WO2006013360A1

WO2006013360A1 - Freeze-drying apparatus

Info

Publication number: WO2006013360A1
Application number: PCT/GB2005/003041
Authority: WO
Inventors: John Landon
Original assignee: Micropharm Limited
Priority date: 2004-08-03
Filing date: 2005-08-03
Publication date: 2006-02-09
Also published as: GB0417309D0

Abstract

A method of freeze-drying a product comprising the steps of: a) placing a closeable vial (1) containing the product in a container comprising: i) a tray (2); ii) a lid (7); and iii) a membrane (31); wherein the lid (7) and tray (2) co-operate to form a hermetic seal therebetween and the lid (7) can move relative to the tray (2) to effect closure of the closeable vials (1); b) freeze-drying the product; and c) moving the lid (7) and tray (2) relative to one another to effect closure of the closeable vials (1). Containers for use in the method are also provided.

Description

FREEZE-DRYING APPARATUS

Ther present application relates to a method and apparatus for freeze-drying a product under sterile conditions.

Background

Freeze-drying (also known as "lyophilisation") of an aqueous solution of product involves freezing followed by removal of most of the water by direct sublimation from the solid (ice) to the gaseous (water vapour) phase (Murgatroyd 1997a).

Freeze-drying of pharmaceutical products is usually undertaken in vials but may be carried out in ampoules or in bulk. Sterility is essential when freeze-drying medicinal products intended for systemic use in order to avoid bacterial or other contaminants. Similar conditions apply if the product to be freeze-dried must be contained because it is, for example, potentially hazardous. For example, it is known that large numbers of influenza viruses or Escherichia coli can be lost from a vial during freeze-drying as a result of ablation due to particle entrapment in the sublimating water vapour, with severe contamination of the inside of the freeze-dryer and immediate environment (Adams, 1991a).

Conventional freeze-drying is undertaken in a pharmaceutical cleanroom with an HVAC air supply. The room must be kept scrupulously clean and must be assessed regularly as to the particulate and bacterial counts. The chamber of conventional freeze-dryers opens directly into the cleanroom while the bulk of the instrument extends outside to facilitate maintenance and avoid adding to the particle count. The freeze-dryer must be cleaned thoroughly and sterilised prior to use. This is usually undertaken in place with sterilisation being best achieved by the use of steam for several hours. This, in turn, requires a suitable steam generator and a source of sterile water. Filling must be carried out in a grade A environment (as defined in Annex I of E. C. Guide to Good Manufacturing Practice). This is generally achieved by filling within a grade A isolator within a grade C cleanroom. Care in the transfer of the trays containing the product from the isolator to the shelves of the freeze-drying chamber is essential to avoid the risk of contamination. Any risk could be overcome by the use of a grade A pharmaceutical cleanroom but this is seldom available. Thus reliance is usually placed on transferring trays via a laminar flow cabinet or, better, within a sterile, small second isolator that can be fitted directly to the door of the freeze-drying chamber.

Once within the chamber, the product is subjected to a freeze-drying cycle lasting at least 24 hours. This comprises its cooling then freezing (4 to 8 hours) followed by primary (~14 hours) and then secondary (-10 hours) drying under a vacuum of -0.4 torr (Adams, 1991b). Primary drying involves the removal of free moisture from the frozen product by sublimation. In freeze-drying of an aqueous solution, this involves the removal of around 90% of the water molecules present. Secondary drying involves the removal of most of the bound moisture from the frozen product by a combination of desorption and evaporation. In freeze-drying of an aqueous solution, this involves the removal of a further 5 to 8% of the water molecules which are adsorbed to or structurally integrated with non-crystallisable solute. Thus primary and secondary drying involves the rapid migration of around 97% of all the water molecules present in the frozen product.

Meanwhile, following completion of the freeze-drying cycle and removal of the product, parts of the instrument and especially its condenser coils are covered by thick layers of ice. Thus the freeze-dryer must be defrosted, usually by passing large volumes of water through the condenser to waste, then cleaned thoroughly, and finally steam sterilised and cooled prior to reuse. Cleaning and sterilisation must be especially rigorous following the freeze-drying of a potentially hazardous product since contamination of the freeze-dryer is virtually certain. If it is intended to freeze-dry potentially hazardous products as well as conventional products, a second, separate cleanroom and a second freeze-dryer are required.

Prior art containers

A number of freeze-dryer containers exist in the art. One such container, which enables sterile freeze-drying in an unclean environment, is described by Taylor et ah, 1978. The base of the container described in Taylor et al is fitted with two biological filters made from pleated glass fibre which ensure containment and prevent contamination while only slightly impeding vapour flow. Vial closure under vacuum in the freeze-dryer is effected by the use of a vial stopper closure plate, which moves independently of the lid and which is depressed by five plungers. Four spring loaded fasteners secure the lid to the box.

However, the container described in Taylor et al is unduly complex and cumbersome and would be expensive to manufacture in large numbers and in a range of sizes. It is not readily adaptable to different freeze-dryers or different sizes of vial. Further, there are several sites where leakages could occur including the two filters and five plungers.

US patent number 5,309,649 (Boehringer Mannheim) describes a process and container for freeze-drying under sterile conditions. Freeze-drying of a product in bulk was undertaken in a bag, tube or trough comprising part synthetic resin and part a hydrophobic, porous, micro-organism-impermeable, water vapour-permeable membrane. The container described in US 5,309,649 is suitable only for bulk product and not for vials or ampoules, which is a severe disadvantage.

