GB2640885A - Reactor apparatus - Google Patents

Abstract

A reactor apparatus comprising a reactor vessel 32, media conditioning circuit 34 and a fluid flow controller 36 wherein the fluid flow controller allows the flow of fluid through the reactor vessel to be reversed. The external fermentation broth loop has conditioning modules 50 52 that may alter pH, temperature, gases, or remove waste products. The fluid flow controller may reverse the flow automatically at intermittent periods, and may be a pump or valve. In an alternative configuration, there is a media and cell separation device 42 44 that retains cells inside the reactor vessel, allowing circulation of reaction media between the bioreactor and media loop. The cells may be separated from the media through a filter or membrane, or mechanically separated by gravity or centrifugation. Additionally, there may be several reactor vessels that are connected in a cascading (430, Fig 5) or parallel (330, Fig 5) arrangement. A method of operating the reactor apparatus is also disclosed.

Description

REACTOR APPARATUS

This invention relates to reactor apparatuses and methods of operating a reactor apparatus, particularly in relation to bioreactor apparatuses.

The stirred tank bioreactor is the most commonly used and available type of bioreactor for research and industrial biomanufacturing. It consists of a single complex vessel that contains cells suspended in a liquid media, with integrated apparatuses for conditioning of the media (including temperature, dissolved oxygen, pH control and sugar/nutrient replenishment). Figure 1 shows an example of a stirred tank bioreactor with a media replenishment device 12, a pH balancing device 14, a sensing device 16, an air sparger device 18, a mixing device 20 and a cooling/heating jacket 22.

According to a first aspect of the invention, there is provided a reactor apparatus comprising: a reactor vessel (e.g. a culture vessel); a media conditioning circuit in fluid communication with the reactor vessel so as to permit circulation of reaction media (e.g. culture media) between the media conditioning circuit and the reactor vessel, wherein the media conditioning circuit includes at least one media conditioning module configured to, in use, condition the reaction media within the media conditioning circuit; and a fluid flow controller configured to, in use, selectively reverse a flow direction of the reaction media through the reactor vessel.

In embodiments of the invention, the fluid flow controller may be configured to, in use, automatically and selectively reverse the flow direction of the reaction media through the reactor vessel. Preferably the reversal of the flow direction of the reaction media through the reactor vessel is timed. For example, the fluid flow controller may be configured to, in use, automatically and selectively reverse the flow direction of the reaction media through the reactor vessel intermittently or at regular intervals.

The reversal of the flow direction of the reaction media through the reactor vessel may be carried out using various types of fluid flow controllers. For example, the fluid flow controller may include at least one pump (e.g. mechanical pump, such as a peristaltic pump) and/or at least one valve (e.g. solenoid valve) configured to, in use, selectively reverse the flow direction of the reaction media through the reactor vessel.

In a preferred embodiment of the invention, the media conditioning circuit and fluid flow controller may be arranged to, in use, maintain a same flow direction of the reaction media through the media conditioning circuit in different flow directions of the reaction media through the reactor vessel. This means that the flow direction of the reaction media through the media conditioning circuit will stay the same regardless of the chosen flow direction of the reaction media through the reactor vessel.

Preferably the media conditioning circuit comprises the fluid flow controller. In alternative embodiments, it is envisaged that the fluid flow controller may be separate from the media conditioning circuit.

In further embodiments of the invention, the reactor apparatus may include at least one media and cell separation device configured to, in use, retain cells inside the reactor vessel and permit circulation of the reaction media between the media conditioning circuit and the reactor vessel.

According to a second aspect of the invention, there is provided a reactor apparatus comprising: a reactor vessel; a media conditioning circuit in fluid communication with the reactor vessel so as to permit circulation of reaction media between the media conditioning circuit and the reactor vessel, wherein the media conditioning circuit includes at least one media conditioning module configured to, in use, condition the reaction media within the media conditioning circuit; and at least one media and cell separation device configured to, in use, retain cells inside the reactor vessel and permit circulation of the reaction media between the media conditioning circuit and the reactor vessel.

In embodiments of the invention, the reactor vessel may comprise the at least one media and cell separation device. The at least one media and cell separation device may include a selectively permeable barrier, such as a filter (such as a stainless steel filter and/or a microfilter) and/or a membrane. Alternatively or additionally, the at least one media and cell separation device may include a mechanical separation device configured to, in use, mechanically separate the cells from the reaction media. Such a mechanical separation device may be configured to, in use, mechanically separate the cells from the reaction media by, for example, centrifugal separation, gravity separation or acoustic separation.

The media conditioning circuit may include a single media conditioning module. Alternatively, the media conditioning circuit may include a plurality of media conditioning modules, each of which is configured to, in use, condition the reaction media. It was not essential for the media conditioning modules to simultaneously condition the reaction media. For example, the media conditioning modules may be arranged to, in use, sequentially and/or independently condition the reaction media. This may be achieved by distributing the media conditioning modules along the media conditioning circuit to, in use, sequentially and independently condition the reaction media. In other examples, a plurality of media conditioning modules may be arranged to, in use, condition respective parallel flows of the reaction media. This may be achieved by configuring the media conditioning circuit to provide parallel flow paths of the reaction media, each of the plurality of media conditioning modules arranged along a respective one of the parallel flow paths that, in use, carries a portion of the reaction media.

