KR910001723B1 - Linear and cyclic analogue of alpha-msh - Google Patents

Abstract

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Description

Novel alpha-MSH linear and cyclic fragment analogs

The present invention relates to novel alpha-MSH linear and annular fragment analogs, which are not known in the art, and to methods for stimulating melanocytes in vertebrates by topical application of the analogs and compositions useful for topical application.

In vertebrates, the color of skin, hair and feathers is generally determined by the number and distribution of color-expressing cells (eg melanocytes), which are genetically controlled. Melanocytes in mammals are located in the basal layer of the epidermis, ie, the junction of the skin and the epidermis and hair follicles. It is regulated by the activity of the agent. Depending on the activity of tyrosinase, eumelanin (brown-black) or phaeomelanin (yellow-red) pigments accumulate in organelles and are completely melaninized before melanosomes, more specifically, umelano Primelanosomes, known as moths and pamelanosomes, are known to be involved in the expression of color [Fitzpatrick, TBY Hori, K. Toda, M. Seiji, Jap. J. Derm. 79: 278 (1969)]. Melanosomes are secreted by cells known as cytokine secretion in keratinocytes around the skin or in cells in the center of growing hair.

Although melanin synthesis and hair morphology are genetically expressed, follicular melan0genesis and hair color changes in some mammals have alpha-melanotropin (alpha) with the following linear tridecape peptide structure: Known as melanocyte stimulating hormone, eg hormonally controlled by alpha-MSH).

Ac-Ser-Tyr-Ser-Met-Glu-His-Phe-Arg-Trp-Gly-Lys-Pro-Val-NH ₂

This hormone is known to be derived from high molecular weight precursor protein, proopiomelanocortin, secreted into the pars intermedia of the pituitary gland, which stimulates adenylide cyclase activity and tyrosinase activity to produce melanin. [Hadley. M. E. C. B. Howard, V. J. Hruby, T. K. Sawyer and Y. C. S. Young's pigment cells 6: 323 (1980)].

In humans, alpha-MSH is found only in the fetal pituitary gland and not in adults. In adults, the degree of melanin production is genetically determined and exists as a constituent. When the amount of melanin is high or low, the degree of melanin synthesis is directly dependent on ultraviolet stimulation, or sunlight. Sunlight may cause the skin to darken as the production of melanin increases.This reaction may be a progressive adaptation to protect humans against the aging and variability of ultraviolet light. Exposure to low-intensity ultraviolet light may lead to melanin biosynthesis in the skin. This causes whitening of the skin to be less effective, and a reduced UV protection effect that allows the skin to absorb more radiation. Although adults do not synthesize alpha-MSH in the pituitary gland, human melanocytes are involved in the hormone and its racemization.

In humans, hypopigmentation of the skin is a local disorder of melanin formation in melanocytes, but the cause of such hypopigmentation has not been identified.

About 1% of the world's population suffers from dysfunction due to low pigment formation. Although alpha-MSH and some alpha-MSH analogs are known to blacken skin upon subcutaneous injection in amphibians and alpha-MSH to blacken skin during intramuscular injection in adrenalectomized humans [Lerner, AB, and JS McGuire, NEJ Med. 270: 539 546 (1964)], this type of administration is not suitable for the usage necessary to obtain and maintain the desired effect repeatedly. Until the present invention has been completed, no suitable method for treating this hypopigmentation disorder is known.

Thus, the present invention was completed by discovering that some analogs of alpha-MSH can be effectively administered to the skin, and these administered compounds make melanocytes an active form for stimulating melanogenesis. In addition, according to the present invention, narcosis (pityriasis), ringworm (alba), tinea versiocolor, vitiligo, spontaneously occurring idiopathic guttate hypomelanosis and depigmental nevus (nevus depigmentosus) It may now be possible and convenient to use topical compositions containing alpha-MSH analogs to normalize disorders due to hypopigmentation, such as hypopigmentation, which later causes inflammation. In addition, topical use of alpha-MSH analogs can blacken gray hair due to aging. In addition, the topical use of this analogue will not only make the skin black, making the skin of the animal highly commercially viable, but it will also be possible to blacken the skin even under UV irradiation or lack of sun light.

All proteins related to alpha-MSH isolated from the pituitary gland were studied, and as an active site, Met ⁴ -Glu ⁵ -His ⁶ -Phe ⁷ -Arg ⁸ -Trp ⁹ -Gly ¹⁰ sequence (ie, methionine-glutamic acid-histidine) -Phenylalanine-arginine-tryptophan-glycine). This heptopeptide sequence is the active site of hormones produced from proopiomelanocortin.

Based on the structural relationship of alpha-MSH (and its analogs and fragments) to the biological activity of alpha-MSH using in vitro frog skin biological assays, the active site of alpha-MSH is It was found to represent the sequence of active sites of suitably defined Ac-Met-Glu-His-Phe-Arg-Trp-Gly-NH ₂ . Synthesis of this fragment (alpha-MSH _4-10 ) shows a weak agonist compared to the original hormone when tested for the degree of stimulation of melfk nin cells in frog skin. Prior to the completion of the method of the present invention, no changes in amino acids at the 5 (Glu) and 10 (gly) positions have been studied. We found evidence from the related literature (see US Pat. Nos. 4,457,864 and 4,485,039, and US Pat. App. It was found that analogs of alpha-MSH can be prepared. In addition, the substitution of phenylalanine at position 7 with d-phenylalanine, the opposite of its enantiomer, yields a more active analogue with increased activity compared to the two known (frog and lizard skin) physiological activities. have. Table 1 below summarizes the present invention.

TABLE 1

partial fragments of alpha-MSH and their physiological activity in frog (Rana pipiens) skin and lizard (anoris carorensis) skin

In addition, as a result of studying the activity relationship, two important factors due to structural differences are found. That is, (1) the substitution of methionine, an amino acid having an oxidative side chain, with Nle, a non-oxidizing amino acid, a structural isomer of methionine, increases the physiological activity of the peptide fragment by about 10-fold, and (2) the L-Phe at position 7 is D- Substitution with Phe and the presence of Lys at position 11 increased the physiological activity of alpha-MSH.

