WO1993021918A1

WO1993021918A1 - Fat emulsion containing fatty acid ester of rhizoxin

Info

Publication number: WO1993021918A1
Application number: PCT/JP1993/000505
Authority: WO
Inventors: Masafumi Hisaoka; Atsushi Kurihara; Atsuko Mizota; Seigo Ueda; Wataru Kato; Tomowo Kobayashi; Kazuhiko Sasagawa
Original assignee: Sankyo Company, Limited
Priority date: 1992-04-28
Filing date: 1993-04-20
Publication date: 1993-11-11
Also published as: AU3905993A

Abstract

A fat emulsion comprising (a) a fatty acid ester of rhizoxin, which is represented by general formula (I) and has a mean particle diameter of 200-950 nm; wherein n represents 7 to 17, (b) at least one fat emulsion base selected from among vegetable oils and mono-, di- and tri-glycerides of synthetic and semisynthetic medium- and long-chain fatty acids, and (c) at least one emulsifying agent selected from among phospholipids and nonionic surfactants. When administered to a living organism, the fat emulsion exhibits a high tendency to accumulate in a cancer tissue, thus exhibiting an excellent anticancer effect.

Description

' ' Specification

[Title of the Invention] Fat emulsion containing rhizoxin fatty acid ester

[Technical field】

TECHNICAL FIELD The present invention relates to a fat emulsion containing an anticancer substance, and more particularly, to a preparation which remarkably improves the anticancer activity of rhizoxin / fatty acid ester.

[Background technology]

In recent years, in order to perform drug treatment more rationally, active efforts have been made to formulate drugs to enhance the efficacy and safety of drugs. When administered to a living body, these preparations can control the release of the drug from the preparation, prevent inactivation of the drug in the body, and provide selective distribution of the drug to the site of action. It is desirable to have characteristics such as the ability to retain the drug locally at the site. One of such preparations is to use a carrier. In other words, this is a method of changing the pharmacokinetics of the drug contained in accordance with the pharmacokinetic characteristics of the carrier.

Carriers currently being studied include ribosomes, microcapsules, microspheres, fat emulsions, polymer complexes, and many others. Among them, the fat emulsion is easy to include the fat-soluble compound and is relatively easy to prepare, and it is possible to prepare fat particles having different average particle diameters according to the purpose. Formulation with a particle size is easy. Therefore, steroids (Japanese Patent Application Laid-Open No. 57-16818), Prossu Grandin's (Japanese Patent Application Laid-Open No. Hei 1-157094, Japanese Patent Application Laid-Open No. Hei 1-157095) and Fat emulsions such as anticancer agents (Japanese Patent Application Laid-Open No. 59-122422) have been reported, and a certain improvement in therapeutic effect has been recognized.

Fat emulsions of anticancer drugs impart selectivity to anticancer drugs because existing anticancer drugs have low selectivity for cancer and have a strong cell killing effect, but also cause damage to normal cells and develop side effects. It is a preparation prepared for the purpose of However, its selective specificity has not yet been fully effective, and complete treatment or prevention of cancer with chemotherapy has not been realized. The present inventors have found that the anti-cancer drug obtained by selecting lysoxine fatty acid ester as the anti-cancer agent and the fat emulsion described below as a fat emulsion has a significantly higher anti-cancer effect than the fat emulsion of the conventional anti-cancer drug ⁵ '. The present invention was found to have an effect and low toxicity, and completed the present invention.

The fat emulsion of the present invention,

(a) Average 'particle strength at ¾00-950nm

—General formula

(In the formula, n represents? To Γ7.) A rhizoxin 'fatty acid ester represented by

(b) vegetable oils and at least one fat emulsion base selected from mono-, di- or triglycerides of Z or synthetic or semi-synthetic medium or long chain fatty acids,

(c) at least one emulsifier selected from phospholipids and Z or a nonionic surfactant \

The present invention relates to a fat emulsion comprising

Rhizoxin is represented by the following formula and is known in the literature (Journal of Antibiotics (J. Antibiotics), Vol. 37, pp. 354-362, 1989). The compound is known to have anti-tumor activity.

