OA20747A

OA20747A - A stable parenteral dosage form of cetrorelix acetate.

Info

Publication number: OA20747A
Application number: OA1202200163
Authority: OA
Inventors: Rajamannar Thennati; Jaydip JOSHI; Rakesh Thummar; Sudeep AGRAWAL; Subhas Balaram Bhowmick; Arunkumar YADAV
Original assignee: Sun Pharmaceutical Industries Limited
Priority date: 2019-10-24
Filing date: 2020-10-23
Publication date: 2023-02-24

Abstract

The present invention relates to a stable parenteral dosage form with a ready-to-inject sterile stable aqueous solution of cetrorelix acetate. The invention also relates to an injection device prefilled with the ready-to-inject sterile stable aqueous solution of cetrorelix acetate. The present invention relates a method of inhibiting premature luteinizing hormone surges in women undergoing controlled ovarian stimulation comprising a stable parenteral dosage form with a ready-to-inject sterile stable aqueous solution of cetrorelix acetate.

Description

A STABLE PARENTERAL DOSAGE FORM OF CETRORELIX ACETATE

Field of the Invention

The present invention relates to a stable parentéral dosage form with a ready-toinject, stérile, stable, aqueous solution of cetrorelix acetate. The invention also relates to an injection device prefilled with the ready-to-inject, stérile, stable, aqueous solution of cetrorelix acetate. The present invention relates to a method of inhibitîng prématuré luteinîzing hormone surges in women undergoing controlled ovarîan stimulation comprising a stable parentéral dosage form with a ready-to-inject, stérile, stable, aqueous solution of cetrorelix acetate.

Background of the Invention

Cetrorelix is gonadotropin releasing hormone antagonist (GnRH antagonist) acetyl-D-3- (2'-naphÎyl)-alanine-D-4-chlorophenylalanine-D-3-(3'-pyridyÎ)-alanine-Lserine-L-tyrosine-D-citruline-L-leucine-L-arginine-L-proline-D-alanine-amide (C70H92CIN17O14) having the below formula. It is a decapeptide with a terminal acid amide group. It acts by blocking the action of GnRH upon the pituitary, thus rapidly suppressing the production and action of leutinizing hormone and follicle stimulating hormone.

Aqueous solutions of peptides are required for parentéral administration. However, aqueous solutions of peptides such as cetrorelix are susceptible to Chemical dégradation. They are also prone to aggregatîon whereby the turbidity or cioudiness of the solution încreases on storage.

The first product on the market was Cetrotide®. It is avai labié as a lyophilized powder în glass vials containing 0.25 mg or 3 mg of cetrorelix. A prefilled glass syringe having 1 ml or 3 ml of stérile water for injection is provided separately and the solution is prepared only prior to injection. Therefore, the first product solved the probiem of dégradation in aqueous solution simply by avoiding preparing a dosage form containing an aqueous solution that needed to be stored over time. Instead the water was removed and a lyophilizate was prepared to avoid înstabilîty problems. However, this solution to the probiem has clear disadvantages - (1) expensive and time consuming process; (2) product is not ready-to-inject and requires reconstitution before administration; and (3) reconstituted solution is stable only for a short period of time. Cetrotîde® thus did not fulfil a need for a ready-to-inject aqueous solution.

US 7,718,599 discloses that aqueous solutions of cetrorelix were prone to aggregatîon. Under a polarized light microscope, liquïd crystalline structures were observed. To cetrorelix acetate solutions (2.5 mg/ml), gluconic acid was added, whereby at concentrations of gluconic acid less than 0.07%, resulting in a pH of 3.7, aggregatîon was seen within 2 days. Similar failure was reported when the pH was more than 3.7. When the concentration of gluconic acid was increased to 0.71%, resulting in a pH of 3.1, the aggregatîon was seen in 12 days indicating that higher concentrations of gluconic acid and thus lower pH led to improvement. The disadvantage of the method îs that the degree of resolution of the probiem of aggregatîon is dépendent on the gluconic acid concentration and with more gluconic acid the pH decreases. However, US 7,718,599 did not report the effect of pH on the Chemical stabilîty of cetrorelix. Moreover, there were no formulations where aggregatîon was not seen during long term storage stability studîes. US 2013/0303464 discloses a ready-to-use aqueous préparation of cetrorelix comprising cetrorelix acetate, glacial acetic acid, a tonicîty adjusting agent and water for injection. A suitable pH was illustrated by working examples where the pH was about 3. The preferred pH according to the invention was pH 2.8 to 3.5.

US 7,214,662 discloses aqueous solutions of peptides including cetrorelix acetate and suggested solutions to the probiem of aggregatîon. It taught that carboxylîc acids and especially hydroxycarboxylic acids, preferably gluconic acid, in combination with a surfactant reduces aggregatîon. The use of carboxylîc acid according to US 7,214,662 resulted in a low pH such as pH 2.5 to 3.

Description of the Invention

The object of the présent invention is to provide a parentéral dosage form comprising a ready-to-inject stérile stable aqueous solution of cetrorelix acetate. Another object of the invention is to provide an injection device pre-filied with the stérile stable aqueous solution of cetrorelix acetate. The terni “ready-to-inject” as used herein refers to a ready-to-inject, stérile, stable, aqueous solution of cetrorelix acetate which is suitable for direct subcutaneous or intramuscular administration, i.e., it is ready-to-inject and there is no requirement of reconstitution or dilution before injection. More particularly, it is another objective that the stérile stable aqueous solution of cetrorelix acetate dispensed in an injection device be ready-to-inject, not only be physically stable in terms of control on aggregation or turbidity development but also be chemically stable such that impurities remain low while the parentéral dosage fonn is stored on the shelf and until it is injected into the patient subcutaneously or întra-muscularly.

Dégradation of peptides can lead to génération of other peptides and/or peptide derîvatives which may themselves hâve pharmacological activity. Therefore the objective more particularly was to develop an appropriate method to separate individual impurities and quantify them. The objective was to limit the concentration of such impurities. The inventors discovered a High Performance Liquîd Chromatographie (“HPLC”) method which gave separate peaks for several impurities which were here before not reported in the prior art. Whereas the prior art advocated low pH values to decrease the tendency for agglomération, the inventors found with the use of theîr HPLC method that in the parentéral dosage form of the présent invention, a pH of 3 to 5 was optimal for Chemical stability in terms of încreases in level of impurities over a periodof tîme and also the aqueous solution of cetrorelix acetate couid be prepared at this higher pH without agglomération problems.

