JPS58162562A - Lhrh nonapeptide and decapeptide analog - Google Patents

Abstract

(57) [Summary] This bulletin contains application data before electronic filing, so abstract data is not recorded.

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to novel and highly potent nonapeptide and decapeptide derivatives of LHRH.

Luteinizing hormone (LB) and follicle stimulating hormone (FSH)
) is a release system produced in the hypothalamus.
g) Released from the pituitary gland under the action of the hormone LklltH. Lli and FSH act on the gonads to stimulate the synthesis of steroid hormones and the maturation of gametes. The pulsatile release of LHRH and the consequent release of LH and FSH regulates the reproductive cycle in livestock and humans.

Furthermore, LHRH influences the placenta and directly the gonads to stimulate the release of chorionic gonadotropin (46G).

Antagonists of LHRH have been shown to be effective in regulating Lee precision. Such antagonists block h ovulation and suppress male spermatogenesis.

Related to these effects, J& and female accessory organs (aec
It is accompanied by suppression of the normal circulation of sex steroids of gonadal origin, including a decrease in mm of aaaory orgas. In livestock, this effect promotes weight gain in the presence of plenty of food, stimulates diapulsion in pregnant animals, and generally acts as a chemical sterilizer.

The natural hormone that releases the hormone LHRE is a decapeptide consisting of naturally occurring amino acids (having the L-conformation, except for the achiral (aehiraL) amino acid glycine). Its sequence is as follows: Pro -Gly-NH.

Many 7 analogues (α9αlog) of this natural product η
have been studied, and a large number of these have shown that K has insufficient biological activity for clinical utility. Certain selected modifications have been shown to reduce the effects of biologically active agents. Koi also costs 1 and the effect improvement is 6th place IJ
! It is obtained by changing 4 groups from G111 to D-amino acids.

In addition to agonists, analogues that are competitive antagonists to LHRIi have been produced; all of these require deletion of the histidine residue at the 2-position or the addition of li1.
e.

176:933 (1972). Generally, the D-amino acid present at that position in the sequence appears to give the best activity; Rags, R, W, A, at al.
, J.

Mad, Chmtn,, 1? :1016 (1974
).

We show that adding a modification at the 6-position, which is linked to the above-mentioned agonist activity, without modification at the 2-position, increases the antagonist activity of the analogue modified at the 2-position. Bgatia, C, w, atall,
J, Mad, Chat), l 8: 12
47 (197B) HRiver, J. at
al, , P#gtidgs, 1976, 4
2? p. Editiovra da L'Uns
Bruzallgs, Balg
isvm (1976).

On the basis of the above-mentioned two main modifications that make LIi and RIi antagonists a useful family, the historical improvement in anti-antagonist activity has already been demonstrated in the modified peptides at the 1, 3 and 3 positions. Or 10-position modified KL tute m lead L5 @ CQ, 1), Hlat al.

Pegtides 1978 #12.462,
k;ditio%#da l'U*i*aratt
g dg Hr%zmL1ms, BaLgi*
m (1976) Tomi R4viar, J.E., a.
tal, LifeSci, 23:869 (11
1? lS) 5Dstta, A, S,.

et al., Biocham tiiophys,
Rag, Comm5tn.

81:382 (197B), #%apprehensionpkdeim
a, J,.

at al, Bioehem, H4olphys
, Res, Comrun.

BS: TO~9 (1978).

N-acylation of the amino acid at the 1-position also helps in the tL tail;
.

Chemical Coo, (1979).

Since the antagonist functions by competing with LHRH against the appropriate receptor V, the natural peptide II-LJ+
In order to do so, high dosages of these compounds are in order.

In this regard, it would be particularly desirable to produce antagonists with a very high degree of potency.

The known analog rod sets require relatively long-haired compounds, increasing the potential for serious problems for Takanobu and other creators.

Therefore, it exhibits a much higher degree of biological activity than previously described and is clinically safe for small objects and humans. There is a need to produce viable LHRH antagonists.

In the present invention, the dalysin residue at the 6-position can be replaced with a carbocyclic or multiple lI
& cyclic side chain and a series of non-naturally occurring 17-lipophilic amino acids by 1)K to facilitate substitution at the 2-position (i.e., conversion of the peptide into an antagonist system). shall be valid. The present invention relates to novel and highly useful nonapeptide and decapeptide analogues of LHRli. History 1
．． Improvements by substitution at the 3 and/or 10-positions are disclosed herein. Furthermore, the present invention relates to various uses of these compounds and pharmaceutical compositions therefor. A set of aspects of the present invention encompasses methods of making the novel electrical compounds described above.

The present invention is based on the formula % (1) [wherein, Based on 1. I! - a carbocyclic aryl-containing group selected from the group consisting of phenyl, naphthyl, anthryl, l-fluorenyl, phenanthryl, biphenylyl and benzhydryl; or h) substituted with three or more linear lower alkyl groups; cyclohexyl, perhydronaphthyl, perhydrobiphenylyl.

Perhydro-2,2-diphenylmethyl.

and adamantyl; or C) the following structural formula ζ in which AI and l/ are independently selected from the group consisting of hydrogen, lower alkyl, chlorine, and bromine, and GFi is selected from the group consisting of nitrogen, sulfur, nitrogen and sulfur.

a heterocyclic aryl-containing group selected from the groups represented by;

A is L-pyroglutamyl, D-pyroglutamyl, N-acyl-L-prolyl, N-acyl-D-prolyl, N-
Acyl-1D-IJ f ) Phanyl, N-acyl-
An aminoacyl residue selected from the group consisting of D-phenylalanyl, N-acyl-D-9-no-10-phenylalanyl, and UN-acyl-X, where X is as defined above.

BflD--yenylalanyl% D-p-no\rophenylalanyl, 2.2-y phenylglycyl, and an aminoacyl residue selected from the group consisting of X), where X is as defined above Yes.

G is L-)Lyptophanyl, D-Tryptophanyl, D
An aminoacyl residue selected from the group consisting of -phenylalanyl, and UX, where X has the same meaning as above.

b" is glycinamide or f-4-Nnoi-R'.

where R1 is lower alkyl, cycloalkyl, fluoro-lower alkyl or t-IJH-C-NH-R, where R3 is hydrogen or lower alkyl. This invention relates to nonapeptide and decapeptide Tsuru conductors. W at residue 01 of the L-histidyl residue present at the 2-position of LfiRH as known and defined in the present specification.
The conversion is at public expense to convert the peptide into a L Ii RH antagonist. Glycyl Th1k at position 6 of LHRH
Substitution of hi, defined as X, with one of hi significantly improves the antagonist effect.

The In,m groups disclosed herein at the 1.3 and 10-positions are of further assistance in enhancing antagonist IR activity.

Abbreviation and foot prosthesis! 8, and to facilitate the tfA version of the present invention, IUPAC-IUB Biochemical Nomenclature Committee.

Biochgrniatry, 11, 1726 (
Conventional abbreviations are used for the various common amino acids as generally accepted in peptide chemistry as recommended by (1972). These represent L-amino acids, with the exception of achiral amino acids, and further exclude achiral unnatural or natural amino acids, otherwise designated as D-, and also LflRHI/
1.1.2.3.6 and 1 for those normally found
Amino acids substituted at position 0 are also excluded. The sequences of all peptides referred to in this BA specification are delivered according to generally accepted conventions, with the N-terminal amino acid on the left and the C-terminal amino acid on the right.

The abbreviation "NAc", when used as a prefix to an amino acid residue, refers to an N-acylaminocy residue.

Certain other abbreviations are also useful in describing the invention.

The present invention uses substitutions with non-naturally occurring amino acids. Among these, those commonly used are as follows.
naphthyl) -D-NcLl(IID-alanyl3-(2-naphthyl)-D-ryal(2)D-alanyl a-(9-hao 7x=l) D-g-ha0-
Pha-D-alanyl a-(t-fluorophe L)-p-to'-Pk
a-yl)-D-alanyl 3-(p-chlorpheny D-p-CL-Phal)-D-alanyl 3-(W-ob-yx D-p-Br-P
hanyl)-D-alanyl 3-(2,3,4,5,D-Me5Pha6-bentamethylphenyl)-D-'alanyl 3-(2,4,6-) D-TMPlimethylphenyl) - D-alanyl 3-(benzimidazo D-BIA-2-yl)
-D-alanyl 3-(4,5,6,7D-TBA-tetrahydrobenzimidazol-2-yl) -D-alanyl Other residues used in the present introduction are abbreviated to 2,2-diphenyldalicyl. DPG, and 1 -ISJHC-NliR'' (wherein R3 is hydrogen or lower alkyl), - No. aza G l 1.IIVHR, and group -NHCH, C1i.

is a group represented by the abbreviation -NHEt.

For further convenience, the amino acid sequence of LHkH is (pio)Gls
t, -H4s-Trp-8ar-7'yr-(yly-
Since it is shown as , is abbreviated by indicating its position as a shoulder, followed by LHRH as the parent body.

