WO2013189800A1

WO2013189800A1 - Cisatracurium and beta-cyclodextrin derivative compositions

Info

Publication number: WO2013189800A1
Application number: PCT/EP2013/062119
Authority: WO
Inventors: Pablo CID; Sergio RUIZ
Original assignee: Farmhispania S.A.
Priority date: 2012-06-22
Filing date: 2013-06-12
Publication date: 2013-12-27
Also published as: WO2013189800A9; GB201211153D0

Abstract

A solid pharmaceutical composition comprising cisatracurium or a pharmaceutically acceptable salt thereof, and a sulfoalkylether-β-cyclodextrin derivative.

Description

Cisatracurium and β-cyclodextrin derivative compositions

The present invention relates to pharmaceutical compositions comprising cisatracurium, or a salt thereof, and a derivative of β-cyclodextrin, a process for the preparation of such a composition and its use in medicine.

Cisatracurium, 5-[3-[(lR,2R)-l-[(3,4-dimethoxyphenyl)methyl]-6,7- dimethoxy-2-methyl-3,4-dihydro-lH-isoquinolin-2-yl]propanoyloxy]pentyl 3- [(lR,2R)-l-[(3,4-dimethoxyphenyl)methyl]-6,7-dimethoxy-2-methyl-3,4-dihydro- lH-isoquinolin-2-yl]propanoate, is a neuromuscular-blocking drug or skeletal muscle relaxant in the category of non-depolarizing neuromuscular-blocking drugs, used adjunctively in anesthesia to facilitate endotracheal intubation and to provide skeletal muscle relaxation during surgery or mechanical ventilation. Cisatracurium is one of the ten isomers of the parent molecule, atracurium.

Cisatracurium is commercialized as the besylate salt, compound of Formula I, in an aqueous solution for injection under the brand of NIMBEX®. NIMBEX® is a sterile, non-pyrogenic aqueous solution provided in 5 mL, 10 mL, and 20 mL vials. To slow down its degradation during storage, the pH is adjusted to from 3.25 to 3.65 with benzenesulfonic acid. Both 5 mL and 10 mL vials contain

cisatracurium besylate (CIS), equivalent to 2 mg/mL cisatracurium.

According to its Monograph, NIMBEX® slowly loses potency with time at rate of approximately 5% per year under refrigeration (5°C). NIMBEX® should therefore be stored under refrigeration (2° to 8°C) and protected from light to preserve potency. The rate of loss in potency increases to approximately 5% per month at 25°C. Upon removal from refrigeration to room temperature storage conditions (25°C), NIMBEX® has then to be used within 21 days, even if returned to a refrigerator. The stability therefore of cisatracurium is a problem. One of the main factors affecting the instability of cisatracurium is that it spontaneously degrades at physiological pH via Hofmann elimination (Rapid Commun Mass Spectrom 9 (14), 1457-1464). Because atracurium undergoes Hofmann elimination as a primary route of chemodegradation, one of the major metabolites from this process is laudanosine, a tertiary amino alkaloid reported to be a modest CNS stimulant with epileptogenic activity (Anesthesiol 63 (6): 577-578) and cardiovascular effects such as hypotension and bradycardia (Eur J Anaesthesiol 19 (7), 466-473). Laudanosine decreases the seizure threshold, and thus it can induce seizures if present at sufficient threshold concentrations (Eur J Anaesthesiol 19 (7), 466-73). Taking this into account, it is preferable to obtain pharmaceutical compositions of cisatracurium which are stable versus Hofmann degradation, in order to obtain low levels of laudanosine.

Neuromuscular blocking agents have been used in the operating room for years as an adjunct to anaesthetics and are now utilized in intensive care units for the management of critically ill patients. Train of Four (TOF) testing, performed to measure the degree of neuromuscular blockade using a peripheral nerve stimulator, can assist the practitioner to administer high quality care, given very challenging clinical situations. The goal is to ensure that the patient is adequately paralyzed with neuromuscular blocking agents (NMBA) using TOF monitoring. Because neuromuscular blockade prevents the patient from moving, an advanced airway and ventilation must be established prior to their administration. Use of NMBAs is especially relevant to facilitate endotracheal intubation under special circumstances, such as relaxation of laryngeal muscles to facilitate passage of endotracheal tube or to facilitate mechanical ventilation and improve gas exchange in patients who cannot be managed with sedation, analgesia and ventilator parameter manipulation alone.

