Systems and methods for affinity cell selection

Abstract

Biological fluid processing system for cell affinity selection and method of operating the system. The system has a fluid processor that operates a fluid circuit having a process vessel, a source vessel filled with a biological fluid, a separation chamber configured to separate the biological fluid from the source vessel into at least two volumes of material, and a selection column connected with the fluid processor. Also disclosed are modified cells for treating a disease, ailment, or medical condition in a patient. The modified cells are produced using a biological fluid processing system. A separation chamber is configured to separate the biological fluid from the source vessel into at least two volumes of material, and a process vessel is used to collect treated cells from the separation chamber. A selection column is connected with the fluid circuit for further cell affinity selection processing of the cells.

Description

[0001] Cross-references to related applications

[0002] This application claims the benefits of U.S. Provisional Application No. 63 / 613,500, filed December 21, 2023, and U.S. Provisional Application No. 63 / 685,540, filed August 21, 2024, the contents of which are incorporated herein by reference in their entirety. Technical Field

[0003] This disclosure generally relates to apparatus and methods for selecting affinity cells. More specifically, this disclosure relates to fluid processing systems for handling biological fluids and performing cell selection. Even more specifically, this disclosure relates to systems and methods for modifying cells, modified cells produced by such fluid processing systems, and systems and methods for using such cells to treat a variety of diseases, ailments, or medical conditions. Background Technology

[0004] The handling of biological fluids (such as blood or blood components) typically involves the use of reusable processing equipment (“hardware”) and a disposable fluid loop adapted to be installed in or otherwise associated with the reusable equipment. The fluid loop typically includes a container, such as a plastic bag, and associated tubing that defines a flow path through the loop. The disposable fluid loop may also include one or more separation devices, wherein the biological fluid / cells may be separated into two or more components, washed, or otherwise processed.

[0005] Separation devices can be based on centrifugation, membrane separation, or other procedures (e.g., using a magnetic field) to separate biological fluids, for example, to select and / or separate target cells. Certain systems may use combinations of these separation devices, such as those described in U.S. Publications Nos. 2017 / 0315121 and 2018 / 0172685 and U.S. Patent No. 11,478,755, which are incorporated herein by reference in their entirety. Summary of the Invention

[0006] This disclosure provides systems and methods for affinity cell selection in biological fluids.

[0007] Systems and methods for affinity cell selection are disclosed. The system includes a fluid handling system using a disposable fluid loop with a cell selection column, which can be configured for manual or automated use of the cell selection column. The system is capable of preparing single-harvest (or other heterogeneous cell populations) of material for cell selection by automatically washing, incubating, and adjusting the volume / concentration of the cell material. The system further facilitates the removal of target / non-target cell fractions by adding a selection reagent to the cell selection column and subsequently washing / resuspending the target / non-target fractions at a desired cell concentration for downstream use.

[0008] The method is highly configurable, allowing operators to define custom values ​​and processing parameters to accommodate a wide range of applications, including: different cell starting materials, different target / non-target cell populations, purity / recovery profiles, and reagent selection.

[0009] In a first aspect, a biofluid processing system for cell affinity selection is provided, the biofluid processing system comprising a fluid processor having reusable hardware for operating a disposable fluid loop connected to a source container and having a biofluid; a separation chamber configured to separate the biofluid from the source container into at least two volumes of material; a processing container for collecting washed and harvested cells from the separation chamber; and a selection column connected to the fluid loop for further cell affinity selection processing of the cells from the processing container.

[0010] In a second aspect, a method is provided for operating a biofluid processing system for cell affinity selection, the method comprising: obtaining a biofluid processing system including a fluid processor that operates a disposable fluid loop connected to a source container having biofluid and configured to separate biofluid from the source container into at least two volumes of material; a processing container connected to the fluid processor along a fluid path; and a selection column connected to the fluid loop. The method further includes performing a pretreatment state, the pretreatment state further comprising: selecting a program scheme, performing program setup, mounting the disposable fluid loop to the fluid processor; attaching a plurality of containers to the fluid processor; running a fluid loop solution prime including the separation chamber; attaching the source container to the fluid processor; and running a fluid loop source prime. The method further includes a processing state, which further includes: treating a biofluid from the source container in a separation chamber via the disposable fluid loop, including one or more cycles of washing, harvesting, and diluting the biofluid; rinsing the source container; diluting the harvested cells; collecting the harvested cells in the processing container; draining the solution from the selection column; adding an antibody to the selection column; transferring the harvested cells from the processing container to the selection column; transferring a non-target fraction containing unbound cells suspended in the selection column to waste; diluting the material in the selection column; refilling the separation chamber for processing the material in the selection column; loading the target fraction from the selection column into the separation chamber; washing and harvesting the cells from the selection column; harvesting the selected cells as the final product in at least one final container; and a post-processing state, which further includes: sealing the at least one final container and removing the final product; and removing the disposable fluid loop.

[0011] In a third aspect, a method is provided for operating a biofluid processing system for cell affinity selection, wherein the system includes a fluid processor that operates a disposable fluid loop connected to a source container filled with biofluid and configured to separate biofluid from the source container into at least two volumes of material; a processing container connected to the fluid processor along a fluid path; and a selection column connected to the fluid loop. The method includes a pretreatment state, a processing state, and a post-treatment state. The pretreatment state further includes: selecting a program scheme; setting up a program; mounting the disposable fluid loop to the fluid processor; attaching at least a first solution container containing a separation / processing buffer, a second solution container containing a resuspension buffer, and a third solution container containing a release buffer to the fluid processor; performing a fluid loop solution pre-fill; attaching the source container to the fluid processor; and performing a fluid loop source pre-fill. The processing state further includes: processing the biofluid from the source container in the separation chamber via the disposable fluid loop, including one or more cycles of washing, harvesting, and diluting the biofluid; rinsing the source container; diluting the harvested cells; collecting the harvested cells in the processing container; draining the buffer solution from the selection column; diluting the material in the selection column by adding an antibody to the selection column; incubating with the antibody in the selection column; transferring the harvested cells from the processing container to the selection column; transferring the non-target fraction containing unbound cells suspended in the selection column to waste; diluting the material in the selection column by transferring biotin to the selection column; incubating; refilling the separation chamber for processing the material in the selection column; loading the target fraction from the selection column into the separation chamber; washing and harvesting the cells processed in the separation chamber; rinsing the source container; harvesting the selected cells as the final product in at least one final container; and diluting the harvested cells in the final container. The post-processing state further includes: sealing the at least one final container and removing the final product, and removing the disposable fluid loop.

[0012] In a fourth aspect, modified cells are provided for treating a patient's disease, ailment, or medical condition, wherein the modified cells are produced using a biofluid processing system comprising a fluid processor having reusable hardware for operating a disposable fluid loop connected to a source container and having a biofluid; a separation chamber configured to separate the biofluid from the source container into at least two volumes of material; a processing container for collecting washed and harvested cells from the separation chamber; and a selection column connected to the fluid loop for further cell affinity selection of the cells from the processing container.

[0013] In a fifth aspect, modified cells are provided for treating a patient's disease, ailment, or medical condition, wherein the modified cells are produced using a method comprising: obtaining a biofluid processing system comprising a fluid processor that operates a disposable fluid loop, the disposable fluid loop being connectable to a source container having biofluid and configured to separate biofluid from the source container into at least two volumes of material; a processing container connected to the fluid processor along a fluid path; and a selection column connected to the fluid loop; entering a pretreatment state, the pretreatment state further comprising: selecting a program scheme, setting a program, mounting the disposable fluid loop to the fluid processor; attaching a plurality of containers to the fluid processor; running a fluid loop solution pre-charge containing the separation chamber; attaching the source container to the fluid processor; and running a fluid loop source pre-charge; entering a processing state, the processing state further comprising: circulating fluid in the separation chamber. The biological fluid from the source container is processed through the disposable fluid loop, including one or more cycles of washing, harvesting, and diluting the biological fluid; rinsing the source container; diluting the harvested cells; collecting the harvested cells in the processing container; draining the solution from the selection column; adding an antibody to the selection column; transferring the harvested cells from the processing container to the selection column; transferring the non-target fraction containing unbound cells suspended in the selection column to waste; diluting the material in the selection column; refilling the separation chamber for processing the material in the selection column; loading the target fraction from the selection column into the separation chamber; washing and harvesting the cells from the selection column; harvesting the selected cells as the final product in at least one final container; and performing a post-processing state, the post-processing state further including: sealing the at least one final container and removing the final product; and removing the disposable fluid loop.

[0014] In a sixth aspect, modified cells are provided for treating a patient's disease, ailment, or medical condition, wherein the modified cells are generated using a method of operating a biofluidic processing system for cell affinity selection, wherein the system includes a fluid processor that operates a disposable fluid loop connected to a source container filled with biofluid and configured to separate the biofluid from the source container into at least two volumes of material; a processing container connected to the fluid processor along a fluid path; and a selection column connected to the fluid loop. The method includes a pretreatment state, a processing state, and a post-treatment state; the pretreatment state further includes: selecting a program scheme; program setup; mounting the disposable fluid loop onto the fluid processor; attaching at least a first solution container containing a separation / processing buffer, a second solution container containing a resuspension buffer, and a third solution container containing a release buffer to the fluid processor; performing fluid loop solution pre-filling; attaching the source container to the fluid processor; and performing fluid loop source pre-filling. The processing state further includes: processing the biofluid from the source container in the separation chamber via the disposable fluid loop, including one or more cycles of washing, harvesting, and diluting the biofluid; rinsing the source container; diluting the harvested cells; collecting the harvested cells in the processing container; removing the buffer solution from the selection column; diluting the material in the selection column by adding an antibody to the selection column; incubating with the antibody in the selection column; transferring the harvested cells from the processing container to the selection column; and removing unbound cells containing cells that remain suspended in the selection column. The process includes: transferring non-target fractions to waste; diluting the material in the selection column by transferring biotin into the selection column; incubation; refilling the separation chamber for processing the material in the selection column; loading the target fractions from the selection column into the separation chamber; washing and harvesting the cells processed in the separation chamber; rinsing the source container; harvesting the selected cells as the final product in at least one final container; and diluting the harvested cells in the final container. The post-processing state further includes: sealing the at least one final container and removing the final product; and removing the disposable fluid loop. Attached Figure Description

[0015] Figure 1 This is a perspective view of one implementation of a fluid handling system for processing biological fluids and performing affinity cell selection.