인 첫번째 유사체 실험에서, 이것은 개구리 피부의 활성에서는 원래의 alpha-MSH 호르몬보다 약 10 내지 20배의 활성을 나타냈고 도마뱀 피부의 활성에서는 약 2배의 alpha-MSH 활성을 나타내었다. 실험 이전에 환상 alpha-MSH 유사체의 구조와 합성은 alpha-MSH 활성부위 (His⁶-Phe⁷-Arg⁸-Trp⁹)의 중심에 β-턴 구조(Beta-Turn Structure)를 고려한 것이며 alpha-MSH의 생리활성에 대한 구조의 중요성을 고려한 것이다. 이런 환상 alpha-MSH유사체에 대한 구조적인 활성 연구결과, 여러가지의 결론을 얻었다. 즉, (1) 4위치(Met)와 10위치(Gly)사이의 시스테인을 구조 이성질체로 치환시키면 멜라닌세포에서의 분비능이 개구리 피부활성에서는 약 10배, 도마뱀 피부활성은에서는 약 2배 이상 향상되었으며, (2) 이 환상 유사체에서의 Phe을 D-Phe로 치환시키면 개구리 피부에서는 2배, 도마뱀 피부활성은 4배가 증가되었고, (3) 11위치에 Lsy가 존재하는 유사체는 이것이 없는 유사체보다 더욱 높은 활성을 나타내는 유사체이며, (4) 23개의 원자로 구성된 환으로부터 디설파이드 결합에 의하여 환 크기를 축소하거나 확장시키면 생리활성이 심하게 줄어든다는 것이다. 환상 alpha-MSH 유사체에 대한 연구 결과는 다음 표 2에 기재하였다.In addition, a number of structurally defined alpha-MSH compounds as well as linear alpha-MSH analogues were synthesized to measure physiological activity.

In the first analog experiment, it showed about 10-20 times more activity than the original alpha-MSH hormone in frog skin activity and about 2 times alpha-MSH activity in lizard skin activity. Prior to the experiment, the structure and synthesis of cyclic alpha-MSH analogues were considered to beta-turn structure in the center of the alpha-MSH active site (His ⁶ -Phe ⁷ -Arg ⁸ -Trp ⁹ ) and alpha-MSH Considering the importance of the structure of physiological activity. Structural activity studies on these cyclic alpha-MSH analogs yielded various conclusions. In other words, (1) the substitution of cysteine between 4-position (Met) and 10-position (Gly) with structural isomers resulted in a secretion of melanocyte secretion by about 10 times in frog skin activity and about 2 times in lizard skin activity. (2) Substitution of Phe with D-Phe in this annular analog increased twofold in frog skin and four times in lizard skin activity, and (3) analogs with Lsy at position 11 were higher than those without it. It is an analogue that shows activity, and (4) reducing or expanding the ring size by disulfide bond from a ring consisting of 23 atoms significantly reduces the bioactivity. The results of the studies on the cyclic alpha-MSH analogs are shown in Table 2 below.

TABLE 2

In vitro measurement of the activity of cyclic alpha-MSH on frog and lizard skin activity

* Biological activity was measured by the linear alpha-MSH administration-response curve.

* Ring size = the number of bonds that form one of the peptides.

Maa = 2-mercaptoacetic acid

Mpa = 3-mercaptopropionic acid

Hcy = homocysteine

In order to further investigate the activity relationship according to the structure of the alpha-MSH _4-10 fragment, the present inventors focused on the substitution effect of the 5 and 10 positions, so that the melanotropic activity of many linear alpha-MSH _4-10 analogs which have not been known so far. We reviewed the structural requirements for. Based on the results obtained in the linear alpha-MAH analogue, another structurally limited alpha-MSH analogue was studied. In short, 4 to 10 fragments and 4 to 13 fragments of alpha-MSH having the following general structure were prepared.

Wherein the X _xx amino acid is glutamic acid (Glu) or aspartic acid (Asp) (ie, one of monoaminodicarboxylic acids) and the Y _yy amino acid is lysine, oronithine, 2,4-diaminobutyl acid (Dab ) Or basic amino acids such as 2,3-diaminopropanoic acid (Dpr).

The following Ac-alpha-MSH _4-10 linear analogue of Table 3 was synthesized by a solid-phase protein synthesis method using p-methylbenzhydralamine resin as a solid support, and conventionally, for alpha-melanotropin and its analogues, Purification by the method associated with the method used.

TABLE 3

Structure of alpha-melanotropin analog

Linear analogs of alpha-MSH fragments are prepared according to the following examples.

Example 1

Preparation of Ac- [Nle ⁴ , D-Phe ⁷ , Lys ¹⁰ , Gly ¹¹ ] -alpha-MSH _1-13 NH ₂ .

To the title compound using an excess of 3 times the amino acid in the solid phase peptide synthesis method on a p- methylbenzamide Hydral amine resin (pMBHA resin 2.0g, 0.7mmole NH ₂ / resin ^{g) N α -Boc-Val (} "Boc The word "means t-butyloxycarbonyl". The resin is washed with dichloromethane and neutralized after 90 minutes, and then the amino group is acetylated with a mixture of acet anhydride and pyridine. The amino group that did not react on the dendrite is searched by ninhydrin reaction test after 30 minutes. The method of joining each amino acid residue in the growing peptide bond consists of the following steps; (1) washed 4 times with 2 ml of 30 ml of CH ₂ Cl ₂ , and (2) treated with 30 ml of 48% trifluoroacetic acid dissolved in dichloromethane containing 2% of anisole for 5 minutes. This was further treated for 20 minutes to separate the Boc group, (3) 30 ml of two times, neutralized and stirred for 2 minutes each, (5) washed 4 times with 30 ml of dichloromethane for 2 minutes, and (6) 5 ml of 2.4 ml of N-hydroxybenzotriazole (N-Box-N ^im Tos) in which a solution of three-fold Boc amino derivatives (2.1 mmol in this case) dissolved in dichloromethane and HOBt in a concentration of 1 mmole / ml in DMF Except for His, N ^im Tis means n-imidazole tosyl), then add 2.4 ml of DCC so that the concentration of DCC in DMF is Immole / ml, then stir the mixture for 2-3 hours (DMF is used as the binding solvent in the case of Trp, Arg and His). (7) Once complete binding (negative for ninhydrin reaction), 30 ml of deckle It washed three times with two minutes each, and Romero Tan, 8 minutes and washed four times with dichloromethane and then washed for 2 minutes in 100% ethanol 3ml, (9) 30ml.

