JPH05194260A - Peptide composition - Google Patents

Abstract

PURPOSE: To obtain a medicinal composition containing a calcitonin and having potent hypocalcemic effect through its transmucosal administration. CONSTITUTION: This medicinal composition contains, as active ingredient, isocaproyl [Ala<7> ,Aib<10> ,<17> ,Lys(For)<11> ,<18> ,Lys(desoxyfructosyl)<24> ]-salmon calcitonin-(5-32) at 2μg-30mg per unit dose mode, and also may contain, in combination with the active ingredient, bile acid, taurocholic acid, chenodeoxycholic acid, amino acid salt, etc. Transmucosal administration of the above calcitonin to a rhesus monkey with a body weight of 5-7kg exhibits at least -9% hypocalcemic effect in the blood. This composition is also effective in nasal and rectum administration, being usable for treating Paget's disease, hypercalcemia, osteoporosis, etc.

Description

Detailed Description of the Invention

[0001]

TECHNICAL FIELD The present invention relates to peptides (calcitonins) having a calcium lowering action.

[0002]

Although generally applicable to peptides, the development of suitable and effective formulations for the administration of calcitonin has many problems. Being a peptide, calcitonin is easily degraded by gastric juice upon oral administration.
Moreover, they can cross the mucous membranes of the body with great difficulty and only inefficiently, whether in the stomach, intestines, oral cavity, nose or rectum (transmucosal route).

[0003]

The present invention relates to a pharmaceutical composition containing calcitonin as a pharmacologically active substance and having an increased calcium lowering effect in blood after transmucosal administration. In one embodiment, the invention provides at least 9 when administered to a rhesus monkey weighing 5-7 kg by the transmucosal route.
%, For example -15%, -21% or -25% of blood calcium-lowering effect, isocaproyl- [Al
a ⁷ , Aib ¹⁰ , ¹⁷ , Lys (For) ¹¹ , ¹⁸ , Lys
(Desoxyfructosyl) ²⁴ ] -salmon calcitonin-
A pharmaceutical composition comprising (5-32) is provided. The present invention is
It is based on the results described in later examples. This effect is stronger than the previously published effects for calcitonin compositions administered by the same route and is therefore novel. Oral is preferred as the transmucosal route, but administration by the nasal and rectal routes is also effective. The discovery of a strong hypocalcemic effect makes it possible to administer calcitonin-containing compositions by the transmucosal route. To date, this has not been practical due to the low stability of chemical compounds and / or the resorbability of known compositions.

In particular, the present invention provides that each of the rhesus monkeys weighing 5-7 kg, when orally administered in an amount of 0.1, 0.5, 1 or 10 mg, is at least -9%, for example -15.
%, -21% or -25% of blood calcium lowering effect, isocaproyl- [Ala ⁷ , Aib ¹⁰ , ¹⁷ ,
There is provided a pharmaceutical composition containing Lys (For) ¹¹ , ¹⁸ , Lys (desoxyfructosyl) ²⁴ ] -salmon calcitonin- (5-32).

This material has the formula: N-α-isocaproyl-Ser-Thr-Ala-Val-Leu-Aib-
Lys- (For) -Leu-Ser-Gln-Glu
-Leu-Aib-Lys (For) -Leu-Gln
-Thr-Tyr-Pro-Lys (1-deoxy-D
-Fructosyl) -Thr-Asn-Thr-Gly-
It has Ser-Gly-Thr-Pro-NH ₂ (For = formyl). Formula: isocaproyl- [Ala ⁷ ,
Aib ¹⁰ , ¹⁷ , Lys (For) ¹¹ , ¹⁸ , Lys (desoxyfructosyl) ²⁴ ] -salmon calcitonin- (5-3
2) is a shortened form and is hereinafter referred to as Compound A. Compound A, along with its nature, is described in British Published Patent Application GB2218102A, the contents of which are incorporated herein by reference. Its manufacture is described in Example 11 of the application. Compound A is indicated for use in the treatment of Paget's disease, hypocalcemia and osteoporosis. Compound A as defined above for use in the present invention is in free form or in the form of a pharmaceutically acceptable salt or in the form of a complex or solvate, eg in the form of a pharmaceutically acceptable acid addition salt. Good. Such salts and complexes are known and are as active and tolerated as the free form. Suitable acid addition salt forms for use in accordance with the present invention include, for example, acetate. Compound A differs from the known compound salmon calcitonin in structure and chemical properties. It is shorter by 4 amino acids and lacks the cyclic disulfide structure typical of natural calcitonin. The pharmacological profile of Compound A is not exactly the same as salmon calcitonin, but both compounds produce a hypocalcemic effect, eg when injected. Here, the present inventors have found that Compound A is sufficiently potent to be used in a pharmaceutical composition for oral (stomach) or rectal administration, for example, when nasal administration or a reabsorption enhancer is present. I found that.

