Stable lyophilized pharmaceutical composition of sovateltide

Abstract

The present invention discloses a lyophilized pharmaceutical composition of Sovateltide for parenteral administration. More specifically, the invention discloses a composition comprising Sovateltide stabilized with lactobionic acid 10 and polyvinyl pyrrolidone (Kollidon PF 12), as well as a process for its preparation, which offers improved stability and reduced impurities.

Description

[0001] “STABLE LYOPHILIZED PHARMACEUTICAL COMPOSITION OF SOVATELTIDE”

[0002] FIELD OF THE INVENTION

[0003] The present invention relates to a lyophilized pharmaceutical composition of Sovateltide for parenteral administration. More specifically, the invention discloses a composition comprising Sovateltide stabilized with lactobionic acid and polyvinyl pyrrolidone (Kollidon PF 12), as well as a process for its preparation, which offers improved stability and reduced impurities.

[0004] BACKGROUND OF THE INVENTION

[0005] The Endothelin family consists of Endothelins (ETs) and their receptors. Endothelins are potent vasoconstrictor peptides that play a critical role in regulating systemic and peripheral vascular systems. The first endothelin peptide was discovered and isolated from the culture supernatant of porcine endothelial cells by Yanagisawa et al. in 1988. Among the endothelins, Endothelin-1 (ET-1), a 21 -amino-acid peptide with two disulfide bonds derived from endothelial cells, exhibits the most potent and enduring vasoconstrictor activity among known peptides. Additionally, two other isopeptides, Endothelin-2 (ET-2) and Endothelin-3 (ET-3), have been subsequently discovered. These three peptides of the endothelin family mediate various biological responses in both cardiovascular and non-cardiovascular systems by binding to two different receptor subtypes: ETA (selective for ET-1 and ET-2) and ETB (non-selective).

[0006] Sovateltide, also known as IRL-1620, is a synthetic analog of Endothelin-1 (ET-1). Unlike ET-1, which acts as an agonist for both endothelin A (ETAR) and endothelin B (ETBR) receptors, Sovateltide selectively agonizes ETBR receptors. Structurally, Sovateltide is a 15-amino-acid peptide, also referred to as PMZ-1620 or SPI-1620, with the chemical name succinyl-(Glu(9), Ala(11,15))-endothelin-1(8-21). Its sequence is represented as Suc-Asp-Glu-Glu-Ala-Val-Tyr-Phe-Ala-His-Leu-Asp-Ile-Ile-Trp.

[0007]

[0008] Sovateltide (Tycamzzi™) is being developed as a neural progenitor cell therapeutic agent for various medical conditions, including acute cerebral ischemic stroke (ACIS), hypoxic-ischemic encephalopathy (HIE), spinal cord injuries, and Alzheimer's disease. In May 2023, Sovateltide was approved in India as a 30pg intravenous injection for the treatment of cerebral ischemic stroke within 24 hours of stroke onset.

[0009] WO2024057296 disclosed lyophilized pharmaceutical composition of Sovateltide with mannitol as freeze drying agent and di sodium citrate dehydrate as buffering agent wherein the composition has a pH of about 7.0 to 8.5, an osmolarity between 240 to 310mOsm / L when reconstituted and total impurity not more than 2% or any unspecified impurity not more than 0.5% or D-His-sovateltide impurity not more than 1%.

[0010] The currently available formulation of Sovateltide is a lyophilized vial containing mannitol and disodium citrate dihydrate. This product requires storage at 2°C to 8°C. However, it demonstrates significant instability under accelerated storage conditions (40°C, 75% relative humidity), as applicable for Zone IV-B of the Indian market. Specifically, the impurity profile reveals a total impurity rise of approximately 6.51% within one month under these conditions. Furthermore, once reconstituted and diluted for infusion, the solution must be used immediately, highlighting limitations in its stability.Given these challenges, there exists a critical need for a stable pharmaceutical composition of Sovateltide. Such a composition would not only address its instability under accelerated storage conditions but also provide a longer shelf life and improved usability. A stable formulation would ensure enhanced therapeutic efficacy, better compliance with storage requirements, and increased accessibility in diverse geographical regions with varying climatic conditions.

[0011] Summary of the Invention

[0012] It is the object of the present invention to provide a stable Sovateltide composition for parenteral administration.

[0013] The other object of the invention is to provide a stable and pharmaceutically acceptable reconstituted solution of Sovateltide.

