Title:  Method of preservation of vaccines with polybiguanide

United States Patent:  6,403,363

Assignee:  Medeva Europe Limited (London, GB)

Filed:  July 12, 2001

Foreign Application Priority Data:  Mar 20, 1997[GB] (9705740)

A method for preventing or reducing bacterial contamination of a viral vaccine is disclosed. The method comprises adding an effective preserving amount of a polybiguanide-containing preservative composition to a solution containing vaccine virus or virus antigen. The method is particularly useful in preventing or reducing bacterial contamination of process solutions involved in the manufacture of influenza vaccines.

SUMMARY OF THE INVENTION

In a first aspect, the principles of the invention provide a method of preventing or reducing bacterial contamination of a viral vaccine, wherein the method comprises adding to a solution containing virus or virus antigen, an effective amount of a preservative composition containing polybiguanide.

Accordingly, in a first aspect, the invention provides a method of preventing or reducing bacterial contamination of a viral vaccine, which method comprises adding to a solution containing vaccine virus or virus antigen an effective preserving amount of a preservative composition containing polybiguanide.

In another aspect, the invention provides a preserved viral vaccine composition comprising a solution containing vaccine virus or virus antigen and an effective preserving amount of a preservative composition containing polybiguanide.

In one embodiment of the invention, the solution may be the final form of the vaccine immediately prior to or after filling into dosage containers such as vials, ampoules and the like, particularly multi-dose containers.

In another embodiment, the solution may be a process solution, the term "process solution" as used herein referring to any solution containing the vaccine virus or viral antigens derived from the vaccine virus up to the point at which the vaccine is filled into dosage containers.

Examples of "process solutions" are solutions of virus harvested from the medium in which the virus has been grown, such as the allantoic fluid from eggs, or the supernatant from a cell culture. Other examples include solutions containing disrupted virus and free antigens such as surface antigens, as well as partially purified and purified solutions of antigens. The term "process solution" can also include the viral growth medium itself, the preservative being added to the medium at the beginning of or during the growth phase of the virus.

The vaccine can contain a single strain of virus, or antigens from a single strain of virus, or it can contain a blend of antigens from different viral strains. For example, in the case of influenza vaccines, the vaccine can contain antigens from one or more strains of influenza A together with antigens from one or more strains of influenza B.

The polybiguanide is typically a polymeric compound containing the repeating unit:

--[NH--C(NH)--NH--C(NH)--NH--R]_n --

wherein R is a divalent hydrocarbon chain, preferably having at least 2 carbon atoms; and n represents the number of repeating units and is at least 2.

More usually, n is in the range from 3 to 20, for example from 4 to about 16, preferably from 5 to 12, and more preferably from 5 to 7. The values given for n are the average values, since a given solution of a polybiguanide will frequently contain a mixture of molecules of differing chain lengths. For example, a solution of a polybiguanide can contain molecules in which n ranges from 1 to about 40. A preferred polybiguanide is one in which the average value for n is 5.5.

The molecular weight of the polymer, excluding the weight of any acid present in the form of acid addition salts of the polymer, may be up to 12,000 or more but is usually less than 5000, and is preferably in the range from about 750 to about 3000.

A presently preferred polybiguanide is poly(hexamethylene) biguanide, the INN for which is polyhexanide. One example of a proprietary product including poly(hexamethylenebiguanide) is "Cosmocil CQ" (RTM) manufactured by Zeneca PLC.

The polybiguanide can be presented in the form of an acid addition salt, and examples of such salts include the salts formed with hydrochloric acid, sulphuric acid, phosphoric acid and acetic acid, a presently preferred salt being the hydrochloride.

The amount of polybiguanide, as defined above, is an effective preserving amount, i.e. an amount which is at least sufficient to preserve the solution. Thus the amount is at least sufficient to provide a biostatic effect, but preferably is an amount sufficient to have a biocidal effect. The amount of polybiguanide is generally selected so as to give at least a one log reduction in the level of microbial contaminants during five hours. More preferably the amount is selected so as to give at least a two log reduction in microbial contaminants over 5 hours. The concentration of the polybiguanide, for example poly (hexamethylenebiguanide), typically can be in the range of about 0.0001% to 0.1% w/v, and preferably is less than 0.1% w/v. More usually, the concentration will be in the range from about 0.0005% to 0.05% w/v. A particularly preferred concentration is approximately 0.002% w/v.

The type and amount of preservative are chosen such that whilst being active against non-viral microbial components of the vaccine process solutions, they do not adversely affect the ability of the virus to replicate or have an adverse effect on the quantities and properties of the viral antigens.

The method of the present invention is particularly applicable to enveloped viruses, i.e. viruses having a lipid bi-layer. Examples of such viruses include paramyxoviruses such as Sendai virus, orthomyxoviruses such as influenza viruses, toga viruses such as Semliki forest virus, and rhabdoviruses such as vesicular stomatitis virus.

In one preferred embodiment of the invention, there is provided a method of preventing or reducing bacterial contamination of an influenza vaccine during manufacture, which method comprises adding to a process solution containing influenza vaccine virus or influenza viral antigen (and in particular surface antigens such as haemaggiutinin and neuraminidase) an effective preserving amount of a polybiguanide.

The method of the invention is particularly useful for preventing microbial contamination of influenza vaccines of the "Fluvirin" type referred to above.

Claim 1 of 28 Claims

We claim:

1. A method of preventing or reducing bacterial contamination of a viral vaccine, said method comprising adding an effective preserving amount of a preservative composition containing a polybiguanide to a solution containing vaccine virus or virus antigen, wherein the polybiguanide is a polymeric compound comprising a plurality of repeating units, each repeating unit being represented by the formula:

--[NH--C(NH)--NH--C(NH)--NH--R]_n --

wherein R is a divalent hydrocarbon chain and the value of n in each repeating unit is in the range from 2 to 40.

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