A further example of a container for use in sterile freeze-drying is the LYOGU ARD^® tray (W. L. Gore and Associates, Livingston, UK). The top of the tray is made from GORE- TEX^® expanded PTFE (polytetrafluoroethylene) membrane, which was developed specifically for lyophilisation. The membrane and tray are welded together thermally with the tray having a thin, flexible base which conforms closely to the drying shelf to ensure efficient temperature transfer. A fill port in one corner of the tray enables the introduction of from 500ml to 1800ml of product for freeze-drying in bulk. However, the LYOGU ARD^® tray can only be used once and is expensive. Moreover, it is not suitable for either vials or ampoules. Furthermore, if one wishes to store the freeze-dried product in the LYOGU ARD^® tray for a prolonged period, each tray must be placed inside a foil barrier pouch to prevent degradation of the dried product by ambient moisture or by oxygen or carbon dioxide.

A vacuum box system is also produced by Usifroid, the French Freeze-dryer Company. The Usifroid vacuum box comprises a metal external tray with a gasket around its rim, which can be sealed hermetically by a closely fitting metal lid. Product is placed in an internal tray either in bulk or in ampoules and this is placed in the external tray and the lid put in place with four stoppers in the "open" position. The whole box is then placed in the freeze-drier. After completion of the freeze-drying cycle, while still under vacuum, the stoppers are depressed to their "closed" position by lowering the shelves of the freeze- dryer such that the box is now completely sealed. After release of the vacuum within the freeze-dryer, the box is removed and transferred to a sterile area while its contents are still under vacuum. Only then are the stoppers lifted to break the vacuum, the lid removed and access gained to the product. If ampoules have been used, these are then sealed.

The Usifroid vacuum box is unduly complex. Since initial transfer to the freeze-dryer and the freeze-drying cycles are conducted with the stoppers in the "open" position, neither containment of the product nor prevention from contamination can be ensured - at least from a regulatory standpoint. There is a paucity of groups in Europe able to undertake sterile lyophilisation of small numbers (a few thousand or less) of vials for research purposes or clinical trials. Consequently costs are high, delays common and small groups must often send products abroad for freeze-drying. There are several reasons for this, including technical complexity, with the consequent need for highly experienced staff and rigorous quality control. Among the most important is cost: it takes more than a year and a substantial amount of money to establish, validate and obtain regulatory approval for even a small facility. This is explained by the need to site freeze-drying in pharmaceutical cleanrooms; to segregate freeze-drying of conventional products from those that are potentially hazardous and must be contained; the number of steps involved; and the several expensive items of equipment required, all of which must meet the requirements of Good Manufacturing Practice ("GMP"). The high cost of freeze-drying usually limits its use to high value products such as labile biological materials, especially if sent to or from tropical and/or developing countries where it is difficult to ensure a cold chain.

The present invention provides a method for freeze-drying a product which, i) enables products to be freeze-dried whilst preventing the risk of their contamination by, for example, microorganisms in the external environment; ii) prevents the product from contaminating the freeze-dryer and external environment, which is particularly important where the product contains potentially hazardous products, such as infectious agents; and iii) enables the closure of vials containing lyophilised product whilst still under vacuum and prior to their removal from the freeze-dryer. The invention also provides containers for use in a method according to the invention and components of the container for use in a method according to the invention.

Use of a method according to the present application improves the steps of sterile transfer of the filled vials and loading into a clean, sterile freeze-dryer; sealing under vacuum; sterile unloading of vials from the freeze-dryer and sterilisation of the freeze-dryer. The steps of cooling, freezing and primary drying in the freeze-drying cycle are also preferably improved. Moreover, the use of a method according to the present application overcomes the problems associated with the high costs of prior art methods.

Summary of the invention

Accordingly, the invention provides a method of freeze-drying a product comprising the steps of:

a) placing a closeable vial containing the product in a container comprising:

i) a tray;

ii) a lid; and

iii) a membrane;

wherein the lid and tray co-operate to form a hermetic seal therebetween and the lid can move relative to the tray to effect closure of the closeable vials;

b) freeze-drying the product; and

c) moving the lid and tray relative to one another to effect closure of the closeable vials.

The invention further relates to containers for use in the method recited above. For example, the invention provides a container for freeze-drying a product comprising:

i) a tray;

ii) a lid; and

iii) a membrane; wherein in use the lid and tray cooperate to form a hermetic seal therebetween and the entire lid can move relative to the tray to effect closure under vacuum of the closeable vials within the container.

Alternatively, the invention provides a container for freeze-drying a product comprising:

i) a tray adapted to hold vials; and

ii) a membrane;

wherein the membrane forms a lid and in use the lid and the tray cooperate to form a hermetic seal therebetween and at least a part of the lid can move relative to the tray to effect closure under vacuum of closeable vials within the container.

The invention will be described further by way of example only, with reference to the following figures in which:

Figures Ia and Ib show a partial cross-sectional view of a container containing a vial wherein the container is located in a freeze-drying chamber;

Figures 2a, 2b and 2c show partial cross-sectional views of configurations of seal;

Figure 3a shows a plan view from above of a lid having holes; Figure 3b shows a plan view from below of a lid wherein a membrane covers the holes; and

Figure 4a shows a partial cross-sectional view of a container having a membrane for a lid; Figure 4b shows a plan view of a clamp for forming a seal with the container of Figure 4a; and Figure 4c shows a partial cross-sectional view of the depression in the tray into which the clamp of Figure 4b may fit.

