JPH0338255B2 - - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a novel aqueous solution for nasal administration containing polypeptides as active ingredients. More specifically, it is an aqueous liquid preparation containing physiologically active polypeptides such as calcitonin and insulin, and hydroxypropylcellulose, and having a specific viscosity, and when administered intranasally, the polypeptides are efficiently released into the nasal cavity. This invention relates to an aqueous solution for nasal administration that is absorbed through mucous membranes.

Peptide hormones such as insulin and calcitonin have large molecular weights and are easily degraded by proteolytic oxygen such as pepsin, trypsin, or chymotrypsin, so they are difficult to absorb when administered orally and cannot exert effective pharmacological effects. Currently, administration is being carried out.

However, since administration by injection is painful, various other administration methods have been attempted.

For example, rectal administration using suppositories using salicylic acid derivatives such as sodium salicylate, sodium 3-methoxysalicylate, and 5-methoxysalicylic acid as absorption enhancers (J.Pharm.
Pharmacol., 33, 334, (1981)). Another method is intratracheal administration (Diabetes, 20,
552, (1971)) Eye drop administration (Proceedings of the Diabetes Society, 237,
(1974)) are being considered.

However, since all of these methods require a higher dose than injection and have the disadvantage that absorption tends to fluctuate, very few methods have reached practical use yet.

On the other hand, as an attempt at intranasal administration, a liquid preparation for intranasal administration of insulin or calcitonin using a surfactant as an absorption enhancer is known (Diabetes, 27, 296, (1978);
89619; Japanese Unexamined Patent Publication No. 130820/1989).

Furthermore, a liquid preparation for nasal administration of insulin containing cyclodextrin is also known (Japanese Unexamined Patent Application Publication No. 1986-1999).
Publication No. 189118).

The present inventors have conducted extensive research with the aim of obtaining polypeptide formulations for nasal administration that can efficiently absorb polypeptides such as calcitonin through the nasal mucosa and are suitable for intranasal administration. As a result, it was surprisingly found that an aqueous liquid composition containing polypeptides and hydroxypropylcellulose and having a viscosity within an appropriate range sufficiently achieved the above-mentioned purpose. We have discovered that it is possible to do so, and have arrived at the present invention.

That is, the present invention provides a nasal nasal preparation having a viscosity in the range of 2 to 4000 cP at 25° C., which contains physiologically active polypeptides and hydroxypropyl cellulose in an aqueous base suitable for application to the nasal mucosa. It is an aqueous solution for administration.

As the physiologically active polypeptides used in the present invention, polypeptides having a molecular weight in the range of 300 to 300,000 are preferred because they are easily absorbed through the nasal mucosa.

Preferred specific examples of physiologically active polypeptides include the following. For example, calcitonin, insulin, angiotensin,
Vasopressin, desmopressin, feripressin, protirelin, luteinizing hormone-releasing hormone, corticotropin, prolactin, somatropin, thyrotropin, luteinizing hormone, kallikrein, parathyrin, glucagon, oxytocin, gastrin, sexylene, serum gonadotropin, growth hormone, erythropoietin, Peptide hormones such as angiotensin, urogastrone, renin and chemically modified compounds thereof;
interferon, interleukin, transferrin, histaglobulin, macrocortin,
Physiologically active proteins such as blood coagulation factor and chemically modified compounds thereof; vaccines such as pertussis vaccine, diphtheria vaccine, tetanus vaccine, influenza vaccine, and lymphocytosis factor and fibrillar hemagglutination factor. Among these, peptide hormones are particularly preferred, and among the peptide hormones, calcitonin, insulin, luteinizing hormone-releasing hormone, desmopressin, vasobrecin, and oxytocin are particularly preferred. More preferred are calcitonin and insulin.

The aqueous solution for nasal administration of the present invention contains hydroxypropyl cellulose as well as the above-mentioned physiologically active polypeptides, and has an appropriate viscosity, so that when administered into the nasal cavity, it first spreads onto the nasal mucosa. Although they are moderately dispersed, compared to when a simple aqueous solution for intranasal administration is administered into the nasal cavity, the flow is slower and they remain at a moderate level, prolonging the time it takes for them to disappear from the nasal cavity. This is thought to be the reason for the high absorption of polypeptides. In the present invention, in order to impart viscosity to the aqueous solution for nasal administration and to promote absorption, conventional absorption enhancers such as surfactants may be used, particularly by selecting hydroxypropyl cellulose. , polypeptides are absorbed to such an extent that they can exhibit sufficient pharmacological effects even when not used. Hydroxypropyl cellulose also has low irritation to the nasal mucosa and has no unpleasant odor, making it extremely suitable for intranasal administration.

