PARENTERAL FORMULATIONS OF MELPHALAN
Background of the Invention
Melphalan, also known as L-phenylalanine mustard, phenylalanine mustard, L-PAM, or L-sarcolysin, is a phenylalanine derivative of nitrogen mustard. Melphalan is a bifunctional alkylating agent that is active against selected human neoplastic diseases. The molecular formula is C13H18CI2N2O2 and the molecular weight is 305.20. The structural formula is:
^ NH2
(ClCH2CHa)3N—(T ))— ChE— -C- - - COOH
^ ' H
U.S. Pat. No. 4,997,651 to Stephen et al., discloses a two-component pharmaceutical formulation of Melphalan comprising freeze-dried Melphalan hydrochloride and a solvent-diluent comprising of citrate, propylene glycol and ethanol.
U.S patent application No. 2013/0131174 to Jose et al., discloses a solid lyophilized composition of Melphalan hydrochloride.
U.S patent application 2014/0221488 to James et al., is directed to a method for conditioning a subject for whom a stem cell transplantation has been indicated, comprising administering a pharmaceutical composition comprising Melphalan and a cyclodextrin derivative.
The commercial injectable formulation of Alkeran® is supplied as a sterile, nonpyrogenic, freeze-dried powder. Each single-use vial contains Melphalan hydrochloride equivalent to 50 mg Melphalan and 20 mg povidone. Alkeran is reconstituted using the sterile diluent containing sodium citrate 0.2 g, propylene glycol 6.0 mL, ethanol (96%) 0.52 mL and water for Injection to a total of 10 mL. The Alkeran® to be infused must be diluted to not more than 0.45 mg/ml in normal saline and infused over 15 minutes.

The commercially available product of Melphalan contains propylene glycol in the formulation. Further the concentration of the infusion liquid should not be greater than 0.45 mg/mL. However the presence of propylene glycol in the formulation often presents acceptability limitations and also the concentration of the infusion liquid of 0.45 mg/mL may have limitations such as higher infusion volume.
The inventors of the present invention have developed a propylene glycol free parenteral formulation of Melphalan which also has the advantage of low administering volume. Further the invention formulation also exhibits improved stability compared to Alkeran.
Summary of the invention
One object of the invention provides a stable, parenteral formulation of Melphalan comprising of Melphalan and cyclodextrin or its derivative thereof, wherein the formulation is free of propylene glycol.
Another aspect of the invention is to provide a stable, lyophilized formulation of Melphalan which can be administered in low volume as compared to the marketed formulations.
Another aspect of the invention is to provide stable parenteral formulation of Melphalan comprising of Melphalan, cyclodextrin(s), one or more solvents, bulking agents, buffering agents and optionally other pharmaceutically acceptable excipients thereof.
Yet another aspect of the invention provides concentration of Melphalan in the infusion liquid.
Yet another aspect of the invention provides weight ratio of cyclodextrin to Melphalan in the formulation.

Detailed description of the Invention
In the context of this invention "Melphalan" refers to the pharmaceutically acceptable salts, solvates, hydrates and anhydrous forms thereof, preferably Melphalan Hydrochloride.
The term "infusion liquid" as used herein means that the drug solution is suitable for direct administration to the patient. The infusion liquid is obtained by reconstitution of the lyophilized powder with the diluent provided and which is further diluted with intravenous infusion fluids such as saline (0.9% NaCl), 5% dextrose solution, Ringer's solution, lactated Ringer's solution and the like.
Melphalan formulation developed according to the present invention provides smaller infusion volume with improved stability. Each vial of the invention formulation contains 50mg of Melphalan. The formulation has concentration in the range of 5mg/ml to 20mg/ml, when reconstituted with the diluent of invention, which when further diluted with normal saline has concentration in the range of 0.7 mg/mL to about 5 mg/mL. The recommended infusion volume of the invention formulation is around 7 ml to 50 ml as compared to 78.2 ml of the formulations known in the art, for palliative treatment of patients with multiple myeloma, considering body surface area (BSA) of 2.2m . A reduction in the infusion volume helps to reduce the time required for administration to the patient. This is particularly advantageous as it aids to reduce the pain in administration, reduce the fluid burden, ability to administer highest dose in small volume and further helps in saving the in-patient time in hospital setting. The concentration of the infusion liquid of the invention formulation ranges from about 0.7 mg/mL to about 5 mg/mL of Melphalan.
One embodiment of the present invention provides propylene glycol free parenteral formulations of Melphalan.