[0016] Figure 2 Is with Figure 1 A block diagram of the controller used in conjunction with other components of the system.

[0017] Figures 3A to 3CThis shows the operation. Figure 1 A flowchart detailing the steps of the system and the methods for incorporating systems therein. Detailed Implementation

[0018] The following provides a more detailed description of the apparatus and methods of using the apparatus according to this disclosure. It should be understood that the following description of specific apparatuses and methods is intended to be exemplary and not to exhaustively list all possible variations or applications. Therefore, the scope of this disclosure is not intended to be limiting and should be understood to cover variations or implementations that would occur to those skilled in the art.

[0019] Figure 1 An exemplary fluid handling system 10 may include many of the properties and structures present in the system shown in U.S. Patent No. 11,478,755, which is owned by the applicant Fenwal, Inc. and is incorporated herein in its entirety.

[0020] Turning Figure 1 This illustration shows one embodiment of a fluid processing system 10 for handling biological fluids, characterized by affinity cell selection. System 10 includes a fluid processor 12 connectable to a source container 14 filled with biological fluid and configured to separate the biological fluid from the source container 14 into at least two material streams. "Biofluid" includes, but is not limited to, blood and blood components, and "cells" or "biological cells" include, but are not limited to, blood cells such as red blood cells, white blood cells, and platelets. Processor 12 is connected to an additional fluid processing container 16, which may be of flexible or rigid walls. The processing container 16 may initially be empty and rest on a tray or base plate 18 of a housing 20. Housing 20 may be open or may include a door or lid providing a closed state. The tray 18 allows for containment, mixing by shaking, etc., and may include clamps to help hold the container 16 in place on the tray 18. Container 16 is also positioned along a fluid path 22.

[0021] Processor 12 includes a disposable fluid loop (also referred to as a set or kit) 24, which is used in combination with reusable processing equipment or hardware 26. Fluid loop 24 can be a “closed” system or loop, where the system’s interior (e.g., flow paths, containers, etc.) is not exposed to or “open” to the external environment. Fluid loop 24 can be referred to as closed even when additional containers are attached to it (e.g., before or during a program). Fluid loop 24 includes fluid paths 22.

[0022] Control unit (or controller) 28 (see Figure 2The system 10 is coupled to a processor 12, which includes a fluid loop 24 and hardware 26. The controller 28 is configured to operate the processor 12 and hardware 26 according to a program or process to cause the system 10 to produce or generate products. For example, the system 10 may consist of a fluid processing system paired with a cell selection column 30. The cell selection column 30 may be integrated into or added to the fluid loop 24. As further discussed herein, the system 10 is capable of preparing single-harvest (or other heterogeneous cell populations) of material for cell selection by automatically washing, incubating, and adjusting the volume / concentration of the cell material. The system 10 can also facilitate the removal of target / non-target cell fractions by adding a selection reagent to the cell selection column 30 and subsequently washing / resuspending the target / non-target fractions at a desired cell concentration for downstream use. Target cells may initially be separated from the biofluid by the processor 12.

[0023] like Figure 1 As shown, a disposable fluid circuit 24 can be connected to a source container 14 for fluids, particularly biological fluids. In this example, the disposable fluid circuit 24 includes a separation chamber 31, exemplified as a rotating membrane separator, for processing the fluid received from the source container 14. The flow of fluid from the source container 14, through the rotating membrane separator 31, and to one or more containers (e.g., processing container 16) is achieved using first and second syringes 32, 34, which are in fluid communication with the source container 14, the rotating membrane separator (or simply the rotating membrane) 31, the processing container 16, and the cell selection column 30. The syringes 32, 34 may also be in fluid communication with a plurality of other containers 36, 38, 40.

[0024] For example, container 36 is initially empty and can be used to receive waste generated during processing; container 38 may contain processing or resuspension buffers or reagents; and container 40 may contain release buffers or reagents, such as biotin release agents. Selective column 30 may initially contain matrix or resin material and may contain storage buffers, if desired, to maintain the wetting of such material.

[0025] Within fluid loop 24, fluid flow between containers 14, 16, 36, 38, 40, rotating membrane 31, syringes 32, 34, and cell selection column 30 is controlled using a flow control box 42, which can be connected to each of the foregoing via tubing (or "line"). Additionally, box 42 may contain internal flow paths partially defined by multiple individual channels or tubing, which may be contained within and defined by a structure (e.g., housing) of box 42. Channels may be connected at multiple selectable joints that control fluid flow from one channel to another. These selectable joints may also be referred to as valves, valve stations, or clamps, as they provide controlled pathways between channels. Box 42 may also contain sensor stations through which sensors can be correlated with the flow paths within box 42 to determine flow characteristics, such as pressure. Preferably, the length of each line and channel is kept as short as possible to further minimize the internal volume of fluid loop 24.

[0026] like Figure 1 As illustrated in the examples, the separation chamber or rotating membrane 31 and syringes 32, 34 can be integrally formed as part of the housing 42 (i.e., integrated with the housing 42) to further reduce the tubing volume associated with the fluid circuit or kit 24. According to other embodiments, the rotating membrane 31 and / or syringes 32, 34 can be attached to the remainder of the fluid circuit 24 during use, as may be the case with one or more of the containers 14, 16, 36, 38, 40. Similarly, as... Figure 1 As illustrated in the example, containers 36 and 16 can be integrally formed with box 42.

[0027] like Figure 1 As shown, the reusable hardware component (or simply reusable hardware) 26 includes a driver 43 for the rotary membrane separator 31, injection pumps 44, 46 for each corresponding injector 32, 34, and a control box interface 48 associated with the flow control box 42 when the fluid circuit 24 is provided on the hardware 26 (e.g., mounted on the hardware 26). The box interface 48 includes actuators and sensors associated with the clamps and sensor stations of the flow control box 42, and is configured to operate the clamps or sense the characteristics of the fluid passing through them.

[0028] Reusable hardware 26 is coupled to controller 28, which is configured to control the operation of system 10, for example, using the following text regarding... Figures 3A to 3C The flowchart in the document explains the operating method. For example... Figure 2As shown, controller 28 may include microprocessor 50, which may include multiple physical and / or virtual processors. According to other embodiments, controller 28 may include one or more circuits designed to perform the actions described herein. Therefore, controller 28 may include microprocessor 50 and other circuitry or circuit systems. Additionally, controller 28 may include at least one memory 52. ​​Instructions for programming microprocessor 50 may be stored on at least one memory 52 associated with microprocessor 50. The at least one memory 52 may include one or more tangible, non-transitory computer-readable memories storing computer-executable instructions that, when executed by microprocessor 50, cause microprocessor 50 to perform one or more actions as described herein.

[0029] As described above, controller 28 can be coupled (i.e., directly or indirectly) to devices of reusable hardware 26, such as rotary membrane actuator 43, first syringe pump 44, second syringe pump 46, and cartridge interface 48. Controller 28 can operate each of these devices, each of which can be a combination of other devices or equipment, to allow fluid to flow through fluid loop 24 associated with hardware 26, for example, to allow fluid to flow from source container 14 through rotary membrane 31 and ultimately into processing container 16.

[0030] For example, controller 28 may be programmed to perform a process or procedure according to a scheme, such as washing the specific cells contained in the biofluid within source container 14 before directing the specific cells to processing container 16. Controller 28 may also be programmed to perform other actions, such as testing fluid circuit 24, pre-filling fluid circuit 24, rinsing portions of circuit 24 after washing has been performed, and adding other components to the cell-containing fluid or completing other steps before dispensing the fluid to processing container 16.

[0031] For example Figure 2 As illustrated in the examples, controller 28 may be coupled to one or more of the aforementioned structures or devices, for example, to receive information (e.g., in the form of signals) from these structures or to provide commands (e.g., in the form of signals) to these structures to control their operation. As illustrated in the examples, controller 28 may be coupled to at least one input device 54 to receive information from that device, and to at least one output device 56 to provide information to that device. At least one input device 54 may comprise a plurality of different devices according to the embodiments described herein. For example, input device 54 may include a keyboard or keypad through which a user provides information and / or instructions to controller 28.

[0032] Alternatively, the input device 54 can be a touchscreen, for example, which can be used in conjunction with an output device 56 in the form of a video display. For example, Figure 1 The embodiments illustrated herein include a touch-sensitive display 58 mounted to the front panel of the housing 60 of the fluid processor 12 of system 10. The input device may also include a reader or scanner, such as a barcode reader, scanner, or RFID reader 62. According to other embodiments, the input device 54 may be in the form of a computer device that allows the cell processing system 10, including the controller 28, to communicate with other processing systems via a local area network (whether via wires, cables, etc., or wirelessly), or via a local area network, wide area network, or the Internet, with other cell processing systems or other computer devices (e.g., servers). According to such embodiments, the input device may alternatively include an internal transmitter / receiver device.