Hydroxypropyl cellulose (HPC) used in the present invention includes, for example, HPC with a viscosity of 3.0 to 5.9 cP in a 2% aqueous solution at 20°C, and HPC with a viscosity of 6.0 to 10.0 cP.
HPC, 150~400cP HPC, 1.000~4.000cP
Examples include HPC. These HPCs may be used alone or in combination.

Aqueous bases suitable for application to the nasal mucosa for use in the present invention include water, saline or buffer solutions. Typical buffers used as buffer solutions include, for example, phosphate buffers, acetate buffers, tartrate buffers, succinate buffers, citrate buffers, lactate buffers, etc. .

HPC and polypeptides as described above are added to these aqueous bases to obtain a predetermined viscosity, that is, 25
It is an aqueous solution having a viscosity of 2 to 4000 cP at °C.
The viscosity of the aqueous liquid is preferably 2 to 2000 cP,
Furthermore, 5 to 1000 cP is preferable. By setting the viscosity within such a specific range, an aqueous solution for nasal administration with excellent absorption of polypeptides can be obtained.

The aqueous solution for nasal administration of the present invention can be produced by a method known per se. For example, polypeptides can be dissolved in water prepared to have a desired viscosity by dissolving hydroxypropylcellulose;
It can be produced by dissolving, suspending, or emulsifying it in physiological saline or a buffer solution.

The final pH of the aqueous solution for nasal administration of the present invention is approximately
3.5 to 7.5, more preferably about 5.0 to 7.5. Within this range, the irritation will be low when the solution is administered intranasally. The desired PH is achieved, for example, by the presence of acids, bases or buffers as described above. Acids used for pH adjustment include:
For example, inorganic acids (e.g., hydrochloric acid, boric acid, phosphoric acid,
carbonic acid, bicarbonate, etc.), amino acids or organic acids (such as monocarboxylic acids, oxycarboxylic acids,
Examples of bases include polycarboxylic acids).
Examples include sodium hydroxide, potassium hydroxide, sodium hydrogen carbonate, and sodium carbonate.

In order to improve the physical properties, appearance, odor, etc. of the aqueous solution for nasal administration of the present invention, known coloring agents, preservatives, antiseptics, stabilizers, flavoring agents, etc. may be added as necessary. A tensioning agent or the like may be added. Coloring agents include copper chlorophyll, β-carotene, Red No. 2, Blue No. 1, etc. Preservatives include ascorbic acid, erythorbic acid and their salts; preservatives include roseoxybenzoic acid ester, phenol, chloride, etc. benzethonium,
Benzalkonium chloride, etc.; stabilizers include, for example, human serum albumin, mannitol, sorbitol, etc.; flavoring agents, include menthol, citrus flavor, etc.; and tonicity agents include glucose, etc.

Furthermore, in the present invention, polypeptides can be absorbed at sufficient concentrations without the use of commonly used absorption enhancers such as surfactants.

A method for administering the aqueous solution for nasal administration of the present invention includes, for example, placing the solution in a nasal dropper, spray container, or similar container suitable for intranasal application of such a solution, and dropping it into the nasal cavity. Alternatively, there is a method of administering the drug by spraying, or a method of administering the solution into the nasal cavity by using an insert that allows easy and quantitative administration of the solution into the nasal cavity.

Although the amount of polypeptide administered varies depending on the type or severity of the disease, the appropriate amount of the solution is approximately 0.05 to 0.5 ml per administration.

As detailed above, the aqueous solution for nasal administration of the present invention, which has a specific viscosity and is composed of hydroxypropyl cellulose and polypeptides, has low toxicity, low irritation to the nasal mucosa, and almost no unpleasant odor. Highly suitable for intranasal administration without
In cases where continuous multiple administration is required, patients can easily perform home therapy by themselves, and physiologically active polypeptides can be efficiently absorbed through the nasal mucosa at sufficient concentrations to obtain therapeutic effects. It has various advantages.

The present invention will be explained below with reference to Examples, but the present invention is not limited to these in any way.

Example 1 An aqueous solution for nasal administration of the present invention was obtained as follows.

(a) Hydroxypropylcellulose (20℃, 2%
Viscosity in aqueous solution is 1000~4000cP) 1g
25 by dissolving in 100ml saline
A solution with a viscosity of about 100 (cP) at °C was obtained.
Next, 0.5 mg of salmon calcitonin (4000 MRC units/mg) was added and dissolved in 5 ml of this solution to obtain a homogeneous physiological saline solution. The physiological saline solution obtained in this way is 0.1 mg
Contains (400 MRC units)/ml salmon calcitonin.