[0014] The invention provides a Sovateltide composition for parenteral administration comprising an active component, Sovateltide, and stabilizing agents.

[0015] In accordance with the above, the present invention provides a lyophilized pharmaceutical composition of Sovateltide for parenteral administration with improved stability and reduced impurities, using lactobionic acid and polyvinyl pyrrolidone as stabilizing agents.

[0016] In an aspect, the prseent invention provides a stable lyophilized pharmaceutical composition comprising;

[0017] i. Sovateltide

[0018] ii. lactobionic acid and

[0019] iii. polyvinyl pyrrolidone.

[0020] The lactobionic acid and polyvinyl pyrrolidone are used as stabilizing agents in the composition.

[0021] The lyophilized pharmaceutical composition of the present invention optionally comprises the pH adjuster.In an aspect, the pH of the composition in bulk solution and prior to lyophilization is adjusted at pH ranging between 7.0 to 8.5.

[0022] In an aspect, the present invention provides a stable lyophilized pharmaceutical composition comprising;

[0023] i. Sovateltide as active ingredient;

[0024] ii. lactobionic acid as stabilizing agent;

[0025] iii. polyvinyl pyrrolidone as stabilizing agent;

[0026] iv. pH adjuster

[0027] wherein pH of the bulk solution is adjusted to 7.0-8.5 prior to lyophilization; and wherein said composition contains NMT 0.5% D-Histidine Sovateltide impurity.

[0028] In another aspect, the Sovateltide as active ingredient is present in an amount of 30pg.

[0029] In another aspect, the stabilizing agents selected from lactobionic acid and polyvinyl pyrrolidone are present in an amount of about 40mg respectively.

[0030] In yet another aspect, the present invention provides a stable reconstituted liquid composition comprising Sovateltide as active ingredient and lactobionic acid and polyvinyl pyrrolidone as stabilizing agents and 0.9% sodium chloride solution. Sovateltide is present in the reconstituted liquid composition in the concentration of 0.0001% to 0.001% w / v, preferably 0.0006%w / v.

[0031] The lyophilized composition of the present invention or reconstituted liquid composition has D- Histidine Sovateltide impurity NMT 0.5%, and total impurity NMT 2%, more preferably NMT 1%

[0032] In yet another aspect, the present invention provides a process for increasing the stability and reducing the impurities of lyophilized pharmaceutical composition of Sovateltide with lactobionic acid and polyvinyl pyrrolidone as stabilizing agents and maintaining the reconstituted solution at a pH range of 7.0 to 8.5.DETAILED DESCRIPTION OF THE INVENTION

[0033] The invention will now be described in detail in connection with certain preferred and optional embodiments, so that various aspects thereof may be fully understood and appreciated.

[0034] The term ‘composition / formulation’ referred interchangeably in the entire description refers and means to the lyophilized pharmaceutical compositions of Sovateltide for parenteral administration.

[0035] The term "freeze-dried or lyophilized" powder, cake, or preparation, as used herein, refers to any solid substance produced through the lyophilization or freeze-drying of an aqueous solution. Preferably, a lyophilized preparation is obtained by freeze-drying a solution composed of aqueous solvents.

[0036] The term “stable lyophilized pharmaceutical composition” refers to a pharmaceutical formulation prepared by freeze-drying (lyophilization) that maintains its physical, chemical, and biological integrity during manufacturing, storage, and reconstitution. Stability in this context encompasses resistance to degradation, such as chemical decomposition, physical changes (e.g. aggregation, precipitation), or loss of potency, ensuring the composition retains its intended efficacy, safety, and shelflife under specified storage conditions.

[0037] Sovateltide a synthetic analog of Endothelin-1 (ET-1). Unlike ET-1 is a 15-amino-acid peptide, The D-His-sovateltide impurity (D-Histidine sovateltide) is a variant where the naturally occurring L-histidine amino acid at a specific position in the peptide chain is replaced by its stereoisomer, D-histidine. This impurity is crucial in pharmaceutical context. The present invention provides the lyophilized pharmaceutical composition containing Sovateltide that contains NMT 0.5% of said D-His-sovateltide impurity ensuring the stability and improved shelf life of the composition.The present invention relates to lyophilized pharmaceutical composition containing Sovateltide and to the method for preparing such composition.