[0033] The controller 28 may also be coupled to alternative devices, such as dispensers for automated injection, for example, dispensers for automatically injecting antibodies into cell selection columns 30. As another example, the controller 28 may be coupled to a device for agitating the fluid in the processing container 16. To allow agitation of the fluid in the processing container 16, the housing 20 may be mounted on a shaft 64 suspended from a motor located within the housing 60 of the processor 12. The motor can rotate the shaft 64 in opposite directions about the axis of the shaft. Thus, the fluid in the processing container 16, resting on the tray 18 of the housing 20, can be agitated in an oscillatory manner.

[0034] Mounting the housing 20 on the shaft 64 also allows the processing container 16 to be angled relative to a horizontal position, rather than being configured such that the container is horizontal. For example, the tray 18 of the housing 20 can be oriented at a 30-degree angle relative to a horizontal position to influence the fluid in the processing container 16. Of course, this can also be achieved in a separate form from the housing 20 by providing a mechanism (e.g., a pivot) that allows the surface of the housing 20 (or tray 18) to be oriented at a variable angle or a mechanism (e.g., an inclined plane) that allows it to be oriented at a specific angle.

[0035] System 10 can be automated or operated automatically. That is, system 10 (and in particular controller 28) can be programmed to perform the processing steps of the treatment method without requiring extensive operator / user intervention. Of course, even in the automated system of this disclosure, it will be understood that user activities may be involved, including loading disposable fluid loops and inputting processing parameters. Additional manual steps may also be required, such as connecting additional fluid sources or containers, and there may be opportunities to reconfigure certain steps in the process. However, the reusable device of system 10 can process biological fluids through the disposable loops described below without extensive user intervention.

[0036] Generally, a detailed description of the method of operating the system begins with a method comprising multiple steps within stages or states including preprocessing 100, processing 200, and postprocessing 300. Flowcharts representing some alternative embodiments of the steps in these states are shown in the figures, for example... Figure 3A Preprocessing 100 Figure 3B Processing 200 and Figure 3C Post-processing 300.

[0037] The user can first activate (e.g., turn on) hardware 26. Hardware 26, in conjunction with controller 28, can perform self-calibration checks, including checking pumps 44, 46, and other components. Similar self-calibration checks can be performed relative to a specific device (e.g., an input or output device) or when the user activates hardware 26.

[0038] like Figure 3A As shown, preprocessing 100 begins with scheme selection at box 102. In this step, the fluid processing system 10 displays a list of schemes from which to select to run the cell processing procedure, for example on touchscreen 58. The relevant scheme parameters for this step may include the corresponding scheme ID and scheme description.

[0039] Next, at box 104, the method includes program setup. During program setup for the selected scheme, system controller 28 prompts the user to input information, which may include, for example, identifiers, consumables, and source composition related to the specific selected program to be executed. Such information can be input via input device 54. For this step 104, the relevant scheme parameters may include more specific information depending on the program, such as: the required program ID, user ID, and source ID, and, for example, identification of target cells, cell components, component retention flags (for secondary washing and post-selection washing), primary set reference number, primary set confirm reference configuration, solution 1 name, solution 1 reference number, solution 2 name, solution 2 reference number, solution 3 name, solution 3 reference number, and the maximum fill volume (mL) of the final product bag. In addition, the controller may prompt the user to input or modify process parameters related to the functionality of hardware 26 using input device 54, including, for example, but not limited to, the volume of cell suspension to be processed, the number of cycles to be executed, etc.

[0040] Preprocessing 100 then includes a kit installation step at box 106. This involves the controller 28 of the fluid handling system 10 prompting the user to install the disposable fluid circuit 24 (also referred to as a disposable kit, cassette, or box) onto the hardware 26 of the processor 12 of the fluid handling system 10. During this step, the fluid handling system 10 loads the fluid circuit 24 and seals it in place. Once the kit 24 is installed, an installation check is performed at box 108. In this step, the system 10 checks the correct installation of multiple components of the disposable kit 24 and verifies the integrity of the kit.

[0041] In the illustrated example, pretreatment 100 continues at frame 110 with the attachment of solution 1, where system 10 prompts the user to attach solution 1 to disposable kit 24. Solution 1 may be a separation / processing buffer or reagent, which may be held in a container (e.g., container 38). Relevant protocol parameters for this step may include the volume of solution 1 being attached. When attaching the solution, the connection, such as a tube-to-container connection, may be made via a pin connector, a sterile connector, or other suitable connection means.

[0042] The next step at box 112 is the attachment of solution 2, where system 10 prompts the user to attach solution 2 to disposable kit 24. Solution 2 may be, for example, a resuspension buffer or a reagent, such as a salt solution, which may be held in a container (e.g., container 40). Relevant protocol parameters for this step may include the volume of the attached solution 2.

[0043] Figure 3A The next step shown is the attachment of solution 3 at box 114. In this step, system 10 prompts the user to attach solution 3 to disposable kit 24. Solution 3 can be, for example, a release buffer or reagent, one example of which can be a solution containing biotin, which can be held in a container (e.g., container 40). Relevant protocol parameters for this step may include the connection point of solution 3 and the volume of solution 3 to be attached. System 10 may also be configured alternatively to attach solution 3 prior to another step in the procedure. Depending on the desired treatment, optional steps may include adding additional solutions. If applicable, system 10 may prompt the user to attach a container of such additional solution to disposable kit 24.

[0044] Once the user has connected the appropriate solution, pretreatment 100 continues with solution pre-filling at box 116. The controller 28 of the fluid handling system 10 pre-fills multiple portions of the fluid loop or disposable kit 24 to (1) check for proper installation of the lines and (2) purge air from the lines and rotating membrane separator 31. In one exemplary embodiment, the fluid loop 24 may be pre-filled with brine, but other biocompatible aqueous solutions may also be used.

[0045] The next step at box 118 is to attach the source, where the controller 28 of system 10 prompts the user to attach the source product to kit 24. The source product may be a biological fluid / cell recently obtained through single-collection, leukocyte separation or other heterogeneous cell populations, material refrigerated overnight or previously frozen, etc., and may be held, for example, in source container 14.

[0046] Following this is source prefill at box 120, whereby the system prefills the line leading to the source product in source container 14 to purge air from the line. Optionally, this state can also be used to pre-dilute the source product, if desired. Relevant protocol parameters for this step may include source prefill volume (mL) and source prefill flow rate (mL / min). At this stage, the method may include source prefill pause at box 122, where system 10 may be configured to pause upon completion of source prefill step 120 to agitate or mix the source product container 14 before transitioning to the next state. Therefore, relevant protocol parameters for this step may include a post-prefill pause configuration and a post-prefill pause text.

[0047] After completion Figure 3A After the preprocessing step 100 illustrated in the example, the method enters the processing state 200, as shown below. Figure 3B As shown in the image. Figure 3B Including circled annotations 1 to 6, which correspond to Figure 3C The diagrams shown are related to the processing path that will be followed, such as when to perform a recurring group of operations or to continue to another stage of the processing.

[0048] Process 200 begins at frame 202 with source loading (for the initial wash cycle). In this step 202, the source product is drawn into the rotary membrane separator 31, for example through a port on the source product container 14 and through tubing and using a first syringe 32 associated with the syringe pump 44. Thus, the controller 28 transfers the biological fluid / cells from the source container 14 to its rotary membrane separator 31 via a fluid loop or kit 24 by operating one or more syringe pumps 44, 46. In a similar manner, the wash medium can be delivered from its container 38 to the rotary membrane separator 31 via the fluid loop 24.

[0049] At frame 210, during the initial wash, cells from the source product accumulate in the annular space of the rotating membrane separator 31 until (1) the source container is empty (in Figure 3B (Identified by the circled figure 2: the source container is empty) or (2) the number of cells in the rotating membrane separator 31 reaches the pre-configured capacity (in Figure 3BThe diagram in Figure 1, marked with a circle, indicates the maximum packed-cell volume (PCV%). When cells are loaded into the separation chamber or rotating membrane 31, the supernatant (filtrate) passes through the membrane and is aspirated into the first syringe 32. It will be understood that the biological cells may be collected at the harvest point in box 212 or harvested in the process container 16, while the supernatant is separated and removed to the waste container 36. Relevant protocol parameters for this step may include the rotating membrane separator loading speed (RPM), source inlet flow rate (mL / min), and maximum packed-cell volume (PCV) (%).

[0050] Processing 200 continues with further source loading 202 and washing 210 (for initial washing), wherein, with cells suspended in the annular space of a rotating membrane separator 31, solution 1 is aspirated into the rotator of separator 31 using a first syringe 32. The remaining supernatant and washing buffer pass through the rotating membrane 31 and are aspirated into the first syringe 32. Relevant protocol parameters for this step may include the initial rotator washing configuration, the initial washing solution, the initial rotator washing volume (mL), and the initial rotator washing flow rate (mL / min). It will also be understood that this step can be repeated with additional washing solutions. For example, when processing fresh apheresis material for selection, platelets are activated by washing solution 2. In this case, the suspension is first washed with solution 1 to transfer the platelets to the filtrate, and then immediately washed with solution 2 to resuspend the cells in the culture medium required for subsequent processing steps.

[0051] Following step 210, at frame 212 is harvest (for the initial wash), where additional cells within the annular space of the rotating membrane separator 31 are aspirated from the rotator using solution 1 using a second syringe 34 and transferred to a process container (e.g., container 16 in the illustrated example) or to an alternative treatment container. If the source container 14 is not empty, the system 10 returns to the source loading (for the initial wash) state (e.g., ...). Figure 3B (The source container is not empty, as indicated by the circular symbol 3).