(b) Hydroxypropylcellulose (20℃, 2%
Viscosity in aqueous solution is 1000-4000cP) 2g
A solution with a viscosity of about 1000 (cP) at 25°C was obtained by dissolving in 100 ml of distilled water. Next, 0.6 mg of salmon calcitonin (4000 MRC units/ml) was added to 3 ml of this solution and dissolved to obtain a homogeneous solution. In this case, bubbles may be generated during dissolution, but these bubbles are
It is easily eliminated by ultrasonication or centrifugation. The aqueous solution thus obtained contains 0.2 mg (800 MRC units)/ml salmon calcitonin.

(c) Hydroxypropylcellulose (20℃, 2%
Viscosity in aqueous solution is 1000~4000cP) 1g
For dissolving in 100 ml of isotonic phosphate buffer
A solution with a viscosity of about 100 (cP) at 25°C was obtained.
Next, 0.4 mg of salmon calcitonin (4000 MRC units/mg) was added to 5 ml of this solution and dissolved to obtain a homogeneous solution. The solution thus obtained is 0.08 mg (320 MRC units)/mg
Contains salmon cartonin.

(d) Hydroxypropylcellulose (20℃, 2%
Viscosity in aqueous solution is 1000~4000cP) 1g
A solution with a viscosity of about 100 (cP) at 25°C was obtained by dissolving in 100 ml of isotonic phosphate buffer with pH 7.4. Then add insulin to 10ml of this solution.
A homogeneous solution was obtained by adding and dissolving 157 mg (25.5 units/mg). The solution thus obtained contained 15.7 mg (approximately 400 units)/ml of insulin.

(e) Hydroxypropylcellulose (20℃, 2%
Viscosity in aqueous solution is 1000~4000cP) 1g
A solution with a viscosity of about 1000 (cP) at 25°C was obtained by dissolving in 100 ml of isotonic phosphate buffer with pH 7.4. Then add insulin to 10ml of this solution.
A homogeneous solution was obtained by adding and dissolving 157 mg (25.5 units/mg). The solution thus obtained contains 15.7 mg (approximately 400 units)/ml insulin.

(f) Hydroxypropylcellulose (20℃, 2%
25℃ by dissolving 1g (viscosity in aqueous solution is 1000-4000cP) in 100ml of physiological saline.
A solution with a viscosity of about 100 (cP) was obtained. Then add 157 mg (25.5 mg) of insulin to 10 ml of this solution.
unit/mg) and mixed well to obtain a suspension. The suspension thus obtained contains 15.7 mg (approximately 400 units)/ml insulin.

(g) Place the calcitonin-containing solutions shown in (a), (b), and (c) and the insulin-containing solutions shown in (d), (e), and (f) into appropriate nasal containers. Thus, a calcitonin and/or insulin preparation for nasal administration to humans, which can be administered to humans in an amount of about 0.05 to 0.0 ml at a time, was obtained.

Example 2 () An aqueous calcitonin solution for nasal administration (for animal experiments) of the present invention was obtained as follows.

(a) Hydroxypropylcellulose (20℃, 2
% viscosity in aqueous solution is 1000-4000cP)2
By dissolving g in 100ml of distilled water
An aqueous solution with a viscosity of about 1000 (cP) at 25°C was obtained. Next, 97.5 μg of salmon calcitonin (4000 MRC units/ml) was added and dissolved in 3 ml of this aqueous solution to obtain a homogeneous solution. If bubbles were generated during dissolution, the bubbles were removed by ultrasonication or centrifugation to obtain a homogeneous solution. The aqueous solution thus obtained contains 32.5 μg (130 MRC units)/ml of salmon calcitonin.

(b) Hydroxypropyl cellulose (20℃, 2
% viscosity in aqueous solution is 1000-4000cP)1
By dissolving g in 100ml of distilled water
An aqueous solution with a viscosity of about 1000 cP at 25°C was obtained. Next, 97.75 μg of salmon calcitonin (4000 MRC units/mg) was added and dissolved in 3 ml of this aqueous solution to obtain a homogeneous solution. The aqueous solution thus obtained was 32.58 μg.
Contains (130.3 MRC units)/ml of salmon calcitonin.

(c) Hydroxypropyl cellulose (20℃, 2
% viscosity in aqueous solution 1000~4000cP) 0.4
By dissolving g in 100ml of distilled water
An aqueous solution with a viscosity of about 10 cP at 25°C was obtained.
Next, 97.65 μg of salmon calcitonin (4000 MRC units/ml) was added to 3 ml of this solution and dissolved to obtain a homogeneous solution. The aqueous solution obtained in this way is 32.55μg
Contains (130.2 MRC units)/ml of salmon calcitonin.