[0038] In an embodiment, the present invention provides a stable lyophilized pharmaceutical composition comprising;

[0039] i. Sovateltide

[0040] ii. lactobionic acid; and

[0041] iii. polyvinyl pyrrolidone.

[0042] The lactobionic acid and polyvinyl pyrrolidone are used as stabilizing agents in the composition.

[0043] The lyophilized pharmaceutical composition of the present invention optionally comprises the pH adjuster.

[0044] In an aspect, the pH of the composition in bulk solution and prior to lyophilization is adjusted at pH ranging between 7.0 to 8.5.

[0045] In an embodiment, the present invention relates to a stable lyophilized pharmaceutical composition for parenteral administration comprising

[0046] i. Sovateltide as active ingredient;

[0047] ii. lactobionic acid as stabilizing agent;

[0048] iii. polyvinyl pyrrolidone as stabilizing agent;

[0049] iv. pH adjusters

[0050] wherein pH is adjusted between 7.0 to 8.5 in bulk solution before lyophilisation; and

[0051] wherein said composition contains NMT 0.5% D- Histidine Sovateltide impurity.

[0052] In an embodiment, the present invention discloses a stable lyophilized pharmaceutical composition for parenteral administration comprising;

[0053] i. Sovateltide in an amount of 30μg;

[0054] ii. lactobionic acid in an amount of 40mg;

[0055] iii. polyvinyl pyrrolidone in an amount of 40mg;

[0056] iv. pH adjuster q,s;

[0057] wherein pH of bulk solution is adjusted to 7.0-8.5 before lyophilization; andwherein said composition contains NMT 0.5% D- Histidine Sovateltide impurity.

[0058] In an embodiment, said composition is a lyophilized powder cake.

[0059] In another embodiment, the present invention relates to a reconstituted liquid composition for parenteral administration comprising;:

[0060] i. Sovateltide, in a concentration ranging from 0.0001% to 0.001% w / v; ii. Stabilizing agent, lactobionic acid in an amount ranging from 0.5% to 1.0% w / v);

[0061] iii. Stabilizing agent, polyvinyl pyrrolidone, in an amount ranging from 0.5% to 1.0% w / ); and

[0062] iv. 0.9%sodium chloride solution;

[0063] v. pH adjuster;

[0064] wherein the pH of said reconstituted composition ranges between 7.0 to 8.5; and wherein said composition contains NMT 0.5% D- Histidine Sovateltide impurity.

[0065] In an embodiment, Sovateltide is present in the reconstituted solution preferably in the concentration of 0.0006% w / v which is equivalent to 30 μg per vial.

[0066] In an embodiment, the stabilizer lactobionic acid is present in the reconstituted solution preferably in the concentration of 0.8% w / v.

[0067] In another embodiment, the stabilizer polyvinyl pyrrolidone is present in the reconstituted solution preferably in the concentration of 0.8% w / v

[0068] The pH may be adjusted, for example with the use of 1.0 N hydrochloric acid or 1.0 N sodium hydroxide solution.

[0069] In an embodiment, the inclusion of lactobionic acid and polyvinyl pyrrolidone as stabilizers addresses the stability challenges associated with Sovateltide underaccelerated storage conditions. The formulation ensures reduced impurity levels, improved shelflife, and enhanced stability.

[0070] The lyophilized composition of the present invention or reconstituted liquid composition has D- Histidine Sovateltide impurity NMT 0.5%, and total impurity NMT 2%, more preferably NMT 1%

[0071] In yet another embodiment, the present invention discloses a process for the preparation of stable lyophilized pharmaceutical composition of Sovateltide comprising the following steps;

[0072] i. Preparing an aqueous solution of Lactobionic acid and polyvinyl pyrrolidone (Kollidon PF 12) in water for injection;

[0073] ii. Adjusting the pH of the solution of step (i) between 8.5-9.0 with IN Sodium hydroxide solution;

[0074] iii. Cooling the solution of step (ii) between 2-8 °C and maintaining at said temperature;

[0075] iv. Dissolving suitable quantity of Sovateltide active ingredient in to the solution of step (iii);

[0076] v. Adjusting pH of the solution between 7.0-8.5 with 1 N Sodium hydroxide solution and making up the volume with water for injection;

[0077] vi. Filtering the solution of step (v) and filling in vials followed by freeze drying to obtain the lyophilized product.