It is desirable that neuromuscular blocking agents show a regular behavior when it comes to control of the depth of relaxation allowing a decrease in the total dose of the NMBA and a faster and more complete spontaneous recovery from neuromuscular blockade, without recurrence, after cessation of the infusion. A greater neuromuscular blocking potency has a positive effect in surgical procedures since the required time for carrying out some procedures, such as endotracheal intubation, may be substantially reduced.

Spontaneous recovery from neuromuscular block occurs through

redistribution, buffered diffusion, or metabolism of the neuromuscular blocking agent (NMBA) administered. Recurarization is an undesirable phenomenon of recurrence of neuromuscular block that consists on residual neuromuscular relaxation in the immediate post-operative period. Accumulation of NMBA in the intravenous line may lead to recurarization after flushing the line in the recovery room (Murphy GS, Brail SJ. Anesthesia Analgesia. Residual

neuromuscular block: lessons unlearned). Residual block may be associated with serious adverse events, such as respiratory depression, pharyngeal dysfunction, hypoxemia and prolonged length of stay in the recovery room (Beaussier M, Boughaba MA. Residual neuromuscular blockade. Ann Fr Anesth Reanim

2005;24: 1266-74).

Cyclodextrins are cyclic oligosaccharides with the capability of forming non- covalent inclusion complexes with a variety of molecules adapted to the size of their cavity. The association is generally of apolar character, which leads to the preferred inclusion of hydrophobic structural motifs. Chemically, the cyclodextrins are formed by at least six D-glucopyranosyl units attached by β-(1, 4) glucosidic bonds. The three commercially available natural cyclodextrins, namely α-, β-, and γ- cyclodextrins (with 6, 7, or 8 glucose units respectively), differ in their ring size and solubility. Chemically, modified cyclodextrin derivatives have been prepared with a view to extend the physicochemical properties and inclusion capacity of the parent cyclodextrins. α-, β-, and γ-cyclodextrins are all generally recognized as safe by the FDA.

Cyclodextrins and their pharmaceutical applications have been reviewed by

Stella and Rajewski (Pharmaceutical Research, 14, 556-567, 1997). The effects on solubility and stability of the addition of cyclodextrins or cyclodextrin derivatives to aqueous solutions of steroid hormones have been described in the art. In the Journal of Pharmaceutical Sciences (1992), 81.(8), 756-61 , Albers, E. et. al, disclose a reduction in the rate of degradation of steroid esters by inclusion of 2- hydroxypropyl-[beta]-cyclodextrin. Loftsson T. et al, International Journal of Pharmaceutics (1993), 98(1-3), 225-30, describe the stabilizing effects of 2-hydroxypropyl-[beta]- cyclodextrin, reducing the degradation of medroxyprogesteron acetate and megestrol acetate in buffered solutions from 2.5 to 4 times.

However, in a review article on pharmaceutical applications of cyclodextrins by Loftsson and Brewster (J. Pharm. Sci. 85, 1017- 1025, 1996) it is emphasized that cyclodextrin interactions with labile drug molecules can either have stabilizing or destabilizing effects, depending on the particular combination of drug and cyclodextrin.

WO2001040316A1 discloses the use of a γ-cyclodextrin derivative for the manufacture of a medicament for the reversal of drug-induced neuromuscular block. Sugammadex is a modified γ-cyclodextrin, with a lipophilic core and a hydrophilic periphery. Sugammadex (Bridion®) is an agent for reversal of neuromuscular blockade of the agent rocuronium in general anaesthesia. Sugammadex's binding encapsulation of rocuronium is one of the strongest among cyclodextrins and their guest molecules.

In WO2008/065142, pharmaceutical compositions in the form of an aqueous solution for parenteral administration comprising rocuronium bromide and β- cyclodextrin complexes are described.

US2007/0087999 discloses solid compositions of lyophilized taxoid, hydrophilic polymers and cyclodextrins for parenteral administration purposes, wherein the invention is focused in the improvement of the solubility of the composition for reducing the reconstitution time.

Patent CN1376463 describes an eye drop pharmaceutical composition for topical ocular use which includes a cyclodextrine and disulfiram, where the disulfiram has an improved solubility. The disulfiram and the cyclodextrine are mixed in water and lyophilized. The final product is obtained by formulating a lyophilized powder with water ready for eye aplication. This composition is for treatment of cataracts and aims to increase the solubility of disulfiram.