Detailed description of the invention

The invention provides a method of freeze-drying a product comprising the steps of: a) placing a closeable vial containing the product in a container comprising:

i) a tray;

ii) a lid; and

iii) a membrane;

wherein the lid and tray co-operate to form a hermetic seal therebetween and the lid can move relative to the tray to effect closure of the closeable vial;

b) freeze-drying the product; and

c) moving the lid and tray relative to one another to effect closure of the closeable vial.

A method according to the present invention preferably provides a dry (1-3% water), shelf- stable, active, readily soluble product of acceptable appearance.

One or usually more vials may be contained in the container used in the method of the present invention.

The closeable vials are closed whilst under vacuum by movement of the lid relative to the tray. Preferably the lid moves relative to the tray in a direction perpendicular to the openings, i.e., parallel to the axis of the openings of the one or more vials, which leads to the closure of the vials. Preferably, the entire lid moves relative to the tray to effect closure of the vials. The method does not encompass the use of a container wherein closure of the vial is effected by a plunger mechanism which moves independently of the lid.

The lid should be able to move relative to the tray by a sufficient distance to permit closure of the or each closeable vial located in the tray. Preferably, the lid is able to move by at least 10 mm relative to the tray. More preferably, the lid is able to move by between 10 and 15 mm relative to the tray. Preferably, the lid is moved relative to the tray by compression between two shelves in the freeze-drying chamber thereby closing the one or more vials whilst under vacuum. For example, Figures Ia and Ib show a vial (1) in a tray (2) located in a freeze-drying chamber having an upper shelf (4) and a lower shelf (11). Movement of the upper shelf (4) in the freeze-drying chamber towards the lower shelf (11) in a direction perpendicular to the opening (6) of the vial (1) results in movement of the lid (7), which acts on the stopper (8) thus closing the vial. The hermetic seal formed by the seal (12) acting between the lid (7) and the tray (2) is maintained throughout this movement.

Preferably, closure of the vial is achieved by depressing a stopper (8) in the vial. Preferably, the stopper (8) is a ventilated stopper having an "open" position (illustrated in Figure Ia) and a "closed" position (illustrated in Figure Ib) and movement of the lid (7) relative to the tray (2) depresses the stopper (8) into the vial (1) from the "open" position to the "closed" position. In the "open" position, the stopper is partly inserted in the opening (6) of the vial (1) but allows the passage of water vapour out through the stopper (8). However, in the "closed" position, the stopper hermetically seals the vial. The stoppers are preferably made from rubber but may alternatively be made from any other material which is suitable for hermetically sealing the individual vials and which can endure the conditions required for sterilisation and freeze-drying.

The stoppers are preferably brought into contact with the lid as the lid is depressed towards the opening of each vial. Alternatively, the lid may contact the stopper via an adapter member which may be associated with the stopper or the lid and which may be integral thereto. The adapter may alternatively be located in the container between the stopper and lid but not in contact with either until the shelves of the freeze-drying apparatus are compressed. Use of a method of the invention avoids the risk that the ventilated stoppers freeze to the upper shelf during closure and are then pulled out of the vials when the shelves are separated again, as occurs occasionally during conventional freeze-drying.

After closure of the vial in the freeze-dryer under vacuum, the vacuum can be released, the container unloaded and the vials oversealed in a non-sterile area.

The closure of the closeable vial containing the lyophilised product whilst still under vacuum and prior to removal of the vial from the freeze-dryer prevents the risk of contamination of the product by, for example, microorganisms in the external environment and prevents the product from contaminating the freeze-dryer and external environment. The ability to contain the product is particularly important where the product contains potentially hazardous products, such as infectious agents. Therefore, the use of a method of the present invention means that it is not necessary to release the vacuum in a sterile area. However, as the outside of the vials may be contaminated, the vacuum should preferably be released in a containment area.

As mentioned above, prior to closure of the vials, the product in the vials should be kept in a sterile environment and thus it is important that environmental contaminants cannot enter the container. It is also important that any potentially hazardous products in the container do not escape into the external environment or contaminate the freeze-dryer. However, water vapour that escapes from the product during the freeze-drying cycle must still be allowed to leave the container. A method according to the present invention enables these aims to be achieved by the use of a membrane in combination with the presence of a hermetic seal between the lid and the tray. A membrane for use in a container according to the present application is permeable to water vapour and atmospheric gases in order to allow vapour molecules to exit the container through the membrane.

Preferably, the membrane should also be impermeable to bacteria, viruses and other microorganisms that may be present in the external environment. This ensures that contaminants from the external environment do not enter the container and contaminate the product. However, as the membrane is permeable to water vapour and atmospheric gases, the vapour molecules are still able to exit the container through the membrane. More preferably, the membrane should also be impermeable to other agents that may be present in the external environment, but for practical purposes, the most important environmental contaminants of relevance are bacteria.

Examples of the range of non-hazardous products that may be freeze-dried and which it is desirable to protect from contamination from the external environment include killed microorganisms as vaccines; human blood products such as clotting factors (e.g., factor VIII, etc); animal blood products such as polyclonal antibodies and their fragments (e.g., antivenoms, anti drugs such as Digibind™, etc.); monoclonal antibodies; enzymes such as L-asparaginase and catalase; and labile molecules such as vitamins and hormones.

In particularly preferred embodiments, a membrane for use in a method according to the present invention is impermeable to potentially hazardous molecules, particularly those that may be present in the product and which it is desirable to contain within the container. The range of potentially hazardous products is numerous and diverse. They include infectious agents (bacteria, viruses and fungi), for example, which may be present in attenuated vaccines, in blood, plasma or serum or in other biological samples such as bone or cornea. They may also include potentially sensitising molecules such as penicillin and other antibiotics, and highly toxic compounds such as the tetrameric enzyme L- asparaginase, cytotoxic drags such as cysplatin and botulinum and other toxins. A further example of potentially hazardous products are live microorganisms for seed culture (inocula), for example, bacteria, viruses or yeasts.

