Reinforcing member for cell culture bioreactor

Abstract

The present invention relates to a bioprocess bag (118) comprising a bag wall defining an enclosed volume for holding a biological material. The bag wall comprises at least one inlet port (142) and at least one outlet port (146). The bioprocess bag (118) further comprises a tube structure (400) comprising a first open end portion proximal to the bag wall and a second distal end portion, the tube extending into the enclosed volume, and the tube structure (400) comprises a reinforcement portion (402) proximal to the first open end portion.

Description

Technical Field

[0001] This invention relates to novel bioprocessing bags for cell culture with enhanced internal structures. More particularly, this invention relates to improved bioprocessing bags and related structures, including enhanced and improved tube features. Background Technology

[0002] Cell therapy is a new but rapidly expanding field of biotechnology, involving the administration of autologous or heterologous cells to perform therapeutic effects in the body. Cell therapy involves several mandatory stages from cell collection to cell injection into the patient. Cell culture for cell therapy is performed in a cleanroom environment. Cell culture and cleanrooms have many mandatory regulations, such as particle size and number in the cleanroom, the number of patient samples processed at one time, the number of instruments per unit, and the requirements for a sterile environment.

[0003] Early systems for cell culture were stand-alone, required large spaces, and could not process multiple patient samples simultaneously. Procedures for cell culture used in cell therapy involve significant human intervention, which can contaminate cell cultures and impair cell growth, especially for small cell cultures.

[0004] Single-use bioreactors or fermenter bags have been developed for cell therapy. The use of single-use bioreactor bags in cell culture reduces batch changeover time by eliminating time-consuming cleaning and equipment validation, resulting in higher production volumes. Figure 1 A single-use bioreactor system is shown in U.S. Patent 9,109,193 entitled “Continuous Perfusion Bioreactor System”. Figure 1 As described more fully in the '193 patent, and briefly herein. Certain aspects include an apparatus 100 comprising a vessel 114, which, in the illustrated embodiment, is a reusable support structure (e.g., a stainless steel tank) that surrounds and houses a container 118. The apparatus 100 may optionally include an environmentally closed housing 120 that surrounds a portion of the vessel 114. The container 118 may be configured as a collapsible bag (e.g., a polymer bag). The collapsible bag 118 may be fluid-proof to allow it to contain a liquid 122, which may contain reactants (e.g., certain solid objects), media, and / or other components necessary to perform the desired process (such as chemical, biochemical, and / or biological reactions). The collapsible bag 118 may also be configured such that the liquid 122 remains substantially in contact only with the collapsible bag during use and not with the support vessel 114, allowing the support structure to be reused without cleaning.

[0005] Figure 1 Optional inlet port 142 and optional outlet port 146 are also shown, which may be formed in container 118 and / or reusable support structure 114 and may facilitate the convenient introduction and removal of liquids and / or gases from the container. These ports may be located in any suitable position relative to container 118. For example, for some devices including sprayers, the container may include one more gas inlet port located at the bottom portion of the container. Pipes may be connected to the inlet ports and / or outlet ports to form, for example, conveying lines and harvesting lines for introducing liquids and removing liquids from the container, respectively. One or more connectors 164 may be located at the top portion of container 118 or at any other suitable position. Connectors 164 may include openings, pipes, and / or valves for adding liquids, gases, etc., or withdrawing liquids, gases, etc., from container 118, each of which may optionally include a flow sensor and / or a filter (not shown). Optionally, connectors 164 may be in fluid communication with gas inlet and outlet ports 165.

[0006] For a system comprising multiple sprayers, the control system 134 may be operatively associated with each of the sprayers and configured to operate the sprayers independently of each other. In some embodiments, the support structure 114 and / or container 118 may also include one or more ports 154 for sampling, analysis (e.g., determining the amount of dissolved gas and / or pH in a liquid) or for other purposes. These ports may be aligned with one or more access ports 156 of an optional environmental enclosure 120.

