High concentration cell packaging and shipping

Abstract

This invention relates to methods and products for packaging and shipping therapeutic cells for cell therapy.

Description

[Technical Field] [Cross-reference to related applications] This application claims priority to U.S. Provisional Application No. 62 / 848,230, filed May 15, 2019, the entire contents of which are incorporated herein by reference. This invention relates to methods and products for packaging and shipping therapeutic cells for cell therapy. [Previous Technology] Cell therapy involves administering cells, such as stem cells or pluripotent cells, to patients, and has been used to treat various conditions. Stem cell or pluripotent cell lines are cell types capable of self-renewal and differentiation into various specialized cell types through mitotic cell division. Accordingly, these cell lines have the potential to treat a wide range of diseases and injuries, including neurological trauma, malignancies, genetic diseases, heme disorders, and immunodeficiency. However, studies have shown that if stem cells (e.g., placental blood cells or umbilical cord blood cells) are not processed rapidly after freezing, functional progenitor cells will be lost (Ivanovic et al., Transfusion, 2011 Sep; 51(9): 2044-5). Therefore, logistical issues often hinder the application of these cells. For example, stem cells (such as umbilical cord blood cells) are routinely cryopreserved in storage facilities (e.g., cell banks) after collection and transported from these facilities to hospitals when needed. This cryopreservation process involves cooling to sub-zero temperatures (typically 77 K or -196 °C, the boiling point of liquid nitrogen) to preserve cells or tissues, and therefore carries certain risks. For example, the preserved cells may be damaged during freezing or thawing to room temperature. These risks are particularly severe for stem cells or pluripotent cells, as one of the most important aspects of transplantation of these cells is the number of viable stem cells / pluripotent cells and their developmental potential at the time of transplantation. For this reason, stem cell / pluripotent cell lines are transported using conventional cryogenic preservation methods for the shortest possible time. In fact, overnight transport on dry ice or in liquid nitrogen transporters is industry standard and requires extremely careful temperature monitoring. However, this practice still cannot eliminate the risks. Furthermore, the standard protocol for cryopreservation involves mixing cells and cryoprotectants to achieve a final concentration of 10⁶ to 10⁷ cells per milliliter. This is because cells frozen at lower or higher concentrations tend to have lower viability (Kielberg et al., Cryopreservation of Mammalian Cells-Protocols, Tech Note No. 14, 2010 Thermo Fisher Scientific Inc., https: / / assets.thermofisher.com / TFS-Assets / LSG / Application-Notes / D19575.pdf). However, therapeutic applications of these cells often require much larger quantities and concentrations exceeding 10⁷ cells / ml. Therefore, clinicians must further process these cells to increase the concentration, creating additional logistical and compliance issues. Therefore, long-distance (e.g., intercontinental) transportation of stem cells is extremely expensive and impractical. Consequently, processes or methods for transporting stem cells at high concentrations at room temperature / ambient temperature are needed. [Summary of the Invention] This invention addresses the aforementioned needs in multiple ways. In one embodiment, the present invention provides a therapeutic composition comprising (i) about 1 x 10⁷ to 1 x 10⁹ / ml (e.g., 2 x 10⁷ to 1 x 10⁹ / ml, 5 x 10⁷ to 1 x 10⁹ / ml, 1 x 10⁸ / ml, 2 x 10⁸ / ml, 3 x 10⁸ / ml, 4 x 10⁸ / ml, 5 x 10⁸ / ml) of therapeutic cells and (ii) a medically acceptable carrier solution. The medically acceptable carrier solution contains (a) about 25 to 30 mM (e.g., 26 to 28 mM and 27 mM) of acetate and about 20 to 25 mM (e.g., 21 to 24 mM and 23 mM) of gluconate and (b) has an osmotic pressure of about 270 to 320 mOsmol / L (e.g., 280 to 310, 280 to 300, 290 to 300 and about 294 to 295 mOsmol / L). The medically acceptable carrier solution may have a sodium concentration of 126 to 154 mEq / L. In some specific embodiments, the medically acceptable carrier solution may contain one or more of the following: about 120 to 160 mM (e.g., 130 to 150 and 140 mM) of Na+, about 3 to 7 mM (e.g., 4 to 6 and 5 mM) of K+, about 1.0 to 2.0 mM (e.g., 1.2 to 1.8 and 1.5 mM) of Mg2+, and about 90 to 110 mM (e.g., 95 to 100 and 98 mM) of Cl-. In some specific examples, the medically acceptable carrier solution is free of Ca2+ or lactate or both. In one example, the medically acceptable carrier solution contains approximately 140 mM Na+, approximately 5 mM K+, approximately 1.5 mM Mg2+, approximately 98 mM Cl-, approximately 27 mM acetate, and approximately 23 mM gluconate. In this case, the medically acceptable carrier solution may contain approximately 90 mM sodium chloride (NaCl), approximately 5 mM potassium chloride (KCl), approximately 1.5 mM magnesium chloride (MgCl2‧6H2O), approximately 27 mM sodium acetate trihydrate (C2H3NaO2‧3H2O), and approximately 23 mM sodium gluconate (C6H11NaO7). The therapeutic composition or the medically acceptable carrier solution may have a pH of about 4.0 to 8.0 (e.g., about 5.5 to about 8.0, about 6.0 to about 7.5, about 6.0 and about 7.4). The therapeutic composition is free of DMSO or contains trace amounts of DMSO (i.e., 0.5% or less). The therapeutic composition may contain about 0.5% to about 5% (e.g., about 1% to about 5%, about 1% to 3%, about 