In most preferred embodiments, a membrane for use in a method according to the present invention is impermeable to bacteria, viruses and other microorganisms that may be present in the external environment and is impermeable to potentially hazardous molecules whilst being permeable to water vapour and atmospheric gases.

In relative terms to water and atmospheric gases, bacteria are enormous and are excluded

by any material with a pore size of 0.2 μm or less. Thus, the membrane preferably has a

pore size of 0.2 μm or less. If other potential contaminants are present in the external

environment, such as viruses, the pore size of the membrane is adapted to take this into account. Further, in embodiments in which the product being freeze-dried contains potentially hazardous molecules that are smaller than bacteria, it will be necessary to use a membrane with a pore size that, will prevent these smaller products from exiting the container.

A membrane for use in a method according to the present invention will also preferably be sterilisable and/or have low extractables and particulates and/or be compatible with the product. Preferably, the membrane will be reusable such that it is not damaged by repeated freeze-drying cycles/sterilisation but this is not essential provided that the membrane is inexpensive to manufacture and easy to obtain. Advantageously, a membrane for use in a container according to the present application has all of these characteristics. Thus a method according to the present application may make use of different membranes according to the nature of the product being freeze-dried and/or according to the environment in which the freeze-drying is taking place.

Materials which are suitable for use as a membrane include semi-permeable paper such as pleated glass fibre paper (e.g., No. 6, Mk 2 dust filter available from Leyland and

Birmingham Rubber Co., Leyland, Preston, Lanes), cellulose and usual cellulose derivatives such as cellulose acetate; films of polymer compounds such as polypropylene or PTFE (polytetrafluoroethylene); films of sterilisation paper according to German

Industrial Standard DDSf 58 953; and Goretex and similar membranes. However, a membrane for use in a method according to the invention may be made of other materials provided that it has the characteristics listed above that are necessary for its use.

A method of the invention involves using a container comprising a membrane which is exposed to both the inside and the outside of the container so that water vapour and atmospheric gases can escape from the container. The membrane may be situated in one or more of the sides of the container, in the base of the container and/or in the lid of the container. Preferably, the membrane is situated in the lid. The amount of membrane that is exposed to both the inside and the outside of the container is a sufficient amount so as not to significantly impede the flow of water vapour. For example, the amount of membrane that is exposed may make up at least 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 10%, 15% or 20% of the surface area of the container.

The ability of a container according to the present invention to prevent contamination and ensure containment when freeze-drying a product, depends upon maintaining an hermetic seal between the tray and its lid during the transfer of the container from the sterile filling unit to the freeze-dryer and during the freeze-drying cycle until closure of the vials whilst still under vacuum. The presence of the hermetic seal between the lid and the tray ensures that no air can penetrate the seal. The term "hermetic" as used herein means "airtight". The hermetic seal between the lid and the tray also ensures containment of the product and prevents contamination of the product.

Freeze-drying will always be a relatively complex procedure and will require highly experienced staff. However, use of a method according to the present application will simplify matters significantly. It is difficult to incorporate a large item of equipment, such as a freeze-dryer, into a cleanroom. Working within pharmaceutical cleanrooms also involves gowning up and constant checks as to the integrity of the air supply and freedom from particulates and environment contaminants. Working within the confined space of an isolator sited in a cleanroom adds to the problems. Thus working in an isolator in a cleanroom is uncomfortable and the need to wear gloves impairs performance. It follows that, ideally, as few steps as possible in freeze-drying should need a sterile environment.

Currently, any group freeze-drying conventional and hazardous products requires a separate cleanroom and freeze-dryer dedicated to each. It is mandatory to prevent a hazardous material from contaminating the freeze-dryer and immediate environment in which a medicinal product for systemic use will be freeze-dried subsequently. When pathogens or hazardous products have to be freeze-dried it is unwise to rely on the primary container (that is the vial and stopper) or the medium formulation as the sole mechanism for preventing contamination of the inside of the freeze-dryer.

There is no way of avoiding the need for sterile facilities while filling vials, ampoules or in bulk. However, the use of a membrane and hermetic seal, as required by the present method, enables hazardous products to be contained and protects products in the container against contamination and thus removes the need for use of a separate cleanroom and freeze-dryer. Thus a single freeze-drying facility will suffice for both and the use of a method according to the present invention is of especial benefit to groups who freeze-dry both conventional and potentially hazardous products.

Further, the use of a method according to the present invention allows the transfer of the container to or from the freeze-dryer to be undertaken through the normal environment without the need for a transfer isolator or laminar flow cabinet. Once freeze-dried, the vials can be sealed while still under vacuum and thus the container need not be opened in a sterile facility when vials are used. However, if the vials contain hazardous product, the container should be opened in a containment area since the outside of the vials will be contaminated. The container is then preferably decontaminated with, for example, a weak solution of formaldehyde immediately upon opening.

Thus use of a method according to the invention for freeze-drying a product in vials enables all steps other than filling to be carried out in an open laboratory even when hazardous products are involved. It greatly simplifies matters that the freeze-dryer can be sited in a general purpose, non-sterile laboratory.