^One fatty acid ester represented by the following formula is a known compound described in JP-A-62-87.

As the lysoxine fatty acid ester in the present invention, n = 7 to; L7 is preferred, more preferably n = 9 to 17, and most preferably n = 13 to 17. It is.

The fat emulsion of the present invention may have any average particle diameter in the range of 200 to 950 nm, but is preferably 200 to 600 nm, more preferably 200 to 400 nm.

Examples of the fat emulsion base that forms the fat emulsion of the present invention include vegetable oils such as soybean oil, cottonseed oil, sesame oil, safflower oil, and corn oil; mono-, di-, or triesters of medium- or long-chain fatty acids of glycerin ( For example, monoglycera cabronate Single glycerides such as amide, caprylic acid monoglyceride, caproic acid diglyceride, caprylic acid diglyceride, dioctyldecyl triglyceride, caprylic acid triglyceride, medium chain fatty acid triglyceride (trade name Coat Ace MT), hard fats (Mixed glycerides such as pharmazol) can be used singly or as a mixture, and preferably, medium-chain fatty acid triglyceride (trade name: COLOACE) MT), and the concentration of the fat emulsion base is preferably 1 to 30% (V / V), more preferably 5 to 20% (V / V).

As the emulsifier for forming the fat emulsion of the present invention,

Phospholipids such as phosphatidylcholine (also known as lecithin), phosphatidylethanolamine, phosphatidylserine, etc. ·

Polyoxyethylene castor oil esters, such as polyoxyethylene castor oil 50, boroxyethylene castor oil 60; polyoxyethylene hydrogenated castor oil 50, polyoxyethylene hydrogenated castor oil 60 (HC 0 Polyoxyethylene hard castor oil esters such as 60); polyoxyethylene sorbitan fatty acid esters such as polyoxyethylene sorbitan monostearate and polyoxyethylene sorbitan trioleate; polyoxyethylene [42]ポリ Polyoxypropylene [67] glycol, polyoxystyrene [160] polyoxypropylene [301 polyoxypropylene-boroxypropylene copolymer such as glycol; polyoxystyrene stear Polyoxyethylene fatty acid esters, such as polyoxyethylene glycol, polyoxyethylene laurate; Nonionic surfactants such as polyoxyxylene sterol ethers such as ethylene cholesterol ether and polyoxycholestanol ether can be used, and preferred are polyoxyxylene hardened castor oil esters. , Also, ? The concentration of the precipitating agent is preferably from 0.1 to 12¾ (WA), more preferably from 0.01 to L¾ (W / V).

In addition, emulsifying auxiliaries, stabilizers, tonicity agents, pH regulators, drug stabilizing agents (antioxidants), etc. can be used according to the conditions. Xymethylcellulose 'sodium, polyvinylpyrrolidone Examples of suitable stabilizers include glycerin S. Examples of suitable tonicity agents include budou sugar and salt. Examples of suitable pH regulators include various acid bases (eg, hydrochloric acid, sodium phosphate, sodium acetate). , Sodium hydroxide).

The fat emulsion of the present invention can be diluted with an appropriate amount of water after preparation, and the dilution factor is not particularly limited as long as the emulsion is stable, but is preferably 2 to 100 times.

The preferred combination of the composition of the fat emulsion is 0.01-1¾W / V for the rhizoxin fatty acid ester, 0.01-1¾W / V for the emulsion, and 30¾V / V for the emulsifying base.

