Methods for allogeneic hematopoietic stem cell transplantation

Abstract

Various embodiments of the present disclosure provide therapeutic compositions and related methods comprising Tregs and Tcons for improving hematopoietic stem cell transplantation, including methods for enhancing protection from graft-versus-host disease while maintaining an effective immune response, such as a graft-versus-tumor immune response.

Description

[Background technology]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit of priority to U.S. Provisional Patent Application No. 63 / 331,709, filed April 15, 2022, and U.S. Provisional Patent Application No. 63 / 445,979, filed February 15, 2023, each of which is incorporated by reference in its entirety for all purposes.

[0002] background For example, patients with hematological malignancies such as leukemia and lymphoma who do not achieve first remission or who experience refractory relapses are rarely cured with standard chemotherapy. Myeloablative allogeneic hematopoietic cell transplantation (alloHCT) improves survival in these patients. Myeloablative alloHCT involves chemotherapy or radiation to remove or destroy the tissue in the bone that is causing the malignancy. The patient then receives hematopoietic cells, including hematopoietic stem and progenitor cells (HSPCs) derived from the donor's blood. However, alloHCT has a major drawback in that it frequently results in graft-versus-host disease (GVHD). GVHD is a condition in which the transplanted donor's peripheral blood stem cells perceive the patient's body as foreign, and the donor cells attack the patient's tissues (e.g., skin, GI tissue, liver tissue, and lung tissue), resulting in numerous complications, many of which are severe and can result in morbidity and mortality. Thus, there is a need for improved methods and therapies for hematopoietic cell transplantation that reduce the incidence and severity of GVHD, including reducing morbidity and mortality. Summary of the Invention

[0003] Quick Overview Certain aspects of the present disclosure relate to multi-component medical treatments administered to a human subject in need thereof. In some embodiments, the multi-component treatment comprises: (a) a CD45+ cell line comprising hematopoietic stem progenitor cells (HSPCs) and granulocytes; + A solution containing a first population of cells, +(b) a solution containing a cell population enriched for regulatory T cells (Treg); (c) a solution containing up to about 10% of the first population of cells containing granulocytes; + a solution containing a third population of cells, + A third population of cells is at least about 20% CD3 + (d) a solution comprising one or more doses of a prophylactic agent for graft-versus-host disease (GVHD).

[0004] In some embodiments that may be combined with any of the preceding embodiments, the GVHD prophylactic agent comprises tacrolimus and / or its analogs and derivatives, which may be formulated according to various embodiments for oral or intravenous administration to a human subject, or other administration or delivery methods known in the pharmaceutical arts, including, for example, intramuscular, transdermal, nasal, buccal, and intravaginal administration. In some embodiments that may be combined with any of the preceding embodiments, tacrolimus inhibits CD45 + and in an amount that maintains a target blood level of at least about 3 ng / ml of blood in the human subject for at least about 20 days after administration of the third population of cells, + and / or in an amount that maintains a target blood level of about 4 ng / ml or greater for at least about 40 days after administration of the third population of cells. + In some cases, which may be combined with any of the preceding embodiments, tacrolimus may be administered in an amount that maintains a target blood level of about 4 ng / ml or greater for at least about 40 days after administration of the third population of cells. + In some embodiments that may be combined with any of the preceding embodiments, tacrolimus is administered in an amount that maintains a target blood level of up to about 10 ng / ml for at least 30 days after administration of the third population of cells. + for at least about 60 days after administration of the third population of cells, +for at least about 90 days after administration of the third population of cells, + The third population of cells is administered for up to about 150 days after administration, and the CD45 + In some embodiments that may be combined with any of the preceding embodiments, the CD45 + The first population of cells comprises at least about 0.5% granulocytes, at least about 1% granulocytes, up to about 5% granulocytes, up to about 3% granulocytes, up to about 3% monocytes, up to about 2% monocytes, up to about 0.5% lymphocytes, up to about 2% lymphocytes, at least about 15% granulocytes, at least about 20% granulocytes, up to about 35% granulocytes, up to about 30% granulocytes, up to about 25% granulocytes, at least about 15% monocytes, at least about 20% monocytes, up to about 35% monocytes, up to about 30% monocytes, up to about 25% monocytes, at least about 0.5% NK cells, and / or at least about 2% NK cells. In some embodiments that may be combined with any of the preceding embodiments, CD45 + A third population of cells contains at least about 0.1% CD34 + cells, or approximately 0.2% to approximately 20% CD34 +In some embodiments that may be combined with any of the preceding embodiments, the multicomponent medical treatment further comprises a conditioning regimen, wherein the conditioning regimen is administered prior to any of the above-listed components (a) through (d). For example, the conditioning regimen is administered about 2 to about 10 days prior to any of (a) through (d). In some embodiments that may be combined with any of the preceding embodiments, the conditioning regimen is a myeloablative conditioning regimen. In some embodiments that may be combined with any of the preceding embodiments, the conditioning regimen comprises at least three conditioning reagents, wherein at least one conditioning reagent is thiotepa. In some embodiments that may be combined with any of the preceding embodiments, the myeloablative conditioning regimen comprises at least one dose of thiotepa, e.g., at least about 5 milligrams of thiotepa per kilogram of the human subject's actual or ideal body weight, or at least about 10 milligrams of thiotepa per kilogram of the human subject's actual or ideal body weight. In some embodiments that may be combined with any of the preceding embodiments, the transplant conditioning treatment comprises one or more doses of busulfan, fludarabine, and thiotepa. In some embodiments that may be combined with any of the preceding embodiments, the one or more doses include about 5 milligrams to about 12 mg of thiotepa per kg of the human subject's actual or ideal body weight, about 7 to about 11 mg of busulfan per kg of the human subject's actual or ideal body weight, and about 10 mg of thiotepa per kg of the human subject's actual or ideal body weight per m of body surface area. 2 In some embodiments that may be combined with any of the preceding embodiments, the CD45 +The first population of cells contains less than about 5 EU of endotoxin per ml of solution, less than about 1 EU of endotoxin per ml of solution, and / or less than about 0.5 EU of endotoxin per ml of solution. In some embodiments that may be combined with any of the preceding embodiments, the Treg-enriched cell population contains less than about 5 EU of endotoxin per ml of solution, less than about 1 EU of endotoxin per ml of solution, and / or less than about 0.5 EU of endotoxin per ml of solution. In some embodiments that may be combined with any of the preceding embodiments, the CD45 + The third population of cells contains less than about 5 EU of endotoxin per ml of solution, less than about 1 EU of endotoxin per ml of solution, and / or less than about 0.5 EU of endotoxin per ml of solution. In some embodiments that may be combined with any of the preceding embodiments, the HSPCs are CD34 + In some embodiments that may be combined with any of the preceding embodiments, Tregs are CD4 + CD25 + CD127dimor CD4 + FOXP3 + In some embodiments that may be combined with any of the preceding embodiments, the Treg-enriched cell population is CD45 + cells, e.g., CD45 + In some embodiments that may be combined with any of the preceding embodiments, the Treg-enriched cell population comprises about 1 x 10 cells per kilogram of actual or ideal body weight of a human subject. 5 ~Approx. 1×10 7 Tregs, or approximately 5 x 10 per kilogram of actual or ideal body weight in a human subject. 5 ~Approx. 4×10 6 In some embodiments that may be combined with any of the preceding embodiments, the HSPCs comprise CD45 Tregs. + The first population of cells is approximately 5 x 10 per kilogram of ideal body weight for a human subject. 5 ~Approx. 2×10 7In some embodiments that may be combined with any of the preceding embodiments, the HSPCs comprise CD45 + The third population of cells is approximately 1 x 10 per kilogram of actual or ideal body weight of a human subject. 5 ~Approx. 1×10 7 containing Tcon or CD45 + The third population of cells is approximately 5 x 10 per kilogram of actual or ideal body weight of a human subject. 5 ~about 5x10 6 In some embodiments that may be combined with any of the preceding embodiments, + The first population of cells, and the Treg-enriched population of cells, expresses CD45 + In some embodiments that may be combined with any of the preceding embodiments, the first dose of the one or more doses of GVHD prophylactic agent is administered prior to the third population of cells. + It is administered after the administration of the third population of cells.

[0005] Another aspect of the present disclosure is a method of treating a human subject diagnosed with a hematological malignancy. In some embodiments, the method comprises administering to the human subject a CD45 + a solution containing a first population of cells, a solution containing a cell population enriched for regulatory Tregs, CD45 + In some embodiments, the method further comprises administering a solution comprising a third population of cells and one or more doses of a GVHD prophylactic agent. + a solution containing a first population of cells, a solution containing a cell population enriched for regulatory Tregs, CD45 + The solution comprising the third population of cells and the solution comprising one or more doses of a GVHD prophylactic agent are as defined according to any of the multi-component medical treatments disclosed herein.

[0006] In some embodiments that may be combined with any of the preceding embodiments, the hematological malignancy may correspond to one or more of acute lymphocytic leukemia, acute myeloid leukemia, chronic myeloid leukemia, multiple myeloma, lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, myelodysplastic syndrome, myelofibrosis, and blastic plasmacytoid dendritic cell neoplasm (BPDCN). In some embodiments that may be combined with any of the preceding embodiments, the human subject is administered CD45 + A first population of cells, a Treg-enriched cell population, and CD45 + In some embodiments that may be combined with any of the preceding embodiments, a third population of cells is injected. + The third population of cells is CD45 + may be administered at least about 12 hours after the first population of cells, + The third population of cells is CD45 + about 24 to about 96 hours after the first population of cells, + The third population of cells is CD45 + Approximately 36 to 60 hours after the first population of cells is administered, and CD45 + The third population of cells is administered at least about 12 hours after the Treg-enriched cell population and expresses CD45 + The third population of cells is administered about 24 to about 96 hours after the Treg-enriched cell population, and / or is CD45 + The third population of cells is administered about 36 to about 60 hours after the Treg-enriched cell population. + The human subject does not develop GVHD higher than stage 2 within about 100 days of administration of the third population of cells, and the human subject is + and the human subject does not develop GVHD greater than Stage 2 within about 180 days or about 200 days after administration of the third population of cells, and the human subject is +and the human subject does not develop GVHD higher than stage 2 within about one year of administration of the third population of cells. In some embodiments that may be combined with any of the preceding embodiments, the human subject has been previously treated for or is concurrently treated for a hematological malignancy. In some embodiments that may be combined with any of the preceding embodiments, the GVHD prophylactic agent is tacrolimus (and / or analogs and derivatives thereof) and is initially administered to the human subject at about 0.03 mg / kg / day of the human subject's actual or ideal body weight, or the tacrolimus is a CD45 + The first administration is about 12 hours to about 24 hours after administration of the third population of cells. In some embodiments that may be combined with any of the preceding embodiments, the dose of tacrolimus may begin tapering about 90 days after the first dose is administered to the human subject, or the dose of tacrolimus begins tapering about 45 days after the first dose is administered to the human subject. In some embodiments that may be combined with any of the preceding embodiments, the CD45 + A first population of cells, a Treg-enriched cell population, and CD45 + The third population of cells is obtained from a single donor in one day or over several days. In some embodiments that may be combined with any of the preceding embodiments, at least one mobilized donated peripheral blood is collected from the donor, or up to two mobilized donated peripheral blood is collected from the donor. In some embodiments that may be combined with any of the preceding embodiments, at least one of the mobilized donated peripheral blood is CD34 +The mobilized donated peripheral blood is processed and selected to enrich for T cells and Tregs. In some embodiments that may be combined with any of the preceding embodiments, the time to process and select one or more of the mobilized donated peripheral blood is less than about 35 hours, the time to process and select one or more of the mobilized donated peripheral blood is less than about 30 hours, the time to process and select one or more of the mobilized donated peripheral blood is less than about 25 hours, the time to process and select one or more of the mobilized donated peripheral blood is up to about 35 hours, and / or the time to process and select one or more of the mobilized donated peripheral blood is up to about 25 hours. In some embodiments that may be combined with any of the preceding embodiments, one or more of the mobilized donated peripheral blood is processed and selected using one or more immune-separation particles (ISPs). For example, the ISPs include affinity reagents, such as immunomagnetic separation particles, which may be antibodies each conjugated to an iron-containing particle. In some embodiments, the affinity reagent comprises a plurality of CD34 reagents (e.g., anti-CD34 antibodies) that bind to one or more CD34 receptors on HSPCs. In some embodiments that may be combined with any of the preceding embodiments, the average number of ISPs per HSPC in the HSPC cell population is less than about 20,000, the average number of ISPs per HSPC in the HSPC cell population is about 10,000 or less, and / or the average number of ISPs per HSPC in the HSPC cell population is about 1500 to about 20,000. In some embodiments that may be combined with any of the preceding embodiments, the affinity reagent comprises a plurality of CD25 reagents (e.g., anti-CD25 antibodies) that bind to one or more CD25 receptors on Tregs. In some embodiments that may be combined with any of the preceding embodiments, the average number of ISPs per Treg cell in the Treg population is about 4000 or less, or the average number of ISPs per Treg cell in the Treg population is about 1500 to about 2500. In some embodiments that may be combined with any of the preceding embodiments, the mobilized donated peripheral blood cells are CD45 +The first population of cells is selected to contain up to about 10% granulocytes. In some embodiments that may be combined with any of the preceding embodiments, the mobilized donated peripheral blood cells are selected to contain CD45 + In some embodiments that may be combined with any of the preceding embodiments, the mobilized donor-donated peripheral blood cells are selected to contain up to about 7% granulocytes. + In some embodiments that may be combined with any of the preceding embodiments, the mobilized donor-donated peripheral blood cells are selected to contain up to about 4% monocytes. + In some embodiments that may be combined with any of the preceding embodiments, the mobilized donor-provided peripheral blood cells are sorted to include at most about 10% CD25 monocytes, such that the Treg-enriched population includes at most about 0.1% monocytes. - In some embodiments that may be combined with any of the preceding embodiments, the cells are selected to contain CD45 + a first population of cells, a Treg-enriched cell population, and / or CD45 + The third population of cells is allogeneic to the human subject. + a first population of cells, a Treg-enriched cell population, and / or CD45 + The third population of cells is obtained from an HLA-matched donor to the human subject. + a first population of cells, a Treg-enriched cell population, and / or CD45 + The third population of cells is obtained from a donor who is HLA-mismatched to the human subject. + a first population of cells, a Treg-enriched cell population, and / or CD45 + The third population of cells is obtained from a donor who is haploidentical to the human subject.+ A third population of cells is invariant natural killer T cells (iNKT), e.g., CD3 + Vα24Jα18 + In some embodiments that may be combined with any of the preceding embodiments, the population of iNKT is about 5 x 10 per kilogram of actual or ideal body weight of the human subject. 2 In some embodiments that may be combined with any of the preceding embodiments, the population of iNKT comprises more than about 5x10 iNKT per kilogram of actual or ideal body weight of a human subject. 2 ~approx. 1x10 7 In some embodiments that may be combined with any of the preceding embodiments, the present invention comprises a CD45 + A third population of cells is memory T cells (Tmem), e.g., CD3 + CD45RA - CD45RO + In some embodiments that may be combined with any of the preceding embodiments, the population of Tmems is about 3 x 10 per kilogram of actual or ideal body weight of a human subject. 5 In some embodiments that may be combined with any of the preceding embodiments, the population of Tmems is greater than about 3x10 per kilogram of actual or ideal body weight of a human subject. 5 ~approx. 1x10 9 Contains Tmem.

[0007] Other aspects of the present disclosure relate to multi-component medical treatments in which a human subject experiences a reduced risk and / or severity of adverse events associated with the multi-component medical treatment compared to a similar method in which Tcon but not Tregs are administered, or a multi-component medical treatment in which a human subject experiences a reduced risk and / or severity of adverse events associated with the method compared to a similar method in which Tcon but not Tregs are administered.

[0008] In some embodiments that may be combined with any of the preceding embodiments, the adverse event is acute GVHD (aGVHD), which may include stage 2 or higher aGVHD. In some embodiments that may be combined with any of the preceding embodiments, the adverse event is chronic GVHD (cGVHD), which may be moderate to severe cGVHD. In some embodiments that may be combined with any of the preceding embodiments, the human subject does not have cGVHD about one year after receiving the cell population. In some embodiments that may be combined with any of the preceding embodiments, the adverse event is a recurrence of malignancy in the human subject. In some embodiments that may be combined with any of the preceding embodiments, the human subject does not have a recurrence of malignancy about one year after receiving the dosing regimen. In some embodiments that may be combined with any of the preceding embodiments, the human subject underwent myeloablative conditioning prior to administration of any of the cell populations, and the adverse event is associated with the myeloablative conditioning. In some embodiments that may be combined with any of the preceding embodiments, the method further includes providing instructions for use (IFU), where the IFU includes instructions for administering the cell population to the patient. In some embodiments that may be combined with any of the preceding embodiments, the IFU also includes instructions for administering one or more pharmaceutical agents or pharmaceutical compositions to the patient.

[0009] Another aspect of the present disclosure relates to a method of transplanting conventional T cells (Tcon) as part of a treatment regimen for a hematological malignancy, which method reduces the risk and / or severity of adverse events associated with the treatment regimen. In some embodiments, the method includes administering to the patient a population of regulatory T cells (Tregs), including Tregs and a liquid suspending the Tregs; or administering to the patient a heterogeneous cell population including lymphocytes, granulocytes, monocytes, and a liquid suspending the cells. In some embodiments, at least about 30% of the lymphocytes comprise Tcon, and following administration of the cell population, the patient experiences a reduced risk and / or severity of adverse events compared to a patient with a hematological malignancy who received Tcon but not Tregs.

[0010] Another aspect of the present disclosure relates to a method of transplanting a cell population into a human patient as part of a therapeutic regimen for a hematological malignancy, the method reducing the risk and / or severity of adverse events associated with the therapeutic regimen. In some embodiments, the method includes providing a population of hematopoietic stem progenitor cells (HSPCs) administered to the patient, the population of HSPCs comprising HSPCs and a liquid suspending the HSPCs; providing a population of regulatory T cells (Tregs) administered to the patient, the population of Tregs comprising Tregs and a liquid suspending the Tregs; and providing a heterogeneous cell population administered to the patient, the heterogeneous cell population comprising lymphocytes, granulocytes, monocytes, and a liquid suspending the cells. In some embodiments, at least about 30% of the lymphocytes comprise conventional T cells (Tcon), and following administration of the cell population, the patient experiences a reduced risk and / or severity of adverse events compared to a patient with a hematological malignancy who received the Tcon cell population but not the Treg cell population.