Although closure under vacuum is not possible when freeze-drying in ampoules or in bulk, the presence of the membrane and the hermetic seal will ensure protection of the freeze- dried product while the container is being transferred from the freeze-dryer back to a sterile environment prior to opening. Thus, when the product is in bulk or in ampoules, it is necessary to open the container within sterile facilities. Ampoules are then closed by heat sealing. However, bulk products or products in open ampoules can be stored in a general purpose area after freeze-drying provided they are kept in the container according to the present invention. However, the duration of their storage should be kept to a minimum since moisture and atmospheric gases can reach the lyophilised product and cause its deterioration.

A significant proportion of the freeze-drying cycle time in conventional freeze-drying is concerned with cleaning and sterilising the plant and freezing the product. As it is not necessary to sterilise the freeze-dryer before or after use, nor to provide a steam generator and a system for providing copious volumes of sterile water as in conventional freeze- drying techniques, the use of a method according to the invention minimises operating time for freeze-drying leading to enhanced utilisation and decreased costs. For example, not only does it save several hours of steam sterilisation time but also the prolonged period (~6 hours) required for the instrument to cool down. Further, avoidance of steam sterilisation also means that the freeze-drying chamber does not have to withstand temperatures of -127⁰C and overcomes certain health and safety issues such as the need for a door locking device to prevent the chamber being opened while still under positive pressure. Thus a less expensive instrument may suffice.

Use of a method according to the invention enables the primary and secondary drying times to be similar to conventional primary and secondary drying times that are used when the vials are placed directly into the freeze-dryer rather than inside a container.

Freezing in a conventional freeze-dryer is inefficient and slow as it is dependent largely on conduction via several layers that seldom make perfect contact. Conduction proceeds through the metal of the cooling shelves, the base of the tray, the glass bottom of the vial and then up through the product starting at its base. Further, for a significant period of time, the pharmaceutical freeze-dryer is utilised only as a freezing cabinet, thereby reducing the economic effectiveness of the plant. Pharmaceutical products dispensed into vials, as may be used in a method according to the present invention, can be prefrozen in a cold chamber, which has the advantage of maximising throughput. However, problems associated with maintaining product cleanliness have previously limited the appeal of external prefreezing procedures. The use of a method according to the present invention overcomes such problems by allowing freezing to be undertaken in a separate refrigeration unit if desired. Thus, being more efficient, the freezing time is greatly reduced. For example, the rate of nucleation (on which ice formation depends) in pure water at -3O⁰C is only 15 X lO¹ nuclei-second.m³ compared with 4.3 x 10¹⁵ nuclei-second.m³ at -4O⁰C.

In addition to shortening freezing time, a separate unit allows freezing to occur in a way which also shortens the time required to complete primary drying (sublimation). As mentioned above, in a plate freezer, the heat transfer is through the bottom of the product with the ice front rising and sweeping the water-soluble components towards the top of the containing vial in the form of a plug of gradually increasing concentration. This can form a 'skin' near the surface which is poorly permeable to water vapour and, therefore, markedly impairs primary drying. Thus, even under ideal conditions, more than 60% of the impedance to vapour flow during drying is due to the product itself and, especially, the dry layer on its surface. This contrasts with, for example, the freezing of a pond which involves convection and radiation as well as conduction and starts at the surface. Use of a separate refrigeration unit would also facilitate a technique known as heat annealing (Adams, 1991(b)) which is designed to produce a more open ice matrix and cause some solutes to crystallise that, otherwise, do not when using standard freeze-drying techniques. A typical heat annealing cycle involves freezing the product to -4O⁰C, then warming to - 1O⁰C before recooling to -4O⁰C.

Further, the temperature in a separate refrigeration unit is easier to control, maintain and record, which provides a significant regulatory advantage. Thus, the method according to the present invention may additionally employ the step of pre-freezing the product. Pre-freezing may be accomplished using a separate refrigeration unit, which is designed to ensure controlled, rapid freezing.

A simpler freeze-dryer having shelves which do not need to reach such low temperatures as those of conventional freeze-dryers, where there will be no requirement for steam sterilisation and/or where the product will not be subjected to positive pressures, may also be used in a method according to the present invention.

Thus the use of a method according to the present invention offers a number of significant advantages over the use of methods available in the prior art. It reduces the time taken to complete the freeze-drying of a product and, thereby, increases throughput and further reduces costs. It also enables a reduction in the cost of the facilities and equipment needed to undertake conventional, small scale sterile freeze-drying in vials. Moreover, it simplifies the process.

Containers

The invention also encompasses containers for use in a method according to the invention and components of the containers. A container according to the present application is suitable for holding one or more vials containing the product. However, the container may alternatively or additionally be used to hold product in bulk or in ampoules.

In one embodiment, a container for freeze-drying a product comprises:

i) a tray;

ii) a lid; and

It is important that the hermetic seal is maintained during the movement of the lid relative to the tray. Examples of preferred configurations of seal that allow such movement of the lid are illustrated in Figure 2. These can be likened, in many respects, to the seal that operates between the barrel and plunger of a syringe and are described below:

(i) Figure 2a shows a seal (gasket) (20) clamped over the entire upper ridge (21) of the tray (2). Such a seal is easy to fit onto any tray without modifying the tray itself and enables the seal to remain airtight whether the lid is to be depressed within or outside the tray;

(ii) Figure 2b shows a seal (22) placed around the upper and outer side (23) of the tray (2). For example, this may be somewhat similar to a thick rubber band. Preferably, a groove (26) or ridge on the tray's outer surface is present to keep the seal secure when the lid (7) is depressed; or

(iii) Figure 2c shows a seal (24) placed around the outer side of the lid (7). Preferably, a groove (27) or ridge on the lid's outer surface is present to keep the seal secure when the lid is depressed. This type of configuration has the advantage that the tray need not be modified. However, under this option, the lid (7) moves down inside the tray (2) and thus a rim (25) is preferably also present around the periphery of the inside of the tray (2) to prevent the lid 'snagging' the most peripheral vials (or ampoules). The rim may be a separate component or may form an integral part of the tray.