A novel impurity discovered by the inventors was Impurity A represented by the compound of Formula I gîven below:

Formula I

Impurity B is characterized to hâve a structure represented by the compound of

Formula H given below:

Formula II.

Impurity D is characterized to hâve a structure represented by the compound of

Formula III given below:

Impurity F is characterized to hâve a structure represented by the compound of Formula IV given below:

Formula IV.

The prior art considered low pH of 3.0 to be the optimum pH for stability; however, the présent invention found that at pH values of 2.5 to 3.0 advocated by the prior art, the ievel of Impurity A increases significantiy upon storage ofthe solution at 25°C/60% relative 15 humidity.

None of the prior art identifîed the compounds of formula I, II, III and IV, i.e. Impurities A, B, D and F respectively.

The présent invention found that not only could the stable aqueous solution of cetrorelix acetate be prepared at a pH 3 to 5 without agglomération problems but also the level of Impurity A and total impurities were well controlled and remain at low concentrations upon storage of the parentéral dosage form at 25°C/60%RH for at least I month, at least 2 months, at least 3 months, or at least 6 months. The parentéral dosage form could also be stored at 2 to 8°C with good stabilîty for at least 24 months.

In one aspect, the présent invention provides a parentéral dosage form comprising a stable aqueous solution comprising:

(i) cetrorelix or a pharmaceutically acceptable sait thereof; and (ii) an impurity of Formula I in an amount less than 5% w/v of cetrorelix base.

(Formula 1).

Preferably, the parentéral dosage form comprises impurity of Formula I in an amount less than 4% w/v of cetrorelix base. More preferably, the parentéral dosage form comprises impurity of Formula I in an amount less than 3% w/v of cetrorelix base. More preferably, the parentéral dosage form comprises impurity of Formula 1 in an amount less than 2% w/v of cetrorelix base. More preferably, the parentéral dosage form comprises impurity of Formula 1 in an amount less than 1 % w/v of cetrorelix base.

The parentéral dosage form further comprises an osmotic agent and water for injection.

In a préférable aspect, the présent invention provides a parentéral dosage form comprising a stable aqueous solution comprising;

(1) cetrorelix or a pharmaceutically acceptable sait thereof; and (ii) an impurity of Formula I in an amount less than 1% w/v of cetrorelix base,

Formula 1.

In another aspect, the présent invention provides a parentéral dosage form comprising a stable, aqueous solution comprising:

(i) cetrorelix or a pharmaceutically acceptable sait thereof; and (ii) an impurity of Formula I în an amount less than 1% w/v of cetrorelix base,

Formula L

In another aspect, the présent invention provides a parentéral dosage form comprising a ready-to-inject, stérile, stable, aqueous solution comprising:

(i) cetrorelix or a pharmaceutically acceptable sait thereof, (ii) an organic acid to adjust the pH in the range of 3 to 5, (iii) Impurity A, a decapeptide of formula I, in an amount less than 1 % w/v of cetrorelix base

(iv) an osmotic agent; and (v) water for injection.

In one embodiment, the invention provides a parentéral dosage form comprising a ready-to-înject stérile, stable, aqueous solution consisting of:

(i) cetrorelix or a pharmaceutically acceptable sait thereof, (ii) an organic acid to adjust the pH in the range of 3 to 5, (iii) Impurity A, a decapeptide of formula I, in an amount less than 1% w/v of cetrorelix base

Formula I, (iv) an osmotic agent, and (v) water for injection.

The parentéral dosage form comprising the ready-to-înject stérile, stable aqueous solution of cetrorelix according to the présent invention remains physically and chemically stable when stored at 2 to 8°C for at least 1 month, at least 3 months, at least 6 months, at least 12 months, at least 18 months, or at least 24 months; or at room température (25°C/60%R.H) for at least 1 month, at least 3 months, or at least 6 months.

Preferred embodîments of the stable parentéral dosage form can be labelled with a shelf life at 2 to 8°C of at least 24 months or of 24 months. More preferred embodiments of the parentéral dosage form can be labelled with a shelf life of at least 6 months or of 6 months at room température (25°C/60%RH) storage condition.

The concentration of decapeptides of formula I (Impurity A) remains in the range of 0.001% to 1.0%, preferably 0.05 to 0.5 % by weight of cetrorelix base, single maximum unknown impurity remains less than 0.5% by weight of cetrorelix base and total impurity remains not more than 3.5 % by weight of cetrorelix base upon storage at 2 to 8°C for at least 1 month, at least 2 months, at least 3 months, at least 6 months, at least 12 months, at least 18 months or at least 24 months and/or at room température (25°C/60%RH) for at least 1 month, at least 2 months, at least 3 months, or at least 6 months.

The parentéral dosage form comprising the ready-to-inject stérile aqueous solution of cetrorelix according to the present invention is physically stable with no aggregation, gel formation or précipitation of the aqueous solution during the shelf-life. The aggregation or gel formation can be determined by measuring the cloudiness or turbidity of the solution. It is measured in FTU unit (Formazin Turbidity Unit) or NTU unit (Nephelometric Turbidity Unit).

The test is performed according to the protocol described in European Pharmacopoeia 9.0. The solution is said to be free of any aggregation or gel formation if the cloudiness/turbidity value is less than or equal to 8 FTU/NTU. The higher the FTU/NTU values the higher the cloudiness or turbidity in the solution and vice-versa. The NTU values of the ready-to-inject, parentéral dosage form according to the present invention remains less than 2 NTU, preferably less than 1 NTU, more preferably less than 0.5 NTU, initial!y and upon long term storage of the dosage form at 2 to 8°C for at least 1 month, at least 2 months, at least 3 months, at least 6 months, at least 12 months, at least 18 months or at least 24 months and/or at room température (25°C/60%RH) for at least 6 months. Thus, there occurs no aggregation, gel formation or précipitation of the aqueous solution during the shelf-life. Also, there occurs no substantîal increase in vîscosïty of the solution upon storage.