That is, for example, G111 at the 6-position is D −N a l (
Sequence 8910 in which H4s at position 21 and 2 is replaced with n-ph- is (D-Ph a”, D-Na J(21・)buR
B and the sequence hAc-Pro-D-Pha-Trp-8er-Tyr
-D-HaJ (21 is (NAc-Pro”, D-P
ha”, D-Na L (2i.

−NHEt”) LHRH.

As used in this EJI specification, a "confectionery-acceptable salt" refers to a salt that retains the desired biological activity of the compound, but does not impart undue toxicity. Examples of such salts include (α) inorganic acids such as hydrochloric acid, hydrobromic acid, bk
Acid cycloaddition I organic acids produced from acids, phosphoric acid, nitric acid, etc. An example is vinegarff! 2. Oxal vinegar, tartaric acid, succinic acid, maleic acid, fumaric acid, gluconic acid, citric acid, malic acid, alcorbine e, benzoic acid, tannic acid, pamoic acid, alginic acid, polyglutamine, naphthalene sulfonic acid, Polyvalent cations such as nesium, aluminum, copper, cobalt, nickel, cadmium, etc., or N, Δ'-sopenzyl ethylene or a combination of (C) (Q) and (b), such as a zinc tannate salt.

What is "lower alkyl" as used herein?

iIl linear or branched unsaturated hydrogen group having 1 to 4 carbon atoms 1 such as methyl, ethyl, n-propyl, isopropyl, bamboo-ethyl, isobutyl, 8#C-butyl and t
# Regarding it-butyl; the term "cycloalkyl group" refers to broken saturated hydrogen groups having 3 to 6 carbon atoms, such as cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl; the term ``fluoro-lower alkyl'' refers to , a lower alkyl group in which one or more hydrogen atoms are substituted with fluorine.

For example, trifluoromethyl, pentafluoroethyl, 2
, 2,2-trifluoroethyl, etc.

As used herein, "naphthyl" refers to l- and 2-naphthyl; "Anthryl" refers to l-12- and 9-
Anthril 5-mi; "Fluorenyl" 1-2-13-
Includes 14- and 9-fluorenyl; "phenanthryl" includes 2-13- and 9-phenanthryl; and "adamantyl" includes 1- and 2-adamantyl.

An "N-acyl" amino@residue is an aginoic acid residue in which the amino group 11 is linked by an amide bond to a carboxylic acid residue; this carboxylic acid residue is a lower alkylcarboxylic acid or benzoic acid residue. It is.

Compounds that are preferred embodiments of the present invention are:

A is L- (pyro) GLsb, D- (pio) Gji,
NAe-D-Phg, NAc-4J-JVajll
, ΔAc −D−Naj12! , NAc-D-p
-No10-Pha, NAe-D-MgsPha%N
Ac-D-TMP, NAc-D-BIA.

B is selected from the group consisting of D-Pha, D-p-Ul-Ph- and DPG; C' is selected from the group consisting of NAc-D-TBA, and I'dAc-L-Pro; L-Trp, DJ'rp, D-Phg, D
-Nag[11, D-Nag (21% D-p-halo PAg, L)-Ma5Pha, D-1'MP,
Select a death consisting of D-HIA and D-1'HA; X is D-IJal(1), 1)-Nal(2), D
-p-Hao-Pnichi 5D-Ma57'A#% D-TM
The number is selected from the group consisting of P, D-BIA and D-TBA, and b is selected from the group consisting of G I INH and AzaGlyNHH.

A more preferable specific example is as follows.

A is L-(pyro)Glu, D-(pyro)Gis, N
Ac-1)-Ph# and □IVAc-L-Pro Kara fx; 6e+, twist? Found out.

Novo is D-Pha, l)-'p-halo ph- and DP
The group consisting of G is chosen.

C is selected from the group consisting of L-Trp, D-Trp, D-Nag (11) and D-N a l (2!).

λ is D-Nal (21, selected from the group consisting of D-TMP, D-BIA and D-TBA).

E is G l 11NB! and AzaGlyNHR.

A specific example suitable for the east is.

A is z, -(pio)Gin, D-(pio)Glw, NA
selected from the group consisting of a-D-Pha, and NAc-L-Pro; B is D-Pha%D-p-C; 1-Pha,
and DPG; t: KaL-Tr p zly=Da'llr p,
The group consisting of D-Ha ltt+ and D-Nal 12+ is selected;
lyNHR.

A most particularly preferred embodiment is the following retroactivity: (pio)Gl
s-DPG-TrlJ-8g r-Tyr-L)-1
Val 121-Law-Arg-Pro-GLyN
H,, abbreviated as (L)PG”, D-)Ja l 121@
) LtiRH; (pio)Gls-1)-Pha-D-
Tr7p-Sar-Tyr-D-NaL (21-Leg
-Arg-Pro-GLyNH, (D-Phg
”, D-Trp Hatake, D-Na tt21
') LIiMHS(Pio)G l w-D-Ph
a -D-ha Jf2+-8g r-1'yr-D-
Nag(2)-Law-Arg-Pro-GlyhHb
Abbreviated as (D-Ph-heavy, D-NaLi3N,
D-Na1(2F)LIiRH; NAe-L-Pro-D-Pha-L)-1'rp-S
ar-Tyr-D-Nal(2+-Law-Arg-P
ro-GLyNH,,%(NAe-L-Pro”
, D-Pk-weight % D-1'rp Hatake,
D-Nal 121”) LHRHB and NAc-L)-Phg-D-yp-CL-Phe-
D-Trp-8# r-Tyr-1)-hta l
121-Law-Arg-Pro-GIINH,
, abbreviated as (NAc-D-Pha", D-'I-ni l-Ph gfu, l)-'l'r p",
L)-Ha l (2+') LHRH; and 1
The 0-position glycinamide is azaGlyNH.

Deli! The corresponding peptides have been tested.

In all of the above embodiments, the compounds may also be prepared as the corresponding H pharmaceutically acceptable salts.

In all embodiments where NAc is present, an N-acetyl residue is a preferred embodiment thereof.

The compounds of the present invention, and especially their salts, exhibit low potency and long-lasting LHR1i antagonist activity.

The axial measure of potency (9o t aney ) is C
orbin, A., and Battia, C.)%',
, k;ndocri*aRan, Comtnm
, 2: I by the method of l (1975)
Ability to inhibit ovulation in rats as rated Ph.
alpa, C1P, et al., Endocrinolog
y.

100:1526 (197?), the ability to completely block LH release and ovulation in rabbits.

Other bioassays used for LHRH antagonism and for compounds of the invention (binα5says)
is as follows: (α) In vivo in rats. Blocking of LHRH-induced fc, 7I' sit and LH release; Vil
chgt-Martingt, J.A., at al.
, bndocrinology.

96:1130 (1975>, and (b) Radioimmunoassay [Valg, 14', gtal
, .

k;ndocritrology, 91:56
2 (1972)], the dispersed fc subcephalic rod Ni1 lobe: inhibition of the release of LH and FSH by j-treated cultures.

Antagonist action and usefulness Primary 41h4 t-E from the antagonist action of this compound
- Contraception of female females that can be carried out; - Suppression or delay of ovulation; Induction of parturition; Synchronization of ovulation; Suppression of oestrus; Growth promotion in difficult animals; - Roux orysis ( 1utaolyais), induction of menstruation; - M initiator during the first 3 months of pregnancy; - Treatment for endometrial stria; Treatment for breast cancer and cysts; - Multilamellar ovary syndrome
treatment for uterine cancer; treatment for benign prostatic hyperplasia and prostate cancer; - contraception in males; treatment of diseases based on excessive gonadal hormone production in either sex; function in male nutritional-producing animals. - Control of bleeding during proestrus in dogs; Control of signs of withdrawal. Aspects of the present invention with respect to special uses for the above-mentioned compounds relate to their usefulness, in particular in the prevention of ovulation and treatment of endometriosis in females, and in the prevention of spermatogenesis and treatment of prostatic hyperplasia in males. be.

In practicing the methods of the invention, a subject in need or desiring treatment is effectively administered a compound of the invention or a pharmaceutical composition containing the same.

These compounds or compositions may be administered in a variety of ways depending on the intended use: oral, parenteral (including subcutaneous, intramuscular and intravenous), intravaginal (particularly for contraceptive use), rectal. Administration can include intrabuccal (including tongue administration), transdermal or intranasal administration. The most appropriate mode of administration for a treatment depends on the active ingredient, the subject, and the judgment of the practitioner. The compounds or compositions can also be administered in sustained release, 11-pack, or insert formulations, such as those described above.

Generally, for the aforementioned uses, the active ingredient will be about 0.01-
10/Kf/day, preferably about 0.1-1. OW
It is advantageous to administer in an amount of 9/Kf body weight/day. This administration can be done once daily in divided doses or in sustained release form to obtain the most effective results.

Dosages and dietary regimens for administration of the present compounds and compositions will depend on the needs of the individual subject being treated.

The type of treatment, degree of suffering or need and of course the judgment of the practitioner! ! Be sure to rely on lr. Generally administered parenterally.

Moreover, it requires less dosage than other administration methods that depend on absorption capacity.