The number and type of media conditioning modules may vary depending on the requirements of the reactor apparatus. The at least one media conditioning module may include, but is not limited to, at least one of: * a pH control module for controlling a pH level of the reaction media; * a temperature control module for controlling a temperature of the reaction media. The temperature control module may include, but is not limited to, a heat exchanger, a shell and tube heat exchanger, a heating jacket, a chiller, a cooler and/or a cooling jacket; * a gas exchange module for introducing gas into the reaction media and/or removing gas from the reaction media. The gas exchange module may include, but is not limited to, an aerator, a sparger and/or an oxygenator; * a media reconstitution module for adding nutrients to the reaction media; * a media removal module for removing waste reaction media from the media conditioning circuit; * a media recycling module for recycling at least part of the reaction media in the media conditioning circuit; * a media reservoir for introducing fresh reaction media into the media conditioning circuit; * a waste removal module for removing waste media or waste products (metabolites, toxins) from the reaction media; * a mixer module for mixing the reaction media; * a mechanical agitation module for agitating the reaction media; * a sensor module for sensing a condition of the reaction media; The sensor module may include, but is not limited to, a pH sensor, a temperature sensor, a nutrient level sensor, a flow rate sensor and/or a gas concentration sensor; * a measurement module for measuring a condition of the reaction media; * a heat recovery module for recovering waste heat from one or more components of the media conditioning circuit; * a harvesting module for harvesting reaction product(s) from the culture media.

It will be appreciated that the sensor module may be combined with any other one of the media conditioning modules. For example, the pH control module may include the pH sensor, and/or the temperature control module may include a temperature sensor, and/or the gas exchange module may include a gas concentration sensor, and/or the media reconstitution module may include a nutrient level sensor.

The number and arrangement of reactor vessels may vary depending on the requirements of the reactor apparatus. The reactor apparatus may include a single reactor vessel or a plurality of reactor vessels. When a plurality of reactor vessels is used, the reactor vessels may be arranged in a cascading or parallel arrangement to, in use, permit a cascading or parallel flow of the reaction media through the reactor vessels. In some embodiments of the invention, the reactor vessels may include a combination of a cascading arrangement of reactor vessels and a parallel arrangement of reactor vessels.

The number and arrangement of media conditioning circuits may vary depending on the requirements of the reactor apparatus. The reactor apparatus may include a single media conditioning circuit or a plurality of media conditioning circuits. When a plurality of media conditioning circuits is used, the media conditioning circuits may be arranged in a parallel arrangement to, in use, permit a parallel flow of the reaction media through the media conditioning circuits.

In still further embodiments of the invention, the reactor apparatus may include at least one sterile selectively permeable barrier configured to permit the passage of the reaction media therethrough. The media conditioning circuit may be arranged in fluid communication with the reactor vessel via the at least one sterile selectively permeable barrier. In such embodiments, the sterile selectively permeable barrier may be a filter (such as a stainless steel filter and/or a microfilter) and/or a membrane.

The reactor apparatus may be used in a variety of cell culture applications. For example, the reactor vessel may be a suspension cell culture vessel or an adherent cell reactor vessel.

According to a third aspect of the invention, there is provided a method of operating a bioreactor system according to any one of the first and second aspects of the inventions and their embodiments, the method comprising the step of circulating the reaction media between the media conditioning circuit and the reactor vessel.

The features and advantages of the preceding aspects of the invention and their embodiments apply mutatis mutandis to the third aspect of the invention and its embodiments.

The method may include the step of selectively reversing a flow direction of the reaction media through the reactor vessel. This step may include timing the reversal of the flow direction of the reaction media through the reactor vessel, for example, to take place intermittently or at regular intervals. Furthermore, this step may include maintaining a same flow direction of the reaction media through the media conditioning circuit in different flow directions of the reaction media through the reactor vessel.

The method may include the step of separating cells from the reaction media so as to retain the cells inside the reactor vessel and permit circulation of the reaction media between the media conditioning circuit and the reactor vessel. This step may include separating the cells from the reaction media using a selectively permeable barrier and/or mechanically separating the cells from the reaction media by, for example, centrifugal separation, gravity separation or acoustic separation.

The method may include operating the at least one media conditioning module to condition the reaction media, which may include, but is not limited to, at least one of: * controlling a pH level of the reaction media; * controlling a temperature of the reaction media; * introducing gas into the reaction media and/or removing gas from the reaction media; * adding nutrients to the reaction media; * removing waste reaction media from the media conditioning circuit; * recycling at least part of the reaction media in the media conditioning circuit; * introducing fresh reaction media into the media conditioning circuit; * removing waste media or waste products from the reaction media; * mixing the reaction media; * mechanically agitating the reaction media; * sensing a condition of the reaction media; * measuring a condition of the reaction media; * recovering waste heat from one or more components of the media conditioning circuit; * harvesting reaction product(s) from the culture media.

It will be appreciated that the use of the terms "first" and "second", and the like, in this patent specification is merely intended to help distinguish between similar features, and is not intended to indicate the relative importance of one feature over another feature, unless otherwise specified.

Within the scope of this patent application it is expressly intended that the various aspects, embodiments, examples and alternatives set out in the preceding paragraphs, and the claims and/or the following description and drawings, and in particular the individual features thereof, may be taken independently or in any combination. That is, all embodiments and all features of any embodiment can be combined in any way and/or combination, unless such features are incompatible. The applicant reserves the right to change any originally filed claim or file any new claim accordingly, including the right to amend any originally filed claim to depend from and/or incorporate any feature of any other claim although not originally claimed in that manner.