Patent application CN101084896 discloses freeze drying compositions of atracurium. The composition adopts atracurium or its pharmaceutically acceptable salt or optical isomer as active ingredient and also comprises sugar and organic acid. According to its description, these compositions have improved water-solubility and improved storing stability.

Taking into account the stringent storage conditions required for the commercial composition of cisatracurium (NIMBEX®), there is thus a great need in the art for pharmaceutical compositions of cisatracurium with improved stability which can be stored at room temperature (25°C) while maintaining an adequate shelf-life and without having a negative effect on its neuromuscular blocking activity.

This problem is solved by cistracurium compositions according to the present invention. This object is achieved by providing an improved stability solid composition comprising cisatracurium or a pharmaceutically acceptable salt thereof and a sulfoalkylether-β-cyclodextrin derivative.

It has been surprisingly found that compositions of cisatracurium according to the present invention do not require storage at low temperature. Whilst the use of cyclodextrins to enhance solubility and as stabilising agents is generally known, there was no reason to expect that the use of a sulfoalkylether-β-cyclodextrin with cisatracurium as claimed here, specifically in a solid state, would enhance thermal stability.

Surprisingly, the presence of a β-cyclodextrin in the compositions according to the present invention not only does not cause binding encapsulation of cisatracurium but also improves its behavior as a neuromuscular blocking agent.

Summary of Invention It is an object of the present invention to provide a neuromuscular blocker or skeletal muscle relaxant composition which overcomes the drawbacks of the prior art. Especially a composition shall be provided showing an improved thermal stability, a competitive neuromuscular blocking activity, and a regular behavior and duration.

Thus, viewed from one aspect the invention provides a solid pharmaceutical composition comprising cisatracurium or a pharmaceutically acceptable salt thereof and a sulfoalkylether-β-cyclodextrin derivative. Viewed from another aspect the invention provides a lyophilised solid pharmaceutical composition comprising cisatracurium or a pharmaceutically acceptable salt thereof and a sulfoalkylether-β-cyclodextrin derivative.

Viewed from another aspect the invention provides a process for preparing a lyophilised composition of cisatracurium or a pharmaceutically acceptable salt thereof and a sulfoalkylether-β-cyclodextrin derivative comprising the steps of: a) dissolving cisatracurium or a pharmaceutically acceptable salt thereof, and a sulfoalkylether-β-cyclodextrin derivative, together or separately, in acidified water, and if necessary combining the two solutions;

b) optionally adjusting the pH of the solution to a pH of from 3.6 to 4.0, preferably 3.8, with an acid, wherein a preferred acid is benzenesulfonic acid; c) freeze-drying the solution to obtain a lyophilized composition, e.g. a powder.

Viewed from another aspect, the present invention provides a lyophilized powder obtainable by a process as hereinbefore defined.

The present invention also provides a pharmaceutical composition as herein before defined for use in medicine, for example as a neuromuscular blocking composition.

Viewed from another aspect the invention provides a method of treating a patient in need of such treatment with cisatracurium or a pharmaceutically acceptable salt thereof comprising:

(I) reconstituting the pharmaceutical composition as hereinbefore defined in water for injections;

(II) injecting an effective amount of cisatracurium or a pharmaceutically acceptable salt thereof into the patient in order to cause, inter alia, a neuromuscular block.

Detailed Description of Invention The pharmaceutical compositions comprising cisatracurium, or a

pharmaceutically acceptable salt thereof, and a sulfoalkylether-P-cyclodextrin derivative according to the present invention are solid compositions. More preferably, the solid is in the form of a powder such as a free flowing powder.

These compositions are suitable for reconstitution at the moment of use.

β-Cyclodextrin is a cyclic oligosaccharide containing seven α-(1-4)-linked D-glucopyranose units. The term sulfoalkylether-β-cyclodextrin derivative means β- cyclodextrin which has been derivatised at one or more of its 2-, 3- and 6-hydroxyl functions to introduce an anionic (C_2-6alkylene)-SO₃- substituent. Such derivatives are disclosed in US Patent 5,134,127 (University of Kansas). A sulfoalkylether-β- cyclodextrin derivative may either be a single well defined derivative or may be a mixture of derivatives obtained by random derivatisation of the β-cyclodextrin hydroxyl functions with a certain molecular excess of an alkylating reagent, such as for example an (C_2-6alkyl)sultone in the presence of a base, for instance by procedures described in US 6,153,746 (Pfizer Inc). Such randomly derivatised sulfoalkylether-β-cyclodextrin derivatives are characterized by a degree of substitution, meaning the average number of derivatised hydroxyl groups per molecule. The degree of substitution may range from about 1 to about 8

sulfoalkylether groups per β-cyclodextrin.