Generally, the seal will be made of rubber or other suitable elastic material. The exact type of seal used will depend upon the conditions used in the freeze-drying cycle and the sterilisation procedure. For example, if the seal needs to cope with temperatures as low as -5O⁰C and steam or some other form of sterilisation, suitable material that can withstand these temperatures should be chosen for the seal.

The membrane is preferably located in the lid. Alternatively, the membrane may be located elsewhere in the container, such as in the base of the container and/or in one or more of the walls of the container. Further, the container may comprise a membrane in one, two or more different locations, for example, in the lid, in the lid and base or in the lid, walls and base. The membrane may form part or all of one or more of the lid, base and/or walls. Where the membrane forms all of the lid, the membrane is preferably sufficiently rigid to enable vial closure to be effected upon compression between the shelves of the freeze-dryer.

The membrane preferably covers one or more holes in the container which allow the free passage of water vapour and atmospheric gases out of the container. The hole(s) may be located in the lid or the tray (e.g., in the top, side walls or base of the container). The number, size and spatial arrangement of the one or more holes should allow a sufficient area for water vapour to escape from the container. Preferably, the arrangement of the one or more holes also enables an equal exposure of each vial to the one or more exit holes.

Preferably, the lid comprises one or more holes covered by the membrane. For example, Figure 3 a shows a lid (7) as viewed from the outside of the container, which comprises holes (30). Figure 3b represents a view of the lid (7) from the inside of the container and shows that the holes (30) are covered by a membrane (31), which is attached to the inner surface of the lid (7). In this embodiment, the number, size and spatial arrangement of the one or more holes should also ensure the strength and rigidity of the lid essential for the closure mechanism. Where the product is contained in vials, these one or more holes are preferably significantly smaller than the size of stoppers used to close the vials to ensure that the stoppers themselves do not pass through the one or more holes. For example, a lid of typical size 255 x 355 mm may comprise around 120 holes each having a diameter of about 10mm. For example, the lid may comprise one 10mm diameter hole per 750 mm .

Preferably, the membrane is present on the internal surface of the container, for example the internal surface of the lid. The membrane is preferably attached to the container by heat welding. It is preferable to avoid the use of adhesive for regulatory reasons. Welding the membrane to the internal surface of the container improves the appearance of the container and ensures close contact of the two during the freeze-drying cycle and especially during primary and secondary drying when the chamber is under vacuum.

In an alternative embodiment, the invention provides a container for freeze-drying a product comprising:

i) a tray adapted to hold vials; and

ii) a membrane;

wherein the membrane forms a lid and in use the lid and the tray cooperate to form a hermetic seal therebetween and at least a part of the lid can move relative to the tray to effect closure under vacuum of the closeable vials within the container.

Thus, in this alternative embodiment the membrane itself forms the lid. The membrane is preferably flexible and thus movement of the lid relative to the tray is possible by depressing part of the membrane, thereby effecting closure of the vials. Alternatively, if a rigid membrane is used, the entire membrane lid can move relative to the tray. Thus this alternative embodiment preferably encompasses the use of membrane lids that are sufficiently flexible to enable the vials to be closed fully whilst still in the freeze-dryer under vacuum by depressing part of the membrane but also encompasses the use of rigid membrane lids that can move in their entirety relative to the tray, thereby effecting closure of the vials whilst under vacuum.

The membrane for use in this alternative embodiment forms an airtight seal with the tray. The membrane is also preferably sufficiently strong to prevent breakage of the membrane during transport of the container to and from the freeze-drying apparatus and during the effecting of closure of the vials. The membrane should also preferably be available in sheets.

Preferably, the vials are filled, partially stoppered in the open position and loaded into the tray and then the sheet of membrane is placed lightly over the top of the vials.

Where a flexible membrane is used, a hermetic seal is preferably formed between the lid and the tray by folding the membrane down over the sides of the tray and fixing it there in such a way as to ensure an airtight seal between the tray and the membrane. For example, the membrane can be sealed hermetically to the sides of the tray by means of an elastic ring. Because it is not necessary in this alternative embodiment where a flexible membrane is used for the entire lid to move relative to the tray, a clamp (e.g., made from metal) could be used which, if required, could fit into a depression around the tray. Such a clamp will preferably be resistant to repeated freeze-drying cycles, easy to sterilise and suitable for repeated use.

Figure 4a shows an example of a container (40) according to this embodiment comprising a tray (2) containing vials (1) having stoppers (8) in their "open" position. The membrane

(41) is placed over the tops of the stoppers and forms the lid of the container. A clamp

(42) is used to effect an airtight seal between the membrane (41) and the sides of the tray (2). The clamp of Figure 4a is shown in more detail in Figure 4b and a method of fixing the clamp to the tray is illustrated further in Figure 4c. The inside dimensions of the clamp (42) are slightly smaller than the outer dimensions of the tray (2) such that the clamp (42) fits within a depression (43) in the tray (2). The clamp (42) is hinged at hinges (44) such that it can be opened to allow the clamp (42) to be placed around the tray (2) in the depression (43). Once the clamp (42) is in position, a clip (45) fastens the opened ends of the clamp (46) together to hold the clamp (42) in place.