The protected peptide resin corresponding to the title compound is obtained as the (addition to) forming the combination of the following N ^α -Boc amino acids (or derivatives): N ^α -Boc amino acids, the N ^α -Boc-Pr0g, N ^α -Boc-Gly, N ^α -Boc-Lys- (N ^ε- 2-ClZ), N ^α -Boc-N ⁱ -For-Trp, N ^α -Boc-N ^g -Tos-Arg, N ^α -Boc- D- Phe, N ^α -Boc-N ^im -Tos-His (“N ^g ” means N-guanidino, “N _o ” is N-indolyl, 2-ClZ is 2 chlorobenzoyloxycarbinol Etc.). Boc-His (N ^im -Tos) -D -Phe-Arg (N ^g -Tos) -Trp- (N ⁱ -For) -Lys (N ^ε -2ClZ) -Gly-Pro-Val-p-MBHA The resin is separated into four parts. One portion of the protected peptide resin (1.0 g, 0.5 mmol) was N-Boc-γ-Bzl-Glu, N ^α -Boc-Nle, N ^α -Boc-Ser (O-Bzl), N ^α -Boc-Tyr (O-2 BrZ), N ^α -Boc-Ser (O-Bzl) (“2BrZ” means 2-bromobenzyloxycarbinol) are combined and then converted to the protected title peptide chain.

After the last amino acid was bound, the N ^α -Boc protecting group was removed and the amino group was neutralized, and the protected peptide was obtained by N ^α -acetylating with 10 times excess of N-acetyl-imidazole dissolved in 20 ml of dichloromethane. Peptide resin, ie Ac-Ser- (O-Bzl) -Tyr (O-2-BrZ) -Ser (O-Bzl) -Nle-Glu (γ-Bzl) -His (N ^im -Tos) -D -Phe -Arg (N ^g -Tos) -Trp (N ⁱ -For) -Lys (N ^ε- 2-ClZ) -Gly-Pro-Val-p-MBHA resin was vacuum dried and protected peptide resin (1.0 g) Was separated from the resin using liquid HF, the volatiles were evaporated in vacuo at 0 ° C., then the dried product was washed with ethyl ether (3 × 30 ml) and extracted with 30% aqueous HOAC (3 × 30 ml). ) Lyophilize. The peptide powder (530mg) was separated into two parts (260mg each), one of which was dissolved in 1.5ml of ammonium acetate buffer solution (pH 4.5), filtered through a cartridge filter, and then a CMC column (2.0 × 30.0cm). It was placed on top of and eluted with 250 ml of NH ₄ OAc, 0.01 M (pH 4.5), 0.1 M (pH 6.8) and 0.2 M (pH 6.8), respectively. The main peak appeared at 280nm, and the fractions between 0.1 and the first half of 0.2M NH ₄ OAc were collected and lyophilized to obtain 142.3 mg of white powder. After 80.0 mg of peptide powder was purified by preparative HPLC, main peaks were collected and lyophilized to obtain 63 mg of the title peptide.

Example 2

Preparation of Ac- [Nle ⁴ , Asp ⁵ , D-Phe ⁷ , Lys ¹⁰ , GIy ¹¹ ] -alpha-MSH _1-13 NH ₂ .

The title compound is N ^α -Boc-Asp (β-Bzl), N ^α -Boc-Nle, N ^α -Boc-Ser (O-BZl), N ^α -Boc-Tyr (0-2-BrZ), N 1.0 g (0.5 mmol) of N Boc-His (N ^im -Tos) -D-Phe-Arg- (N ^g -Tos) -Trp () by stepwise binding of ^α- Boc-Ser- (O-Bzl) It is prepared from N ⁱ -For) -Lys- (N ^ε- 2-Clz) -Gly-Pro-Val-p-MBHA resin.

Each combination is performed in the same manner as in Example 1. Acetylation of the protected tridecappapeptide resin is performed with a solution of 10-fold excess of N-acetylimidazole in dichloromethane after neutralization and deprotection of the N-terminal Boc group (5 h). Ac-Ser (O-Bzl) -Tyr (O-2-BrZ) -Ser (O-Bzl) -Nle-Asp (β-Bz1) -His (N ⁱ -Tos) -D-Phe-Arg- (N ^g- Tos) -Trp (N ⁱ -For) -Lys (N ^ε- 2-Clz) -Gly-Pro-Val-p-MBHA resin was vacuum dried to obtain 1.8 g of protected peptide resin. 1.0 g of protected peptide resin is separated by liquid HF, the volatiles are evaporated, washed with diethyl ether (3 x 30 ml), extracted with 30% aqueous HOAc (3 x 30 ml) and lyophilized. . Some of the crude tridecapeptide (200 mg) was dissolved in 1.5 ml of NH ₄ OAc buffer (pH 4.5), filtered through a cartridge filter and placed on top of a CMC column (2.0 × 3.0 cm) as described in Example 1 NH ₄ OAc is discontinuously gradient eluted. Pectide isolated after lyophilization was 152 mg. The obtained crude 100 mg of the peptide was further purified by HPLC to obtain 67 mg of the title compound.

Example 3

Preparation of Ac- [Nle ⁴ , D-Phe ⁷ , Lys ¹⁰ , Gly ¹¹ ] -alpha-MSH _4-13 NH ₂ .

The title peptide was sequentially combined with N ^α -Boc-Blu (γ-Bzl) and N ^α -Boc-Nle using solid phase technique, followed by 1.0 g (0.5 mmol) of Boc-His (N ^im -Tos)- D-Phe-Arg (N ^g -Tos) -Trp (N ⁱ -For) -Lys (N ^ε 2-ClZ) -Gly-Pro-Val-p-MBHA resin. Each binding reaction is carried out as in the above-described example, except that the N-terminal acetylation is carried out (1 hour) by 1: 1 excess of acetic anhydride-pyridine dissolved in dichloromethane. Peptide 4 is a white powder obtained in the yield of 22% by purification in the same manner as in Example 2.

Example 4

Preparation of AC- [Nle ⁴ , ASP ⁵ , D-Phe ⁷ , Lys ¹⁰ , Gly ¹¹ ] -alpha-MSH _4-13 NH ₂ .

The title peptide is 1.0 g (0.5 mmol) Boc-His (N ^im -Tos) -D-Phe-Arg by sequential binding of N ^α -Boc-Asp (β-Bzl) and N ^α -Boc-Nle. It was prepared from (N ^g -Tos) -Trp (N ⁱ -For) -Lys (N ^ε- 2-ClZ) -Gly-Pro-Val-p-MBHA resin. Protective peptide resins are prepared using the method described in Example 1. 130 mg of the purified peptide was purified by CMC chromatography as in Example 1 to obtain 53 mg of pure peptide.