Preferred embodiments of the invention will now be described, by way of non-limiting examples, with reference to the accompanying drawings in which: Figure 1 shows a conventional stirred tank bioreactor; Figure 2 shows a reactor apparatus according to a first embodiment of the invention; Figure 3 shows a reactor apparatus according to a second embodiment of the invention; Figure 4 shows a reactor apparatus according to a third embodiment of the invention; Figure 5 shows a reactor apparatus according to a fourth embodiment of the invention; Figure 6 shows a reactor apparatus according to a fifth embodiment of the invention; and Figure 7 shows a reactor apparatus according to a sixth embodiment of the invention.

The figures are not necessarily to scale, and certain features and certain views of the figures may be shown exaggerated in scale or in schematic form in the interests of clarity and conciseness.

The following embodiments of the invention are described with reference to a bioreactor apparatus comprising a suspension cell reactor vessel that contains culture media, but it will be appreciated that the following embodiments apply mutatis mutandis to other types of reactor apparatuses, reactor vessels and reaction media.

Biomanufacturing will have an ever-increasing role in sustainable manufacturing of medicines, materials, food, fuel, chemicals and more. At the core of biomanufacturing is the bioreactor, where cells are grown and the biological product is cultivated. To date biomanufacturing and bioreactors yield high value low volume products, with high R&D expenditure, long R&D timelines and high technical risk for process development and scaling.

Existing bioreactors provide low yield, are based on inefficient processes and are difficult to scale. This results in resource intensive, costly processes that are difficult to commercialise. This means that important and powerful biological innovations will not progress successfully out of the lab to commercial production. Most development efforts to overcome these problems is focussed on biological solutions but little attention is given to the underlying hardware and technology. System analysis of existing bioreactor systems reveals underlying conflicting requirements which inherently limit yield and scalability.

The inventor has developed a bioreactor apparatus that decouples media conditioning from the cell culture vessel, thereby enabling all functions required for cell growth to be independently controlled and optimised while providing low stress, optimal conditions for cell growth. In particular, the configuration of the bioreactor apparatus according to the invention provides an environment with minimal shear stress conditions for cell growth, optimised oxygen transfer to culture media, optimised heat transfer from culture media and optimised media nutrient conditions while providing consistent conditions independent of culture vessel size which facilitates predictable scale-up and scale-down of the reactor apparatus.

The invention will increase production yield, reduce production cost, improve resource efficiency and reduce the time to reach commercial production for biomanufacturing processes. The versatility, modularity and scalability of the invention enables the use of new cell types and the production of new products that are incompatible with existing bioreactors, thus enabling biomanufacturing of new sustainable biomanufactured products that cannot be produced currently.

The invention is compatible with batch and continuous biomanufacturing systems. The invention is applicable in a wide variety of industries, such as the food, cultivated meat, microbial fermentation and pharmaceutical industries.

Exemplary embodiments of the novel reactor apparatus are described as follows.

A bioreactor apparatus according to a first embodiment of the invention is shown in Figure 2 and is designated generally by the reference numeral 30.

The reactor apparatus 30 comprises a suspension cell culture vessel 32, a media conditioning circuit 34 and a fluid flow controller 36.

The suspension cell culture vessel 32 provides an enclosed environment with controlled conditions for promoting cell growth. This includes the provision of a culture media inside the culture vessel 32. The culture media provides a media in which the cells can grow. The culture media provides oxygen and nutrients to the cells while accepting carbon dioxide and byproducts from the cells. The culture vessel 32 may be in the form of a tank, jar or any other container. The culture vessel 32 may be made of, e.g., glass, polymers, metals, metal alloys, or combinations thereof. The culture vessel 32 comprises a media outlet 38 and a media inlet 40, which in the embodiment shown are in the form of openings at the ends of the culture vessel 32. The media outlet 38 permits flow of the culture media out of the culture vessel 32, while the media inlet 40 permits flow of the culture media into the culture vessel 32. By reversing the flow of culture media through the culture vessel 32, the media outlet 38 can be configured as a media inlet to permit flow of the culture media into the culture vessel 32, while the media inlet 40 can be configured as a media outlet to permit flow of the culture media out of the culture vessel 32. The culture media is typically water or water-based but may be formed of other liquids.

The culture vessel 32 comprises a media and cell separation device in the form of first and second filters 42,44 (e.g. membrane microfilters), which are arranged to trap and thereby retain cells inside the culture vessel 32. Particularly, the first filter 42 is arranged at or combined with the media outlet 38 to prevent cells from flowing through the media outlet 38, while the second filter 44 is arranged at or combined with the media inlet 40 to prevent cells from flowing through the media inlet 40. The first filter 42 may be integrated with the media outlet 38 to form a first filter housing, the second filter 44 may be integrated with the media inlet 40 to form a second filter housing, and the first and second filter housings are preferably arranged inside the culture vessel 32. An average pore size of the filters 42,44 is set (e.g. in the range of 2pm to 10pm for microbial (e.g. yeast, fungi, bacteria) and mammalian/animal/stem cell applications, potentially larger for other applications such as microcarrier-based applications) to allow culture media to flow through the filters 42,44 while preventing cells from flowing through the filters 42,44. Alternatively or additionally, the at least one media and cell separation device may include a mechanical separation device configured to, in use, mechanically separate the cells from the culture media by, for example, centrifugal separation, gravity separation or acoustic separation.