Preferred sulfoalkylether-β-cyclodextrin derivatives for use in the invention are the sulfobutylether-β-cyclodextrin derivatives (SBE-β-CDs). In an especially preferred embodiment, the compositions of the invention comprise a

sulfobutylether-β-cyclodextrin having an average of about 7 sulfobutylether substituents per cyclodextrin molecule (annotated as: sulfobutylether 7-β- cyclodextrin, SBE7-β-CD), for example with a general Formula (II), wherein R is - (CH₂)₄SO₃Na or -H. Sulfobutylether 7-β-cyclodextrin is commercialized as Captisol® and Advasep 7.

One of the possible representations of the structure of Captisol® could be a product of Formula (III).

preferred embodiment the solid composition is a lyophilized powder. Surprisingly, the inventors believe that the interaction between the sulfoalkylether-β-cyclodextrin and cisatracurium established during the freeze drying cycle increases the stability of the obtained composition.

A preferred embodiment according to compositions of the present invention encompasses pharmaceutical compositions wherein the molar ratio of cisatracurium or salt thereof to sulfoalkylether-β-cyclodextrin derivative is from 1 : 1 to 1 : 10, for example 1 :2, preferably 1 :3 to 1 :6, more preferably from 1 :4 to 1 :5. Ratios of approximately 1 :4 are especially preferred.

The pharmaceutical composition of the invention should contain an effective amount of the cisatracurium or salt thereof. Dosages will be familiar to the skilled worker and will depend on the patient. In a preferred embodiment according to the present invention,

pharmaceutical compositions of above comprise cisatracurium in the form of a salt, wherein a preferred salt is the benzenesulfonate (besylate) salt.

In order to prepare compositions according to the present invention, it is preferred to start with an acidic aqueous solution at an appropriate pH such as from 3.0 to 4.5. Optionally, the pH of the starting solution may be adjusted with a base, preferably NaOH aqueous solution, for example NaOH IN. The solution of the sulfoalkylether-β-cyclodextrin derivative and the solution cisatracurium or a salt thereof, in acidified water could be prepared all together or separately. Preferably, both solutions are prepared separately and subsequently mixed.

In a preferred embodiment, the acidic solution used in the process is prepared by dissolving benzenesulfonic acid in water for injection.

The acidified solutions containing cisatracurium or a salt thereof and/or the sulfoalkylether-β-cyclodextrin derivative respectively may both have a pH in the range 3.0 to 4.5. Preferably, both pH's are approximately the same.

The two solutions can be mixed in an appropriate ratio. If necessary, pH can then be adjusted, preferably using benzenesulfonic acid in water for injection. The pH before freeze drying is ideally 3.6 to 4.0, preferably 3.7 to 3.9, more preferably 3.8. This might be achieved by adjustment of the pH after mixing or by making sure that the pH of the solutions before mixing is correct.

The processes for preparing a pharmaceutical compositions according to the present invention may optionally include a step before the freeze-drying process, which involves filtration of the acidic solution of cisatracurium and cyclodextrin in order to sterilize it.

Thereafter, or after formulation in acidified water, the solution can by freeze dried to form a solid. The solid obtained at this point may have less than 6 wt% water, preferably less than 5 wt%.

The term lyophilization when used in the context of the present invention refers to the drying process freeze-drying. The product obtained by this process, in the context of the present invention, is named lyophilized powder. Freeze drying can be carried out conventionally using well known apparatus. It will be appreciated therefore that the compostion of the invention might comprise an acid buffer, especially benzenesulfonic acid.

In a preferred embodiment, pharmaceutical compositions according to the present invention are suitable for reconstitution in injectable form.

The compositions of the invention can be stored at room temperature (25 °C) without loss of activity for prolonged periods, e.g. at least 2 months, hi general, it is envisaged that the solution of the invention performs as well at room temperature storage as NIMBEX stored in a refrigerator.

The compositions of the invention may exceed the NMBA performance of NIMBEX. Preferably the compositions of the invention will show improvement in the onset and/or duration of neuromuscular block. The compositions of the invention may also or alternatively show improved maintenance properties in comparison with NIMBEX, such as reduced maintenance dose (2nd bolus) and/or longer time between maintenance doses. The compositions of the invention may also or alternatively show improved recovery times and/or improved recovery characteristcs, for example, faster or more regular recovery profile.