The parentéral dosage form comprising the ready-to-inject, stérile, stable, aqueous solution of cetrorelix according to the present invention contains cetrorelix acetate at a concentration ranging from 0.26 mg/ml to 0.28 mg/ml, which amount is équivalent to 0.25 mg/ml of cetrorelix base. Preferably, cetrorelix acetate is present in the ready-to-inject stérile, stable aqueous solution at a concentration équivalent to 0.25 mg/ml of cetrorelix base.

In one embodiment, the parentéral dosage form comprising the ready-to-inject stérile, stable aqueous solution of cetrorelix according to the présent invention comprises a pH adjusting agent at a concentration sufficient to adjust the pH in the range of 3 to 6.

In a preferred embodiment, the parentéral dosage form comprising the ready-toinject, stérile, stable aqueous solution of cetrorelix according to the présent invention comprises an organic acid as a pH adjusting agent at a concentration sufficient to adjust the pH in the range of 3 to 5, more preferably in the range of 4 to 4.5. The pH ofthe ready-to-inject stérile, stable aqueous solution according to the présent invention may be for example, 3, 3.05, 3.10, 3.15, 3.20, 3.25, 3.30, 3.35, 3.40, 3.45, 3.5, 3.55, 3.60, 3.65, 3.70, 3.75, 3.80, 3.85, 3.90, 3.95, 4.00, 4.05, 4.10, 4.15, 4.20, 4.25, 4.30, 4.35, 4.40, 4,45, 4.50, 4.55, 4.60, 4.65, 4.70, 4.75, 4.80, 4.85, 4.90, 4.95, 5.00, 5.05, 5.10, 5.15, 5.20, 5.25, 5.30, 5.35, 5.40, 5.45, 5.50, 5.55 and 6 or intermediate ranges thereof.

The organic acid may be selected from any parenterally acceptable organic acid soluble in water but is preferably acetic acid, more preferably lactic acid. For example, lactic acid may be used in the ready-to-inject stérile aqueous solution according to the présent invention ata concentration ranging from about 0.013 mg/ml to 0.53 mg/ml, preferably in amount ranging from about 0.033 mg/ml to about 0.53mg/ml; and intermediate ranges thereof.

Preferably, according to the présent invention, the ready-to-inject stérile, stable aqueous solution of cetrorelix comprise cetrorelix (base) and organic acid in a weight ratio ranging from 5 0.47 ; 1 to 19.23; I, preferably in a weight ratio ranging from about 0.47:1 to 7.57:1, more preferably in a weight ratio ranging from about 1.56:1 to 7.57:1 and intermediate ranges thereof.

The parentéral dosage form comprising the ready-to-inject stérile, stable aqueous solution of cetrorelix according to the présent invention comprises an osmotic agent or tonicity adjusting agent, in amounts suitable to adjust the osmolality of the solution in the range of about 250-375 mOsm/kg, preferably 270-330 mOsm/kg. The osmotic agent that may be used in the aqueous solution according to présent invention is selected from, but not limited to, mannitol, glycerol, sorbitol, sodium chloride, potassium chloride, dextrose.

sucrose, and the like and mixtures thereof.

According to one preferred embodiment, the osmotic agent is mannitol and it may be used in the aqueous solution in an amount ranging from about 40.0 mg/ml to 60.0 mg/ml, preferably in an amount ranging from about 50,0 mg/ml to 58.0 mg/ml. In one preferred embodiment, the osmotic agent is mannitol and it is used in the ready-to-inject stérile aqueous solution in an amount of about 55.0 mg/ml.

The ready-to-inject, stérile, aqueous solution of the parentéral dosage form of the présent invention does not contain lactic acid in the form of its dérivatives, polymer or copolymers such as poly lactic acid or polylactic-co-glycolic acid. Preferably, lactic acid is used as a sole pH adjusting agent. In preferred embodiments, the ready-to-inject stérile, aqueous solution is free of any surfactant, such as tween 80, polysorbates, poloxamers, spans and the like. The ready-to-inject stérile, aqueous solution of the parentéral dosage form avoîds use of surfactants, complexing agents, préservât!ve oranti-oxidants for solubiîization or stabilization. In certain embodiments, the solution is free of complexing agents like cyclodextrins, free of co-solvents such as alcohols or glycols and is also free ofpreservatives and antioxidants.

In another aspect, the présent invention provides the stérile, aqueous solution of cetrorelix acetate as above which remains stable for at least 1 month, preferably for at least 3 months and more preferably for at least 6 months at 25°C température and 60 % relative humidity.

In yeî another aspect, the présent invention provides the stérile, aqueous solution of cetrorelix acetate as above which remains stable for at least 1 month, preferably for at least 3 months, more preferably for at least 6 months, even more preferably for at least 12 months or 18 months, and most preferably for at least 24 months at 2-8°C.

The stable parentéral dosage form comprising the ready-to-inject, stérile, stable aqueous solution of cetrorelix according to the présent invention is suitable for administration by subeutaneous route or intra-muscular route. The ready-to-inject, stérile, stable aqueous solution is suitable for direct subeutaneous administration, Le., it is readyto-inject or ready-to-self-administer and there is no requirement of reconstitution or dilution before use. The ready-to-inject, stérile, stable aqueous solution according to the présent invention does not involve lyophilization.

The stable parentéral dosage form of the présent invention is suitable for selfadministration and enables the patient to self-administer a small volume of the aqueous solution subcutaneously. The volume of the ready-to-inject stérile, aqueous solution of cetrorelix fi lied in the réservoir of the injection device ranges from about 0.5 ml to 10.0 ml, preferably 1.0 ml to 2.0 ml, more preferably 1.0 ml. According to one of the preferred embodiments, the ready-to-inject, stérile, stable, aqueous solution of cetrorelix is filled in the réservoir of the injection device in volume of 1.0 ml. Preferably the parentéral dosage form according to the présent invention is suitable for administerîng a single dose of cetrorelix acetate. In one embodiment, the parentéral dosage form comprises a fill volume of about l .0 ml of aqueous solution of cetrorelix acetate suitable for self-administration as a single dose. In some embodiment, the parentéral dosage form may comprise aqueous solution of cetrorelix at a fill volume of about 10.0 ml, suitable for multiple dose administration.

