JPS6067498A - Novel peptide, drug and manufacture - 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 provides natural LH-R11 (p-Glu-11is-T
rp-3er-Tyr-Gly-Leu-Arg-P
Look at the amino acid sequence of ro-Gly-Nllzj' d-
The present invention relates to novel peptides and peptide derivatives that act as antagonists of luteinizing hormone-releasing hormone (luteinizing hormone-releasing hormone).

The invention further relates to a process for the preparation of novel peptides and to medicaments containing these substances as drug components.

Various attempts have already been made over the years to produce selectively effective LH-R11 antagonists by modification of the original L11-R0 molecule. This j;
The advantage of the antagonist is that it is useful not only in the endocrine field but also in the treatment of various cancers.
This is explained by the fact that J can be used.

Particularly active LH-RH antagonists are yo[! It is disclosed in Tsuba Patent Publication No. 81877.

A new peptide has now been discovered that has an interesting LH-1il+ antagonistic effect and is at least as active as the known peptides mentioned above. This peptide is often easier and cheaper to produce and is not associated with certain side effects like known peptides.

The new peptide has general formula 1: X-R'-R2-R3-3
er-Tyr-R'-R5-R6~R'-I+'-N
l12 (wherein, X is hydrogen or lower anru(1,-6C)
The groups R1 and R2 are the same or different, and D-11al(2)
, D-Bal (3), D-Nal (1,), D-Nal
(2), D-Phe or phenyl moiety substituted with one or more halogens, argyl (1-4
C), alkoxy (1-4C) or nitro group, R3 is D-Tr+), D-Bat (2), D-B
al(3), D-11al(1), D-Na2(2) or D-Pal (However, at least one of R1 or R3 is D-Bal(2) or D~Ba1(3) disorder) ), R4 is D-Arg, D-Lys, D-homoArg or D-alkyl(1-4C)-homoArg
, R5 is L-Leu.

L-Met, or L-Cys, L-3er or L-homoSer alkyl (i-4C) or +, t 7 s, -
' ) Iy 7 Lucyl (7-LOC) 31- Chil,
R8 is L-Lys or L-Arg, R7 is L-Pro
Or 1. - deaprolyl, and R8 is D-ΔIa or G
ly). The present invention further extends to salts of the peptide θ).

Peptides and peptide derivatives according to formula I and their acid addition salts are produced by methods commonly applied to peptide synthesis. The conventional method for producing the compound involves bonding the desired amino acid by condensation through homogenization or, for example, from the so-called ``li\IIIJ'', and then the resulting peptide is treated with its protective group and/or solid (1''). This is to release it.

Condensation with homogenization is carried out as follows (1, I).

a) condensing, in the presence of a condensing agent, an amino acid or peptide with a free carboxyl group and other protective reactive group with an amino acid or peptide with a free amino group and another protected reactive group; b) an activated carboxyl group; C) condensation of an amino acid or peptide with a free amino group and an optionally protected other reactive group with an amino acid or peptide bearing a free amino group and an optionally protected other reactive group; and optionally protected other reactive groups are condensed with amino acids or peptides having activated amino acids and other optionally protected reactive groups.

Activation of the carboxyl group is particularly carried out by converting the carboxyl group into an acid halokenide, an anodide, anhydride, imidazolate,
or activated esters such as N-hytroquine-succinimite, N-hydroxy-benztriazole or p-
This can be accomplished by converting to a nitrophenyl ester.

The amino group can be activated by converting it to a phosphite amide or by using the phosphorazo method-1.

A suitable method for the condensation reaction is described in “The Peptid
es”.

volume 1.1965 (Academic Pr.
ess), by E, 5chr6derand K,
These are the carbodiimide method, the AND method, the mixed anhydride method, and the activated ester method, as described in Li1bke.