The term Captisol®, when used in the context of the present invention, refers to sulfobutylether-7-β-cyclodextrin.

In the context of the present invention, HPLC analysis were made using a Modular apparatus for liquid chromatography WATERS, comprising pump and autoinjector (model W2695) and detector two wavelengths (model 2487) or PDA detector (model 2996), with an Empower's data acquisition and processing system. Chromatographic methods and assessment of related substances are described in the European Pharmacopoeia's monograph for Atracurium besilate v. 7.0.

The term Recovery (%), when used in the context of the present invention, refers to the ratio between the assay solution and theoretical assay based on API weight.

The term Total Impurities (%), when used in the context of the present invention, refers to the sum of the all the impurities determined by HPLC.

In the context of the present invention, neuromuscular blocking agents

(NMBA) refer to a group of drugs that prevents motor nerve endings from exciting skeletal muscle. TOF is an usual mode of stimulation for clinical monitoring of

neuromuscular junction (Ali HH, et. al. Stimulus frequency in the detection of neuromuscular block in humans. Br J Anaesth 1970; 42:967-78.). Four successive stimuli are given at a frequency of 2 Hz, potentially eliciting 4 twitches (T1-T4). The ratio T4:T1 indicates the degree of neuromuscular block. Non-depolarizing NMBAs produce a decrease in magnitude of the first twitch compared with a pre- relaxant stimulus, and a progressive reduction in magnitude of T1-T4. The number of elicited twitches indicates the degree of receptor occupancy. Disappearance of T4, T3, T2, Tl corresponds to 75%, 80%, 90% and 100% occupancy. With recovery of neuromuscular function the twitches appear in the reverse order.

Accepted values for TOF count are: a) 1 twitch for tracheal intubation, b) 1-2 twitches during established anaesthesia and c) 3-4 twitches before reversal of neuromuscular blockade is attempted.

Time to Tl (>90% block) is a measure of the onset time for neuromuscular block. Faster onset of neuromuscular block may be indicated by a shorter time to Tl. This may have various advantages such as allowing a more rapid intubation, shorter overall time under anaesthetic, reduced overall dosage, etc. In practice, a second bolus dose is administered when the TOF monitor shows signs of muscle recovery, i.e. when T3 is reached, in order to maintain a surgical level of relaxation between T2-T3. Second bolus time is a measure of the maintenance of

neuromuscular block. The second bolus dose is the standard maintenance dose of relaxation effect for muscle receptor occupancy and typically may be approximately 25% of the initial dose. An increased second bolus time may have advantages in terms of the overall dose administered to maintain neuromuscular block for a certain period. A regular second bolus time also may be advantageous in providing improved control of the level of neuromuscular block.

In the context of the present invention, spontaneous recovery refers to a physical condition in which muscle relaxant activity has vanished and the subject is able to maintain itself breathing without ventilatory support and with no need of reversal agents. As for the second bolus time, a more regular and therefore more predictable recovery time may be advantageous. In the context of the present invention, Solid-state Carbon- 13 NMR refers to a kind of Carbon- 13 nuclear magnetic resonance spectroscopy (CI 3 NMR), characterized by the presence of anisotropic (directionally dependent) interactions.

In the context of the present invention, Thermogravimetric analysis (TG) refers to the thermal analysis method in which changes in physical and chemical properties of materials are measured as a function of increasing temperature (with constant heating rate), or as a function of time (with constant temperature and/or constant mass loss).

The invention will now be described with reference to the following non limiting examples and figures.

Brief Description of the drawings

Figure 1. HPLC Recovery (%) of cisatracurium/Captisol® (25°C) composition and NIMBEX® (5°C) vs time.

Figure 2. Impurities (%) by HPLC of cisatracurium/Captisol® (25°C) composition and NIMBEX® (5°C) vs time.

Figure 3. TG comparative: captisol®, CIS and cisatracurium/captisol® lyophilized powder (1 :4).

Figure 4. TG comparative: cisatracurium/captisol®: lyophilized powder (1 :4), physical mixture (1 :4) and CIS.

Figure 5. Solid-state C13 NMR of cisatracurium/captisol®: lyophilized powder (1 :4) vs. physical mixture (1 :4).

Figure 6. Solid-state C13 NMR of cisatracurium/captisol® lyophilized powder: overlapped (1 :4) (1 :2) (1 : 1).

Figure 7. Solid-state CI 3 NMR of cisatracurium.

