DEVICE ASSEMBLY FOR PRODUCING BIOCONJUGATES

Abstract

A device assembly for producing bioconjugates, in particular antibody-drug conjugates, including a conjugation unit for performing a bioconjugation reaction in a medium, a first filtration unit for separating precipitates and/or agglomerates, and a second filtration unit for performing an ultrafiltration and/or a diafiltration process. The first filtration unit is arranged in a flow path between the conjugation unit and the second filtration unit. The device assembly further includes a single control unit for controlling the transfer of medium from the conjugation unit through the first filtration unit to the second filtration unit and for controlling the ultrafiltration and/or diafiltration process.

Claims

1. A device assembly for producing bioconjugates, in particular antibody-drug conjugates, the device assembly comprising: a conjugation unit for performing a bioconjugation reaction in a medium; a first filtration unit for separating precipitates and/or agglomerates; a second filtration unit for performing an ultrafiltration and/or a diafiltration process, the first filtration unit being arranged in a flow path between the conjugation unit and the second filtration unit; and the device assembly further comprising a single control unit for controlling the a transfer of the medium from the conjugation unit through the first filtration unit to the second filtration unit and for controlling the ultrafiltration and/or the diafiltration process.

2. The device assembly according to claim 1, characterized in that a first group of components of the device assembly coming into contact with the medium are configured as single-use components made from one or more materials of a first group of materials consisting of: high-density polyethylene (HDPE); polyamide (PA); polybutylene terephthalate (PBT); polytetrafluoroethylene (PTFE); polypropylene (PP); polyethylene terephthalate (PET); ceramics; glass; and in that a second group of components of the device assembly coming into contact with the medium are configured as single-use components made from one or more of the following materials: low-density polyethylene (LDPE); (reinforced) platinum cured silicone (Si(Pt)); thermoplastic elastomer (TPE); thermoplastic polyolefin (TPO); thermoplastic vulcanizate (TPV); ethylene propylene diene monomer rubber (EPDM); silicone; a material from the first group of materials.

3. The device assembly according to claim 1, characterized in that the conjugation unit comprises a flexible conjugation bag and a first bag holder holding the conjugation bag.

4. The device assembly according to claim 3, characterized in that the conjugation bag has a bottom portion and an outlet formed therein, the first bag holder having a bottom surface and a draining opening formed at a lowest point of the bottom surface, the bottom portion of the conjugation bag resting on the bottom surface of the first bag holder such that the outlet of the conjugation bag is located at the draining opening of the first bag holder.

5. The device assembly according to claim 3, characterized in that the first bag holder includes a double-walled containment, the conjugation unit further comprising a temperature control system for controlling the temperature of the medium in the conjugation bag, the temperature control system including a heat transfer fluid flowing through the double-walled containment of the first bag holder.

6. The device assembly according to claim 4, characterized in that a pH sensor is arranged in the conjugation bag.

7. The device assembly according to claim 1, characterized in that the conjugation unit comprises a controllable draining valve and the first filtration unit comprises a transfer pump for transferring the medium from the conjugation unit through the first filtration unit to the second filtration unit.

8. The device assembly according to claim 1, characterized in that the first filtration unit comprises a filter device with a membrane having a pore size of less than 0.8 μm.

9. The device assembly according to claim 5, characterized in that the second filtration unit comprises a single-use loop-assembly for recirculating the medium in the ultrafiltration and/or the diafiltration process which can be mounted to and/or dismounted from a basic structure of the device assembly as a whole, together with a single-use conjugation bag and a single-use recirculation bag and other single-use components of the device assembly.

10. The device assembly according to claim 9, characterized in that the single-use loop-assembly includes a recirculation pump, at least one sensor, tubing and connectors.

11. The device assembly according to claim 10, characterized in that the recirculation bag has a bottom portion and an outlet formed therein, the recirculation bag being held in a second bag holder having a bottom surface and a draining opening formed at a lowest point of the bottom surface, the bottom portion of the recirculation bag resting on the bottom surface of the second bag holder such that the outlet of the recirculation bag is located at the draining opening of the second bag holder.