In another aspect, the present invention provides isocaproyl- [Ala ⁷ , Aib ¹⁰ , ¹⁷ , Lys (For)].
^{Medicament containing 11} , ¹⁸ , Lys (desoxyfructosyl) ²⁴ ] -salmon calcitonin- (5-32) in combination with bile acid or a salt thereof, such as a basic amino acid, for example, with a metal or ammonium salt A composition is provided.

Bile acid or its salt has a transmucosal reabsorption promoting action. Preferred bile acids are of formula I:

[Chemical 1] Wherein R ₁ is α- or β-OH, R ₂ is H or OH, R ₃ is H or α-OH, and R ₄ is O.
H or NH. R ₅ (In the formula, R ₅ is —CH ₂ —COOH.
Or —CH ₂ —CH ₂ —SO ₃ H). Bile acids are known in the art.

The preferred bile acids are R ₁ = α-OH, R ₂ = H, R ₃ = α-OH and R ₄ = NH.
A compound of formula I, wherein R ₅ is OH, wherein R ₅ is-
When CH ₂ --COOH the acid is glycocholic acid; when R ₅ is --CH ₂ --CH ₂ --SO ₃ H the acid is taurocholic acid. Other preferred bile acids are R ₁
= Α- or β-OH, R ₂ = H, R ₃ = H
And R ₄ = OH, wherein R ₁ = 7α-
When OH, the bile acid is chenodeoxycholic acid; when R ₁ = 7β-OH, the bile acid is ursodeoxycholic acid. Is R ₁ = α-OH, a R ₂ = H, a R ₃ = α-OH, when an R ₄ = OH, the bile acid is cholic acid. Preferred bile acids are chenodeoxycholic acid and taurocholic acid. The metal salt is preferably an alkali metal salt, such as the sodium salt. The basic amino acid is preferably arginine, ornithine,
Lysine, or delta-oxylysine. A preferred acid addition salt is a salt of lysine and taurocholic acid or chenodeoxycholic acid (Tauro-Lys or Ch respectively).
eno-Lys).

One of the most preferred salts is taurocholic acid lysinate. The present inventors have found that, for example, taurocholate lysinate salt is particularly useful as an accelerator for the calcium lowering action of compound A.

Based on the findings set forth in Example 2 below and in the accompanying Table I, the present invention relates to isocaproyl [Ala ⁷ , Aib ¹⁰ , ¹⁷ , Lys (For)] as the active ingredient.
^There is provided a pharmaceutical composition comprising ¹¹ , ¹⁸ , Lys (desoxyfructosyl) ²⁴ ] -salmon calcitonin- (5-32) and bile acid, the composition comprising 3 mg of an active ingredient in a rhesus monkey weighing 5-7 kg. Oral administration of the corresponding dose, 2 after administration
100% relative biological activity when measured over 4 hours
It shows a relative calcium lowering bioactivity (strength) of at least 200% compared to the same composition without bile acid, defined as%. The area under the curve (AUC) showing the calcium lowering effect measured during 24 hours after oral administration was measured by AUC of the composition according to the present invention and the control composition containing no reabsorption enhancer under the same conditions. The strength of the relative calcium decrease is calculated by comparing with.

The improvement in the hypocalcemic effect is due, for example, to the elevated plasma levels of compound A. Plasma level AU
C can also be calculated. Based on the experiments described in Example 2 below and its results described in Table II, the present invention shows that isocaproyl [Ala ⁷ , Ai as the active ingredient.
^b10,17 , Lys (For) ^11,18 , Lys (desoxyfructosyl) ²⁴ ] -salmon calcitonin- (5-32) and a bile acid. The same composition without acid addition salt, defined as a relative bioavailability of 100% when orally administered to a 7 kg rhesus monkey in an amount equivalent to 3 mg of the active ingredient and measured over 24 hours after administration. Shows a relative bioavailability of at least 200% of the active ingredient compared to.

