CZ697A3 - Therapeutical human albumin with low bonding capacity to aluminium - Google Patents

Abstract

There is described a therapeutic human albumin having a very low capacity for the absorption of aluminium, during storage in a glass receptacle, wherein the final composition of the albumin solution adjusted to be stable and isotonic at a concentration of 5, 20 or 25% of protein in an aqueous medium, preferably, or to any other therapeutic concentration acceptable for intravenous administration, has a citrate content in the final albumin composition equal to or less than 0.5 mM (millimolar) and, preferably, less than 0.037 mM (millimolar). <IMAGE>

Description

The present invention relates to a therapeutic human albumin composition having low aluminum binding capacity. The property of this composition is the low absorption and fixation capacity of polyvalent metals eluted from the vessel or container in which the solution is placed.

The present invention is mainly used in the field of therapeutic plasma proteins for intravenous administration and is particularly suitable for albumin solutions, although it may be used for other proteins with similar properties.

BACKGROUND OF THE INVENTION

An essential feature that any pharmaceutical composition must have is its stability until the expiration date.

In order to achieve long stability, possible structural or functional changes of the active ingredient are determined. Sometimes it is the degradation or change of product that leads to the formation of undesirable toxic compounds. Their maximum levels are given by Pharmacopea or other legal standards.

In the case of plasma proteins for intravenous administration and in the specific case of certain polluting metals, such as aluminum, the concentration must be as low as possible, taking into account the toxicity of the product which is capable of causing, in traces, disorders such as disorders, which are referred to as Alzheimer's type senile dementia, breakdown encephalopathy, etc. [Klein GL: Aluminum in Parenteral Products: Medical Perspective on Large and Smalys Volume Parenterals, Journal of Parenteral Science and Technology 43 (3). 120 (1989)].

Toxicity studies show that aluminum administered intravenously may accumulate in tissues and organs (brain), has a toxic effect on patients with renal dysfunction due to insufficient clearance.

For this reason, the European Health Authority has determined the level of aluminum in intravenous albumin to be less than 200 ppb (µg / l) in solutions having a 5, 20 or 25% (w / w) protein concentration.

Naturally, the maximum aluminum concentration limit in albumin must not be determined immediately after preparation of the product, but must be maintained for the expiration period.

One factor that commonly limits the useful time of therapeutic human albumin is the increase in aluminum concentration over time. This increase in aluminum concentration is due to the absorption of this metal from the glass containers used to store albumin.

It is known that glass containers used as product containers contain considerable amounts of aluminum that can be extracted from the crystal lattice [Hoiberg Ch. P .: Aluminum in Parenteral Products: Overwiev of Chemistry Concerns and Regulators Actions, Journal of Parenteral Science and Technology 43 (3), 127 (1989).].

Commercial ampoules made of neutral or borosilicate glass (type I), which have low alkaline extractability, contain more aluminum than standard surface treated silicate glasses (ammonium salts or sulfur oxides: type II glass), so that alkalinity is not directly associated with potential release aluminum glass. It is therefore preferred to use standard Type II commercial glass rather than Type I.

Conventional technology does not precisely solve the problem of increasing aluminum during storage, especially at higher temperatures, which are temperatures from 25 to 35 ° C [Quagliaro D.A. et al., Aluminum in Albumin for Injection, Journal of Parenteral Science and Technology 42 (6). 187 (1988)].

In order to achieve aluminum levels below 200 ppb (gg / l) in the albumin solution, not only immediately after preparation but also at the end of its life, albumin is prepared so that it does not come into contact with exogenous aluminum contamination at all times. any step of fractionating human plasma with ethanol at reduced temperature or according to the Cohn method (Cohn Y. et al., J. Amer. Chem. Soc. 68. 459 (1946).) (use of aluminum-free or low-aluminum clarification filters; Pre-washed acidic media, freshly prepared acidic and alkaline reagents When using fraction V (according to Cohn's counting) as the starting material for albumin recovery, an ultrafiltration dialysis technique is used to reduce aluminum levels to less than 200 ppb. g / l). These procedures were first introduced in 1987 by the Scottish National Blood Transfusion Service [McBay W.E. et al., Aluminum and Human Albumin Solutions, British Medical Journal 295. 1062 (1987).], then Millipore in cooperation with the Swiss Red Cross (internal communication) and a similar method was later described by Kabim (WO 91/00290) .

These procedures were based on the elution of polyvalent metals from albumin during dialysis by ionic metal substitution. The amount of metal ion, such as aluminum, is reduced by diafiltration through membranes trapping 10,000 to 30,000 molecular weights that capture proteins (having a higher molecular weight) and allow the passage of ions or molecules that are smaller than the pores of the membranes.