Example 5

Preparation of AC- [Nle ⁴ , D-Phe ⁷ , Lys ¹⁰ ] -alpha-MSH _4-10 NH ₂ .

2.7 g of p-methylbenzhydrylamine resin (0.7mmole NH ₂ / g resin) was suspended in dichloromethane and washed three times with 30 ml of dichloromethane, and the washed resin was dried in dichloromethane before the solvent was filtered. Stir for hours. Neutralize the stretched MBHA resin and bind amino acids as in Example 1. The following amino acid derivatives are bonded to the resin in the bonding order; N ^α -Boc-D-Lys (N ^ε- 2-ClZ), N ^α -Boc-Trp- (N ⁱ -For), N ^α -Boc-Arg (N ^g -Tos), N ^α -Boc-D -Phe, N ^α -Boc-His- (N ^im -Tos).

The obtained Boc-His (N ^im -Tos) -D-Phe-Arg (N ^g -Tos) -Trp (N ⁱ -For) -Lys (N ^α- 2-ClZ) -p-MBHA resin was separated into two parts. do. The separated portion of the resin was combined with N ^α -Boc-Glu (γ-Bzl) and N ^α -Boc-Nle in turn, and the obtained peptide resin was deboxylated and the N-terminus was dichloromethane. Acetylated with 2 times the acetic anhydride-pyridine mixture dissolved in 1: 1 for 1 hour. The resulting protected peptide resin was washed with dichloromethane and dried in vacuo to give 2.1 g. 1.7 g of the protected peptide resin is separated by anhydrous liquid HF-anisole-dithioethane (17 ml HF, 2 ml anisole, 1 ml dithioethane). After evaporating the volatiles, the dried and washed peptides were washed three times with 30 ml of anhydrous diethyl ether and extracted three times with 30 ml of 30% aqueous HOAc, followed by 700 mg of crude peptide from an aqueous precipitate of lyophilized and lyophilized peptides. Got. 300 mg of the crude peptide was dissolved in 2 ml of NH ₄ OAc buffer solution (pH 4.5), followed by filtration using a cartridge fill, and chromatography on a CMC column. Main peak was collected and lyophilized to obtain 172 mg of white powder peptide. 110 mg of crude peptide was purified by HPLC to give 50 mg of pure title compound.

Example 6

Preparation of AC- [Nle ⁴ , D-Phe ⁷ , Lys ¹⁰ ] -alpha-MSH _4-10 NH ₂ .

The title compound, a protected peptide resin, combines N ^α -Bco-Asp (β-Bzl) and N ^α -Boc-Nle in turn to 1.8 g of Boc-His (N ^im -Tos) -D-Phe-Arg- It was prepared from (N ^g -Tos) -Trp (N ⁱ -For) -Lys (N ^ε- 2-ClZ) -p-MBHA. Each binding reaction was carried out in the same manner as described above, using three times excess N ^α -Boc amino acids (or derivatives), 2.4 times DCC, and 2.4 times HOBt. After binding the last amino acid, the N ^α -Boc protecting group is removed and the amino group is neutralized, followed by acetylation for 1 hour in a double excess of an acetic anhydride-pyridine mixture dissolved in 1: 1 in dichloromethane. Ac-Nle-Asp (β-Bzl) -His (N ^im -Tos) -D-Phe-Arg (N ^g -Tos) -Trp- (N ⁱ -For) -Lys (N ^ε- 2-ClZ)- The p-MBHA resin was washed with dichloromethane and dried in vacuo to give 2.1 g. 1.5 g of the protected peptide resin was isolated by liquid HF and subjected to the same process as in Example 5, and then 112 mg of pure peptide obtained by CMC chromatography was purified by HPLC to obtain 64 mg of the title compound.

Example 7

Preparation of AC- [Nle ⁴ , D-Phe ⁷ , Dab ¹⁰ ] -alpha-MSH _4-10 NH ₂ .

Peptide resins protected with the title compound were N ^α -Boc-Dab- (Nγ-Z) and N-Boc -Asp (β-Bzl), respectively N ^α -Boc-Lys (N ^ε -2CLZ) and N ^α- Replace with Boc-Glu (γ-Bzl) Ac-Nle-Asp (β-Bzl) -His- (N ⁱⁿ -Tos) -D-Phe-Arg (N ^g -Tos) -Trp (N ⁱ -For)- It was prepared in the same manner as in Example 5 except that Dab (Nγ-Z) -p-MBHA resin was obtained. By separating by the method described in Example 5 and preparing a peptide resin, a white powdery peptide 12 was obtained in a yield of 36%.

Example 8

Preparation of AC- [Nle ⁴ , Asp ⁵ , D-Phe ⁷ , Dpr ¹⁰ ] -alpha-MSH _4-10 NH ₂ .

Peptide resins protected with the title compounds replaced N ^α -Boc-Dpr (N ^β -Z), N ^α -Boc-Glu (γ-Bzl) instead of N ^α -Boc-Lys (N ^ε 2-Clz). Instead, use N ^α -Boc-Asp (β-Bzl) to use Ac-Nle-Asp (β-Bzl) -His (N ⁱⁿ -Tos) -D-Phe-Arg (N ^g -Tos) -Tpr (Ni Synthesis was carried out as in Example 5, except that -For) -Dpr (N ^β -Z) -MBHA resin was prepared. Peptides were separated from the resin in the same manner as in Example 5, and the desired peptide was obtained as a white powder through several processes (yield 36%).

Example 9

Preparation of AC- [Nle ⁴ , Lys ¹⁰ ] -alpha-MSH _4-10 NH ₂ .

The protected peptide resin to the title compound is N ^α -Boc-D-Phe instead of N ^α using -Boc-Phe Ac-Nle-Glu (γ-Bzl) -His (N in -Tos) -Phe-Arg Synthesis was carried out by the method of Example 5, except that (N ^g -Tos) -Trp (N ⁱ -For) -Lys (N ^epsilon- 2-CIZ) -p-MBHA resin was prepared. Peptides were isolated and subjected to various processes to yield the title compound in 40% yield.

Example 10

Preparation of AC- [Nle ⁴ , Asp ⁵ , Lys ¹⁰ ] -alpha-MSH _41-0 -NH ₂ .

