High potency stable formulations of vaginal lactobacillus

Abstract

This invention provides for a dry preserved formulation of Lactobacillus suitable for administration to people as a pro-biotic or live biotherapeutic treatment where the formulation is stable, has high potency, and contains no animal-derived excipients.

What is claimed is: 
     
         1 . An aqueous bacterial suspension of vaginal  Lactobacillus  species, having no animal-derived excipients, where the suspension results from a combination of a cell pellet of vaginal  Lactobacillus  species with an aqueous preservation medium consisting essentially of:
 (i) trehalose at between 5-20%, w/v;   (ii) xylitol at between 2-9%, w/v;   (iii) sodium ascorbate 0.5-1.5%, w/v; and   (iv) sodium phosphate at between 10-50 mM.   
     
     
         2 . The bacterial suspension of  claim 1 , wherein the aqueous preservation medium optionally comprises sodium glutamate at between 0-5%. 
     
     
         3 . The bacterial suspension of  claim 1 , wherein the vaginal  Lactobacillus  species has the ability to produce greater than 0.5 ppm of hydrogen peroxide under effective culture conditions. 
     
     
         4 . The bacterial suspension of  claim 1 , wherein the vaginal  Lactobacillus  species is selected from the species consisting of  Lactobacillus crispatus, Lactobacillus jensenii  and  Lactobacillus gasseri.    
     
     
         5 . The bacterial suspension of  claim 1 , wherein the aqueous preservation medium consists essentially of:
 (i) trehalose at between 5-15%, w/v;   (ii) xylitol at between 2-7%, w/v;   (iii) sodium ascorbate 0.5-1.0%, w/v; and   (iv) sodium phosphate at between 10-30 mM.   
     
     
         6 . The bacterial suspension of  claim 5 , wherein the aqueous preservation medium optionally comprises sodium glutamate at between 0-5%. 
     
     
         7 . The bacterial suspension of  claim 1 , wherein the suspension is lyophilized to yield a dry powder. 
     
     
         8 . The dry powder of  claim 7 , wherein the powder has a water activity value of less than 0.220. 
     
     
         9 . The dry powder of  claim 8 , wherein the powder is combined with an inactive excipient at a ratio of powder: excipient of between 1:1 and 1:10 w/w. 
     
     
         10 . The dry powder of  claim 9 , wherein the excipient is maltodextrin. 
     
     
         11 . The dry powder of  claim 9 , wherein the ratio of powder: excipient is between 1:1 and 1:5 w/w. 
     
     
         12 . The dry powder of  claim 7 , wherein the powder is contained within a plastic housing designed for vaginal administration. 
     
     
         13 . A method of preserving  Lactobacillus  spp. under dry conditions without animal-derived excipients, the method comprising:
 (i) obtaining an aqueous suspension of vaginal  Lactobacillus  species having a cell concentration between 10 8  to 10 13  per ml;   (ii) centrifuging the solution to form a bacterial cell pellet; and   (iii) resuspending the bacterial cell pellet in an aqueous preservation medium consisting essentially of:
 (a) trehalose at between 5-20%, w/v; 
 (b) xylitol at between 2-9%, w/v; 
 (c) sodium ascorbate 0.5-1.5%, w/v; and 
 (d) sodium phosphate at between 10-50 mM; 
   
       where the weight ratio of cell pellet wet weight (grams) to preservation media (mL) is between 1:1 and 1:5 grams of cell pellet to milliliter of preservation media to yield a bacterial suspension. 
     
     
         14 . The method of  claim 13 , wherein the aqueous preservation medium optionally comprises sodium glutamate at between 0-5%. 
     
     
         15 . The method of  claim 13 , wherein the vaginal  Lactobacillus  species has the ability to produce greater than 0.5 ppm of hydrogen peroxide under effective culture conditions. 
     