[0052] It will be understood that the steps of source loading 202, washing 210, and harvesting 212 can be repeated until, for example, source container 14 is empty. Relevant protocol parameters for step harvesting 212 (for the initial washing) may include harvest volume (mL) and harvest flow rate (mL / min).

[0053] The subsequent step at box 204 is source rinsing (for the initial wash). System 10 then returns to the source loading state at box 208 to process (wash and harvest) the rinse solution with remaining cells. However, system 10 can be configured to include a source rinsing pause at box 206 after transferring solution 1 to source container 14, allowing the user to mix or agitate the rinse solution in source container 14 to enhance the ability to capture any remaining cells trapped in source container 14. The text displayed on screen 58 can be configurable to allow this step to be added. The relevant protocol parameters for this step may include source rinsing pause configuration and source rinsing pause text. If a source rinsing pause is used, system 10 then proceeds to the source loading step at box 208, which processes (washes and harvests) the rinse solution with remaining cells.

[0054] If source container 14 eventually becomes empty, system 10 will transition to step dilution 1 at box 214. In step dilution 1, system 10 dilutes the cell suspension in treatment container 16 with solution 1 immediately after harvesting cells from the rotating membrane separator 31. Relevant protocol parameters for step dilution 1 may include dilution 1 volume (mL) and dilution 1 flow rate (mL / min).

[0055] Following step 1 dilution at box 214, the process can proceed to the next step at box 216, namely, pausing 1 dilution. During pausing 1 dilution, system 10 can be configured to pause when the 1 dilution state is complete. The relevant protocol parameters for this pausing 1 dilution may include pausing 1 dilution configuration, pausing 1 dilution text, 1 dilution sample volume (mL), and antibody volume (mL).

[0056] Following the dilution pause at 216, the storage buffer is drained from the select column at box 218. Therefore, in this specific example, the buffer solution initially held in select column 30 can be drained, leaving the matrix resin material. Note that if a buffer solution that does not require pre-wetting is used, this step can be skipped. Subsequently, at box 220, the select column can be washed / rinsed to prepare select column 30 for further processing.

[0057] The next step is dilution 2 at box 222 (transferring Ab to the selected column). This is Figure 3B The procedure shown includes a point where the user can add the antibody directly to the selection column 30. For example, the user can aseptically inject the antibody fragment (FAB) solution into the selection column 30 manually. According to other embodiments, the antibody can be automatically introduced into the selection column 30.

[0058] Next, at box 224, is incubation. In this example, the incubation is antibody incubation, where the matrix material in selection column 30 is incubated together with the added antibody. The duration of incubation can be configured by the user via at least one input device 54. Next, at box 226, is the transfer of washed cells to the selection column, where the cell suspension collected in processing container 16 is transferred to selection column 30.

[0059] The next step, in box 228, is the incubation process. The duration of incubation can be configured by the user, for example, via input device 54. Relevant program parameters may include the incubation time.

[0060] After incubation at box 228, the procedure can proceed to box 240 for the step of loading target fractions from the selection column (e.g.) Figure 3B The circular symbol 6 indicates negative selection only, or the process can proceed to the next step at box 230, which is to transfer the non-target fraction to waste.

[0061] Processing 200 continues at frame 230 with the step of transferring the negative fraction to waste. In this step 230, the unbound cells remaining in suspension are removed from the selection column 30 via a second syringe 34. The removed cells are then transferred to the waste container 36. The number of negative selections can be configured by the user using input device 54. Additional negative selections may not be configured (e.g., ...). Figure 3B (Identified by circular symbol 5: No additional selection). Alternatively, if an additional negative selection is configured, the system proceeds to box 232 for dilution 3, and again dilutes and resuspends the remaining cells in selection column 30 with solution 1, re-incubates, and repeats the transfer of the negative fraction to waste. Relevant protocol parameters at box 230 may include the number of negative selections. As indicated, dilution 3 is provided at box 232. Relevant protocol parameters may include dilution 3 flow rate (mL / min), dilution 3 solution, and dilution 3 volume (mL).

[0062] At box 234, dilution 4 (transferring the release agent to the selection column) transfers the release agent (e.g., biotin) to selection column 30. Incubation follows at box 236. This incubation may be provided, for example, with a release buffer, in which the cell suspension in selection column 30 is incubated together with solution 3. The duration of incubation can be configured by the user via at least one input device 54. Relevant protocol parameters may include incubation time. Using such an incubation state is desirable based on the release time associated with the release agent (e.g., the release time associated with the affinity of biotin in selection column 30).

[0063] System 10 then proceeds to box 238 for vortex refill. In this step, system 10 refills a section of the fluid circuit or disposable kit 24, for example, with solution 2 and uses a second syringe 34 to purge air from the fluid circuit 24. Relevant protocol parameters for connecting solution 2 may include the volume of solution 2 connected, while relevant protocol parameters for vortex refill may include the selected post-refill volume (mL) and the selected post-source refill flow rate (mL / min).

[0064] System 10 then proceeds to box 240 to load target fractions from the selection column. Using the first syringe 32, the cell suspension is aspirated through the source port into the rotator of the rotating membrane separator 30. Cells accumulate in the annular space of the rotator until (1) the selection column 30 is empty (e.g., ...). Figure 3B (Identified by circular symbol 2: the source container is empty), in which case the next step is to flush the selected column at box 242 (discussed below), or (2) the number of cells in the rotating membrane 31 reaches the pre-configured capacity (e.g. Figure 3B In the diagram marked by circle 1 (reaching maximum annular PCV %), the next step is washing at box 248. When cells are loaded into the rotator of the rotating membrane separator 31, the supernatant (filtrate) passes through the membrane and is aspirated into the first syringe 32. Relevant protocol parameters for loading target fractions from the selection column after such selection may include the post-selection rotator loading speed (RPM), the post-selection source inlet flow rate (mL / min), and the post-selection maximum annular PCV (5%).

[0065] When column 31 is empty (e.g.) Figure 3B When the source container is empty (as indicated by circular symbol 2), the method proceeds to box 242 for a selection column rinse. This is a post-selection column rinse, where, when selection column 31 is empty, solution 2 is transferred at box 238 to selection column 30 to recover any remaining cells. The system returns to the post-selection target fraction loading state to process the rinse solution with remaining cells. Relevant protocol parameters for this source rinse step may include the post-selection source rinse configuration and the post-selection source rinse volume (mL).

[0066] At box 244, system 10 reaches a post-selection column rinse pause. This is a post-selection column rinse pause, where, after transferring solution 2 to selection column 31, system 10 can be configured to pause to allow the user to mix or agitate the rinse solution in selection column 31 to capture any remaining trapped cells. The text displayed on display 58 is configurable, for example, via input device 54. Relevant protocol parameters for this step may include the post-selection column rinse pause configuration and the post-selection column rinse pause text. This continues at box 246 from the target fraction loading at selection column, preparing for further washing at box 248.

[0067] The next step is washing (with fresh culture medium) at box 248, whether due to the cell number in the rotator reaching the pre-configured capacity (e.g.) Figure 3B The circular diagram 1 indicates whether the target fraction is loaded from the selected column at box 240 (reaching the maximum annular PCV%) or from a further rinsing step and box 246. With cells suspended in the annular space of the rotating membrane separator 31, solution 2 is aspirated into the separation chamber 31 using the first syringe 32. The remaining supernatant and wash buffer pass through the membrane and are aspirated into the first syringe 32. Relevant protocol parameters for this step may include the post-selection rotating membrane wash configuration, the post-selection wash volume (mL), and the post-selection wash flow rate (mL / min).

[0068] Processing 200 continues harvesting (to the final container) at frame 250. In this harvesting step, cells within the annular space of the rotating membrane separator 31 are aspirated from the rotator with solution 2 using a second syringe 34 and transferred to the final product container 66, which is a user-connected container (bag, culture vessel, or other container) connected to the fluid path 22. If column 30 is not empty (e.g....), Figure 3B If the source container is not empty (as indicated by circular symbol 3), then system 10 will return to the step of loading the target fraction from the selected column at box 240. If the selected column 30 is empty, system 10 will transition to the final dilution (fresh medium) state at box 252. Relevant protocol parameters for this step may include harvest volume (mL) and harvest flow rate (mL / min).

[0069] In the further final dilution (fresh culture medium) step at box 252, the system immediately dilutes the cell suspension in the final product container 66 connected to fluid path 22 with solution 2 after harvesting cells from the rotator of rotating membrane 31 to achieve a target final product volume or target final concentration. Relevant protocol parameters for this step may include final dilution flow rate (mL / min), final dilution input method, final dilution concentration, final dilution volume (mL), and final dilution air-chase volume (mL).

[0070] System 10 then switches to post-processor 300, such as Figure 3C As shown, this includes sealing / removing the final product at box 302. In this step, system 10 prompts the sealing and removal of the final product container 66 attached to fluid path 22. A subsequent procedure summary step may be at box 304, where the system displays a summary of the procedure, for example, on display screen 58. Finally, at box 306, kit removal occurs, and system 10 prompts the user to remove the disposable fluid loop or kit 24 from the fluid handling hardware 26.

[0071] According to certain implementation schemes, such as Figure 1 As shown, the sealing and removal of the final product can be performed in a product container 66. Alternatively, as... Figure 1 As shown, the system may also provide additional tubes and containers to distribute the cell-selected product equally into a plurality of smaller product containers 66A, having a volume suitable for administration or delivery to a patient. Such product containers may also be configured as delivery containers, such as syringes.