A historical aspect of the present invention relates to pharmaceutical compositions containing a compound of the present invention as an active ingredient in admixture with a pharmaceutically acceptable non-toxic support. As mentioned above, such compositions can be administered non-tactilely (subcutaneously, intramuscularly or intravenously).
For administration, 1% i ffJ in the form of a liquid solution or suspension.
For internal or rectal administration, especially in the form of creams and herbal medicines; orally or! For intranasal administration, especially in the form of tablets or capsules; or for intranasal administration, especially as a powder.

It can be prepared in the form of truth drops or aerosols.

The compositions can be administered in one convenient half-dosage form and can be administered by methods well known in the pharmaceutical arts, such as Rgtnington'
aPharmacastical 5science.

Mack Publishing Company,
Haaton, F.A.

1, 1970. Formulations for parenteral administration include sterile water or saline, polyalkylene glycols as usual excipients. The oil may contain polyethylene glycols, oils of chemical origin, hydrogenated naphthalenes, and the like.

Formulations for intravaginal or direct administration, such as herbal medicines, may contain as excipients such as polyalkylene glycols, petrolatum, cocoa butter, etc. as excipients. The formulation for inhalation administration is solid and ψ
Rie can contain lactose as an excipient,
Alternatively, for administration in the form of drops, it may be an aqueous or oily solution. For buccal administration 1 Typical excipients include sugar, calcium stearate, magnesium stearate, pregelatinized powder, and the like.

It is particularly useful to provide a subject with a compound of the invention in % administration over an extended period of time, for example once a week to once a year!
I want it. Various sustained release, depot or insert dosage forms are available. For example, the dosage form may contain pharmaceutically acceptable non-toxic salts of the present compounds with low solubility in body fluids, such as Fi(α)Gj
Durable acids such as phosphoric acid, sulfuric acid, citric acid, tartaric acid, tannic acid, pamoic acid
d), alginic acid, polyglutamine l! #, naphthalene mono- or di-sulfonic acid, polygalacturon I
I <90LVQα1acturonicαaid)
Acid addition salts with etc.; (b) polyvalent metal cations;

Examples include zinc, calcium, bismuth, and NonojJum.

Also #'2. with magnesium, aluminum, copper, cobalt, nickel, cadmium, etc. For example, salts with organic cations generated from N,N'-dipenzoleethylenediamine or ethylenesoamine; or
One example of the combination salt of 6) may include a zinc tannate salt. Additionally, a compound of the present invention, or preferably a relatively insoluble salt such as those described immediately above, can be formulated into a compound such as aluminum monostearate with plum oil, which is commonly used for injections. Particularly suitable salts include zinc salts, zinc tannate salts, pamoate salts, and the like. of other ridges.

An example of an injectable sustained-release depot formulation is U.S. Automobile Specialty Ff No. 1? ? Sweetening compounds or salts dispersed or encapsulated in slowly degrading, non-toxic, non-antigenic polymers such as those described in US Pat. More particularly for use in animals, compounds such as those described above or preferably relatively insoluble salts may be incorporated into cholesterol matrix silastic pellets (ailaatia p.
may be formulated in alletm). Historical sustained-release, depotable or intercalated formulations 1, such as liposomes, are well known in the literature;
Dgkk-de, Ino,.

New York, 1 9 7 g.

References regarding compounds of the LHRH type are, for example, US Pat. No. 4,010,125.

Polypeptides of the invention can be synthesized by techniques known to those skilled in the peptide art.

Many existing techniques are based on solid-phase peptide lactate, J. M., Stgwart and J., 77, Young.
.

5olid Phase Peptirh 5ynth
esis, h, HlFreeman Co, ,
Sa% Fratseisco, 1969.

Prgaa (New York), 1973. and II 5chroda for the classical solution synthesis method.
ra%dA.

Lubka, The Pgptidea, Vow,
l, Acαd-inertia-CPress (New Y
ork), 196 B is superior.

In general, these methods consist of sequentially adding one or more amino acids or appropriately preserved amino acids to a growing peptide chain. Usually, either the amino or the t-ruboxyl group of the first amino acid is anchored with a suitable anchoring group. This protected amino acid or deconductorized amino acid is then attached to an inert solid support, or ii the next amino acid with one (amino or carboxyl) group suitably preserved is attached to the amide bond. It can be utilized in solution by addition under conditions suitable for formation. The protecting group is then removed from this newly added amino acid residue, followed by the addition of the next amino@ (suitably a protected amino acid) and the process repeated. After all desired amino acids have been attached in the proper order.

Any remaining protecting groups (and solid support) are subsequently or simultaneously removed to produce the final polypeptide. According to a simple modification of this common method.

For example, by combining a protected tripeptide with a suitably protected dipeptide (under conditions that do not racemize the asymmetric center) and, after removal of the protective @ group, forming a pentapeptide, one or more amino acids can be combined. can be added to the growing chain all at once.

Particularly preferred methods for producing compounds of the invention include homologous peptide synthesis methods.

In this particularly preferred method, the α-amino function of the amino acid is capacitated with an @ or a tomb-sensitive group.

Such protecting groups are stable under the conditions of production of the peptide, but are easily removed without racemization of either the intercalation of the growing peptide chain or the chiral center incorporated therein. You should have good qualities. Appropriate Houmoto Fi
t-Butyroxycarbonyl (BoC), penpoxoxycarbonyl (Cbg), biphenylisoglubiloxycargenyl, t-amyloyloxycargenyl, isolylnyloxycargenyl? Nyl, α, at/methyl-3,5-dimethoxybenzyloxycarbonyl, O-nitrophenylsulfenyl, 2-cyano-1-1 thyroxycarbonyl, 9
-Fluorenylmethyloxycarbonyl is t-butyroxycarbonyl (BOC) in %.

Particularly preferred side@4 stock groups are dinitro to arginine, p-toluenesulfonyl, 4-methoxybenzenesulfonyl, C; bz% Boc and (j7 damanthyloxycarbonyl; Brompencyl, xycarbonyl, 2゜6-nochlorobensol,
Isopropyl, cyclohexyl, cyclopentyl and acetyl; 7 benzol and tetrahydropyranyl for serine; about 7 benzol and tetrahydropyranyl for histidine: bensol, p-toluenesulfonyl and 2,4-dunitrophenyl.

The C-terminal amino acid gold is attached to a suitable isotropic support. Solid supports iL useful in the synthetic methods described above.

It is a substance that is inert to the reagents and reaction conditions of the stepwise condensation and radical reactions and incurable to the medium used. Suitable facepiece supports L1 include chloromethylpolystyrene-divinylbenzene polymers, hydroxymethyl-polystyrene-divinylbenzene polymers, and especially chloromethyl-polystyrene-1tidivinylbenzene members. G of the compound
In the special case where the -terminus is a glycinamide, a particularly important carrier is P, Rivaillg et al., H
alv, Chim, Acta, .

54.2772 (1971), it is the benzhydrylamino-polystyrene-novinylbenzene Chlormethylpolystyrene-divinylbenzene to v8om (D k combination II, N
Amino acids, especially IJoc-amino@purple, as cesium, tetramethylammonium, triethylammonium, l,5-zoadebicyclo-5゜4.019ndec-5-ene, or similar salts, and in ethanol, acetonitrile, In N,N-methylformamide, etc., cesium salt on the bottom and chloromethyl resin in the cesium, and about 40 to 60 U on the outside,
Q4 is produced by reacting at about 50C for about 12 to 48 hours, preferably about 24 hours. N-Boa-amino acid benzhydrylamine a
Coupling to J IIK via intervening coupling of A/, AI'-socyclohexylcarposoimide (77(a(J)/1-hydroxybenzotriazole (HBi'), dichloromethane or l) ME', Preferably in an intoxicant of dichloromethane [1], about lθ~50C1, preferably 25
C. for about 2 to about 24 hours, preferably about 12 hours. Conjugation of serially stored aminodes can be carried out in automatic polypeptide synthesizers well known in the art. What is the difference between N and Erosion? IJ may be carried out at about room temperature in the presence of trifluoroacetic acid in methylene chloride, hydrogen chloride in dioxane, hydrogen chloride in acetic acid, or other strong acid solutions, preferably a 50% solution of trifluoroacetic acid in dichloromethane. can. Each conserved amino acid is preferably introduced in an excess of about 25 molar, with a bond of about 1? , is nochloromethane, zochloromethane/D
The binder is usually 1) CC in dichloromethane, but N,N'-SOIN-
Propyl carbodiimide and others 1) Carboimide may be used alone in the presence of di-μH H1'% N-hydroxysuccinimide, other N-hydroxyimides or oximes. other.

Protected amino acid active esters (eg p-nitrophenyl, pentafluorophenyl 7) or symmetrical anhydrides may also be used.