1% to 2.5% or about 1%) of serum or serum albumin. Examples include human serum or human serum albumin (HSA). The therapeutic composition may be available at temperatures in the range of about 1 to 10°C, about 2 to 8°C, or about 3 to 5°C. Preferably, the composition is available at a temperature of about 4°C. Examples of therapeutic cells include monocytes, umbilical cord blood cells, hematopoietic stem cells, mesenchymal stem cells, embryonic stem cells, peripheral blood cells, bone marrow cells, or placental blood cells. In some specific examples, the therapeutic composition or cell line contains CD13+, CD34+, or CD134+ cells. In one specific example, the cells may be frozen and thawed, for example, cells obtained from a blood bank. In this case, the composition may contain about 10 to 100 U / ml, for example, 10, 15, 20, 25, 30, 40, 50, 60, 70, 80, 90, or 100 U / ml of DNAse (e.g., human DNAse). Alternatively, the cells may be obtained fresh from a donor and not frozen, in which case DNAse is not essential and the composition may not contain DNAse. In the second embodiment, the present invention provides a packaging product containing the above-described components in a container comprising a substrate; the substrate having a polymer. In some examples, the polymer may be polytetrafluoroethylene (PTFE), perfluoroalkoxy polymer (PFA), fluorinated ethylene propylene (FEP), polyvinylidene fluoride (PVDF), polyethylene, or polyvinyl chloride (PVC) with low friction or non-stick properties. The polymer may also be other biologically suitable polymers, such as ultra-low density polyethylene, low-density polyethylene (LDPE), linear low-density polyethylene (LLDPE), high-density polyethylene (HDPE), coaxially oriented polypropylene (COPP), biaxially oriented polypropylene (BOPP), polyethylene terephthalate (PET), such as nylon, ethylene-vinyl alcohol polymer (EVOH), and metallized polyimide resins thereof. The packaged product can be in any suitable shape, including but not limited to bags, syringes, or vials for syringes. In one example, the product is pre-filled with pluripotent cells for clinical use. The pluripotent cells can be stem cells such as hematopoietic stem cells or mesenchymal stem cells. The composition may contain peripheral blood cells, umbilical cord blood cells, or bone marrow cells. The present invention provides a method for manufacturing the aforementioned packaging product. The method includes the following steps: (a) providing a composition containing cells (e.g., pluripotent cells); (b) providing a container comprising a substrate, wherein the substrate comprises a polymer; (c) placing the composition within the container; and (d) sealing the container. In a third embodiment, the present invention provides a method for storing or transporting therapeutic cells, such as pluripotent cells or monocytes. The method comprises (i) providing the therapeutic composition; and (ii) storing or transporting the composition at a temperature in the range of 1 to 10°C for about 24 to 96 hours, for example, about 24 to about 72 hours. In some specific examples, the method includes the steps of providing the packaged product and delivering the packaged product to a recipient, such as a messenger, agent, or receiving hospital personnel. During this delivery step, the temperature can be in the range of 1 to 10°C, for example, 1 to 7°C, 2 to 6°C, 3 to 5°C, or about 4°C. Using this method, the cells can be delivered within 12 to 96 hours, for example, at least 24, 36, 38, 60, 72, 84, or 96 hours. In one instance, the therapeutic component is stored or transported for about 72 hours. Upon delivery, the pluripotent cells or monocytes may have a total connate cell number (TNC) recovery rate of more than 40% (e.g., 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, and 95%). Furthermore, upon delivery, the pluripotent cells may have a viability rate of more than 60% (e.g., 65%, 70%, 75%, 80%, 85%, 90%, and 95%) (e.g., as determined by the AO / PI method disclosed herein). In one specific example, when delivered, the pluripotent cell or monocyte may have at least 0.25% CD34+CD45+ cells, for example, 0.25%, 0.3%, 0.35%, 0.4%, 0.45%, 0.5%, 0.55%, 0.6%, 0.65%, 0.7%, 0.75%, 0.8%, 0.85%, 0.9%, 0.95%, 1.0%, 1.1%, 1.15%, 1.2%, 1.3%, 1.4%, 1.5%, 1.6%, 1.7%, 1.8%, 1.9%, or 2.0%. In another specific example, when delivered, the pluripotent cell or monocyte may have at least 0.10% CD133+ cells, for example, 0.10%, 0.25%, 0.3%, 0.35%, 0.4%, 0.45%, 0.5%, 0.55%, 0.6%, 0.65%, 0.7%, 0.75%, 0.8%, 0.85%, 0.9%, 0.95%, 1.0%, 1.1%, 1.15%, 1.2%, 1.3%, 1.4%, 1.5%, 1.6%, 1.7%, 1.8%, 1.9%, or 2.0%. In another specific example, upon delivery, the pluripotent or mononuclear cell is capable of forming a large number of colony-forming units (CFUs) per plate. For example, as shown in Figure 1, after being stored or transported at about 2 to about 8°C for more than 72 hours, the cell is capable of forming at least 30 CFUs per 3 x 10⁴ seeded cells, for example, 40, 50, 60, 70, 80, 90, 95, 100, or 110. The above values ​​can be determined according to methods known in the technical field to which the invention pertains or the methods disclosed in the following embodiments. Details of one or more specific embodiments of the present invention are set forth in the following detailed description. Other features, objectives, and advantages of the present invention will become apparent from the specification and the claims. [Simplified Explanation of the Diagram] Figures 1A, 1B, 1C and 1D are a set of diagrams showing the results of stability studies of cells after storage or transportation in saline or the composition of the present invention at room temperature or 4°C.