     
         16 . The method of  claim 13 , wherein the vaginal  Lactobacillus  species is selected from the species consisting of  Lactobacillus crispatus, Lactobacillus jensenii  and  Lactobacillus gasseri.    
     
     
         17 . The method of  claim 13 , wherein the aqueous preservation medium consists essentially of:
 (a) trehalose at between 5-15%, w/v;   (b) xylitol at between 2-7%, w/v;   (c) sodium ascorbate 0.5-1.0%, w/v; and   (d) sodium phosphate at between 10-30 mM.   
     
     
         18 . The method of  claim 17 , wherein the aqueous preservation medium optionally comprises sodium glutamate at between 0-5%. 
     
     
         19 . The bacterial suspension of  claim 13 , wherein the bacterial suspension is lyophilized to yield a dry powder. 
     
     
         20 . The dry powder of  claim 13  wherein the powder has a water activity value of less than 0.220. 
     
     
         21 . The dry powder of  claim 20 , where the dry powder is combined with an inactive excipient at a ratio of powder: excipient of between 1:1 and 1:10 w/w. 
     
     
         22 . The dry powder of  claim 21 , wherein the excipient is maltodextrin. 
     
     
         23 . The dry powder of  claim 21 , wherein the ratio of powder:excipient is between 1:1 and 1:5 w/w. 
     
     
         24 . A method of treating abnormal vaginal microbiota in a woman comprising the steps of:
 (i) selecting a woman having a diagnosis of abnormal vaginal microbiota;   (ii) administering an antibiotic in an amount effective to reduce the level of abnormal vaginal microbiota;   (iii) following step ii, administering a dry powder derived from an aqueous bacterial suspension of vaginal  Lactobacillus  species with no animal-derived excipients where the suspension results from a combination of a cell pellet of vaginal  Lactobacillus  species with an aqueous preservation medium consisting essentially of:
 (a) trehalose at between 5-20%, w/v; 
 (b) xylitol at between 2-9%, w/v; 
 (c) sodium ascorbate 0.5-1.5%, w/v; and 
 (d) sodium phosphate at between 10-50 mM. 
   
     
     
         25 . The method of  claim 24 , wherein the aqueous preservation medium optionally comprises sodium glutamate at between 0-5%. 
     
     
         26 . The method of  claim 24 , wherein step ii includes daily administration of antibiotic for between 2 and 7 days and wherein step iii begins at any time between two days before the completion of antibiotic administration and two days after the administration of antibiotic in step ii ends. 
     
     
         27 . The method of  claim 24 , wherein the vaginal  Lactobacillus  species has the ability to produce greater than 0.5 ppm of hydrogen peroxide under effective culture conditions. 
     
     
         28 . The method of  claim 24 , wherein the vaginal  Lactobacillus  species is selected from the species consisting of  Lactobacillus crispatus, Lactobacillus jensenii  and  Lactobacillus gasseri.    
     
     
         29 . The dry powder of  claim 24 , wherein the dry powder has a water activity value of less than 0.220. 
     
     
         30 . The dry powder of  claim 29 , wherein the dry powder is combined with an inactive excipient at a ratio of powder: excipient of between 1:1 and 1:10 w/w. 
     
     
         31 . The dry powder of  claim 30 , wherein the excipient is maltodextrin. 
     
     
         32 . The dry powder of  claim 30 , wherein the ratio of powder:excipient is between 1:1 and 1:5 w/w 
     
     
         33 . The method of  claim 24 , wherein the cell pellet is combined with an aqueous preservation medium consisting essentially of:
 (a) trehalose at between 5-15%, w/v;   (b) xylitol at between 2-7%, w/v;   (c) sodium ascorbate 0.5-1.0%, w/v; and   (d) sodium phosphate at between 10-30 mM.   
     
     
         34 . The method of  claim 33 , wherein the aqueous preservation medium optionally comprises sodium glutamate at between 0-5%. 
     
     
         35 . The method of  claim 24 , wherein the antibiotic is clindamycin, metronidazole, or tinidazole.