At the end of the solid phase assembly, the fully protected polypeptide is removed from the resin. If the assembly to the resin support is in the benzyl ester form, the quaternary can be prepared by aminolysis with an alkylamine or fluoroalkylamine in the case of peptide F with a zololine C-terminus, or by aminolysis with an alkylamine or fluoroalkylamine for a peptide with a glycine C-terminus. For example, in ammonia/methanol or ammonia/ethanol, the amine content is 1'l'f, about lθ~50C1, preferably about 25U, about 12~2
41-'F, preferably about 18 hours. In addition, peptides can be used, for example, ester paternity with methanol, carp, etc.
It can be removed from the resin by aminolysis. The preserved peptide can be purified at this point by silica gluchromatography.

Removal of the s(:j & stock group) from the polypeptide by treatment with anhydrous liquid hydrogen fluoride, for example in the presence of anisole or other carbonium scavenger.

by treatment with arsenic/pyridine lead bodies, by treatment with tris(trifluoroacetyl)boride and triple oroacetic acid, by reduction with hydrogen and palladium on carbon or evolivinylpyrrolidone, or by reduction with i-form sodium in ammonia. Preferably from about -1O to +IOC
, preferably using liquid hydrogen fluoride and anisole for about 15 minutes to 1 hour f+U, preferably about 30 minutes at a temperature of about oH. In the case of glycine-terminated peptides for benzhydrylamine side bends, by using liquid hydrogen fluoride and anisole'fr as described above.

Both steps of resin cleavage and Il>t, &- groups can be combined in one step. The principally secured polypeptide is then processed in a series using any or all of the following types: Purified by chromatography: acetate salt is ionized by oxidation; unconducted polystyrene-subvinylbenzene (Sarie is Amh
Hydrophobic absorption chromatography using carboxymethylcellulose; ion exchange chromatography using carboxymethyl cellulose;
Partition chromatography or cocurrent separation according to 25; performance liquid chromatography (HPLC), especially reverse phase HPLC with octyl- or octadecylsilyl-silica-bonded phase column packings 1, 2, 3 or 6- When assembled with a racemic amino acid in position, the diastereomeric nonapeptide or decapeptide final product is separated and the desired peptide incorporating a D-amino acid in the appropriate position is separated in one chromatographic step as described above. It is also ladle-made.

Separation of peptides with a C-terminal azadalicin amide is preferably carried out by the classical pezotide bath synthesis method using known peptide intermediates. This is Example 3
It is described in more detail in .

That is, at other R points, the present invention is as follows.

The stock base and optionally covalently attached solid support are removed from the protected polypeptide to obtain a compound of formula (R) or a salt thereof, and optionally, a compound of formula (11) is prepared. (h) converting the salt of the compound of formula (1) into a pharmaceutically acceptable salt; or (C) converting the salt of the compound of formula (1) into a pharmaceutically acceptable salt; (1) Free /
Compounds of the present invention and their production W comprising decomposition into IJ peptides: A method for producing a scientifically acceptable salt.

The following examples describe how one skilled in the art may more fully understand and put into practice the present invention. The v6 examples should not be viewed as limiting the scope of the invention, but are merely representative illustrations of the invention.

Production example A Anhydrous ethanol (distilled from magnesium ethoxide) 0
．． into an oven-dried flask containing 1 t.

1.52 f of sodium gold maple was added. When the evolution of hydrogen ceased, 10.21 P of 2-acetamido-2-cyanide [% ethyl] and 13.26 t of 2-20-methylna-7-talene were added to the solution. The solution was refluxed for 1 hour. Heated and then cooled. The ethanol was removed under reduced pressure and the residue was taken up in ethyl t-acetate. The organic phase was diluted with 5 ml of water.
Washed twice with Q+d and once with 50 m of saturated sodium chloride, and dried over magnesium sulfate. The solution was filtered and the W4 medium was distilled off under reduced pressure.

The residue was hydrolyzed by refluxing for 2 hours in 100% concentrated hydrochloric acid.

The hydrolyzed and decomposed mixture was cooled and the crude product precipitate was filtered. The crude product was redissolved in 0.5 t of hot water treated with activated carbon and treated with hot water, and the solution p1! was adjusted to 6 with concentrated ammonium hydroxide. The precipitate was crushed with 0'i force and transferred under vacuum to obtain pure 3-(2-
11.3f of D,L-alanine (S7til)-D,L-alanine was obtained.

Instead of 2-bromomethylnaphthalene, the chemically used l-bromomethylnaphthalene.

9-10 methyl anthracene.

9-bromomethylfluorene.

2-bromomethylfluorene, 2-7P rommethyl anthracene.

l-bromomethylanthracene.

α-Chlorindurene.

4-bromomethylbiphenyl, l-bromomethylamemankun.

3-bromomethylphenanthrene.

l-chloromethyl-2,4,6-tri(n-butyl)
benzene, and 1-coolmethyl-2,3,4,5,6-pentamethylbenzene.

The primary amino acid 3-(1-naphthyl)-D,L-alanine, melting point 185
~187C.

3-(9-anthryl)-D,L-alanine, 831 points 290C (HCl salt).

3-(9-fluorenyl)-D,L-alanine.

3-(2-fluorenyl)-D,L-alanine, 7 points 264-269U.

3-(2-anthryl)-D,L-alanine.

3-(1-anthryl)-D,L-alanine.

3-(2,4,6-drimethylphenyl)-D.

L-alanine, melting point 235-237C.

3-(4-biphenylyl)-D,L-alanine.

One point 290υ.

3-(1-adamantyl)-D,L-alanine.

3-(3-7 dinanthryl)-D, L-alanine, 3-
(2,4,6-tri(%-butyl)phenyl)-D
, L-alanine, and 3-(2,3,4,5,6-pentamethylphenyl)
-D, L-alanine.

were obtained respectively.

Production Example B 1.1-diphenylethylene 1B, 29. Mel α-methoxy N-benzyloxycar? Nilglycinate 2F
h, 3f, and 1.5 t of 2-naphthalenesulfonic acid,
The anhydrous benzene 300-medium biao solution was refluxed for 2 days. The crude product was then converted from CB, ct, to C; H, Cl, /Et
It was purified on a silicic acid column using a # medium gradient of OAe (1 g: 1).

This purified methyl 2-(1-(2,2-zo7-enzylenyl)]-]A'-benziloxica was used to convert nildarisine into KOH1α9F in 10% aqueous methanol 35 (ld).
was hydrolyzed to the corresponding acid with a solution of The obtained assembly machine was mixed with 95% ethanol (10%) containing 1 Bm in concentrated H.
3-(2
,2-diphenylmethyl)-D,L-alanine 2.4f
I got it.

Production Example C 3-(2-naphthyl)-D,L-alanine 129f(D
, IMNaOH120-medium solution was cooled over acetic anhydride. Adjust pH to 2 with concentrated HUE.

The collected precipitate was filtered. This solid was recrystallized from 60% aqueous ethanol to yield N-acetyl-3-(2-naphthyl)-D,L-alanine 1L2f.

Anhydrous methanol 2 of this N-acetylamino acid 15f
Boron trifluoride Nieteller) was added to the 40d medium bath and the mixture was refluxed for 1 hour. Evaporate the alcohol and add 200ml of water.

The solution was extracted with ethyl acetate. The organic layer was washed with aqueous base and acid, dried over MQSO, filtered, and evaporated to give an oil. This oil was crystallized from ethyl acetate/hexane to obtain methyl N, melting point 79-80C.
-acetyl-3-(2-naphthyl)-D,L-alanine) 142t was obtained.

Chemically equivalent to 3-(2-naphthyl)-D,L-alanine.

3-(l-su7tyl)-D, L-alanine, 3-(2-
fluorenyl)-D,L-alanine.

3-(2-anthryl)-D,L-alanine.

Methyl N-acetyl- 3-(l-naphthyl)-L), L-alaninate, mp 97.5-98C.

Methyl N-acetyl-3-(2-fluorenyl)-D,
L-alaninate, melting point 170-17 IC.

Methyl N-acetyl-3-(2-anthryl)-D,L
-alaninate, and methyl N-acetyl-3-(2,2-diphenylmethyl)-47,L-alaninate, melting point 113-114C.

I swallowed it.

Production example D Cy'methyl sulfoxide 300d, 1MkLC112
Methyl N-acetyl-3-(2-naphthyl)-D,L-alanine) a6 in a mixture of 0 ml and Btθ780-
The ro bath solution was treated with aIMKcl 3-medium enzyme subtiliain 3161119. The pH was then adjusted to 7 by automatic titration with 0.2M NaOH using a rasometer pHstat (ATAT).
maintained. After 30 minutes, 70 ml of Na0Ii solution was consumed and the hydrolysis was stopped. This bath solution was mixed with NaHCO, 12
The mixture was made basic with F and extracted with ethyl acetate. The organic layer contained methyl N-acetyl-3-(2-naphthyl)-D-anthunate. Eaves 6> Crystallization from ethyl/hexane gave a yellow solid with h points 80-81t,'.