The culture vessel 32 may include an air vent, preferably a filtered air vent. The culture vessel 32 may include a sampling outlet for online or offline sampling of product from the culture vessel 32. The sampling may be carried out continuously, intermittently or at regular intervals. The culture vessel 32 may include a port for cell or biomass harvesting. The harvesting may be carried out continuously, intermittently or at regular intervals.

The bioreactor apparatus 30 may comprise one or more level sensors, such as an optical sensor, to detect a level of the culture media in the culture vessel 32. The bioreactor apparatus may include a heater for directly heating the culture vessel 32, a stirrer (e.g. a magnetic stirrer) for directly stirring the culture media in the culture vessel 32, and a weighing scale for weighing the culture vessel 32. The bioreactor apparatus 30 may optionally include an aerator or sparger for direct gas injection into the culture vessel 32.

The media conditioning circuit 34 is arranged in fluid communication with the media outlet 38 and media inlet 40 of the culture vessel 32 so as to create a media circulation loop comprising the media conditioning circuit 34 and the culture vessel 32. In use, culture media flows in the media circulation loop by exiting the media outlet 38 of the culture vessel 32, into and through the media conditioning circuit 34 and entering the media inlet 40 of the culture vessel 32, thereby circulating the culture media between the media conditioning circuit 34 and the culture vessel 32. Such fluid communication between the media conditioning circuit 34 and the media outlet 38 and media inlet 40 can be achieved by use of fluid conduits, pipes, tubes, channels and the like. As stated above, by reversing the flow of culture media through the culture vessel 32, the media outlet 38 can be configured as a media inlet to permit flow of the culture media into the culture vessel 32, while the media inlet 40 can be configured as a media outlet to permit flow of the culture media out of the culture vessel 32.

The media conditioning circuit 34 includes a plurality of media conditioning modules. Each media conditioning module is configured to, in use, condition the culture media flowing within the media conditioning circuit 34. In the embodiment shown, the media conditioning modules are distributed along the media conditioning circuit 34 so that, in use, the media conditioning modules sequentially and independently condition the culture media.

Non-limiting examples of suitable media conditioning modules are described through the specification. In the embodiment shown in Figure 2, the culture media entering the media conditioning circuit 34 first flows past a sensor module 46, a pump 48, a gas exchange module 50, a pH control module 52, a media reconstitution module 54, a temperature control module 56, another pump 48 and another sensor module 46 in that order. It will be appreciated that, in all embodiments of the invention, the sequence of modules in the media conditioning circuit 34 may vary and that different numbers and types of modules may be used in the media conditioning circuit 34. The media conditioning circuit 34 may include multiple of the same media conditioning module for increased capacity or capability.

Each sensor module 46 may include a single sensor or multiple sensors for sensing at least one condition of the culture media. Each sensor module 46 may include, but is not limited to, a pH sensor, a temperature sensor, a glucose sensor, a flow rate sensor and/or a gas concentration sensor. This allows one or more conditions of the culture media to be sensed and/or measured, and the resultant sensing signals can be used to control the other media conditioning modules. The first sensor module 46 may be used to sense at least one condition of the culture media before it undergoes media conditioning by the other media conditioning modules. The second sensor module 46 may be used to sense at least one condition of the culture media after it undergoes media conditioning by the other media conditioning modules. The sensing and/or measurement by the sensor modules 46 can be carried out continuously, intermittently or at regular intervals. Multiple sensors may be distributed along the media conditioning circuit 34 instead of being concentrated in a single sensor module 46.

The pumps 48 are configured to drive the flow of the culture media through the media conditioning circuit 34. The flow rate of the culture media through the media conditioning circuit 34 may be controlled depending on the media conditioning capabilities of the media conditioning modules and/or the condition requirements of the culture media and/or the volume of the culture vessel 32. Preferably the conditions of the culture media in the culture vessel 32 should be regulated to be as consistent as possible through control over the flow rate of the culture media through the media conditioning circuit 34. The flow rate of the culture media through the media conditioning circuit 34 may be in the range of 0.05 to 10 vessel volumes per minute, such as for microbial applications, and may be in the range of 0.01 to 1 vessel volumes per minute, such as for mammalian/animal/stem cell applications. Each pump 48 may be a mechanical pump, such as a peristaltic pump. Each pump 48 may be driven by a motor. The number of pumps may increase or decrease depending on the flow requirements of the media conditioning circuit 34.

The gas exchange module 50 is for introducing gas into the culture media and/or removing gas from the culture media. The gas exchange module 50 may include, but is not limited to, an aerator, a sparger and/or an oxygenator. This allows the gas concentration of the culture media to be controlled at a level that promotes cell growth in the culture vessel 32.

The pH control module 52 is configured to control a pH level of the culture media by enabling introduction of acid, base and/or CO2 into the culture media. This allows the pH of the culture media to be controlled at a level that promotes cell growth in the culture vessel 32.

The media reconstitution module 54 is for adding nutrients to the culture media. The nutrients may include, but is not limited to, carbon sources, sugars (e.g. glucose), proteins, fats, lipids, nitrogen sources, trace elements, inorganic salts, mineral salts and/or growth factors (e.g., vitamins, pH regulators). This allows the nutrients in the culture media to be controlled at a level that promotes cell growth in the culture vessel 32.

The temperature control module 56 is for controlling a temperature of the culture media. The temperature control module 56 may include, but is not limited to, a heat exchanger (such as an in-line heat exchanger), a shell and tube heat exchanger, a heating jacket, a chiller, a cooler and/or a cooling jacket.