[0072] Therefore, improved fluid handling systems for processing biological fluids and performing cell selection are discussed herein. The descriptions provided above, as well as the other aspects provided below, are intended for illustrative purposes and are not intended to limit the scope of this disclosure to any particular method, system, device, or apparatus described herein.

[0073] It will be understood that operating the biofluidic system 10 includes treating or modifying target cells, cell isolation, concentration and / or washing, cell modification (including but not limited to genetic modification) or other suitable forms of cell treatment. As used herein, the phrases “cell modification” or “modified” or “treated” refer to a process in which target cells are isolated from other cells and subsequently mixed with a modification solution, reagent, or used with a carrier as described below.

[0074] In one implementation, the modified cells generated by the blood processing system include chimeric antigen receptor T-cells (CAR-T) cells. These CAR-T cells can be used for several applications and therapeutic treatments of a variety of diseases, ailments, or conditions. For example, CAR-T cells can be used to treat solid tumors such as anal / rectal, epithelial, ovarian, breast, fallopian tube, endometrial, pancreatic, colorectal, lung, and / or gastrointestinal tumors. CAR-T cells can also be used to treat melanoma, multiple myeloma, acute lymphoblastic leukemia, acute myeloid lymphoma, non-Hodgkin lymphoma, diffuse large B-cell lymphoma, small cell lung cancer, neuroblastoma, glioblastoma, prostate cancer, hemophilia, sickle cell disease, lupus erythematosus, lupus nephritis, myasthenia gravis, autoimmune diseases, soft tissue sarcoma, osteosarcoma, hepatocellular carcinoma, graft-versus-host disease, and rheumatoid arthritis. CAR-T cells can also be used in kidney transplant patients. Therefore, this disclosure provides modified CAR-T cells generated by a blood processing system for the treatment of diseases or conditions selected from: solid tumors, such as anal / rectal, epithelial, ovarian, breast, fallopian tube, endometrial, pancreatic, colorectal, lung, and / or gastrointestinal tumors; melanoma, multiple myeloma, acute lymphoblastic leukemia, acute myeloid lymphoma, non-Hodgkin lymphoma, diffuse large B-cell lymphoma, small cell lung cancer, neuroblastoma, glioblastoma, prostate cancer, hemophilia, sickle cell disease, lupus erythematosus, lupus nephritis, myasthenia gravis, autoimmune diseases, soft tissue sarcoma, osteosarcoma, hepatocellular carcinoma, graft-versus-host disease, rheumatoid arthritis, and kidney transplant-related injuries.

[0075] Different modification strategies can be used in the blood processing system to generate CAR-T cells, and a variety of methods can be implemented in the preparation of CAR-T cells for any of the aforementioned applications or therapeutic treatments. Vectors can be used in the blood processing system (particularly the carrier delivery module) to generate CAR-T cells, such as mRNA, shRNA, siRNA, saRNA, SeekRNA, polymers, proteins and peptides, antibodies, viruses (including but not limited to lentivirus (LV), adeno-associated virus (AAV)), marker molecules, small molecules, virus-like particles (VLPs), transposons / transposases (sleeping beauty, TcBuster, PiggyBac), transcription activator-like effector nucleases (TALENs), zinc finger nucleases (ZFNs), base editors, leader editors, programmable addition via site-specific targeting element (PASTE), CRISPR / Cas, and CRISPR RNPs. Therefore, in some embodiments, this disclosure provides modified CAR-T cells generated using the above-described vector for the treatment of diseases, ailments, or conditions as shown in Tables 1 and 2:

[0076] Table 1

[0077]

[0078] Table 2

[0079]

[0080] The embodiments 1 to 59 listed in Tables 1 and 2 are applicable not only to modified CAR-T cells, but also to any other modified cell type that can be produced by a biofluidic processing system.

[0081] Therefore, the “modified cells” used herein are cells produced by a biofluidic processing system using the carriers (Table 2) of embodiments 35 to 59. The modified cells are also provided for the treatment of diseases, ailments, or conditions according to embodiments 1 to 34 (Table 1).

[0082] In one implementation, the modified cells are modified chimeric antigen receptor natural killer (CAR-NK) cells.

[0083] In another embodiment, the modified cells are modified chimeric antigen receptor monocytes (CAR-M) cells.

[0084] In another embodiment, the modified cell is a modified, engineered T-cell receptor (TCR) cell.

[0085] In another implementation, the modified cells are modified, engineered B cells.

[0086] In another embodiment, the modified cells are modified tumor-infiltrating lymphocytes (TILs).

[0087] In another implementation, the modified cells are modified induced pluripotent stem (iPSC) cells.

[0088] In another embodiment, the modified cells are modified invariant natural killer T (iNKT) cells.

[0089] In another embodiment, the modified cells are modified mesenchymal stem (MSC) cells.

[0090] In another embodiment, the modified cells are modified dendritic cells.

[0091] In another implementation, the modified cells are modified hematopoietic stem cells / stem (CD34+) cells.

[0092] Modified cells can be generated and administered by a blood processing system in a variety of environments, wherein the modified cells are generated using any of the aforementioned carriers and administered in any of the aforementioned applications or therapeutic treatments. For example, the blood processing system can generate and / or administer modified cells within a patient treatment room / facility (inpatient or outpatient). The blood processing system can generate and / or administer modified cells within a cell processing laboratory. The cell processing laboratory can be located in a hospital facility, a non-hospital facility, or a commercial manufacturing facility (centralized or decentralized). The blood processing system can generate and / or administer modified cells within a sterile preparation suite. The sterile preparation suite can be located in a hospital facility, a non-hospital facility, an academic facility, or a commercial manufacturing facility (centralized or decentralized).

[0093] While the modified cells generated by this system are described as being used to treat specific conditions and patient groups, it should be understood that they can be applied to other conditions and / or patient groups, including subgroups of said patient groups (i.e., patients having the same characteristics characterizing a specific patient group but with additional characteristics not shared by all patients in that patient group), larger patient groups encompassing said patient groups (i.e., patient groups having characteristics that include those broadly defined to characterize a specific patient group), and entirely different patient groups (i.e., patients having characteristics that exclude them from a specific patient group). Without departing from the scope of this disclosure, the modified cells can also be applied at various doses, administration regimens, and routes of administration.

[0094] It should be understood that the implementation schemes disclosed herein can be combined with each other in any conceivable combination.

[0095] Other aspects

[0096] Aspect 1. A biofluid processing system for cell affinity selection, the biofluid processing system comprising a fluid processor having reusable hardware for operating a disposable fluid loop connected to a source container and having a biofluid; a separation chamber configured to separate the biofluid from the source container into at least two volumes of material; a processing container for collecting washed and harvested cells from the separation chamber; and a selection column connected to the fluid loop for further cell affinity selection processing of the cells from the processing container.

[0097] Aspect 2. The system of Aspect 1, wherein the fluid processor further comprises a controller configured to operate the reusable hardware when the disposable fluid loop is mounted on the reusable hardware and connected to the source container, the processing container and the selection post.

[0098] Aspect 3. The system described in Aspect 2, wherein the controller further comprises at least a microprocessor and a memory.

[0099] Aspect 4. The system of any one of Aspects 1 to 3, wherein the fluid processor further comprises an input device and an output device connected to the controller.

[0100] Aspect 5. The system of any one of Aspects 1 to 4, wherein the reusable hardware of the fluid processor further comprises a driver for a rotating membrane separator in the separation chamber.

[0101] Aspect 6. The system of any one of Aspects 1 to 5, wherein the reusable hardware of the fluid processor further includes a pump and cartridge interface that engages with the fluid loop.

[0102] Aspect 7. The system of any one of Aspects 1 to 6, wherein the fluid circuit further comprises a single final product container.

[0103] Aspect 8. The system of any one of Aspects 1 to 7, wherein the fluid circuit further comprises a plurality of final product containers.

[0104] Aspect 9. The system of any one of Aspects 1 to 8, wherein the selection column initially comprises a matrix or resin material.

[0105] Aspect 10. The system of any one of Aspects 1 to 9, wherein at least three additional containers are connected to the fluid loop, the at least three additional containers comprising a first solution container containing a separation / processing buffer, a second solution container containing a resuspension buffer, and a third solution container containing a release buffer.

[0106] Aspect 11. The system of any one of Aspects 1 to 10, wherein the controller is coupled to a separator driver, at least two injection pumps and a control box interface for a rotary membrane separator for the separation chamber, the controller being configured to selectively operate the separator driver, the injection pumps and the box interface to provide a procedure for using the selection column according to a scheme.

[0107] Aspect 12. A method of operating a biological fluid processing system for cell affinity selection, the method comprising:

[0108] A biofluid processing system is obtained, the biofluid processing system comprising a fluid processor that operates a disposable fluid loop connected to a source container having biofluid and configured to separate the biofluid from the source container into at least two volumes of material; a processing container connected to the fluid processor along a fluid path; and a selection column connected to the fluid loop.

[0109] The process includes a pre-processing state, which further includes: selecting a program scheme, setting the program, installing the disposable fluid circuit on the fluid processor; attaching multiple containers to the fluid processor; running a fluid circuit solution pre-charge containing the separation chamber; attaching the source container to the fluid processor; and running a fluid circuit source pre-charge.

