FREEZE-DRIED POWDER CONTAINING 2-[(3-AMINOPROPYL)AMINO]ETHANETHIOL AND ITS USE FOR PREPARING A THERMOGEL

Abstract

A freeze-dried powder for preparing a thermogel including: from 1% to 35% of 2-[(3-aminopropyl)amino]ethanethiol or one of its pharmaceutically acceptable salt; from 40% to 85% of one or more poloxamer; and from 0.1% to 20% of one or more carbohydrate compound.

Claims

1. A freeze-dried powder for preparing a thermogel comprising: from 1% to 35% of 2-[(3-aminopropyl)amino]ethanethiol or one of its pharmaceutically acceptable salt; from 40% to 85% of one or more poloxamer; and from 0.1% to 20% of one or more carbohydrate compound.

2. A freeze-dried powder according to claim 1, wherein it contains from 5% to 30% of 2-[(3-aminopropyl)amino]ethanethiol.

3. A freeze-dried powder according to claim 1, wherein it contains from 45% to 80% of poloxamer.

4. A freeze-dried powder according to claim 3, wherein it contains from 60% to 75% of poloxamer.

5. A freeze-dried powder according to claim 1, wherein poloxamer is chosen as being Poloxamer 407 or Poloxamer 188.

6. A freeze-dried powder according to claim 1, wherein it contains a mixture of Poloxamer 407 and Poloxamer 188.

7. A freeze-dried powder according to claim 1, wherein it contains from 0.5% to 15% of a carbohydrate compound.

8. A freeze-dried powder according to claim 7, wherein it contains from 1% to 10% of carbohydrate compound.

9. A freeze-dried powder according to claim 1, wherein carbohydrate compound is chosen among sugars.

10. Freeze-dried powder according to claim 9, wherein carbohydrate compound is chosen among mannitol, lactose, sucrose, trehalose, sorbitol, glucose or raffinose.

11. Freeze-dried powder according to claim 10, wherein carbohydrate compound is chosen as being sorbitol or mannitol.

12. A freeze-dried powder according to claim 1, wherein it further contains one or more of the following ingredients: flavouring agent; and/or high-intensity sweetener.

13. A process for preparing a freeze-dried powder according to claim 1, the process comprising the following steps: hydrolysing amifostine into amifostine thiol under acidic conditions; preparing a solution containing the excipients by: dissolving and mixing firstly all excipients presented in the formulation, except the poloxamer(s); cooling the temperature before adding gradually the polaxamer(s) to get a homogenous excipient solution; mixing the obtained amifostine thiol solution with the obtained excipient solution; freeze-drying the obtained solution; and nitrogen inerting before sealing.

14. A process for preparing a thermogel comprising the reconstitution of the freeze-dried powder according to claim 1 with an aqueous solution.

15. A thermogel obtainable by a process according to claim 14.

16. A thermogel according to claim 15 for treating or protecting mucosal or cutaneous tissue from damage associated with anticancer therapy.

Description

EXAMPLE 1—FREEZE-DRIED COMPOSITION

1.1—Process for Preparing Freeze-Dried Composition

[0051] A solution of amifostine at 500 mg/ml is prepared in hydrochloric acid 4M and heated in a water-bath at 60° C. for 1 hour to obtain the amifostine thiol solution.

[0052] A solution containing the excipients is prepared by: [0053] dissolving and mixing firstly all excipients presented in the formulation, except the poloxamers, in a bottle containing water under magnetic agitation; [0054] cooling the temperature of the obtained solution by placing the bottle in ice and keeping it cold for next step; and [0055] adding poloxamers (Kolliphor) gradually, under magnetic agitation, until complete solubilisation.

[0056] The obtained solution of amifostine thiol and the excipient solution are mixed at required proportion to get the solution at target composition and concentration.

[0057] Solution is then distributed in 20 ml vials and is freeze-dried using the program reported in Table 1 below.

TABLE-US-00001 TABLE 1 Freeze-drying Program Product Ramp Temp. Pressure (° C./ Hold Step (° C.) (μbar) min) (H) Loading 5 / / / Freezing Segment 1 −5 / 0.5 1 Freezing Segment 2 −40 / 1 5 Primary drying −18 200 0.03 48 Secondary drying 20 50 NA 4

[0058] The obtained freeze-dried powder is then inerted under nitrogen manually during 30 seconds before sealing using rubber stopper and aluminium crimp.

1.2—Freeze-Dried Powders 1 and 2

[0059] Composition of freeze-dried powders 1 and 2 (LP1 and LP2) reported in Table 2 below have been prepared according to the process disclosed above (example 1.1).

TABLE-US-00002 TABLE 2 LP1 and LP2 Freeze-dried Freeze-dried powder 1—LP1 powder 2—LP2 Ingredient (% w/w) (% w/w) Amifostine thiol 15.8% 16.2% Kolliphor P407 45.5% 46.7% Kolliphor P188 17.8% 18.2% Sorbitol  5.1% — Mannitol —  2.6% Hydrochloric acid  4.4%  4.5% Phosphoric acid 11.5% 11.8%

[0060] The obtained white freeze-dried powders LP1 and LP2 have good appearance and well-formed cake.

EXAMPLE 2—PREPARATION OF THERMOGEL

[0061] To reconstitute freeze-dried powders LP1 and LP2 to thermogel, 5 ml of water are introduced to the sealed vial through the stopper using a needle and a syringe.