The fluid flow controller 36 comprises a valve arrangement which may comprise a single valve or a plurality of valves. The or each valve may be a solenoid valve. The valve arrangement is configured to enable selective reversal of the flow direction of the culture media through the culture vessel 32 while maintaining the same flow direction of the culture media through the media conditioning circuit 34. In one flow direction of the culture media through the culture vessel 32, the culture media exits the culture vessel 32 via the media outlet 38 and enters the culture vessel 32 via the media inlet 40. In the other flow direction of the culture media through the culture vessel 32, the culture media exits the culture vessel 32 via the media inlet 40 and enters the culture vessel 32 via the media outlet 38.

The configuration of the valve arrangement may vary. For example, the valve arrangement may include a pair of 3-way fittings. Each 3-way fitting couples the media outlet 38 and media inlet 40 to a respective end of the media conditioning circuit. Each 3-way fitting is coupled to at least one valve that selectively opens and closes the flow paths in the 3-way fitting so that the respective end of the media conditioning circuit 34 is fluidly connected to either the media outlet 38 or the media inlet 40 at any given time. The operation of the valves associated with the 3-way fittings are coordinated to ensure that one end of the media conditioning circuit 34 is fluidly connected to one of the media outlet 38 and media inlet 40 while the other end of the media conditioning circuit 34 is fluidly connected to the other of the media outlet 38 and media inlet 40.

The reversal of the flow direction of the culture media through the culture vessel 32 may be automatic and may be timed to take place intermittently or at regular intervals.

This may be achieved by, for example, using a programmable controller (such as a programmable logic controller) to control the operation of the valve arrangement.

High flow rates of the culture media through the media circulation loop are beneficial to ensure regular replenishment of the depleted culture media and thereby continuous supply of replenished culture media to the culture vessel 22 to maintain optimal cell growth conditions in the culture vessel 32. The reversal of the flow direction enables an alternating backflow of the culture media through the culture vessel 32 to clean the filters 42,44 to prevent blockage resulting from accumulation of cells and other products at the filters 42,44 while maintaining a constant high flow rate of the culture media through the media conditioning circuit 34 for conditioning purposes.

A bioreactor apparatus according to a second embodiment of the invention is shown in Figure 3 and is designated generally by the reference numeral 130. The bioreactor apparatus of Figure 3 is similar in configuration and operation to the bioreactor apparatus of Figure 2, and like features share the same reference numerals.

The bioreactor apparatus of Figure 3 differs from the bioreactor apparatus of Figure 2 in that the bioreactor apparatus of Figure 3 includes the following additional features: a media removal module 58; a media recycling module 60; a media reservoir 62; a heat recovery module 64; and a sterile filter configuration 68, 70.

The media removal module 58 is configured to bleed waste culture media from the media conditioning circuit 34. Useful reaction products 66 are continuously removed from the waste culture media by a harvesting module in a downstream process. The media recycling module 60 removes waste products from the waste culture media (e.g. by a filtration device) and recycles the waste culture media by re-introducing the waste culture media back into the media conditioning circuit 34. The media reservoir 62 may be used to introduce fresh culture media into the media conditioning circuit 34 as necessary. It is envisaged that, in some embodiments of the invention, there is no media recycling in the media conditioning circuit 34 and/or the media conditioning circuit 34 is configured to receive fresh culture media from the media reservoir 62 to directly replace used culture media.

The heat recovery module 64 is for recovering waste heat from one or more components of the media conditioning circuit 34. In the embodiment shown, the heat recovery module 64 is configured to recover waste heat from motors associated with the pumps 48 and the temperature control module. Optionally the heat recovery module 64 may be configured to recover waste heat from control and sensor electronics associated with the bioreactor apparatus 130. The heat recovery may be carried out in various ways, such as the use of heat exchange elements to absorb the waste heat.

The sterile filter configuration includes sterile filters 68 respectively separating the media outlet 38 and media inlet 40 from the media conditioning circuit 34. In the embodiment shown, the sterile filters 68 are arranged between the media outlet 38 and the fluid flow controller 36, and between the media inlet 40 and the fluid flow controller 36. The sterile filters 68 are configured (e.g. by setting an average pore size) to allow the culture media to flow therethrough while preventing contaminants from passing therethrough. In this way the media conditioning circuit 34 is arranged in fluid communication with the culture vessel 32 via the sterile filters 68 which prevent contaminants from the media conditioning circuit 34 from entering the culture vessel 34. The sterile filter configuration may include a filter cleaning device 70 for cleaning the sterile filters 68, which may involve denaturing the sterile filters 68 and/or flushing the sterile filters 68 to remove accumulated cells and other products.

It will be appreciated that the bioreactor apparatus may exclude one or more of these additional features. It will be appreciated that one or more of these additional features may be implemented in any one of the embodiments described in this specification.

A bioreactor apparatus according to a third embodiment of the invention is shown in Figure 4 and is designated generally by the reference numeral 230. The bioreactor apparatus of Figure 4 is similar in configuration and operation to the bioreactor apparatus of Figure 2, and like features share the same reference numerals.