Furthermore, in Example 4, the plasma level of Compound A was measured from the composition further containing an orally administered reabsorption enhancer, and the plasma level of Compound A was the same as that of the same composition administered intravenously without a reabsorption enhancer. Compared. Absolute bioavailability could be calculated in this experiment. Based on the results set forth in Table III below, the present invention provides isocaproyl [Ala ⁷ , Aib ¹⁰ , ¹⁷ , Lys (For) ¹¹ , ¹⁸ , L
The present invention provides a pharmaceutical composition comprising ys (desoxyfructosyl) ²⁴ ] -salmon calcitonin- (5-32) and a reabsorption enhancer, wherein the composition provides 200 μg of the active ingredient to Wistar rats weighing 280-300 g. When compared to the same composition without reabsorption enhancer, the absolute bioavailability was defined as 100%, when administered at a comparable dose orally and measured over 5 hours after administration, at least ng. It shows an absolute bioavailability of 1500% at h / ml.

Based on the results set forth in paragraph 1 of Example 3, Compound A is resorbable nasally and without a resorption promoter, and as described in Example 1, It is concluded that it is resorbable orally with a resorption enhancer. Based on the results of Example 3, the present invention further provides
Rhesus macaques weighing 5-7 kg were treated with isocaproyl [Ala
⁷ , Aib ¹⁰ , ¹⁷ , Lys (For) ¹¹ , ¹⁸ , Lys (desoxyfructosyl) ²⁴ ] -salmon calcitonin- (5-
32) Relative bioavailability is 1 when nasally administered in an amount equivalent to 80 mg and measured for 8 hours after administration.
There is provided a pharmaceutical composition which exhibits a relative bioavailability of at least 200% compared to a control solution containing salmon calcitonin as active ingredient, defined as 00%.

In British patent application GB 2218098A it was disclosed that a pharmaceutical composition containing salmon calcitonin and taurocholate lysinate exhibits improved local tolerability when administered intranasally. They had a bioavailability of the same order as that obtained by nasal administration of compositions containing salmon calcitonin and the known reabsorption enhancer sodium taurocholate. However, it cannot be expected from this prior document that a composition having good calcium lowering activity and good tolerance as obtained by the composition according to the present invention, and having little interaction with food can be obtained. Will. Furthermore, it was not expected that Compound A, which is chemically significantly different from salmon calcitonin, would give such good transmucosal passage and bioavailability results.
This pharmaceutical composition may, if desired, be manufactured in a conventional manner using suitable auxiliary substances for the intended route of administration, eg oral, nasal or rectal route.

The present invention further comprises mixing calcitonin with an auxiliary substance, for example, if a reabsorption improving effect of this composition for oral use is desired, further bile acids or salts thereof, such as metal or basic amino acids. And a salt thereof, which is mixed with a salt thereof to prepare a pharmaceutical composition.
Such compositions may be formulated according to conventional techniques, eg for enteral administration, in conventional dosage forms such as capsules, tablets, suppositories, dispersions, syrups, elixirs, suspensions or solutions. Can be manufactured into the following dosage forms. Suitable pharmaceutical diluents or carriers are described in the examples below. These include, for example, water, lactose and suitable preservatives, suspending agents, wetting agents, to provide sophisticated, palatable pharmaceutical preparations.
It may be a granulating and disintegrating agent, a binder, a lubricant.

If a liquid carrier is present in the nasal composition according to the invention, ie in the liquid composition according to the invention, this is preferably aqueous, but suitable for nasal mucosa physiology. It can also be selected from the non-aqueous solvents that are acceptable. The liquid carrier is preferably water, saline, such as saline, or an aqueous buffer.

If a solid carrier is present, ie in the solid nasal composition according to the invention, it may be, for example, water-insoluble, sparingly water-soluble, water-absorbable, water-swellable, gel-forming or water-soluble. .. Examples of such carriers include, for example, optionally crosslinked synthetic or semi-synthetic polymers such as polyacrylates such as sodium polyacrylate,
Same potassium or ammonium, polylactic acid, polyglycolic acid, copolymer of lactic acid and glycolic acid, polyvinyl alcohol, polyvinyl acetate, copolymer of vinyl alcohol and acetate, carboxyvinyl polymer, polyvinylpyrrolidone and polyethylene glycol; cellulose, Cellulose such as microcrystalline cellulose and α-cellulose, and cellulose derivatives such as methyl cellulose, ethyl cellulose, hydroxyethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, sodium carboxymethyl cellulose and ethyl hydroxyethyl cellulose; α-β- or γ- Dextrins such as cyclodextrin, dimethyl-β-cyclodextrin and dextrin; natural dextrins Starches such as starch and their derivatives, such as hydroxyethyl or hydroxypropyl starch and carboxymethyl starch; polysaccharides, such as dextran, cross-linked dextran, pullulan, alginic acid and salts, hyaluronic acid and salts, pectic acid and salts, phytic acid and Phytin; saccharoses such as D-mannitol, glucose, lactose, fructose, inositol, sucrose and amylose; amino acids such as glycine and taurine; polyamino acids such as polyglutamic acid, polyaspartic acid, polyglycine and polyleucine; Proteins such as casein, gelatin,
Gelatin derivatives such as succinyl gelatin, chitin and chitosan; gums such as gum arabic, gum tragacanth and glucomannan and phospholipids; and mixtures thereof.