On the other hand, compounds according to formula I can be prepared by solid phase method (Merrif 1e
ld, JAmer, Chem, Soc, 85.214
9 (1963)) 1. : It can be generated. Starting from the side with the amino acid bond (j, the terminal carboxyl group) of the peptide to be produced, the method requires a solid support on which the reactive groups are present or to which the reactive groups can be attached. For example, it can be a copolymer of dichlorostyrene and sichnylhenzene with reactive chloromethyl groups, or a polymeric carrier rendered reactive with hydroxymethyl or hensylamine groups.

For example, when using a carrier carrying chloromethyl groups,
The first α-amino-protected amino acid (in this case D-Ala or GIy) is attached to the carrier via an ester linkage to yield the compound of formula below.

Amino acid solid support (wherein 1? is an α-amino-protecting group) When a support having a hensylamine group is used, binding is carried out through an amide bond to obtain the following compound.

After removal of the R group, the next α-amino-protected amino acid (in this case L-Pro or I7-diaprolyl) can be attached, and after removal of the protecting group of the α-amino group, the next amino acid can be force-conjugated. , and so on, similar combinations are obtained.

After synthesis of the desired amino acid sequence, the complete peptide is removed from the carrier. This removal can be carried out by hydrolysis, alcohol rinsing, amitrilysis or hydrogenation, depending on the type and nature of the carrier and the desired peptide derivative. -Gly-Nll,), it is preferable to employ a carrier and removal method that directly generates such amide groups.

Reactive groups which are not to take part in the condensation reaction are protected by groups which are suitably removed in a non-destructive manner, for example by hydrolysis or reduction. Thus, the carboxyl group' is right-protected, for example by esterification with methanol, ethanol, tert-butanol, henzine alcohol or p-nitrohenyl alcohol.

Groups which can effectively protect amino groups are generally anol groups, such as anol groups derived from aliphatic, aromatic, araliphatic or heterocyclic carboxylic acids, such as acedel, benzoyl or pyrenecarpoquinol, or anol derived from a carboxylic acid (e.g. etquin carbonyl, pennologinocarbonyl, t-pudyloxycarponyl or p-medyloxybenoloxycarponyl group, or an acid derived from a sulfonic acid, e.g. benzenesulfonyl group or toluenesulfonyl group,
Other groups may likewise be used, for example substituted or unsubstituted aryl or aralkyl groups such as hensyl or triphenylmedyl, or groups such as ortho-nitro-phenyl-sulfenyl and 2-benzoyl-1-methylvinyl. Ill get it.

The ε-amino antagonist of lysine and the guanidino group of arginine may be similarly protected. Common protecting groups in this case are the tertiary butyloxycarbonyl or tonol group for lysine, and the nitro or MBS group for alkynine. When using solid phase synthesis, it is also advisable to protect the (additional) hydroxyl groups of serine and dironone.

Accordingly, the general synthesis used to produce such peptides of general formula I involves using one or more crystalline and/or solid supports of the general formula]: X'-1+'-R2-1+3-3er(
R”)-Tyr(1(10)44(1)-R5-R(+
(+n-R'-R8-Y (in the formula, 1 (kan, It',
R'', R5, R? and R8 are synonymous with the definition in 1.
Xl has the same meaning as X, but may also be an N-protecting group, 1
(1) (same meaning as 1?), but further includes an R8-protected D-phosphorus group, NG+-5 protected D-arginyl, NG-protected D-homoarginyl or NG-protected noalkyl-homoalkynyl group. RG(11 has the same meaning as R6, but may also be an R5-protected linol or NG-protected arginyl group, I(° and +7'' are hydrogen or a protecting group, respectively, and Y is NII, parentheses, or after removal Ni12
A solid support that can be converted into a group (provided that if the group is Y or II H2 group, then the group XI, R4 (l), j (a (+), B9 or c) (at least one of z+ is a protecting group or(
'' containing a protecting group), and then the resulting peptide (where X is hydrogen) is optionally unnuated and/or the resulting peptide is converted into an acid addition salt. It consists of converting into .