[0011] In some embodiments that may be combined with any of the preceding embodiments, the cell populations are administered to the patient by intravenous infusion. In some embodiments that may be combined with any of the preceding embodiments, each cell population is provided as a separate cell population and is derived from a single human blood donor. In some embodiments that may be combined with any of the preceding embodiments, the adverse event is acute graft-versus-host disease (aGVHD), e.g., Stage 2 or higher aGVHD. In some embodiments that may be combined with any of the preceding embodiments, the patient does not have Stage 2 or higher aGVHD about 180 days after receiving the cell population. In some embodiments that may be combined with any of the preceding embodiments, the adverse event is chronic graft-versus-host disease (cGVHD). In some embodiments that may be combined with any of the preceding embodiments, the patient does not have cGVHD about one year after receiving the cell population. In some embodiments that may be combined with any of the preceding embodiments, the adverse event is moderate to severe cGVHD. In some embodiments that may be combined with any of the preceding embodiments, the patient is free of moderate to severe cGVHD about one year after administration of the cell population. In some embodiments that may be combined with any of the preceding embodiments, the adverse event is a recurrence of the patient's malignancy. In some embodiments that may be combined with any of the preceding embodiments, the patient is free of a recurrence of the malignancy about one year after administration of the cell population. In some embodiments that may be combined with any of the preceding embodiments, the adverse event includes graft-versus-host disease (GVHD) and a recurrence of the patient's malignancy. In some embodiments that may be combined with any of the preceding embodiments, the patient is free of GVHD and a recurrence of the malignancy one year after administration of the cell population. In some embodiments that may be combined with any of the preceding embodiments, at least one of the cell populations contains less than about 5 EU of endotoxin per mL of each suspension. In some embodiments that may be combined with any of the preceding embodiments, the patient has undergone a myeloablative conditioning regimen prior to administration of the cell population, and the adverse event is associated with the myeloablative conditioning regimen. In some embodiments that may be combined with any of the preceding embodiments, the adverse event includes a recurrence of a malignant tumor or an infection in the patient.In some embodiments that may be combined with any of the preceding embodiments, the heterogeneous cell population comprises about 0.2 to about 2.0 percent hematopoietic stem and progenitor cells. In some embodiments that may be combined with any of the preceding embodiments, the hematologic malignancy is leukemia, acute leukemia, acute myeloid leukemia (AML), acute lymphocytic leukemia (ALL), mixed phenotype acute leukemia (MPAL), chronic myeloid leukemia (CML), multiple myeloma, lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, myelodysplastic syndrome, myeloproliferative syndrome, myelofibrosis, or blastic plasmacytoid dendritic cell neoplasm (BPDCN). In some embodiments that may be combined with any of the preceding embodiments, the gene expression level of Treg cells correlates with cells harvested from the donor within about 60 hours prior to administration to the patient. In some embodiments that may be combined with any of the preceding embodiments, the number of Treg cells in the Treg population is approximately equal to the number of Tcon cells in the heterogeneous cell population. In some embodiments that may be combined with any of the preceding embodiments, the Treg cells in the Treg population inhibit activation of conventional T cells in the heterogeneous cell population by up to approximately 20 percent of the patient's healthy tissue. In some embodiments that may be combined with any of the preceding embodiments, the patient's peripheral blood has a CD4+ upregulation level up to about 100 days after administration of the cell population compared to healthy human subjects who did not receive the cell population. +In some embodiments that may be combined with any of the preceding embodiments, at least about 50% of the cells in the HSPC cell population are colony-forming units. In some embodiments that may be combined with any of the preceding embodiments, at least one of the cell populations has an elevated amount of granulocyte colony-stimulating factor compared to unmobilized blood. In some embodiments that may be combined with any of the preceding embodiments, at least one of the cell populations is a heterogeneous cell population. In some embodiments that may be combined with any of the preceding embodiments, at least one of the cell populations has a plurality of immunoseparation particles (ISPs) bound to receptors on cells of the cell population. In some embodiments that may be combined with any of the preceding embodiments, the plurality of ISPs are immuno-magnetic separation particles. In various embodiments, the plurality of ISPs comprises an antibody conjugated to an iron-containing particle. In some embodiments that may be combined with any of the preceding embodiments, the population of Tcon is administered at least about 12 hours after the HSPC population. For example, the population of Tcon is administered about 24-96 hours after the population of HSPCs, or about 36-60 hours after the population of HSPCs. In some embodiments that may be combined with any of the preceding embodiments, the population of Tcon is administered at least about 12 hours after the population of cells comprising Tregs. For example, the population of Tcon is administered about 24-96 hours after the population of cells comprising Tregs, or about 36-60 hours after the population of cells comprising Tregs. In some embodiments that may be combined with any of the preceding embodiments, the methods disclosed herein further include administering to the patient a single graft-versus-host disease (GVHD) prophylaxis (GVHDPA) comprising tacrolimus (tacrolimus GHVDPA) for up to about 180 days, wherein the tacrolimus GHVDPA is administered to maintain tacrolimus concentrations in the patient's blood above a threshold during that period, and the risk and / or severity of GHVD is significantly reduced.In some embodiments that may be combined with any of the preceding embodiments, the threshold is greater than about 4 ng of tacrolimus per ml of patient blood, or the threshold is greater than about 5 ng of tacrolimus per ml of patient blood. In some embodiments that may be combined with any of the preceding embodiments, tacrolimus GHVDPA is administered to maintain a tacrolimus concentration in the patient's blood below an upper threshold during the period. In some embodiments that may be combined with any of the preceding embodiments, the upper threshold is less than about 10 ng of tacrolimus per ml of patient blood. In some embodiments that may be combined with any of the preceding embodiments, the patient has a reduced risk of at least one of malignant tumor recurrence, infection, or renal failure. In some embodiments that may be combined with any of the preceding embodiments, the patient does not develop GVHD within about 30 days of administering Tcon, does not develop GVHD within about 100 days of administering Tcon, does not develop GVHD within about 180 days of administering Tcon, and / or does not develop GVHD within about one year of administering Tcon. In some embodiments that may be combined with any of the preceding embodiments, tacrolimus graft-versus-host disease (GVHD) prophylaxis (GVHDPA) may be administered intravenously or orally to the patient. In some embodiments that may be combined with any of the preceding embodiments, administration of tacrolimus graft-versus-host disease (GVHD) prophylaxis (GVHDPA) is initiated about 12 to 24 hours after administration of Tcon. In some embodiments that may be combined with any of the preceding embodiments, tacrolimus GHVDPA is administered for up to about 90 days, and for up to about 60 days. In some embodiments that may be combined with any of the preceding embodiments, tacrolimus GHVDPA is initially administered to the patient at about 0.03 mg / kg of the patient's actual or ideal body weight / day. In some embodiments that may be combined with any of the preceding embodiments, the dose of tacrolimus GVHDPA administered to the patient begins to taper about 90 days after the first dose is administered to the patient, or begins to taper about 45 days after the first dose is administered to the patient.In some embodiments that may be combined with any of the preceding embodiments, the method further includes administering a myeloablative conditioning regimen to the patient prior to administration of any of the cell populations, the conditioning regimen comprising administering at least one conditioning agent to the patient. In some embodiments that may be combined with any of the preceding embodiments, the patient does not receive any irradiation as part of the myeloablative conditioning regimen. In some embodiments that may be combined with any of the preceding embodiments, the at least one conditioning agent is administered about 2 to about 10 days prior to administration of any of the cell populations. In some embodiments that may be combined with any of the preceding embodiments, the at least one conditioning agent is administered about 5 days prior to administration of any of the cell populations. In some embodiments that may be combined with any of the preceding embodiments, the at least one conditioning agent comprises thiotepa. In some embodiments that may be combined with any of the preceding embodiments, the dose of thiotepa administered to the patient ranges from about 5 to about 10 mg per kilogram of the patient's actual or ideal body weight. In some embodiments that may be combined with any of the preceding embodiments, the at least one conditioning agent comprises busulfan and fludarabine. In some embodiments that may be combined with any of the preceding embodiments, the doses of thiotepa, busulfan, and fludarabine administered to a patient are about 10 mg per kilogram of the patient's actual or ideal body weight, about 9.6 mg per kilogram of the patient's actual or ideal body weight, and about 1 m of body surface area, respectively. 2 In some embodiments that may be combined with any of the preceding embodiments, the method further includes providing instructions for use (IFU), wherein the IFU includes instructions for administering the cell population to the patient. In some embodiments that may be combined with any of the preceding embodiments, the IFU also includes instructions for administering one or more pharmaceutical agents or pharmaceutical compositions to the patient.

[0012] Another aspect of the present disclosure relates to a method of transplanting a cell population into a human patient as part of a treatment regimen for a hematological malignancy. In some embodiments, the method comprises administering to the patient a population of hematopoietic stem progenitor cells (HSPCs; the population of HSPCs includes HSPCs and a liquid suspending the HSPCs); administering to the patient a population of regulatory T cells (Tregs), the population of Tregs including Tregs and a liquid suspending the Tregs; and administering to the patient a heterogeneous cell population, the heterogeneous cell population including lymphocytes, granulocytes, monocytes, and a liquid suspending the cells, wherein the heterogeneous cell population comprises at least about 30 % of the lymphocytes comprise conventional T cells (Tcon); and administering to the patient a single graft-versus-host disease (GVHD) prophylaxis (GVHDPA) comprising tacrolimus (tacrolimus GHVDPA) for up to approximately 180 days, wherein the tacrolimus GHVDPA is administered to maintain a tacrolimus concentration in the patient's blood above a threshold during that period, wherein the risk and / or severity of GHVD associated with the treatment regimen for the hematologic malignancy is significantly reduced. Another aspect of the present disclosure is CD45 + a first container containing a solution containing a first population of cells, the CD45 + In some embodiments, the kit includes a second container containing a solution comprising a third population of cells, and a third container containing a solution comprising a cell population enriched for regulatory T cells (Tregs). + A solution containing a first population of cells, CD45 + The solution comprising the third population of cells and the solution comprising the Treg-enriched cell population are as defined according to any multi-component medical treatment or method disclosed herein. In some embodiments, the kit further comprises a fourth container comprising a GVHD prophylactic agent. In some embodiments that may be combined with any of the preceding embodiments, the kit further comprises instructions for practicing any of the methods disclosed herein.

[0013] Another aspect of the present disclosure is (a) CD34 +(b) one or more reagents for selecting regulatory T cells (Tregs) from the mobilized peripheral blood composition; (c) a high number of CD3 + and (d) a solution comprising one or more doses of a graft-versus-host disease (GVHD) prophylactic agent. In some embodiments, the kit further comprises instructions for practicing any of the methods disclosed herein.

[0014] Another aspect of the present disclosure relates to a method of transplanting a population of conventional T cells (Tcon) into a human subject without eliciting a stage 2 or higher graft-versus-host disease (GVHD) response up to about 100 days after transplantation. In some embodiments, the method comprises: (i) administering a heterogeneous cell population comprising lymphocytes, granulocytes, and monocytes, wherein at least about 30% of the lymphocytes comprise conventional T cells (Tcon); and (ii) administering a population of regulatory T cells (Treg). In some embodiments, the heterogeneous cellular component and / or the Treg population comprises less than about 5 EU / ml of endotoxin.

[0015] Another aspect of the present disclosure relates to a method of treating a human subject, comprising: (a) administering a plurality of cell populations, the plurality of cell populations comprising: (i) a population of hematopoietic stem progenitor cells (HSPCs); (ii) a cell population comprising regulatory T cells (Tregs); and (iii) a population of conventional T cells (Tcons); and (b) administering a single graft-versus-host disease (GVHD) prophylactic agent for less than about 120 days. In some embodiments, the HSPC population comprises less than about 2% CD3 + Contains cells.

[0016] Another aspect of the present disclosure relates to a method of treating a human subject in need thereof, the method comprising administering to the human subject at least two pharmaceutical compositions, the pharmaceutical compositions being selected from: (a) a pharmaceutical composition comprising a population of hematopoietic stem progenitor cells (HSPCs); (b) a pharmaceutical composition comprising a population of regulatory T cells (Tregs); and (c) a pharmaceutical composition comprising a population of conventional T cells (Tcon). In some embodiments, pharmaceutical compositions (a), (b), and (c) each contain less than about 5 EU / ml endotoxin, and in a group of 100 human subjects administered two or more pharmaceutical compositions, fewer than 15 human subjects develop a stage 2 or higher graft-versus-host disease (GVHD) response within about 30 days of receiving the pharmaceutical composition comprising the Tcon population.

[0017] Another aspect of the present disclosure relates to a method of transplanting a population of conventional T cells (Tcon) into a human subject without initiating a stage 2 or higher graft-versus-host disease (GVHD) response up to about 100 days after transplantation. In some embodiments, the method includes (i) administering a solution comprising a population of conventional T cells (Tcon), and (ii) administering a solution comprising a population of regulatory T cells (Treg). In some embodiments, the population of Tcon has been cryopreserved for at least about 4 hours, and the solution comprising the population of Tcon and the solution comprising the population of Treg contain less than about 5 EU of endotoxin per ml of solution.

[0018] Another aspect of the present disclosure relates to a method of treating a hematological malignancy in a human subject in need thereof, the method comprising administering to the human subject: (a) a population of hematopoietic stem progenitor cells (HSPCs); (b) a population of regulatory T cells (Tregs); and (c) a population of conventional T cells (Tcon). In some embodiments, the population of HSPCs and the population of Tregs are administered before the population of Tcon, and the peripheral blood of the human subject exhibits elevated numbers of Tregs up to about 100 days after administration of the three cell populations, compared to a healthy human subject that did not receive the three cell populations.

[0019] Another aspect of the present disclosure relates to a method of transplanting a population of conventional T cells (Tcon) into a human subject without initiating a stage 2 or higher graft-versus-host disease (GVHD) response up to about 100 days after transplantation. In some embodiments, the method includes (i) administering a population of conventional T cells (Tcon), and (ii) administering a population of regulatory T cells (Treg). In some embodiments, the population of Tcon is administered at least about 12 hours after the population of Treg is administered, and the population of Tcon and the population of Treg contain less than about 5 EU / ml of endotoxin.

[0020] Certain aspects of the present disclosure relate to methods of treating a human subject diagnosed with a hematological malignancy, the methods comprising administering to the human subject (a) CD45 containing hematopoietic stem progenitor cells (HSPCs); + (b) a solution containing a first population of cells; (c) a solution containing a cell population enriched for regulatory T cells (Treg); + a solution containing a third population of cells, + The third population of cells is CD3 + and (d) a solution containing one or more doses of a graft-versus-host disease (GVHD) prophylactic agent, wherein the solution contains CD45 + A first population of cells, a Treg-enriched cell population, and CD45 + The third population of cells is obtained from a single allogeneic donor with at least one HLA mismatch to the human subject, and the non-relapse mortality rate of the human subject is reduced and / or the overall survival of the human subject is increased following administration, compared to a corresponding human subject undergoing standard myeloablative allogeneic hematopoietic stem cell transplant (alloHSCT) from a donor with at least one HLA mismatch to the corresponding human subject.