These carriers can also be present in the oral composition according to the present invention. Preferred carriers improve contact between the nasal composition and the nasal mucosa or promote diffusion of the drug from the nasal composition to the nasal mucosa, e.g., prolong the residence time of the nasal composition in the nasal cavity, and And / or carriers that reduce the distance between the drug or composition and the mucosa.

A preferred solid support is polyacrylate,
Sodium carboxymethyl cellulose, starch and its derivatives, alginic acid and salts, hyaluronic acid and salts, pectic acid and salts, gelatin and its derivatives, gums, polylactic acid and its copolymers, polyvinyl acetate, celluloses and its derivatives, Coated celluloses,
Crosslinked dextran, more preferably polylactic acid and copolymers thereof, polyvinyl acetate, celluloses and derivatives thereof, coated celluloses and crosslinked dextran. Particularly preferred is microcrystalline cellulose.

For reasons of stability, for example, the liquid compositions according to the invention preferably have a mildly acidic pH, for example 3.7. The required acidity can be conveniently achieved, for example, by the addition of a buffer or an acid such as HCl or other suitable mineral or organic acid, eg phosphoric acid.

The liquid and solid compositions according to the invention further comprise further additives such as antioxidant preservatives such as benzalkonium chloride, methyl p-hydroxybenzoate and p-.
Alkyl p-hydroxybenzoates such as propyl hydroxybenzoate (paraben), or sodium methylmercury thiosalicylate (thiomersal) can also be included.

According to another aspect, the present invention provides a pharmaceutical composition adapted for nasal administration, containing Compound A and a liquid or solid carrier in the absence of a resorption enhancer. The term enhancer is used, for example, by interacting with mucosal components and / or by increasing mucosal permeability.
By compound is meant to be present primarily to increase absorption across the nasal mucosa. Suitable accelerators are eg EP-
Choline esters disclosed in A-214898, such as acylcarnitines, aldoses and glucosamines disclosed in EP-A-215697, such as E
Ascorbates and salicylates disclosed in PA-37943, such as the α-cyclodextrins disclosed in EP-A-94157, such as EP-A-1739.
Pyroglutamates disclosed in US Pat. No. 90, eg US4
No. 476116, chelating agent, polyacrylic acid gel base, sodium glycyrrhetinate, sodium caprate, ammonium tartrate, glycyrrhizinate, for example sodium or ammonium glycyrrhizinate disclosed in EP-A-327756, glycine Or it includes γ-aminolevulinic acid.

In the solid composition for nasal cavity according to the present invention, the particle size of the carrier, for example, the constituent component containing a cellulose carrier is
It can be 5 to 500μ, preferably 10 to 250μ, and more preferably 20 to 200μ.
The liquid composition according to the invention can be prepared by intimately mixing the compound A in a liquid carrier, optionally with further ingredients. The solid composition for nasal cavity according to the present invention can be produced by a conventional method. Compound A can be mixed with carrier particles, eg polymeric bases or cellulosic products, in a conventional manner, optionally together with further components as described above, eg absorption promoters or disclosed surfactants. Compound A may be a solution, eg an aqueous solution or an alcoholic solution, when mixed with carrier particles and the solvent is evaporated, eg by freeze-drying or spray-drying. Such drying can be carried out under conventional conditions. Alternatively, the mixture can be compressed or granulated and then micronized and / or sieved. Compositions for nasal administration may be prepared in conventional powder or liquid form as aerosols, which are then administered, especially from a nebulizer or an evaporator. A metered valve system can deliver a unit dose. The pharmaceutical composition according to the invention is conveniently prepared for oral or nasal administration.