Where a rigid membrane is used, a seal that allows movement of the entire lid relative to the tray, for example, as described previously and shown in Figure 2, should be used.

After establishing a hermetic seal between the lid and the tray, the tray can then be transferred safely via a non-sterile environment into a non-sterilised freeze-dryer. Where a flexible membrane is used, the stoppers and tops of the vials preferably extend above the sides of the tray so that the stoppers can be inserted fully into the vials by compression between shelves of the freeze-drying chamber following freeze-drying while still under vacuum.

A container for use in a method according to the present application may additionally comprise terminals to allow thermocouples to record temperature changes of the product in one of the vials. Preferably, these thermocouples are present in the lid of the container.

The lid may also be transparent to permit viewing of the product, preferably of the product in the vials, through the chamber door during the freeze-drying cycle and their closure. This requires that the membrane, when it forms part of the lid, is transparent, or very small in area as compared with the lid.

A tray for use with a container according to the present invention is preferably constructed from a strong, light material that conducts temperature changes rapidly from the freeze- dryer shelves to the product being freeze-dried. The tray also preferably has a flat smooth base to ensure maximum contact with the cooling shelves. The tray may be made from any material with suitable mechanical and thermal conductivity properties, e.g., steel. Alternatively, the tray may be constructed of aluminium or of various plastics. Aluminium has ideal thermal properties. However, it is not as strong mechanically as stainless steel and warps and distorts when heat sterilised. It also corrodes when in contact with saline and products may stick unless the aluminium has been Teflon coated. Thus steel is preferred despite its poorer thermal qualities. Most preferably, stainless steel such as pharmaceutical grade, polished stainless steel is used, as specified by GMP. However, plastic trays may also be used despite having inferior thermal conduction properties to metal trays as they may be much less expensive to produce.

The dimensions of a tray of a container according to the present application may vary, since vials and ampoules come in many different sizes and since the surface area of the freeze-dryer shelves and the distance between them may vary markedly, sometimes even within a single company. Clearly, the dimensions of a container according to the present application may vary depending on the freeze-dryer that it is for use with. The size of the trays will be dictated by their ease of handling and weight when fully loaded. Thus the tray is available in a wide range of sizes, both with respect to height and surface area, to fit all existing freeze-dryers and to be suitable for all sizes of vials and ampoules.

After filling with the required volume of product, the vials (or ampoules) are usually carefully loaded into trays for transfer to the freeze-dryer. When a tray has been filled completely with vials or ampoules there is no danger that any will fall during transfer.

However, there is a real risk of falling when only a few are placed on the tray. Thus the tray may additionally comprise a barrier placed around the vials or ampoules. The barrier may form a wall around the periphery of the one or more vials or ampoules contained in the tray. Individual barriers may be used for individual vials or ampoules or groups of vials or ampoules. Alternatively, the barrier may comprise a rack comprising compartments each capable of holding one or more, preferably one, vial. Thus the barrier can be of varying size and/or shape to cope with different numbers of vials or ampoules and, thereby, stabilise and locate them as desired. The barrier is preferably made from metal or plastic. The barrier may be a separate component or may form an integral part of the tray.

The use of a barrier as described above will prevent the vials or ampoules from falling during transfer. The barrier may also be used to alter the position of the vials or ampoules in relation to the membrane. Preferably, the barrier allows each vial to be positioned in equal juxtaposition to the membrane.

The components of a container according to the present application comprise at least a lid and a tray, as the membrane may form the lid. The lid and the tray for use with a container according to the present invention each individually form a separate aspect of the invention. Thus the application also relates to lids as described herein for use with a container according to the present application and trays as described herein for use with a container according to the present application.

The components of a container according to the present application should be sufficiently strong for their purpose. They are preferably also simple and relatively inexpensive to manufacture. Preferably, the components of a container according to the present application have a smooth surface to facilitate cleaning. Preferably, the components of a container of the present application are reusable. In order to be reusable, the components must be resistant to repeated freeze-drying and to autoclaving and/or sterilisation with steam (or, possibly, hydrogen peroxide) without bending, bubbling or other damage. Since some products may be freeze-dried in bulk within the container, its constituents also preferably have low extractables and particulates, are compatible with the products and are available in a wide range of sizes.

Example

An exemplary operation of sterile freeze-drying using the container of the present application is described. A pharmaceutical cleanroom with HVAC air supply is used to house the grade A isolator used for sterile filling. As with conventional freeze-drying, the cleanroom is scrupulously cleaned and assessed and the isolator is sterilised with hydrogen peroxide prior to use. The freeze-dryer need not open into the cleanroom and can be sited in any clean non-sterile area, such as a general purpose laboratory. Although it must be cleaned thoroughly before use, it never needs to be sterilised, thereby avoiding the need for a steam generator and a source of copious volumes of sterile water.

The vials are generally filled within a purpose designed grade A isolator sited in a grade C/D cleanroom. The isolator and its contents are sterilised using a hydrogen peroxide

generator. The product for lyophilisation is pumped into the isolator via a 0.2 μm filter to

ensure its sterility. Also placed in the isolator are vials (after being washed and depyrogenated in an oven); sterilised trays into which the vials will be placed; and the closure system, including ventilated rubber stoppers (after being washed to remove particulates and then sterilised in an autoclave or by gamma irradiation).

After delivering accurately into each vial the required volume of product, the vials are placed in the trays and the rubber stoppers introduced by hand in their "open" position (Figure Ia). The lids are placed carefully over the trays and lowered gently until just in contact with the ventilated stoppers. At this time, the hermetic seal between the lid and the tray is now present and thus the contents of the vials are fully protected against environmental contamination and any potentially hazardous products are contained within the container.