[0110] The process further includes: treating a biofluid from the source container in a separation chamber via the disposable fluid loop, including one or more cycles of washing, harvesting, and diluting the biofluid; rinsing the source container; diluting the harvested cells; collecting the harvested cells in the processing container; draining the solution from the selection column; adding an antibody to the selection column; transferring the harvested cells from the processing container to the selection column; transferring a non-target fraction containing unbound cells suspended in the selection column to waste; diluting the material in the selection column; refilling the separation chamber for processing the material in the selection column; loading the target fraction from the selection column into the separation chamber; washing and harvesting the cells from the selection column; harvesting the selected cells as the final product in at least one final container; and

[0111] The process includes a post-processing phase, which further includes sealing the at least one final container and removing the final product; and removing the disposable fluid loop.

[0112] Aspect 13. The method of aspect 12, wherein the treatment uses only negative selection relative to the target cells.

[0113] Aspect 14. The method of aspect 12 or 13, wherein the processor is configured to further perform positive selection incubation and positive grade separation relative to the target cells.

[0114] Aspect 15. The method of any one of Aspects 12 to 14, wherein the final product is equally divided into a plurality of smaller containers before the fluid loop is removed.

[0115] Aspect 16. The method of any one of Aspects 12 to 15, wherein the processor transfers the negative fraction from the selection column to waste, and then dilutes the material in the selection column with a release buffer.

[0116] Aspect 17. The method of any one of Aspects 12 to 16, wherein the release buffer is biotin.

[0117] Aspect 18. The method of any one of Aspects 12 to 17, wherein the processor further comprises a controller configured to selectively operate the processor to provide, according to a scheme, a procedure for using the separation chamber, the processing container, and the selection column.

[0118] Aspect 19. A method of operating a biofluid processing system for cell affinity selection, wherein the system comprises a fluid processor that operates a disposable fluid loop connected to and configured to separate the biofluid from the source container into at least two volumes of material; a processing container connected to the fluid processor along a fluid path; and a selection column connected to the fluid loop, the method comprising:

[0119] Preprocessing state, processing state, and postprocessing state;

[0120] The pretreatment state further includes: selecting a program scheme; setting the program; installing the disposable fluid loop on the fluid processor; attaching at least a first solution container containing a separation / processing buffer, a second solution container containing a resuspension buffer, and a third solution container containing a release buffer to the fluid processor; running fluid loop solution pre-charge; attaching the source container to the fluid processor; and running fluid loop source pre-charge;

[0121] The processing state further includes: processing the biofluid from the source container in the separation chamber via the disposable fluid loop, including one or more cycles of washing, harvesting, and diluting the biofluid; rinsing the source container; diluting the harvested cells; collecting the harvested cells in the processing container; removing a buffer solution from the selection column; diluting the material in the selection column by adding an antibody to the selection column; incubating with the antibody in the selection column; transferring the harvested cells from the processing container to the selection column; transferring the non-target fraction containing unbound cells suspended in the selection column to waste; diluting the material in the selection column by transferring biotin to the selection column; incubating; refilling the separation chamber for processing the material in the selection column; loading the target fraction from the selection column into the separation chamber; washing and harvesting the cells processed in the separation chamber; rinsing the source container; harvesting the selected cells as the final product in at least one final container; and diluting the harvested cells in the final container; and

[0122] The post-processing state further includes: sealing the at least one final container and removing the final product, and removing the disposable fluid loop.

[0123] Aspect 20. The method of aspect 19, wherein the fluid processor further comprises a controller configured to selectively operate the fluid processor to provide, according to a scheme, a procedure for using the separation chamber, the processing container, and the selection column.

[0124] Aspect 21. Modified cells for treating a patient’s disease, ailment, or medical condition, characterized in that the modified cells are produced using a biofluid processing system comprising a fluid processor having reusable hardware for operating a disposable fluid loop connected to a source container and having a biofluid; a separation chamber configured to separate the biofluid from the source container into at least two volumes of material; a processing container for collecting washed and harvested cells from the separation chamber; and a selection column connected to the fluid loop for further cell affinity selection of the cells from the processing container.

[0125] Aspect 22. The modified cells of aspect 21, wherein the target cells are selected from chimeric antigen receptor T cells, chimeric antigen receptor natural killer cells, chimeric antigen receptor monocytes, modified T cell receptors, modified B cells, tumor-infiltrating lymphocytes, induced pluripotent stem cells, mesenchymal stem cells, dendritic cells and hematopoietic stem cells.

[0126] Aspect 23. The modified cell of any one of Aspects 21 to 22, wherein the target cell is modified using a vector selected from: lentivirus, adeno-associated virus, virus-like particles, transposons / transposases, transcription activator-like effector nucleases, zinc finger nucleases, mRNA, shRNA, siRNA, SeekRNA, CRISPR / Cas, base editors, leader editors, and programmable additions implemented with site-specific targeting elements.

[0127] Aspect 24. The modified cells of any one of Aspects 21 to 23, wherein the modified cells are produced in an environment selected from: a patient treatment room in an inpatient facility, a patient treatment room in an outpatient facility, a cell processing laboratory in a hospital facility, a cell processing laboratory in a non-hospital facility, a cell processing laboratory in a centralized commercial production facility, a cell processing laboratory in a decentralized commercial production facility, a sterile preparation suite in a hospital facility, a sterile preparation suite in a non-hospital facility, a sterile preparation suite in an academic facility, a sterile preparation suite in a centralized commercial production facility, and a sterile preparation suite in a decentralized commercial production facility.

[0128] Aspect 25. The modified cells of any one of Aspects 21 to 24, wherein the disease, ailment or medical condition is selected from solid tumors, melanoma, multiple myeloma, acute lymphoblastic leukemia, acute myeloid leukemia, non-Hodgkin's lymphoma, diffuse large B-cell lymphoma, small cell lung cancer, kidney transplantation, neuroblastoma, glioblastoma, prostate cancer, hemophilia, sickle cell disease, lupus erythematosus, lupus nephritis, myasthenia gravis, autoimmune diseases, soft tissue sarcoma, osteosarcoma, hepatocellular carcinoma, graft-versus-host disease and rheumatoid arthritis.

[0129] Aspect 26. Modified cells for treating a patient's disease, ailment, or medical condition, characterized in that the modified cells are produced using a method comprising: obtaining a biofluid processing system comprising a fluid processor operating a disposable fluid loop connected to a source container having biofluid and configured to separate biofluid from the source container into at least two volumes of material; a processing container connected to the fluid processor along a fluid path; and a selection column connected to the fluid loop; entering a pretreatment state, the pretreatment state further comprising: selecting a program scheme, setting a program, mounting the disposable fluid loop to the fluid processor; attaching a plurality of containers to the fluid processor; running a fluid loop solution pre-fill containing the separation chamber; attaching the source container to the fluid processor; and running a fluid loop source pre-fill; entering a processing state, the processing state further comprising: passing through the separation chamber via the A disposable fluid loop processes a biofluid from the source container, including one or more cycles of washing, harvesting, and diluting the biofluid; rinsing the source container; diluting the harvested cells; collecting the harvested cells in the processing container; draining the solution from the selection column; adding an antibody to the selection column; transferring the harvested cells from the processing container to the selection column; transferring a non-target fraction containing unbound cells suspended in the selection column to waste; diluting the material in the selection column; refilling the separation chamber for processing the material in the selection column; loading the target fraction from the selection column into the separation chamber; washing and harvesting the cells from the selection column; harvesting the selected cells as the final product in at least one final container; and performing a post-processing state, the post-processing state further including: sealing the at least one final container and removing the final product; and removing the disposable fluid loop.

[0130] Aspect 27. The modified cells described in Aspect 26, wherein the target cells are selected from chimeric antigen receptor T cells, chimeric antigen receptor natural killer cells, chimeric antigen receptor monocytes, modified T cell receptors, modified B cells, tumor-infiltrating lymphocytes, induced pluripotent stem cells, mesenchymal stem cells, dendritic cells, and hematopoietic stem cells.

[0131] Aspect 28. The modified cell of any one of Aspects 26 to 27, wherein the target cell is modified using a vector selected from: lentivirus, adeno-associated virus, virus-like particles, transposons / transposases, transcription activator-like effector nucleases, zinc finger nucleases, mRNA, shRNA, siRNA, SeekRNA, CRISPR / Cas, base editors, leader editors, and programmable additions implemented with site-specific targeting elements.

[0132] Aspect 29. The modified cells of any one of Aspects 26 to 28, wherein the modified cells are produced in an environment selected from: a patient treatment room in an inpatient facility, a patient treatment room in an outpatient facility, a cell processing laboratory in a hospital facility, a cell processing laboratory in a non-hospital facility, a cell processing laboratory in a centralized commercial production facility, a cell processing laboratory in a decentralized commercial production facility, a sterile preparation suite in a hospital facility, a sterile preparation suite in a non-hospital facility, a sterile preparation suite in an academic facility, a sterile preparation suite in a centralized commercial production facility, and a sterile preparation suite in a decentralized commercial production facility.

[0133] Aspect 30. The modified cells of any one of Aspects 26 to 29, wherein the disease, ailment, or medical condition is selected from solid tumors, melanoma, multiple myeloma, acute lymphoblastic leukemia, acute myeloid leukemia, non-Hodgkin's lymphoma, diffuse large B-cell lymphoma, small cell lung cancer, kidney transplantation, neuroblastoma, glioblastoma, prostate cancer, hemophilia, sickle cell disease, lupus erythematosus, lupus nephritis, myasthenia gravis, autoimmune diseases, soft tissue sarcoma, osteosarcoma, hepatocellular carcinoma, graft-versus-host disease, and rheumatoid arthritis.