12. The device assembly according to claim 11, characterized in that the second bag holder includes a double-walled containment the second filtration unit further comprising a temperature control system for controlling the temperature of the medium in the recirculation bag, the temperature control system including a heat transfer fluid flowing through the double-walled containment of the second bag holders.

13. The device assembly according to claim 10, characterized in that a conductivity sensor is arranged in the recirculation bag.

14. The device assembly according to claim 10, characterized in that the single-use loop-assembly includes controllable valves.

15. The device assembly according to claim 9, characterized in that a dip tray is arranged below the single-use loop-assembly, the dip tray having a bottom surface and a draining means formed at a lowest point of the bottom surface.

16. The device assembly according to claim 1, characterized in that the second filtration unit comprises at least one cross-flow filter cassette held in a filter press.

17. The device assembly according to claim 12, characterized in that the control unit controls at least one of the following: temperature of the heat transfer fluid flowing through the double-walled containment of the first bag holder; temperature of the heat transfer fluid flowing through the double-walled containment of the second bag holder; controllable valves; a transfer pump; the recirculation pump.

18. The device assembly according to claim 1, characterized in that all components of the device assembly, except for an operating panel coupled to the control unit, are mounted on a trolley.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0035] Further features and advantages of the invention will become apparent from the following description and from the accompanying drawing to which reference is made. In the drawings:

[0036] FIG. 1 shows a device assembly for producing bioconjugates according to the invention without the most parts of a single-use loop-assembly; and

[0037] FIG. 2 shows another view of the device assembly of FIG. 1 with the single-use loop-assembly;

[0038] FIG. 3 shows a single-use conjugation bag and equipment used in the device assembly;

[0039] FIG. 3a shows a detail of the conjugation bag;

[0040] FIG. 4 shows a contactless single-use stirrer of the impeller system of the device assembly; and

[0041] FIG. 5 shows a single-use recirculation bag and equipment used in the device assembly.

DETAILED DESCRIPTION

[0042] As shown in FIG. 1, an entire device assembly 10 for producing bioconjugates is built on a trolley 12 and movable as a whole. In the device assembly 10 three dedicated units for performing three unit operations in a bioconjugate production process are combined. The first unit is a conjugation unit for performing a bioconjugation reaction in a medium. The second unit is a filtration unit for separating precipitates and/or agglomerates from the medium containing the bioconjugate. The third unit is a filtration unit for performing an ultrafiltration and/or a diafiltration process for purifying and concentrating the bioconjugate product. These three units will be described in detail below.

[0043] The conjugation unit comprises a first bag holder 14 for accommodating a flexible single-use conjugation bag 16 (shown separately in FIG. 3). The first bag holder 14 includes a double-walled or double-jacketed containment 18 and a transparent glass door 20. The containment 18 together with the glass door 20 laterally surround a receiving space. The glass door 20 enables easy access and makes the receiving space visible to the operator. While its upper side is open, the first bag holder 14 has a conical or otherwise inclined bottom surface 22 with a draining opening 24 formed at the lowest point of the bottom surface 22. The first bag holder 14 further includes a load cell 26 for measuring the weight of the medium in the conjugation bag 16. The load cell 26 is connected to a central control unit 28 of the device assembly 10 that will be described in more detail later.

[0044] The conjugation bag 16 (see FIG. 3) is a 3D-shaped sterilized single-use bag made from a flexible foil material. The volume of the conjugation bag 16 and the dimensions of the first bag holder 14 are adapted to each other so that the filled conjugation bag 16 takes a stable position in the receiving space. A typical working volume of the conjugation bag 16 is 20 L. Of course, other volumes are possible.

[0045] Regardless of the size, a bottom portion of the conjugation bag 16 rests on the bottom surface 22 of the first bag holder 14 such that a lower outlet port of the conjugation bag 16 is located directly above or in the draining opening 24. The lower outlet port is connected to a conjugation outlet hose line (draining line) 30 through the draining opening 24. The conjugation outlet hose line 30 can be opened and blocked by a draining valve 32, preferably a pinch valve, which is controlled by the control unit 28. Since the lower outlet port is placed at the lowest point of the bottom surface 22, the conjugation bag 16 can be completely emptied by gravity without any active draining means when the conjugation outlet hose line 30 is opened by the draining valve 32.