The protected peptide resin to the title compound is N ^α using the N ^α -Boc-Phe-D-Phe, and instead ^{-Boc, N α -Boc-Glu (γ-} Bzl) for N ^α -Boc-Asp (β- Bzl) as prepared by the Ac-Nle-Asp (β- Bzl) -Phe-Arg (N g -Tos) N ε ((N -For) -Lys -Trp -2-ClZ) -p-MBHA resin substituted Synthesis was carried out by the method described in Example 5 except for the one.

Peptides were separated from the resin, then the protecting group was removed and purified by the method described above to give the title compound as a white powder product (yield 37%).

The physiological activity of the linear analogues prepared according to the method of the examples described above is shown in Table 4, where "P" represents increased "activity" ("+" = present, "-" = none) and "ND" Indicates that the sample was not specifically tested.

TABLE 4

Results from structurally restricted alpha-MSH analogs have been incorporated into studies using other types of restricted analogs, structurally cyclic alpha-MSH analogs.

Solid phase peptide synthesis of cyclic melanotropin peptide analogs was done using conventional synthetic techniques. In summary, N ^α -tert-butyloxycarbonyl (Boc) -protected amino acids and their derivatives were dissolved in p-methylbenz hydrylamine resin and 3-fold excess of Boc-protected amino acid derivatives in DMF at a concentration of 1 mmole / ml. 2.4-fold excess N-hydroxybenztriazole (HOBt) solution (except for His), bound using 2.4-fold excess dicyclohexylcarbodiimide (DCC) dissolved in DMF at a concentration of 1 mmol / ml It is. The coupling reaction was carried out in dichloromethane for about 1 to 3 hours, and the reaction result was examined by ninhydrin reaction or chloranyl test and, if necessary, repeated. The reactive side chains of amino acids are as follows: Lys to 2,4-dichlorobenzyloxycarbonyl, Orn, Dab and Dpr to benzyloxycarbonyl, Trp to formyl, Arg and His to tosyl, Glu and Asp Is protected by benzyl ester. Separation of the N ^α -Boc protecting group is performed by treating twice for 5 minutes and for 20 minutes with 48% trifluoroacetic acid containing 2% anisole dissolved in dichloromethane.

The method of binding each amino acid residue to a growing peptide bond consists of several steps: (1) wash with CH ₂ Cl ₂ (4 × 30 ml, 1 min / wash) and (2) protected Boc for 5 minutes with 48% TFA dissolved in CH ₂ Cl ₂ containing 2% of anisole and 20 Remove in each step by treating twice for minutes, then (3) wash with CH ₂ Cl ₂ (2 × 30 ml) and (4) neutralize with 10% diisopropylethylamine dissolved in CH ₂ Cl ₂ . (2 × 30 ml. 3 min / wash), (5) washed with CH ₂ Cl ₂ (3 × 30 ml, 2 min / wash), and then (6) Boc protected amino acid derivative dissolved in 20 ml of CH ₂ Cl ₂ . (In case of Trp, Arg and His, except for the use of DMF instead of CH ₂ Cl ₂ due to solubility problems), add HOBt and DCC in turn and stir for 1 to 3 hours, then (7) CH Wash with ₂ Cl ₂ (3 × 30 ml, 2 min / wash) and (8) wash with 100% EtOH (3 × 30 ml, 2 min / wash).

The after the last amino acid binding to test that the bond is complete remove the N ^α -Boc protecting group and the amino group neutralized, and then dissolved in the N- acetyl imidazole or a 10-fold excess CH ₂ Cl ₂ was dissolved in CH ₂ Cl ₂ Acetic anhydride: Acetylated with a mixture of pyridine 1: 1 (2 times excess for 1 hour). Peptides are unprotected from the protected and separated from the resin by treatment with liquid HF (10 ml per gram of resin) containing 10% anisole and 8% 1,2-dithioethane for 45 minutes at 0 ° C. After evaporating the volatiles in vacuo, the free peptide is washed with diethyl ether or ethyl acetate (3 x 30 ml), extracted with a 30% aqueous solution of acetic acid (3 x 30 ml) and distilled with water (3 x 30 ml). . The aqueous extracts were combined and lyophilized to obtain a white powdered peptide. Each peptide was purified by column chromatography using cationic carboxymethyl cellulose (CMC) resin. Chromatography was discontinuous using 250 ml of 0.01 M NH ₄ OAc (pH 4.5), 250 ml of 0.01 M NH ₄ OAc (pH 6.8), and 250 ml of 0.2 M NH ₄ OAc (pH 6.8), respectively. Gradient eluted. The final fraction of 0.01 M NH ₄ OAc (pH 6.8) and the fraction of the main peak (detected at 280 nm) eluted during the first half of 0.1 M NH ₄ OAc (pH 6.8) buffer were collected and lyophilized to obtain a white powder. Purified peptides were obtained.

Release of gradient 31. 0. Zoperperk eluted during the first half of the OIM.

The structure of the cyclic alpha-melanotropin analog that is part of the present invention is described in Table 5 below.

TABLE 5

Structure of Cyclic Alpha-Melanotropin Analogues

The cyclic protein analogs of the present invention are prepared according to the following examples.

Example 11

의 제조

Manufacture

The protected peptide resin to the title compound, N ^α -Boc- Val-p-MBHA resin, and then to the (N ^α -Boc-Val of 0.7mmole) as a starting material ^α -Boc- N-protected amino acid of 1.0g ( The order of addition) is prepared by combining sequentially; ^{N α -Boc-Pro, N α} -Boc-Gly, N α -Boc-Lys (N ε -2, 4- CLZ), N α -Boc- Trp (N i -For), N α -Boc-Arg ^{- (N g -Tos), N} α -Boc-D-Phe, N α -Boc- His (N in -Tos), N α -Boc-Glu (γ-Bzl), N α -Boc-Nle.

After binding the last amino acid, the N ^α -Boc protecting group was removed and the amino group was neutralized, followed by a 10-fold excess of N-acetylimidazole (6 to 8 hours) dissolved in dichloromethane or acetic anhydride in dichloromethane; Ac-Nle-Glu (γ-Bzl) -His (N ⁱⁿ -Tos) -D-Phe-, a peptide resin protected by acetylation of a 2-fold excess (l to 2 hours) of pyridine dissolved in 1: 1 Arg (N ^g -Tos) -TrP- (N ⁱ -For) -Lys (N ^ε -2, 4-ClZ) -Gly-Pro-Val-p-MBHA resin was obtained. 1.0 g (0.6 mmole) of the protected peptide resin was treated with 10 ml of anhydrous HF containing 1 ml of anisole and 0.8 ml of 1,2-dithioethane for 45 minutes at 0 ° C. The mixture of dried product obtained by treatment with HF, anisole and 1, 2-dithioethane and evaporated in vacuo is washed three times with 30 ml of diethyl ether and the protein is extracted three times with 30 ml of 30% acetic acid.