[0021] In some embodiments, the hematological malignancy is selected from leukemia, acute leukemia, acute myeloid leukemia (AML), acute lymphocytic leukemia (ALL), mixed phenotype acute leukemia (MPAL), chronic myelogenous leukemia (CML), multiple myeloma, lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, myelodysplastic syndrome, myeloproliferative syndrome, myelofibrosis, and blastic plasmacytoid dendritic cell neoplasm (BPDCN). In some embodiments that may be combined with any of the preceding embodiments, the human subject is administered CD45 + A first population of cells, a Treg-enriched cell population, and CD45 + In some embodiments that may be combined with any of the preceding embodiments, a third population of cells is injected. + The third population of cells is CD45 + In some embodiments that may be combined with any of the preceding embodiments, the first population of cells is administered at least about 12 hours after the first population of cells. + The third population of cells is CD45 + In some embodiments that may be combined with any of the preceding embodiments, the first population of cells is administered about 24 to about 120 hours after the first population of cells. + The third population of cells is CD45 + In some embodiments that may be combined with any of the preceding embodiments, the first population of cells is administered about 36 to about 72 hours after the first population of cells. + The third population of cells is administered at least about 12 hours after the Treg-enriched cell population. + The third population of cells is administered about 24 to about 120 hours after the Treg-enriched cell population. + The third population of cells is administered about 36 to about 72 hours after the Treg-enriched cell population. In some embodiments that may be combined with any of the preceding embodiments, the HSPCs are CD34 + In some embodiments that may be combined with any of the preceding embodiments, CD45 +The first population of cells is approximately 5 x 10 per kilogram of actual or ideal body weight of a human subject. 5 ~Approx. 2×10 7 In some embodiments that may be combined with any of the preceding embodiments, Tregs comprise CD4 HSPCs. + CD25 + In some embodiments that may be combined with any of the preceding embodiments, the Treg-enriched cell population is CD45 + cells, including CD45 + In some embodiments that may be combined with any of the preceding embodiments, the Tregs are CD4 + CD25 + CD127dimor CD4 + FOXP3 + In some embodiments that may be combined with any of the preceding embodiments, the Treg-enriched cell population is about 1 x 10 per kilogram of actual or ideal body weight of a human subject. 5 ~Approx. 1×10 7 In some embodiments that may be combined with any of the preceding embodiments, the Tregs include CD45 + The third population of cells is about 1 x 10 per kilogram of the human subject's actual or ideal body weight. 5 ~Approx. 1×10 7 In some embodiments that may be combined with any of the preceding embodiments, the human subject comprises CD45 + In some embodiments that may be combined with any of the preceding embodiments, the human subject does not develop GVHD higher than stage 2 within about 100 days of administration of the third population of cells. + In some embodiments that may be combined with any of the preceding embodiments, the human subject does not develop GVHD greater than Stage 2 within about 180 days or about 200 days of administration of the third population of cells. +and does not develop GVHD higher than stage 2 within about one year of administration of the third population of cells. In some embodiments that may be combined with any of the preceding embodiments, the human subject has been previously treated for or is concurrently treated for a hematological malignancy. In some embodiments that may be combined with any of the preceding embodiments, the GVHD prophylactic agent is tacrolimus and is initially administered to the human subject at about 0.03 mg / kg of the human subject's actual or ideal body weight / day. In some embodiments that may be combined with any of the preceding embodiments, the GVHD prophylactic agent is CD45 + The first administration is about 12 to about 24 hours after administration of the third population of cells. In some cases that may be combined with any of the preceding embodiments, tacrolimus is administered to inhibit CD45 + In some cases that may be combined with any of the preceding embodiments, tacrolimus is administered in an amount that maintains a target blood level of at least about 3 ng / ml for at least about 20 days after administration of the third population of cells. + In some cases that may be combined with any of the preceding embodiments, tacrolimus is administered in an amount that maintains a target blood level of about 4 ng / ml or greater for at least about 40 days after administration of the third population of cells. + The administration of the third population of cells is continued for at least about 60 days, at least about 90 days, at least about 120 days, or up to about 160 days after administration of the third population of cells. In some embodiments that may be combined with any of the preceding embodiments, the administration of the GVHD prophylactic agent begins to taper about 90 days after the initial administration of the GVHD prophylactic agent. In some embodiments that may be combined with any of the preceding embodiments, the method further comprises administering about 1000 mg of mycophenolate mofetil (MMF). In some embodiments that may be combined with any of the preceding embodiments, MMF is administered to inhibit CD45 +The first administration is about 12 to about 24 hours after the administration of the third population of cells. In some embodiments that may be combined with any of the preceding embodiments, the administration of MMF begins to taper about 30 days, about 35 days, about 40 days, about 41 days, about 42 days, about 43 days, about 44 days, about 45 days, about 46 days, about 47 days, about 48 days, about 49 days, or about 50 days after the first administration of MMF. In some embodiments that may be combined with any of the preceding embodiments, the single allogeneic donor with at least one HLA mismatch is unrelated to the human subject. In some embodiments that may be combined with any of the preceding embodiments, the single allogeneic donor with at least one HLA mismatch is related to the human subject. In some embodiments that may be combined with any of the preceding embodiments, the method further includes collecting one or more or two or more mobilized donated peripheral blood samples from the donor. In some embodiments that may be combined with any of the preceding embodiments, the method further includes collecting up to two mobilized peripheral blood donations from the donor. In some embodiments that may be combined with any of the preceding embodiments, the donated peripheral blood is mobilized with granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), stem cell factor (SCF), SDF-1 antagonist, CXCR4 antagonist (e.g., POL6326, BKT-140, TG-0054, NOX-A12), Mozovir® (plerixafor), CXCR2 ligand (e.g., GROβ), sphingosine-1-phosphatase (S1P) agonist (e.g., SEW2871), VCAM / VLA-4 inhibitor (e.g., BIO5192), proteosome inhibitor (e.g., bortezomib), parathyroid hormone, hypoxia-inducible factor (HIF) stabilizer (e.g., FG-4497), and combinations thereof. In some embodiments that may be combined with any of the preceding embodiments, at least one of the mobilized donated peripheral blood is CD34 +The cells are treated and selected using one or more immunoseparation particles (ISPs) to enrich for Tregs. In some embodiments that may be combined with any of the preceding embodiments, one or more ISPs comprise an affinity reagent, optionally an immunomagnetic separation particle, optionally an antibody, each antibody conjugated to an iron-containing particle. In some embodiments that may be combined with any of the preceding embodiments, the average number of ISPs per HSPC in the HSPC cell population is about 20,000 or less. In some embodiments that may be combined with any of the preceding embodiments, the average number of ISPs per HSPC in the HSPC cell population is about 1,000 to about 20,000. In some embodiments that may be combined with any of the preceding embodiments, the affinity reagent comprises a plurality of CD25 reagents that bind to one or more CD25 receptors on Tregs. In some embodiments that may be combined with any of the preceding embodiments, the average number of ISPs per Treg cell in the Treg population is about 4,000 or less. In some embodiments that may be combined with any of the preceding embodiments, the average number of ISPs per Treg cell in the Treg population is about 1500 to about 2500. In some embodiments that may be combined with any of the preceding embodiments, the at least one HLA mismatch is at an allele selected from the following: HLA-A, HLA-B, HLA-C, DRB-1, and any combination thereof. In some embodiments that may be combined with any of the preceding embodiments, the allogeneic donor with at least one HLA mismatch is 6 / 8 HLA-mismatched to the human subject or 7 / 8 HLA-mismatched to the human subject. In some embodiments that may be combined with any of the preceding embodiments, the allogeneic donor is 7 / 8 HLA-mismatched to the human subject. In some embodiments that may be combined with any of the preceding embodiments, the allogeneic donor that is 7 / 8 HLA-mismatched to the human subject has a mismatch at HLA-A. In some embodiments that may be combined with any of the preceding embodiments, the allogeneic donor that is 7 / 8 HLA-mismatched to the human subject has a mismatch in HLA-B.In some embodiments that may be combined with any of the preceding embodiments, the allogeneic donor is 7 / 8 HLA-mismatched to the human subject and has a mismatch in HLA-C. In some embodiments that may be combined with any of the preceding embodiments, the allogeneic donor is 7 / 8 HLA-mismatched to the human subject and has a mismatch in HLA-DRB1. In some embodiments that may be combined with any of the preceding embodiments, the allogeneic donor has at least one HLA mismatch with the human subject and has a mismatched HLA allele because the allogeneic donor is homozygous for that HLA allele and the human subject is heterozygous for that HLA allele. In some embodiments that may be combined with any of the preceding embodiments, the allogeneic donor has at least one HLA mismatch with the human subject and has a mismatched HLA allele because the allogeneic donor is heterozygous for that HLA allele and the human subject is homozygous for that HLA allele. In some embodiments that may be combined with any of the preceding embodiments, the allogeneic donor having at least one HLA mismatch with the human subject has a mismatched HLA allele because both the allogeneic donor and the human subject are heterozygous for that HLA allele. In some embodiments that may be combined with any of the preceding embodiments, the method further includes a conditioning treatment, wherein the conditioning treatment is administered before any of (a)-(d). In some embodiments that may be combined with any of the preceding embodiments, the conditioning treatment is administered about 2 days to about 10 days before any of (a)-(d). In some embodiments that may be combined with any of the preceding embodiments, the conditioning transplant treatment is a myeloablative conditioning transplant treatment. In some embodiments that may be combined with any of the preceding embodiments, the myeloablative conditioning transplant treatment includes at least three conditioning transplant treatment reagents, wherein at least one conditioning transplant treatment reagent includes thiotepa. In some embodiments that may be combined with any of the preceding embodiments, the myeloablative conditioning transplant treatment includes one or more doses of busulfan, fludarabine, and thiotepa. In some embodiments that may be combined with any of the preceding embodiments, the dose of one or more of busulfan, fludarabine, and thiotepa is about 5 milligrams to about 12 mg of thiotepa per kg of the human subject's actual or ideal body weight, about 7 to about 11 mg of busulfan per kg of the human subject's actual or ideal body weight, and about 10 mg of busulfan per kg of the human subject's actual or ideal body weight per m of body surface area. 2 Each contains approximately 100 to 200 mg of fludarabine.

[0022] Another aspect of the present disclosure relates to a multi-component medical treatment administered to a human subject in need thereof, the multi-component treatment comprising: (a) a CD45+ cell line containing hematopoietic stem progenitor cells (HSPCs) and granulocytes; + a solution containing a first population of cells, the CD45 + (b) a solution containing a cell population enriched for regulatory T cells (Treg); (c) a solution containing up to about 10% of the first population of cells containing granulocytes; + a solution containing a third population of cells, + A third population of cells is at least about 20% CD3 + (d) a solution comprising one or more doses of a graft-versus-host disease (GVHD) prophylactic agent, wherein the solution comprises CD45 + A first population of cells, a Treg-enriched cell population, and CD45 + The third population of cells has at least one HLA mismatch with the human subject and, when administered, expresses CD45 + A first population of cells, a Treg-enriched cell population and CD45+ The third population of cells reduces non-relapse mortality in the human subject and / or increases overall survival of the human subject compared to non-relapse mortality and / or overall survival in a matched human subject who has undergone standard myeloablative allogeneic hematopoietic stem cell transplantation (alloHSCT) with at least one HLA mismatch to the matched human subject.

[0023] In some embodiments that may be combined with any of the preceding embodiments, the GVHD prophylactic agent is tacrolimus. In some embodiments that may be combined with any of the preceding embodiments, the one or more doses of the GVHD prophylactic agent are administered intravenously. + and tacrolimus in an amount that maintains a target blood level of at least about 3 ng / ml for at least about 20 days after administration of the third population of cells. In some embodiments that may be combined with any of the preceding embodiments, the treatment further comprises a conditioning transplant treatment. In some embodiments that may be combined with any of the preceding embodiments, the conditioning transplant treatment is a myeloablative conditioning transplant treatment. In some embodiments that may be combined with any of the preceding embodiments, the myeloablative conditioning transplant treatment comprises at least three conditioning transplant treatment reagents, wherein at least one conditioning transplant treatment reagent comprises thiotepa. In some embodiments that may be combined with any of the preceding embodiments, the myeloablative conditioning transplant treatment comprises doses of one or more of busulfan, fludarabine, and thiotepa, the doses being about 5 milligrams to about 12 mg of thiotepa per kg of the human subject's actual or ideal body weight, about 7 to about 11 mg of busulfan per kg of the human subject's actual or ideal body weight, and about 10 mg of busulfan per m of body surface area. 2 In some embodiments that may be combined with any of the preceding embodiments, the HSPCs are CD34 + In some embodiments that may be combined with any of the preceding embodiments, Tregs are CD4 + CD25 + In some embodiments that may be combined with any of the preceding embodiments, the Treg-enriched cell population is CD45+ cells, and + In some embodiments that may be combined with any of the preceding embodiments, the Tregs are CD4 + CD25 + CD127dimor CD4 + FOXP3 + In some embodiments that may be combined with any of the preceding embodiments, HSPCs containing CD45 + The first population of cells is about 5 x 10 per kilogram of the human subject's actual or ideal body weight. 5 ~Approx. 2×10 7 In some embodiments that may be combined with any of the preceding embodiments, the Treg-enriched cell population comprises about 1 x 10 HSPCs per kilogram of the human subject's actual or ideal body weight. 5 ~Approx. 1×10 7 In some embodiments that may be combined with any of the preceding embodiments, the Tregs include CD45 + The third population of cells is about 1 x 10 per kilogram of the human subject's actual or ideal body weight. 5 ~Approx. 1×10 7 In some embodiments that may be combined with any of the preceding embodiments, + The first population of cells, and the Treg-enriched population of cells, expresses CD45 + In some embodiments that may be combined with any of the preceding embodiments, the first dose of the one or more doses of GVHD prophylactic agent is formulated for administration prior to administration of the third population of cells. + In some embodiments that may be combined with any of the preceding embodiments, the treatment further comprises about 1000 mg of mycophenolate mofetil (MMF). In some embodiments that may be combined with any of the preceding embodiments, the MMF is a CD45 +The allogeneic donor is formulated for administration about 12 to about 24 hours after administration of the third population of cells. In some embodiments that may be combined with any of the preceding embodiments, the single allogeneic donor with at least one HLA mismatch is unrelated to the human subject. In some embodiments that may be combined with any of the preceding embodiments, the single allogeneic donor with at least one HLA mismatch is related to the human subject. In some embodiments that may be combined with any of the preceding embodiments, the at least one HLA mismatch is in an allele selected from the following: HLA-A, HLA-B, HLA-C, DRB-1, and any combination thereof. In some embodiments that may be combined with any of the preceding embodiments, the allogeneic donor with at least one HLA mismatch is 6 / 8 HLA-mismatched to the human subject or 7 / 8 HLA-mismatched to the human subject. In some embodiments that may be combined with any of the preceding embodiments, the allogeneic donor is 7 / 8 HLA-mismatched to the human subject. In some embodiments that may be combined with any of the preceding embodiments, the allogeneic donor is 7 / 8 HLA-mismatched to the human subject and has a mismatch in HLA-A. In some embodiments that may be combined with any of the preceding embodiments, the allogeneic donor is 7 / 8 HLA-mismatched to the human subject and has a mismatch in HLA-B. In some embodiments that may be combined with any of the preceding embodiments, the allogeneic donor is 7 / 8 HLA-mismatched to the human subject and has a mismatch in HLA-C. In some embodiments that may be combined with any of the preceding embodiments, the allogeneic donor is 7 / 8 HLA-mismatched to the human subject and has a mismatch in HLA-DRB1. In some embodiments that may be combined with any of the preceding embodiments, the allogeneic donor has at least one HLA mismatch with the human subject and has a mismatched HLA allele because the allogeneic donor is homozygous for that HLA allele and the human subject is heterozygous for that HLA allele.In some embodiments that may be combined with any of the preceding embodiments, an allogeneic donor with at least one HLA mismatch to a human subject has a mismatched HLA allele because the allogeneic donor is heterozygous for that HLA allele and the human subject is homozygous for that HLA allele. In some embodiments that may be combined with any of the preceding embodiments, an allogeneic donor with at least one HLA mismatch to a human subject has a mismatched HLA allele because both the allogeneic donor and the human subject are heterozygous for that HLA allele.

[0024] Another aspect of the present disclosure relates to a method of treating a human subject diagnosed with a hematological malignancy, the method comprising: (a) a myeloablative conditioning regimen comprising one or more doses of busulfan, fludarabine, and thiotepa; (b) a CD45+ / CD ... + (c) a solution containing a cell population enriched for regulatory T cells (Treg); (d) a solution containing a CD45 + a solution containing a third population of cells, + The third population of cells is CD3 + and (e) a solution comprising one or more doses of a graft-versus-host disease (GVHD) prophylactic agent, wherein the pre-transplant treatment is administered about 2 to about 10 days prior to any of (b) through (e). In some embodiments that may be combined with any of the preceding embodiments, the dose of one or more of busulfan, fludarabine, and thiotepa is about 5 milligrams to about 12 mg of thiotepa per kg of the human subject's actual or ideal body weight, about 7 to about 11 mg of busulfan per kg of the human subject's actual or ideal body weight, and about 10 mg of busulfan per kg of the human subject's actual or ideal body weight. 2In some embodiments that may be combined with any of the preceding embodiments, the human subject's GVHD- and relapse-free survival (GFRS) is increased by at least about 3.5-fold after administration, overall survival is increased by at least about 1.25-fold after administration, and / or non-relapse mortality is reduced by at least about 60% after administration, compared to a corresponding human subject undergoing standard allogeneic myeloablative hematopoietic stem cell transplant (alloHSCT).

[0025] Another aspect of the present disclosure relates to a method of treating a human subject diagnosed with a hematological malignancy, the method comprising administering to the human subject (a) CD45 containing hematopoietic stem progenitor cells (HSPCs); + (b) a solution containing a first population of cells; (b) a solution containing a cell population enriched for regulatory T cells (Treg); (c) a CD45 + a solution containing a third population of cells, + The third population of cells is CD3 +and (d) a solution comprising one or more doses of a graft-versus-host disease (GVHD) prophylaxis agent, wherein, compared to a corresponding human subject undergoing standard myeloablative allogeneic hematopoietic stem cell transplant (alloHSCT), the human subject's GVHD- and relapse-free survival (GFRS) is increased by at least about 3.5-fold after administration, overall survival is increased by at least about 1.25-fold after administration, and / or non-relapse mortality is reduced by at least about 60% after administration. In some embodiments that may be combined with any of the preceding embodiments, the multicomponent medical treatment further comprises a myeloablative transplant conditioning regimen comprising doses of one or more of busulfan, fludarabine, and thiotepa. In some embodiments that may be combined with any of the preceding embodiments, the transplant conditioning regimen is administered about 2 to about 10 days before any of (a)-(d). In some embodiments that may be combined with any of the preceding embodiments, the dose of one or more of busulfan, fludarabine, and thiotepa is about 5 milligrams to about 12 mg of thiotepa per kg of actual or ideal body weight of a human subject, about 7 to about 11 mg of busulfan per kg of actual or ideal body weight of a human subject, and about 10 mg of busulfan per kg of actual or ideal body surface area of ​​a human subject. 2 Each contains approximately 100 to 200 mg of fludarabine.