This is firstly converted into free amino acids and then N
-Hoe Converted to amino acid: Methyl N-acetyl-3-(2-naphthyl)-D-7 laninate 2.5 t
NoroN l1C160-Medium bath liquid to 120-130C 3
The mixture was heated for a while and cooled to room temperature. The white precipitate produced was collected and 12A/H contained It, 050
from at, re-MJj by neutralizing with Δ1i40H to pH 6 and drying under vacuum to a melting point of 242-24.
1.2t of 3-(2-naphthyl)-D-alanine of 4C was obtained: (α) D” 2 &6 @(e-0-5-Ll
i@L Ol B) @3-(2-naphthyl)-D
-% of Alanine tAINaOH55#It, H1O10
#El! ! The solution was stirred in a mixture of 20 yd. 1
．． After 5 hours, 0.3v of di-tart-ethyl socarnate was added and the mixture was brought to room temperature. Solids were removed by p-filtration and solution F was concentrated at 50+dj.

The aqueous solution was adjusted to pH 804 with NaH804 and extracted with ethyl acetate.

The organic layer was washed with 54NaH8O water and saturated salt solution. The ethyl acetate solution was dried over magnesium sulfate, filtered and concentrated to give an oil. This oil was crystallized from ether/hexane to obtain N -Bo'c 3 with a melting point of 90-91C.
- (!-S7til)-D-AnsylBt was obtained; (
a)”-ateo (CmlIo, 8, M a OH)
.

Methyl N-acetyl-3-(2-su7nal)-ri, L
-Methyl N-acetyl-3-(1-naphthyl) is chemically used instead of anthunate.

L-Anthnate.

Methyl N-acetyl-3-(2-7'luorenyl)-L
), L-Anthunate.

Methyl N-acetyl-3-(2-anthryl)-ly, L
-anthunate, and methyl N-acetyl-3-(2,2-sophenylmethyl)-D,L-anthunate.

Replicating the steps above using i.b.r (and then the corresponding free amino acid followed by N-BoC-3-(1-naphthyl)-17-alanine.

Melting point 92-93C, [9)% 54.8° (C content 0.5° MAIES).

N-Hoc-3-(2-fluorenyl)-D-alanine, melting point 161-163c (decomposed), N-Bee-3
-(2-anthryl)-D-alanine, and N-Boa -3-(2,2-sophenylmethyl) -
D-7 Rani y% melting point 153-154c.

The N-Boc amino acid was obtained.

3-(2-su7thyl)-D-7 in the Pearl hydrogenation equipment
52:yo, 85 t, 100 t of 2M hydrochloric acid, and 0.85 t of Adams catalyst (PtO,) were added. bath liquid,
II, 601 b/in” for 20 hours in a bar hydrogenator. The mixture was heated to dissolve the white precipitate and filtered through diatomaceous earth. The solution was concentrated under reduced pressure and By freeze-drying from water, 3-(2-
perhydronaphthyl) -1J-alanine 0.82 as a white solid of 230-232c per point.

Konomono ik, lNNaOH12#;e, dissolved in a mixture of 5 parts of water and 15 parts of disxane, klgoo, 14V
and tart-butyl oxide) 0.85
? Processed with. 1 hour on OC and 2b on 25C “Stupid.

The suspension was filtered and concentrated to dryness under reduced pressure.

Dissolve the residue in water, wash with diethyl ether, and
Acidified to pH 2' with s'4. The defunct aqueous layer was extracted three times with ethyl acetate, the extracts were combined and MQS
O, t#, dried, filtered and concentrated to N-BoC-3-
(2-perhydronaphthyl)-D-alanine 0.75
t was obtained as a colorless oil.

This substance 6o, zr was dissolved in tetrahydrofuran 5-,
1ml fresh until the light yellow color persists! ! ll! It can be treated with diazomethane under OC. The reaction was stopped with acetic acid eF (1-), the solvent was removed and the residue was dissolved in ethyl acetate 751 and water 751
It was distributed between 5a+Z. The organic layer was diluted with 5% NaH (:03
% water, 5% l5ltxH804, water, saturated Δacl solution and dried over MQSO. The solution was filtered, concentrated under reduced pressure and placed on a preparative thin layer chromatography plate (75
0 thickness no silica 5'h, 20X20σ). The laminate was developed with dichloromethane/ethyl acetate (18/1) to remove the @ of the product. The silica glue from the production zone was filtered onto a glass filter in dichloromethane/ethyl acetate (
9:l) and concentrated solution F to give methyl N-BOO-
3-(2-perhydronaphthyl)-D-anninate 0
．． 1F was obtained as a light yellow oil.

This substance has 2 at the 2-position of the nohydronaphthalene nucleus.
It was obtained as a mixture of two isomers. These noastereomeric mixtures were prepared using ethyl acetate/hexane (1:9).
High performance liquid chromatography □(
Column: Lichrosorb silica column 60.5 micron) allowed separation and hydrolysis to the free acid N-Hoc-3-(2-perhydronaphthyl)-D-alanine.

In place of 3-(2-su7tyl)-D-alanine, use the chemically equivalent amount of rjf.

3-(l-su7tyl)-D-alanine.

3-(2,2-sophenylmethyl)-D-alanine, 3-(2,4,6-)limethylphenyl)-D.

L-alanine.

3-(4-biphenylyl)-D,L-alanine.

3-(2,4,6-)li(ethyl)phenyl)-
, D, L-alanine, and 3-(2,3,4,5,6-bentamethyl7enyl)-
D, L-alanine.

By repeating the above one stock using
c-3-(1-perhydronaphthyl)-D-annine.

N-hoe-a-(perhydro-2,2-sophenylmethyl)-D-alanine, 8'-Boc-3-(2,4,6-drimethylcyclohexyl)-, L-alanine.

N-Hoc-3-(perhydro-4-biphenylyl)-
D, L-alanine, N-Boc-3-(2,4,6-tri(n-butyl)
cyclohexyl)-D, L-alanine, and IV-BO
N-BoC amino acids of C-3-(2,314#5#6-pentamethylcyclohexyl)-D, L-alanine, respectively, were obtained.

The preparation was dissolved in a mixture of 4.9 ft of ethanol, 50 sd of distilled water, and 50 #110 of distilled water. The pB of this solution was brought to 7 with aqueous cesium bicarbonate. The solvent was then removed under vacuum.

After drying under vacuum for 18 hours, the residue was dissolved in anhydrous DMFtso.
- dissolved in Then chloromethylated polystyrene-1
% novinylbenzene (Merrifiald) resin (equivalent to 25 mmol of chloride) was added. The mixture was incubated at soC for 24 hours, filtered and the resin was
Washed sequentially with MF, water and ethanol. This resin was dried under vacuum for 3 i to obtain BaC-Gly-0-resin 2.
8.34ft, obtained.

Example 1 Boc-Glu-0-resin (Lab System
s company) 7.29F (0.8 mmol) was added. This resin was then sequentially loaded with amino acids according to the following synthetic nology: 1 root 1 HICl, 1 wash 1.5 min at C° - 4 Cfl, C - 3 washes 1 .5 minutes 5 10 tidoethyl 2 times 1.5 minutes Amine 7ch, ct.

6 Ch, Cal, wash 3 times 1.5
Minutes 7 N -BoC-ami L times Added noic acid bath solution 8N, N'-dicyclo 1 time Added hexylcal diimide bath solution 11 C1i, washed 11 times 3 times 1.
5 minutes 12 Ethanol wash 3 times L5 minutes 13 H, T, wash 31! jI
After 15 minutes, 1-13 completes the conjugation cycle for one amino acid and completes one reaction.E., Kais-de et al.

Anal, Biochgtn,,, 34.595 (
It can be known from ninhydrin η (1970).

2. Add the resin. ．． A 5 molar excess of each protected 1 was combined with the next amino acid and DCC. That is, the resin is Hoe-Pro-OB, 3.01FBoa-Arg in a continuous Vr binding cycle.
(Toshik)-OH, 599? Boa -Law-
Treated with OH, If, 0.3.499.

The tetrapeptide kIN obtained at this point was divided into small fractions. 1.08F fraction in successive cycles - Boc-D-Na 1121-OB, α31! ! f
Hoc-'l'yr (2,6-di-ohbenzyl)-
OH%0.44 tBoC-8at (pennol)-
OB, 0.295tHoc-1'rp-U chisel, 0.
304fBoa-D-Phg -011,0,265r
Combined with pyroglutamine powder, 0.129F to the east.

The resin was removed from the reaction chamber and washed with CB, CI,
Polypeptide resin protected by drying under vacuum 1.66
I got t. The protected peptide was treated with methanol saturated with ammonia oc for 24 hours at room temperature.
Then it was removed from the resin. The resin beads were filtered and washed with methanol and DMk.
The protected peptide was removed by removing the solvent from the solution under vacuum.
9f was obtained as a yellow oil.

Subsequently, in a Kal-F reactor, anhydrous liquid HFt in the presence of anisole (scavenger) was heated at 0°C [1
The protecting group was removed by treatment for 0 minutes. HF was evaporated in vacuum FK to form (PyroJ-Gin-D-Phe-
Trp-8s r-Tyr-D-Na l (2)-L
The residue as the HF salt of aw-Arg-Pro-GlyNH, was washed with ether. The residue was then extracted with ice vinegar. The acetic acid extract was lyophilized to obtain a crude substance α5fff.