The aqueous peptide extract was then lyophilized to give 325 mg of crude Ac-Nle-Glu-His-D-Phe-Arg-Trp-Lys-Lys-Gly-Pro-Val-NH ₂ peptide on white powder. 150 mg of crude Ac- [Nle ⁴ , D-Phe ⁷ , Lys ¹⁰ , Gly ¹¹ ] -alpha-MSH _4-13 NH ₂ is dissolved in _2-4 ml of 0.01 M NH ₄ OAc (pH 4.5) and carboxymethylcellulose column Chromatography was performed by discontinuous gradient eluting with 0.01 M (pH 4.5), 0.01 M and 0.2 M NH ₄ OAc (pH 6.8) over (2.0 × 25.0 cm), respectively. The main peak detected at 280 nm was eluted in the first half of 0.1 M NH ₄ OAc (pH 6.8) buffer, and lyophilized to obtain 104 mg of white powder. Vydac 218TP15 using Ac- [Nle ⁴ , D-Phe ⁷ , Lys ¹⁰ , Gly ⁿ ] -alpha-MSH _4-13 NH ₂ purified by CMC column using 0.1% trifluoroacetic acid buffer and acetonitrile as organic emollient Further purification was by HPLC using -16 C ₁₈ RP (25 cm x 25 mm) semi-automatic column. l00 mg of peptide was purified by HPLC to obtain 74 mg of pure Ac- [Nle ⁴ , D-Phe ⁷ , Lys ¹⁰ , Gly ¹¹ ] -alpha-MSH _4-13 NH ₂ peptide. 40 mg of pure Ac- [Nle ⁴ , D-Phe ⁷ , Lys ¹⁰ , Gly ¹¹ ] -alpha-MSH _4-13 NH ₂ samples were dissolved in 1 ml of 5% aqueous HCl solution and diethylalnoethylcellulose (hydrochloroic acid). Elution peak at 280 nm was investigated by chromatography on column (1.0 × 15.0 cm), eluting with 100 ml of 5% aqueous HCl solution.

를 P₄폴리아크릴아미드컬럼(80.0cm)상에서 30% HOAc를 사용하여 탈염시켜 정제한 다음, 반자동식 HPLC로 더욱 정제하여 16mg의 환상 폴리펩타이드

를 얻었다.Peptide peaks were collected and lyophilized to obtain 35 mg of Ac- [Nle ⁴ , D-Phe ⁷ , Lys ¹⁰ , Gly ¹¹ ] -alpha-MSH _4-13 NH ₂ xHCl salt. Peptide salt was dissolved in 3 ml of dry DMF and secondary free amine (distilled ninhydrin under reduced pressure), anhydrous K ₂ HPO ₄ was added to the pephydr solution dissolved in DMF, and the reaction mixture was dissolved in ice salt bath. After cooling to 0 ° C. and 17 μl of diphenylphosphonyl azide are added, the reaction mixture is stirred at 0 ° C. and the entire reaction flask is transferred to a 12 ° C. cold room. After stirring the reaction mixture at 12 ° C. overnight, the reaction was completed by HPLC (Vydac column, 25.0 cm × 4.6 mm) using trifluoroacetic acid / CH ₃ CN as eluent. The ninhydrin reaction is also used to investigate the complete ring formation. After stopping the reaction with 10% HOAc aqueous solution

Was purified by desalting with 30% HOAc on P ₄ polyacrylamide column (80.0 cm) and then further purified by semi-automated HPLC to give 16 mg of cyclic polypeptide.

Got.

Example 12

의 제조.

Manufacturing.

The title compound is N ^α -Boc-Lys (N ^α -2, 4-Cl ₂ Z) -P-MBHA resin (N ^α -Boc-Lys (N ^ε -2,4-Cl ₂ Z) of 1.0mmole) It is prepared by a 2.0g as a starting material, a screen-protected product of the title peptide resin then of N ^α - are prepared after sequential combination with (in order of addition) a protected amino ^{acid; N α -Boc-Trp (N} i For); N ^α -Boc-Arg (N ^g -Tos); N ^α -Boc-D-Phe; N ^α -Boc-His (N ⁱⁿ -Tos).

얻었다.The resulting protected peptide resin, ie N ^α -Boc -His (N ⁱⁿ -Tos) -D-Phe-Arg (N ^g -Tos) -Trp (Ni-For) -Lys (N ^ε -2,4, Cl ₂ Z) -p-MBHA resin is cut in two parts. 1.4 g (0.5 mmole) of the protected pentapeptide resin was converted to the protected peptide peptide after binding of N ^α -Boc-Glu (γ-Bzl) and N ^α -Boc-His-Nle. After binding the last amino acid, the N ^α -Boc protecting group was removed, the amino group was neutralized, and then acetylated by the method described in Example 11 to protect the peptide resin Ac-Nle-Glu (γ-Bzl) -His (N ^in- Tos) -D-Phe-Arg (N ^g -Tos) -Trp (N ⁱ -For) -Lys (N ^ε −2,4, Cl ₂ Z) -P-MBHA resin was obtained. 1.0 g of the protected peptide resin was separated by liquid HF, and the peptide was separated by 356 mg of crude-Ac- [Nle ⁴ , D-Phe ⁷ , Lys ¹⁰ ] -alpha-MSH _4- , a white powder using the method of Example 11. ₁₀ NH ₂ peptide was obtained. 100.0 mg of the peptide was purified by the method described in Example 1 to obtain 65 mg of pure Ac- [Nle ⁴ , D-Phe ⁷ , Lys ¹⁰¹ ] -alpha-MSH _4-10 NH ₂ peptide purified by HPLC. 40 mg of this pure peptide was 13 mg of pure purified by HPLC using the same method as in Example 11.

Got it.