[0026] In some embodiments that may be combined with any of the preceding embodiments, the hematological malignancy is selected from leukemia, acute leukemia, acute myeloid leukemia (AML), acute lymphocytic leukemia (ALL), mixed phenotype acute leukemia (MPAL), chronic myelogenous leukemia (CML), multiple myeloma, lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, myelodysplastic syndrome, myeloproliferative syndrome, myelofibrosis, and blastic plasmacytoid dendritic cell neoplasm (BPDCN). In some embodiments that may be combined with any of the preceding embodiments, the human subject is administered CD45 + A first population of cells, a Treg-enriched cell population, and CD45 + In some embodiments that may be combined with any of the preceding embodiments, a third population of cells is injected.+ The third population of cells is CD45 + In some embodiments that may be combined with any of the preceding embodiments, the first population of cells is administered at least about 12 hours after the first population of cells. + The third population of cells is CD45 + In some embodiments that may be combined with any of the preceding embodiments, the first population of cells is administered about 24 to about 120 hours after the first population of cells. + The third population of cells is CD45 + In some embodiments that may be combined with any of the preceding embodiments, the first population of cells is administered about 36 to about 72 hours after the first population of cells. + The third population of cells is administered at least about 12 hours after the Treg-enriched cell population. + The third population of cells is administered about 24 to about 120 hours after the Treg-enriched cell population. + The third population of cells is administered about 36 to about 72 hours after the Treg-enriched cell population. In some embodiments that may be combined with any of the preceding embodiments, the HSPCs are CD34 + In some embodiments that may be combined with any of the preceding embodiments, CD45 + The first population of cells is approximately 5 x 10 per kilogram of actual or ideal body weight of a human subject. 5 ~Approx. 2×10 7 In some embodiments that may be combined with any of the preceding embodiments, Tregs comprise CD4 HSPCs. + CD25 + In some embodiments that may be combined with any of the preceding embodiments, the Treg-enriched cell population is CD45 + cells, including CD45 + In some embodiments that may be combined with any of the preceding embodiments, the Tregs are CD4 + CD25+ CD127dimor CD4 + FOXP3 + In some embodiments that may be combined with any of the preceding embodiments, the Treg-enriched cell population is about 1 x 10 per kilogram of actual or ideal body weight of a human subject. 5 ~Approx. 1×10 7 In some embodiments that may be combined with any of the preceding embodiments, the Tregs include CD45 + The third population of cells is about 1 x 10 per kilogram of the human subject's actual or ideal body weight. 5 ~Approx. 1×10 7 In some embodiments that may be combined with any of the preceding embodiments, the human subject comprises CD45 + In some embodiments that may be combined with any of the preceding embodiments, the human subject does not develop GVHD higher than stage 2 within about 100 days of administration of the third population of cells. + In some embodiments that may be combined with any of the preceding embodiments, the human subject does not develop GVHD greater than Stage 2 within about 180 days or about 200 days of administration of the third population of cells. + and does not develop GVHD higher than stage 2 within about one year of administration of the third population of cells. In some embodiments that may be combined with any of the preceding embodiments, the human subject has been previously treated for or is concurrently treated for a hematological malignancy. In some embodiments that may be combined with any of the preceding embodiments, the GVHD prophylactic agent is tacrolimus and is initially administered to the human subject at about 0.03 mg / kg of the human subject's actual or ideal body weight / day. In some embodiments that may be combined with any of the preceding embodiments, the GVHD prophylactic agent is CD45 + The first administration is about 12 to about 24 hours after administration of the third population of cells. In some cases that may be combined with any of the preceding embodiments, tacrolimus is administered to inhibit CD45 +In some cases that may be combined with any of the preceding embodiments, tacrolimus is administered in an amount that maintains a target blood level of at least about 3 ng / ml for at least about 20 days after administration of the third population of cells. + In some cases that may be combined with any of the preceding embodiments, tacrolimus is administered in an amount that maintains a target blood level of about 4 ng / ml or greater for at least about 40 days after administration of the third population of cells. + The administration of the third population of cells is continued for at least about 60 days, at least about 90 days, at least about 120 days, or up to about 160 days after administration of the third population of cells. In some embodiments that may be combined with any of the preceding embodiments, administration of the GVHD prophylactic agent begins to taper off about 90 days after the initial administration of the GVHD prophylactic agent. In some embodiments that may be combined with any of the preceding embodiments, the method further comprises collecting one or more, or two or more, mobilized donated peripheral blood samples from the donor. In some embodiments that may be combined with any of the preceding embodiments, the method further comprises collecting up to two mobilized donated peripheral blood samples from the donor. In some embodiments that may be combined with any of the preceding embodiments, the donated peripheral blood is mobilized with granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), stem cell factor (SCF), SDF-1 antagonist, CXCR4 antagonist (e.g., POL6326, BKT-140, TG-0054, NOX-A12), Mozovir® (plerixafor), CXCR2 ligand (e.g., GROβ), sphingosine-1-phosphatase (S1P) agonist (e.g., SEW2871), VCAM / VLA-4 inhibitor (e.g., BIO5192), proteosome inhibitor (e.g., bortezomib), parathyroid hormone, hypoxia-inducible factor (HIF) stabilizer (e.g., FG-4497), and combinations thereof. In some embodiments that may be combined with any of the preceding embodiments, at least one of the mobilized donated peripheral blood is CD34 +The cells are treated and selected using one or more immunoseparation particles (ISPs) to enrich for Tregs. In some embodiments that may be combined with any of the preceding embodiments, one or more ISPs comprise an affinity reagent, optionally an immunomagnetic separation particle, optionally an antibody, each antibody conjugated to an iron-containing particle. In some embodiments that may be combined with any of the preceding embodiments, the average number of ISPs per HSPC in the HSPC cell population is about 20,000 or less. In some embodiments that may be combined with any of the preceding embodiments, the average number of ISPs per HSPC in the HSPC cell population is about 1,000 to about 20,000. In some embodiments that may be combined with any of the preceding embodiments, the affinity reagent comprises a plurality of CD25 reagents that bind to one or more CD25 receptors on Tregs. In some embodiments that may be combined with any of the preceding embodiments, the average number of ISPs per Treg cell in the Treg population is about 4,000 or less. In some embodiments that may be combined with any of the preceding embodiments, the average number of ISPs per Treg cell in the Treg population is about 1500 to about 2500.

[0027] Another aspect of the present disclosure relates to a multicomponent medical treatment administered to a human subject in need thereof, the multicomponent treatment comprising: (a) a myeloablative conditioning regimen comprising one or more doses of busulfan, fludarabine, and thiotepa; (b) a CD45+ cell line comprising hematopoietic stem and progenitor cells (HSPCs); + (c) a solution containing a cell population enriched for regulatory T cells (Treg); (d) a solution containing a CD45 + a solution containing a third population of cells, + The third population of cells is CD3 + (b) a solution containing conventional T cells (Tcon); and (e) a solution containing one or more doses of a graft-versus-host disease (GVHD) prophylactic agent, wherein the transplant conditioning treatment is formulated to be administered about 2 to about 10 days before any of (b) to (e).

[0028] In some embodiments that may be combined with any of the preceding embodiments, the dose of one or more of busulfan, fludarabine, and thiotepa is about 5 milligrams to about 12 mg of thiotepa per kg of actual or ideal body weight of a human subject, about 7 to about 11 mg of busulfan per kg of actual or ideal body weight of a human subject, and about 10 mg of busulfan per kg of actual or ideal body surface area of ​​a human subject. 2 In some embodiments that may be combined with any of the preceding embodiments, the GVHD prophylactic agent is tacrolimus. In some embodiments that may be combined with any of the preceding embodiments, the one or more doses of the GVHD prophylactic agent are + and tacrolimus in an amount that maintains a target blood level of at least about 3 ng / ml for at least about 20 days after administration of the third population of cells. + In some embodiments that may be combined with any of the preceding embodiments, Tregs are CD4 + CD25 + In some embodiments that may be combined with any of the preceding embodiments, the Treg-enriched cell population is CD45 + cells, and + In some embodiments that may be combined with any of the preceding embodiments, the Tregs are CD4 + CD25 + CD127dimor CD4 + FOXP3 + In some embodiments that may be combined with any of the preceding embodiments, HSPCs containing CD45 + The first population of cells is about 5 x 10 per kilogram of the human subject's actual or ideal body weight. 5 ~Approx. 2×10 7 In some embodiments that may be combined with any of the preceding embodiments, the Treg-enriched cell population comprises about 1 x 10 HSPCs per kilogram of the human subject's actual or ideal body weight. 5 ~Approx. 1×107 In some embodiments that may be combined with any of the preceding embodiments, the Tregs include CD45 + The third population of cells is about 1 x 10 per kilogram of the human subject's actual or ideal body weight. 5 ~Approx. 1×10 7 In some embodiments that may be combined with any of the preceding embodiments, + The first population of cells, and the Treg-enriched population of cells, expresses CD45 + In some embodiments that may be combined with any of the preceding embodiments, the first dose of the one or more doses of GVHD prophylactic agent is formulated for administration prior to administration of the third population of cells. + It is formulated for administration after administration of the third population of cells.

[0029] Another aspect of the present disclosure is (a) about 1.0 x 10 per kilogram of body weight of a human subject receiving the product. 5 ~Approx. 1.0×10 8 CD34 + Hematopoietic stem progenitor cells (HSPC), approximately 1.5×10 7 ~Approx. 1.5×10 10 HSPCs, or approximately 5.0 x 10 5 ~Approx. 5.0×10 8 A dose of isolated CD45 HSPCs containing + a first single-dose transfer bag containing a first population of cells, + (b) the first population of cells is formulated with an excipient at a neutral pH; and (b) about 1.0 x 10 cells per kilogram of body weight of a human subject receiving the product. 5 ~Approx. 2.0×10 7 Fresh CD4 + CD25 + CD127 dim Regulatory T cells (Treg), approximately 1.5 x 10 7 ~Approx. 3.0×10 9 fresh Tregs, or approximately 5.0 × 10 5 ~Approx. 1.0×10 8 A dose of fresh Tregs containing isolated CD45+ a second single-dose transfer bag containing a second population of cells, the isolated CD45 + the second population of cells is formulated with an excipient at a neutral pH; and (c) about 1.0 x 10 cells per kilogram of body weight of a human subject receiving the product. 5 ~Approx. 4.0×10 7 Normal CD3 + T cells (Tcon), approximately 1.5×10 7 ~Approx. 6.0×10 9 Tcon, or approximately 5.0 x 10 5 ~Approx. 2.0×10 8 Isolated CD45 containing a dose of Tcon + a third single-dose transfer bag containing a third population of cells, the third population comprising the isolated CD45 + The third population of cells relates to multi-component cell therapy products, including those formulated with excipients at neutral pH, where the excipients include one or more cryoprotectants.

[0030] Another aspect of the present disclosure relates to a method of treating a human subject diagnosed with a hematological malignancy, the method comprising administering to the human subject (a) an isolated CD45 + (b) a solution containing a first population of cells; (b) isolated CD45 T cells, including regulatory T cells (Tregs); + (c) a solution containing a second population of cells; (d) isolated CD45 + a solution containing a third population of cells, the solution comprising the isolated CD45 + The third population of cells is CD3 + and (d) a solution containing one or more doses of a graft-versus-host disease (GVHD) prophylactic agent, wherein the isolated CD45 + A first population of cells, a Treg-enriched cell population, and isolated CD45 +The third population of cells is obtained from a single allogeneic donor with at least one HLA mismatch to the human subject, and the non-relapse mortality rate of the human subject is reduced following administration, and / or the overall survival of the human subject is increased following administration, compared to a corresponding human subject who has undergone standard myeloablative allogeneic hematopoietic stem cell transplantation (alloHSCT) from a donor with at least one HLA mismatch to the corresponding human subject. Another aspect of the present disclosure relates to a multi-component medical procedure of the present disclosure for use in treating a human subject diagnosed with a hematological malignancy. Another aspect of the present disclosure relates to the use of a multi-component medical procedure of the present disclosure in the manufacture of a medicament for treating a human subject diagnosed with a hematological malignancy.

[0031] Another aspect of the present disclosure relates to a multi-component medical treatment administered to a human subject in need thereof, the multi-component treatment comprising: (a) isolated CD45 hematopoietic stem progenitor cells (HSPCs) and granulocytes; + a solution containing a first population of cells, the isolated CD45 + (b) isolated CD45 T cells containing regulatory T cells (Tregs), up to about 10% of the first population of cells. + (c) a solution containing a second population of cells; (d) isolated CD45 + a solution containing a third population of cells, the solution comprising the isolated CD45 + A third population of cells is at least about 20% CD3 + (d) a solution comprising one or more doses of a graft-versus-host disease (GVHD) prophylactic agent, wherein the isolated CD45 + A first population of cells, a Treg-enriched cell population, and isolated CD45 + The third population of cells has at least one HLA mismatch with the human subject and, when administered, produces isolated CD45 + A first population of cells, a Treg-enriched cell population, and isolated CD45 +The third population of cells reduces non-relapse mortality in the human subject and / or increases overall survival of the human subject compared to non-relapse mortality and / or overall survival in a matched human subject who has undergone standard myeloablative allogeneic hematopoietic stem cell transplantation (alloHSCT) and has at least one HLA mismatch with the matched human subject.

[0032] Another aspect of the present disclosure relates to a method of treating a human subject diagnosed with a hematological malignancy, the method comprising: (a) a myeloablative conditioning regimen comprising one or more doses of busulfan, fludarabine, and thiotepa; (b) administering to a subject an isolated CD45+ antibody containing hematopoietic stem and progenitor cells (HSPCs); + (c) a solution containing a first population of cells; (d) an isolated CD45 T cell population containing regulatory T cells (Tregs); + (d) a solution containing a second population of cells; (e) isolated CD45 + a solution containing a third population of cells, the solution comprising the isolated CD45 + The third population of cells is CD3 + and (e) a solution comprising one or more doses of a graft-versus-host disease (GVHD) prophylactic agent, wherein the pre-transplant treatment is administered about 2 to about 10 days prior to any of (b) through (e). Another aspect of the present disclosure relates to the multi-component medical treatment of any of the preceding embodiments for use in treating a human subject diagnosed with a hematological malignancy. Another aspect of the present disclosure relates to the use of the multi-component medical treatment of any of the preceding embodiments in the manufacture of a medicament for treating a human subject diagnosed with a hematological malignancy.

[0033] Another aspect of the present disclosure relates to a method of treating a human subject diagnosed with a hematological malignancy, the method comprising administering to the human subject: a) an isolated CD45 containing hematopoietic stem progenitor cell (HSPC) + a) a solution containing a first population of cells; b) isolated CD45 T cells containing regulatory T cells (Tregs); + a solution containing a second population of cells; c) isolated CD45 + a solution containing a third population of cells, the solution comprising the isolated CD45+ The third population of cells is CD3 + and a) a solution comprising one or more doses of a graft-versus-host disease (GVHD) prophylactic agent, wherein the human subject's GVHD- and relapse-free survival (GFRS) is increased by at least about 3.5-fold after administration, overall survival is increased by at least about 1.25-fold after administration, and / or non-relapse mortality is reduced by at least about 60% after administration, compared to a corresponding human subject who has undergone standard myeloablative allogeneic hematopoietic stem cell transplantation (alloHSCT). Another aspect of the present disclosure relates to the multicomponent medical treatment of any of the preceding embodiments for use in treating a human subject diagnosed with a hematological malignancy. Another aspect of the present disclosure relates to the use of the multicomponent medical treatment of any of the preceding embodiments in the manufacture of a medicament for treating a human subject diagnosed with a hematological malignancy.

[0034] Another aspect of the present disclosure relates to a multicomponent medical treatment administered to a human subject in need thereof, the multicomponent treatment comprising: a) a myeloablative conditioning regimen comprising one or more doses of busulfan, fludarabine, and thiotepa; b) an isolated CD45+-containing hematopoietic stem progenitor cell (HSPC) therapy; + a solution containing a first population of cells; c) isolated CD45 T cells containing regulatory T cells (Tregs); + a solution containing a second population of cells; d) isolated CD45 + a solution containing a third population of cells, the solution comprising the isolated CD45 + The third population of cells is CD3 + and e) a solution comprising one or more doses of a graft-versus-host disease (GVHD) prophylactic agent, wherein the transplant conditioning treatment is formulated to be administered about 2 to about 10 days before any of (b) to (e).

[0035] Another aspect of the present disclosure relates to a method of preparing a multi-component cell therapy product, the method comprising: a) obtaining an HSPC-mobilized peripheral blood apheresis from a donor allogeneic to a human subject receiving the product; b) combining a first apheresis sample with one or more cryoprotectants and cryopreserving the first apheresis sample, wherein the first apheresis sample contains about 1.0 x 10 to about 4.0 x 10 conventional CD3 per kilogram of body weight of the human subject receiving the product. + c) sorting the second apheresis sample using one or more immunoseparation particles (ISPs) specific for CD34 to obtain a CD34-enriched cell population and a CD34-depleted cell population, the CD34-enriched cell population comprising about 1.0 x 10 to about 1.0 x 10 CD34 cells per kilogram of body weight of the human subject receiving the product; + hematopoietic stem progenitor cells (HSPCs), about 5.0 x 10 to about 5.0 x 10 HSPCs, or about 1.5 x 10 to about 1.5 x 10 HSPCs; d) sorting the CD34-depleted cell population with one or more ISPs specific for CD25 to obtain a CD25-enriched cell population and a CD25-depleted cell population; and e) sorting the CD25-enriched cell population with one or more ISPs specific for CD4 and one or more ISPs specific for CD127, and obtaining a CD4 + and selecting a population of CD127dim cells, and + and CD127 dim The cell population is approximately 1.0 x 105 to approximately 2.0 x 107 CD4 cells per kilogram of body weight of the human subject receiving the product. + CD25 + CD127 dimRegulatory T cells (Tregs), about 5.0 x 10 to about 1.0 x 10 isolated fresh Tregs, or about 1.5 x 10 to about 3.0 x 10 isolated fresh Tregs, wherein the multi-component cell therapy product comprises Tcon, HSPCs, and Tregs.

[0036] Another aspect of the present disclosure relates to a method of preparing a multi-component cell therapy product, the method comprising: a) obtaining an HSPC-mobilized peripheral blood apheresis from a donor allogeneic to a human subject receiving the product; b) sorting the first apheresis sample with one or more immunoseparation particles (ISPs) specific for CD3 to obtain a CD3-enriched cell population and a CD3-depleted cell population, the CD3-enriched cell population comprising between about 1.0 x 10 and about 4.0 x 10 conventional CD3 per kilogram of body weight of the human subject receiving the product. + and c) sorting the second apheresis sample using one or more ISPs specific for CD34 to obtain a CD34-enriched cell population and a CD34-depleted cell population, the CD34-enriched cell population comprising about 1.0 x 10 to about 1.0 x 10 CD34 cells per kilogram of body weight of the human subject receiving the product. + hematopoietic stem progenitor cells (HSPCs), about 1.5 x 10 to about 1.5 x 10 HSPCs, or about 5.0 x 10 to about 5.0 x 10 HSPCs; d) sorting the CD34-depleted cell population with one or more ISPs specific for CD25 to obtain a CD25-enriched cell population and a CD25-depleted cell population; and e) sorting the CD25-enriched cell population with one or more ISPs specific for CD4 and one or more ISPs specific for CD127, and obtaining a CD4 + and CD127 dim and selecting a cell population of CD4 + and CD127 dimThe cell population is approximately 1.0 x 105 to approximately 2.0 x 107 CD4 cells per kilogram of body weight of the human subject receiving the product. + CD25 + CD127 dim Regulatory T cells (Tregs), about 1.5 x 10 to about 3.0 x 10 Tregs, or about 5.0 x 10 to about 1.0 x 10 Tregs, wherein the multicomponent cell therapy product comprises Tcon, HSPCs, and Tregs.

[0037] Another aspect of the present disclosure relates to a method of preparing a multi-component cell therapy product, the method comprising: a) obtaining an HSPC-mobilized peripheral blood apheresis from a donor allogeneic to a human subject receiving the product; b) obtaining from a first apheresis sample about 1.0 x 10 to about 4.0 x 10 conventional CD3 per kilogram of body weight of the human subject receiving the product; + and cryopreserving a cell population comprising T cells (Tcon), about 1.5 x 10 to about 6.0 x 10 Tcon, or about 5.0 x 10 to about 2.0 x 10 Tcon; c) sorting the second apheresis sample with one or more immunoseparation particles (ISPs) specific for CD25 to obtain a CD25-enriched cell population and a CD25-depleted cell population, and sorting the CD25-depleted population with one or more ISPs specific for CD34 to obtain a CD34-enriched cell population and a CD34-depleted cell population, wherein the CD34-enriched cell population comprises about 1.0 x 10 to about 1.0 x 10 CD34 per kilogram of body weight of the human subject receiving the product. + hematopoietic stem progenitor cells (HSPCs), comprising about 1.5 x 10 to about 1.5 x 10 HSPCs, or about 5.0 x 10 to about 5.0 x 10 HSPCs; d) sorting the CD25-enriched cell population using one or more ISPs specific for CD4 and one or more ISPs specific for CD127, and + and CD127 dim and selecting a cell population of CD4 + and CD127 dimThe cell population is approximately 1.0 x 105 to approximately 2.0 x 107 CD4 cells per kilogram of body weight of the human subject receiving the product. + CD25 + CD127 dim Regulatory T cells (Tregs), about 1.5 x 10 to about 3.0 x 10 Tregs, or about 5.0 x 10 to about 1.0 x 10 Tregs, wherein the multicomponent cell therapy product comprises Tcon, HSPCs, and Tregs.