[0046] The conjugation unit comprises an impeller system (not visible in FIGS. 1 and 2). At the bottom of the first bag holder 14 a magnetic drive causes a contactless stirrer 34 (shown separately in FIG. 4), which is placed loosely inside the conjugation bag 16, to lift from a receptacle 35 and rotate. In FIG. 3a the magnetic coupling 36 between the magnetic drive and the stirrer 34 can be seen. The stirring of the medium ensures a homogeneous mixture throughout the conjugation reaction. The contactless stirrer 34 produces only little abrasion inside the conjugation bag 16. However, other impeller systems can be used as well, e.g. an impeller system including a driven shaft penetrating into the conjugation bag 16 through a sealed opening. The impeller system is controlled by the control unit 28.

[0047] As can be seen in FIGS. 3 and 3a, the conjugation bag 16 is provided with several ports 38 for connecting supply lines 40 through which the fluids/solutions necessary for the conjugation reaction are fed into the conjugation bag 16. Peristaltic pumps 44 (see FIGS. 1 and 2) controlled by the control unit 28 are used to supply the fluids/solutions as required from reservoirs 42 or other sources through the supply lines 40, which are connected to the ports 38 of the conjugation bag 16 on the one hand, and to the reservoirs 42 or the other sources on the other hand, via sterile connectors 46.

[0048] The conjugation bag 16 is equipped with a single-use pH sensor 48 (probe). Other single-use sensors for detecting parameters of interest in connection with the conjugation reaction can be provided as well. The sensors are connected to the control unit 28.

[0049] The conjugation unit also comprises a first temperature control system coupled to the control unit 28 for controlling the temperature of the medium in the conjugation bag 16. A heat transfer fluid can be pumped through the double-walled containment 18 of the first bag holder 14 to cool or heat the medium in the conjugation bag 16 as desired. A temperature sensor transmits the current temperature value to the control unit 28. The temperature sensor can be arranged at the bottom surface 22 of the first bag holder 14, for example.

[0050] Further, a first sterile venting filter 50 (see FIG. 1) for venting the conjugation bag 16 is fixed to a mount above the conjugation bag 16 and connected to a venting port of the conjugation bag 16.

[0051] The conjugation outlet hose line 30 is part of a flow path extending between the conjugation bag 16 and a recirculation bag 52 of a second filtration unit for performing an ultrafiltration and/or a diafiltration process which will be described later. A peristaltic transfer pump 54 and a single-use filter device 56 are integrated into this flow path, forming a first filtration unit of the device assembly 10.

[0052] The general type and the specific configuration of the filter device 56 can be chosen according to the given requirements of the separation process. In the embodiment shown in FIG. 2 the filter device 56 is a filter capsule having a multi-layer membrane made of one or more of the following materials: nylon; polytetra-fluoroethylene (PTFE); polypropylene (PP) fleece. The membrane may also be a stabilized cellulose based membrane, e.g. a Hydrosart® high performance membrane.

[0053] The membrane of the filter device 56 has a pore size of less than 0.8 μm. Preferably, the filter device 56 is a sterile filter with a pore size of about 0.2 μm or less, which is yet larger than the size of the bioconjugate formed in the medium of the conjugation bag 16. Thus, the filter device 56 is capable of effectively separating precipitates and/or agglomerates (or other unwanted lumps) from the medium containing the bioconjugate that have also formed unintentionally during the conjugation reaction in the conjugation bag 16.

[0054] The second filtration unit of the device assembly 10 comprises the already mentioned recirculation bag 52 (shown separately in FIG. 5) which is held in a second bag holder 58. The recirculation bag 52 and the conjugation bag 16 are of the same or a similar type and size.