Compound A, isocaproyl- [Ala ⁷ ,
Aib ¹⁰ , ¹⁷ , Lys (For) ¹¹ , ¹⁸ , Lys (desoxyfructosyl) ²⁴ ] -salmon calcitonin- (5-3
The exact dose of 2) can be ascertained in routine animal or clinical studies. For example, this dose can be ascertained by bioavailability comparison studies with other formulations of known therapeutic effect, which dose, when stable, results in a therapeutically effective level of drug Selected to produce plasma levels. Generally, the dose of Compound A with bile acid for oral or rectal absorption will be about half or sufficient of the dose of A in a comparable formulation for the same mode of administration but without the reabsorption enhancer. Is one.

Isocaproyl- [Ala ⁷ , Ai
b ^10,17 , Lys (For) ^11,18 , Lys (desoxyfructosyl) ²⁴ ] -salmon calcitonin- (5-32) The preferred daily dose is about 2 μg for oral or rectal or nasal application. It is about 30 mg, for example 0.1 to 20 mg. Conveniently the composition is in unit dose form or, preferably for oral use, for example about 0.1 to 300, preferably 50 to 200, especially up to about 100 mg of bile acid (exact amount thereof). Can be divided into unit doses, each containing, for example, depending on the mode of administration. Briefly, isocaproyl- [Ala ⁷ , Aib ¹⁰ , ¹⁷ ,
The weight ratio of Lys (For) ¹¹ , ¹⁸ , Lys (desoxyfructosyl) ²⁴ ] -salmon calcitonin- (5-32) is, for example, from about 1: 1 to about 1: 5000 for oral or rectal administration. For example, preferably 1: 2 to 1:
2000. Reabsorption enhancers, such as bile acids, are used as excipients in nasal pharmaceutical compositions, for example, from 0.1 to 1
It can be used in an amount of 0, preferably 0.7 to 1.5% (by weight), especially in oral and rectal compositions in amounts of 1 to 75%, and in oral compositions especially 20 to 75%. , For example 40 to 75% (by weight). A preferred composition is eg Example 2
Compound A 10 mg and bile acid 200 mg as described in 1.
In the form of a lysine salt or in equivalent amounts in the form of a metal salt, for example the sodium salt.

Compound A administered by the composition of the present invention
The dose will of course depend on the type of disease to be treated, the desired frequency of administration and the type of effect desired. The invention is particularly
There is provided a pharmaceutical composition according to the invention for use in the treatment of Paget's disease, hypercalcemia or osteoporosis.

The invention further provides a method of administering a compound to a patient in need of treatment with Compound A which comprises administering to the patient an effective amount of a pharmaceutical composition according to the invention.

[0028]

【Example】

Example 1 Capsules for oral application 1A Compound A 0.1-20 mg Taurocholate lysinate salt 100 mg Microcrystalline cellulose 50 mg Lactose 50 mg For each animal in the group of rhesus monkeys weighing approximately 5-7 kg,
The capsules of Example 1A and placebo capsules containing 0.1, 0.5, 1, 2 and 10 mg of Compound A were administered in 6 separate trials. Animals were fasted for 1 day prior to treatment. 0, 1, 2, 3, 4, 5, 6, 7, 8, 12 and 2
The level of calcium ion in the blood was measured after 4 hours. This calcium is from the International Symposium "Calcitonin 1984", Milan, October 2-4, 1984, Current Clinical Practical Series 42,
Exerta Medica
It was measured by a calcium ion analyzer (ICA 2 radiometer) according to the method described in Selected Short Communications published in a). The results are plotted in the accompanying Figure 1, which shows the good calcium-lowering effect of Compound A, which can be used, for example, in the treatment of osteoporosis. n represents the number of animals in the test.

In another test, changes in total calcium content in blood were measured using monkeys after administration of the oral capsules of Example 1A containing 1 mg of compound A and placebo capsules. The same test was repeated with capsules without taurocholate lysinate salt. Compound A was demonstrated to produce a more potent calcium lowering effect in the presence of taurocholate lysinate salt (Appended Figure 2). This result shows that when orally administered to rhesus monkeys weighing 5 to 7 kg in an amount equivalent to 0.1; 0.5; 1 or 10 mg of the active ingredient, isocaproyl [Al
a ⁷ , Aib ¹⁰ , ¹⁷ , Lys (For) ¹¹ , ¹⁸ , Lys
(Desoxyfructosyl) ²⁴ ] -salmon calcitonin-
Each of the pharmaceutical compositions containing (5-32) is -9%;-
15%; -21% or -25% is shown to have a calcium lowering effect. This calcium lowering effect is an effect on blood calcium ion levels.