[0038] Another aspect of the present disclosure relates to a method of preparing a multi-component cell therapy product, the method comprising: a) obtaining an HSPC-mobilized peripheral blood apheresis from a donor allogeneic to a human subject receiving the product; b) combining a first apheresis sample with one or more cryoprotectants and cryopreserving the first apheresis sample, wherein the first apheresis sample contains between about 1.0 x 10 and about 4.0 x 10 conventional CD3 per kilogram of body weight of the human subject receiving the product. + c) sorting the second apheresis sample with one or more immunoseparation particles (ISPs) specific for CD25 to obtain a CD25-enriched cell population and a CD25-depleted cell population, and sorting the CD25-depleted population with one or more ISPs specific for CD34 to obtain a CD34-enriched cell population and a CD34-depleted cell population, wherein the CD34-enriched cell population comprises about 1.0 x 10 to about 1.0 x 10 CD34 per kilogram of body weight of the human subject receiving the product. + hematopoietic stem progenitor cells (HSPCs), comprising about 1.5 x 10 to about 1.5 x 10 HSPCs, or about 5.0 x 10 to about 5.0 x 10 HSPCs; d) sorting the CD25-enriched cell population using one or more ISPs specific for CD4 and one or more ISPs specific for CD127, and + and CD127 dimand selecting a cell population of CD4 + and CD127 dim The cell population is approximately 1.0 x 105 to approximately 2.0 x 107 CD4 cells per kilogram of body weight of the human subject receiving the product. + CD25 + CD127 dim Regulatory T cells (Tregs), about 1.5 x 10 to about 3.0 x 10 Tregs, or about 5.0 x 10 to about 1.0 x 10 Tregs, wherein the multicomponent cell therapy product comprises Tcon, HSPCs, and Tregs.

[0039] Another aspect of the present disclosure relates to a method of preparing a multi-component cell therapy product, the method comprising: a) obtaining an HSPC-mobilized peripheral blood apheresis from a donor allogeneic to a human subject receiving the product; b) sorting the first apheresis sample with one or more immunoseparation particles (ISPs) specific for CD3 to obtain a CD3-enriched cell population and a CD3-depleted cell population, the CD3-enriched cell population comprising between about 1.0 x 10 and about 4.0 x 10 conventional CD3 per kilogram of body weight of the human subject receiving the product. + and c) sorting the second apheresis sample using one or more ISPs specific for CD25 to obtain a CD25-enriched cell population and a CD25-depleted cell population, and sorting the CD25-depleted population using one or more ISPs specific for CD34 to obtain a CD34-enriched cell population and a CD34-depleted cell population, wherein the CD34-enriched cell population comprises about 1.0 x 10 to about 1.0 x 10 CD34 cells per kilogram of body weight of the human subject receiving the product. +hematopoietic stem progenitor cells (HSPCs), comprising about 1.5 x 10 to about 1.5 x 10 HSPCs, or about 5.0 x 10 to about 5.0 x 10 HSPCs; d) sorting the CD25-enriched cell population using one or more ISPs specific for CD4 and one or more ISPs specific for CD127, and + and CD127 dim and selecting a cell population of CD4 + and CD127 dim The cell population is approximately 1.0 x 105 to approximately 2.0 x 107 CD4 cells per kilogram of body weight of the human subject receiving the product. + CD25 + CD127 dim Regulatory T cells (Tregs), about 1.5 x 10 to about 3.0 x 10 Tregs, or about 5.0 x 10 to about 1.0 x 10 Tregs, wherein the multicomponent cell therapy product comprises Tcon, HSPCs, and Tregs.

[0040] Another aspect of the present disclosure relates to a method of preparing a multi-component cell therapy product, the method comprising: a) obtaining an HSPC-mobilized peripheral blood apheresis from a donor allogeneic to a human subject receiving the product; b) cryopreserving a first apheresis sample, the first apheresis sample containing about 1.0 x 10 to about 4.0 x 10 conventional CD3 per kilogram of body weight of the human subject receiving the product. + T cells (Tcon), about 1.5 x 10 to about 6.0 x 10 Tcon, or about 5.0 x 10 to about 2.0 x 10 Tcon; c) about 1.0 x 10 to about 1.0 x 10 CD34 per kilogram of body weight of a human subject receiving the product. + and d) selecting from the second apheresis sample a CD34-enriched cell population comprising hematopoietic stem progenitor cells (HSPCs), about 1.5 x 10 to about 1.5 x 10 HSPCs, or about 5.0 x 10 to about 5.0 x 10 HSPCs; and +and selecting a CD25-enriched cell population from the second apheresis sample, wherein the cell population is CD4 + and CD127 dim The cell population is approximately 1.0 x 105 to approximately 2.0 x 107 CD4 cells per kilogram of body weight of the human subject receiving the product. + CD25 + CD127 dim Regulatory T cells (Tregs), about 1.5 x 10 to about 3.0 x 10 Tregs, or about 5.0 x 10 to about 1.0 x 10 Tregs, wherein the multicomponent cell therapy product comprises Tcon, HSPCs, and Tregs.

[0041] Another aspect of the present disclosure relates to a method of preparing a multi-component cell therapy product, the method comprising: a) obtaining an HSPC-mobilized peripheral blood apheresis sample from a donor allogeneic to a human subject receiving the product; and b) obtaining from the apheresis sample about 1.0 x 10 to about 1.0 x 10 conventional CD34 cells per kilogram of body weight of the human subject receiving the product. + A first sample containing hematopoietic stem progenitor cells (HSPCs), about 1.5 x 10 to about 1.5 x 10 HSPCs, about 5.0 x 10 to about 5.0 x 10 HSPCs, and about 1.0 x 10 to about 2.0 x 10 CD4 per kilogram of body weight of a human subject receiving the product. + CD25 + CD127 dim a second sample containing regulatory T cells (Tregs), about 1.5 x 10 to about 3.0 x 10 Tregs, or about 5.0 x 10 to about 1.0 x 10 Tregs, and about 1.0 x 10 to about 4.0 x 10 conventional CD3 per kilogram of body weight of a human subject receiving the product. + and selecting a third sample comprising T cells (Tcon), about 1.5 x 10 to about 6.0 x 10 Tcon, or about 5.0 x 10 to about 2.0 x 10 Tcon, wherein the multicomponent cell therapy product comprises Tcon, HSPCs, and Tregs.

[0042] Another aspect of the present disclosure relates to a multi-component cell therapy product or multi-component medical treatment made or prepared by the method of any of the preceding embodiments.

[0043] Another aspect of the present disclosure relates to a method of treating a human subject having or suspected of having acute leukemia in complete remission, the method comprising administering to the human subject: a) a CD34 + Isolated CD45 containing hematopoietic stem progenitor cells (HSPCs) + a) a solution containing the first population of cells; b) fresh CD4 + CD25 + CD127 dim Isolated CD45 containing regulatory T cells (Tregs) + a solution containing a second population of cells; and c) conventional CD3 + Isolated CD45 containing T cells (Tcon) + and administering a multicomponent cell therapy comprising a solution comprising a third population of cells. In some embodiments, the multicomponent cell therapy is a multicomponent cell therapy product of the present disclosure or a multicomponent medical procedure of the present disclosure. Another aspect of the present disclosure relates to a multicomponent cell therapy of the present disclosure for use in treating a human subject having or suspected of having acute leukemia in complete remission. Another aspect of the present disclosure relates to the use of a multicomponent cell therapy of the present disclosure in the manufacture of a medicament for treating a human subject having or suspected of having acute leukemia in complete remission.

[0044] Another aspect of the present disclosure relates to a method of treating a human subject having or suspected of having active leukemia, the method comprising administering to the human subject: a) a CD34 + Isolated CD45 containing hematopoietic stem progenitor cells (HSPCs) + a) a solution containing the first population of cells; b) fresh CD4 + CD25 + CD127 dim Isolated CD45 containing regulatory T cells (Tregs) + a solution containing a second population of cells; and c) conventional CD3 + Isolated CD45 containing T cells (Tcon)+ and a solution comprising a third population of cells. In some embodiments, the multicomponent cell therapy is a multicomponent cell therapy product of the present disclosure or a multicomponent medical procedure of the present disclosure. Another aspect of the present disclosure relates to a multicomponent cell therapy of the present disclosure for use in treating a human subject having or suspected of having active leukemia. Another aspect of the present disclosure relates to the use of a multicomponent cell therapy of the present disclosure in the manufacture of a medicament for treating a human subject having or suspected of having active leukemia.

[0045] Another aspect of the present disclosure relates to a method of treating a human subject having or suspected of having acute leukemia refractory to first-line therapy or acute leukemia with minimal residual disease, the method comprising administering to the human subject: a) a CD34 + Isolated CD45 containing hematopoietic stem progenitor cells (HSPCs) + a) a solution containing the first population of cells; b) fresh CD4 + CD25 + CD127 dim Isolated CD45 containing regulatory T cells (Tregs) + a solution containing a second population of cells; and c) conventional CD3 + Isolated CD45 containing T cells (Tcon) + a solution comprising a third population of cells. In some embodiments, the multicomponent cell therapy is a multicomponent cell therapy product of the present disclosure or a multicomponent medical procedure of the present disclosure. Another aspect of the present disclosure relates to a multicomponent cell therapy of the present disclosure for use in treating a human subject having or suspected of having acute leukemia refractory to first-line therapy or acute leukemia with minimal residual disease. Another aspect of the present disclosure relates to use of a multicomponent cell therapy of the present disclosure in the manufacture of a medicament for treating a human subject having or suspected of having acute leukemia refractory to first-line therapy or acute leukemia with minimal residual disease.

[0046] Another aspect of the present disclosure relates to a method of treating a human subject having or suspected of having low-risk acute myeloid leukemia (AML), the method comprising administering to the human subject: a) administering to the human subject a CD34 + Isolated CD45 containing hematopoietic stem progenitor cells (HSPCs) + a) a solution containing the first population of cells; b) fresh CD4 + CD25 + CD127 dim Isolated CD45 containing regulatory T cells (Tregs) + a solution containing a second population of cells; and c) conventional CD3 + Isolated CD45 containing T cells (Tcon) + and a solution comprising a third population of cells. In some embodiments, the multicomponent cell therapy is a multicomponent cell therapy product of the present disclosure or a multicomponent medical procedure of the present disclosure. Another aspect of the present disclosure relates to a multicomponent cell therapy of the present disclosure for use in treating a human subject having or suspected of having low-risk acute myeloid leukemia (AML). Another aspect of the present disclosure relates to the use of a multicomponent cell therapy of the present disclosure in the manufacture of a medicament for treating a human subject having or suspected of having low-risk acute myeloid leukemia (AML).

[0047] Another aspect of the present disclosure relates to a method of treating a human subject having or suspected of having high-risk acute myeloid leukemia (AML) in complete remission, the method comprising administering to the human subject: a) a CD34 + Isolated CD45 containing hematopoietic stem progenitor cells (HSPCs) + a) a solution containing the first population of cells; b) fresh CD4 + CD25 + CD127 dim Isolated CD45 containing regulatory T cells (Tregs) + a solution containing a second population of cells; and c) conventional CD3 + Isolated CD45 containing T cells (Tcon) +and administering a multicomponent cell therapy comprising a solution comprising a third population of cells. In some embodiments, the multicomponent cell therapy is a multicomponent cell therapy product of the present disclosure or a multicomponent medical procedure of the present disclosure. Another aspect of the present disclosure relates to a multicomponent cell therapy of the present disclosure for use in treating a human subject having or suspected of having high-risk acute myeloid leukemia (AML) who is in complete remission. Another aspect of the present disclosure relates to the use of a multicomponent cell therapy of the present disclosure in the manufacture of a medicament for treating a human subject having or suspected of having high-risk acute myeloid leukemia (AML) who is in complete remission.

[0048] Another aspect of the present disclosure relates to a method of treating a human subject having or suspected of having chronic myeloid leukemia (CML), the method comprising administering to the human subject: a) administering to the human subject a CD34 + Isolated CD45 containing hematopoietic stem progenitor cells (HSPCs) + a) a solution containing the first population of cells; b) fresh CD4 + CD25 + CD127 dim Isolated CD45 containing regulatory T cells (Tregs) + a solution containing a second population of cells; and c) conventional CD3 + Isolated CD45 containing T cells (Tcon) + and a solution comprising a third population of cells. In some embodiments, the multicomponent cell therapy is a multicomponent cell therapy product of the present disclosure or a multicomponent medical procedure of the present disclosure. Another aspect of the present disclosure relates to a multicomponent cell therapy of the present disclosure for use in treating a human subject having or suspected of having chronic myeloid leukemia (CML). Another aspect of the present disclosure relates to the use of a multicomponent cell therapy of the present disclosure in the manufacture of a medicament for treating a human subject having or suspected of having chronic myeloid leukemia (CML).

[0049] Another aspect of the present disclosure relates to a method of treating a human subject having or suspected of having a high-risk myelodysplastic syndrome, the method comprising administering to the human subject: a) a CD34 +Isolated CD45 containing hematopoietic stem progenitor cells (HSPCs) + a) a solution containing the first population of cells; b) fresh CD4 + CD25 + CD127 dim Isolated CD45 containing regulatory T cells (Tregs) + a solution containing a second population of cells; and c) conventional CD3 + Isolated CD45 containing T cells (Tcon) + and administering a multicomponent cell therapy comprising a solution comprising a third population of cells. In some embodiments, the multicomponent cell therapy is a multicomponent cell therapy product of the present disclosure or a multicomponent medical procedure of the present disclosure. Another aspect of the present disclosure relates to a multicomponent cell therapy of the present disclosure for use in treating a human subject having or suspected of having a high-risk myelodysplastic syndrome. Another aspect of the present disclosure relates to the use of a multicomponent cell therapy of the present disclosure in the manufacture of a medicament for treating a human subject having or suspected of having a high-risk myelodysplastic syndrome.

[0050] Another aspect of the present disclosure relates to a method of treating a human subject having or suspected of having a therapy-related myelodysplastic syndrome and / or a secondary myelodysplastic syndrome, the method comprising administering to the human subject: a) a CD34 + Isolated CD45 containing hematopoietic stem progenitor cells (HSPCs) + a) a solution containing the first population of cells; b) fresh CD4 + CD25 + CD127 dim Isolated CD45 containing regulatory T cells (Tregs) + a solution containing a second population of cells; and c) conventional CD3 + Isolated CD45 containing T cells (Tcon) +and administering a multicomponent cell therapy comprising a solution comprising a third population of cells. In some embodiments, the multicomponent cell therapy is a multicomponent cell therapy product of the present disclosure or a multicomponent medical procedure of the present disclosure. Another aspect of the present disclosure relates to a multicomponent cell therapy of the present disclosure for use in treating a human subject having or suspected of having a therapy-related myelodysplastic syndrome and / or a secondary myelodysplastic syndrome. Another aspect of the present disclosure relates to the use of a multicomponent cell therapy of the present disclosure in the manufacture of a medicament for the treatment of a human subject having or suspected of having a therapy-related myelodysplastic syndrome and / or a secondary myelodysplastic syndrome.

[0051] Another aspect of the present disclosure relates to a method of treating a human subject having or suspected of having a myeloproliferative syndrome, the method comprising administering to the human subject: a) CD34 + Isolated CD45 containing hematopoietic stem progenitor cells (HSPCs) + a) a solution containing the first population of cells; b) fresh CD4 + CD25 + CD127 dim Isolated CD45 containing regulatory T cells (Tregs) + a solution containing a second population of cells; and c) conventional CD3 + Isolated CD45 containing T cells (Tcon) + and a solution comprising a third population of cells. In some embodiments, the multicomponent cell therapy is a multicomponent cell therapy product of the present disclosure or a multicomponent medical procedure of the present disclosure. Another aspect of the present disclosure relates to a multicomponent cell therapy of the present disclosure for use in treating a human subject having or suspected of having a myeloproliferative syndrome. Another aspect of the present disclosure relates to the use of a multicomponent cell therapy of the present disclosure in the manufacture of a medicament for treating a human subject having or suspected of having a myeloproliferative syndrome.

[0052] Another aspect of the present disclosure relates to a method of treating a human subject having or suspected of having a poor prognosis non-Hodgkin's lymphoma that is not suitable for autologous hematopoietic cell transplantation (HCT), the method comprising administering to the human subject: a) a CD34+ Isolated CD45 containing hematopoietic stem progenitor cells (HSPCs) + a) a solution containing the first population of cells; b) fresh CD4 + CD25 + CD127 dim Isolated CD45 containing regulatory T cells (Tregs) + a solution containing a second population of cells; and c) conventional CD3 + Isolated CD45 containing T cells (Tcon) + and a solution comprising a third population of cells. In some embodiments, the multicomponent cell therapy is a multicomponent cell therapy product of the present disclosure or a multicomponent medical procedure of the present disclosure. Another aspect of the present disclosure relates to a multicomponent cell therapy of the present disclosure for use in treating a human subject having or suspected of having a non-Hodgkin's lymphoma with a poor prognosis who is not suitable for autologous hematopoietic cell transplantation (HCT). Another aspect of the present disclosure relates to the use of a multicomponent cell therapy of the present disclosure in the manufacture of a medicament for the treatment of a human subject having or suspected of having a non-Hodgkin's lymphoma with a poor prognosis who is not suitable for autologous hematopoietic cell transplantation (HCT).

[0053] Another aspect of the present disclosure relates to a method of treating a human subject having or suspected of having multiple sclerosis, the method comprising administering to the human subject: a) a CD34 + Isolated CD45 containing hematopoietic stem progenitor cells (HSPCs) + a) a solution containing the first population of cells; b) fresh CD4 + CD25 + CD127 dim Isolated CD45 containing regulatory T cells (Tregs) + a solution containing a second population of cells; and c) conventional CD3 + Isolated CD45 containing T cells (Tcon) +and a solution comprising a third population of cells. In some embodiments, the multicomponent cell therapy is a multicomponent cell therapy product of the present disclosure or a multicomponent medical procedure of the present disclosure. Another aspect of the present disclosure relates to a multicomponent cell therapy of the present disclosure for use in treating a human subject having or suspected of having multiple sclerosis. Another aspect of the present disclosure relates to the use of a multicomponent cell therapy of the present disclosure in the manufacture of a medicament for treating a human subject having or suspected of having multiple sclerosis.

