COLLAGENASE FORMULATIONS AND METHODS OF PRODUCING THE SAME
Disclosed herein are improved collagenase-containing formulations and methods of preparing the same. The collagenase-containing formulations comprise a collagenase, about 30 mM to about 240 mM of a disaccharide, about 50 mM to about 800 mM of mannitol, and about 6 mM to about 10 mM of a Tris-HCl. Lyophilized and reconstituted formulations are also provided.
1 . A formulation comprising:
a collagenase;
about 30 mM to about 240 mM of a disaccharide;
about 50 mM to about 800 mM of mannitol; and
about 6 mM to about 10 mM of a Tris-HCl.
2 . The formulation of claim 1 , wherein the collagenase comprises a collagenase I.
3 . The formulation of claim 2 , wherein the collagenase I comprises the amino acid sequence of SEQ ID NO: 1.
4 . The formulation of claim 1 , wherein the collagenase comprises a collagenase II.
5 . The formulation of claim 4 , wherein the collagenase II comprises the amino acid sequence of SEQ ID NO: 2.
6 . The formulation of claim 1 , wherein the collagenase comprises a mixture of collagenase I and collagenase II.
7 . The formulation of claim 6 , wherein the collagenase I comprises the amino acid sequence of SEQ ID NO: 1 and the collagenase II comprises the amino acid sequence of SEQ ID NO: 2.
8 . The formulation of claim 7 , wherein the collagenase is collagenase Clostridium histolyticum (CCH).
9 . The formulation of claim 1 , wherein the disaccharide comprises sucrose or trehalose.
10 . The formulation of claim 1 , wherein the pH of the formulation is about 7.8 to about 8.8.
11 . The formulation of claim 1 , wherein the formulation comprises:
CCH;
about 60 mM sucrose;
about 225 mM mannitol; and
about 10 mM Tris-HCl,
wherein the formulation has a pH of about 8.5.
12 . The formulation of claim 1 , further comprising a surfactant comprising polysorbate 20, polysorbate 80, or poloxamer 188.
13 . The formulation of claim 12 , comprising from about 0.01% to about 2% of the surfactant.
14 . The formulation of claim 13 , comprising about 0.02% of the surfactant.
15 . The formulation of claim 1 , wherein the formulation is liquid.
16 . A lyophilized formulation comprising:
a collagenase;
a disaccharide;
mannitol; and
Tris-HCl.
17 . The lyophilized formulation of claim 16 , wherein the collagenase comprises a collagenase I.
18 . The lyophilized formulation of claim 17 , wherein the collagenase I comprises the amino acid sequence of SEQ ID NO: 1.
19 . The lyophilized formulation of claim 16 , wherein the collagenase comprises a collagenase II.
20 . The lyophilized formulation of claim 19 , wherein the collagenase II comprises the amino acid sequence of SEQ ID NO: 2.
21 . The lyophilized formulation of claim 16 , wherein the collagenase comprises a mixture of collagenase I and collagenase II.
22 . The lyophilized formulation of claim 21 , wherein the collagenase I comprises the amino acid sequence of SEQ ID NO: 1 and the collagenase II comprises the amino acid sequence of SEQ ID NO: 2.
23 . The lyophilized formulation of claim 21 or 22 , wherein the collagenase is collagenase Clostridium histolyticum (CCH).
24 . The lyophilized formulation of claim 16 , wherein the disaccharide comprises sucrose or trehalose.
25 . The lyophilized formulation of claim 16 , wherein, prior to lyophilization, the formulation comprises:
CCH;
60 mM sucrose;
225 mM mannitol; and
10 mM Tris-HCl, and
having a pH of about 8.5.
26 . The lyophilized formulation of claim 16 , wherein the lyophilized formulation is stable at pressures above 380 μbar.
27 . The lyophilized formulation of claim 26 , wherein the lyophilized formulation is stable at a pressure of about 4000 μbar.
28 . The lyophilized formulation of claim 16 , wherein the lyophilized formulation is stable at:
(a) 2-8° C. for at least 36 months;
(b) 25° C./60% relative humidity for at least 36 months;
(c) 40° C./75% relative humidity for at least 6 months; or
(d) any combination of (a) to (c).
29 . The lyophilized formulation of claim 16 , wherein the lyophilized formulation is formed by a method comprising:
freezing the formulation at a temperature between about −25° C. and −55° C. to form a frozen formulation; and
drying the frozen formulation at a temperature between about 25° C. and about 50° C. to form the lyophilized formulation.
30 . The lyophilized formulation of claim 29 , wherein the lyophilized formulation is formed by a method comprising:
freezing the formulation at a single temperature between about −25° C. and −55° C. to form a frozen formulation; and
drying the frozen formulation at a single temperature between about 25° C. and about 50° C. to form the lyophilized formulation.
31 . The lyophilized formulation of claim 16 , wherein the lyophilized formulation is formed by a lyophilization method that is performed for less than 72 hours.
32 . The lyophilized formulation of claim 16 , wherein the lyophilized formulation is formed by a lyophilization method that is performed at a pressure of between about 380 μbar to about 4000 μbar.
33 . The lyophilized formulation of claim 16 , in a unit-dose vial, multi-dose vial, cartridge, or syringe.
34 . A reconstituted formulation comprising:
a collagenase;
a disaccharide;
mannitol;
Tris-HCl;
calcium chloride; and
sodium chloride.
35 . The reconstituted formulation of claim 34 , wherein the collagenase is collagenase Clostridium histolyticum (CCH).
36 . The reconstituted formulation of claim 35 , wherein the disaccharide comprises sucrose or trehalose.
37 . The reconstituted formulation of claim 34 , wherein the reconstituted formulation is isotonic to human blood.
38 . A kit comprising:
a container comprising the lyophilized formulation of claim 16 ; and
a container comprising a sterile diluent comprising calcium chloride and sodium chloride.