[0007] In some embodiments, the device 100 may include one or more connection ports 180 for interconnecting the interior of the reusable support structure 114 (e.g., gap 132) to the interior of the second device. Alternatively or additionally, the device may include one or more connection ports 182 adapted to connect the interior of container 118 (e.g., interior 56) to the interior of the second device. These ports may facilitate the transfer of material from interior 56 to the second device or another suitable container (e.g., a sealed bag). Transfer may be achieved, for example, by pumping material through a tube (e.g., by peristaltic pumping or by applying positive pressure to the inlet), by using gravity, and / or by applying a vacuum.

[0008] The apparatus 100 may optionally include a mixing system, such as an impeller 151 positioned within a container 118, which can be rotated (e.g., about a single axis) using a motor 152 that may be located outside (or inside) the container. The mixing system may be controlled by a control system 134. Optionally, the container and / or support structure may include a utility tower 150 that facilitates interconnection between one or more devices within the container and / or support structure and one or more pumps, controllers and / or electronics (e.g., sensor electronics, electronic interfaces, and pressurized gas controllers) or other devices. Such devices may be controlled using the control system 134.

[0009] Figure 2 Another disposable bioreactor system is shown in U.S. Patent 7,629,167, filed June 6, 2005, entitled “Disposable Bioreactor System and Methods”. Figure 2 As described more fully in the '167 patent, and briefly herein. As shown, the bioreactor 200 includes one or more ports 202 for adding or removing gas and / or fluid from the bioreactor. A harvest port or discharge port 204 is generally located at the bottom of the bag, allowing gravity to be used to guide the contents out of the bioreactor. Probes and / or sensors 206 may be integral with the side of the bioreactor, allowing the sensors and / or probes to be disposable. In one embodiment of the invention, the sensor / probe may be an optical probe that visually presents the output. Thus, the sensor / probe port 206 can be used to visually monitor the status of the sensor / probe.

[0010] A portion of the mixing system can be integrated with the bioreactor. Specifically, such as... Figure 2 As shown, a portion of the mixing system included in the bioreactor may include a portion 208 of the mixing system—an impeller plate and an impeller hub. The impeller plate is connected to a motor drive system to provide power to the impeller and also provides a seal between the motor and the interior of the bioreactor. Some embodiments of the invention provide one or more specific mixing systems that offer an inexpensive method for agitating the contents of the bioreactor. Such mixing systems may utilize materials such as HDPE (high-density polyethylene) and / or other gamma-radiative biocompatible plastics. One or more components of the mixing system may be manufactured by machining blocks of material, but may also be molded and / or cast.

[0011] Figure 3Another disposable bioreactor system is shown in U.S. Patent 9,550,969 entitled “Flexible Bag for Cultivation of Cells”. Figure 3 As described more fully in the '969 patent, and briefly herein. As shown, an inflatable bioreactor bag 1 for cell culture comprises a top sheet 2 and a bottom sheet 3 of flexible material joined together to form two end edges 4 and two side edges 5, wherein one or more baffles 6 extend from the bottom sheet 3 in a region where the shortest distance to either of the two end edges 4 (i.e., the closest end edge) is greater than approximately one-quarter of the shortest distance D between the two end edges 4. The bag may be generally rectangular, in which case the shortest distance to either of the end edges 4 (the closest one) will never be greater than D / 2 for any point on the bottom sheet 3. Thus, the baffle 6 may extend from the bottom sheet 3 in a region where the shortest distance to the end edge 4 is between approximately one-quarter and one-half of the shortest distance D between the end edges 4 (i.e., between D / 4 and D / 2).

[0012] Bag 1 is pivotally mounted to base 9 about a movable axis 7 generally parallel to end edges 4. The movable axis 7 may be located below bag 1, and the bag may be mounted on a support 8, for example, where the distance between each edge 4 and the projection of the movable axis 7 onto the bottom sheet 3 is approximately equal to D / 2. A suitable pivotable support mounted on the movable axis may be, for example, WAVE Bioreactor™ Systems (GE Healthcare). The flexible material of the top sheet 2 and bottom sheet 3 may be a polymer material, such as a plastic film or laminate having a thickness, for example, in the range of 50-500 micrometers. In addition to one or more polymer materials, the laminate may include, for example, a bath layer, which may be a polymer or an inorganic oxide or metal. In particular, the top sheet 2 may be transparent to ensure visibility inside the bag. The top sheet 2 and bottom sheet 3 are defined relative to their position during bag use, i.e., during use, the top sheet 2 is positioned above the bottom sheet 3. The top and bottom pieces can also be distinguished in that the port 11 is preferably located on the top piece 2, providing a smooth outer surface for the bottom piece 3 to rest against the support 8. The side edge 5 may be longer than the end edge 4.

