Fermentation media free of animal-derived components for production of diphtheria toxoids suitable for human vaccine use

A diphtheria toxoid obtainable by a process comprising:

• (i) growing a culture of a strain of Corynebacterium diphtheriae expressing a diphtheria toxin in a fermentation medium free of animal-derived components;
• (ii) purifying the diphtheria toxin from the fermentation medium to obtain a diphtheria toxin solution that was at least 90% pure;
• (iii) adjusting the concentration of diphtheria toxin in the diphtheria toxin solution to at least 3000 Lf/mL to obtain a concentrated solution;
• (iv) adding to the concentrated solution (a) an amino acid at a final concentration of no more than 0.025 M and (b) formaldehyde at a final concentration in the range of 0.75-1% to obtain a detoxification solution;
• and
• (v) incubating the detoxification solution to obtain the diphtheria toxoid.

A diphtheria toxoid for use in human vaccination, obtained by a process comprising:

• (i) growing a strain of Corynebacterium diphtheriae expressing a diphtheria toxin or a derivative thereof in at least 100 L of a fermentation medium that is free from animal-derived components, optionally wherein the fermentation medium comprises yeast extract;
• (ii) purifying the diphtheria toxin or derivative from the fermentation medium to obtain a purified diphtheria toxin or derivative, wherein the purified toxin or derivative is at least 90% pure;
• (iii) adding formaldehyde to the purified diphtheria toxin or derivative;
• and
• (iv) incubating the purified diphtheria toxin or derivative from step (iii) to obtain the diphtheria toxoid,
• wherein the diphtheria toxoid is cross-linked by formaldehyde to at least one component of the fermentation medium.

A diphtheria toxoid for use in human vaccination, obtained by a process comprising:

• (i) growing a culture of a strain of Corynebacterium diphtheriae expressing a diphtheria toxin in a fermentation medium free of animal-derived components;
• (ii) purifying the diphtheria toxin from the fermentation medium to obtain a diphtheria toxin solution that was at least 90% pure;
• (iii) adjusting the concentration of diphtheria toxin in the diphtheria toxin solution to at least 2000 Lf/mL to obtain a concentrated solution;
• (iv) adding to the concentrated solution (a) an amino acid at a final concentration of no more than 0.025 M and (b) formaldehyde at a final concentration in the range of 0.75-1% to obtain a detoxification solution;
• and
• (v) incubating the detoxification solution to obtain the diphtheria toxoid,
• wherein the diphtheria toxoid is cross-linked by formaldehyde to at least one component of the fermentation medium.

A diphtheria toxoid obtainable by a process comprising:

• (i) growing a strain of Corynebacterium diphtheriae expressing a diphtheria toxin or a derivative thereof in at least 100 L of a fermentation medium that is free from animal-derived components, optionally wherein the fermentation medium comprises yeast extract;
• (ii) purifying the diphtheria toxin or derivative from the fermentation medium to obtain a purified diphtheria toxin or derivative, wherein the purified toxin or derivative is at least 90% pure;
• (iii) adding formaldehyde to the purified diphtheria toxin or derivative;
• and
• (iv) incubating the purified diphtheria toxin or derivative from step (iii) to obtain the diphtheria toxoid.