The bioreactor apparatus of Figure 4 differs from the bioreactor apparatus of Figure 2 in that, in the embodiment shown in Figure 4: * the culture media entering the media conditioning circuit 34 first flows past a pump 48, a temperature control module 56 (e.g., in the form of an in-line heat exchanger), a gas exchange module 50, a pH control module 52, a media reconstitution module 54, a mixing module 72 and a sensor module 46 in that order; and * the bioreactor apparatus omits the fluid flow controller 36 in the form of the valve arrangement.

The media reconstitution module 54 may be additionally configured as or combined with a waste removal module for removing metabolites and toxins from the culture media and/or removing waste media from the culture media.

The mixing module 72 is for mixing the culture media flowing therethrough. The mixing module 72 may include a static mixer and/or a dynamic mixer, such as an impeller.

Optionally the pump 48 may be configured to drive the flow of the culture media through the media conditioning circuit 34 and the culture vessel 32 in both directions. In this way the pump acts as a fluid flow controller that selectively reverses a flow direction of the culture media through the culture vessel 22. Similarly, the embodiments of Figures 2 and 3 may omit the valve arrangement, and their pumps may be configured to drive the flow of the culture media through the media conditioning circuit 34 and the culture vessel 32 in both directions.

A bioreactor apparatus according to a fourth embodiment of the invention is shown in Figure 5 and is designated generally by the reference numeral 330. The bioreactor apparatus of Figure 5 is similar in configuration and operation to any one of the bioreactor apparatuses of Figures 2, 3 and 4, and like features share the same reference numerals.

The bioreactor apparatus of Figure 5 differs from the bioreactor apparatuses of Figures 2, 3 and 4 in that the bioreactor apparatus of Figure 5 includes a plurality of culture vessels 22 arranged in a parallel arrangement. In use, the culture media flows in parallel through the culture vessels 22, and the parallel flows of the culture media combine into a single flow of the culture media through the media conditioning circuit 24.

A bioreactor apparatus according to a fifth embodiment of the invention is shown in Figure 6 and is designated generally by the reference numeral 430. The bioreactor apparatus of Figure 6 is similar in configuration and operation to any one of the bioreactor apparatuses of Figures 2, 3 and 4, and like features share the same reference numerals.

The bioreactor apparatus of Figure 6 differs from the bioreactor apparatuses of Figures 2, 3 and 4 in that the bioreactor apparatus of Figure 6 includes a plurality of culture vessels 22 arranged in a cascading (or series) arrangement. In use, the culture media flows sequentially through the culture vessels 22.

A bioreactor apparatus according to a sixth embodiment of the invention is shown in Figure 6 and is designated generally by the reference numeral 430. The bioreactor apparatus of Figure 6 is similar in configuration and operation to any one of the bioreactor apparatuses of Figures 2, 3 and 4, and like features share the same reference numerals.

The bioreactor apparatus of Figure 6 differs from the bioreactor apparatuses of Figures 2, 3 and 4 in that the bioreactor apparatus of Figure 6 includes a plurality of media conditioning circuits 34 arranged in a parallel arrangement. In use, the culture media flows in parallel through the media conditioning circuits 34, and the parallel flows of the culture media combine into a single flow of the culture media through the culture vessel 22.

In embodiments of the invention, the bioreactor apparatus may include at least one media conditioning device that, in use, directly conditions the culture media inside the culture vessel. The at least one media conditioning device may be integrated with the culture vessel. For instance, the at least one media conditioning device may include, but is not limited to, a pH control device, a temperature control device, a mechanical agitator (e.g. a stirrer or an impeller) and/or an aerator or sparger.

The bioreactor apparatus of the invention confers at least the following benefits: * The media conditioning circuit decouples the process control and media conditioning from the culture vessel: o The media bioreactor apparatus can be designed to enable the media conditioning to be carried out wholly or partially external to the culture vessel; o The media conditioning can be performed, controlled and designed independently from the cell growth process in the culture vessel; o The efficiency of the media conditioning is increased due to the removal of constraints imposed by the cell growth condition requirements of the culture vessel; o Stress-inducing media conditioning processes are removed from the culture vessel; * The contents of the culture vessel are mixed and maintained in suspension through the energy of the recirculating flow of the culture media: o Low shear and efficient mixing through fluidic mixing caused by the flow of the recirculating culture media, rather than high stress mechanical mixing or bubble mixing; o Controlled variability of the mixing through control over the flow of the recirculating culture media. The mixing may vary over the duration of the same cell growth process and between different cell growth processes.

o The fluidic mixing may be combined with a gentle direct mechanical mixing inside the culture vessel; * The media and cell separation is preferably integrated into the cell culture vessel: o Media and cell separation is conducted within the culture vessel, rather than outside the culture vessel; o Microfilter membranes are low cost, structurally simple and provide high flux while preventing cells from passing therethrough; o Other methods of media/cell separation could be used instead of or in conjunction with microfilter membranes (for instance centrifugal, gravity, acoustic separation, spinning filter etc.); * Recirculating flow direction of culture media into/out of the culture vessel is intermittently or periodically reversed. The flow of the culture media into/out of the culture vessel is through filters: o The reversal of the flow direction of the culture media through the culture vessel provides regular cleaning of the filters through alternating backflow; o On the other hand, using a single flow direction of the culture media through the culture vessel risks a blocking accumulation of cells at the filters; * The media conditioning circuit includes specialised, distributed media conditioning units along the flow path of the culture media: o Media conditioning is achieved in individual media conditioning modules that are distributed along the flow path of the culture media, rather than in a single complex vessel with inefficient and poor process control (as in conventional bioreactor systems); o The separation of the media conditioning modules from the culture vessel reduces the complexity of the interactions between the media conditioning modules from the culture vessel. This results in a highly modular and versatile bioreactor apparatus that can be readily scaled while maintaining optimal media conditioning performance and optimal cell growth conditions inside the cell culture vessel. This is because the capacity and/or capability of individual media conditioning modules can be independently optimised and scaled together with scaling of culture vessel size; * Measurement and sensing can be carried out external to the culture vessel: o Process measurements and sensing are taken partially or exclusively in the external media conditioning circuit and outside of the culture vessel; o This means that the measurement and sensing are of continuously flowing culture media rather than a point measurement inside the culture vessel; * Sterile filters preferably separate the culture vessel from the external media conditioning circuit: o This reduces the risk of contamination from the external media conditioning circuit o This reduces the sterilisation requirements of the components outside of the culture vessel and thereby simplifies the overall cost and design of the bioreactor apparatus; o The sterile filters continuously clean the culture media from any bacterial/microbial contamination as the culture media is continuously passed through the sterile filters.