Acid addition salts are obtained by aliquoting the peptide from the desired acidic medium;
Veptides may be converted to acid addition salts by reaction with acids such as Br, phosphoric acid, sulfuric acid, acetic acid, maleic acid, tartaric acid, citric acid or polyglutamic acid.

N-anlu derivatives, whether N-terminal lower anol (1-8C) derivatives (see definition of . Next, this anol group is used as a protecting group for peptide synthesis. The desired anol derivative is thus produced directly.

On the other hand, it is also possible to introduce a desired anol group by unnulating the peptide using conventional methods.

The N-anol group used preferably has 4 carbon atoms.
11 alkanoyl or alkenoyl groups, such as acedel, propanoyl or acrylyl groups.

The peptide according to the present invention has Ll(-R1
One with antagonistic characteristics, its boldness Takumi - Atsushi
It can be used for both ml and medical purposes. Peptides according to formula I can be used to block the undesirable effects of Lll-R1+. Preference for LH-1i11 (7
A negative effect is particularly seen in hormone-dependent tumors such as benign or malignant D1j statutor tumors and tumors of the breast, ovary, and testis. Acne and amenorrhea can also be considered as undesirable effects of Lll-R11.

The peptides according to the invention can be used for ovulation regulation and therefore in fertility control, for example as intercourse or post-coital contraception 111 and for oestrus synchronization in animals.

The peptides according to the invention are free from the allergic side effects and edema formation that often limit the use of highly active tifl conventional peptides.

The peptides according to the invention may be administered orally, parenterally (dermally or intramuscularly), sublingually, intranasally, rectally or vaginally. Parenteral and intranasal administration are preferred.

, ``Na Island?, - Pepki V 14hZ #: I
/ l-1rl -f R-l, 〆8 suitable for parenteral or intranasal administration) and thus solutions, (optionally by microencapsulation)
It is in the form of suspended fA liquid, emulsion and spray.

the peptide is mixed with suitable auxiliaries or fillers;
They may be used in forms suitable for oral or sublingual administration, such as pills, tablets and dragees. Additionally, the peptides can be administered rectally or vaginally in the form of suppositories.

When administered parenterally, the peptide or peptide derivative according to the invention preferably ranges from 1 μg to 1 μg per 10 kg body weight.
One smoker is administered. For intranasal administration, the preferred dose is 10 per dose! :8j The amount is 5 μg to 10 μg or more, especially 10 μg to 1 μg per kg of body weight. Human administration (
The suitable dosage for parenteral or intranasal administration is
1 to 1001'g for oral, rectal and vaginal administration;
The dosage is generally 10-1000 times higher.

The moieties used alone or in combination with other suitable moieties in the peptides of the invention preferably include (a) R':
There may be a halogen or alkyl group in the ortho and/or meta position of the phenyl group.
(b) R3 has a D-Bal(3) moiety, (c) R'
l is D-8rg, D-homoΔrg or (ma1)-noalkyl(1-4C)-homoArg moiety, (d) R' is L-Arg moiety, (e) R7 is L-Pro (f) R'' is a D-Ala moiety, (g)

The D-Ba 1 moiety present in position 1 or 2 of the molecule is
] or the D-Bat(2) portion, while the 1)-1,la1 portion at position 3 is preferably the D-Bat(2) portion as described above.
-Bal(3) portion.

The abbreviations used in the examples, the present specification, and the claims below refer to the following amino acid groups.

p -G l u -PyroglutamylLys -Lynyl11is -HistidylArg =ArginylGIy -GlynolPro -ProlylMet =MethionylSer = -SerylLeu -Leuynol 1'yr -TyronorPhe = -PhenylalanylAla -Alanyl'l'rp =) Liptophil Ba1(2) -3-(penzodien-2-yl)-alanyl Ba1(3) =3-(benzodien-3-yl)alanyl Pal =3-(pyridyl)-alanyl Pa
1(3) =3-(3-pyridyllualanylttal(
1) -3-(1-naphthyl)-alanyl Na1(2)
-3-(2-naphthyl)-alanyl 11ar(El)
2 = N', N-diethylhomoarginyl 1'hz
Cydiaprolyl Cys (Bzl) -nsteinyl-henonyl ether Cys (C113) - nosteinyl-medyl ether Cys -nsteinyl.