Since the contents of partially stoppered vials (or of ampoules or in bulk where used) are now totally safe within the container of the present invention, the container can be removed from the isolator and cleanroom into a non-sterile environment. Normally, they will be transferred directly to the freeze-dryer which, itself, is in a non-sterile site. Alternatively, the container can be stored for prolonged periods prior to freeze-drying, especially if kept at -2O⁰C. This also makes possible the initial freezing stage of the freeze- drying cycle in a separate refrigeration unit.

Once the container has been placed on the shelves of the freeze-drying chamber, the cycle is similar to that of conventional freeze-drying since the membrane does not impede vapour flow.

Once completed and while still under vacuum, the vials are closed by lowering the upper shelves which, in turn, press down the lid to depress fully the rubber ventilated stoppers into the vials (see Figure Ib). Closure while still under vacuum avoids the freeze-dried product being exposed to the moisture, oxygen and carbon dioxide in the atmosphere, all of which may impair its activity. It also ensures the protection of the product within the vial from contaminants in the environment and of the environment against potentially hazardous products in the vial.

As with conventional freeze-drying, after releasing the vacuum, the container with freeze- dried product in vials is removed to a non-sterile setting for their initial assessment and final oversealing of the vials with a crimped metal ring.

After final stoppering/sealing the vials/ampoules are subject to 100% integrity testing, various microbiological procedures (pyrogenicity, sterility and bioburden), measurement of their moisture content and activity and determination of their reconstitution time and the clarity of the reconstituted product. They are then labelled and stored prior to distribution and use.

Following completion of the freeze-drying cycle, the freeze-dryer is defrosted and cleaned thoroughly. It does not need to be sterilised even when used to freeze-dry potentially hazardous products. However, in the latter instance, the inside of the container and the outside of the vials will generally have been contaminated. Thus the container should be opened only in a suitable containment area and subjected immediately to sterilisation with, for example, alcohol or formalin.

It will be understood that the invention has been described above by way of example only and that further embodiments and alternatives are included within the scope of the application.

References:

Murgatroyd, K., 'The freeze-drying process', Good Pharmaceutical Freeze-drying Practice, Ed. P. Cameron, Interpharm Press Inc., Buffalo Grove, IL, USA₅ Chapter 1, pages 1-58,

1997(a);

Murgatroyd, K., 'The freeze-dryer and freeze-dryer design', Good Pharmaceutical Freeze- drying Practice, Chapter 2, pages 59-124, 1997b;

Adams, G.D.J., 'The loss of substrate from a vial during freeze-drying using Escherichia coli as a trace organism', J. Chem. Tech. Biotech., 52: 511-518, 1991(a);

Adams, G.D.J., 'Freeze-frying of biological materials', Drying Technology, 9:891-925, 1991 (b);

Taylor, R., 'Sterile freeze drying in an unclean environment', J. appl. Chem. Biotechnol., 1978, 28, 213-216;

E.C. Guide to Good Manufacturing Practice, published 1998, see 'Manufacture of Sterile Medicinal Products' Annex I.

Claims

1. A method of freeze-drying a product comprising the steps of:

a) placing a closeable vial containing the product in a container comprising:

i) a tray;

ii) a lid; and

iii) a membrane;

b) freeze-drying the product;

2. A container for freeze-drying a product comprising:

i) a tray;

ii) a lid; and

iii) a membrane;

wherein in use the lid and tray cooperate to form a hermetic seal therebetween and the entire lid can move relative to the tray to effect closure under vacuum of the closeable vials within the container.

3. A container according to claim 2, wherein the hermetic seal is formed by a seal over the entire upper ridge of the tray.

4. A container according to claim 2, wherein the seal is placed around the upper and outer sides of the tray.

5. A container according to claim 4, wherein a groove or ridge on the tray's outer surface is present to keep the seal secure when the lid is depressed.

6. A container according to claim 2, wherein the seal is placed around the outer side of the lid.

7. A container according to claim 6, wherein a groove or ridge on the lid's outer surface is present to keep the seal secure when the lid is depressed.

8. A container according to claim 6 or claim 7, wherein a rim around the periphery of the inside of the tray is present to prevent the lid 'snagging' the most peripheral vials.

9. A container according to any one of claims 2 to 8, wherein the container comprises one or more holes covered by the membrane.

10. A container according to claim 9, wherein the one or more holes are in the lid.

11. A container for freeze-drying a product comprising:

i) a tray adapted to hold vials; and

ii) a membrane;

12. A container according to claim 11, wherein the membrane is flexible and movement of the lid relative to the tray is effected by depressing part of the membrane, thereby effecting closure of the vials.

13. A container according to claim 11 or claim 12, wherein the seal is formed between the lid and the tray by folding the membrane down over the sides of the tray and fixing it there to ensure an airtight seal between the tray and the membrane.

14. A container according to claim 13, wherein the seal is formed using an elastic ring.

15. A container according to claim 13, wherein the seal is formed using a clamp.

16. A container according to claim 15, wherein the clamp fits into a depression around the tray.

17. A container according to any one of claims 11 to 16, wherein the stoppers and tops of the vials extend above the sides of the tray.

18. A container according to any of claims 2-9 or 11, wherein the lid is a rigid membrane and the entire Kd can move relative to the tray to effect closure under vacuum of closeable vials within the container.

19. A tray for use with a container according to any one of claims 2 to 18.

20. A lid for use with a container according to any one of claims 2 to 18.