Modified collagen

Abstract

The present invention relates to a modified collagen obtainable by providing isolated collagen; freezing the isolated collagen; dehydrating the frozen collagen; and maturing the dehydrated collagen. Also disclosed are methods of preparing the modified collagen and uses thereof.

1 .- 26 . (canceled) 
     
     
         27 . A composition comprising matured collagen, wherein the matured collagen comprises collagen stored at a temperature of at least about 40° C. for a period of at least one week or at a temperature of about 2-8° C. for about 1-3 years. 
     
     
         28 . The composition of  claim 27 , wherein the matured collagen comprises the collagen stored at a temperature of about 40° C. to about 65° C. for a period of at least one week. 
     
     
         29 . The composition of  claim 27 , wherein the matured collagen comprises the collagen stored at a temperature of at least 65° C. for a period of at least one week. 
     
     
         30 . The composition of  claim 27 , wherein the matured collagen comprises the collagen stored at a temperature of about 2-8° C. for about 1-3 years 
     
     
         31 . The composition of  claim 27 , further comprising gentamicin, bupivacaine or a pharmaceutically acceptable salt thereof. 
     
     
         32 . The composition of  claim 31 , wherein the composition comprises bupivacaine HCl. 
     
     
         33 . The composition of  claim 31 , wherein the composition comprises about 25% w/w to about 57% w/w bupivacaine HCl. 
     
     
         34 . The composition of  claim 31 , wherein the composition exhibits a release profile substantially corresponding to the following pattern:
 less than about 60% of the bupivacaine is released in about 45 minutes;   less than about 70% of the bupivacaine is released in about 60 minutes;   less than about 90% of the bupivacaine is released at about 120 minutes; and   when dissolution tested using a Dissolution Apparatus Type II in 500 mL of PBS buffer (pH 7.4, bath temperature 37° C.) and stirred at 50 rpm for 24 hours.   
     
     
         35 . The composition of  claim 27 , wherein a 0.9% dispersion of the composition has a viscosity of less than 500 cP at a temperature of 40° C. when tested for viscosity by a method comprising:
 preparing a 0.9% dispersion from the composition; and 
 measuring the viscosity of the dispersion using a Brookfield Digital Rheometer DV-III at a constant shear rate of 15 s −1  at a temperature of 40° C. 
 
     
     
         36 . The composition of  claim 27 , wherein the composition absorbs about 60 mg of water when tested for water uptake by:
 weighing a 1.5 cm×4 cm sample of the composition in a dry state to obtain a dry weight;   inserting the sample in water for injection for about 10 minutes;   weighing the sample after 10 minutes of insertion in water to obtain a wet weight; and   
       wherein the difference between the wet weight and dry weight is the amount of water absorbed. 
     
     
         37 . The composition of  claim 27 , wherein the composition swells to a thickness of less than 60 μm when tested for swelling by:
 measuring the thickness of a 1.5 cm×4 cm sample of the composition in dry state; 
 inserting the sample in water for injection for about 10 minutes; 
 measuring the thickness of the sample after 10 minutes of insertion in water to obtain thickness in a wet state; and 
 
       wherein swelling is the difference between the thickness in the wet state and dry state. 
     
     
         38 . The composition of  claim 27 , wherein the composition degrades in about 25 minutes when tested for degradation by a method comprising:
 (a) placing a 4.5 cm×4.5 cm sample of the composition in 15 mL of 0.2N phosphate buffer (pH 7.4 with CaCl 2 );   (b) adding 0.5 mL of 10 mg/mL Collagenase Type IA-S to the solution in step (a);   (c) agitating the solution of step (b) using a shaking water bath at 37° C. at 120 rpm for 60 minutes; and   (d) measuring the degradation of the sample every 5 minutes throughout step (c).   
     
     
         39 . The composition of  claim 27 , wherein about 50% of the composition is degraded in about 15 to 20 minutes upon when tested for degradation by a method comprising:
 (a) placing a 3.1 cm×3.1 cm sample of the composition in 15 mL of 0.2N phosphate buffer (pH 7.4 with CaCl 2 );   (b) adding 100 μl of 10 mg/mL Collagenase Type IA-S to the solution in step (a); and   (c) measuring the degradation of the sample after 5, 10, 15, 25, 40, 60, and 90 minutes.   
     
     
         40 . A composition comprising matured collagen prepared by a method comprising the steps of:
 (a) providing isolated collagen;   (b) freezing the isolated collagen;   (c) dehydrating the frozen collagen; and   (d) maturing the dehydrated collagen, wherein the maturing step comprises storing the dehydrated collagen at a temperature of at least 40° C. for at least one week or at a temperature of about 2-8° C. for about 1-3 years.   
     