() In order to compare with the aqueous solution for nasal administration of the present invention, the comparative solution shown below was obtained.

(d) A homogeneous solution was obtained by adding and dissolving 98.8 μg of salmon calcitonin (4000 MRC units/mg) in three portions of distilled water. The aqueous solution thus obtained was 32.6μg
Contains (130.4 MRC units)/ml of salmon calcitonin.

Example 3 Nasal administration experiment of aqueous calcitonin preparation in domestic rabbits The aqueous calcitonin preparation prepared in Example 2 (a) to (d) was administered into the nasal cavity of Japanese white native breed male domestic rabbits (weight 2.5 to 3.5 kg). Administer 0.975 to 0.978 μg (approximately 3.9 MRC units)/30 μg/Kg, and administer 30 μg before and after administration.
Blood was collected from the ear vein of the rabbit at minutes, 1 hour, 2 hours, 4 hours, and 6 hours.

Intranasal administration was performed without anesthesia, that is, under normal conditions, and the administration device used was a 1 ml syringe with a micropipette tip attached to the tip to enable quantitative administration of the viscous solution. . The serum calcium concentration was measured before and after administration, and the absorbability of calcitonin from the nasal mucosa was investigated. Serum calcium was measured using a calcium measurement kit manufactured by Yatron. The results are shown in FIG. 1 as the degree of decrease (%) in calcium value relative to the serum calcium value before administration of the aqueous calcitonin preparation. 1st
The values shown in the figure are average values for 8 to 9 domestic rabbits.
As is clear from FIG. 1, the aqueous calcitonin preparation for nasal administration of the present invention has high calcitonin absorption.

Example 4 Nasal administration experiment of aqueous insulin preparation in dogs Injected into the nasal cavity of a male beagle dog (weight 9.2 to 10.6 kg) in the same manner as in (d), (e), and (f) of Example 1. An aqueous insulin preparation prepared for the experiment was administered at 3 units/10μ/Kg, and blood was collected from the forearm vein before and after administration. Glyose concentration in plasma was measured by a method using orthotoluidine (Clinical Chemistry).
Chemistry) 8, 215 (1962)). Intranasal administration was performed as described in Example 3. Figure 2 shows changes in plasma glucose in terms of plasma drop rate (%), and all values are average values for 3 to 4 beagle dogs. For comparison, the results of changes in plasma glucose concentration when 3 units/10μ/Kg of insulin dissolved in isotonic phosphate buffer with pH 7.4 and suspended in physiological saline were respectively administered nasally. Also second
Indicated by a broken line in the figure. As is clear from FIG. 2, the aqueous solution for nasal administration of the present invention, in which hydroxypropylcessulose is added to an aqueous base to give it appropriate viscosity, has high insulin absorption.

Example 5 A solution with a viscosity of about 1000 (eP) at 25°C was obtained by dissolving 2 g of hydroxypropyl cellulose (viscosity of 2% aqueous solution at 20°C: 1000 to 4000 cP) in an isotonic vinegar buffer. Ta. Then add 20 vasopressin (70-100 units/mg) to 3 ml of this solution.
A homogeneous solution was obtained by adding and dissolving mg. The aqueous solution thus obtained is approximately 467-667
Contains units/ml of vasopressin. By placing the obtained vasopressin-containing liquid in a suitable nasal container, an aqueous liquid for human nasal administration, which can be administered in an amount of about 0.05 to 0.1 ml at a time, was obtained.

Example 6 A solution having a viscosity of about 1000 (cP) at 25°C was obtained by dissolving 2 g of hydroxypropyl cellulose (viscosity of 2% aqueous solution at 20°C: 1000 to 2000 cP) in physiological saline. Then this solution 4
A homogeneous solution was obtained by adding and dissolving 10 mg of luteinizing hormone-releasing hormone in 1 mL of the solution. The aqueous solution thus obtained contains 2.5 mg/ml of luteinizing hormone-releasing hormone and can be administered at a time from about 0.05 to
An aqueous solution for human nasal administration, which can be administered in 0.1 ml, was obtained.

Example 7 By dissolving 40 g of hydroxypropylcellulose (viscosity of 2% aqueous solution at 20°C from 1000 to 2000 cP) and 1.5 mg of desmopressin acetate in 10 ml of isotonic acetate buffer, the viscosity at 25°C was approximately 10
A homogeneous solution of (cP) was obtained. The aqueous solution thus obtained contains 150 μg/ml of desmopressin acetate, and is an aqueous solution for human nasal administration that can be administered in a dose of about 0.05 to 0.1 ml at a time by placing it in a suitable nasal container. I got it.

