JPH0133449B2 - - Google Patents

Description

[Detailed description of the invention]

The present invention relates to stabilized frozen or lyophilized live measles vaccines. A live measles vaccine is a vaccine whose main ingredient is a live, attenuated measles virus, and when administered to a person susceptible to measles, the recipient acquires semi-permanent immunity. Since this vaccine aims at active immunization with a live virus, it must maintain sufficient viral activity (infectivity) during the period after manufacture until it is used for inoculation. However, during the manufacturing process (cryopreservation, freeze-drying, etc.) and in the freeze-dried state of the final product, the live measles virus tends to become inactivated over time and the titer (infectivity) of the vaccine tends to decrease. . In particular, there is a remarkable tendency for the titer to decrease during the freeze-drying process and during the storage of the freeze-dried product, creating a major bottleneck in the production, transportation, and storage of this vaccine. As a countermeasure,
Although a method of adding lactose-glutamate as a stabilizer (Japanese Patent Publication No. 22271/1971) has been reported, none of the stabilizers proposed so far have been satisfactory. Based on this technical background, the present invention provides a frozen or freeze-dried live measles vaccine whose titer is stable for a long period of time. That is, the present invention provides a live measles vaccine solution containing approximately
0.5-5% hexa-tetrahydric sugar alcohol and about 1
It is a live measles vaccine that contains ~7% disaccharide and is frozen or freeze-dried. The live measles vaccine stock solution in the present invention includes:
It contains live attenuated measles virus as an active ingredient and is based on the "Biological Product Standards [Ministry of Health and Welfare Notification (July 1970)
17th) No. 263] may be used. The present invention provides the above live measles vaccine stock solution with approximately
0.5-5% hexa-tetrahydric sugar alcohol and about 1
By freezing or freeze-drying the mixture containing ~7% of both disaccharides, a remarkable stabilizing effect is achieved, and such an effect cannot be achieved when using only either sugar alcohol or sugar. . The hexa- to tetravalent sugar alcohol may be either chain or cyclic. Examples of hexavalent sugar alcohols include sorbitol, inositol, mannitol, dulcitol, allitol, talitol, glucitol, and iditol, and examples of pentahydric alcohols include xylitol, adonitol, and arabitol. Furthermore, examples of the tetravalent sugar alcohol include erythritol and threitol. Although these sugar alcohols exist in stereoisomers and racemic forms, they can be used without distinction in the present invention. Further, these sugar alcohols may be used alone or in combination of two or more. As the disaccharide, for example, lactose, sucrose, etc. are conveniently used. The content of hexa- to tetravalent sugar alcohol in the vaccine of the present invention is approximately 0.5-5% based on the total amount of the vaccine solution subjected to freezing or lyophilization (all percentages herein are expressed as weight/volume percentages). ) preferably about 1 to 3%;
The preferred content of disaccharides is about 1 to 7%, especially about 4 to 6%, based on the total amount of the vaccine solution. Approximately 0.5-5% 6-4 in the vaccine of the present invention
The titer stabilizing effect of the coexistence of a sugar alcohol of about 1 to 7% and about 1 to 7% of a disaccharide can be further enhanced by the coexistence of human serum albumin (hereinafter abbreviated as HSA). this
The concentration of HSA is approximately 0.1 to 1%, especially approximately 0.2%, relative to the vaccine solution to be frozen or freeze-dried.
~0.5% is preferred. In the case of freeze-dried vaccines, by adding dextran and gelatin as excipients to the vaccine solution and performing the freeze-drying process, the appearance shape becomes carmelized during freeze-drying, and a beautiful freeze-dried product can be obtained. Obtainable. The preferred concentration of dextran and gelatin is about 0.1 to 1%, especially about 0.2 to 1%, respectively, relative to the vaccine solution.
It is 0.5%. These stabilizers and excipients are dissolved in advance in an aqueous solvent (water, physiological saline, phosphate buffer, etc.), mixed with the live attenuated measles vaccine stock solution to the above concentration, and then frozen or Subjected to freeze drying. In this case, it may be stored frozen at a temperature of -40° C. or lower, then thawed and immediately freeze-dried, or it may be stored frozen again, then thawed and subjected to freeze-drying. The effects of the present invention will be explained in more detail below using experimental examples. Experimental Example 1 Fetal primary cultured cells obtained from 10-day-old embryonated chicken eggs that are not infected with infectious diseases were inoculated with live attenuated measles virus (Schwartz strain), and TCM
-199 medium at 28 to 32°C for 7 to 14 days, and the resulting virus suspension is purified by a conventional method to obtain a live attenuated measles virus vaccine stock solution. The above-mentioned virus suspension and vaccine stock solution were subjected to various tests based on the above-mentioned biological product standards and confirmed to be suitable. The attenuated live measles virus vaccine stock solution contains the additives shown in Table 1 below and has a titer of 10 ⁵ TCID ₅₀ /
ml of vaccine solution was prepared, immediately dispensed into vials (2 ml capacity), and freeze-dried by a conventional method. For each freeze-dried product, the stability of the titer before and after the freeze-drying process and during storage (4°C) for 2 years after freeze-drying was measured by an infectious titer measurement test. The results were as shown in Table 1. In addition, in the following description, the titer is
Express as log ₁₀ TCID ₅₀ /ml.

【table】

[Table] Experimental Example 2 The raw attenuated measles virus vaccine obtained in Experimental Example 1 contained the additives shown in Table 2 and 0.3% each of dextran and gelatin.
A vaccine solution with a titer of 5.0 was prepared, and Experimental Example 1
When freeze-dried in the same manner as above, a freeze-dried product with a beautiful appearance was obtained. For each freeze-dried product, the titer was measured before and after the freeze-drying process and after 1.5 years of storage (4°C) after freeze-drying. The results were as shown in Table 2.

[Table] Experimental Example 3 From the live attenuated measles virus vaccine stock solution obtained in Experimental Example 1, a vaccine solution containing the additives shown in Table 3 and 0.3% each of dextran and gelatin and having a titer of 5.0 was prepared. Freeze-drying was carried out in the same manner as in Experimental Example 1. For each freeze-dried product, the titer was measured before and after the freeze-drying process and after 1.5 years of storage (4°C) after freeze-drying. The results were as shown in Table 3.

[Table] Experimental Example 4 From the live attenuated measles virus vaccine stock solution obtained in Experimental Example 1, the vaccine solutions listed in the left column of Table 4 (Samples No. 1 to 12: all titers 5.0) were prepared. After repeating freezing and thawing five times for each sample, it was frozen at -70°C for one year, and after thawing, it was further freeze-dried. The changes in titer were as shown in Table 4. In addition, the circle mark in the table indicates that the vaccine liquid contains the component.

【table】

【table】

Claims (1)

[Scope of Claims] 1. A live measles vaccine prepared by freezing or freeze-drying a live measles vaccine solution containing about 0.5 to 5% of a hexa- to tetravalent sugar alcohol and about 1 to 7% of a disaccharide. 2. The live measles vaccine according to claim 1, which further contains human serum albumin. 3. The live measles vaccine according to claim 1 or 2, further comprising dextran and gelatin.