis (Dog No. 8807).
[0113] 3) Evaluation of Skin Thickness
[0114] The thickness of the epidermis was increased significantly
(p<0.05 in both Dog 8807 and Dog 8808).
[0115] No. 8807: 17.0.+-.1.9 .mu.m.fwdarw.26.8.+-.5.9 .mu.m
(mean.+-.SD)
[0116] No. 8808: 24.6.+-.3.6 .mu.m.fwdarw.42.0.+-.9.0 .mu.m
[0117] The composition for prophylaxis and therapy of dermal aging
as provided by this invention is usually applied in the form of a
preparation for external application (e.g. lotion, ointment, patch,
liniment, aerosol, suspension, emulsion, etc.) or an external
powder (e.g. an enzyme facial cleanser) but may also be used in the
conventional pharmaceutical dosage forms such as powders, fine
granules, granules, tablets, sugar-coated tablets, parenteral
preparations, inhalants, microcapsules, capsules, suppositories,
solutions, syrups and so on. Furthermore, it can be used in such
formulations as facial cleansers, facial wash-off/emolient
preparations, and bath preparations. Where necessary, a diluent or
disintegrator (e.g. sucrose, lactose, starch, crystalline
cellulose, low-substitution-degree hydroxypropylcellulose,
synthetic aluminum silicate, etc.), a binder (e.g. cellulose,
methylcellulose, hydroxypropylcellulose,
hydroxypropylmethylcellulose, polypropylpyrrolidone,
polyvinylpyrrolidone, gelatin, gum arabic, polyethylene glycol,
etc.), a coloring agent, a sweetener and a lubricant (e.g.
magnesium stearate), among others, can be additionally dispersed in
such formulations.
[0118] The level of use of the composition for prophylaxis and
therapy of dermal aging according to this invention depends on the
species of substance used, symptoms and other factors but,
generally speaking, the recommended dose of an external dosage
form, in terms of the concentration of the substance having human
leukocyte elastase inhibitory activity or a pharmaceutically
acceptable salt thereof, can be judiciously selected from the range
of about 0.001.about.20%, preferably about 0.01.about.10%.
* * * * *