[0054] Another aspect of the present disclosure relates to a method of treating a human subject having or suspected of having a hematological malignancy with a multi-component cellular therapy, the method comprising: a) administering to a subject a CD34 + Isolated CD45 containing hematopoietic stem progenitor cells (HSPCs) + a) administering a first population of cells; b) administering an isolated CD45 + After administration of the first population of cells, isolated fresh CD4 + CD25 + CD127 dim Isolated CD45 containing regulatory T cells (Tregs) + and c) administering a second population of isolated CD45 cells. + About 12 hours to about 120 hours after administration of the second population of cells, conventional CD3 + Isolated CD45 containing T cells (Tcon) + and administering a third population of cells, wherein the multicomponent cell therapy comprises administering an isolated CD45 + First population of cells, isolated CD45 + A second population of cells, and isolated CD45 + and a third population of cells. Another aspect of the present disclosure relates to a multi-component cell therapy of the present disclosure for use in treating a human subject having or suspected of having a hematological malignancy. Another aspect of the present disclosure relates to the use of a multi-component cell therapy of the present disclosure in the manufacture of a medicament for the treatment of a human subject having or suspected of having a hematological malignancy.

[0055] Another aspect of the present disclosure is (a) about 1.0 x 10 per kilogram of body weight of a human subject receiving the product. 5 ~Approx. 1.0×10 8 CD34 + Hematopoietic stem progenitor cells (HSPC), approximately 1.5×10 7 ~Approx. 1.5×10 10 HSPCs, or approximately 5.0 x 10 5 ~Approx. 5.0×10 8 A dose of isolated CD45 HSPCs containing + a first single-dose transfer bag containing a first population of cells, + (b) the first population of cells is formulated with an excipient at a neutral pH; and (b) about 1.0 x 10 cells per kilogram of body weight of a human subject receiving the product. 5 ~Approx. 2.0×10 7 Fresh CD4 + CD25 + CD127 dim Regulatory T cells (Treg), approximately 1.5 x 10 7 ~Approx. 3.0×10 9 freshly isolated Tregs, or approximately 5.0 x 10 5 ~Approx. 1.0×10 8 1 dose of isolated fresh Tregs containing isolated CD45 + a second single-dose transfer bag containing a second population of cells, the isolated CD45 + the second population of cells is formulated with an excipient at a neutral pH, and optionally the first single-dose transfer bag and the second single-dose transfer bag are the same transfer bag; and (c) about 1.0 x 10 cells per kilogram of body weight of the human subject receiving the product. 5 ~Approx. 4.0×10 7 Normal CD3 + T cells (Tcon), approximately 1.5×10 7 ~Approx. 6.0×10 9 Tcon, or approximately 5.0 x 10 5 ~Approx. 2.0×10 8 Isolated CD45 containing a dose of Tcon +a third single-dose transfer bag containing a third population of cells, the third population comprising the isolated CD45 + The third population of cells relates to multi-component cell therapy products, including those formulated with excipients at neutral pH, where the excipients include one or more cryoprotectants. In some embodiments that may be combined with any of the preceding embodiments, a) isolated CD45 + The first population of cells is about 1.0 x 10 per kilogram of body weight of the human subject receiving the product. 5 HSPCs, approximately 2.0 x 10 per kilogram of body weight of the human subject receiving the product 5 HSPCs, approximately 3.0 x 10 per kilogram of body weight of the human subject receiving the product 5 HSPCs, approximately 4.0 x 10 per kilogram of body weight of the human subject receiving the product 5 or more than about 5.0 x 10 HSPCs per kilogram of body weight of the human subject receiving the product 5 HSPCs or more, approximately 6.0 x 10 per kilogram of body weight of the human subject receiving the product 5 7.0 x 10 HSPCs or more per kilogram of body weight of the human subject receiving the product 5 HSPCs, approximately 8.0 x 10 per kilogram of body weight of the human subject receiving the product 5 HSPCs or more, approximately 9.0 x 10 per kilogram of body weight of the human subject receiving the product 5 or more than about 1.0 x 10 HSPCs per kilogram of body weight of the human subject receiving the product 6 or more than about 1.5 x 10 HSPCs per kilogram of body weight of the human subject receiving the product 6 HSPCs, approximately 2.0 x 10 per kilogram of body weight of the human subject receiving the product 6 HSPCs, approximately 2.5 x 10 per kilogram of body weight of the human subject receiving the product 6 HSPCs, approximately 3.0 x 10 per kilogram of body weight of the human subject receiving the product 6 or more than about 3.5 x 10 HSPCs per kilogram of body weight of the human subject receiving the product6 HSPCs, approximately 4.0 x 10 per kilogram of body weight of the human subject receiving the product 6 or more than about 4.5 x 10 HSPCs per kilogram of body weight of the human subject receiving the product 6 or more than about 5.0 x 10 HSPCs per kilogram of body weight of the human subject receiving the product 6 or more than about 5.5 x 10 HSPCs per kilogram of body weight of the human subject receiving the product 6 HSPCs or more, approximately 6.0 x 10 per kilogram of body weight of the human subject receiving the product 6 HSPCs or more, approximately 6.5 x 10 per kilogram of body weight of the human subject receiving the product 6 7.0 x 10 HSPCs or more per kilogram of body weight of the human subject receiving the product 6 7.5 x 10 HSPCs or more per kilogram of body weight of the human subject receiving the product 6 HSPCs, approximately 8.0 x 10 per kilogram of body weight of the human subject receiving the product 6 or more than approximately 8.5 x 10 HSPCs per kilogram of body weight of the human subject receiving the product 6 HSPCs or more, approximately 9.0 x 10 per kilogram of body weight of the human subject receiving the product 6 or more than about 9.5 x 10 HSPCs per kilogram of body weight of the human subject receiving the product 6 or more than about 1.0 x 10 HSPCs per kilogram of body weight of the human subject receiving the product 7 or more than about 1.5 x 10 HSPCs per kilogram of body weight of the human subject receiving the product 7 HSPCs, approximately 2.0 x 10 per kilogram of body weight of the human subject receiving the product 7 HSPCs, approximately 2.5 x 10 per kilogram of body weight of the human subject receiving the product 7 HSPCs, approximately 3.0 x 10 per kilogram of body weight of the human subject receiving the product 7 or more than about 3.5 x 10 HSPCs per kilogram of body weight of the human subject receiving the product 7HSPCs, approximately 4.0 x 10 per kilogram of body weight of the human subject receiving the product 7 or more than about 4.5 x 10 HSPCs per kilogram of body weight of the human subject receiving the product 7 or more than about 5.0 x 10 HSPCs per kilogram of body weight of the human subject receiving the product 7 or more than about 5.5 x 10 HSPCs per kilogram of body weight of the human subject receiving the product 7 HSPCs or more, approximately 6.0 x 10 per kilogram of body weight of the human subject receiving the product 7 HSPCs or more, approximately 6.5 x 10 per kilogram of body weight of the human subject receiving the product 7 7.0 x 10 HSPCs or more per kilogram of body weight of the human subject receiving the product 7 7.5 x 10 HSPCs or more per kilogram of body weight of the human subject receiving the product 7 HSPCs, approximately 8.0 x 10 per kilogram of body weight of the human subject receiving the product 7 or more than approximately 8.5 x 10 HSPCs per kilogram of body weight of the human subject receiving the product 7 HSPCs or more, approximately 9.0 x 10 per kilogram of body weight of the human subject receiving the product 7 or more than about 9.5 x 10 HSPCs per kilogram of body weight of the human subject receiving the product 7 HSPCs or approximately 1.0 x 10 per kilogram of body weight of the human subject receiving the product 8 and / or b) a dose of isolated CD45 HSPCs. + The first population of cells was approximately 5.0 x 10 5 More than 6.0 x 10 HSPCs 5 More than 7.0 x 10 HSPCs 5 More than HSPCs, approximately 8.0 x 10 5 More than 9.0 x 10 HSPCs 5 More than 1.0 x 10 HSPCs 6 More than 1.5 x 10 HSPCs 6 More than HSPCs, approximately 2.0 x 10 6 More than HSPCs, approximately 2.5 x 10 6More than 3.0 x 10 HSPCs 6 More than HSPCs, approximately 3.5 x 10 6 More than HSPCs, approximately 4.0 x 10 6 More than HSPCs, approximately 4.5 x 10 6 More than 5.0 x 10 HSPCs 6 More than 5.5 x 10 HSPCs 6 More than 6.0 x 10 HSPCs 6 More than 6.5 x 10 HSPCs 6 More than 7.0 x 10 HSPCs 6 More than 7.5 x 10 HSPCs 6 More than HSPCs, approximately 8.0 x 10 6 More than HSPCs, approximately 8.5 x 10 6 More than 9.0 x 10 HSPCs 6 More than 9.5 x 10 HSPCs 6 More than 1.0 x 10 HSPCs 7 More than 1.5 x 10 HSPCs 7 More than HSPCs, approximately 2.0 x 10 7 More than HSPCs, approximately 2.5 x 10 7 More than 3.0 x 10 HSPCs 7 More than HSPCs, approximately 3.5 x 10 7 More than HSPCs, approximately 4.0 x 10 7 More than HSPCs, approximately 4.5 x 10 7 More than 5.0 x 10 HSPCs 7 More than 5.5 x 10 HSPCs 7 More than 6.0 x 10 HSPCs 7 More than 6.5 x 10 HSPCs 7 More than 7.0 x 10 HSPCs 7 More than 7.5 x 10 HSPCs 7 More than HSPCs, approximately 8.0 x 10 7 More than HSPCs, approximately 8.5 x 10 7 More than 9.0 x 10 HSPCs 7 More than 9.5 x 10 HSPCs 7 More than 1.0 x 10 HSPCs 8 More than 1.5 x 10 HSPCs 8More than HSPCs, approximately 2.0 x 10 8 More than HSPCs, approximately 2.5 x 10 8 More than 3.0 x 10 HSPCs 8 More than HSPCs, approximately 3.5 x 10 8 More than HSPCs, approximately 4.0 x 10 8 More than HSPCs, approximately 4.5 x 10 8 More than 5.0 x 10 HSPCs 8 More than 5.5 x 10 HSPCs 8 More than 6.0 x 10 HSPCs 8 More than 6.5 x 10 HSPCs 8 More than 7.0 x 10 HSPCs 8 More than 7.5 x 10 HSPCs 8 More than HSPCs, approximately 8.0 x 10 8 More than HSPCs, approximately 8.5 x 10 8 More than 9.0 x 10 HSPCs 8 More than 9.5 x 10 HSPCs 8 More than 1.0 x 10 HSPCs 9 More than 1.5 x 10 HSPCs 9 More than HSPCs, approximately 2.0 x 10 9 More than HSPCs, approximately 2.5 x 10 9 More than 3.0 x 10 HSPCs 9 More than HSPCs, approximately 3.5 x 10 9 More than HSPCs, approximately 4.0 x 10 9 More than HSPCs, approximately 4.5 x 10 9 More than 5.0 x 10 HSPCs 9 More than 5.5 x 10 HSPCs 9 More than 6.0 x 10 HSPCs 9 More than 6.5 x 10 HSPCs 9 More than 7.0 x 10 HSPCs 9 More than 7.5 x 10 HSPCs 9 More than HSPCs, approximately 8.0 x 10 9 More than HSPCs, approximately 8.5 x 10 9 More than 9.0 x 10 HSPCs 9 More than 9.5 x 10 HSPCs 9More than 1.0 x 10 HSPCs 10 HSPCs or approximately 1.5 × 10 10 and / or c) a dose of HSPCs containing isolated CD45 + The second population of cells is about 1.0 x 10 per kilogram of body weight of the human subject receiving the product. 5 or more isolated fresh Tregs, approximately 2.0 x 10 per kilogram of body weight of the human subject receiving the product. 5 or more isolated fresh Tregs, approximately 3.0 x 10 per kilogram of body weight of the human subject receiving the product. 5 or more isolated fresh Tregs, approximately 4.0 x 10 per kilogram of body weight of the human subject receiving the product. 5 or more isolated fresh Tregs, approximately 5.0 x 10 per kilogram of body weight of the human subject receiving the product. 5 or more isolated fresh Tregs, approximately 6.0 x 10 per kilogram of body weight of the human subject receiving the product. 5 or more isolated fresh Tregs, approximately 7.0 x 10 per kilogram of body weight of the human subject receiving the product. 5 or more isolated fresh Tregs, approximately 8.0 x 10 per kilogram of body weight of the human subject receiving the product. 5 or more isolated fresh Tregs, approximately 9.0 x 10 per kilogram of body weight of the human subject receiving the product. 5 or more isolated fresh Tregs, approximately 1.0 x 10 per kilogram of body weight of the human subject receiving the product. 6 or more isolated fresh Tregs, approximately 1.5 x 10 per kilogram of body weight of the human subject receiving the product. 6 or more isolated fresh Tregs, approximately 2.0 x 10 per kilogram of body weight of the human subject receiving the product. 6 or more isolated fresh Tregs, approximately 2.5 x 10 per kilogram of body weight of the human subject receiving the product. 6 or more isolated fresh Tregs, approximately 3.0 x 10 per kilogram of body weight of the human subject receiving the product. 6or more isolated fresh Tregs, approximately 3.5 x 10 per kilogram of body weight of the human subject receiving the product. 6 or more isolated fresh Tregs, approximately 4.0 x 10 per kilogram of body weight of the human subject receiving the product. 6 or more isolated fresh Tregs, approximately 4.5 x 10 per kilogram of body weight of the human subject receiving the product. 6 or more isolated fresh Tregs, approximately 5.0 x 10 per kilogram of body weight of the human subject receiving the product. 6 or more isolated fresh Tregs, approximately 5.5 x 10 per kilogram of body weight of the human subject receiving the product. 6 or more isolated fresh Tregs, approximately 6.0 x 10 per kilogram of body weight of the human subject receiving the product. 6 or more isolated fresh Tregs, approximately 6.5 x 10 per kilogram of body weight of the human subject receiving the product. 6 or more isolated fresh Tregs, approximately 7.0 x 10 per kilogram of body weight of the human subject receiving the product. 6 or more isolated fresh Tregs, approximately 7.5 x 10 per kilogram of body weight of the human subject receiving the product. 6 or more isolated fresh Tregs, approximately 8.0 x 10 per kilogram of body weight of the human subject receiving the product. 6 or more isolated fresh Tregs, approximately 8.5 x 10 per kilogram of body weight of the human subject receiving the product. 6 or more isolated fresh Tregs, approximately 9.0 x 10 per kilogram of body weight of the human subject receiving the product. 6 or more isolated fresh Tregs, approximately 9.5 x 10 per kilogram of body weight of the human subject receiving the product. 6 More than one freshly isolated CD4 + CD25 + CD127 dim Tregs, approximately 1.0 × 10 7 or more isolated fresh Tregs, approximately 1.5 x 10 per kilogram of body weight of the human subject receiving the product. 7or more than about 2.0 x 10 isolated fresh Tregs per kilogram of body weight of the human subject receiving the product. 7 and optionally, the Tregs are selected from the group consisting of FOXP3 + and / or d) isolated CD45 + The second population of cells was approximately 5.0 x 10 5 More than 6.0 x 10 freshly isolated Tregs 5 More than 7.0 x 10 freshly isolated Tregs 5 More than 8.0 x 10 freshly isolated Tregs 5 More than 9.0 x 10 freshly isolated Tregs 5 More than 1.0 x 10 freshly isolated Tregs 6 More than 1.5 x 10 freshly isolated Tregs 6 More than 2.0 x 10 freshly isolated Tregs 6 More than 2.5 x 10 freshly isolated Tregs 6 More than 3.0 x 10 freshly isolated Tregs 6 More than 3.5 x 10 freshly isolated Tregs 6 More than 4.0 x 10 freshly isolated Tregs 6 More than 4.5 x 10 freshly isolated Tregs 6 More than 5.0 x 10 freshly isolated Tregs 6 More than 5.5 x 10 freshly isolated Tregs 6 More than 6.0 x 10 freshly isolated Tregs 6 More than 6.5 x 10 freshly isolated Tregs 6 More than 7.0 x 10 freshly isolated Tregs 6 More than 7.5 x 10 freshly isolated Tregs 6 More than 8.0 x 10 freshly isolated Tregs 6 More than 8.5 x 10 freshly isolated Tregs 6 More than 9.0 x 10 freshly isolated Tregs6 More than 9.5 x 10 freshly isolated Tregs 6 More than 1.0 x 10 freshly isolated Tregs 7 More than 1.5 x 10 freshly isolated Tregs 7 More than 2.0 x 10 freshly isolated Tregs 7 More than 2.5 x 10 freshly isolated Tregs 7 More than 3.0 x 10 freshly isolated Tregs 7 More than 3.5 x 10 freshly isolated Tregs 7 More than 4.0 x 10 freshly isolated Tregs 7 More than 4.5 x 10 freshly isolated Tregs 7 More than 5.0 x 10 freshly isolated Tregs 7 More than 5.5 x 10 freshly isolated Tregs 7 More than 6.0 x 10 freshly isolated Tregs 7 More than 6.5 x 10 freshly isolated Tregs 7 More than 7.0 x 10 freshly isolated Tregs 7 More than 7.5 x 10 freshly isolated Tregs 7 More than 8.0 x 10 freshly isolated Tregs 7 More than 8.5 x 10 freshly isolated Tregs 7 More than 9.0 x 10 freshly isolated Tregs 7 More than 9.5 x 10 freshly isolated Tregs 7 More than 1.0 x 10 freshly isolated Tregs 8 More than 1.5 x 10 freshly isolated Tregs 8 More than 2.0 x 10 freshly isolated Tregs 8 More than 2.5 x 10 freshly isolated Tregs 8 More than 3.0 x 10 freshly isolated Tregs 8 More than 3.5 x 10 freshly isolated Tregs 8More than 4.0 x 10 freshly isolated Tregs 8 More than 4.5 x 10 freshly isolated Tregs 8 More than 5.0 x 10 freshly isolated Tregs 8 More than 5.5 x 10 freshly isolated Tregs 8 More than 6.0 x 10 freshly isolated Tregs 8 More than 6.5 x 10 freshly isolated Tregs 8 More than 7.0 x 10 freshly isolated Tregs 8 More than 7.5 x 10 freshly isolated Tregs 8 More than 8.0 x 10 freshly isolated Tregs 8 More than 8.5 x 10 freshly isolated Tregs 8 More than 9.0 x 10 freshly isolated Tregs 8 More than 9.5 x 10 freshly isolated Tregs 8 More than 1.0 x 10 freshly isolated Tregs 9 More than 1.5 x 10 freshly isolated Tregs 9 More than 2.0 x 10 freshly isolated Tregs 9 More than 2.5 x 10 freshly isolated Tregs 9 ≥ 10 freshly isolated Tregs, or approximately 3.0 x 10 9 and optionally, the Tregs are selected from the group consisting of FOXP3 + and / or e) isolated CD45 + The third population of cells is about 1.0 x 10 per kilogram of body weight of the human subject receiving the product. 5 or more than about 2.0 x 10 per kilogram of body weight of the human subject receiving the product 5 or more Tcon, approximately 3.0 x 10 per kilogram of body weight of the human subject receiving the product 5 or more than about 4.0 x 10 per kilogram of body weight of the human subject receiving the product 5or more than about 5.0 x 10 per kilogram of body weight of the human subject receiving the product 5 or more than about 6.0 x 10 per kilogram of body weight of the human subject receiving the product 5 or more Tcon, approximately 7.0 x 10 per kilogram of body weight of the human subject receiving the product 5 or more Tcon, approximately 8.0 x 10 per kilogram of body weight of the human subject receiving the product 5 or more Tcon, approximately 9.0 x 10 per kilogram of body weight of the human subject receiving the product 5 or more than about 1.0 x 10 per kilogram of body weight of the human subject receiving the product 6 or more Tcon, approximately 1.5 x 10 per kilogram of body weight of the human subject receiving the product 6 or more than about 2.0 x 10 per kilogram of body weight of the human subject receiving the product 6 or more than about 2.5 x 10 per kilogram of body weight of the human subject receiving the product 6 or more Tcon, approximately 3.0 x 10 per kilogram of body weight of the human subject receiving the product 6 or more Tcon, approximately 3.5 x 10 per kilogram of body weight of the human subject receiving the product 6 or more than about 4.0 x 10 per kilogram of body weight of the human subject receiving the product 6 or more Tcon, approximately 4.5 x 10 per kilogram of body weight of the human subject receiving the product 6 or more than about 5.0 x 10 per kilogram of body weight of the human subject receiving the product 6 or more Tcon, approximately 5.5 x 10 per kilogram of body weight of the human subject receiving the product 6 or more than about 6.0 x 10 per kilogram of body weight of the human subject receiving the product 6 or more Tcon, approximately 6.5 x 10 per kilogram of body weight of the human subject receiving the product 6 or more Tcon, approximately 7.0 x 10 per kilogram of body weight of the human subject receiving the product 6 or more Tcon, approximately 7.5 x 10 per kilogram of body weight of the human subject receiving the product6 or more Tcon, approximately 8.0 x 10 per kilogram of body weight of the human subject receiving the product 6 or more Tcon, approximately 8.5 x 10 per kilogram of body weight of the human subject receiving the product 6 or more Tcon, approximately 9.0 x 10 per kilogram of body weight of the human subject receiving the product 6 or more Tcon, approximately 9.5 x 10 per kilogram of body weight of the human subject receiving the product 6 or more than about 1.0 x 10 per kilogram of body weight of the human subject receiving the product 7 or more Tcon, approximately 1.5 x 10 per kilogram of body weight of the human subject receiving the product 7 or more than about 2.0 x 10 per kilogram of body weight of the human subject receiving the product 7 or more than about 2.5 x 10 per kilogram of body weight of the human subject receiving the product 7 or more Tcon, approximately 3.0 x 10 per kilogram of body weight of the human subject receiving the product 7 or more Tcon, approximately 3.5 x 10 per kilogram of body weight of the human subject receiving the product 7 or more than 4.0 x 10 per kilogram of body weight of the human subject receiving the product 7 and / or f) isolated CD45 + The third population of cells was approximately 5.0 x 10 5 More than 6.0 x 10 Tcon 5 More than 7.0 x 10 Tcon 5 More than 8.0 x 10 Tcon 5 More than 9.0 x 10 Tcon 5 More than 1.0 x 10 Tcon 6 More than 1.5 x 10 Tcon 6 More than 2.0 x 10 6 More than 2.5 x 10 Tcon 6 More than 3.0 x 10 6 More than 3.5 x 10 Tcon 6 More than 4.0 x 10 Tcon 6More than 4.5 x 10 Tcon 6 More than 5.0 x 10 6 More than 5.5 x 10 Tcon 6 More than 6.0 x 10 Tcon 6 More than 6.5 x 10 Tcon 6 More than 7.0 x 10 Tcon 6 More than 7.5 x 10 Tcon 6 More than 8.0 x 10 Tcon 6 More than 8.5 x 10 Tcon 6 More than 9.0 x 10 Tcon 6 More than Tcon, about 9.5 x 10 6 More than 1.0 x 10 Tcon 7 More than 1.5 x 10 Tcon 7 More than 2.0 x 10 7 More than 2.5 x 10 Tcon 7 More than 3.0 x 10 7 More than 3.5 x 10 Tcon 7 More than 4.0 x 10 Tcon 7 More than 4.5 x 10 Tcon 7 More than 5.0 x 10 7 More than 5.5 x 10 Tcon 7 More than 6.0 x 10 Tcon 7 More than 6.5 x 10 Tcon 7 More than 7.0 x 10 Tcon 7 More than 7.5 x 10 Tcon 7 More than 8.0 x 10 Tcon 7 More than 8.5 x 10 Tcon 7 More than 9.0 x 10 Tcon 7 More than 9.5 x 10 Tcon 7 More than 1.0 x 10 Tcon 8 More than 1.5 x 10 Tcon 8 More than 2.0 x 10 8 More than 2.5 x 10 Tcon 8 More than 3.0 x 10 8More than 3.5 x 10 Tcon 8 More than 4.0 x 10 Tcon 8 More than 4.5 x 10 Tcon 8 More than 5.0 x 10 8 More than 5.5 x 10 Tcon 8 More than 6.0 x 10 Tcon 8 More than 6.5 x 10 Tcon 8 More than 7.0 x 10 Tcon 8 More than 7.5 x 10 Tcon 8 More than 8.0 x 10 Tcon 8 More than 8.5 x 10 Tcon 8 More than 9.0 x 10 Tcon 8 More than 9.5 x 10 Tcon 8 More than 1.0 x 10 Tcon 9 More than 1.5 x 10 Tcon 9 More than 2.0 x 10 9 More than 2.5 x 10 Tcon 9 More than 3.0 x 10 9 More than 3.5 x 10 Tcon 9 More than 4.0 x 10 Tcon 9 More than 4.5 x 10 Tcon 9 More than 5.0 x 10 9 More than 5.5 x 10 Tcon 9 Tcon or more, or approximately 6.0 × 10 9 Contains one or more doses of Tcon.