[0055] In a bottom portion of the recirculation bag 52 a lower outlet port is connected to a recirculation outlet hose line (draining line) 60 which can be opened or closed by a draining valve 62, preferably a pinch valve (see FIG. 2), controlled by the control unit 28. Further, the recirculation bag 52 has several ports 64, one of which is for receiving the filtrate of the medium that passes through the filter device 56 of the first filtration unit. Other ports 64 of the recirculation bag 52 enable the supply or exchange of buffers or other fluids/solutions during the UF/DF filtration process. Similar to the conjugation bag 16, hose lines 66 are connected to the ports 64 of the recirculation bag 52 on the one hand, and to reservoirs or other sources or to further tubing of the device assembly 10 on the other hand, via sterile connectors 46.

[0056] Instead of or in addition to a single-use pH sensor as used in the conjugation bag 16, the recirculation bag 52 is equipped with a single-use conductivity sensor 68. Other single-use sensors for detecting parameters of interest in connection with the filtration process running in the second filtration unit can be provided as well. The sensors are connected to the control unit 28.

[0057] Like the conjugation bag 16 and the recirculation bag 52, also the first bag holder 14 and the second bag holder 58 have a similar design, including—inter alia—a draining opening 24 at the bottom surface 22 for the recirculation outlet hose line 60 and a load cell 26 connected to the control unit 28. A second sterile venting filter 70 (see FIG. 1) for venting the recirculation bag 52 is fixed to a mount above the recirculation bag 52 and connected to a venting port of the recirculation bag 52. The temperature of the medium in the recirculation bag 52 is controlled by a second temperature control system similar to the first temperature control system. However, an impeller system is not required for the medium in the recirculation bag 52.

[0058] The recirculation outlet hose line 60 is part of a tubing of the second filtration unit leading to one or more single-use filter cassettes 72 held in a filter press 74. The filter cassettes 72 are self-contained units configured for cross-flow filtration, in particular for an ultrafiltration or diafiltration process. Several filter cassettes 72 can be connected in parallel or in series.

[0059] The type, number and arrangement of filter cassettes 72 can be chosen according to the given requirements of the UF/DF process. The nominal molecular weight limit (NMWL) or the molecular weight cutoff (MWCO) of the membranes of the filter cassettes 72 is adapted to the size and weight of the bioconjugate so that the unbound small-sized and lighter components of the medium are washed out to the permeate side while the larger-sized and heavier bioconjugate is kept in the retentate. The total membrane area of the filter cassette(s) 72 preferably ranges from 0.1 to 1.4 m.sup.2.

[0060] The second filtration unit further comprises a single-use recirculation pump 76, tubing, connectors, sensors 78 and further controllable valves 80, preferably pinch valves, enabling the blocking and unblocking of a variety of flow paths. Such flow paths include the path for recirculating the medium in the second filtration unit, and can further include paths for supplying buffer to the recirculation bag 52, discharging waste medium, flushing etc. The sensors 78 are pressure sensors and/or flow meters arranged in the feed, permeate and retentate lines of the tubing. An optional UV probe can be arranged in the permeate line for UV spectroscopy in order to detect any product in the permeate line in case of a defective membrane. Like the conductivity sensor 68 in the recirculation bag 52, the sensors 78 transmit their measured values or signals to the control unit 28. The pinch valves 80 are all controlled by the control unit 28.

[0061] Thus, the second filtration unit is capable of performing a complete UF/DF process for purifying and concentrating the bioconjugate, controlled by the control unit 28 without human interaction.

[0062] A special characteristic of the second filtration unit is that the recirculation pump 76, the tubing, the connectors 46, the sensors 78 and the valves 80 used in the UF/DF process are single-use components configured as a preassembled unit, hereinafter referred to as single-use loop-assembly. The complete single-use loop-assembly can be mounted to and dismounted from a basic structure of the device assembly 10 as a whole.

[0063] The pre-configured tubing design of the single-use loop assembly ensures a minimum dead volume (hold-up volume), e.g. below 300 mL. This is very important in view of the high monetary value of the bioconjugate product.