Among the fat emulsions of the present invention, preferred are:

1) Fat emulsion with an average particle size of 200-600nm

2) Fat emulsion with average particle size of 200-400nm

3) Fat emulsion using medium chain fatty acid triglyceride as the base of fat emulsion

4) Fat S-type emulsion using boroxyxylene hardened castor oil ester as emulsifier

5) Fat emulsion wherein n is 13 to 17

6) the average particle size is 910nm,

'' Luminous emulsion with medium-chain fatty acid triglyceride as the base of lunar fat emulsion, boroxyethylene hard castor oil ester as emulsifier, and _n of 13 to 17

7) The average particle size is 600nm,

Medium-chain fatty acid triglyceride as a fat emulsion base, boroxyethylene hard castor oil ester as an emulsifier, and n is 13 to 17

8) The average particle size is 380mn,

Luminous fat emulsion using medium-chain fatty acid triglyceride as the base of the fat emulsion, polyoxyethylene hydrogenated castor oil ester as the emulsifier, and n of 13 to 17

9) the average particle size is 221nm,

Medium-chain fatty acid triglyceride is used as a fat emulsion base and poly is used as an emulsifier. Fatty emulsion with _η of 13 to 17 using oxyethylene hydrogenated castor oil ester

Can be given.

The fat emulsion of the present invention can be produced, for example, by the method described in tff.

After dissolving rhizoxin 'fatty acid ester, emulsifier, and other necessary auxiliary components (such as the emulsifying auxiliary described above) in the fat emulsion base, add an appropriate amount of water, and use a high-pressure emulsifier for an appropriate time ( Fatty emulsions having various average particle diameters can be prepared by emulsification.

As a high-pressure emulsifier used for emulsification, a microfluidizer (maximum processing pressure 1

600 kg / cm microfluidic clay, Mantongo mono-lin type injection emulsifier (maximum processing pressure 700 kg / cm ² , Manton Gorin ').

[Example]

Hereinafter, the present invention will be described specifically with reference to examples.

(Example 1)

Rhizoxin palmitate (Compound with η = 15: hereafter referred to as “Compound 1 J.”) 500 mg, butylhydroxytoluene 50 mg, Nichol HC060 (Polyoxyethylene hydrogenated castor oil ester, Nikko Chemicals Co., Ltd.) L O Omg was added to 1 Oml of Coto Ace MT (medium-chain fatty acid triglyceride, manufactured by Nisshin Oil Co., Ltd.), and dissolved by stirring with a magnetic stirrer. The total volume was adjusted to 10 O ml, and the mixture was subjected to oiling three times at a pressure of 140 kg / cm ² using a high-pressure emulsifier Microfluidizer M-120E to obtain a fat emulsion having an average particle diameter of 910 runs. .

(Example 2)

A fat emulsion having an average of 600 was obtained in the same manner as in Example 1 except that the number of emulsifications described in Example 1 was changed to 5.

(Example 3)

500 mg, 50 mg of butylhydroxytoluene and 50 mg of Nichol HC060 were added to Coat Ace MT 10 ml, and the mixture was stirred and dissolved with a magnetic stirrer. Distilled water for injection is added to this solution to make the total volume 100 ml, Using a pressure emulsifier, a microfluidizer M-120E, milking was performed five times at a pressure of 85 ° kg / cm ² to obtain a fat emulsion having an average particle diameter of 380 nm.

(Example 4)

A fat emulsion having an average particle size of 22 lrnrn was obtained in the same manner as in Example 3, except that the amount of 2'sochol HC060 described in Example 3 was changed to 500 mg.

(Comparative Example 1)

A fat emulsion having an average particle diameter of 99 nm was obtained in the same manner as in Example 3, except that the amount of Nikkor HC060 described in Example 3 was changed to 400 mg.

(Comparative Example 2)

A fat emulsion having an average particle diameter of 69 nm was obtained in the same manner as in Example 3, except that the amount of Nikkor HC060 described in Example 3 was changed to 1200 mg.

[The invention's effect]

By setting the average particle size of the fat emulsion of the present invention to 200-950 nm, the anticancer effect of rhizoxin / fatty acid ester can be remarkably enhanced.