[0013] The advantage of centering the baffle 6 is that when the bag is partially filled with cell suspension, inflates, and oscillates around axis 7, virtually all of the cell suspension will repeatedly pass over the baffle. This increases the stirring intensity and improves gas exchange at the air-liquid interface, while the stirring remains gentle enough not to cause any damage to the fragile cells.

[0014] In some embodiments, at least one of the baffles 6 (such as two baffles) is tubular. One advantage of this is that the baffles can be made flexible enough to allow easy packaging and storage of folded bags before use, yet rigid enough to withstand hydrodynamic forces during bag handling. Another advantage is that the tubular structure allows material to be transported through the baffles into or out of the bag. The tubular baffles can be made, for example, of elastomeric materials that allow the structure to fold during bag packaging, but are flexible enough to give full recovery of the open tubular shape when the bag is filled and / or inflated. Elastomeric materials can be, for example, cross-linked silicone rubber or other vulcanized rubber materials. If tubes with thick walls and / or high rigidity are used, the tubular baffles can also serve as pillars, keeping the top and bottom sheets of the bag separate before and during bag inflation.

[0015] Improved bioreactor or fermenter bags that can withstand sterilization, packaging, shipping, setup, and use remain necessary. Summary of the Invention

[0016] This invention includes aspects related to improved bioprocess bags and related structures, including reinforced and improved tube features. In one aspect, the invention relates to an improved bioprocess bag comprising: a bag wall defining a closed volume for retaining biological material, the bag wall including at least one inlet port and at least one outlet port; and a tube structure including a first open end near the bag wall and a second distal end, the tube extending into the closed volume, and the tube structure including a reinforcing portion near the first open end. The reinforcing section may include a pattern of sections raised relative to the outer surface of the tube, and the raised sections contact each other when the tube structure is bent. This contact prevents the tube structure from curling during shipping, installation, or operation.

[0017] In another aspect, the present invention relates to a bioprocess bag comprising: a bag wall defining a closed volume for maintaining biological material, the bag wall including at least one inlet port and at least one outlet port; and a tube structure including a first open end proximate to the bag wall and a second closed end distal to the bag wall, the tube extending into the closed volume, the tube structure including a predominantly cylindrical inner wall and a notch extending at least a portion of the length of the inner wall, wherein the notch defines a channel allowing air to flow out when a probe is inserted into the tube structure.

[0018] These aspects of the bioprocess bag may include other features used in mixers, bioreactors, and other related applications. Those features may include an impeller, heater, and gas outlet. The tube structure may be adapted to receive probes, such as probes with thermocouples or resistance temperature detectors (RTDs). Alternatively or additionally, the tube structure may be adapted to receive a spray bar. It is contemplated that any number of tube structures according to embodiments of the invention may be used together with or in combination with other tube structures in a single bioprocess bag. The tube structure may include a predominantly cylindrical inner wall and a notch extending at least a portion of the length of the inner wall. The notch may be adapted to define a channel allowing airflow when a probe is inserted into the tube structure.

[0019] In one aspect, the invention includes a reinforced probe housing comprising: a distal portion having a first outer diameter and a first inner diameter having a defined thickness, the inner diameter being adapted to receive a probe, the probe housing having a closed end adjacent to the distal portion; and a reinforcing portion sharing the same inner diameter as the distal portion, wherein the outer diameter of the reinforcing portion includes raised sections that provide reinforcement by contacting each other when the probe housing is bent, the probe housing having an open end adjacent to the reinforcing portion, the open end being adapted to receive a probe. The reinforced probe housing may be included in a bioprocess bag or used independently. The probe housing may be adapted to receive a thermocouple or a resistance temperature detector (RTD). Attached Figure Description

[0020] Figure 1 An example of a prior art single-use large-scale bioreactor system is shown.