[0064] Below the single-use loop-assembly a dip tray 82 is arranged. The dip tray 82 has a bottom surface and a draining means, e.g. a ball valve, formed at a lowest point of the bottom surface. Thus, in case of a leakage, the leaking fluid is collected in the dip tray 82 and can be guided to a waste tank positioned below the draining means.

[0065] The device assembly 10 further comprises an operating panel 84, preferably a touchscreen, which is part of the control unit 28. With the operating panel 84 an operator can make all selections and adjustments that are necessary before and after the bioconjugate production process, which itself is performed highly automatically. The operating panel 84 also serves to inform the operator of the current settings and status of the process and allows interaction, if necessary.

[0066] In the embodiment shown in FIGS. 1 and 2 the operating panel 84 is fixed to a mount of the device assembly 10. According to another embodiment, the operating panel 84 is a separate unit, wirelessly connected to the control unit 28 of the device assembly 10 or by a long enough cable, allowing a positioning of the operating panel 84 remote from the rest of the device assembly 10. The remote positioning of the operating panel 84 makes it possible to arrange the rest of the device assembly 10 in a restricted area, especially in an isolated room (isolator) and/or under a fume hood. If the possibility of human manipulation at the device assembly 10 is still required or desired, the device assembly 10 (without the operating panel 84) can be arranged in a glovebox.

[0067] All of the components that come into contact with toxic medium (wetted components) are dedicated single-use components that will not be used again and therefore need not be cleaned and sterilized after use. The wetted components are made from materials that are especially resistant to the following solvents which could be used in connection with the conjugation reaction: dimethylacetamide (DMAc), dimethyl sulfoxide (DMSO), dimethylformamide (DMF), propylene glycol (PG), acetonitrile (ACN), n-methyl-2-pyrrolidone (NMP).

[0068] A first group of the wetted components, especially wetted components of the conjugation unit, are resistant to a 100% concentration of the solvents (i.e. the pure solvents). This first group includes a toxin supply line 86 which leads from the reservoir 42 (here a bottle), containing toxin solved in 100% solvent, to the conjugation bag 16.

[0069] Since the mixing with other fluids/solutions during the conjugation reaction and the UF/DF filtration process dilutes the solvent, a second group of the wetted components, especially the wetted components of the second filtration unit, only have to be resistant to at least a 20% concentration of the solvents. This also applies to the other supply lines 40 because they are connected to the conjugation bag 16 below the liquid level so that instant dilution occurs there.

[0070] The wetted components of the first and second groups may be made from one or more of the following materials (100% resistant): high-density polyethylene (HDPE); polyamide (PA); polybutylene terephthalate (PBT); polytetrafluoro-ethylene (PTFE); polypropylene (PP); polyethylene terephthalate (PET); ceramics; glass. The wetted components of the second group may also be made from one or more of the following materials (20% resistant): low-density polyethylene (LDPE); (reinforced) platinum cured silicone (Si(Pt)); thermoplastic elastomer (TPE); thermoplastic polyolefin (TPO); thermoplastic vulcanizate (TPV); ethylene propylene diene monomer rubber (EPDM); silicone.

[0071] In the following, operation of the device assembly 10 in a bioconjugate production process is described.

[0072] Before the device assembly 10 can be used, the single-use components are provided and mounted to the basic structure of the device assembly 10. In particular, the conjugation bag 16 and the recirculation bag 52 are placed in the respective bag holders 14, 58 and integrity tested, or the other way round. The single-use loop-assembly, including the recirculation pump 76, the tubing, the connectors 46, the sensors 78 and the valves 80 used in the UF/DF process, is mounted, step-by-step or as a whole, to the basic structure of the device assembly 10. The necessary connections between the single-use loop-assembly and the conjugation bag 16 and the recirculation bag 52 and any further connections are made via sterile connectors 46.

[0073] After the preparations, the operator starts the bioconjugate production process at the operating panel 84. The conjugation bag 16 is filled with the fluids/solutions necessary to start the conjugation reaction. The reaction conditions are monitored, and the peristaltic pumps 44 controlled by the control unit 28 supply fluids/solutions as required while the conjugation reaction takes place. The temperature in the conjugation bag 16 is adjusted via the first temperature control unit.