[0078] In yet another embodiment, the present invention discloses the process of freeze drying comprising:

[0079] i. freezing the solution filled in vials at a temperature below -45°C and maintaining said temperature for at least 3 hours; ii. primary drying the frozen solution of step (a) under vacuum from 50 mtorr to 300 mtorr and at a temperature between -30°C and 15°C, and maintaining said conditions for at least 50 hours; andiii. secondary drying the primary dried frozen solution of step (b) under vacuum from 10 mtorr to 50 mtorr and at a temperature between 15°C and 40°C, and maintaining said conditions for at least 8 hours.

[0080] In yet another embodiment, the lyophilized formulation of Sovateltide of the present invention is subjected to accelerated degradation studies at 40°C with relative humidity of 75% moisture for six months and compared with the Tycamzzi™ composition, wherein present lyophilized pharmaceutical composition of Sovateltide exhibits improved stability and reduced impurities over Tycamzzi™.

[0081] The freeze dried Sovateltide when reconstituted with 5 ml of sodium chloride 0.9% w / v contains final drug concentrate of 6 mcg / ml.

[0082] In an embodiment, the Osmolarity of lyophilized pharmaceutical composition after reconstitution with 5 ml 0.9% w / v sodium chloride was found to be between 250 mOsmol to 350 mOsmol.

[0083] In an embodiment, the present invention relates to a method of treating neurological disorder or disease comprising administering the lyophilized pharmaceutical composition to a patient in need thereof.

[0084] Other features and embodiments of the invention will become apparent by the following examples which are given for illustration of the invention rather than limiting its intended scope. Various changes and modifications to the disclosed embodiments will be apparent to those skilled in the art.

[0085] Example 1 (a): Freeze dried composition of Sovateltide

[0086] Sr No Ingredients

[0087] 1. Sovateltide 30 mcg

[0088] 2. Lactobionic acid 40 mg

[0089]

[0090] 3. Polyvinyl pyrollidone (Kollidon PF 12) 40 mg

[0091] 4. 1 N Sodium Hydroxide solution q.s

[0092] 5. Water for Injection I. P 3.0 ml

[0093]

[0094] Example 1 (b): Process for the manufacture of a stable freeze-dried pharmaceutical composition.

[0095] a) Preparing an aqueous solution of Lactobionic acid and polyvinyl pyrrolidone (Kollidon PF 12) in water for injection;

[0096] b) adjusting the pH of the Solution prepared in step (a) between 8.5 to 9.0 with 1 N Sodium hydroxide solution and cooling the solution between 2°C to 8°C and maintaining the temperature throughout the manufacturing; c) Dissolving Sovateltide API in the Solution obtained in step (b);

[0097] d) adjusting the pH of the Solution prepared in step (c) between 7.0 to 8.5 with 1 N Sodium hydroxide solution and making up the volume to batch size with water for injection;

[0098] e) filtering the solution obtained in step (d) and filling the vials;

[0099] f) freezing the Solution filled in vials obtained in step (e); and

[0100] g) freeze drying the frozen solution filled in vials obtained in step (f). Example 1 (C): Stability

[0101] Stability Data for Sovateltide for Injection (30 mcg / vial)

[0102] The stability of present composition of Sovateltide for Injection (30 mcg / vial) were compared to the innovator's formulation. Stability studies were conducted under different storage conditions, and stress testing was performed to assess the robustness of the formulation. The results are summarized below:

[0103] Long-term Stability Study

[0104] Present invention Formulation:

[0105] Tests Initial 2-8°C 25°C

[0106] 3 Months 6 Months 3 Months 6 Months Assay 101.70% 101.56% 101.61% 101.00% 101.14%

[0107]

[0108] D-Histidine 0.44% 0.43% 0.46% 0.42% 0.49% Sovateltide

[0109] Single max Nil Nil Nil Nil Nil Impurity

[0110] Total impurities 0.44% 0.43% 0.46% 0.42% 0.49%

[0111]

[0112] Example 2a: Stability of Tycamzzi™ marketed Formulation:

[0113] Tests Initial 2-8°C 25°C

[0114] 3 Months 6 Months 3 Months 6 Months Assay 99.70% 98.26% 97.92% 97.41% 97.15% D-Histidine 0.22% 0.45% 0.48% 0.87% 0.96% Sovateltide