Implementation Method

Claims

1. A therapeutic composition comprising (i) about 10⁹ / ml of umbilical cord blood cells; and (ii) a medically acceptable carrier solution, which (a) contains about 25 to 30 mM acetate and about 20 to 25 mM gluconate, and (b) has an osmolarity of about 270 to 320 mOsmol / L, wherein, This therapeutic composition does not contain DMSO.

2. The therapeutic composition as described in claim 1, wherein, The medically acceptable carrier solution contains one or more of the following: approximately 120 to 160 mM Na+, approximately 3 to 7 mM K+, approximately 1.0 to 2.0 mM Mg2+, and approximately 90 to 110 mM Cl-.

3. The therapeutic composition as described in claim 2, wherein, The medically acceptable carrier solution is free of Ca2+ or lactate or both.

4. The therapeutic composition as described in claim 2, wherein, The medically acceptable carrier solution contains approximately 140 mM Na+, approximately 5 mM K+, approximately 1.5 mM Mg2+, approximately 98 mM Cl-, approximately 27 mM acetate, and approximately 23 mM gluconate.

5. The therapeutic composition as described in claim 4, wherein, The medically acceptable carrier solution contains approximately 90 mM sodium chloride (NaCl), approximately 5 mM potassium chloride (KCl), approximately 1.5 mM magnesium chloride (MgCl2˙6H2O), approximately 27 mM sodium acetate trihydrate (C2H3NaO2˙3H2O), and approximately 23 mM sodium gluconate (C6H11NaO7).

6. The therapeutic composition as described in claim 1, wherein, The medically acceptable carrier solution contains 126 to 154 mEq / L of sodium.

7. The therapeutic composition as described in any one of claims 1 to 6, wherein, The medically acceptable carrier solution has a pH of 5.5 to 8.

0.

8. The therapeutic composition as described in any one of claims 1 to 6, wherein, The umbilical cord blood cell line is CD34+.

9. A packaged product comprising: a composition as described in any one of claims 1 to 8, and a container containing the composition and comprising a substrate, wherein, The substrate contains polymers.

10. The packaged product as described in claim 9, wherein, The container is a bag, tube, syringe, or vial for use with syringes.

11. The packaged product as described in claim 10, wherein, The container is sealed.

12. A method for storing or transporting pluripotent cells, comprising (i) providing a therapeutic composition as described in any one of claims 1 to 8 or a packaged product as described in any one of claims 9 to 11, and (ii) storing or transporting the composition at a temperature in the range of 1 to 10°C for about 24 to 96 hours.

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