The previously reported fat emulsion containing an anticancer drug (Japanese Patent Application Laid-Open No. 59-122324) has been studied only for its effect on an intraperitoneal transplantation model of ascites liver cancer. However, in this evaluation system, the development of new blood vessels found in human cancer tissues is unknown, and the selection specificity is unknown. Originally, in order to examine the selective specificity of a fat emulsion, it is important to use a solid cancer model that more closely resembles human cancer and to evaluate the efficacy of an anticancer drug in a tumor state in which new blood vessels have developed.

Therefore, the present inventors studied the anticancer effect using the following tumor tissues. In other words, in solid malignant tumors, when the tumor mass grows to a certain size, new blood vessels having characteristics different from those of normal tissue capillaries develop. Since this new blood vessel is a blood vessel having an incomplete endothelium, it can pass through even a large molecule substance to reach a cancer tissue. On the other hand, many of the capillaries in normal tissues have blood vessels with continuous endothelium and cannot pass large molecular substances.] (Critical Reviews in Therapeutic Drug Carrier Systems, 6, Issue 3, 193, (1989)). As described above, there is a structural difference between the two blood vessels, and it is thought that the preparation of a fat emulsion having a certain size will make a significant difference, and the tumor selectivity will be obtained. available.

Thus, the antitumor activity of the compounds of the present invention having various particle sizes was examined.

(Test Example 1)

(1) Sample preparation-Fat emulsions prepared in Examples 2, 3 and 4, and Comparative Examples 1 and 2 (average particle diameters are 910 nm, 600 nra, 380, 221 nm, 99 nm and 69 rim) were used as samples 1, 2, 3, 4, and 1, respectively. In addition, as a positive control of the anticancer effect, compound 110 O mg. Butyl hydroxytoluene 10 mg and nicole HC06060 Omg were dissolved in N, N-dimethylacetamide to make the total amount 1 O O mg. The solution was diluted to an appropriate dilution with a physiological saline solution to prepare a sample X. A physiological saline solution containing no rhizoxin 'fatty acid ester was used as control sample 0.

(2) Test animals

Three to six BDF _L mice (female, 9 weeks old, weighing 19 to 22 g) were used as one group, and one group was used per sample.

(3) In vivo anti-cancer effects in tumor-bearing mice

1 × 10 ⁶ Z 576 fibrosarcomas of mouse experimental cancer were implanted subcutaneously in the right axilla of each animal group consisting of 6 mice. The day of transplanting the cancer cells was set to day 0, and each sample was intravenously administered once 13 days after that (daylS: a time when new blood vessels were observed), and the tumor size and the survival of the mice after the administration were observed. The same treatment was performed for the control sample 0.

The anticancer activity was evaluated based on (a) the size of the tumor mystery on day, (b) the survival rate of tumor-bearing mice (ILS%: Increase in Life Span), and (c) complete cure on day 120. did. The tumor and the survival rate were calculated from the following equations. Major axis + minor axis *

Tumor diameter =--one

Two Median days of survival for sample treated group

Survival rate = (111) Median days of survival in the control sample-administered group The anti-cancer activity of each sample was examined at various doses and compared at the obtained optimal dose. That is, the highest dose that did not cause mouse s' death within 10 days after sample administration and reduced body weight to less than 15% was defined as the optimal dose.

Table 1 shows the results of the anticancer activity at that time. -

[Table 1] Antitumor activity against M5076 fibrosarcoma tumor-bearing mice

S formula fee Survival days on average day44 Survival rate On dayl20 Particle size Dose Average tumor diameter Completely cured case

(nm) (mg / kg) imm) (days) (ILS%)

1 910 54 3.0〉 120.0〉 224 3/6

2 600 60 0.8〉 120.0> 224 5/6

3 380 40 1.5〉 109.0> 195 3/6

4 221 15 1.1> 120.0> 224 3/6

99 99 4.5 9.4 62.5 69 0/6

Π 69 6.0 12.0 65.5 77 0/6

X 6.0 17.7 60.0 62 0/6

0 37.0 0/6

* 5/6 force ^ death by day44 **: Median The tumor diameters of Samples 1, 2, 3, 4 and i-administered group were smaller than that of Sample X which was a solution, and an excellent antitumor effect was observed. In particular, mice that administered samples 1, 2, 3, and 4 exhibited long-lived mice, and three out of six mice showed complete disappearance of tumor aneurysm.