Fruit-1 In the following examples, when the structure of an amino acid is not shown, it means the form I.

Example 1, Ac-D-Bal(3)-D-p,'CI
Phe-1)-'rp-3er1'yr-D-Arg
-Leu-Δrg-Pro-D-Ala-NHp.

Ac-D-Bal (3) of 621” (], mmol)
-D-p, cI -Pl+e-1)-1'rp-011
and Ig (~1 mmol) of H-3er-Tyr-1)
-Arg-l, eu-Arg-Pro-D-Ala-N
11. Add 7 pl+ of the solution to the 10% DMI) solution with
Sufficient N-edylmorpholine was added to make (=J).

Thereafter, the mixture was cooled to 0°C and 1-human (1-human triacyl (1,5 mmol)
and 230''%+ (1, Imno+I, J) were added continuously with quinlecarponoimide for one hour without stirring.

The reaction mixture was stirred at 5° C. for 3 hours and at room temperature for 16 hours.

Thereafter, Synchro/Bexylurea was removed for 15 minutes, and the p solution was evaporated. The thus obtained 411 was stirred with 2511-identified ethyl acetate, and a portion of the crude precipitate was collected. The substance obtained in this way is 1. , Buo)l/
11.0 (1/I) to avoid converting the decapeptide to the acetate form.
y'ex). The ion exchanger was treated with p, the p solution was lyophilized, and the product thus obtained was further purified by column chromatography (SiO2, eluent butanol-pyrinone-acetic acid-water (8:o, 750.25:1): + Purified.

Fractions containing the desired material were evaporated and then lyophilized. 650 n'i of product were thus obtained. Butanol/pyridine/acetic acid/water (410,7510,2
5/1) Rf = 0.53 (Si02), αD (C
=0.5, tumedelformamide)--3.7°.

Cold retro J ζ The following materials were produced in a similar manner.

1, Ac-D-Bal(2)-D-p, cl Pbe-
D-'I'rp-8er-Tyrl) to rg-Leu-
Herg-Pro-D-Ala-Nllt2, Ac-D
-p, cl Pl+e-D-p, cl Pbe-D-B
a[3)-3er-Tyr-D-Arg-Leu-8r
g-Pro-I) more a-Nl123, Ac-D-B
al(2)-D-p, cI Phe-D-Bal(3)
-3cr-Ty+゛ D-^rg-Leu-Δrg-P
ro-D-81a-NIL4, II-D-Bal(3
)-D-1)-CI Pbe-D-Trp-3er-'
l'yr-D-A rg-Leu-A rg-P
ro-D-Ala-N1125, Ac-D-
Nal(2)-D-1), CI Phe-D-Bal(
3)-3er-1'yr-D-Arg-Leu-/lr
g-Pro-D-Ala-Nll6, Δc-D-p, L
ert, butyl Pbe-D-p, cl Phe-
1)-11al(3)-3er-Tyr-D-Arg-
Leu-Arg-Pro-D-Ala-Nll.