[0134] Aspect 31. Modified cells for treating a patient's disease, ailment, or medical condition, characterized in that the modified cells are produced using a method of operating a biofluidic processing system for cell affinity selection, wherein the system comprises a fluid processor that operates a disposable fluid loop connected to a source container filled with biofluid and configured to separate biofluid from the source container into at least two volumes of material; a processing container connected to the fluid processor along a fluid path; and a selection column connected to the fluid loop, the method comprising: a pretreatment state, a processing state, and a posttreatment state; the pretreatment state further comprising: selecting a program scheme; setting a program; mounting the disposable fluid loop on the fluid processor; attaching at least a first solution container containing a separation / processing buffer, a second solution container containing a resuspension buffer, and a third solution container containing a release buffer to the fluid processor; performing fluid loop solution pre-filling; attaching the source container to the fluid processor; and performing fluid loop source pre-filling; the processing state further comprising: processing the biofluid from the source container in a separation chamber via the disposable fluid loop. The process includes one or more cycles of washing, harvesting, and diluting the biofluid; rinsing the source container; diluting the harvested cells; collecting the harvested cells in the processing container; removing a buffer solution from the selection column; diluting the material in the selection column by adding an antibody to the selection column; incubating the material in the selection column with the antibody; transferring the harvested cells from the processing container to the selection column; transferring the non-target fraction containing unbound cells suspended in the selection column to waste; diluting the material in the selection column by transferring biotin to the selection column; incubation; refilling the separation chamber for processing the material in the selection column; loading the target fraction from the selection column into the separation chamber; washing and harvesting the cells processed in the separation chamber; rinsing the source container; harvesting the selected cells as the final product in at least one final container; and diluting the harvested cells in the final container; and the post-processing state further includes sealing the at least one final container and removing the final product, and removing the disposable fluid loop.

[0135] Aspect 32. The modified cells of aspect 31, wherein the target cells are selected from chimeric antigen receptor T cells, chimeric antigen receptor natural killer cells, chimeric antigen receptor monocytes, modified T cell receptors, modified B cells, tumor-infiltrating lymphocytes, induced pluripotent stem cells, mesenchymal stem cells, dendritic cells and hematopoietic stem cells.

[0136] Aspect 33. The modified cell of any one of Aspects 31 to 32, wherein the target cell is modified using a vector selected from: lentivirus, adeno-associated virus, virus-like particles, transposons / transposases, transcription activator-like effector nucleases, zinc finger nucleases, mRNA, shRNA, siRNA, SeekRNA, CRISPR / Cas, base editors, leader editors, and programmable additions implemented with site-specific targeting elements.

[0137] Aspect 34. The modified cells of any one of Aspects 31 to 33, wherein the modified cells are produced in an environment selected from: a patient treatment room in an inpatient facility, a patient treatment room in an outpatient facility, a cell processing laboratory in a hospital facility, a cell processing laboratory in a non-hospital facility, a cell processing laboratory in a centralized commercial production facility, a cell processing laboratory in a decentralized commercial production facility, a sterile preparation suite in a hospital facility, a sterile preparation suite in a non-hospital facility, a sterile preparation suite in an academic facility, a sterile preparation suite in a centralized commercial production facility, and a sterile preparation suite in a decentralized commercial production facility.

[0138] Aspect 35. The modified cell of any one of Aspects 31 to 34, wherein the disease, ailment or medical condition is selected from solid tumors, melanoma, multiple myeloma, acute lymphoblastic leukemia, acute myeloid leukemia, non-Hodgkin lymphoma, diffuse large B-cell lymphoma, small cell lung cancer, kidney transplantation, neuroblastoma, glioblastoma, prostate cancer, hemophilia, sickle cell disease, lupus erythematosus, lupus nephritis, myasthenia gravis, autoimmune diseases, soft tissue sarcoma, osteosarcoma, hepatocellular carcinoma, graft-versus-host disease and rheumatoid arthritis.

Claims

1. A biofluid processing system for cell affinity selection, the biofluid processing system comprising a fluid processor having reusable hardware for operating a disposable fluid loop connected to a source container and having a biofluid; a separation chamber configured to separate the biofluid from the source container into at least two volumes of material; a processing container for collecting washed and harvested cells from the separation chamber; and a selection column connected to the fluid loop for further cell affinity selection treatment of the cells from the processing container.

2. The system of claim 1, wherein the fluid processor further comprises a controller configured to operate the reusable hardware when the disposable fluid loop is mounted on the reusable hardware and connected to the source container, the processing container and the selection post.

3. The system of claim 2, wherein the controller further comprises at least a microprocessor and a memory.

4. The system of any one of claims 1 to 3, wherein the fluid processor further comprises an input device and an output device connected to the controller.

5. The system of any one of claims 1 to 4, wherein the reusable hardware of the fluid processor further comprises a driver for the rotary membrane separator in the separation chamber.

6. The system of any one of claims 1 to 5, wherein the reusable hardware of the fluid processor further comprises a pump and cartridge interface that engages with the fluid loop.

7. The system of any one of claims 1 to 6, wherein the fluid loop further comprises a single final product container.

8. The system of any one of claims 1 to 7, wherein the fluid circuit further comprises a plurality of final product containers.

9. The system of any one of claims 1 to 8, wherein the selection column initially comprises a matrix or resin material.

10. The system of any one of claims 1 to 9, wherein at least three additional containers are connected to the fluid loop, the at least three additional containers comprising a first solution container containing a separation / processing buffer, a second solution container containing a resuspension buffer, and a third solution container containing a release buffer.

11. The system of any one of claims 1 to 10, wherein the controller is coupled to a separator driver, at least two injection pumps and a control box interface for a rotary membrane separator in the separation chamber, the controller being configured to selectively operate the separator driver, the injection pumps and the box interface to provide a procedure for using the selection column according to a scheme.

12. A method of operating a biological fluid processing system for cell affinity selection, the method comprising: A biofluid processing system is obtained, the biofluid processing system comprising a fluid processor that operates a disposable fluid loop, the disposable fluid loop being connectable to a source container having biofluid and configured to separate the biofluid from the source container into at least two volumes of material; and a processing container connected to the fluid processor along a fluid path; And a selection column, which is connected to the fluid circuit; The process includes a pre-processing state, which further includes: selecting a program scheme, setting the program, installing the disposable fluid circuit on the fluid processor; attaching multiple containers to the fluid processor; running a fluid circuit solution pre-charge containing the separation chamber; attaching the source container to the fluid processor; and running a fluid circuit source pre-charge. The process further includes: treating a biofluid from the source container in a separation chamber via the disposable fluid loop, including one or more cycles of washing, harvesting, and diluting the biofluid; rinsing the source container; diluting the harvested cells; collecting the harvested cells in the processing container; draining the solution from the selection column; adding an antibody to the selection column; transferring the harvested cells from the processing container to the selection column; transferring a non-target fraction containing unbound cells suspended in the selection column to waste; diluting the material in the selection column; refilling the separation chamber for processing the material in the selection column; loading the target fraction from the selection column into the separation chamber; washing and harvesting the cells from the selection column; harvesting the selected cells as the final product in at least one final container; and The process includes a post-processing phase, which further includes sealing the at least one final container and removing the final product; and removing the disposable fluid loop.

13. The method of claim 12, wherein the treatment uses only negative selection relative to the target cells.

14. The method of claim 12 or 13, wherein the processor is configured to further perform positive selection incubation and positive grade separation relative to the target cells.

15. The method of any one of claims 12 to 14, wherein the final product is equally divided into a plurality of smaller containers before the fluid loop is removed.

16. The method of any one of claims 12 to 15, wherein the processor transfers the negative fraction from the selection column to waste, and then dilutes the material in the selection column with a release buffer.

17. The method of any one of claims 12 to 16, wherein the release buffer is biotin.

18. The method of any one of claims 12 to 17, wherein the processor further comprises a controller configured to selectively operate the processor to provide, according to a scheme, a procedure for using the separation chamber, the processing container, and the selection column.

19. A method of operating a biofluid processing system for cell affinity selection, wherein the system includes a fluid processor that operates a disposable fluid loop connected to a source container filled with biofluid and configured to separate the biofluid from the source container into at least two volumes of material; A processing container connected to the fluid processor along the fluid path; And a selection column connected to the fluid circuit, the method comprising: Preprocessing state, processing state, and postprocessing state; The pretreatment state further includes: selecting a program scheme; setting the program; installing the disposable fluid loop on the fluid processor; attaching at least a first solution container containing a separation / processing buffer, a second solution container containing a resuspension buffer, and a third solution container containing a release buffer to the fluid processor; running fluid loop solution pre-charge; attaching the source container to the fluid processor; and running fluid loop source pre-charge; The processing state further includes: processing the biofluid from the source container in the separation chamber via the disposable fluid loop, including one or more cycles of washing, harvesting, and diluting the biofluid; rinsing the source container; diluting the harvested cells; collecting the harvested cells in the processing container; removing a buffer solution from the selection column; diluting the material in the selection column by adding an antibody to the selection column; incubating with the antibody in the selection column; transferring the harvested cells from the processing container to the selection column; transferring the non-target fraction containing unbound cells suspended in the selection column to waste; diluting the material in the selection column by transferring biotin to the selection column; incubating; refilling the separation chamber for processing the material in the selection column; loading the target fraction from the selection column into the separation chamber; washing and harvesting the cells processed in the separation chamber; rinsing the source container; harvesting the selected cells as the final product in at least one final container; and diluting the harvested cells in the final container; and The post-processing state further includes: sealing the at least one final container and removing the final product, and removing the disposable fluid loop.

20. The method of claim 19, wherein the fluid processor further comprises a controller configured to selectively operate the fluid processor to provide, according to a scheme, a procedure for using the separation chamber, the processing container, and the selection column.