Example 2 Capsules for oral application The following compositions were compared by the procedure described in Example 1. 2A mg Compound A 3 Taurocholate lysinate salt 200 Microcrystalline cellulose 50 Magnesium stearate 25 Lactose 25 2B Compound A 3 Chenodeoxycholic acid lysinate salt 200 Microcrystalline cellulose 50 Magnesium stearate 25 Lactose 25 2C Compound A 3 Taurocholate linate salt 200 Starch microspheres 50 Microcrystalline cellulose 50 Magnesium stearate 25 Lactose 25 2D Compound A 3 Chenodeoxycholic acid lysinate salt 200 Starch microspheres 50 Microcrystalline cellulose 50 Magnesium stearate 25 Placebo composition (capsule) mg Compound A 3 Microcrystals Cellulose 100

After the oral administration of the composition to rhesus monkeys, the calcium lowering effect of the simultaneous administration of taurocholic acid lysinate salt and chenodeoxycholic acid lysinate salt appeared to be comparable. The first salt shown produced a stronger effect. As shown in the accompanying Table I and Table II figures, starch microspheres have a small enhancing effect when combined with both salts. The results obtained show that an oral preparation containing taurocholic acid lysinate or chenodeoxycholic acid lysinate had a composition (placebo) of compound A containing no cholic acid addition salt (10%
Defined as 0%), and a relative calcium-lowering bioactivity of 567 and 534%, respectively, and 586 and 45, respectively.
It is shown to give an excellent bioavailability (plasma concentration) of Compound A of 3%. In these tests,
Plasma levels of Compound A were measured using a specific RIA. The detection limit was 0.025 ng / ml.

Example 3 Nasal Composition The calcium lowering effect of the following composition on calcium ions was measured by the method described in Example 1. 3A Spray solution Compound A 440 or 4400 or 8800 μg Sodium chloride 7.5 mg Benzalkonium chloride 0.1 mg HCl 0.1N (until pH 3.7) 9.5 mg Distilled water 1.0 ml Gas N ₂ proper amount (about 40, 400 or 800 μg / activation 3B powder Compound A 40 μg Microcrystalline cellulose 30 mg (about 40 μg / capsule) Control salmon calcitonin (SCT) spray solution Salmon calcitonin (SCT) 440 μg (= 2200 IU) sodium chloride 7.5 mg Benzalkonium 0.1 mg HCl 0.1N Add appropriate amount to pH 3.7 Distilled water up to 1.0 ml Gaseous N ₂ appropriate amount (about 40 μg = 200 IU / activation) Placebo spray solution Sodium chloride 7.5 mg Benzalkonium chloride 0 .1 mg HC Distilled water 1.0ml gaseous N ₂ qs to 0.1N qs addition pH 3.7 (approximately 90 [mu] l / activation)

1. Administration of increasing amounts of spray solution 3A is shown in FIG. 3 [volume-response relationship of ionized calcium in blood (mean + .- SEM) after intranasal administration to rhesus monkeys (n = 4)].
The results are plotted in. They show a good calcium-lowering effect of Compound A of -21% compared to placebo. The maximum calcium lowering effect of SCT (control) after administration of a total of 80 μg of SCT was −
6% or less. 2.80μg (40μg in each nostril) weight 5-7k
g of rhesus macaque over a total of 24 hours,
Calcium ion of compositions 3A and 3B and control spray solution was measured to show 555% bioactivity for composition 3A and 760% for composition 3B (hypocalcemia) compared to 100% control solution (SCT). The effect was calculated. 3. When Compound A was administered to a rhesus monkey weighing 5-7 kg in an amount of 80 μg, the plasma levels of Compound A from Compositions 3A and 3B and the SCT level of the control spray solution were measured over a total of 8 hours, SCT1
Compositions 3A to 316%, Composition 3 compared to 00%
A relative bioavailability of 702% was calculated from B.

Example 4 Male Wistar rats (n = 6) weighing 280-300 g were treated with 500 μl of saline solution containing 200 μg of Compound A with or without a given amount of resorption promoter (control). It was administered by intubation into the oral cavity. 15 minutes before drug administration and 10, 30, 60, 120, 180
And after 300 minutes, a 0.7 ml blood sample was taken from the jugular vein of each conscious animal. All samples were analyzed by using RIA for salmon calcitonin after measuring cross-reactivity to compound A. The drug content of all doses was controlled by HPLC analysis. Absolute bioavailability (BA) after oral administration was calculated according to the following equation:

[Equation 1]

The pharmacokinetic parameters calculated according to the use of specific resorption enhancers are shown in Table III. From Table III, Example 4A, attached, the absolute bioavailability of Compound A with resorption enhancer compared to 100% control composition containing Compound A and no resorption enhancer was 3930. Seems to be up to%.