? , Ac-D-Bal(3)-D-p, cl Pbe
-D-Trp-3er゛l'yr-1)homo-Ar
g-Leu-Arg-Pro-D-Ala-Nllp8
, Ac-D-Bat(2)-D-p, cl Pbe-
1)-Trp-3cr-'l'yr-D11ar (El
)2-Leu-Arg-Pro-D-Ala-Nllp
9, Ac-D-p, cl Phe-D-p, CI Ph
e-D-Bal(3)-3er'-Tyr-D-Lys
-Leu-Pro-D-Ala-Nl210, Ac-
D-p, cl Inohe-D-p, cI Pbe-D-
Bal(2)-3er-Tyr-D-Arg-Leu-
Pro-D-Ala-NHtll, Ac-D-Nal (
2)-D-p, cl Phe-D-Bal(3)-3e
r-Tyr-l)-11ar(Et), -Leu-Pr
o-D-Ala-Nllp12, Ac-D-p, cI
Pbe-D-1), CI Phe-D-Ba[3)-8
er-Tyr-D-11ar(Et), -Leu-Pr
o-D-Ala-NHz13, Ac-D-Bal (2)
-D-1), CI Pho-D-Pal(3)-3er
-Tyr-D-11ar(IiL)2-Leu-Pro
-D-Ala-Nl12゜Example 3. Δc-D-Na
l(2)-D-p, cl Phe-D-Bal(3)-
3er-Tyr-D-Arg-Met-Arg-Pro
-D-Δ1a-Nl12゜642' fly (1 mmol) of Ac-D-Nal (2)-D-p, cl Pbe-D-
Bal(3)-0H and l g (= l m m o
l) If-3er-Tyr-D-Arg-λlet
, -Arg-Pro-D-81a-Nll, (Boc-
to 3e-T)'r D-Arg-Met Arg-Pr
obtained by removing the N-protecting group from the o-D-Ala-Nl12 fragment via trifluoroacetic acid) - Eggplant l - Sufficient toethylmorpholin was added. After cooling the mixture to 5°C, 20 mmol of hydroxybencitriazole (= 1, 5 mmol,
quinolcarponoimide ("
: 1.1 mmol) was added.

The reaction mixture was stirred at 5° C. for 3 hours and at room temperature for 16 hours.

After that, quinolurea was removed to the formed synchro,
The p solution was concentrated by evaporation. 2) The oil obtained by
After ffi stirring with 511+1' ethyl acetate and -combination, the crude product was precipitated and drained. The material thus obtained was mixed with t of 25 rr&, Bu0II/IItO (1/I
) and convert the decapeptide to the acetate form (
1011f acetate type ion exchanger (Dowex). The solution was lyophilized and the resulting product was subjected to column chromatography (S+Ox, eluent: butanol/pyridine/acetic acid/water (Bu/Py/Δ
c/Wa) 8:0.75:0.25:1).

Fractions containing the desired material were concentrated by evaporation and then lyophilized. In this way, 720 pieces of product were obtained. flu/Py/8c/Wa (to 10.75/D, 25
/1) Rf = 0.47 (SiOt), a D (c =
1.0, trimethylformamimide)--6.5°.

Kanseidoguchi = The following decapeptides were produced in the same manner.

1, Ac-D-p, cI Phe-D-p, cI P
be-D-Bal(3)-Set-Tyr-D-Arg
-MeL-Arg-Pro-D-Ala-Nll.

2, Δc-D-Nal(2)-D-p, CI Phe-
D-Bal(3)-3er-Tyr-D-Arg-Cy
S(Cl13)-Arg-PrO-D Ala N11
t3, Ac-D-p, cI Phe-D-p, cI
Phe-D-Bal(3)-8er-Tyr-D-8r
g=Cys(CH3)-Arg-Pro-D-Ala-
NIL24, Ac-D-Bal(2)-D-p, cI
Phe-D-Bal(3)-Set-Tyr-D-A
rg-Met-Arg-Pro7D-Ala-NHz5
Δc-D-Nal(2)-D-p, cl Phe-D
-Bal(3)-3er-Tyr-l)-11ar(I
iL), -McL-Arg-1nro-D-Ala-N
ll6 Δc-D-Nal(2)-D-p, CI I'
hc-D-Bal(3)-3er'-Tyr-D-Ar
g-Met-8rg-Thz-D-Ala-Nllt7
, 8c-D-Nal(2)-D-p', cI Phe
-D-Bal(3)-3er-1'yrD-8rg-3
er(t,Bu)-Arg-PrO-D-Ala-1j
Ill? 8, Δc-D'-Bal(2)-D-P, CI
PhelD-Pal(3)-3er-'I'y+D-8rg-Me-Arg-Pro-D-Ala-Nl12
9, 8c-D-p, cl I'he-D-p, cl
Phe-p-Bal(3)-3θr-Tyr-D-Ar
g-3er(+,,Bu)-Arg-Pro-D-Al
a-Nll.