Example 13

의 제조

Manufacture

을 얻었다.The peptide resin protected with the title compound was 1.4 g (0.5 mmol) of Boc-His (N ⁱⁿ -Tos) -D by sequentially combining N ^α -Boc-Asp (β-Bzl) and N ^α -Boc-Nle. -Phe-Arg (N ^g -Tos) -Trp (N ⁱⁿ -For) -Lys (N ^ε -2, 4-Cl ₂ Z) -p-MBHA resin. Each binding reaction is carried out in the same manner as the binding method described in the general solid-phase peptide methodology. After binding the last amino acid, the N ^α -Boc protecting group was removed and the amino group was removed, followed by acetylation by the method described in Example 11 to protect the peptide resin Ac-Nle-Asp- (β-Bzl) -His- ( N ⁱ -Tos) -D-Phe-Arg (N ^g -Tos) -Trp (N ⁱ -For) -Lys (N ^ε -2, 4-Cl ₂ Z) -P-MBHA resins were obtained. 1.0 g of the peptoid resin sample dried in vacuo was isolated and treated as in Example 11 to obtain 370 mg of crude Ac- [Nle ⁴ , Asp ⁵ , D-Phe ⁷ , Lys ¹⁰ ] -alpha-MSH _4-10 NH ₂ . Got it. A portion of the crude heptapeptide (110 mg) was purified in the same manner as used in Example 11 to obtain 82 mg of the title peptide as a white powder. 40.0 mg of pure Ac- [Nle ⁴ , Asp ⁵ , D-Phe ⁷ , Lys ¹⁰ ] -alpha-MSH _4-10 NH ₂ was cyclized after HPLC purification to obtain 12 mg of pure

Got.

Example 14

의 제조

Manufacture

를 얻었다.N ^α -Boc-Orn (Nγ-Z) is bound to 1.0 g of p-MBHA resin (0.7 mmol / g) using the binding method described in the general solid phase method. After one hour the coupling reaction was stopped, the resin was washed and neutralized and acetylated using a 2-fold excess of a 1: 1 mixture of acet anhydride-pyridine dissolved in dichloromethane on the free amino group in the resinous phase. Then, the following amino acids are in turn bound to the resin; N ^α -Boc-Trp (N ⁱ -For), N ^α -Boc-Arg (N ^g -Tos), N ^α -Boc-D-Phe, N ^α -Boc-His (N ⁱⁿ -Tos), N ^α -Boc-Asp (β Bzl), N ^α -Boc-Nle. Each binding reaction was carried out in the same manner as the binding reaction described in the general solid-phase methodology. After binding the last amino acid, the N ^α -Boc protecting group was removed, the N-terminal amino group was neutralized, and then acetylated to protect the peptide resin Ac-Nle-Asp (β-Bzl) -His- (N ⁱⁿ -Tos)-D-Phe-Arg (N ^g -Tos) -Trp (N ⁱ -For) -Orn (Nγ-Z) -p-MBHA resin was obtained. 1.0 g of the peptide resin dried in vacuo was isolated and treated by the method described in Example 11. 332 mg of crude Ac- [Nle ⁴ , Asp ⁵ , D-Phe ⁷ , Orn ¹⁰ ] -alpha-MSH _4-10 NH ₂ Got. 105 mg of the crude heptapeptide was purified by the method used in Example 11 to obtain 78 mg of the linear powder, white powder. 40.0 mg of crude Ac- [Nle ⁴ , Asp ⁵ , D-Phe ⁷ , Orn ¹⁰ ] -alpha-MSH _4-10 NH ₂ was subjected to the cyclization reaction described in

Got.

Example 15

의 제조.

Manufacturing.

1.0 g of p-MBHA resin (0.7 mmol / g) and N ^α -Boc-Dab (Nγ-Z) were combined using the binding method described in the general solid phase method, and after 1 hour, the binding reaction was stopped and the resin was washed. Then, neutralized and acetylated unreacted amino group in a 1: 1 mixture of acet anhydride: pyridine dissolved in dichloromethane for 1 hour, and then the following amino acids are bound to the resin in turn; N ^α -Boc-Trp (N ⁱ -For), N ^α -Boc-Arg (N ^g -Tos), N ^α -Boc-D-Phe, N ^α -Boc-His (N ⁱⁿ -Tos), N ^α -Boc-Asp (β-Bzl), N ^α -Boc-Nle.

를 13.2mg 얻었다.After binding the last amino acid, the N ^α -Boc protecting group was removed, the N-terminal amino group was neutralized and acetylated in the same manner as described in Example 11 to protect the peptide resin Ac-Nle-Asp (β-Bzl)- His (N ⁱⁿ -Tos) -D-Phe-Arg (N ^g -Tos) -Trp- (N ⁱ -For) -Dab (Nγ-Z) -p-MBHA resin was obtained. 1.0 g of the peptide dried in vacuo was isolated and treated to obtain 318.2 mg of crude Ac- [Nle ⁴ , Asp ⁵ , D-Phe ⁷ , Dab ¹⁰ ] -alpha-MSH _4-10 NH ₂ . 100.0 mg of crude heptapeptide was purified to give 78.2 mg of white peptide, linear peptide. 45.0 mg of pure Ac- [Nle ⁴ , Asp ⁵ , D-Phe ⁷ , Dab ¹⁰ ] -alpha-MSH _4-10 NH ₂ was cyclized and purified as described above to obtain pure

13.2 mg was obtained.

Example 16

의 제조.

Manufacturing.

1.0 g of p-MBHA resin (0.7 mmol / g) and N ^α -Boc-Dap (N ^β -Z) were combined using the binding method described in the general solid phase method, and after 1 hour, the binding reaction was stopped. Wash and neutralize, wash and neutralize unreacted amino groups on dendritic, unreact, dendritic, amino groups on acetic anhydride: 1.1 mixture of pyridine dissolved in dichloromethane for 1 hour. The following amino acids are bound to the resin in staged bonds. The following amino acids are bound to the resin in stepwise bonds; N ^α -Boc-Trp (N ⁱ -For), N ^α -Boc-Arg (N ^g -Tos), N ^α -Boc-D-Phe, N ^α -Boc-His (N ⁱⁿ -Tos), N ^α -Boc-Asp (β-Bzl), N ^α -Boc-Nle.

를 11.3mg 얻었다.After binding the last amino acid, the N-terminal amino acid was neutralized and acetylated as in Example 11 to protect the peptide resin Ac-Nle-Asp (β-Bzl) -His (N ⁱⁿ -Tos) -D- Phe-Arg (N ^g -Tos) -Trp (N ⁱ -For) -Dpr (β-Z) -p-MBHA resin was obtained. 1.0 g of the peptide dried in vacuo was isolated and treated as described in Example 1 to obtain 310.8 mg of crude Ac- [Nle ⁴ , Asp ⁵ , D-Phe ⁷ , Dpr ¹⁰ ) -alpha-MSH _4-10 NH ₂ . Got it. 115.0 mg of crude heptapeptide was purified by the method of Example 11 to obtain 82.5 mg of a linear peptide, a white powder. 38.3 mg of pure Ac- [Nle ⁴ , Asp ⁵ , D-Phe ⁷ , Dpr ¹⁰ ) -alpha-MSH _4-10 NH ₂ was cyclized and purified by the method described above to obtain pure

Obtained 11.3 mg.