Another embodiment of the present invention provides the weight ratio of cyclodextrin to Melphalan in the formulation. The weight ratio of cyclodextrin to Melphalan is in the range of 10:1 to 40:1 (w/w).
The invention relates to stable parenteral formulation of Melphalan comprising:
i. Melphalan,
ii. Cyclodextrin,
iii. Bulking agent,
iv. One or more solvents, wherein the weight ratio of cyclodextrin to Melphalan ranges from about 10:1 to 40:1.
Yet, another embodiment of the invention relates to stable parenteral formulations of Melphalan comprising:
i. Melphalan Hydrochloride,
ii. Cyclodextrin,
iii. Bulking agent,
iv. Buffering agents and/or pH adjusting agents
v. One or more solvents selected from dimethylacetamide, ethanol and water,
wherein the weight ratio of cyclodextrin to Melphalan ranges from about 10:1 to 35:1.
Yet, another embodiment of the invention relates to stable lyophilized parenteral formulations of Melphalan having low infusion volume, compared to the marketed formulations of Melphalan.
Suitable cyclodextrins include but not limited to a, P and y-cyclodextrin and
cyclodextrins modified with alkyl-, hydroxyalkyl-, dialkyl-, and sulfoalkyl-ether
modified cyclodextrins such as methyl or hydroxypropyl P-cyclodextrins (HPpCD),
methyl-and-ethyl-P-cyclodextrin, sulfoalkylether-substituted beta-cyclodextrin,
sulfobutylether-P-cyclodextrin (SBECD) and the like. Preferably sulfobutylether-P-cyclodextrin (SBECD) is used.

Suitable solvents include, but not limited to dimethylacetamide (DMA), dimethyl sulfoxide (DMSO), N-methylpyrrolidone, dimethylisosorbide, ethanol, glycerine, polyethylene alcohol, polyethylene glycols and water. Preferred solvents are ethanol, dimethylacetamide and water.
The bulking agents can be selected from mannitol, glucose, sucrose, maltose, xylitol, starches, sorbitol, lactose, povidone, maltitol, trehalose, glycine, dextrose, dextran, raffinose, polyvinylpyrrolidone (PVP) and the like.
Suitable buffering agents include, but not limited to citrate buffer, glutamate buffer, lactate buffer, carbonate buffer, bicarbonate buffer, gluconate buffer, TRIS buffer, acetate buffer, borate buffer, phosphate buffer, amino acids such as arginine, alanine, histidine, glycine, lysine and corresponding salts and mixtures thereof. Preferred buffering agents are amino acids.
Suitable pH adjusting agents include the following, but are not limited to sodium hydroxide, sodium carbonate, sodium bicarbonate, potassium hydroxide, ammonium carbonate, hydrochloric acid, citric acid, lactic acid, phosphoric acid and the like. Preferred pH adjusting agents are sodium hydroxide and hydrochloric acid.
The pharmaceutical compositions of the present invention may also contain anti-oxidants, stabilizers, cryostabilizers, surfactants, tonicity modifiers, preservatives or any other suitable excipients thereof.
The lyophilized parenteral formulation of Melphalan is reconstituted with the diluent. The diluent may comprise of one or more agents selected from cyclodextrins, solvents, buffering agents, tonicity modifiers, pH adjusting agents, electrolytes, or any other suitable excipient thereof.
Comparative dilution studies were performed to check the stability of the diluted formulations. Melphalan formulation prepared according to the invention was diluted with the proposed diluent and further diluted with 0.9% Normal saline (NS) to get

concentration of 1.5 mg/mL and 3mg/mL. Stability of the diluted product was studied at 0 minutes, lhr and 6hrs. Alkeran® vial (Batch No: P283) was considered as reference for comparative dilution study. Reference product was initially diluted with Alkeran diluent and further diluted with 0.9% normal saline (NS) so as to obtain a concentration of 0.45mg/ml (as prescribed in the package insert of Alkeran). Stability of the diluted product was studied at 0 minutes and lhr. Comparative dilution stability data of the invention formulation with the reference product is summarized in table 1.
The stability of invention formulation exhibited improved stability when compared to reference product. Impurity D (Mono hydroxy impurity) and Impurity G (Dimer impurity) were found to be high in reference product compared to invention formulation. Further total impurities observed in the invention formulation at 1.5 mg/mL and 3mg/mL are significantly less compared to reference product. The infusion volume can also be reduced with the invention formulation as the concentration of the diluted solution (infusion liquid) is 1.5 mg/mL and 3mg/mL.
The infusion volume of the invention formulation required for administration is compared with the infusion volume of the commercially available formulations. Comparison of infusion volume is shown in table 2.

A preferred embodiment of the invention comprises
i. Melphalan Hydrochloride
ii. Bulking agent
iii. Cyclodextrin
iv. Solvents selected from ethanol and water, and other pharmaceutically
acceptable excipients, wherein the concentration of the infusion liquid ranges from about 0.7 mg/mL to about 5 mg/mL of Melphalan.
The following examples further describe certain specific aspects and embodiments of the present invention and demonstrate the practice and advantages thereof. It is to be understood that the examples are given by way of illustration only and are not intended to limit the scope of the invention in any manner.