Figure 8. Solid-state C 13 NMR of captisol®.

Examples - Preparative example 1.

Cisatracurium besylate used in the examples and comparative examples was obtained according to a process described in EP-A-0539470.

Comparative Example 1. Fresh solution according to NEV1BEX® composition.

To ensure perfect condition of the reference product for HPLC essays, a fresh solution was prepared according to the composition of NIMBEX® 2 mg/mL disclosed in the FDA's PDR instead of using a commercial sample, since there is no conclusive information of its storage.

250 mL of cisatracurium besylate solution were prepared by dissolving 704 mg of cisatracurium besylate in an acidic solution, pH: 3.25, of benzenesulfonic acid in water for injection. The final pH was readjusted to 3.2-3.3 with a solution of benzenesulfonic acid 1% by weight in water for injection.

Immediately after its preparation, samples of 5 mL with a concentration of 2 mg/mL were stored at 5°C.

Example 1.

Cisatracurium/β-cyclodextrin derivative lyophilized powder, molar ratio (1 :4)

704 mg of cisatracurium besylate were dissolved in approximately 130 mL of an acidic solution, pH: 3.7-3.8, of benzenesulfonic acid in water for injection. 5,0 g of Captisol® were dissolved in approximately 20 mL of an acidic solution, pH: 3.7-3.8, of benzenesulfonic acid in water for injection. Both solutions were mixed and some acidic solution, pH: 3.7-3.8, of benzenesulfonic acid in water for injection was added up to 250 mL of solution. The pH was readjusted to 3.8 with a solution of benzenesulfonic acid 1% by weight in water for injection just before reaching a total volume of solution of 250 mL. The solution was divided in samples of 5 mL, with a cisatracurium concentration of 2 mg/mL, and was dried by freeze-drying obtaining a lyophilized powder. The samples were stored at room temperature (25°C) and 40°C.

Example 2.

A sample obtained in Example 1, stored at 25°C, was reconstituted with 50 mL of water for injection. The solution obtained was analyzed by HPLC obtaining data of its Essay and its Degradation. A sample of a composition emulating fresh NIMBEX® prepared and stored at 5°C, according to Comparative Example 1, was analyzed by HPLC obtaining data of its Recovery and its Degradation.

Parallel analysis by HPLC for each composition were performed to obtain the data in Table 1, wherein the variation of the Essay and the Degradation of the stored samples with time is showed.

Example 3.

Cisatracurium/β-cyclodextrin derivative lyophilized powder, molar ratio (1 :4)

704 mg of cisatracurium besylate were dissolved in approximately 130 mL of an acidic solution, pH: 3.7-3.8, of benzenesulfonic acid in water for injection. 5,0 g of Captisol® were dissolved in approximately 20 mL of an acidic solution, pH: 3.7-3.8, of benzenesulfonic acid in water for injection. Both solutions were mixed and some acidic solution, pH: 3.7-3.8, of benzenesulfonic acid in water for injection was added up to 250 mL of solution. The pH was readjusted to 3.6 with a solution of benzenesulfonic acid 1% by weight in water for injection just before reaching a total volume of solution of 250 mL. The solution was divided in samples of 5 mL, with a cisatracurium concentration of 2 mg/mL, and was dried by freeze-drying obtaining a lyophilized powder. The samples were stored at room temperature (25°C) and 40°C. After two months of storage at temperature 40°C, a sample was analyzed by HPLC (Recovery: 93% and Total Impurities 5.2%).

Example 4.

With the aim of determining the effect of the compositions disclosed in the patent application CN101084896 with respect to cis-atracurium stability, examples according to this application were done and stored under stability at 40°C. To discard the impact of the method used in the preparation of the samples, Example 2 of CN101084896 was reproduced, one literally, and other following a process according to present invention.

4.a Samples for stability were prepared according to the process of Example 1 of the present invention using the quantities of Example 2 of CN101084896.

709.7 mg cisatracurium besylate (2 mg/mL)

17.9 g Lactose Monohydrate

Ratio: 1 :20 w/w 4.b Samples for stability were prepared according to the process of Example

2 of CN101084896.

629.0 mg cisatracurium besylate

13.1 g Lactose Monohydrate

Ratio: 1 :20 w/w

4.c Samples for stability were prepared according to the process of Example 1 of the present invention using the quantities of Example 5 of CN101084896.

711.7 mg cisatracurium besylate (2 mg/mL)

12.0 g Glucose

Ratio: 1 : 17 w/w

The samples obtained in each example were analyzed by HPLC: Table 2.