In addition, the bioreactor apparatus of the invention has the following characteristics: o Improved process control, production and yield, with minimal shear stress to cells. Fragile cells are grown independently from harsh media conditioning processes. This is particularly beneficial for animal, mammalian and stem cells that are sensitive to shear stress, which means that a wider range of cells can now be used. Also, a wider range of mixing techniques and parameters can be used without concern over stresses to the cells; o Continuous media conditioning and sensing results in improved process control and uniformity of conditions; o Removal of metabolites in the external media conditioning circuit enables higher cell densities; o Continuous temperature control of the recirculating culture media, e.g. via specialised heat exchangers, provide an energy efficient method of improved temperature control. Accurate and precise temperature control is critical for cell growth and viability; o Oxygenation of the culture media happens in the absence of cells, avoiding stress to the cells associated with sparging and collapsing air bubbles in conventional stirred tank bioreactors. Recirculating culture media promotes uniform distribution of oxygenated media throughout the cell culture vessel. The configuration of the invention improves oxygen transfer and enables highly controllable dissolved oxygen profiles; o Continuous pH adjustment of culture media in the absence of cells avoids pH 'hotspots' and enables more uniform pH distribution across the culture vessel.

uniform pH distribution; o Continuous measurement and replenishment of culture media ensures sufficient access to nutrients for all cells throughout the cell culture process; o Reduced water consumption due to less use of fresh culture media and recycling of depleted culture media; o Continuous real time monitoring of the recirculated culture media enables superior control of critical process parameters; o The recirculating culture media provides hydrodynamic energy which mixes the contents of the cell culture vessel whilst avoiding excessive mechanical shear stress to the cells.

In comparison, the conventional stirred tank design has the following characteristics: o Difficulty in scaling up bioreactor apparatus due to complex interaction between components of culture vessel that can result in conflicting requirements, which in turn leads to inflexible and suboptimal scaling with high technical risk; o Growth of fragile cells and harsh media conditioning occur in the same enclosed vessel; o Single point sensing and direct injection of reagents results in uncontrolled process conditions; o Release of metabolites and other waste products in culture vessel limits cell density. Furthermore, traditional means of toxin removal use expensive filtration solutions and/or require pumping of cells which can cause damage to cells; o Water jacket provides heating/cooling via the external surface of the culture vessel. This is an inefficient and ineffective means of temperature control, especially as the size of the vessel increases. This is also an inefficient use of energy; o Air bubbles are directly sparged into the culture vessel. This results in large variations in oxygen concentration across the culture vessel, with highest concentrations near the sparger. The air bubbles provide direct hydrodynamic stress to cells. For fragile animal, mammalian and stem cell processes, such as for cultivated meat, spa rging and bubble collapse damages cells; o Acid and base are added directly into the culture vessel depending on the single point pH sensor readings in the vessel. Addition of acid/base directly into the vessel results in local variations in pH ('hotspots') which can damage cells. Large variation in pH distribution and local spikes in pH also limits yield; o Highly concentrated culture media is added at the start of the process which is depleted over time and periodically replenished through direct injection of nutrients into the culture vessel. This results in large concentration gradients of nutrients throughout the vessel and large variations during the process.

o Stirring the culture media inside the culture vessel, e.g. by using an impeller, provides mechanical shear stress to the cells; o Risk of contamination is high due to various pieces of equipment directly interacting with the culture vessel.

It will be appreciated that the above numerical values are merely intended to help illustrate the working of the invention and are not necessarily limiting on the scope of the invention.

The listing or discussion of an apparently prior-published document or apparently prior-published information in this specification should not necessarily be taken as an acknowledgement that the document or information is part of the state of the art or is common general knowledge.

Preferences and options for a given aspect, feature or parameter of the invention should, unless the context indicates otherwise, be regarded as having been disclosed in combination with any and all preferences and options for all other aspects, features and parameters of the invention.