The biological activity of linear and cyclic alpha-MSH analogs was determined by the ability to stimulate melanosomal secretion using physiological analysis of frogs and lizards in vitro. Analysis of this experiment provides a clear dose-response curve (2.5 × 10 ⁻¹¹ and 4 × 10 ⁻⁴ M) and can detect alpha-MSH minimum concentrations of 10 ⁻¹¹ M. The assay is based on the centrifugal secretion of melanosomes (melanin granules) in dendritic processes that use melanocytes in the skin portion to blacken the skin. All experimental solutions were prepared by serial dilution of the original solution (10 ^-4 M). The frogs (Rans pipiens) used for this measurement were obtained from Kons Scientific, Germantown, WI and the lizards (Anolis carolinsis) from Snake Farm, La Place, LA.

The physiological activity of the cyclic alpha-melanotropin analog is shown in Table 6 below.

TABLE 6

환상 유사체에 있어서 주요한 특성은 도마뱀 피부의 생물학적 분석에서는 낮은 활성을 나타내는 반면 개구리 피부에서는 높은 활성을 나타내고, 또한 최적의 환크기(23개 원자환)와 환크기가 줄어든 것(22개 원자환) 혹은 증가한것(24개 원자환)을 비교시에 활성이 500배 감소한다는 것이다. 5위치와 20위치의 아미노산 사이를 아미드결합 시킴으로써 alpha-MSH를 구조적으로 제한한 것은 본 발명의 일부인 환상 화합물군이다. 환크기가 23개의 원자로된 환인 화합물 19는 개구리 피부 분석에서의 활성에 많은 변화는 없지만 도마뱀 피부에서는 가장 높은 생리활성을 나타낸다. 환상유사체와 그것의 선형전구유사체 사이의 활성 증가치는 10배 정도이다. 환상유사체 18의 활성은 선형유사체와 비교시 큰 영향을 끼치는 것은 아니지만 환상유사체의 증가 활성을 나타내는 중요한 변화를 가진 것이다. C-말단이 Gly-Pro-Val의 트리펩타이드는 화합물 17에서 나타나는 것 같이 그 유사체의 활성에 영향을 끼치지 않는다. 이 환상 락탐펩타이드에서는 선형유사체의 활성과 비교시 환상유사체의 활성이 증가된다는 중요한 특징이 있다.The present invention has revealed the importance of structurally limiting the active site of alpha-MSH or ACTH in the conventional patent documents (US Pat. Nos. 4,457,864 and 4,485,039) which are described collectively by reference. Cycling alpha-MSH at positions 4 and 10 using the pseudo-structural isomeric substitution of Met-4 and GIy-10 with Cys amino acids resulted in a multiple-fold increase in secretion capacity by analogs synthesized in melamine cells. Brought it. The increase in activity of the cyclic analogs compared to the linear precursors of the cyclic analogs was used in the bioactivity assay. The main objective of the present inventors is to obtain analogs with high activity in lizards, as the latter biological assay results in a close relationship with the activity in mammals.

The main characteristics of the cyclic analogs are low activity in the biological analysis of lizard skin, high activity in frog skin, and also the optimal ring size (23 atom rings) and reduced ring size (22 atom rings) or Compared to the increase (24-membered ring), the activity is reduced 500 times. Structural restriction of alpha-MSH by amide bonding between amino acids 5 and 20 is a group of cyclic compounds which are part of the present invention. Compound 19, which has a ring size of 23 atoms, shows the highest physiological activity in lizard skin, although there is no significant change in activity in frog skin analysis. The increase in activity between the cyclic analog and its linear precursor analog is about tenfold. The activity of cyclic analog 18 is not significant compared to the linear analog, but has a significant change indicating the increased activity of the cyclic analog. The tripeptides of the C-terminal Gly-Pro-Val do not affect the activity of the analogs as seen in compound 17. In this cyclic lactam peptide, there is an important characteristic that the activity of the cyclic analog is increased compared with the activity of the linear analog.

Compounds of the present invention are superior to alpha-MSH in terms of activity measured by frog and lizard assays in vivo and in vitro, in vivo persistence affecting such assays, and resistance to degradation by serum enzymes. .

Useful compounds of the present invention can be administered in the middle of the skin and are prepared in suitable compositions determined by the intended method of administration according to methods and processes known in the art. As used herein, the term "intermediate skin" is to be considered in its broadest sense, that is, between the epithelial layers of cells. The word "administration" also refers to topical, oral, nasal and other methods of administration. Compounds suitable for use in the present invention are prepared or synthesized in a dosage form desired by the individual, such as creams, ointments, keels, lotions, tablets, sprays, in a form convenient for use of the respective skin component.

In mass production of these formulations, the compositions can be mixed with known thickening agents, emollients, surfactants, pigments, flavorings, preservatives, fillers and emulsions, all of which are known and which are convenient for the preparation of dermatological preparations. Used. Typically, these inert ingredients make up a large part of the final formulation. The composition is preferably produced slowly or secreted in fine amounts.

Another prominent feature of the compounds of the present invention is that they are advantageous for the substitution of alpha-MSH and [Nle ⁴ ] -alpha-MSH for diagnostic, therapeutic and research purposes. In the area of diagnostic methods, compounds of the present invention, in particular compounds that are bound to or emit gamma rays, or which are radioisotope isotopically determine the location and characteristics of melanocytes on the basis of their relationship with melamine receptors in melanocytes. It turns out to be very suitable for finding things. Serum stability of the compounds of the present invention makes the compounds of the present invention a fundamental competitor in administered selective agent secretion systems, where the skin tissues of interest are known to have high concentrations of melanotropin receptors. Increased activity and relatively high efficacy of the compounds of the present invention in relation to discoloration will double the hormones that stimulate known melanocytes and other biological effects occurring in their synthetic analogs.

Claims (2)

Clear alpha-melanotropin analogues, characterized in that selected from the group consisting of.

Cyclic alpha-melanotropin analogues, characterized in that selected from the group consisting of.