Example 5.

Pharmacodynamic behavior of cisatracurium/p-cyclodextrin derivative compositions - Induced neuromuscular block, maintenance of the neuromuscular block and spontaneous recovery. a) Sample preparation.

Vials with cisatracurium/captisol® molar ratio (1 :4) lyophilized powder were prepared according to example 1 of the present invention.

Vials with cisatracurium/captisol® molar ratio (1 :2) lyophilized powder were prepared according to example 1 of the present invention just modifying the quantity of captisol® .

Vials were mantained at room temperature until their use. The lyophilized powder of each vial was reconstituted with 5 mL of salina before its injection. All the samples showed a high solubility. b) Methodology and materials.

White large pigs, between 16 and 21 kg, were prepared following modifications of the reported methods (Henning, R. Br. J. Pharmacol. 1993, 108, 717-720; Thesleff, S. et al, J Pharmacol. Expt. Ther. 1954, 111, 99-118). Intramuscular premedication consisted of midazolam 0.2 mg/kg, azaperone 2 mg/kg and medetomidine 4 mg/kg, 1 hr before the beginning of surgery. On arrival in the operating room, the electrocardiogram (ECG), haemoglobin oxygen saturation (SpO2) and non-invasive arterial pressure (NIBP) were monitored. An intravenous cannula was inserted into an ear vein and it was maintained with saline at 6 ml/h.

For intubation, an anaesthetic induction mask and sevorane 8% were used.

Once intubated, anaesthesia was only maintained with oxygen flow of 10-15 ml/kg/min and sevoforane 2-3%. for one hour to avoid a potential impact of the premedication on the results of the essays.

After that, a first bolus of NMB A, dose of 0.4 mg/kg, was given. A second bolus, with a ¼ of the initial dose, is administered when the TOF monitor shows signs of muscle recovery (T3). Apneas, CO₂ maintainability of spontaneous breathing and muscle resilience were monitored and recorded during the procedure.

Spontaneous recovery of neuromuscular function was allowed (controlled by TOF). The recovery process evolution was followed for 24 hours.

Equipment used during anaesthesia and surgery:

o 5250RGM Ohmeda® Anaesthesia Monitor with relevant modules. C02 levels, respiratory rate, apnea, tidal volume, airway pressure, CAM, Pulse Oxymetry (Sp02).

o Harvard esophageal temperature sensor.

o Electromyogram Biopac System MP36 with 2 Channels

o 2 TOF- Watch® SX Monitor

o Thermal Blanket c) Results

Data about time to reach Tl, i.e. >90% blocking; time to a second bolus

(maintenance of the neuromuscular block); and time until spontaneous recovery, are shown in the following table:

In all the essays, animal spontaneous recovery was good without recurrence of the neuromuscular block.

Example 6.

Thermogravimetric analysis (TG) and solid-state C13 NMR of

cisatracurium/β-cyclodextrin derivative compositions (lyophilized powder) vs.

physical mixture of cisatracurium and β-cyclodextrin derivative. a) Sample preparation.

Vials with cisatracurium/captisol® lyophilized powder were prepared according to example 1 of the present invention just modifying the quantity of captisol®.

Vials with a physical mixture of cisatracurium/captisol® were prepared, by stirring the two products in a vial until complete homogenization of the sample. b) Methodology and materials.

Equipment used during TG trace measurements:

Instrument: TA instruments SDT Q600 V20.9 Build 20, Pan: Platinum, Gasl : Nitrogen 0.0 ml/min, Gas2: Nitrogen 100.0 ml/min, Method: Ramp 10.00 °C/min to 400.00 °C.

Equipment used during solid-state C13 NMR measurements:

Samples are stored in dried conditions from its delivery to analysis. Samples were weighted and packed into 4 mm Zr02 rotors.

Spectrometer: Broker Avance III 400 MHz., Probe: MAS 4BL CP BB, Temperature: 298K controlled by BCU-Xtreme, Transmitter frequency offset 1H 3.0 ppm, Transmitter frequency offset 13C: 100.0 ppm, Spectral width: 496.8629pm, Number of scans: 3312 (13k), Size of FID: 4096 (4K), FID resolution: 12.21 Hz, Inter scan delay: 4 seconds, Acquisition Time (FID time): 0.041 seconds, Pulse program: cp.av, Cross Polarization Mixing Time: 2500 μs, Cross Polarization Field Strength: 80 KHz, Sample spinning rate: 10 KHz, External reference: adamantine. c) Results and discussion.