The injection device according to the stable, parentéral, dosage form of the présent invention may be selected from, but not limited to, prefïlled syringes, autoinjectors and the like. In one preferred embodiment, the injection device is a prefilied syringe. In another preferred embodiment, the injection device is an autoinjector such as a pen auto-injector. These pre-filled syringes or auto-înjectors are suitable for self-administration or autoinjection of the drug solution by the patients in need thereof, thus providing a user friendly approach.

in one preferred embodiment, the injection device is a prefïlled syringe. The prefïlled syringe comprises the following components: a réservoir such as, for example, a barrel or a cartridge, which stores the aqueous solution; a stalked needle attached at one end of the réservoir; a needle shieid or tip cap that covers the needle and seals the needle tip opening, optionally, a rigid shieid covering the needle shieid or tip cap; a plunger stopper at other end of the réservoir that stoppers and seals the aqueous solution filled in the réservoir; a plunger rod that fits into the plunger stopper and is used to push the plunger stopper along with the solution towards the needle end while administerîng the drug.

In another preferred embodiment, the injection device is an autoinjector. The autoinjector can hâve varied designs. In one preferred design, the autoinjector comprises the following components; a central assembly or body portion that is suitable to hold a pre

II fi lied syringe, the syringe comprising a réservoir such as a barrel or a cartridge which stores the aqueous solution, the réservoir having a stalked needle at one end and a plunger stopper at other end. The central body portion may hâve a clear inspection window through which the solution in the réservoir is visible. The autoinjector further comprises a front assembly having a cap portion that holds a needle shield or tip cap, and it is attachable to the central assembly covering the stalked needle and sealing the needle tip opening. The autoinjector further comprises a rear assembly which comprises a plastic rod with a spring assembly and an activation button. During self-administration of the aqueous solution, first, the cap along with needle shield is removed from the body portion exposing the needle and subsequently after placing the body portion of the autoinjector at the site of administration the activation button is pressed, which pushes the plastic rod with spring assembly towards the plunger stopper which ieads to delivery of the aqueous solution through the needle to the patient.

The réservoir may be a barrel or a cartridge, such as, for example, a barrel of a prefïlied syringe or a cartridge of an auto-injecter. Il may be made up of a material selected from glass, plastic or a polymeric material. In some preferred embodiments, the réservoir is made up of glass, such as USP Type I siliconized glass or non-pyogenic glass material. In other embodiments, the réservoir is made up of a non-glass plastic or polymeric material selected from cycloolefîn polymer, cycloolefin copolymer, polyolefins, styrenepolyolefin based polymers and block co-polymers, polycarbonates and the like. In one preferred embodiment, the réservoir is a non-pyogenic glass barrel of a pre-filied syringe or non-pyogenic glass cartridge of an auto- injector.

In one or more embodiments, the réservoir may hâve a stacked needle at one end. In some other embodiments, the réservoir is needleless and has a luer tîpped lock at one end with provision for attachîng a needle at the leur tip before use. The stalked needle may be made up of stainless Steel. The needle tip is shielded orcovered with a needle shield or tip cap. The réservoir containing the stérile aqueous solution of drug is further sealed with a stopper such as a plunger stopper at the other end. These stoppers, needle shields or tip caps provide a physical and steri 1 ity barrier against exterior environment.

Preferably, the plunger stopper, the needle shield /tip cap or the cap of leur lock is made up of a non-glass component. The non-glass component may be a rubber or elastomeric material such as for example, bromobutyl rubber, chlorobutyl rubber, USP type II rubber, natural rubber made up of poly-cis-l,4-isoprene, styrene butadiene rubber and the like. Other suitable materais include high density polyethylene or low density polyethylene or other plastic materîals. in preferred embodiments, the plunger stopper is made up of bromobutyl rubber and the needle shield or tip cap is made up of natural rubber. The needle shield may further be covered on an outer side by a rigid shield made up of polypropylene. It protects the needle shield from damage and enhances removal of needle shield before injection. The injection device assembly may hâve a plunger rod that attaches to the plunger stopper and is used to push the plunger stopper along with the solution towards the needle end while administering the drug.

Preferably, the ready-to-inject, stérile, stable aqueous solution of cetrorelix is filled in the réservoir of the injection device and stoppered in such a manner that there is substantially no headspace air left inside the réservoir. The aqueous solution in the réservoir always remains in contact with the plunger stopper made up of elastomeric or rubber material during storage. In the case of prefilied syringes having a stalked needle made up of stainless Steel, the needle being covered by a needle shield or tip cap, the aqueous solution remains in contact with the needle and the needle shield or tip cap during storage.

The injection device may optionally be packaged or enclosed in a secondary packaging. The secondary packaging may be a blister pack or an aluminum pouch and/or an opaque carton. A suitable oxygen scavenger may optionally be placed inside the secondary packaging.

The srability testing of the parentéral dosage form is donc by storing the dosage form at2-8°C and at room température (25°C/60 % relative humîdity). During stability testing, the ready-to-inject stérile solution of cetrorelix remains in contact with the plunger stopper and needle shield made up of elastomeric rubber material as well as with the stacked needle made up of stainless Steel. In preferred embodiments, the parentéral dosage fonn comprising the ready-to-inject stérile aqueous solution of cetrorelix according to the présent invention remains physîcally and chemically stable fora period of 1 year, preferably 2 years when stored at 2-8°C and at least for 6 months at room température (25°C, 60 % relative humîdity). The concentration of Impurity A remains less than 1.0 % by weîght of cetrorelix base upon storage of the filled injection device at room température (25°C/60% relative humîdity) for at least 6 months and at 2-8°C for at least 24 t

months. The extrapolated shelf iife of the aqueous solution of cetrorelix determined by Minitab computation for Impurity A considering levels of not more than 1%, is found to be 122 months.

In one aspect, the présent invention relates to a method of inhibiting prématuré luteinizing hormone surges in women undergoing controlled ovarîan stimulation comprising:

a parentéral dosage form comprising: a ready-to-inject stérile, stable aqueous solution comprising:

(i) cetrorelix or a pharmaceutically acceptable sait thereof; and (ii) an impurity of Formula 1 in an amount less than 5% w/v of cetrorelix base.