Claims (25)

1. CLAIMS1. A reactor apparatus comprising: a reactor vessel; a media conditioning circuit in fluid communication with the reactor vessel so as to permit circulation of reaction media between the media conditioning circuit and the reactor vessel, wherein the media conditioning circuit includes at least one media conditioning module configured to, in use, condition the reaction media within the media conditioning circuit; and a fluid flow controller configured to, in use, selectively reverse a flow direction of the reaction media through the reactor vessel.
2. 2. A reactor apparatus according to Claim 1 wherein the fluid flow controller is configured to, in use, automatically and selectively reverse the flow direction of the reaction media through the reactor vessel.
3. 3. A reactor apparatus according to any one of the preceding claims wherein the fluid flow controller is configured to, in use, automatically and selectively reverse the flow direction of the reaction media through the reactor vessel intermittently or at regular intervals.
4. 4. A reactor apparatus according to any one of the preceding claims wherein the fluid flow controller includes at least one pump configured to, in use, selectively reverse the flow direction of the reaction media through the reactor vessel.
5. 5. A reactor apparatus according to any one of the preceding claims wherein the fluid flow controller includes at least one valve configured to, in use, selectively reverse the flow direction of the reaction media through the reactor vessel.
6. 6. A reactor apparatus according to any one of the preceding claims wherein the media conditioning circuit and fluid flow controller are arranged to, in use, maintain a same flow direction of the reaction media through the media conditioning circuit in different flow directions of the reaction media through the reactor vessel.
7. 7. A reactor apparatus according to any one of the preceding claims wherein the media conditioning circuit comprises the fluid flow controller.
8. 8. A reactor apparatus according to any one of the preceding claims including at least one media and cell separation device configured to, in use, retain cells inside the reactor vessel and permit circulation of the reaction media between the media conditioning circuit and the reactor vessel.
9. 9. A reactor apparatus comprising: a reactor vessel; a media conditioning circuit in fluid communication with the reactor vessel so as to permit circulation of reaction media between the media conditioning circuit and the reactor vessel, wherein the media conditioning circuit includes at least one media conditioning module configured to, in use, condition the reaction media within the media conditioning circuit; and at least one media and cell separation device configured to, in use, retain cells inside the reactor vessel and permit circulation of the reaction media between the media conditioning circuit and the reactor vessel.
10. 10. A reactor apparatus according to Claim 8 or Claim 9 wherein the reactor vessel comprises the at least one media and cell separation device.
11. 11. A reactor apparatus according to any one of Claims 8 to 10 wherein the at least one media and cell separation device includes a selectively permeable barrier.
12. 12. A reactor apparatus according to Claim 11 wherein the selectively permeable barrier is a filter and/or a membrane.
13. 13. A reactor apparatus according to any one of Claims 8 to 12 wherein the at least one media and cell separation device includes a mechanical separation device configured to, in use, mechanically separate the cells from the reaction media.
14. 14. A reactor apparatus according to Claim 13 wherein the mechanical separation device is configured to, in use, mechanically separate the cells from the reaction media by centrifugal separation, gravity separation or acoustic separation.
15. 15. A reactor apparatus according to any one of the preceding claims wherein the media conditioning circuit includes a plurality of media conditioning modules, each of which is configured to, in use, condition the reaction media.
16. 16. A reactor apparatus according to Claim 15 wherein the media conditioning modules are arranged to, in use, sequentially condition the reaction media.
17. 17. A reactor apparatus according to Claim 15 or 16 wherein the media conditioning modules are arranged to, in use, independently condition the reaction media.
18. 18. A reactor apparatus according to any one of the preceding claims wherein the at least one media conditioning module includes at least one of: * a pH control module for controlling a pH level of the reaction media; * a temperature control module for controlling a temperature of the reaction media; * a gas exchange module for introducing gas into the reaction media and/or removing gas from the reaction media; * a media reconstitution module for adding nutrients to the reaction media; * a media removal module for removing waste reaction media from the media conditioning circuit; * a media recycling module for recycling at least part of the reaction media in the media conditioning circuit; * a media reservoir for introducing fresh reaction media into the media conditioning circuit; * a waste removal module for removing waste media or waste products (e.g. metabolites, toxins) from the reaction media; * a mixer module for mixing the reaction media; * a mechanical agitation module for agitating the reaction media; * a sensor module for sensing a condition of the reaction media; * a measurement module for measuring a condition of the reaction media; * a heat recovery module for recovering waste heat from one or more components of the media conditioning circuit; * a harvesting module for harvesting reaction product from the culture media. 30
19. 19. A reactor apparatus according to any one of the preceding claims including a plurality of reactor vessels, wherein the reactor vessels are arranged in a cascading or parallel arrangement to, in use, permit a cascading or parallel flow of the reaction media through the reactor vessels.
20. 20. A reactor apparatus according to any one of the preceding claims including at least one sterile selectively permeable barrier configured to permit the passage of the reaction media therethrough, wherein the media conditioning circuit is arranged in fluid communication with the reactor vessel via the at least one sterile selectively permeable barrier.
21. 21. A reactor apparatus according to Claim 20 wherein the sterile selectively permeable barrier is a filter and/or a membrane.
22. 22. A reactor apparatus according to any one of the preceding claims wherein the reactor vessel is a suspension cell reactor vessel or an adherent cell reactor vessel.
23. 23. A method of operating a bioreactor system according to any one of the preceding claims, the method comprising the step of circulating the reaction media between the media conditioning circuit and the reactor vessel.
24. 24. A method according to Claim 23 when dependent from Claim 1 or any claim dependent thereon, the method including the step of selectively reversing a flow direction of the reaction media through the reactor vessel.
25. 25. A method according to Claim 23 or Claim 34 when dependent from Claim 8, Claim 9 or any claim dependent thereon, the method including the step of separating cells from the reaction media so as to retain the cells inside the reactor vessel and permit circulation of the reaction media between the media conditioning circuit and the reactor vessel.