[0021] Figure 2 An example of a disposable bioreactor bag from the prior art is shown.

[0022] Figure 3 An example of a disposable bioreactor bag from another prior art is shown.

[0023] Figure 4 A thermocouple sheath tube with a thermocouple sheath probe inserted therein is shown according to an aspect of the invention.

[0024] Figure 5 A partially enlarged perspective view of the thermocouple bushing according to an aspect of the invention is shown.

[0025] Figure 6 A bent thermocouple sleeve according to an aspect of the invention is shown.

[0026] Figure 7 A cross-section of a portion of the thermocouple bushing according to an aspect of the invention is shown. Detailed Implementation

[0027] The inventors have discovered that several components of the internal structure of bioreactor or fermenter bags are susceptible to damage or detectable damage due to sterilization and packaging, and further damage to these components can occur during transport, setup, or use. Sterilizing the bags with gamma radiation can, in some cases, exacerbate the problem by melting adjacent material components or freezing the material into a bent shape. Regarding the internal portions of the bag, given the need to maintain sterility without compromising the bag structure, these problems are sometimes compounded by the inability to inspect the internal portions. Detection of damage to the internal portions can render the entire bag unusable. The tubular structure can be adapted to receive probes (such as thermocouples or resistance temperature detectors (RTDs)) or spray bars. Alternatively or additionally, the tubular structure can be adapted to introduce or remove material from the bioprocess bag.

[0028] The internal bioreactor bag structures most susceptible to this type of damage, or the damage that is most easily detected, are tubular structures. These tubular structures inside the bag can become bent, rolled up, or folded at some point in the supply chain.

[0029] Thermocouple sheaths obtained in existing bioreactor bags are a particular type of structure that is susceptible to damage. An example of such a thermocouple sheath is described in U.S. Patent 6,599,012 entitled "Thermowell Adapter," the disclosure of which regarding its thermocouple sheath structure is incorporated herein by reference. The thermocouple sheath is a straight tube made of a plastic material that allows the thermocouple sheath to be inserted into the bioreactor bag without disrupting the bag's seal. Typically, the thermocouple sheath is made of molded plastic or rubber material. A rigid insert is used to prevent the tube from folding and self-sealing internally during gamma radiation. In some cases, the rigid insert may break and puncture the tube. Without the rigid insert, the tube may kink and remain sealed during gamma radiation. When the temperature sensor 405 is inserted, it may puncture the tube.

[0030] exist Figure 4 In one embodiment shown, the thermocouple sheath 400 includes a distal portion 401 and a reinforcing portion 402. The distal portion 401 may have a length on the order of approximately 1 inch (2.54 cm), but the distal portion 401 may be longer or shorter depending on the user's needs. The distal portion of the thermocouple sheath 400 is generally thinner than the reinforcing portion 402 and is designed to allow enhanced heat flow from the interior portion of the bioreactor (not shown) to the distal end 401 of the thermocouple sheath when it is inserted into the thermocouple sheath 400, such as... Figure 4 As shown in the diagram, the reinforcing portion 402 is defined by a rib portion 402a, which is spaced apart by separating portions 402b (at intervals). The reinforcing portion 402 provides greater structural stability to the thermocouple socket 400.

[0031] The thermocouple sheath 400 may also include an optional base portion 403 attached to the thermocouple sheath flange 404, which can be used to form a seal with a bioreactor bag (not shown). Alternatively, the thermocouple sheath may be directly molded into the bioreactor bag. When the thermocouple sheath is inserted into the thermocouple sheath 400, the base portion provides additional structural support for the thermocouple sheath.