[0115] Single max 0.23% 0.28% 0.89% 0.45% 0.94% Impurity

[0116] Total impurities 0.70% 0.82% 1.55% 1.67% 2.15%

[0117]

[0118] Example 2b: Stress Stability Study at 40°C

[0119] Summary of Stability at Elevated Temperature

[0120] Tests Present invention Formulation Tycamzzi marketed Formulation

[0121] Initial 40 °C for 1 Initial 40 °C for 1 Month Month Assay 101.70% 99.54% 100.19% 95.33% Total 0.44% 1.93% 0.93% 6.51% impurities

[0122]

[0123] Conclusion: The present invention demonstrates superior stability compared to the Tycamzzi™ marketed Formulation under both refrigerated (2-8°C) and accelerated (25°C and 40°C) conditions.o Assay values for Present invention remain within 101.00% to 101.70% over 6 months, indicating minimal degradation.

[0124] o Impurity levels are consistently lower, with no single impurity exceeding detectable limits.

[0125] Tycamzzi™ marketed Formulation exhibits significant degradation at higher temperatures, with assay values dropping to 95.33% and total impurities rising to 6.51% after 1 month at 40°C.

[0126] The present composition is robust and well-suited for long-term storage and handling, even under stress conditions, making it a significant improvement over the Tycamzzi™ marketed Formulation.

[0127] Example 3a: Composition for scale up batch (Batch size: 8666 vials)

[0128] Sr No Ingredients Qty / Vial

[0129] 1. Sovateltide 30 mcg

[0130] 2. Lactobionic acid 40 mg

[0131] 3. Polyvinyl pyrollidone (Kollidon PF 12) 40 mg

[0132] 4. 1 N Sodium Hydroxide solution q.s

[0133] 5. Water for Injection I. P 3.0 ml

[0134]

[0135] Example 3 (b): Process for the manufacture of a stable freeze-dried pharmaceutical composition for scale up batch.

[0136] a) Preparing an aqueous solution of Lactobionic acid and polyvinyl pyrrolidone (Kollidon PF 12) in water for injection;

[0137] b) adjusting the pH of the Solution prepared in step (a) to between 8.5 to 9.0 with 1 N Sodium hydroxide solution and cooling the solution between temperature of 2°C to 8°C and maintaining the temperature throughout the manufacturing;

[0138] c) Dissolving Sovateltide API in the Solution obtained in step (b);d) adjusting the pH of the Solution prepared in step (c) to between 7.0 to 8.5 with 1 N Sodium hydroxide solution and making up the volume to batch size with water for injection,

[0139] e) filtering the solution obtained in step (d) and filling the vials;

[0140] f) freezing the solution filled in vials obtained in step (e) and

[0141] g) freeze drying the frozen solution filled in vials obtained in step (f).

[0142] Example 3 (C): Stability for scale up batch (Batch size: 8666 vials) Stability Data for Sovateltide for Injection (30 mcg / vial)

[0143] The novel formulation of Sovateltide for Injection (30 mcg / vial) exhibiting improved stability characteristics were compared to the innovator's formulation. Stability studies were conducted under different storage conditions, and stress testing was performed to assess the robustness of the formulation. The results are summarized below:

[0144] Stability Study at different storage conditions:

[0145] Present invention Formulation:

[0146] 3M_2- 3M_2 3M_4 6M_2 6M_25 6M_40 Tests Initial

[0147] 8°C 5°C 0°C -8°C °C °C 1 Descriptio white white white white white white white n lyophili lyophili lyophi lyophi lyophi lyophili lyophili zed zed lized lized lized zed zed cake cake cake cake cake cake cake 2 Related

[0148] substances

[0149] D- Histidine 0.32 0.35 0.31 0.32

[0150] 0.34% 0.34 % 0.35 % Sovateltid % % % %

[0151] e

[0152] Unspecific Nil Nil Nil Nil 0.19% 0.23% 0.29%

[0153]

[0154] d Impurity

[0155] Total 0.32 0.35 0.60 0.71

[0156] 0.34% 0.34 % 0.85 % Impurities % % % %

[0157] 3 Assay 100.43 100.3 100.3 100.3 100.5

[0158] 100.97% 99.26%

[0159] % 9% 6% 8% 5%

[0160]

[0161] The stability study substantiates that the lyophilized composition of the present invention ensures better shelf life and quality compliance under varying storage conditions.