Therefore, isocaproyl- [Al
a ⁷ , Aib ¹⁰ , ¹⁷ , Lys (For) ¹¹ , ¹⁸ , Lys
(Desoxyfructosyl) ²⁴ ] -salmon calcitonin-
There is provided a pharmaceutical composition comprising (5-32) and no reabsorption enhancer, which composition is administered nasally to a rhesus monkey weighing 5-7 kg in an amount corresponding to 80 μg of the active ingredient for 8 hours after administration. When measured, it was compared with a control solution containing salmon calcitonin as an ingredient and having no reabsorption enhancer, where the relative bioavailability was defined as 100%. h /
It shows a relative bioavailability of up to 702% in ml. Whereas the bioavailability of compound A with taurocholic acid lysinate (Example 4C) is better than compound A with chenodeoxycholic acid lysinate (Example 4B),
The resorption activity of chenodeoxycholic acid in the form of its sodium salt when combined with lysine HCl (Example 4A)
Better. Chenodeoxycholic acid sodium salt 1
% (By weight) and 1% of lysine HCl is theoretically 2.4 kg mol of chenodeoxycholic acid lysinate.
And lysine HCl 3.1 gr mol, which means that the presence of additional lysine has a tremendous improving effect on absorption compared to Example 4B.

Therefore, in the presence of bile acid, regardless of whether an acid addition salt of bile acid and basic amino acid is present, or both acids are present in equivalent amounts necessary to form the acid addition salt. It is possible to improve the bioavailability of Compound A. From Example 4D of Table III, chenodeoxycholic acid sodium salt also appears to have a positive effect. After oral application, the salt can be converted to the free chenodeoxycholic acid in the acid medium of the stomach. In contrast to taurocholic acid, chenodeoxycholic acid is slightly soluble in the gastric acid medium. Compound A is added to the acid medium from a composition containing it slowly, and more slowly than when taurocholic acid or its sodium salt is present, when more chenodeoxycholic acid or its sodium salt is present in the composition. Is released to. Therefore, Compound A may be better protected from destructive attack in the acid medium of the stomach when combined with chenodeoxycholic acid or its alkali salts than taurocholic acid or its alkali salts. Thus, from the viewpoint of protecting Compound A, chenodeoxycholic acid or its alkali salt is a preferred reabsorption enhancer for oral administration.

[0038]

Table I. Calcium-lowering activity in rhesus monkeys after oral administration of 3 mg A in capsules of different formulations (n = 4) Main biokinetic parameters Dosage form Maximum mean of Tmax Tmax AUC Relative lowering activity Average large calcium Tmax 0-24h Average biological activity (hr) Lowering activity (hr) (△% * h) Biological activity (△%) (△%) (%) Tauro-Lys Addition- 17.8 10 -16.4 8 -258.9 497 Add (2A) Cheno-Lys Add -17.1 8.8 -16.7 8 -253.1 486 Add (2B) Tauro-Lys + -19.6 12 -19.6 12 -295.6 567 Add starch microspheres (2C) Cheno- Lys + -18.5 10.8 -16.3 8 -278.3 534 Starch microspheres added (2D) Microcrystalline cell -7.2 3 -6.4 4 -52.1 100 Loose added (control) (by definition)

[0039]

Table 2 Table II Plasma levels in rhesus monkeys (n = 4) after oral administration of 3 mg A in capsules with different formulations Key pharmacokinetic parameters Mean average AUC of Tmax of dosage form Cmax Relative Average Mean Cmax Tmax 0-24h Biology (ng / ml) (hr) (ng / ml) (hr) (ng * h / ml) Practical availability (%) Tauro-Lys Supplement 0.470 0.875 0.325 1 0.999 377 Add (2A) Cheno-Lys Add 0.278 1.375 0.275 1.5 0.798 301 Add (2B) Tauro-Lys + 0.539 1 0.430 1 1.552 586 Add starch microspheres (2C) Cheno-Lys + 0.401 1.625 0.339 1 1.201 453 Add starch microspheres (2D) Microcrystalline cell 0.150 1.5 0.090 1.5 0.265 100 Loose added (control) (by definition)