10, Ac-D-p, cl Phe-D-p, cl
Phe-D-Bali13)-6er-Tyr-D-A
rg-Cys(Bzl)-Arg-Pro-D-Ala
-Nll,.

Claims (1)

[Claims] (1) -General formula X-1i'-R2-R3-3er-
Tyr-R'-1i5-Ito-R7-1i''-NI1
2+: where X is hydrogen or low-abortion anol (+-8C) JI
i-1111 and 172 (same or different, #3-
(benzochen-2-yl)-alanyl, D-3-(hensodin-3-yl)-alanyl, D-3-(]-nanaphthyl-alanyl, D-3-(2naphthyl)-alanyl, D-P h e or phenyl moiety as 1f1.!=
I or more C [I replaced by l) -1' h
e, alkyl (]-4C), alkoxy (1-4C
) or nitro bracket, l is D -T rp,
D-3-(henzochen 2-yl)-alanyl,
1)-3-(hensodin-3-yl)-alanyl, 1
) ~3-(1~naphdyl)-alanyl, D-3-(2-
naphthyl)-alanyl or D-pyritulualanyl (with the proviso that it is a 1te1('' or -3-yl)-alanyl group), R'tj, 1)-Art! ,1)-1,,ys,
D-homoArg or D-sialgyl(14c)-homo8rg, R5 is L-Leu, L-AleL, 1. -7
Stein-argyl (1-4C) or phenylargyl (7i0C) ether, L-serine-alkyl (1,-
4C) or phenylalkyl (7-10C) ether or 1. -homoserine alkyl (14C) or phenylalkyl (7-10C), 1--thyl, R'" (Jl,
-1, ys or L-Arg, R7 is L-Pro or I
Archab [J crane and Iri8 are D-Ala or Ga
y) Peptides and their acids: [Salted. (2) Bebuti j. according to claim 1, characterized in that +1' is the amino acid residue Ja%D-Ala. (3) The peptide according to claim 1, wherein 173 is the amino acid residue D-3-(bensodine:1-yl)-alanyl. (4) A drug prepared by mixing the peptide according to claim 1gJ with a pharmaceutical carrier. A method for producing a peptide according to claim 1, which comprises producing a peptide. (6) One or more protecting groups and/or solid carriers are represented by the general formula (1):
r(R10)-R”'-R5-1+'(')-1+'
-R'-Y (wherein I+', R', R3, R', R7 and R8 have the same meanings as defined above, Xl has the same meaning as X, and may also be an N-protecting group, R4 (+1 is synonymous with R4, or is further He-protected D-phosphorus group, NG-protected D-arginyl, NG-protected D-homoalkynyl or (J
N-protected dialkyl-homoarginyl group, 17
6 (II is synonymous with R6, or may further be N5-protected Ritzl or NG-protected arginyl, R9 and RIG
are each hydrogen or a protecting group, and Y is N11. group, or a solid support that can be converted to NII,i after removal (with the proviso that
Y is N11. If it is a group, random X1.174+11, R
c (at least one of +1.17e or RIo is or contains a protecting group), and then the obtained peptide (X is hydrogen) 5. The production method according to claim 5, which comprises optionally unbinding and/or converting the obtained peptide into an acid (=1 salt).