21. Modified cells for treating a patient's disease, ailment, or medical condition, characterized in that... The modified cells are generated using a biofluid processing system comprising a fluid processor having reusable hardware for operating a disposable fluid loop connected to a source container and having biofluid; a separation chamber configured to separate the biofluid from the source container into at least two volumes of material; a processing container for collecting washed and harvested cells from the separation chamber; and a selection column connected to the fluid loop for further cell affinity selection of the cells from the processing container.

22. The modified cell of claim 21, wherein the target cell is selected from chimeric antigen receptor T cells, chimeric antigen receptor natural killer cells, chimeric antigen receptor monocytes, modified T cell receptors, modified B cells, tumor-infiltrating lymphocytes, induced pluripotent stem cells, mesenchymal stem cells, dendritic cells, and hematopoietic stem cells.

23. The modified cell of any one of claims 21 to 22, wherein the target cell is modified using a vector selected from: lentivirus, adeno-associated virus, virus-like particles, transposons / transposases, transcription activator-like effector nucleases, zinc finger nucleases, mRNA, shRNA, siRNA, SeekRNA, CRISPR / Cas, base editors, leader editors, and programmable additions implemented with site-specific targeting elements.

24. The modified cells of any one of claims 21 to 23, wherein the modified cells are produced in an environment selected from: a patient treatment room in an inpatient facility, a patient treatment room in an outpatient facility, a cell processing laboratory in a hospital facility, a cell processing laboratory in a non-hospital facility, a cell processing laboratory in a centralized commercial production facility, a cell processing laboratory in a decentralized commercial production facility, a sterile preparation suite in a hospital facility, a sterile preparation suite in a non-hospital facility, a sterile preparation suite in an academic facility, a sterile preparation suite in a centralized commercial production facility, and a sterile preparation suite in a decentralized commercial production facility.

25. The modified cell of any one of claims 21 to 24, wherein the disease, ailment, or medical condition is selected from solid tumors, melanoma, multiple myeloma, acute lymphoblastic leukemia, acute myeloid leukemia, non-Hodgkin's lymphoma, diffuse large B-cell lymphoma, small cell lung cancer, kidney transplantation, neuroblastoma, glioblastoma, prostate cancer, hemophilia, sickle cell disease, lupus erythematosus, lupus nephritis, myasthenia gravis, autoimmune diseases, soft tissue sarcoma, osteosarcoma, hepatocellular carcinoma, graft-versus-host disease, and rheumatoid arthritis.

26. Modified cells for treating a patient's disease, ailment, or medical condition, characterized in that... The modified cells are produced using a method comprising: A biofluid processing system is obtained, the biofluid processing system comprising a fluid processor that operates a disposable fluid loop connected to a source container having biofluid and configured to separate the biofluid from the source container into at least two volumes of material; a processing container connected to the fluid processor along a fluid path; and a selection column connected to the fluid loop. The process includes a pre-processing state, which further includes: selecting a program scheme, setting the program, installing the disposable fluid circuit on the fluid processor; attaching multiple containers to the fluid processor; running a fluid circuit solution pre-charge containing the separation chamber; attaching the source container to the fluid processor; and running a fluid circuit source pre-charge. The process further includes: treating a biofluid from the source container in a separation chamber via the disposable fluid loop, including one or more cycles of washing, harvesting, and diluting the biofluid; rinsing the source container; diluting the harvested cells; collecting the harvested cells in the processing container; draining the solution from the selection column; adding an antibody to the selection column; transferring the harvested cells from the processing container to the selection column; transferring a non-target fraction containing unbound cells suspended in the selection column to waste; diluting the material in the selection column; refilling the separation chamber for processing the material in the selection column; loading the target fraction from the selection column into the separation chamber; washing and harvesting the cells from the selection column; harvesting the selected cells as the final product in at least one final container; and The process includes a post-processing phase, which further includes sealing the at least one final container and removing the final product; and removing the disposable fluid loop.

27. The modified cell of claim 26, wherein the target cell is selected from chimeric antigen receptor T cells, chimeric antigen receptor natural killer cells, chimeric antigen receptor monocytes, modified T cell receptors, modified B cells, tumor-infiltrating lymphocytes, induced pluripotent stem cells, mesenchymal stem cells, dendritic cells, and hematopoietic stem cells.

28. The modified cell of any one of claims 26 to 27, wherein the target cell is modified using a vector selected from: lentivirus, adeno-associated virus, virus-like particles, transposons / transposases, transcription activator-like effector nucleases, zinc finger nucleases, mRNA, shRNA, siRNA, SeekRNA, CRISPR / Cas, base editors, leader editors, and programmable additions implemented with site-specific targeting elements.

29. The modified cells of any one of claims 26 to 28, wherein the modified cells are produced in an environment selected from: a patient treatment room in an inpatient facility, a patient treatment room in an outpatient facility, a cell processing laboratory in a hospital facility, a cell processing laboratory in a non-hospital facility, a cell processing laboratory in a centralized commercial production facility, a cell processing laboratory in a decentralized commercial production facility, a sterile preparation suite in a hospital facility, a sterile preparation suite in a non-hospital facility, a sterile preparation suite in an academic facility, a sterile preparation suite in a centralized commercial production facility, and a sterile preparation suite in a decentralized commercial production facility.

30. The modified cell of any one of claims 26 to 29, wherein the disease, ailment, or medical condition is selected from solid tumors, melanoma, multiple myeloma, acute lymphoblastic leukemia, acute myeloid leukemia, non-Hodgkin's lymphoma, diffuse large B-cell lymphoma, small cell lung cancer, kidney transplantation, neuroblastoma, glioblastoma, prostate cancer, hemophilia, sickle cell disease, lupus erythematosus, lupus nephritis, myasthenia gravis, autoimmune diseases, soft tissue sarcoma, osteosarcoma, hepatocellular carcinoma, graft-versus-host disease, and rheumatoid arthritis.

31. Modified cells for treating a patient's disease, ailment, or medical condition, characterized in that... The modified cells are generated using a method of operating a biofluid processing system for cell affinity selection, wherein the system includes a fluid processor that operates a disposable fluid loop that can be connected to a source container filled with biofluid and configured to separate the biofluid from the source container into at least two volumes of material. A processing container connected to the fluid processor along the fluid path; And a selection column connected to the fluid circuit, the method comprising: Preprocessing state, processing state, and postprocessing state; The pretreatment state further includes: selecting a program scheme; setting the program; installing the disposable fluid loop on the fluid processor; attaching at least a first solution container containing a separation / processing buffer, a second solution container containing a resuspension buffer, and a third solution container containing a release buffer to the fluid processor; running fluid loop solution pre-charge; attaching the source container to the fluid processor; and running fluid loop source pre-charge; The processing state further includes: processing the biofluid from the source container in the separation chamber via the disposable fluid loop, including one or more cycles of washing, harvesting, and diluting the biofluid; rinsing the source container; diluting the harvested cells; collecting the harvested cells in the processing container; removing a buffer solution from the selection column; diluting the material in the selection column by adding an antibody to the selection column; incubating with the antibody in the selection column; transferring the harvested cells from the processing container to the selection column; transferring the non-target fraction containing unbound cells suspended in the selection column to waste; diluting the material in the selection column by transferring biotin to the selection column; incubating; refilling the separation chamber for processing the material in the selection column; loading the target fraction from the selection column into the separation chamber; washing and harvesting the cells processed in the separation chamber; rinsing the source container; harvesting the selected cells as the final product in at least one final container; and diluting the harvested cells in the final container; and The post-processing state further includes: sealing the at least one final container and removing the final product, and removing the disposable fluid loop.

32. The modified cell of claim 31, wherein the target cell is selected from chimeric antigen receptor T cells, chimeric antigen receptor natural killer cells, chimeric antigen receptor monocytes, modified T cell receptors, modified B cells, tumor-infiltrating lymphocytes, induced pluripotent stem cells, mesenchymal stem cells, dendritic cells, and hematopoietic stem cells.

33. The modified cell of any one of claims 31 to 32, wherein the target cell is modified using a vector selected from: lentivirus, adeno-associated virus, virus-like particles, transposons / transposases, transcription activator-like effector nucleases, zinc finger nucleases, mRNA, shRNA, siRNA, SeekRNA, CRISPR / Cas, base editors, leader editors, and programmable additions implemented with site-specific targeting elements.

34. The modified cells of any one of claims 31 to 33, wherein the modified cells are produced in an environment selected from: a patient treatment room in an inpatient facility, a patient treatment room in an outpatient facility, a cell processing laboratory in a hospital facility, a cell processing laboratory in a non-hospital facility, a cell processing laboratory in a centralized commercial production facility, a cell processing laboratory in a decentralized commercial production facility, a sterile preparation suite in a hospital facility, a sterile preparation suite in a non-hospital facility, a sterile preparation suite in an academic facility, a sterile preparation suite in a centralized commercial production facility, and a sterile preparation suite in a decentralized commercial production facility.

35. The modified cell of any one of claims 31 to 34, wherein the disease, ailment, or medical condition is selected from solid tumors, melanoma, multiple myeloma, acute lymphoblastic leukemia, acute myeloid leukemia, non-Hodgkin's lymphoma, diffuse large B-cell lymphoma, small cell lung cancer, kidney transplantation, neuroblastoma, glioblastoma, prostate cancer, hemophilia, sickle cell disease, lupus erythematosus, lupus nephritis, myasthenia gravis, autoimmune diseases, soft tissue sarcoma, osteosarcoma, hepatocellular carcinoma, graft-versus-host disease, and rheumatoid arthritis.

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