[0056] In some embodiments that may be combined with any of the preceding embodiments, the product further comprises a pharmaceutical composition comprising a dose of the graft-versus-host disease (GVHD) prophylactic agent tacrolimus sufficient to maintain a trough blood level of about 5 ng / mL to about 10 ng / mL in a human subject receiving the product, optionally the pharmaceutical composition comprising a twice-daily dose of tacrolimus in the range of about 0.01 mg per kilogram of body weight of the human subject receiving the product to about 0.50 mg per kilogram of body weight of the human subject receiving the product, and optionally the tacrolimus inhibits CD45+ and in an amount to maintain a target blood level of about 1 ng / mL to about 10 ng / mL in a human subject receiving the product for about 20 days or more, about 25 days or more, about 30 days or more, about 35 days or more, about 40 days or more, about 45 days or more, about 50 days or more, about 55 days or more, about 60 days or more, about 65 days or more, about 70 days or more, about 75 days or more, about 80 days or more, about 85 days or more, about 90 days or more, about 95 days or more, about 100 days or more, about 110 days or more, about 120 days or more, about 130 days or more, about 140 days or more, or about 150 days after administration of the third population of cells.

[0057] In some embodiments that may be combined with any of the preceding embodiments, a) isolated CD45 + First population of cells, isolated CD45 + A second population of cells and / or isolated CD45 +the third population of cells is formulated in a volume ranging from about 5 mL to about 1 L; and / or b) the neutral pH is in the range of about 6.8 to about 7.6; and / or c) the excipient comprises a transport buffer, optionally the transport buffer comprising about 120 to 160 mEq of sodium, and / or the transport buffer comprising a total of about 270 to about 320 mOsmol / L, optionally the transport buffer is selected from phosphate buffered saline (PBS), human serum, PlasmaLyte, and any combination thereof, and optionally the transport buffer further comprises about 0.1% w / v to about 10% and / or d) the first single-dose bag, the second single-dose bag, and the third single-dose bag are polyvinyl chloride (PVC) transfer bags or ethylene vinyl acetate (EVA) transfer bags; and / or e) the one or more cryoprotectants are selected from sorbitol, dimethyl sulfoxide (DMSO), propylene glycol, glycerol, polyvinylpyrrolidone (PVP), polyethylene glycol (PEG), serum, HSA, hetastarch, CRYOSTOR CS2, CRYOSTOR CS5, and CRYOSTOR CS10.

[0058] In some embodiments that may be combined with any of the preceding embodiments, isolated CD45 + First population of cells, isolated CD45 +The second population of cells and the third population of isolated CD45+ cells are derived from an allogeneic donor with at least one HLA mismatch with the human subject receiving the product, optionally the HLA-mismatched donor is unrelated to the human subject receiving the product, or the HLA-mismatched donor is related to the human subject receiving the product, optionally the at least one HLA mismatch is in an allele selected from HLA-A, HLA-B, HLA-C, HLA-DRB1 and any combination thereof, optionally the cells with at least one HLA mismatch are derived from a donor that is 6 / 8 HLA mismatched with the human subject receiving the product, or from a donor that is 7 / 8 HLA mismatched with the human subject receiving the product, optionally the donor is homozygous for the HLA allele while the human subject receiving the product is heterozygous for the HLA allele. and thus, the donor who has at least one HLA mismatch with the human subject receiving the product has a mismatched HLA allele, or the donor who has at least one HLA mismatch with the human subject receiving the product has a mismatched HLA allele because the donor is heterozygous for that HLA allele while the human subject receiving the product is homozygous for that HLA allele, or the donor who has at least one HLA mismatch with the human subject receiving the product has a mismatched HLA allele because both the donor and the human subject receiving the product are heterozygous for that HLA allele, and optionally, the isolated CD45 + First population of cells, isolated CD45 + The second population of cells and the third population of isolated CD45+ cells, when administered, reduce non-relapse mortality in human subjects receiving the product and / or increase overall survival in human subjects receiving the product.

[0059] Another aspect of the present disclosure relates to a method of treating a human subject having or suspected of having a hematological malignancy, the method comprising administering to a subject a multi-component cell therapy product of any of the preceding embodiments, or an isolated CD45 hematopoietic stem progenitor cell (HSPC) comprising: a) a CD45 hematopoietic stem cell (HSPC) comprising: + a) a solution containing a first population of cells; b) isolated CD45 T cells containing regulatory T cells (Tregs); + a solution containing a second population of cells; c) isolated CD45 + a solution containing a third population of cells, the solution comprising the isolated CD45 + The third population of cells is CD3 + and b) a solution containing one or more doses of graft-versus-host disease (GVHD) antibodies, wherein the isolated CD45 + First population of cells, isolated CD45 + A second population of cells, and isolated CD45 + The third population of cells is obtained from a single allogeneic donor with at least one HLA mismatch to the human subject, and wherein after administration, the human subject's non-relapse mortality rate is reduced and / or after administration, the human subject's overall survival is increased compared to a corresponding human subject who has undergone standard allogeneic myeloablative hematopoietic stem cell transplant (alloHSCT) from a donor with at least one HLA mismatch to the corresponding human subject, optionally wherein the hematological malignancy is selected from acute lymphocytic leukemia, acute myeloid leukemia, chronic myeloid leukemia, multiple myeloma, lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, myelodysplastic syndrome, myelofibrosis, and blastic plasmacytoid dendritic cell neoplasm (BPDCN), and optionally wherein the administration includes administering to the human subject isolated CD45 + First population of cells, isolated CD45 + A second population of cells, and isolated CD45 + and injecting a third population of cells, optionally in this case isolated CD45 + A third population of cells was isolated and labeled with CD45 +about 24 to 120 hours after the first population of cells, optionally isolated CD45 + A third population of cells was isolated and labeled with CD45 + and administering the second population of cells about 24 to 120 hours after the first population of cells, optionally in which case the human subject is administered an isolated CD45 + and the human subject does not develop GVHD higher than Stage 2 within about 100 days, about 180 days, about 200 days, or about 1 year after administration of the third population of cells, optionally in which case the human subject has been previously treated or is concurrently treated for the hematological malignancy, and optionally in which case the GVHD prophylactic agent is tacrolimus and is initially administered to the human subject at about 0.03 mg / kg of the human subject's actual or ideal body weight / day, and optionally in which case the GVHD prophylactic agent is isolated CD45 + and optionally, in which case tacrolimus is administered about 12 hours to about 24 hours after administration of the third population of cells, wherein the tacrolimus is administered to the isolated CD45 + and in an amount that maintains a target blood level of at least about 3 ng / ml for at least about 20 days, or about 40 days, after administration of the third population of cells, optionally in which case tacrolimus is administered to the isolated CD45 + and for at least about 60 days, at least about 90 days, at least about 120 days, or up to about 160 days after administration of the third population of cells, optionally in which case administration of the GVHD prophylactic agent begins to taper off about 90 days after the initial administration of the GVHD prophylactic agent, optionally in which case the method further comprises administering about 1000 mg of mycophenolate mofetil (MMF), optionally in which case the MMF is administered to the isolated CD45 +the first administration is about 12 hours to about 24 hours after administration of the third population of cells, and optionally in which case administration of MMF begins to be tapered about 30 days, about 35 days, about 40 days, about 41 days, about 42 days, about 43 days, about 44 days, about 45 days, about 46 days, about 47 days, about 48 days, about 49 days, or about 50 days after the first administration of MMF, and optionally in which case the at least one HLA mismatch is with an allele selected from HLA-A, HLA-B, HLA-C, HLA-DRB1, and any combination thereof, and optionally in which case at least The allogeneic donor with another HLA mismatch is 6 / 8 HLA mismatched to the human subject, or 7 / 8 HLA mismatched to the human subject, optionally in this case, the allogeneic donor with at least one HLA mismatch to the human subject is homozygous for that HLA allele, while the human subject is heterozygous for that HLA allele, and therefore the allogeneic donor with mismatched HLA alleles or at least one HLA mismatch to the human subject is heterozygous for that HLA allele. while the human subject is homozygous for the HLA allele and therefore has a mismatched HLA allele, or an allogeneic donor with at least one HLA mismatch to the human subject has a mismatched HLA allele because both the allogeneic donor and the human subject are heterozygous for the HLA allele, optionally in this case, the method further comprises a conditioning treatment, wherein the conditioning treatment is administered before any of (a) to (d), optionally in this case, the conditioning treatment is a myeloablative conditioning treatment, optionally in this case In some cases, the myeloablative conditioning regimen includes at least three conditioning reagents, at least one of which includes thiotepa; optionally in these cases, the myeloablative conditioning regimen includes a dose of one or more of busulfan, fludarabine, and thiotepa; optionally in these cases, the dose of one or more of busulfan, fludarabine, and thiotepa is about 5 to about 12 mg of thiotepa per kg of the human subject's actual or ideal body weight; about 7 to 11 mg of busulfan per kg of the human subject's actual or ideal body weight;and body surface area per m , 2 and about 100 to about 200 mg of fludarabine per 100 mg of erythropoietin, optionally wherein the human subject's GVHD- and relapse-free survival (GFRS) is increased by at least about 3.5-fold, overall survival is increased by at least about 1.25-fold, and / or non-relapse mortality is reduced by at least about 60% following administration, compared to a corresponding human subject receiving standard myeloablative allogeneic hematopoietic stem cell transplant (alloHSCT). Another aspect of the present disclosure relates to a multicomponent cell therapy product of any of the preceding embodiments, or a multicomponent medical procedure of the present disclosure, for use in treating a human subject having or suspected of having a hematological malignancy. Another aspect of the present disclosure relates to the use of a multicomponent cell therapy product of any of the preceding embodiments, or a multicomponent medical procedure of the present disclosure, in the manufacture of a medicament for treating a human subject having or suspected of having a hematological malignancy.

[0060] Another aspect of the present disclosure relates to a method of preparing a multi-component cell therapy product, the method comprising: a) obtaining an HSPC-mobilized peripheral blood apheresis sample from a donor who is allogeneic to the human subject receiving the product; and b) obtaining about 1.0 x 10 HSPCs per kilogram of body weight of the human subject receiving the product. 5 ~Approx. 1.0×10 8 CD34 + Hematopoietic stem progenitor cells (HSPC), approximately 1.5×10 7 ~Approx. 1.5×10 10 HSPCs, approximately 5.0 x 10 5 ~Approx. 5.0×10 8 a first sample containing HSPCs, about 1.0 x 10 per kilogram of body weight of the human subject receiving the product 5 ~Approx. 2.0×10 7 CD4 + CD25 + CD127 dim Regulatory T cells (Treg), approximately 1.5 x 10 7 ~Approx. 3.0×10 9 Tregs, or approximately 5.0 × 10 5 ~Approx. 1.0×10 8a second sample containing about 1.0 x 10 Tregs per kilogram of body weight of a human subject receiving the product; 5 ~Approx. 4.0×10 7 Normal CD3 + T cells (Tcon), approximately 1.5×10 7 ~Approx. 6.0×10 9 Tcon, or approximately 5.0 x 10 5 ~Approx. 2.0×10 8 and selecting a third sample from the apheresis sample comprising Tcon(s), wherein the multi-component cell therapy product comprises Tcon(s), HSPC(s), and Treg(s), optionally wherein the HSPC(s) are selected by sorting the first sample with one or more ISP(s) specific for CD34 to select a CD34-enriched population, and optionally wherein the Treg(s) are selected by sorting the second sample with one or more ISP(s) specific for CD25, one or more ISP(s) specific for CD4, and / or one or more ISP(s) specific for CD127 to select a CD4 + and CD127 dim and optionally wherein the Tcon is selected by screening a third sample with one or more ISPs specific for CD3, and optionally wherein the third sample is cryopreserved. Another aspect of the present disclosure relates to a multicomponent cell therapy product made by the method of any of the preceding embodiments, optionally wherein the multicomponent cell therapy product is the multicomponent cell therapy product of any of the preceding embodiments.