This washing procedure is readily performed without increasing the concentration of monovalent ions (e.g. sodium chloride), which allows the replacement of polyvalent metals such as aluminum. One way to maintain the concentration of monovalent ions (e.g. sodium chloride) at a constant level is to use my known diafiltration method at a constant volume. Conventional albumin diafiltration methods are used to reduce the amount of aluminum (and other metals) to less than 200 ppb and also to reduce the amount of ethanol and excess salts (sodium chloride or other low molecular weight compounds) that may be present in the fraction In the Cohn fractionation process, which is used as the starting material. In this way, it is possible to reduce the amount of aluminum ions to less than 200 ppb and even to less than 50 ppb by using three volumes of a dialysis solution formed with a monovalent salt (sodium chloride).

However, solutions thus obtained having an aluminum level of less than 200 ppb (Mg / l) gradually increase the contents over time when stored in a final container (type I or II glass) and (stability test) at room temperature (25 to 35 ° C). of aluminum, so that at the time of expiration (for example after 3 years) it easily exceeds the given limit of 200 ppm (/ ig / l).

Thus, there remains a technical solution to this phenomenon of increasing the amount of aluminum during storage or storage of protein solutions, mainly albumin, in glass containers.

It is therefore an object of the present invention to provide an albumin composition having a low aluminum binding capacity that can be stored at higher than normal temperatures (room temperature or higher) or whose shelf life or expiration can be extended.

Prior to the present invention it was generally assumed that the release of aluminum from the container (glass) occurs solely because of the ability of aluminum to fix and transport metals and that it is therefore not possible to solve this problem without altering the composition or surface of the glass used (preparation of non-standard special glasses) .

The investigations carried out by the authors of the invention have made it possible to demonstrate the causal nature of the aluminum content increase phenomenon during storage of albumin in containers, and has made it possible to develop a final composition for human therapeutic albumin which overcomes the aforementioned disadvantages.

SUMMARY OF THE INVENTION

The fresh or isolated human plasma from which the terpeutic albumin is prepared contains an anticoagulant solution in which citrate ions are always present.

The study of citrate distribution in plasma fractions revealed the presence of substantial amounts of this anion in fraction V, which is used as a starting material for the preparation of albumin (from about 10 to 20% (w / w) relative to the original plasma citrate content). separation of fraction V).

This citrate contamination occurs in all albumin solutions prepared by the Cohn method. Typically, the concentration exceeds 1 mmol / l in the final composition of the final product (e.g. with 20 wt% protein) despite the fact that the starting fraction V solution is diafiltered to remove aluminum to less than 200 ppb or even 50 ppb or less .

The inventors have described that the presence of citrate (always present in the fraction V used as starting material) in albumin solutions stored in glass greatly increases the release of aluminum from the glass containers containing these solutions. This effect has been demonstrated experimentally. There is a logarithmic relationship between release rate and citrate concentration. Thus, the amount of citrate should be reduced to an amount lower than that existing in fraction V.

The mechanism of release, although not well known today, is believed to be based on the balance between the albumin-citrate complex. Its dissociation by ionic charge promotes the presence of free citrate, which is able to cause the migration of aluminum from the glass. In the absence of a free citrate anion, albumin alone cannot capture aluminum, although it is nevertheless a good carrier and can strongly associate with any polyvalent metal.

As a result of the inventors' research, it has been discovered that a significant reduction in the increase in aluminum content can be achieved if the final composition of the albumin solution is rendered stable and isotonic at a concentration of 5, 20 or 25% by weight. protein in an aqueous medium, or preferably at any other therapeutic concentration acceptable for intravenous administration, is preferably contained in standard commercial glass (type II) containers (ampoules) and if the citrate concentration in the final albumin composition is preferably 0.5 or less mM (millimolar), preferably less than 0.037 mM.

The product obtained complies with European regulations specifying less than 200 ppb (jug / l) of aluminum and usually less than 50 ppb (gg / l) of aluminum. The product obtained is stable for more than three years at 25 ± 10 ° C without exceeding the maximum aluminum content limits under storage conditions.

The present invention will be explained here based on non-limiting examples of embodiments of the invention.

DETAILED DESCRIPTION OF THE INVENTION

Example 1

Three groups of fresh frozen plasma containing ACD (citric acid / dextrose) as an anticoagulant are fractionated by the Cohn method. Fraction V was obtained. After diafiltration and adjustment to a 20% (w / w) protein concentration in the sterile preparation prepared, each group was dispensed under aseptic conditions into 50 ml ampoules (20 mm neck) of Type II standard glass (BSN) and hermetically will be sealed. They are then subjected to a pasteurization step (10 h at 60 ° C) and accelerated quarantine (14 days at 31 ° C).

The citrate concentration of each dose was then determined. The assay was performed by accelerated release of aluminum in 20% (w / w) packaged albumin that was exposed to 45 ° C. The aluminum content was in all cases determined after different exposure times.

The results obtained are shown in Table 1 and are graphically shown in Figure 1, which compares the aluminum content with the exposure time for each citrate concentration.