[0032] Figure 5 A partially enlarged perspective view of the thermocouple socket 400 is shown. The end portion 401 of the thermocouple socket 400 has a shape that maximizes the thermal exposure of the thermocouple socket (not shown) when inserted into the thermocouple socket 400. Rib portions 402a are preferably cylindrical in construction and spaced apart by separation portions 402b, which may have the same geometry as the end portion. In one aspect, the rib portions 402a are designed to have a length 406, and the separation portions 402b are designed to have a length 407. In the cylindrical construction, the rib portions project outward to a height 408, which is defined by the difference between the outer diameter of the rib portions 402a and the outer diameter of the separation portions 402b. The thermocouple socket 400 has an inner diameter 409 adapted to tightly match the outer diameter of a thermocouple socket probe (not shown).

[0033] The reinforcing portion 402 may be defined by several geometries to achieve one or more objectives of the invention. For example, in one variation, the height 408 of the rib portion 402a may gradually increase in a continuous or stepped manner in the direction from the end portion 401 to the base portion 403. In some cases, a gradual increase in height 408 may result in a more continuous transition from the outer diameter of the end portion 401 to the base portion 403. Various other reinforcing geometries are within the scope of the invention.

[0034] Figure 6 A thermocouple sheath with an end 401 and a base 404 connected to a flange 404 is shown, bent to illustrate the operation of a reinforced portion according to an aspect of the invention. Ribs 402a toward the interior of the bend 410 are configured to contact each other to resist further deformation of the thermocouple sheath 400. In a preferred aspect, the contact between the ribs 402a reinforces the entire structure when the tube is bent. This reinforcement protects the internal conduit 409 of the thermocouple sheath from folding, curling, or other deformation. In this aspect, the length 407 of the spacer portion 402b between adjacent ribs 402a increases toward the exterior of the bend 411.

[0035] Figure 7A cutout of the thermocouple socket 400 is shown, illustrating details of the inner diameter of the thermocouple socket 400 according to an alternative aspect of the invention. As discussed above, the thermocouple socket 400 has an inner diameter 409 adapted to a cylindrical thermocouple probe (not shown). In this aspect, the inner wall of the thermocouple socket 400 is provided with a notch 413, wherein the inner diameter increases to 412 in length 414. The notch formation allows an internal channel for air to pass through during sensor insertion and removal. The notch 413 allows air to escape from the interior of the thermocouple socket 400 when the thermocouple probe is inserted into the thermocouple socket 400. It will be understood that the exact geometry of the notch can vary to achieve one or more objectives of the invention. Furthermore, the notch can be designed to extend from the edge of the thermocouple socket 400 (or optionally the edge of the thermocouple socket base 403) to the end of the thermocouple socket end 401, extending the entire length of the inner diameter of the thermocouple socket cavity. Alternatively, the notch can extend over a defined portion of the inner diameter of the thermocouple socket 400. In one aspect, the notch 413 extends through the reinforcing portion and terminates at a point in the end portion 401. Alternatively, the notch 413 extends to the edge of the reinforcing portion 402. In another aspect, the notch terminates at a point within the reinforcing portion 402.

[0036] It should be understood that the reinforcing portion 402 described above with respect to the thermocouple insert 400 can be applied to any other structure within a bioreactor or fermenter bag. Rib-like structures on the tube can be applied to other applications in single-use bioreactor bags where there is a possibility of kinking and obstruction of fluid flow for the tube. These areas include any internal tubes for fluid transfer, such as internal spray tubes or internal suction tubes that directly deliver fluid into a bulk process fluid. This can be used on top of the bioreactor for discharge filter lines, as these typically require bending to accommodate and support within the heater while still providing a discharge path for condensate back to the reactor. Any additional lines supported and draped on a rigid rod can benefit from small sections of ribbed tube (in which it is bent). This prevents tube fattening, which can cause flow restriction and high-pressure events. Like the thermocouple insert, these portions can be directly molded into the bag or attached to the bag using flanges, as shown with the thermocouple insert above.

[0037] It should be understood that the reinforced tube structure of the present invention will be particularly useful when added to single-use bioreactors. Ribbing along the tube (e.g., the molded thermocouple sheath 400) prevents kinking and self-sealing due to gamma radiation. When used with a thermocouple sheath probe, the temperature sensor portion of the probe is located in a ~1-inch (~2.54 cm) section at the end 401 of the thermocouple sheath, and the thermocouple sheath 400 will have a thin-walled section to facilitate a rapid heat transfer response. An internal channel (or notch 413) will allow air to pass through during the insertion and removal of the temperature sensor 405.