[0162] Example 4: Osmolarity data

[0163] Tycamzzi™ marketed GUFIC’s Formulation Formulation

[0164] Reconstitution 1 vial + 5 ml 0.9% sodium 1 vial + 5 ml 0.9% procedure with diluent chloride Injection sodium chloride Injection (as per PIL of

[0165] Tycamzzi)

[0166] Results +543mOsmol +323mOsmol

[0167]

Claims

We Claim;1. A stable lyophilized pharmaceutical composition for parenteral administration comprising;i. Sovateltide;ii. lactobionic acid;iii. polyvinyl pyrrolidone.

2. The stable composition as claimed in claim 1, wherein lactobionic acid and polyvinyl pyrrolidone are used as stabilizing agents.

3. The stable composition as claimed in claim 1, wherein said composition optionally comprises the pH adjuster.

4. The stable composition as claimed in claim 1, wherein said composition is a lyophilized powder cake.

5. The stable composition as claimed in claim 1, wherein said lyophilized composition or its reconstituted liquid solution contains NMT 0.5% D- Histidine Sovateltide impurity.

6. The stable composition as claimed in claim 1, wherein said lyophilized composition or its reconstituted liquid solution contains NMT 2.0% of the total impurities.

7. A stable lyophilized pharmaceutical composition for parenteral administration comprising;i. Sovateltide as active ingredient;ii. lactobionic acid as stabilizing agent;iii. polyvinyl pyrrolidone as stabilizing agent;iv. pH adjuster;wherein pH of bulk solution is adjusted to 7.0-8.5 before lyophilization; and wherein said composition contains NMT 0.5% D-Histidine Sovateltide impurity and total impurities NMT 2.0%.

8. The stable composition as claimed in claim 7, wherein said composition comprises;i. Sovateltide in an amount of 30μg;ii. lactobionic acid in an amount of 40mg;iii. polyvinyl pyrrolidone in an amount of 40mg;iv. pH adjusterwherein pH of bulk solution is adjusted to 7.0-8.5 before lyophilization; and wherein said composition contains NMT 0.5% D- Histidine Sovateltide impurity and total impurities NMT 2.0%.

9. A reconstituted liquid composition for parenteral administration comprising;i. Sovateltide in a concentration ranging from 0.0001% to 0.001% w / v; ii. Stabilizing agent, lactobionic acid in an amount ranging from 0.5% to 1.0% w / v;iii. Stabilizing agent, polyvinyl pyrrolidone in an amount ranging from 0.5% to 1.0% w / ; andiv. 0.9% sodium chloride solution;wherein pH of the reconstituted liquid composition ranges between 7.0- 8.5; andwherein said reconstituted composition contains NMT 0.5% D- Histidine Sovateltide impurity and total impurities NMT 2.0%.

10. The reconstituted liquid composition as claimed in claim 9, wherein Sovateltide is present in the concentration of 0.0006%w / v equivalent to 30 μg per vial.

11. The reconstituted liquid composition as claimed in claim 9, wherein said reconstituted composition has osmolarity ranging between 250-350 mOsmol.

12. A process for preparation of stable lyophilized pharmaceutical composition of Sovateltide comprising;i. Preparing an aqueous solution of Lactobionic acid and polyvinyl pyrrolidone (Kollidon PF 12) in water for injection;ii. Adjusting the pH of the solution of step (i) between 8.5-9.0 with IN Sodium hydroxide solution;iii. Cooling the solution of step (ii) between 2-8 °C and maintaining at said temperature;iv. Dissolving suitable quantity of Sovateltide in to the solution of step (iii);v. Adjusting pH of the solution between 7.0-8.5 with IN Sodium hydroxide solution and making up the volume with water for injection;vi. Filtering the solution of step (v) and filling in vials followed by freeze drying to obtain the lyophilized product.

13. The process as claimed in claim 12, wherein the process of freeze drying comprising:i. freezing the solution filled in vials at a temperature below -45°C and maintaining said temperature for at least 3 hours;ii. primary drying the frozen solution of step (a) under vacuum from 50 mtorr to 300 mtorr and at a temperature between -30°C and 15°C, and maintaining said conditions for at least 50 hours; andiii. secondary drying the primary dried frozen Solution solution of step (b) under vacuum from 10 mtorr to 50 mtorr and at a temperature between 15°C and 40°C, and maintaining said conditions for at least 8 hours.

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