This crude material was converted to the acetate salt by passing in an aqueous system for 1 hour in AG3X (weakly basic tertiary aquinone resin) which had been converted to the acetate form. By drying the effluent, crude pegtide acetate 045f was obtained as a white solid.

The crude material was washed with running buffer s s % CHpN/s 5%
B, 0 (0.03Af in NH4OAe, pH 4,5)
Lteroprgp Rp-1,8(2
Silk was produced using high performance liquid chromatography using a αgX30ca column (5-40 microns). The main UV absorption (280 gs) peak eluting in approximately 6 column volumes was collected, concentrated to dryness, and lyophilized three times from distilled water to yield pure (pio)Gls-D-Phm- Trp-
8ar-'I'yr-D-Nal(2)-Las-Ar
g-Pro-GLyNH*1[a]”n=-3&
6 (C0,5// (JAa) 1 melting point = 162-
I got 75 for 165°.

Example 2 The same synthetic program as described in Example IK was used to synthesize a congener with the C-terminus Pro-NHCH, C11. Bahrnan @ 90 board synthesizer reaction valley! II, Bee-;Pro-0B (D
Anhydrous-t'/17mu salt and chloromethyl-polystyrene/
116 divinylheisene (Lab Systems,
1%6. ) Nine Boa-Pr produced by reacting with 1 molar ratio of Boa-Pr.

-〇-Resin 113f was loaded. Use nine Boc-Pr.
-〇-The amount of resin is 100%, containing 7 moles of glycerine L4.

The resin was then combined with a 28 molar excess of each of the amino acids and DCC. Namely, resin.

Bee-Arg(Tosyl)-(J#, L@IFBo
g-Law-OR-Ill), &@B? Boa-3
-(11-7thyl)-D-alanine, α94F and l
-Hydroxybenzotriazole, α49t.

N-Boo-0-2-bromopenoloxyal-ni-L-thion L7Sfs and Eoc-8a
t(Pentzl)-(j#, was reacted with LllF.

At this time point Vc, divide the protected polypegtide resin into tt-halves, add its Ht followed by Bog-Trp-Oil, α57t Bee-D-Pha-C) until completion.11. Reacted with α4801 pyroglutamic acid and α21f.

The resin was removed from the reaction vessel and washed with cn, ct,
polypegtide resin protected by drying in vacuum), 2
Obtained 6t.

The protected polypeptide was then treated with ethylamine 25-
Then, it was cleaved from the resin by 18 hours under 2C. Evaporate the methanol and react with a mixture of anisole WLt and redistilled (from cob) anhydrous liquid HF in a Keg-F reactor for 30 min at 0 °C to prepare (pio)Glu-D-. P
ha-1'rp-8et (pairzyl)-Tyr(2-bromopensiloxycarbonyl)-D-Nag (2)-
Law-Arg(tosyl)-Pro-NIICE.

CH, 1,39tf was obtained. Emit VC# under HFf vacuum,
The residue was washed with ether. Crude (pyro)GLs-
D-Pha-1'rp-8ar-Tyr-Lg pulp-Ar
g-Pro-NHCH, CB, α°82t was obtained as an acetic acid salt. The final result of the gorge is Octade Furunrir Seven Norika (Marek, IAghropr) from 40 to SO.
The sample was injected into preparative high performance liquid chromatography using an α9X55 (1+s column) of ap C1).The effluent was 64-α03MNB40Ae/3.
It was 6% acetonitrile. 8i in 4 injections
The entire layer of Mono-Sada 61~ was slightly hidden. Pure pyroglutamyl-D-phenylalanyl-tributophyl-seryl-tyrocle-3-(2
-naphthyl)-D-alanyl-leucyl-arginyl-
Gloline ethylamide tS was obtained as an acetic acid addition salt.

The above-mentioned interlayers can be modified by substituting ethylamine with the chemically ambiguous n-butylamine, cyclohexylamide, cyclohexylamine, trifluoromethylamine, pentafluoroethylamine, and 2.2.2-trifluoroethylamine. By repeating the procedure, the corresponding n-butylamide, cyclohexylamide, cyclohexylamide, trifluoromethylamide, pentafluoroethylamide, and 2.2.2-trifluoromethylamide of the above nonapeptide can be prepared by friend.

Example 3 A compound of formula 1 where E is -NH-5NH-R'' was prepared by classical solution synthesis.

For example, “AzoGL11NH1 is O-NH
-NH-C- NH, the pegutide was prepared as free vegutide or salt according to the following: (Pyro)01%-D-Pka-Trp-Sgr-Tyr-ON m
(PiC1) GLs-D-Pha-Trp-8ar-1'
yr-D-The bond of each fragment is an acyl azide fI
t(J.

Hongal et al., Co1L, Czech, Chafl
&,Ctnm, ,2 @e! 3!13 (-1971
) r (This could be carried out by the DCC/11BT binding method or other non-lacetization fragment ligation methods. Compound (9) is known: A, S, Dstt
a et al., J., Chum, Son.

Parkinl, 1979 @3?9 I and compounds (
1) can be produced in the same manner as in Example 1.

Compound (3) is removed by hydrogenolysis to remove the C& and nitro groups, followed by conjugation with N-Bog-8-<2-su7tyl)-D-alanine using DCC/HBT or other known coupling agents. It is produced from (2) by Similar LllRE
Regarding the synthesis of homologs, see Dm t a et al., supra.

Similarly, by using other amino acids in place 9 of those used to produce (1) and (2) t- the primary one t- was produced = (in the following, NAa is acetamide 6 ) (D-P A N @ Z)-7V a l (I
t)@*afGllNIlj”)LHRH.

(D-Pha”, D-Trj”, D-
NaJ (11)', 7fGlyNRll・)
LERH recruitment (D-Ph-Ge, D-Trp Lu, D-BI
A.
a", D-1'rp".

D-Mal(fl)・AzuGIINH,”・)LHRH
.

(NAa-L-Pro”, DPG Tomi, D-
Tryp”, D-Nag(2)・, AzaGLyNM−・)LHRH.

(NAa-D-Phl, D-Pha", D-Trp
” , D-BIA@, Azo GtlNH, 凰・)
LIIRIi (NAe-D-Phm”, D-p-C1
-Pha", D-1'rp".

D-1'BA・、Aza Gl 111NB、")L
HRII; (NAa-L-Pro”, D-P
hm", D-1'rp", D-TMP・,
-Azo GIINH-・)LERH纂(D-(via)Gj
s', D-PhN, D-Trp”
, D-Nag (ri", AzoGLyNIi,'
・)LHRHI<DPG', D-Nag (2
)・, Aza GIINH-・)LERH; (NAe-L-Pro', D-Pha", D-T
rp”, D-NaL (2)” * 7 de GIw
NH-.)LHRH.

(D-Pha”, D-Trp”, D-
Mal (Ri・, AzoGIINB-・)LHRH.

Example 4 Following the method of Example 1 and making amino acid substitutions from LIiRII at position II! JCD-amino acid ori.

using L-amino acids (in case of lk, diastereomeric peptides were separated during chromatography).

By substituting appropriate amino acids by in-phase synthesis, the following decapegtides were isolated and obtained. This can be characterized as an acetic acid addition salt: (NA- in the following is acetamide) CD-Pka',
D-Nag (ri', D-Nag (31”) LHRH
l(a)"zz--j! ill (Gα4HOAc
), melting point = 1.827184°5 (D-Pha", D-Trp", D-N
ag (g)'' )LHREtCa )7 = -27
,1! (cas/10Aa).

Melting point = 17! -174°! (D-Pha”, D-Trp”, D-B
IJP ) LHRIIB ('NAa-D-Phe',
D-p-C1-PhN, D-Trp”, D-
Nag (2)・)LHHH: (NAe-L-Pro'
, DPG', D-Trp'', D-Nal (2
)') LHRH irises (NAe-D-Pha', D-PkN, D-Trp
”.

D-BIA・)LIIRIi (NAa-D-Ph-凰, D-p-C1-Pha”,
D-Trp'', D-1'BA・)LHRE.

(NAc-L-Pro”, D-Pha”, D
-Trp”.

D-TMP") LIIRIi; (D-(via)Gls', D-Pha",
D-Trp”.

D-N a l (2) ') LHRH1<DPG
' , D-Na L (2)・)LHRllg(NA
a-L-Pro', D-PhN, D-Tryp"
, D-Na l (2) @) LHRE: l: c
l ) ”n = -3486 (to'0.2110Ac)
Melting point = 176-178°.

Example 5 Repeat the method of Example 2 and introduce an amino acid different from LHHH - amino acid or 1ltD.