[0062] After waiting one minute to let water penetrating in the cake, the vial is then gently shake by hand for two minutes to get a clear thermogel.

EXAMPLE 3—STABILITY OF FREEZE-DRIED POWDERS

3.1—Reference

[0063] A freeze-dried powder which does not contain carbohydrate compound and which has been prepared according to process disclosed in example 1.1 is used as a reference (i.e. Ref.) for this evaluation. Composition of said reference freeze-dried powder is reported in the Table 3 below.

TABLE-US-00003 TABLE 3 Reference Reference freeze-dried Ingredient powder—Ref. (% w/w) Amifostine thiol 16.6% Kolliphor P407 47.9% Kolliphor P188 18.7% Hydrochloric acid  4.6% Phosphoric acid 12.1%

3.2—Experimental Protocol

[0064] Vials of LP1, LP2 and Ref. were stored under the following conditions: [0065] 5° C./Ambient RH (i.e. relative humidity); [0066] 25° C./60% RH; and [0067] 40° C./75% RH; [0068] in stability chambers.

[0069] Stability of LP1, LP2 and Ref. under these conditions was assessed after 1-, 3- and 6-month storage.

[0070] Stability testing was performed according to ICH guideline “Q1A(R2) Stability Testing of New Drug Substances and Products”. The following parameters have been assessed: [0071] cake appearance by visual inspection; [0072] solution appearance by visual inspection; [0073] pH using pH-meter; [0074] purity; and impurity by HPLC.

3.3—Results

[0075] Results obtained for LP1, LP2 and Ref. are reported in Tables 4 to 6 below.

TABLE-US-00004 TABLE 4 LP1 stability 1-month 3-months 6-months 25° C./ 40° C./ 25° C./ 40° C./ 25° C./ 40° C./ Storage 65% 75% 65% 75% 65% 75% conditions T.sub.0 5° C. RH RH 5° C. RH RH 5° C. RH RH Cake White White White White White White White White White White appearance cake cake cake cake cake cake cake cake cake cake Solution Clear Clear Clear Clear Clear Clear Clear Clear Clear Clear appearance pH 4.5 4.5 4.5 4.5 4.4 4.5 4.4 4.4 4.5 4.5 Purity (area % 99.3 99.5 99.1 99.2 99.6 99.2 99.2 99.6 99.5 99.2 at 210 nm) Impurity (area % at 210 nm) Amifostine 0.05 0.06 0.10 0.070 0.06 0.10 0.05 0.08 0.08 <0.05 Disulfide 0.36 0.13 0.25 .28 0.18 0.29 0.35 0.16 0.16 0.21

TABLE-US-00005 TABLE 5 LP2 stability 1-month 3-months 6-months 25° C./ 40° C./ 25° C./ 40° C./ 25° C./ 40° C./ Storage 65% 75% 65% 75% 65% 75% conditions T.sub.0 5° C. RH RH 5° C. RH RH 5° C. RH RH Cake White White White White White White White White White White appearance cake cake cake cake cake cake cake cake cake cake Solution Clear Clear Clear Clear Clear Clear Clear Clear Clear Clear appearance pH 4.6 4.5 4.5 4.5 4.4 4.4 4.4 4.4 4.4 4.5 Purity (area % 99.6 99.5 99.4 99.1 99.4 99.3 98.9 99.6 99.4 98.8 at 210 nm) Impurity (area % at 210 nm) Amifostine <0.05 0.06 0.10 0.12 0.07 0.09 0.08 0.08 0.11 0.07 Disulfide 0.13 0.13 0.15 0.18 0.23 0.23 0.41 0.14 0.11 0.18

TABLE-US-00006 TABLE 6 Ref. stability 1-month 3-months 6-months 25° C./ 40° C./ 25° C./ 40° C./ 25° C./ 40° C./ Storage 65% 75% 65% 75% 65% 75% conditions T.sub.0 5° C. RH RH 5° C. RH RH 5° C. RH RH Cake White White White White White White White White White White appearance cake cake cake cake cake cake cake cake cake cake Solution Clear Clear Clear Clear Clear Clear Clear Clear Clear Clear appearance pH 4.6 4.7 4.6 4.6 4.7 4.6 4.6 4.6 4.6 4.7 Purity (area% 99.6 99.5 99.1 99.0 99.6 99.2 98.6 98.9 99.2 95.4 at 210 nm) Impurity (area % at 210 nm) Amifostine <0.05 0.05 0.13 0.12 0.07 0.10 0.11 0.09 0.13 <0.05 Disulfide 0.16 0.15 0.25 0.24 0.17 0.39 0.50 0.23 0.21 2.65

3.4—Conclusion

[0076] Reference formulation Ref. (which does not contain carbohydrate compound) is instable when it is stored at accelerated condition 40° C./75% RH. The key degradation impurity disulfide (RRT 1.25) was particularly increased (>2.6%) after 6-months storage at 40° C./75% RH.

[0077] On the contrary, stability of LP1 and LP2 is excellent, even after 6 months storage at accelerated condition (40° C./75% RH). Purity determination by HPLC has showed low level of the key impurities, amifostine and its disulfide.

[0078] Furthermore, it can be noted that addition of carbohydrate compound, in particular mannitol (see LP2) in freeze-dried composition is beneficial to reconstitution. The reconstitution of the product containing mannitol is fast and no significant change is observed after 6-months storage even at 40° C./75% RH.