According to the information derived from the thermal analysis of both kind of samples, the degree of molecular arrangement in the CIS thermal stabilization was significantly higher in the case of the sample prepared by lyophilization than in the sample prepared by physical mixture. The differences observed between the solid-state 13C NMR spectra of CIS-Captisol physical mixture and CIS-Captisol lyophilized powder are indicative of a different molecular-level ordering, that could be associated with the forming of a molecular complex in the lyophilized powder product.

Figure 4 shows TG spectra of cisatracurium/captisol® (1 :4) lyophilized powder and the physical mixture (1 :4). The spectral profiles substantially differ although both samples contains the same ratio of both components, hence it appears that the two samples have significantly different structures.

Solid-state C13 NMR experiments have shown differences in the chemical shift position of some peaks, as a general procedure on this analytical technique, when comparing physical mixtures vs complexes involving cyclodextrins. Main differences between the physical mixture and the lyophilized powder of the current invention are tabulated in the following Table 4, wherein C designates carbon atoms of the CIS molecule and ** designates carbon atoms of the Captisol® molecule.

Carbon atoms assignment is disclosed in the structure of both compounds (Formula I and Formula III).

There are also highly noticeable changes in the intensity of some peaks, which are clearly different between both spectra, especially a loss of signal in the cisatracurium/captisol® lyophilized spectra around 55 ppm (see Figure 5).

Differences are reported in the following Table 5:

Direct comparison of the solid-state CI 3 NMR spectra of

cisatracurium/captisol® lyophilized powder samples with ratios 1 : 1, 1 :2 and 1 :4 (Figure 6), shows that there are two groups of signals, designated respectively as zone Y and zones G. While intensity of peaks in designated zone Y decrease as expected according to the cisatracurium/captisol® ratio, the changes in intensity of peaks in the designated zone G is observed to occur in a more severe way. That means that there is a differentiation in CrossPolarization transfer mechanism when comparing 1 : 1 and 1 :4 samples in those zone G signals. Notice that samples 1 : 1 and 1 :4 only could be considered equal if observed intensity decrease was the same for each signal, and this is not the case.

According this, solid-state NMR indicates that 1 : 1 and 1 :4 samples have a different molecular arrangement. It is also important to remark that results are in concordance with those obtained after comparing the physical mixture (1 :4) and the lyophilized powder (1 :4) because loss of signal was observed just for the same signals.

Claims

1. A solid pharmaceutical composition comprising cisatracurium or a pharmaceutically acceptable salt thereof, and a sulfoalkylether-β-cyclodextrin derivative.

2. A pharmaceutical composition according to claim 1 , wherein the solid pharmaceutical composition is a lyophilized.

3. A pharmaceutical composition according to claim 2, wherein the solid pharmaceutical composition is a lyophilized powder.

4. A pharmaceutical composition according to any of claims the 1 to 3, wherein the sulfoalkylether-β-cyclodextrin derivative is a mixture of sulfobutylether-β- cyclodextrin derivatives.

5. A pharmaceutical composition according to claim 4 wherein the

sulfobutylether-β-cyclodextrin derivative is sulfobutylether-7-β-cyclodextrin.

6. A pharmaceutical composition according to any one of the claims 1 to 5, wherein the molar ratio of cisatracurium to sulfoalkylether-β-cyclodextrin derivative is from 1 :3 to 1 :6, preferably from 1 :4 to 1 :5.

7. A pharmaceutical composition according to any of the claims 1 to 6, wherein the composition comprises an acid buffer.

8. A pharmaceutical composition according to claim 7, wherein the acid buffer is benzenesulfonic acid.

9. A pharmaceutical composition according to any of the claims 1 to 8, wherein cisatracurium or salt thereof is cisatracurium besylate.

10. A process for preparing a lyophilised composition of cisatracurium or a pharmaceutically acceptable salt thereof and a sulfoalkylether-β-cyclodextrin derivative comprising the steps of:

a) dissolving cisatracurium or a pharmaceutically acceptable salt thereof, and a sulfoalkylether-β-cyclodextrin derivative, together or separately, in acidified water, and if necessary combining the two solutions;

11. A lyophilized powder (for a solid pharmaceutical composition) obtainable by a process according to claim 10.

12. A composition according to any of the claims 1 to 9 and 11 for use in medicine.

13. A composition according to claim 12 for use as neuromuscular blocking agent.