     
         41 . The composition of  claim 40 , wherein step (a) comprises providing an isolated collagen dispersion having a concentration of about 3-30% (w/w) collagen particles. 
     
     
         42 . The composition of  claim 40 , wherein step (d) comprises storing the dehydrated collagen at a temperature of about 40° C. to about 65° C. for a period of at least one week. 
     
     
         43 . The composition of  claim 40 , wherein the maturing step comprises storing the dehydrated collagen at a temperature of at least 65° C. for a period of at least one week. 
     
     
         44 . The composition of  claim 40 , wherein the matured collagen comprises the collagen stored at a temperature of about 2-8° C. for about 1-3 years 
     
     
         45 . The composition of  claim 40 , further comprising gentamicin, bupivacaine or a pharmaceutically acceptable salt thereof. 
     
     
         46 . The composition of  claim 45 , wherein the composition comprises bupivacaine HCl. 
     
     
         47 . The composition of  claim 46 , wherein the composition comprises about 25% w/w to about 57% w/w of bupivacaine HCl. 
     
     
         48 . The drug delivery composition of  claim 45 , wherein the composition exhibits a release profile substantially corresponding to the following pattern:
 less than about 60% of the bupivacaine is released in about 40 minutes;   less than about 70% of the bupivacaine is released in about 60 minutes;   less than about 90% of the bupivacaine is released at about 120 minutes; and   when dissolution tested using a Dissolution Apparatus Type II in 500 mL of PBS buffer (pH 7.4, bath temperature 37° C.) and stirred at 50 rpm for 24 hours.   
     
     
         49 . The composition of  claim 40 , further comprising:
 (e) preparing an aqueous dispersion of the matured collagen of step (d);   (f) degrading the aqueous dispersion; and   (g) dehydrating the aqueous dispersion.   
     
     
         50 . The composition of  claim 49 , wherein the dehydrated collagen of step (c) is milled before step (d). 
     
     
         51 . The composition of  claim 49 , wherein step (d) comprises storing the dehydrated collagen at a temperature of at least 40° C. for a period of at least four weeks; or comprises storing the dehydrated collagen at a temperature of at least 65° C. for a period of at least one week. 
     
     
         52 . The composition of  claim 49 , wherein a 0.9% dispersion of the composition has a viscosity of less than 500 cP at a temperature of 40° C. when tested for viscosity by a method comprising:
 preparing a 0.9% dispersion from the composition; and 
 measuring the viscosity of the dispersion using a Brookfield Digital Rheometer DV-III at a constant shear rate of 15 s −1  at a temperature of 40° C. 
 
     
     
         53 . The composition of  claim 49 , wherein the composition absorbs about 60 mg of water when tested for water uptake by:
 weighing a 1.5 cm×4 cm sample of the composition in a dry state to obtain a dry weight;   inserting the sample in water for injection for about 10 minutes;   weighing the sample after 10 minutes of insertion in water to obtain a wet weight; and   
       wherein the difference between the wet weight and dry weight is the amount of water absorbed. 
     
     
         54 . The composition of  claim 49 , wherein the composition swells to a thickness of less than 60 μm when tested for swelling by:
 measuring the thickness of a 1.5 cm×4 cm sample of the composition in dry state; 
 inserting the sample in water for injection for about 10 minutes; 
 measuring the thickness of the sample after 10 minutes of insertion in water to obtain thickness in a wet state; and 
 
       wherein swelling is the difference between the thickness in the wet state and dry state. 
     
     
         55 . The composition of  claim 49 , wherein the composition degrades in about 25 minutes when tested for degradation by a method comprising:
 (a) placing a 4.5 cm×4.5 cm sample of the composition in 15 mL of 0.2N phosphate buffer (pH 7.4 with CaCl 2 );   (b) adding 0.5 mL of 10 mg/mL Collagenase Type IA-S to the solution in step (a);   (c) agitating the solution of step (b) using a shaking water bath at 37° C. at 120 rpm for 60 minutes; and   (d) measuring the degradation of the sample every 5 minutes throughout step (c).   
     
     
         56 . The composition of  claim 49 , wherein about 50% of the composition is degraded in about 15 to 20 minutes upon when tested for degradation by a method comprising:
 (a) placing a 3.1 cm×3.1 cm sample of the composition in 15 mL of 0.2N phosphate buffer (pH 7.4 with CaCl 2 );   (b) adding 100 μl of 10 mg/mL Collagenase Type IA-S to the solution in step (a); and   (c) measuring the degradation of the sample after 5, 10, 15, 25, 40, 60, and 90 minutes.