Formula I.

Preferably, the stable aqueous solution comprises impurity of Formula I in an amount less than 4% w/v of cetrorelix base. More preferably, the stable aqueous solution comprises impurity of Formula 1 in an amount less than 3% w/v of cetrorelix base. More preferably, the stable aqueous solution comprises impurity of Formula I in an amount less than 2% w/v of cetrorelix base. More preferably, the stable aqueous solution comprises impurity of Formula I in an amount less than 1% w/v of cetrorelix base.

The stable aqueous solution further comprises an osmotic agent and water for 20 injection.

(i) cetrorelix or a pharmaceutically acceptable sait thereof, (ii) Impurity A, a decapeptide of formula I in an amount less than 1% w/v of cetrorelix base,

In one aspect, the present invention relates to a method of inhibîtîng prématuré luteinizing hormone surges in women undergoing controlled ovarian stimulation comprising: a parentéral dosage form comprising: a ready-to-inject stérile, stable aqueous solution comprising:

(i) cetrorelix or a pharmaceutically acceptable sait thereof, (ii) Impurity A, a decapeptide of formula I, in an amount less than 1 % w/v of cetrorelix base.

Formula I.

In one préférable aspect, the present invention relates to a method of inhibiting prématuré luteinizing hormone surges in women undergoing controlled ovarian stimulation comprising: a parentéral dosage form comprising: a ready-to-inject, stérile, stable aqueous solution comprising:

(i) cetrorelix or a pharmaceutically acceptable sait thereof, (ii) an organic acid to adjust the pH in the range of 3 to 5, (iîi) Impurity A, a decapeptide of formula I, in an amount less than 1% w/v of cetrorelix base.

(iv) an osmotic agent, and (v) water for injection.

In another aspect, this disclosure provides a decapeptide of formula I

This compound is termed “Impurity A” herein, as it is an impurity of a cetrorelix solution.

This disclosure also provides a composition comprising a decapeptide of formula 1:

O..NH, HN^NFb

N H ⁰ fR ° f°H ⁰ f H ° L JJ N JH q i H Q f H Q » J H Τι ^N 1Ί T 0 U Mqh ¹ Formula I.

NH JvÇJ * °0^AnY^h H 0

In another aspect the disclosure provides a process to identify the decapeptîde of Formula 1 by HPLC analysis, the process comprising:

a) injectîng a diluent comprising water, acetonitrile and formic acid into the chromatographie System,

b) injecting a System suitability solution comprising cetrorelix acetate, diluent and impurity stock solution and recording the chromatogram,

c) injecting a standard solution comprising cetrorelix acetate and diluent into the chromatographie System,

d) injecting a sample comprising aqueous solution of cetrorelix acetate and placebo préparation into the chromatographie System, and

e) determining the relative rétention time and relative response factor of impurities and cetrorelix acetate with respect to cetrorelix acetate.

This disclosure also provides a decapeptîde of Formula I, identified by HPLC analysis, the process comprising:

1. injecting a diluent comprising water, acetonitrile and formic acid into the chromatographie System,

2. injecting a System suitability solution comprising cetrorelix acetate, diluent and impurity stock solution and recording the chromatogram,

3. injecting a standard solution comprising cetrorelix acetate and diluent into the chromatographie System,

4. injecting a sample comprising aqueous solution of cetrorelix acetate and placebo préparation into the chromatographie System, and

5. determining the relative rétention time and relative response factor of impurities and cetrorelix acetate with respect to cetrorelix acetate,

Hereinafter, the invention wiil be more specifically described by way of Examples. The examples are not intended to Iimit the scope of the invention and are merely used as illustrations.

EX AMPLE IA IDENTIFICATION OF THE DEGRADATION PRODUCT

In order to investigate the dégradation of cetrorelix, peptide related substances of cetrorelix were prepared by the known technique of solid phase peptide synthesis. The synthesis invoived coupling of one amino acid at a time sequentially starting from ctermînal amino acid on a resin. The synthesis of the peptide chain was carried out using the Fluorenylmethyloxycarboyl (Fmoc)ÆButyi (Fmoc/tBu) with N,N’-diisopropyl carbodiimide (DIPC) as the coupling reagent. The Fmoc groups were removed via treatment with 20% piperidine in dimethylformamide. The peptide formed on resin was finally cleaved using trifluoroacetic acid to obtain related substances which were further purified by reverse phase high performance liquid chromatography (RP-HPLC) on a Cl 8 Silica column using a gradient of acetonitrîle/water containîng 0.1% trifluoroacetic acid. The purified peptide related substances were îyophîlized to obtain pure solid form. The structure of these related substances were characterized by Proton NMR, Carbon NMR, Mass spectroscopy and elemental analysis and they were referred to as Impurity A, B, D and F.

Impurity-A: Ac-2-D-Nal-4-CLD-Phe-3-D-Pal-Ser-Tyr-D-Cit-Leu-Arg-Pro-D-Ala-OH (detailed structure depicted as the Compound of Formula I),

Impurity-B: 2-D-Nal-4-Cl-D-Phe-3-D-Pal-Ser-Tyr-D-Cit-leu-Arg-Pro-D-Ala-NH2 (detailed structure depicted as the Compound of Formula II),

Impurity-D: Ac-2-D-Nal-4-Cl-D-Phe-3-D-Pal-Ser-Tyr-D-Cit-Leu-OH (detailed structure depicted as the Compound of Formula HI), and

Impurity-F: Ac-2-D-Nal-4-Cl-D-Phe-3-D-Pal-Ser-Tyr-D-Cît-Leu-Arg-Pro-OH (detailed structure depicted as the Compound of Formula IV).

The dégradation peaks separated on the HPLC column, were identified to be these compounds based on their relative rétention time. The details of the HPLC method is provided in Example IB below;

EXAMPLE IB

Cetrorelix and the identified impurities namely, Impurity A, Impurity B, Impurity D and Impurity F from the aqueous solution sampies were separated on a reverse phase (C-18) column using gradient technique (Column: X-Select CHS Cl8, (150 x 4.6) mm, 2.5μ (by Waters, Ireland, Part No: 186006729), detected and quantîfied by Ultraviolet spectroscopy at 225 nm wavelength. The mobile phase was run at a flow rate of 0.7 ml/min and 1.0 ml/min. The run time of the chromatogram was 150 minutes.