[0038] The reinforced tubular structure of the present invention (and particularly the reinforced thermocouple sheath) can be applied to any disposable bioprocess bag (and particularly those systems made of flexible materials such as plastics). Figure 1-3 Any of the disposable bags shown may be modified to include an enhanced internal tubular structure using embodiments of the invention. Additionally, any of the bags shown may be modified to include an enhanced thermocouple sheath according to aspects of the invention.

[0039] Other embodiments and uses of the invention will become apparent to those skilled in the art upon consideration of the description and practice of the invention disclosed herein. All references cited herein (including all cited patents and patent applications) are specifically and entirely incorporated herein by reference where permissible. It is intended that the description and examples be considered exemplary only, wherein the true scope and spirit of the invention are indicated by the following claims.

Claims

1. A bioprocess bag comprising: a bag wall defining an enclosed volume for holding a biological material, the bag wall comprising at least one inlet port (142) and at least one outlet port (146); and a tube structure comprising a first open end proximate the bag wall and a second distal end, the tube structure extending into the enclosed volume, and the tube structure comprising a reinforced portion (402) proximate the first open end, wherein the reinforced portion (402) comprises a ribbed structure, wherein the height (408) of the ribbed segments gradually increases in a continuous or stepped manner in a direction from a tip portion (401) to a base portion (403).

2. The bioprocess bag of claim 1, wherein, the second distal end is a closed end, and the tube structure is adapted to receive a probe (206) for measuring one or more properties of the biological material.

3. The bioprocess bag of claim 2, wherein, the probe (206) comprises a thermocouple or a resistance temperature detector (RTD).

4. The bioprocess bag according to any one of claims 1 to 3, characterized in that, the tube structure comprises a first open end proximate the bag wall and a second distal end extending into the enclosed volume, the reinforced portion (402) being proximate the first open end.

5. The bioprocess bag according to any one of claims 1 to 3, characterized in that, the tube structure is adapted to receive a spray rod.

6. The bioprocess bag of claim 1, wherein, the ribbed structure comprises a pattern of segments that are raised relative to the outer surface of the tube, and the raised segments contact each other when the tube structure is bent.

7. The bioprocess bag of claim 6, wherein, the contacting of the raised segments prevents the tube structure from kinking.

8. The bioprocess bag according to any one of claims 1 to 3, characterized in that, the tube structure comprises a primarily cylindrical inner wall and a notch (413) extending at least a portion of the length of the inner wall.

9. The bioprocess bag of claim 8, wherein, the notch (413) defines a passageway that allows air to escape when a probe is inserted into the tube structure.

10. The bioprocess bag of any one of claims 1 to 3, wherein, the bag wall comprises one or more of the following: an impeller (151), a heater, and / or a gas outlet.

11. The bioprocess bag according to any one of claims 1 to 3, characterized in that, further comprising a base portion (403) attached to a thermocouple insert collar flange (404) for forming a seal with the bioprocess bag.

12. The bioprocess bag of any one of claims 1 to 3, wherein, further comprising a probe housing, the probe housing comprising; a tip portion (401) having a first outer diameter defining a thickness and a first inner diameter, the inner diameter being adapted to receive a probe (206), the probe housing having a closed end proximate the tip portion (401); a reinforced portion (402) sharing the same inner diameter as the tip portion (401), wherein the outer diameter of the reinforced portion (402) comprises raised segments that provide reinforcement by contacting each other when the probe housing is bent, the probe housing having an open end proximate the reinforced portion (402) adapted to receive the probe (206).

13. The bioprocess bag of claim 12, wherein, the probe (206) comprises a thermocouple or a resistance temperature detector (RTD).

14. The bioprocess bag of claim 12, wherein, the tip portion (401) has a shape that maximizes the exposure of the probe (206) to heat when inserted into the inner diameter.

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