Example 8 By dissolving 80 mg of hydroxypropylcellulose (viscosity of 1000-4000 cP in 2% aqueous solution at 20°C) and 20 mg of oxytocin in 10 ml of isotonic citrate buffer, the viscosity at 25°C was approximately 50 cP.
A homogeneous solution of (cP) was obtained. The aqueous solution thus obtained contained 2 mg/ml oxytocin;
By placing this in a suitable nasal spray container, an aqueous solution for human nasal administration, which can be administered in an amount of about 0.05 to 0.1 ml at a time, was obtained.

Example 9 100 mg of hydroxypropyl cellulose (viscosity of 1000 to 4000 cP in a 20% aqueous solution at 20°C) and 75 mg of interferon (100,000 units/ml) lyophilized with human serum albumin were added to an isotonic phosphate buffer solution of 10
ml, the viscosity at 25℃ is approx.
A homogeneous solution of 100 (cP) was obtained. The aqueous solution thus obtained contains 750,000 units/ml of interferon, and is suitable for human nasal administration, where approximately 0.05 to 0.1 ml can be administered at a time by placing it in a suitable nasal container. An aqueous solution was obtained.

Example 10 3.3 g of hydroxypropylcellulose (20°C,
100ml of 2% aqueous solution (viscosity 150-400cP)
A solution with a viscosity of about 1000 (cP) at 25° C. was obtained by dissolving it in physiological saline. Next, add salmon calcitonin (4000 MRC units/
A homogeneous saline solution was obtained by adding and dissolving 0.5 mg). The saline solution thus obtained yielded 0.1 mg (400 MRC units)/ml of salmon calcitonin.

[Brief explanation of the drawing]

FIG. 1 shows the absorption of calcitonin in terms of serum calcium lowering rate when the aqueous solution of the present invention using calcitonin as a peptide hormone was administered nasally. FIG. 2 shows the absorption of insulin in terms of blood sugar lowering rate when the aqueous solution of the present invention using insulin was administered nasally. In Figure 1, 1 is 2 (w/v)% hydroxypropyl cellulose ((a) of Example 2), 2 is 1
(w/v)% hydroxypropylcellulose ((b) in Example 2, 3 indicates the case where a 0.4(w/v)% hydroxypropylcellulose ((c) in Example 2) aqueous solution was used, 4 indicates the comparison Therefore, the case where distilled water (Example 2 (d)) is simply used as the liquid diluent is shown.
In Figure 2, 1 is an isotonic phosphate buffer with a pH of 7.4 containing 1 (w/v)% hydroxypropylcellulose ((d) in Example 1), and 2 is a solution containing 2 (w/v)% hydroxypropylcellulose. Isotonic phosphate buffer with pH 7.4 ((e) of Example 1), 3 is a physiological saline containing 1 (w/v)% hydroxypropylcellulose (Example 1)
(f)) is used, and 4 is simply PH for comparison.
7.4 shows the case where isotonic phosphate buffer and 5 shows the case where simply physiological saline was used.

Claims (1)

[Claims] 1. A polypeptide having physiological activity and hydroxypropyl cellulose contained in an aqueous base suitable for application to the nasal mucosa, and having a viscosity at 25°C in the range of 2 to 4000 cP. Aqueous solution for nasal administration. 2. The aqueous solution for nasal administration according to claim 1, wherein the polypeptides have a molecular weight of 300,000 to 300,000. 3. The aqueous solution for nasal administration according to claim 1 or 2, wherein the polypeptides are physiologically active proteins of peptide hormones, enzyme proteins, or vaccines. 4. The aqueous solution for nasal administration according to any one of claims 1 to 3, wherein the peptide hormone is calcitonin, insulin, luteinizing hormone, releasing hormone, desmopressin, vasobrecin, oxytocin, or interferon. . 5. The aqueous liquid preparation for nasal administration according to any one of claims 1 to 4, which has a viscosity in the range of 2 to 2000 cP at 25°C. 6. The aqueous liquid preparation for nasal administration according to any one of claims 1 to 4, which has a viscosity in the range of 5 to 1000 cP at 25°C. 7. Any one of claims 1 to 6, wherein the aqueous base is water, physiological saline, or a buffer solution.
Aqueous liquid preparation for nasal administration as described in . 8 Claims 1 to 8 whose PH is 3.5 to 7.5
7. The aqueous solution for nasal administration according to any one of item 7. 9 Claims 1 to 9 whose PH is 5.0 to 7.5
The aqueous solution for nasal administration according to any one of item 7.