Manufacturing process:
(i) Hydrochloric acid was added to the vessel containing water for injection and cooled
to 1°C. (ii) Melphalan Hydrochloride was added to the above solution. (iii)Povidone was added to another vessel containing water for injection and stirred and
the solution was cooled to 1°C. (iv)Povidone solution was added to the solution of step (i) (v) The solution was filtered and filled in vials and freeze dried.
Diluent vial: SBECD was added to water for injection and stirred. Histidine was added to the above solution followed by ethanol and stirred. The obtained solution was filtered and filled in vials.

Manufacturing process:
(i) Hydrochloric acid was added to the vessel containing water for injection and cooled
to 1°C. (ii) Melphalan Hydrochloride was added to the above solution. (iii)Povidone was added to another vessel containing water for injection and stirred and
the solution was cooled to 1°C. (iv)Povidone solution was added to the solution of step (i) (v) The solution was filtered and filled in vials and freeze dried.
Diluent vial: SBECD was added to water for Injection and stirred. Histidine was added to the above solution followed by ethanol and stirred. The obtained solution was filtered and filled in vials.

Manufacturing process:
(i) Hydrochloric acid was added to the vessel containing water for injection and cooled
to 1°C. (ii) Melphalan Hydrochloride was added to the above solution. (iii)Povidone was added to another vessel containing water for injection and stirred and
the solution was cooled to 1°C. (iv)Povidone solution was added to the solution of step (i) (v) The solution was filtered and filled in vials and freeze dried.
Diluent vial: SBECD was added to water for Injection and stirred. L-Arginine was added to the above solution followed by ethanol and stirred. The obtained solution was filtered and filled in vials.
Manufacturing process:
(i) Hydrochloric acid was added to the vessel containing water for injection and cooled
to -1±2°C. (ii) Melphalan Hydrochloride was added to the solution of step (i).

(iii)Povidone was added to another vessel containing water for injection and stirred and
the solution was cooled to 2-8°C. (iv)Povidone solution was added to the solution of step (i). (v) SBECD was added followed by the addition of arginine to the solution and stirred
till a clear solution was obtained. (vi)The solution was filtered and filled in vials and freeze dried.

We claim
Claim 1: A stable parenteral formulation of Melphalan comprising
i. Melphalan or a pharmaceutical^ acceptable salt thereof ii. Cyclodextrin and iii. Solvents,
wherein the concentration of the infusion liquid ranges from about 0.7 mg/mL to about
5 mg/mL.
Claim 2: The formulation of claim 1, further comprising one or more excipients selected from bulking agents, buffering agents and pH adjusting agents.
Claim 3: A stable propylene glycol free, parenteral formulation of Melphalan comprising:
i. Melphalan or a pharmaceutically acceptable salt thereof
ii. Cyclodextrin
iii. Bulking agent
iv. Buffering agent and/or pH adjusting agent and
v. One or more solvents, wherein the weight ratio of cyclodextrin to Melphalan ranges from about 10:1 to 40:1.
Claim 4: The formulation according to claims 1 and 3, wherein cyclodextrin is selected from a, P and y-cyclodextrin and cyclodextrins modified with alkyl-, hydroxyalkyl-, dialkyl-, and sulfoalkyl-ether modified cyclodextrins such as methyl or hydroxypropyl P-cyclodextrins, methyl-and-ethyl-P-cyclodextrin, sulfoalkyl ether-substituted beta-cyclodextrin and sulfobutylether-P-cyclodextrin.
Claim 5: The formulation according to claims 1 and 3, wherein the solvent is selected from dimethylacetamide, dimethyl sulfoxide, N-methylpyrrolidone, ethanol, glycerine, polyethylene glycols and water.

Claim 6: The formulation according to claims 1 and 3, wherein the bulking agent is selected from mannitol, glucose, sucrose, maltose, xylitol, starches, sorbitol, lactose, povidone, maltitol, trehalose, glycine, dextrose, dextran, raffinose and polyvinylpyrrolidone.
Claim 7: The formulation according to claims 1 and 3, wherein the buffering agent is selected from citrate buffer, glutamate buffer, lactate buffer, carbonate buffer, bicarbonate buffer, gluconate buffer, TRIS buffer, acetate buffer, borate buffer, phosphate buffer and amino acids including arginine, alanine, histidine, glycine and lysine.
Claim 8: The formulation according to claims 1 and 3, wherein the pH adjusting agent is selected from sodium hydroxide, sodium carbonate, sodium bicarbonate, potassium hydroxide, hydrochloric acid, citric acid, lactic acid and phosphoric acid.