[0074] The conjugation reaction is terminated when the pH measured by the pH sensor 48 exceeds a predetermined threshold value. Further stop criterions may be involved, like expiry of a set time period or analysis of a sample taken from the conjugation bag 16. The control unit 28 then opens the draining valve 32 and activates the transfer pump 54.

[0075] The medium leaves the conjugation bag 16 through the conjugation outlet hose line 30 and passes through the filter device 56. Thereby the precipitates and/or agglomerates are separated from the filtrate. The filtrate containing the bioconjugate is directed to the recirculation bag 52.

[0076] When all of the filtrate has entered the recirculation bag 52, the UF/DF process for purifying and concentrating the bioconjugate in the second filtration unit is started. The control unit 28 activates the recirculation pump 76 to keep the medium circulating in the single-use loop-assembly with the filter cassettes 72. Here, the permeate with the unbound reactants is guided to a waste tank while the retentate with the bioconjugate is kept in the loop. Since the precipitates and/or agglomerates have been removed by the filter device 56, they cannot clog the membranes of the filter cassettes 72. Buffer is supplied/exchanged as required with the aid of the corresponding valves 80 which are controlled by the control unit 28.

[0077] The UF/DF process is terminated when the conductivity value measured by the conductivity sensor 68 in the recirculation bag 52 exceeds a predetermined threshold value. Further stop criterions may be involved, like expiry of a set time period or evaluation of a UV spectrum measured by the UV probe or analysis of a sample taken from the conjugation bag 16.

[0078] The control unit 28 controls the pinch valves 80 such that the retentate with the purified and concentrated bioconjugate can be extracted from the single-use loop-assembly through a harvest line into a container for further processing. Since the dead volume of the single-use loop-assembly is very low, only a small amount of the valuable medium remains in the loop-assembly.

[0079] Before the single-use loop-assembly, including the connected filter cassettes 72, is removed from the basic structure of the device assembly 10 together with the bags 16, 52, the tubing (including the supply and other hose lines 30, 40, 60, 66, 86), the connectors 46, the sensors 48, 68, 78 and the pinch valves 80 as a whole, the control unit 28 closes all peripheral pinch valves 80 to create a confinement for the residual medium in the loop-assembly. The sealed single-use loop-assembly together with the other single-use components is dismounted and disposed of as a whole, i.e. its components are not separated from each other. This enables a very high safety standard since the toxic medium cannot exit the loop-assembly and come into contact with the operator and the environment.

[0080] The device assembly 10 is particularly useful for preclinical or clinical production of ADCs.

[0081] The device assembly 10 can also be used without the conjugation unit and the first filtration unit, serving as a conventional UF/DF filtration device assembly with a disposable single-use loop-assembly.

LIST OF REFERENCE SIGNS

[0082] 10 device assembly [0083] 12 trolley [0084] 14 first bag holder [0085] 16 conjugation bag [0086] 18 containment [0087] 20 glass door [0088] 22 bottom surface [0089] 24 draining opening [0090] 26 load cell [0091] 28 control unit [0092] 30 conjugation outlet hose line [0093] 32 draining valve [0094] 34 stirrer [0095] 35 receptacle for stirrer [0096] 36 magnetic coupling [0097] 38 ports of conjugation bag [0098] 40 supply lines [0099] 42 reservoirs [0100] 44 peristaltic pumps [0101] 46 connectors [0102] pH sensor [0103] 50 first venting filter [0104] 52 recirculation bag [0105] 54 transfer pump [0106] 56 filter device [0107] 58 second bag holder [0108] 60 recirculation outlet hose line [0109] 62 draining valve [0110] 64 ports of recirculation bag [0111] 66 hose lines [0112] 68 conductivity sensor [0113] 70 second venting filter [0114] 72 filter cassette [0115] 74 filter press [0116] 76 recirculation pump [0117] 78 sensors [0118] 80 controllable valves [0119] 82 dip tray [0120] 84 operating panel [0121] 86 toxin supply line