The following nonapeptide was obtained by using L-amino f81 (in the latter case, separating the noastereomeric peptide during chromatography) and substituting the appropriate amino acid by solid phase synthesis. This could be separated and characterized as an acetic acid addition salt: (N A a is acetamide below) (Pyro)GLw-D-pci-P
ka-1'rp-8ar-1'yr-D-Nal (1ri-Lax-Arg-Pr.

and its ethylamide, ethylamide, cyclozorobylamide, cyclohexylamide, trifluorethylamide, pentafluorethylamide, and 2. ! ,2
-) Rifluorethylamide toad (Pia) GLs-D-P
kg-Try-8ar-Noyr-L)-Nal (
2) -Law-Arg-Pr.

and its ethylamide, dragon-ethylamide, 7-cloglopyramide, cyclohexylamide, trifluoromethylamide, interfluoroethylamide and 2.2.2
-Trifluorethylamide i(bio)GLs-D-D
PG-Trp-'; mr-Tyr-D-NaJ(2)-
Lash-Arg-Pr.

and its ethylamide, ethylamide, cyclozorobilamide, cyclohexylamide, trifluoromethylide, pentafluoroethylamide and 2.2.2
-Trifluoroethylamide-rich NAe-L-Pro-D
-Pha-D-Trp-8et-1'yr-D-NaL
(2>-Law-Arg-Pr.

and its ethylamide, di-ethylamide, cyclozolobylamide, cyclohexylamide, trifluoromethylamide, pentafluoroethylamide and 2.2.2
-Trifluorethylamide 1NAe-D-Pro-D
PG-1'rp-8ar-Tyr-D-TBAIiLg
u-Arg-Pr.

and its ethylamide, exo-ethylamide, cyclozolobylamide, cyclohexylamide, trifluoromethylamide, interfluoroethylamide and 2.2.2
-) Lifluorethylamide 1 (pyro)GLw-D-p
-C1-Pha-D-Phe-8et-Tyr-DT
MP-Las-Arg-Pr.

and its ethylamide, exo-ethylamide, cyclozolobylamide, cyclohexylamide, trifluoromethyland, pentafluoroethylamide and 2.2.2
-Trifluoroethylamide (via) GLs-D-P
hm-D-NaL (2)-8ar-Tyr-D-HI
A-Lgs-Arg-Pr.

And its ethyland, 舊-ethylamide, cyclozorobylamide, to cyclo! ? Fluamide, trifluoromethylamide, pentafluoroethylamide and 2.2.
2-trifluorethylamide; (pio)GLst-D
-Pkm-Trp-8ar-1'yr-D-1Val
(L)-Leg-Arg-Pr.

and its ethylamide, ethylamide, 7-chloropyrado, 7-chlorohexylamide, trifluoromethylide, pentafluoroethyland, and 2.2.2
-Trifluoroethylamide.

Example 6 A, (fo)GLs-D-Pha-1'rp-8
ay-noyr-D-Nag(2)-Law-Arg-
A solution of the hydrogen fluoride salt a1f of Pro-GlyNB (see Example 1) was added to water Sol! jlc and passed through a column of Domz 3 anion exchange resin previously equilibrated with acetic acid and washed with deionized water. The column was flushed with deionized water and the effluent was lyophilized to (pyro)GL.
u-D-Phet-1°rp-8er-Tyr-D-N
ag(2)-Lau-Arg-Pro-Gtv-NH,
, (cl) ”,”-'a & 6℃(c==o,s
, HOAa), melting point 162-165°C, the corresponding vinegar r1! I got a fence.

By repeating the above method using other acids instead of acetic acid during the equilibration of the resin, for example the corresponding salts with acids, hydrobromic acid, sulfuric acid, phosphoric acid, nitric acid, benzoic acid, etc. I was able to get it.

Similarly, acid addition salts of other peptides homologous to LERII described herein could be prepared.

B. In the case of salts with low water solubility, these can be prepared by precipitation from water using the desired acid.

For example, zinc tannate salt-water al-(pyro)Gls-D-P
ha-Trp-8ar-Tyr-D-Nal (Ri-L
A solution of ts-Arg-Pro-G1g-NH, acetate was treated with a solution of 8 da tannic acid in o, 2&MNa011 (10 m).
Add a solution of ZnSO4·'iE, Oj$ Iv in 1-9.

The resulting suspension was diluted with 1 mj of water, and the precipitate was centrifuged. The supernatant solution was decanted, then the residue was washed twice with 11 portions of water by centrifugation of the precipitate and decanting of the supernatant. The sedimentation tank was dried under vacuum to yield 15 Jvt of mixed zinc tannate of the LHRIi homolog named above.

Pamoate salt - Ethanol L611j and 125MNa
In a mixture of UH0,1-(pio) GE%-D-Ph
a-Trp-8mday1”yr-D-Mal (2)-
Las-Arg-Pro-Gly-NH, acetate @0q
(D solution [K, (L25M NaOHCh8-/4 Moic A'/7F11M9 solution was added. The solvent was removed under reduced pressure, the residue was suspended in water, centrifuged,
The supernatant was decanted to 1. The precipitate was washed with H,0L5-, centrifuged, and the supernatant decanted. The precipitate was dried in vacuo F to yield 54 Da of the 9 moate salt of the LHHH congener named above.

In the same manner, Ikeme low water solubility salt tg could be produced.

(20) GLm-D-Pha-D-1'rp-8 as a free base synthesis from free acid addition salt of C0 addition salt
ar-Tyr-D-NaL (2)-Las-Arg-
1)'J acetic acid 30-1- was added to a solution of 50 lv of Pro-GlyNH. The obtained fz liquid was freeze-dried to obtain the above-mentioned acetic acid jfL501: [a)”l)-27
, 11" (cO, 5,, HOAa), #! point: 72-174°C.

Similarly, other acid addition salts of the above-mentioned peptides, such as the 9-acid, hydrobromic acid, etc., can be prepared by substituting other acids (stoichiometrically correct for the peptide) , addition salts with sulfuric acid, phosphoric acid, and nitric acid were obtained.

D. Production of salts with metal cations, such as subfrog salts 0, 25
M NaOHa 4 d, 3 to water and ethanol 1
- in a mixture of (bio) GLs-D-P-Trp
-8ar-Tyr-D-Nali2)-Lau-Arl
-Pro-Gll-NH, Acetate 5oqtDfljll
A solution of Z% SO1·'IH,OIS da in water α2- was added to L. The precipitate was centrifuged and the supernatant was decanted. The precipitate was washed with water ITIL by centrifugation and decanting of the supernatant. This precipitate was dried under vacuum and the previously named LR
The zinc-based 481Igk of the RR homologue was obtained.

In a similar manner, other polyvalent cations such as calcium, bismuth, barium, magnesium.

It was also possible to produce salts with aluminum, copper, cobalt, nickel, and cadmira.

Example 7 In water g5d (Pio) GLm-D-Pha-Trp-
8ar-Tyr-D-Nal (2)-Lms-Ar
A 50 t column of Dowas 1 ('] basic quaternary ammonium anion exchange resin) in which a solution of g-Pro-GL y-NH, acetate was pre-equilibrated with Na01j solution and the counterion hydroxide was used. 6 passed through the inside.

The column was flushed with 1 sl of 15Q water and the effluent was lyophilized to yield 4.5 μl of the corresponding polypeptide as the free base.

Similarly, other acid addition salts of the compounds of the peptides mentioned above, such as those mentioned in Example 6, could also be converted into the corresponding free bases.

Example 8 The following are LIIRH antagonists of the invention, e.g.
) Gls-D-Pha-Try-8ar-Tyr-D-
Nal (2)-Law-Arg-Pro-GlyN
It is a typical pharmaceutical composition containing Ilg as an active ingredient, either by itself or as a pharmaceutically acceptable salt, such as an acetic acid addition salt, a zinc salt, a zinc tannate salt, etc.

A. Tablet formulation for buccal (e.g. sublingual) administration = L L
HRII antagonist 1aOy compressible sugar, USP ga4 Calcium Sunastearate Table ◎ ~ 2, LHRH antagonist saccharide
1 aosy compressible sugar, usp s
asav magnesium stearate i, s
syλ LHRH antagonist 1 drop Mannitol, USP S 15 Magnesium stearate, USP 1.511p Pregelatinized starch, USP 1 (LOO40LHHE antagonist f
it 10.0mg lactose, USP
T 45° Pregelatinized Starch, USP
l&0IIIp Stearin LYZ” Danesium
L5ηa, LHRH antagonist, was dissolved in enough waxy water to form a moist granulation when mixed with the sugar portion of the excipient. After thorough mixing, one granulated product was dried in a moving bed dryer A. The dried granulate was then sieved to break up any large agglomerates and then mixed with the remaining ingredients. The granulation was then sized to the specified tablet weight by standard tabletting.

b. In this manufacturing method, all formulations contain α01% gelatin, IJSP2. The gelatin was first dissolved in an aqueous gelatinization solvent and then in the LHRH cognate.

The remaining steps were the same as (1).

Formulation 4 can also be used as a tablet for oral administration.

B. Long-term active injectable formulation 1gL Long-term active 10M, injectable sesame oil 1 LHRH antagonist
10.01p Aluminum monostearate, USP 2αolf Aluminum monostearate was pressed together with sesame oil and heated to 125C with stirring until a clear yellow solution was formed.Then the autoclave was sterilized in an autoclave, Allowed to cool. Then CLHRH
The congener was added aseptically while being triturated. Particularly suitable LH
RH congeners are salts of low solubility, such as zinc salts, zinc tannate salts, nodamoate salts, and the like. Nira et al. exhibit exceptionally long-term activity.