Mobile phase details:

Mobile Phase A: A mixture of buffer solution as below, with acetonitrile and tetrahydrofuran in the ratio of (700:280:20), degassed by sonication.

Mobile Phase B: A mixture of buffer solution as below, with acetonitrile and tetrahydrofuran in the ratio of (500:480:20), degassed by sonication.

Buffer: 2.5 g of Ammonium dihydrogen orthophsphate and 0.75 g of 1-Octane sulphonic acid sodium sait in 1000 ml water with pH adjusted to 8.0 ± 0.05 using triethylamine.

Diluent: A mixture of water, acetonitrile and formic acid in the ratio of (700:300:1).

Table 1: Details of gradient elution

Time (minutes)	Flow Rate	Mobile Phase A (% v/v)	Mobile Phase B (% v/v)
0	0.7	100	0
65	0.7	100	0
75	0.7	0	100
76	1.0	0	100
135	1.0	0	100
136	0.7	100	0
150	0.7	100	0

Préparation of the stock solution of impurities:

3.125 mg each of Impurity A; Impurity B, Impurity D and Impurity F were taken in a 50 ml volumétrie flask and dissoived in about 5 ml of diluent by sonication, followed by making up the volume using the diluent.

Préparation of svstem suitabilitv solution:

This was prepared by weighing and transferring about 12.5 mg of cetrorelix acetate working standard in ! 00 ml volumétrie flask and dissolving it in about 50 ml of diluent by sonication. followed by addition of about 2 ml of impurity stock solution and making up the volume using the diluent.

Préparation. of thejsiaiidard^siÎliiiÎQn of cetrorelix acetate:

The srandard solution of cetrorelix acetate was prepared by weighing and transferring 20 mg of cetrorelix acetate working standard into 250 ml volumétrie flask and dissolving it in about 50 ml of diluent by sonication and making up the volume with the diluent. Two ml of this solution was transferred into 250 ml volumétrie flask and volume made up to the mark using the diluent with mixing.

Préparation of test solution:

The aqueous solution of cetrorelix acetate from about 10 pre-fïlled syringes of the sample to be tested (prepared according to example as described above) was rnixed in a comaîner. The solution comprises cetrorelix acetate, an organic acid, an osmotic agent and water for injection. Accurately about 5.0 ml of this solution was transferred in 10 ml volumétrie flask and about 3 ml of the diluent was added and the solution was sonicated for 5 minutes with intermediate shaking. Volume made up using the diluent with mîxîng.

The placebo was préparée! by transferring accuraiely about 5.0 ml of placebo solution in 10 ml volumétrie flask, adding about 3 ml diluent and sonicating for 5 minutes with intennediate shaking. Volume made up using the diluent with mixing. 50 microlitres injections in duplicate of diluent as blank were injected into the chromatographie System. Subsequently, the System suitability solution was injected and the chromatogram was recorded. The resolution between Impurity D and Impurity F is not less thau 2.0. Following this, six replicates of standard solution were injected. Subsequently, the sample and placebo préparation were injected into the chromatographie System.

The relative rétention time and relative response factor of cetrorelix acetate and Impurities A. B, D and F with respect to cetrorelix acetate are presented in Table 2.

Table 2;

Naine of i coin pou nd	Rétention Time (minute)	Relative rétention time
\| Cetrorelix	42.3	1.00
î Impurity A	23.5	0.55
' Impurity B	56.8	1.34
Impurity D	16.9	0.39
Impurity F	20.3	0.48

The percentage of Impurities A, B, D, F and unknown impurity was calculated excluding peaks from diluent and placebo. The sum of ail knowu and unknown impurities provided % total impurities.

The % of identified impurities (A. B, D, F) was calculated by following formula:

Al WS 2 10 P 1

---Λ----- JC------JC ---X -----X---AS 250 250 V LC RRF

Where,

Al = Peak response of each known impurity in the chromatogram of test préparation AS = Average peak response of cetrorelix in the chromatogram of standard préparation WS = Weight of cetrorelix acetate working standard in mg

V = Volume of sample taken in ml

P = % potency of cetrorelix working standard (on as is basis) LC = Label claim of cetrorelix in mg per ml (0.25mg/mI) RRF = Relative response factor of each Impurity

The % of Unknown impurity was calculated by following formula

Al WS 2 10 P ---X---X----X--X--AS 250 250 V LC

Where,

Al = Peak response of each unknown impurity in the chromatogram of test préparation AS = Average peak response of cetrorelix in the chromatogram of Standard préparation WS = Weight of cetrorelix acetate workîng standard in mg

V = Volume of sample taken in ml

P = % potency of cetrorelix workîng standard (on as is basis) LC = Label claim of 10 cetrorelix in mg per ml (0.25mg/ml)

The total impurities (%) = Sum of % known împurities and % unknown impurities.

Table 3: Composition

(N

SUN-074

Method of Préparation:

Water for injection was taken at température between 2°C to 8°C in a vessel. Mannîtol was added and dissolved gradually in water for injection with stirring, until a clear solution was obtained. To this cetrorelix acetate was added and dissolved gradually with stirring. The pH of the solution was checked and was adjusted to the pH as mentioned in Table 3 for each example of the invention and comparative examples, usîng specified amount (volume) of 0.1 % w/v lactic acid solution. The volume was made up with water for injection. The solutions were stîrred for 10-15 minutes. The solutions of the Examples were filtered aseptically through a bed of 0.2 micron membrane filter. The solution was aseptically fi lied in the réservoir of injection device, i.e., in the barrel of 1 ml glass syringe with a fil! volume of 1.1 ml. The stacked needle in the barre! was stoppered by elastomeric needie shield. covered by a rigid cap before fillîng. After filling, the glass syringe (barrel) was stoppered with plonger stopper by vacuum stoppering in such a manner that there was substantially no headspace air left inside the syringe. The aqueous solution remains in contact with the plunger stopper made up of rubber, stacked needle made up of staînless Steel and needle shield made up of natural rubber upon storage.