(Experimental example 2)

1 × 10 ⁶ mice / animal fibrosarcoma of mouse experimental cancer were implanted subcutaneously in the right fovea of each group consisting of 3 animals. Let the cancer cell transplant day be day 0, and 13 B later,

On (day 13: a time when a new ^ iflL tube was observed), Samples 2, 4, ィ and X prepared in (1) of Test Example 1 were intravenously administered once. The doses were 60 rag / kg, 15 mg / kg, 4.5 mg / kg, and & mg / kg, respectively, which were the optimal doses at which the anticancer effect of each sample was most strongly exhibited. Six hours after the administration of the sample: at 2, 3, 4 and each time point, cancer tissue was searched from one group of animals and the concentration of lysoxine / palmitate ester in & 濃度 was measured. The results are shown in Figure 1.

Rhizoxin cancer tissue - palmitic acid ester concentration, the sample 2 as compared with the sample X, 4 and I especially ₉ higher was observed trend that forces s in the case of administration, high when given samples 2 and 4 projecting, moreover Showed persistence.

Based on the above results, it is considered that the fat emulsion of lysoxine / fatty acid ester with an average particle size of 200 to 950 nm accumulates in cancer cells at a higher concentration than other fibers when administered to a living body, and is extremely selective. It turned out that it could be an excellent control »j.

[Brief description of the drawings 1

FIG. 1 is a graph showing the accumulation property of a drug on a cancer tissue.

Claims

1.1

1 =

(a) Average particle diameter is 200-950nm

One j¾S expression

(Wherein n represents 7 to 17.) (b) selected from vegetable oils and / or mono-, di- or triglycerides of synthetic or semi-synthetic medium or long chain fatty acids. A fat emulsion comprising at least one fat emulsion base, and (c) at least one emulsifier selected from phospholipids and / or nonionic surfactants.

2. The fat emulsion according to claim 1, having an average particle size of 200-600 ™.

3. The fat emulsion according to claim 1, having an average particle size of 200 to 400ππι.

Four. 2. The fat emulsion according to claim 1, which is a fat emulsion base containing triglyceride fatty acid.

Five. The fat emulsion according to claim 1, wherein the emulsifier is polyoxyethylene hard castor oil ester.

6. The fat emulsion according to claim 1, which has n forces s13 to 17.

7. In claim 1,

The average particle size is 910nm,

A fat emulsion wherein the fat emulsion base is a medium fatty acid triglyceride, the emulsifier is a polyoxyethylene hardened castor ester, and n is 13 to 17. . Claim 1. The average diameter of the retainer is 600 nm,

A fat emulsion wherein the fat emulsion base is a medium-chain fatty acid triglyceride, the emulsifier is polyoxyethylene hard castor oil ester, and n is 13 to 17. . In claim 1,

The average particle size is 80 nm,

Luminous fat emulsion base strength Fat emulsion in which S is a medium-chain fatty acid triglyceride, the emulsifier is polyoxyethylene hydrogenated castor oil ester, and n is 13 to 17. 0. In claim 1,

The average particle mystery is 221 hidden,

A fat emulsion in which the base of lunar fat emulsion is medium triglyceride triglyceride, the emulsifier is polyoxyethylene hydrogenated castor oil ester, and n is 13 to 17. 1. A method for treating or preventing a mammal having a cancer disease, which comprises administering the fat emulsion according to claim 1 to a mammal.