[0040]

Table III Table III Plasma levels in rats after oral administration of 200 μg of A Dosage form AUC 5h CV Cmax CV Tmax CV Absolute coefficient (dose: saline 500 ng h / ml% ng / ml% h % Biology 200 ug A / rat in μl)% + 10.09 101 8.27 79 0.48 157 2.36 3930% 4A Sodium salt of chenodeoxycholic acid 1% + Ricin HCl 1% + 2.32 67 2.32 60 0.48 157 0.55 920% 4B Kenedooxycholic acid ricinate 1% + 6.20 133 2.92 126 0.36 93 1.18 1970% 4C Taurocholic acid 1% + 7.08 163 4.35 113 0.17 0 1.73 2880% 4D chenodeoxycholic acid sodium salt 0.5% Control 1.40 101 0.53 53 0.96 143 0.06 100% (to definition) According to)

[Brief description of drawings]

FIG. 1 is a graph showing a time-dependent change in calcium ion concentration (Example 1) by a capsule containing Compound A and taurocholic acid lysinate.

FIG. 2 is a graph showing a comparison of the effects of compound A in the presence and absence of taurocholic acid (Example 1).

FIG. 3 is a graph of a capacity-response curve in Example 3.

 ─────────────────────────────────────────────────── ─── Continuation of the front page (72) Inventor Gerd Flickr Germany-Day 7813 Staufen, Albert Hoogart-Strasse No. 10 (72) Inventor Jackie von Dercher France, F-68400 Riedisem, Liu Day Brewback No. 29

Claims (17)

[Claims] 1. Isocaproyl [Ala ⁷ , A, which produces a calcium-lowering effect of at least -9% in blood when administered by transmucosal route to rhesus monkeys weighing 5-7 kg.
ib ¹⁰ , ¹⁷ , Lys (For) ¹¹ , ¹⁸ , Lys (desoxyfructosyl) ²⁴ ] -salmon calcitonin- (5-32)
A pharmaceutical composition comprising: 2. Isocaproyl [Ala ⁷ , Aib ¹⁰ , ¹⁷ , Lys (For)] in the presence of bile acid or a salt thereof.
A pharmaceutical composition comprising ^11,18 , Lys (desoxyfructosyl) ²⁴ ] -salmon calcitonin- (5-32). 3. The pharmaceutical composition according to claim 2, wherein the acid is taurocholic acid. 4. The pharmaceutical composition according to claim 2, wherein the acid is chenodeoxycholic acid. 5. The salt according to claims 2-4, wherein the salt is a salt of an amino acid.
The pharmaceutical composition according to any one of 1. 6. The pharmaceutical composition according to claim 5, wherein the amino acid is lysine. 7. The pharmaceutical composition according to any one of claims 2-4, wherein the salt is a metal salt. 8. A pharmaceutical composition according to any one of the preceding claims, adapted for oral administration. 9. A pharmaceutical composition according to any one of the preceding claims, adapted for rectal administration. 10. A pharmaceutical composition according to any one of the preceding claims, adapted for nasal administration. 11. Isocaproyl [Ala ⁷ , Aib ¹⁰ , ¹⁷ , Lys (Fo) in the absence of a resorption promoter.
r) ¹¹ , ¹⁸ , Lys (desoxyfructosyl) ²⁴ ]-
A pharmaceutical composition adapted for nasal administration, comprising salmon calcitonin- (5-32) and a liquid or solid carrier. 12. The pharmaceutical composition according to claim 11, which is in solid form. 13. The method according to claim 1, comprising microcrystalline cellulose.
The pharmaceutical composition according to 2. 14. The pharmaceutical composition according to claim 11, which is in the form of a liquid containing benzalkonium chloride. 15. 2 μg to 30 m per unit dose form
The pharmaceutical composition according to any one of the preceding claims, comprising g of calcitonin. 16. A patient in need of treatment with the compound, which comprises administering an effective amount of the pharmaceutical composition according to any one of the preceding claims, to isocaproyl [Al.
a ⁷ , Aib ¹⁰ , ¹⁷ , Lys (For) ¹¹ , ¹⁸ , Lys
(Desoxyfructosyl) ²⁴ ] -salmon calcitonin-
A method of administering (5-32). 17. A pharmaceutical composition according to any one of the preceding claims, which comprises mixing calcitonin with an auxiliary substance and, if a resorption promoting action is desired, with bile acid or its salts. how to.