The ready-to-inject, aqueous solution of working examples 1 to 9 and comparative examples 10 to 14 were subjected to Chemical analysis at different stages. Initially, the % assay of cetrorelix in the solution before and after filtration was analyzed by the HPLC method described above. The change in the Chemical assay % before and after filtration was determined.

The solutions of the examples contained in the glass syringes were then subjected to storage stabi!ity testing. The % assay, the level of dégradation products like the compounds of formula I, II, III and IV and the level of unknown and total impurities in the filtered solution fi lied in injection device of the parentéral dosage form at initial time point and upon storage at different time points at room température (25°C/60 % relative humidity) and at 2 to 8°C were determined using the high performance liquid chromatographie method described above.

It was found that after 6 months of storage at room température the level of impurities A. B, single maximum unknown impurity and the total impurities remained unchanged or the change was small. Based on this data it is expected that the parentéral dosage form of the présent invention is chemically stable over a long period of time. It

S UN-074 was found that the solutions dîd not exhibit any probîems of agglomération or increase in vîscosity when prepared and when fîlled into the injection device and stored. The data also demonstrated that there was no absorption or adsorptîon of cetrorelix onto or into the components of the device, for instance, the rubber stopper which was in contact with the solution during the period of storage.

The stability results for the stable parentéral dosage form at 25°C/60% RH and 2-

8°C according to the présent invention are provided in Table 4 and Table 5 below:

Table 4

\o O1

Table 5

Total impurity (%)	Time points (monlhs)	24	Ϊ69Ό	.0.387	1 82ΖΌ ;	0.15
oo	OC s	0.31		Ε0ΕΌ
	0.551	0.337	0.242	S*
	££E0	o	0.174	0.136
	,0.161	6900 :	0.144	9ΠΌ
—	0.206	0.087	0.23	0 213
a	O	0.335	c>	O
	O	0.149	o	c>
ut timc points tipon slorage al (2 8^UC) Single masimum unlinowit impurity (%)	00	ce	0.139	0.186	K10
ri	0.109	0.108	0.105	0.107
	0.146	S£T0	0.138	9ΠΌ
	800	S	0.144	9LL0
—	1 0Ό79	!	0.16	^: 0.142
	£110	0.105	0.095	BQL
Observation at diffère Impurity 15 (%) i	24	BQL	—1 θ'	BQL	ND
GO	BQL	BQL	ND	BQL
2!	lÙfJ	BQL	(J N	ND
	ND	ND	ND	ND
	g	ND	ND	ND
-	ND	BQL	ND	ND
o	i ^.055	0.068	0.039	850Ό
Impurity A (%)	rî	0.545	QQ		BQL
oo	0.49	0.171	8900	BQL
—	o	5	0.055	BQL
	81Ό	I	0.03	BQL
	0.08	θ' œ	BQL	ND
-	soo	BQL	BQL	BQL
1		BQL	BQL	BQL	ON
		3.5		4.5

Table 6

Assay of Cetrorelix acetate eq. to Cetrorelix (%)
Storage conditions
	2-8°C	25°C/60%RH
pH	Unfïltered	Initial	IM	2M	3M	6M	12M	18M	24M	IIM	2M	3M	6M
2.5	104.05	103.97	103.03	105.51	105.04	104.54	103.7			102.76	102.9	102.9	99.97
3	103.56	101.H	101.8	104.9	105.42	104	103.5	104.93	104.77	102.09	104.71	105.08	102.36
3.5	103.86	102.51	101.82	104.88	102.88	104.2	103.1	104.13	104.16	103.65	103.62	102.23	103.68
4	103.76	102.96	104	104.17	104.75	104.84	103.3	104.48	103.61	102.58	103.28	103.72	103.39
4.5	102.52	99.56	103.43	103.97	103.57	103.59	102.6	103.97	103.86	101.66	103.62	102.79	102.77
⁵	99.48									99.02	-

The stabîlîty results for additîonal intermédiare pH ranges were studied at different time points upon storage at 25°C/60%RH and 2- 8°C are given in Table 7 below:

Table 7

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Table 8

Assay of Cetrorelix acetate eq. to Cetrorelix (%)
	Storage conditions
	2-8°C	25°C/60%»	Ή
pH	Unfïïtered	Initial	1M	3M	6M	12M	IM	3M	6M
3.1	99.96	98.67	99.12	99.32	98.98	98.94	98.65	97.65	97.52
3.2	100.89	100.21	99.9	100.89	99.61	101.27	100.06	100.38	98.2
3.3	99.96	99.05	98.58	100.03	99.13	100.29	98.97	99.87	98.13
3.4	100.02	98.54	99.59	-	-	-	99.91	99.97	99.03

Table 9 Stability data of cetrorelix acetate Injection 0.25mg/ml, 1 ml PFS at pH 5

Each mL contaîns cetrorelix acetate eq. to cetrorelix 0.25 Mg, Mannitol 54.8 mg, Lactic acid q.s. to pH adjusted 5.0, Water For Injection q.s. to I mL

	Description	Assay of Cetrorelix acetate eq. To Cetrorelix	Reiated Substances
Known Impurities	Unknown Impurities	Total Impurities
Impurity A	Impurity B	High est Unknown Impurity
95.0% to 105.0% of LC	Not more than 1.0%	Not more than 1.0%	Not more than 0.5%	Not more than 3.5%
UNFILTER	*	99.59
INITIAL	A	99.67	BQL (<0.03 5%)	ND	0.131	0.131
’-8°C OTS	1 M	A	98.13	BQL (<0.03 5%)	ND	0.11	0.182
	2 M	A	98.6	ND	ND	0.109	0.208
	3 M		99.98	ND	ND	0.112	0.198
25°C /50 %	1 M	*	98	BQL (<0.03 5%)	ND	0.106	0.106
RH	2 M	*	98.24	0.Û74	ND	0.109	0.369
OTS	3 M	*	98.18	0.18	ND	0.107	0.353

ND: Not Detected; RH — Relative Humidity; BQL: Below Quantifiable limit;

* Clear colorless solution filled in 1 ml PFS

COMPARATIVE EXAMPLES:

Table 11

Cl

COMPARATIVE EXAMPLE 2

An aqueous solution of cetrorelix acetate was prepared as per the disclosure of US 2013/0303464 (Patel et al.). The composition is illustrated below in Table 12:

Table 12

Ingrédients	Quanti ty (mg/m!)
Cetrorelix acetate	0.25
Mannitol	42.0
Glacial Acetic acid	q.s to pH 3.0
Water for injection	1 ml

Method of Préparation: Water for injection was taken at température between 2°C to 8°C în a vessel. Mannitol was added and dissoived gradually in water for injection with stirring, until a clear solution was obtained. To this cetrorelix acetate was added and dissoived gradually with stirring. Glacial acetic acid was then added and the pH of the solution was adjusted to about 3.0. The volume was made up with water for injection. The solution was stirred for 10-15 minutes and subsequently filtered aseptîcally through a bed of 0.2 pm membrane filter (optiscale 47 capsule, Polyethersulfone membrane filter by Mîllipore). The solution was aseptîcally filled in the réservoir of injection device, i.e. in the barrel of 1 ml glass syringe with a fîll volume of 1.1 ml. The stacked needle în the barrel was stoppered by elastomeric needle shield, covered by a rjgid cap before filling. After filling. the glass syringe (barrel) was stoppered with plunger stopper by vacuum stopperîng în such a manner that there was substantîally no headspace air left inside the syringe. The aqueous solution remains in contact with the plunger stopper made up of rubber, stacked needle made up of stainless Steel and needle shield made up of natural rubber upon storage.

The solution of this comparative example (comparative example 2) filled in glass syringe was subjected to storage stability testing, The level of Impurity A, Impurity B and total impurity in the solution were analyzed initially and upon storage at room température (25°C/60 % relative humidity) by high performance liquid chromatographie technique. The results are provided in Table 13 below.

Table 13: Stabiiity resuIts of comparative example 2

Impurity A (%) (25°C/60%RH)	Impurity B (%) (25^ûC/60%RH)	Total impurity (%) (25°C/60%RH)
Time Point (Months)
0	J	6	0	3	6	0	3	6
0.06	0.84	1.77	ND	0.07	0.17	0.99	1.88	2.83

ND: Not detected; RH- Relative Humidity

It was observed that the solution of cetrorelix acetate of US 2013/0303464 (comparative) showed significant increase in the level of Impurity A and total impurity 5 upon storage at room température. Particularly, the level of Impurity A which is a dégradation impurity increases signifïcantly and increases to 1.77% by weight of cetrorelix in 6 months. Also the level of total impurity increases to 2.83% by weight of cetrorelix in 6 months.

In contrast, the parentéral dosage form comprising the ready-to-inject aqueous 10 solution of cetrorelix acetate ofthe présent invention remains stable at room température for a prolonged period of time whereby there occurs substantially no dégradation or increase in level of Impurity A, other impurities or total impurities upon storage and the solution hâve an extrapolated shelf life of more than 24 months.

Claims

1 l. The parentéral dosage form according to claim 8, wherein the injection device is a pen auto-injector.

12. The parentéral dosage form according to claim l, wherein the stable, aqueous solution is stable for at least l month at 25°C température and 60 % relative humidity.

13. The parentéral dosage form according to daims 1, wherein the stable, aqueous solution is stable for at least 3 months at 25°C température and 60 % relative humidity.

14. The parentéral dosage form according to claim 1, wherein the stable, aqueous solution is stable for at least 6 months at 25°C température and 60 % relative humidity

15. The parentéral dosage form according to claim 1, wherein the parentéral dosage form is suitable for subeutaneous use.

16. The parentéral dosage form according to claim 1, wherein the parentéral dosage form is suitable for întramuscular use.

17. A pharmaceutical composition of cetrorelix or a pharmaceutically acceptable sait îhereof, comprising a decapeptîde of formula I:

HN^NH₂NH

Formula I.

18. A decapeptîde of formula I

Formula 1.

19. The decapeptide of claim 18, wherein the decapeptide is identified by HPLC analysis, the process comprising:

a) injecting a diluent comprising water, acetonitrile and formîc acid înto the chromatographie system,

b) injecting a system suitabilîty solution comprising cetrorelix acetate, diluent and impurity stock solution and recording the chromatogram,

20. A process to identify the decapeptide of claim 18 by HPLC analysis, the process comprising:

a) injecting a diluent comprising water, acetonitrile and formic acid into the chromatographie System,

d) injecting a sample comprising aqueous solution of cetrorelix acetate and placebo préparation înto the chromatographie system, and

21. The process of claim 19 or 20, wherein the mobile phase A and B in the HPLC analysis comprises a briffer, acetonîtrile and tetrahydrofuran, and

1. A parentéral dosage form comprising a stable aqueous solution comprising:

(i) cetrorelix or a pharmaceutically acceptable sait thereof; and (ii) an impurity of Formula I in an amount less than 1% w/v of cetrorelix base.

2. The parentéral dosage form as claimed in daim 1, wherein the dosage form is a stérile, stable, aqueous solution.

3. The parentéral dosage form as claimed in daim 1, wherein the dosage form is a stérile, ready-to-infuse dosage form.

4. The parentéral dosage form as claimed in daim 1, wherein the stable, aqueous solution further comprises an organic acid to adjust the pH in the range of 3 to 5.

5. The parentéral dosage form as claimed in daim 4, wherein the stable, aqueous solution further comprises an osmotic agent.

6 A parentéral dosage form according to claim 1, wherein the amount of cetrorelix or a pharmaceutically acceptable sait thereof is 0.25 mg/ml.

7. A parentéral dosage form according to daim 5, wherein the osmotic agent is present in an amount sufficient for osmolality of the solution in the range of 250 to 375 mOsm/Kg.

8. The parentéral dosage form according to claim 1, wherein the parentéral dosage form is a ready-to-inject, stérile, stable aqueous solution present in the réservoir of an injection device.

9. The parentéral dosage form according to claim 8, wherein the injection device is a prefilled syringe.

10. The parentéral dosage form according to claim 8, wherein the injection device is an auto-injecter.

5 wherein the relative rétention time and relative response factor for the decapeptide is determined to be 0.57 and 1.0, respectively.