Table 1

Relationship between final citrate concentration and accelerated release of aluminum and storage time at 45 ° C

concentration citrate concentration of aluminum (t = time in days at (PPb) 45 ° C) t = 120 linear regression (correlation) (g / D (mM) t = 0 t = 30 t = 60 0.21 0.71 20.05 110.3 227.35 446 3.5906.t + 12.42 (r = 0.9992) 0,036 0.12 24.35 75.48 104.25 - 1, 2806.t + 30.26 (r = 0.9922) <0.011 0,037 12.13 12.40 20.43 30.60 0.1442.t + 10.20 (r = 0.8941)

It should be noted that although the original aluminum content (t = 0) is of the same order and very low in all three cases (less than 25 ppb), the same cannot be applied to the citrate concentration which appears to be directly responsible for increase in aluminum content after exposure to elevated temperature (45 ° C).

Linear correlations are significant in all cases (mainly those containing 0.71 and 0.12 mM citrate concentration) and therefore are also important in different aluminum release rate guidelines.

The different aluminum release rates of 3.5906 and 0.1422 ppb / day obtained as a result of the presence of 0.71 or <0.037 mM citrate in albumin show that the release rate is 3.5906 / 0.1422 = 25 times the release rates obtained for the two concentrations citrate.

Example 2

Various portions of the final stabilizer-stabilized albumin (sodium caprylate and sodium tryptophanate) and the isotonic reagent (sodium chloride) to neutral pH were sterilized by filtration.

Each solution was filled under aseptic conditions in two equal portions into 50 ml ampoules (20 mm throat diameter), some of which were made of conventional Type II glass (BSN), others had a special surface treatment (bisulfite). The citrate and aluminum content of the ampoules was determined. The final albumin 3 processed batch formulation had a citrate content less than the quantitative limit of the technique (<0.011 g / L or <0.037 mM). These batches were subjected to an accelerated release of aluminum at 45 ° C. Samples were taken from each batch to determine the release of aluminum over time.

The results are shown in Table 2 below.

Table 2

Accelerated release (45 ° C) of aluminum in albumin depending on the glass of the vial

glass concentration concentration of aluminum linearar- ampoules citrate (t = exposure days 45 ° c) financial co- (m / m) (m / m) t = 0 t = 30 t = 60 t = 90 t = 120 session disi- <0.011 <0.037 12.13 10.73 17.43 13.00 26.13 0,1009.t + ičičitan (± 5.09) ± 0.60 ± 2.75 ± 2.04 +9.83 (r = 0.7655) type II <0.011 <0.037 12.13 12.40 20.43 18.13 30.60 0.1422.t + (BSN) (± 4.24) ± 7.33 ± 4.79 ± 3.83 +10,20 (r = 0.8941)

Taking into account the linear correlations obtained in the aluminum release kinetics of the two groups of vials having different glass surface treatments and the small difference between the slope curves or the release rate, it can be concluded that the type of glass is not a decisive factor for the aluminum release rate beyond provided that the citrate is present at a sufficiently reduced level (close to the detection limit of the technique <0.011 g / l).

Example 3

The same final albumin solution stabilized with sodium caprylate and sodium tryptophanate at 0.016 M concentration in both cases and 20% (w / w) protein concentration containing 0.15 M sodium chloride, pH adjusted to 7.0 ± 0.2, was sterilized by filtration using 0.22pm membrane.

This solution was filled into 50 ml type II glass vials to which known amounts of sodium citrate were added. The mixture was pasteurized at 60 ° C for 10 h. Solutions that were subjected to accelerated release of aluminum were stored at 45 ° C.

The aluminum content of the samples was determined at the times given in Table 3.

Table 3

Accelerated release of aluminum at 45 ° C in 20% (w / w) albumin depending on citrate concentration

concentration citrate aluminum concentration (ppb) (t = exposure time in days at 45 ° C) (g / D (mM) t = 0 t = 30 t = 60 t = 90 t = 150 <0.011 <0.037 9.0 10.0 28.2 31.0 43.6 6.12 1,8 164.2 204.6 330.2 - 721.6 9.51 2.8 161.4 240.6 349 - 886.4 23.1 6.8 165.6 287.2 418.8 - 913.6 40.1 11.8 177.2 276.3 585 "" 1176.0

This table clearly shows the effect of freely added citrate on the ureurized release of aluminum for the same dose of albumin and for the same glass container. This table also shows a significant difference in final albumin levels after 150 days at 45 ° C between albumin solutions at less than 0.037 mM and 1.8 mM citrate, corresponding to 43.6 and 721.6 ppb aluminum, respectively. Thus, a small degree of contamination with citrate or citrate residue greatly accelerates the release of aluminum.

PATENTSERVIS

Claims (1)

Therapeutic human albumin with low aluminum binding capacity, characterized in that the final composition of the albumin solution is modified to be stable and isotonic at 5, 20 or 25% (w / w) protein concentration in an aqueous medium or at any other therapeutic concentration acceptable for intravenous administration, it is prepared in type II glass ampoules and preferably contains 0.5mM or less, preferably less than 0.037mM citrate concentration in the final albumin composition.