LHRH antagonist [ls The microcasselles (0”tsoμ) of the above were suspended in: Dextrose &O%CMC,
Sodium α596 pennoralcohol α9% Tween 80
α1% purified water, sufficient amount 1G (
LO% microcapsules 25 apr were suspended in excipient LQd.

C0 Aqueous solution for intramuscular injection LIIRH antagonist 500° non-antigenic gelatin 511 g Water for injection Dissolve enough gelatin and LHRH antagonist in water for injection to make a total amount of 100 mg.
The solution was then subjected to no mW.

D. Formulation for rectal administration [Herbal excipient LHRH antagonist for rectal administration
WitapaoL H15 with melted paoL H152αoaI LHRH antagonist
and mix well! It was poured into a mold f.

Example 9 Biological activity assay method = lθ 5its, ea% female albino rat) (180F or more) F:, Laboratory conditions for 7 days (light on at 2 a.m.; light: dark; (14:10) K acclimatization.

Daily vaginal washes were collected from each rat from 7:30 am to $e:00 pm for 12 days.

The cytoplasm of vaginal washes was examined microscopically to determine the stage of the estrous cycle. At least 2 prior to the current cycle
Prooestrous (proaa) with a regular 4-day cycle
tro*a) Rats were sorted.

From these selected rats, 10:00 am to 11:00 am
The 20th vaginal wash was taken, and only rats showing typical proestrous washes were assigned to treatment groups.

Rats were treated with a single subcutaneous injection of test compound 100 ~ in propylene glycol/saline vehicle α2-at 12:00 noon during the proestrus 4J session.

The control group received vehicle a2- alone.

The next morning, the treated rats were sacrificed, the oviducts were removed, and the oviducts were removed.
The presence of newly ovulated eggs was examined under a dissecting microscope. The eggs were counted by tearing them open. The nodal cent of female ovulation was determined by the presence or absence of eggs.

Results: Compound A control for rat ovulation
to.

[:N-Aa-L-Pro', D-pj'-Pha
”.

D-Nal (jj)”+6]LHHH0(N-A
e-L-Pro', D-Pka''.

D-1'rp", D-Nag (2)'] LHRH
TO-D-Nal (2)”’ JLHRH70% permission J11 people Shintex (Ni Nis A) Inc.

Claims (1)

[Claims] 1. Formula % Formula % (1) [wherein X (ID-alanyl residue, one water ice on C-3 is α) three or more direct chainbutsu semoku alkyl group lfj
or 6) Acid-converted with three or more straight-chain lower alkyl groups; or C) a saturated carbocyclic group selected from the group consisting of cyclohexyl, hydronaphthyl, perhydrobiphenyl, hydro-2,2-sophenylmethyl, and adamantyl; or C) the following formula % Formula % where A" and A' are independently rL hydrogen, lower alkyl,
Chlorine, and J! , from the group consisting of dt''L, then GFJ, oxygen, nitrogen and sulfur. A heterocyclic aryl-containing group selected from the group consisting of groups represented by: by 1. A is L-piroglu towel, l)-piroglu towel, N-
Acyl-L-prolyl, N-7 sil-D-prolyl, N
-Acyl-D-tryptophanyl, N-7sil-D-phenylalanyl, N+acyl-D-t-halo7enylalanyl, and N-acyl-X (where h is Here, X has the same meaning as described above. BdD-phenylalanyl, D-59-halophenylalanyl, 2,2-nopheylglycyl, and aminoacyl &iM selected from the group consisting of λ, here) O
Well, as mentioned above, 1rJJ Yoshideshi. C is an optional aminoacyl group consisting of L-) liptophanyl, D-) IJ butophanyl, D-phenylalanyl and X, where X is as defined above. E9 Dalicinamide is also fJ-NH-R4. wherein Ry is 4-lower alkyl, cycloalkyl, fluoro-lower alkyl or 1-NH-C-NH-fl'', and R8 is hydrogen or t-lower alkyl; and their pharmaceutically acceptable compounds. 2. A is L-(pio)Glu, D-(pio)Glu. h is a square consisting of NAc-D-Phe and IyAc-L-Pro. b is D-Phg, D -9-haloPhg and L)PG. C is L-'l'rp%D-Tr7p, D-Nag (
1) and D-Nag (2), and λ is selected from the group consisting of D-NaL (2), D-TMP, D-HIA and D-TBA. E is selected from the group consisting of G I INH, and AtaGlyNHR. Special 1. A combination of tuna and tuna, and its pharmaceutically acceptable salts. 3. X is D-! Ife 1rT which is vaL (2)
4it) The compound described in Items 1 and 2 of ``10 Kame'' and its pharmaceutically acceptable salt. 4. The compound according to claim 3, wherein E is G I INH or AzaGlyNH, and its pharmaceutically applicable weir. &(Bio)GLs-DPG-1'r7p-7Set
-1'yr-L)-Nag (2) -Lea-A
rg-Pro-GlyNH,. -D-Nal (2) -Lea-Arg-Pro-
GlyNHl. (Bio)Gin-D-Pha-D-Nag (2)-
8et-Tyr-D-Nag (2)-Law-Arg
-Pro-GLyNH,,NAc-L-Pro-
Li-Ph e-1)-Trp-Be r-Tyr
-D-Nag (2)-Law-Arg-Pro-G
lyNH,,hJAc-D-P-D-p- to 1-P
ha-D-Trp-8er-Tyr-D-Nal (
2) -Lau-Arg-Pro-GlyNH,. NAc -L-Pr o -Dp -Cl -Ph
a -D-Ha l (2)-δat-1'yr-D-
Nag (2)-Lax-Arg-Pro-G111
NB; and NAc-L-Pro-L)-vJ'-Phe-D-Na
l (2) -Sgr-Tyr-D-Nal (2)-
Law-Arg-Pro-GtyN parrot. Compounds and pharmaceutically acceptable salts thereof according to item 4 selected from the group consisting of: 6. The compound according to claim 1, or a pharmaceutically acceptable acid addition salt thereof, to a human subject in need of treatment for a disorder affecting the reproductive system of the subject. A method for the treatment of disorders affecting the reproductive system of a human subject, characterized in that the method comprises administering in a therapeutically effective manner or in the form of a pharmaceutical composition containing a therapeutically effective amount thereof. . 7. Claim m1 for subjects who require contraceptive treatment
characterized in that the compound or its therapeutically acceptable acid addition salt thereof is administered in a therapeutically effective amount or in the form of a pharmaceutical composition containing the therapeutically effective agent. A method of contraception for people who do. & Comprising the therapeutically effective measure of the adduct or its pharmaceutically acceptable acid addition salt as set forth in claim 1, in a mixture with a pharmaceutically acceptable excipient. Pharmaceutical composition. 9, Formula % Formula % (1) [In the formula, X is a D-alanyl residue, and one hydrogen on C-a is. (1) Phenyl, 7-tyl, anthryl, fluorenyl, 7-enanthryl substituted with three or more linear lower alkyl groups. or 6) cyclohexyl, perhydronaphthyl, perhydronaphthyl, perhydro-hydro2 substituted with three or more linear lower alkyl groups; .2-C/phenylmethyl. or C) a saturated carbocyclic group selected from the group consisting of hydrogen, lower alkyl, chlorine, - and AX; or C) the following structural formula: G is substituted with a heterocyclic aryl-containing group selected from the group consisting of oxygen, nitrogen and sulfur; A is L-pyroglutamyl, D-pyroglutamyl, N -Acyl-L-prolyl, N-acyl-D-prolyl, N-
Acyl-D-t. IJ F) 7-anyl, N-acyl-D-phenylalanyl, N-acyl-D-p-halophenylalanyl,
and (#V-acyl-X, where X has the same meaning as above, BFiI)-phenylalanyl, D-yp-halophenylalanyl, 2. 2-y phenylglycyl, and an aminoacyl residue selected from the group consisting of X, where X is as defined above. C#'iL-) is an aminoacyl residue selected from the group consisting of liptophanyl, D-tryptopanyl, D-phenylalanyl, and UX, where X is as defined above, and E is dalicin iP. or -NE-R”,
wherein R1 is lower alkyl, cycloalkyl, fluoro-lower alkyl or -NH-C-NH-R'', and R3 is hydrogen or lower alkyl; and pharmaceutically acceptable salts thereof. In the method for producing , the g4m group and optionally the covalently bonded fc(5) body support are removed from the protected polypeptide to produce the metal compound of the above formula (1) or a salt thereof, or at public expense. At 16, (→
Converting the compound t11!1 of the above formula (1) into a pharmaceutically acceptable salt, (&) converting the salt of the compound of the above formula O) into a pharmaceutically acceptable salt, or (6) The salt of the compound of the above formula (1) is decomposed into the free polypeptide of the above formula 0). A method for producing the compound of formula (1) and a pharmaceutically acceptable salt thereof, which is characterized by the above.