[0061] Another aspect of the present disclosure relates to a method of treating a human subject having or suspected of having acute leukemia in complete remission, active leukemia, acute leukemia refractory to first-line therapy, or acute leukemia with minimal residual disease, the method comprising administering to the human subject a multi-component cell therapy product of any of the preceding embodiments, optionally wherein the acute leukemia is in complete remission with incomplete hematologic recovery, optionally wherein minimal residual disease is present in the human subject, or wherein minimal residual disease is absent in the human subject, optionally wherein the acute leukemia is classified as intermediate-risk to very high-risk acute leukemia, and optionally wherein the acute leukemia is acute myeloid leukemia (AML), acute lymphocytic leukemia (ALL), and / or mixed phenotype acute leukemia (MPAL).

[0062] Another aspect of the present disclosure relates to a method of treating a human subject having or suspected of having low-risk acute myeloid leukemia (AML), high-risk AML in complete remission, or chronic myeloid leukemia (CML), the method comprising administering to the human subject a multi-component cell therapy product of any of the preceding embodiments, optionally wherein the high-risk AML has a complex karyotype with three or more clonal chromosomal abnormalities. and wherein the three or more clonal chromosomal abnormalities are each selected from monosomic karyotypes -5, 5q-, -7, or 7q-, t(11q23, t(9;11), inv(3), t(3,3)t(6;9)t(9;22), normal karyotype with fms-like tyrosine kinase 3 (FLT3)-ITD mutation, and any combination thereof, and optionally wherein the CML is in blast crisis phase, second chronic phase, chronic phase, accelerated, has a history of blast crisis, and / or is treated with one or more first- or second-generation tyrosine kinase inhibitors.

[0013] Another aspect of the disclosure relates to the use of a multi-component cell therapy product of any of the preceding embodiments in the manufacture of a medicament for treating a human subject having or suspected of having low-risk acute myeloid leukemia (AML), high-risk AML in complete remission, or chronic myeloid leukemia (CML).

[0014] Another aspect of the disclosure relates to the use of a multi-component cell therapy product of any of the preceding embodiments in the manufacture of a medicament for treating a human subject having or suspected of having low-risk acute myeloid leukemia (AML), high-risk AML in complete remission, or chronic myeloid leukemia (CML).

[0063] Another aspect of the present disclosure relates to a method of treating a human subject having or suspected of having high-risk myelodysplastic syndrome, therapy-related myelodysplastic syndrome, and / or secondary myelodysplastic syndrome, comprising administering to the human subject a multi-component cell therapy product of any of the preceding embodiments, optionally wherein the human subject has active disease at the time of treatment with the multi-component cell therapy, optionally wherein the human subject has a blast burden of about 10% or less in the bone marrow, and optionally wherein the therapy-related and / or secondary myelodysplastic syndrome is in complete remission and is classified as intermediate-risk to high-risk. Another aspect of the present disclosure relates to a multi-component cell therapy product of any of the preceding embodiments for use in treating a human subject having or suspected of having high-risk myelodysplastic syndrome, therapy-related and / or secondary myelodysplastic syndrome. Another aspect of the present disclosure relates to the use of a multi-component cell therapy product of any of the preceding embodiments in the manufacture of a medicament for treating a human subject having or suspected of having high-risk myelodysplastic syndrome, therapy-related myelodysplastic syndrome and / or secondary myelodysplastic syndrome.

[0064] Another aspect of the disclosure relates to a method of treating a human subject having or suspected of having a myeloproliferative syndrome, the method comprising administering to the human subject the multicomponent cell therapy product of any of the preceding embodiments. Another aspect of the disclosure relates to the multicomponent cell therapy product of any of the preceding embodiments for use in treating a human subject having or suspected of having a myeloproliferative syndrome. Another aspect of the disclosure relates to the use of the multicomponent cell therapy product of any of the preceding embodiments in the manufacture of a medicament for treating a human subject having or suspected of having a myeloproliferative syndrome.

[0065] Another aspect of the present disclosure relates to a method of treating a human subject having or suspected of having a non-Hodgkin's lymphoma with a poor prognosis that is not suitable for autologous hematopoietic cell transplantation (HCT), the method comprising administering to the human subject a multi-component cell therapy product of any of the preceding embodiments. Another aspect of the present disclosure relates to a multi-component cell therapy product of any of the preceding embodiments for use in treating a human subject having or suspected of having a non-Hodgkin's lymphoma with a poor prognosis that is not suitable for autologous hematopoietic cell transplantation (HCT). Another aspect of the present disclosure relates to the use of a multi-component cell therapy product of any of the preceding embodiments in the manufacture of a medicament for treating a human subject having or suspected of having a non-Hodgkin's lymphoma with a poor prognosis that is not suitable for autologous hematopoietic cell transplantation (HCT).

[0066] In some embodiments that may be combined with any of the preceding embodiments, a) isolated CD45 + A third population of cells was isolated and labeled with CD45 + and / or b) isolated CD45 + A third population of cells was isolated and labeled with CD45 + and / or c) the multicomponent cell therapy further comprises a solution comprising one or more doses of a graft-versus-host disease (GVHD) prophylactic agent, optionally wherein the GVHD prophylactic agent is tacrolimus, optionally wherein the tacrolimus is provided in an amount sufficient to maintain a trough blood level of between about 5 ng / mL and about 10 ng / mL in the human subject, optionally wherein the tacrolimus is provided twice daily at a dose ranging from about 0.01 mg per kilogram of body weight of the human subject to about 0.50 mg per kilogram of body weight of the human subject, optionally wherein the tacrolimus is administered to an isolated CD45 + and / or d) an isolated CD45 cell line, wherein the tacrolimus is first administered about 12 hours to about 24 hours after the first administration of tacrolimus, optionally wherein the administration of tacrolimus begins about 90 days after the first administration of tacrolimus.+ The first population of cells is approximately 1.0 x 10 per kilogram of body weight of a human subject. 5 ~Approx. 1.0×10 8 HSPCs, approximately 5.0 x 10 5 ~Approx. 5.0×10 8 HSPCs, or approximately 1.5 x 10 7 ~Approx. 1.5×10 10 and / or e) isolated CD45 + The second population of cells is approximately 1.0 x 10 per kilogram of body weight of a human subject. 5 ~Approx. 2.0×10 7 Tregs, approximately 1.5 × 10 7 ~Approx. 3.0×10 9 Tregs, or approximately 5.0 × 10 5 ~Approx. 1.0×10 8 Tregs, optionally in which the Tregs are FOXP3 + and / or f) isolated CD45 + The third population of cells is approximately 1.0 x 10 per kilogram of body weight of a human subject. 5 ~Approx. 4.0×10 7 Tcon, approximately 5.0 x 10 5 ~Approx. 2.0×10 8 Tcon, or approximately 1.5 x 10 7 ~Approx. 6.0×10 9 and / or g) the human subject comprises an isolated CD45 + and / or h) does not develop GVHD higher than Stage 2 within about 100 days, about 180 days, about 200 days, or one year after administration of the third population of cells; and / or + First population of cells, isolated CD45 +The second population of cells, and the third population of isolated CD45+ cells, are derived from an allogeneic donor with at least one HLA mismatch to the human subject, optionally wherein the HLA-mismatched donor is unrelated to the human subject, or wherein the HLA-mismatched donor is related to the human subject, optionally wherein the at least one HLA mismatch is to an allele selected from HLA-A, HLA-B, HLA-C, HLA-DRB1, and any combination thereof, and optionally wherein the donor with at least one HLA mismatch is 6 / 8 HLA-mismatched to the human subject, or 7 / 8 HLA-mismatched to the human subject. A donor that is HLA mismatched, optionally in this case having at least one HLA mismatch with the human subject, because the donor is homozygous for that HLA allele while the human subject is heterozygous for that HLA allele, or a donor that has at least one HLA mismatch with the human subject, because the donor is heterozygous for that HLA allele while the human subject is homozygous for that HLA allele, or a donor that has at least one HLA mismatch with the human subject, because the donor is heterozygous for that HLA allele while the human subject is homozygous for that HLA allele, and the human subject are both heterozygous for the HLA alleles and therefore have mismatched HLA alleles; and / or i) the non-relapse mortality rate of the human subject is reduced after administration of the multicomponent cellular therapy compared to a matched human subject who has undergone a standard myeloablative allogeneic hematopoietic stem cell transplant (alloHSCT) from a donor with at least one HLA mismatch to the matched human subject; and / or j) the overall survival of the human subject is increased after administration of the multicomponent cellular therapy compared to a matched human subject who has undergone a standard myeloablative allogeneic hematopoietic stem cell transplant (alloHSCT) from a donor with at least one HLA mismatch to the matched human subject;and / or k) the method further comprises collecting one or more, or two or more, mobilized donated peripheral blood samples from the donor, optionally wherein the donated peripheral blood samples are mobilized with granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), plerixafor, or any combination thereof, optionally wherein at least one of the mobilized donated peripheral blood samples is CD34; + and / or l) the method further comprises a conditioning treatment, wherein the conditioning treatment is administered prior to administration of the multicomponent cellular therapy, optionally wherein the conditioning treatment is administered from about 2 days to about 10 days prior to administration of the multicomponent cellular therapy, optionally wherein the conditioning treatment is a total body irradiation-based (TBI-based) treatment, optionally wherein the TBI-based treatment further comprises one or more conditioning treatments, optionally wherein the one or more conditioning treatments are selected from cyclophosphamide, etoposide, thiotepa, and any combination thereof, optionally wherein the conditioning treatment is administered prior to administration of the multicomponent cellular therapy. The conditioning is a myeloablative conditioning regimen, optionally wherein the myeloablative conditioning regimen comprises one or more conditioning reagents, optionally wherein the one or more conditioning reagents are selected from thiotepa, busulfan, fludarabine, cyclophosphamide, antithymocyte globulin (ATG), and any combination thereof, or the myeloablative conditioning regimen comprises a dose of one or more of busulfan, fludarabine, and thiotepa, optionally wherein the dose of one or more of busulfan, fludarabine, and thiotepa is about 5 mg to about 12 mg of thiotepa per kilogram of body weight of the human subject, about 7 to about 11 mg of busulfan per kilogram of body weight of the human subject, and about 10 mg to about 15 mg of cyclophosphamide per square meter of body surface area of ​​the human subject. 2and / or m) the GVHD- and relapse-free survival (GFRS) of the human subject is increased by at least about 3.5-fold after administration of the multicomponent cellular therapy compared to a corresponding human subject receiving a standard myeloablative allogeneic hematopoietic stem cell transplant (alloHSCT); and / or n) the overall survival is increased by at least about 1.25-fold after administration of the multicomponent cellular therapy compared to a corresponding human subject receiving a standard myeloablative allogeneic hematopoietic stem cell transplant (alloHSCT); and / or o) the non-relapse mortality rate is increased by at least about 60% after administration of the multicomponent cellular therapy compared to a corresponding human subject receiving a standard myeloablative allogeneic hematopoietic stem cell transplant (alloHSCT). and / or p) chronic GVHD-free survival is increased following administration of the multicomponent cellular therapy compared to a corresponding human subject given standard myeloablative allogeneic hematopoietic stem cell transplant (alloHSCT), optionally in which the human subject experiences chronic GVHD-free survival of about 1 month or more, about 2 months or more, about 3 months or more, about 6 months or more, about 9 months or more, about 1 year or more, about 1.25 years or more, about 1.50 years or more, about 1.75 years or more, about 2 years or more, about 2.25 years or more, about 2.50 years or more, about 2.75 years or more, about 3 years or more, about 3.25 years or more, about 3.50 years or more, about 3.75 years or more, about 4 years or more, about 4.25 years or more, about 4.50 years or more, about 4.75 years or more, or about 5 years or more following administration of the multicomponent cellular therapy; and / or q) the incidence of primary graft failure or secondary graft failure is reduced following administration of the multicomponent cellular therapy compared to corresponding human subjects given standard myeloablative allogeneic hematopoietic stem cell transplantation (alloHSCT), optionally in which the human subject continues to have a graft failure for about 1 month or more, about 2 months or more, about 3 months or more, about 6 months or more, about 9 months or more, about 1 year or more, about 1.25 years or more, about 1.50 years or more, about 1.75 years or more, about 2 years or more, about 2.25 years or more, about 2.50 years or more, about 2.75 years or more, about 3 years or more, about 3.25 years or more, about 3. and / or r) the incidence, severity, or any combination thereof, of Grade I to Grade IV acute GVHD is reduced after administration of the multicomponent cellular therapy compared to a corresponding human subject who has received standard myeloablative allogeneic hematopoietic stem cell transplant (alloHSCT), optionally in which case the human subject is free from primary or secondary graft failure for more than 50 years, more than about 3.75 years, more than about 4 years, more than about 4.25 years, more than about 4.50 years, more than about 4.75 years, or more than about 5 years. ≥ about 2 months, ≥ about 3 months, ≥ about 6 months, ≥ about 9 months, ≥ about 1 year, ≥ about 1.25 years, ≥ about 1.50 years, ≥ about 1.75 years, ≥ about 2 years, ≥ about 2.25 years, ≥ about 2.50 years, ≥ about 2.75 years, ≥ about 3 years, ≥ about 3.25 years, ≥ about 3.50 years, ≥ about 3.75 years, ≥ about 4 years, ≥ about 4.25 years, ≥ about 4.50 years, ≥ about 4.75 years, or ≥ about 5 years in the absence of Grade I to Grade IV acute GVHD; and / or s) the incidence of steroid-refractory acute GVHD is higher than that of standard bone marrow transplants. or greater than about 1 month, or greater than about 2 months, or greater than about 3 months, or greater than about 6 months, or greater than about 9 months, or greater than about 1 year, or greater than about 1.25 years, or greater than about 1.50 years, or greater than about 1.75 years, or greater than about 2 years, or greater than about 2.25 years, or greater than about 2.50 years, or greater than about 2.75 years, or greater than about 3 years, or greater than about 3.25 years, or greater than about 3.50 years, or greater than about 3.75 years, or greater than about 4 years, or greater than about 5 years, or greater than about 6 years, or greater than about 7 years, or greater than about 8 years, or greater than about 9 years ...and / or t) GVHD- and relapse-free survival (GFRS) is increased after administration of the multicomponent cellular therapy compared to a corresponding human subject receiving standard myeloablative allogeneic hematopoietic stem cell transplant (alloHSCT), optionally wherein the human subject remains free from steroid-refractory acute GVHD for about 4.25 years or more, about 4.50 years or more, about 4.75 years or more, or about 5 years or more after administration of the multicomponent cellular therapy, compared to a corresponding human subject receiving standard myeloablative allogeneic hematopoietic stem cell transplant (alloHSCT), optionally wherein the human subject remains free from steroid-refractory acute GVHD for about 1 month or more, about 2 months or more, about 3 months or more, about 6 months or more, about 9 months or more, about 1 year or more, about 1.25 years or more, about 1.50 years or more, about 1 year or more, after administration of the multicomponent cellular therapy. or about 5 years or more; and / or u) relapse-free survival is increased after administration of the multicomponent cellular therapy compared to a corresponding human subject given standard myeloablative allogeneic hematopoietic stem cell transplant (alloHSCT), optionally in which case the human subject experiences GFRS for about 1 month or more after administration of the multicomponent cellular therapy, or about 5 years or more; and / or v) the incidence, severity, or timing of moderate to severe chronic GVHD, or any combination thereof, is greater than or equal to about 2 months, greater than or equal to about 3 months, greater than or equal to about 6 months, greater than or equal to about 9 months, greater than or equal to about 1 year, greater than or equal to about 1.25 years, greater than or equal to about 1.50 years, greater than or equal to about 1.75 years, greater than or equal to about 2 years, greater than or equal to about 2.25 years, greater than or equal to about 2.50 years, greater than or equal to about 2.75 years, greater than or equal to about 3 years, greater than or equal to about 3.25 years, greater than or equal to about 3.50 years, greater than or equal to about 3.75 years, greater than or equal to about 4 years, greater than or equal to about 4.25 years, greater than or equal to about 4.50 years, greater than or equal to about 4.75 years, or greater than or equal to about 5 years; and / or v) the incidence, severity, or timing of moderate to severe chronic GVHD, or any combination thereof, is greater than or equal to about 2 months, greater than or equal to about 3 months, greater than or equal to about 6 months, greater than or equal to about 9 months, greater than or equal to about 1 year, greater than or equal to about 1.25 years, greater than or equal to about 1.50 years, greater than or equal to about 1.75 years, greater than or equal to about 2 years, greater than or equal to about 2.25 years, greater than or equal to about 2.50 years, greater than or equal to about 2.75 years, greater than or equal to about 3 years or greater than about 1 month, or greater than about 2 months, or greater than about 3 months, or greater than about 6 months, or greater than about 9 months, or greater than about 1 year, or greater than about 1.25 years, or greater than about 1.50 years, or greater than about 1.75 years, or greater than about 2 years, or greater than about 2.25 years, or greater than about 2.50 years, or greater than about 2.75 years, or greater than about 3 years, or greater than about 3.25 years, or greater than about 3.50 years, or greater than about 3.75 years, or greater than about 4 years, or greater than about 4.25 years, or greater than about 4.50 years, or greater than about 4.75 years, or greater than about 5 years, or greater than about 6 years, or greater than about 7 years, or greater than about 8 years, or greater than about 9 years ...or about 5 years or more without severe chronic GVHD; and / or w) the incidence, severity, timing of non-relapse mortality, or any combination thereof, is reduced following administration of the multicomponent cellular therapy, as compared to a corresponding human subject given standard myeloablative allogeneic hematopoietic stem cell transplant (alloHSCT), optionally wherein the human subject remains free from severe chronic GVHD for about 1 month or more, about 2 months or more, about 3 months or more, about 6 months or more, about 9 months or more, about 1 year or more, about 1.25 years or more, about 1.50 years or more, about 1.75 years or more, about 2 years or more, about 2.25 years or more, and / or x) have not experienced a non-relapse mortality for about 2.50 years or more, about 2.75 years or more, about 3 years or more, about 3.25 years or more, about 3.50 years or more, about 3.75 years or more, about 4 years or more, about 4.25 years or more, about 4.50 years or more, about 4.75 years or more, or about 5 years or more; and / or x) Grade 3 or higher infections are reduced after administration of the multicomponent cellular therapy compared to a corresponding human subject given standard allogeneic hematopoietic stem cell transplant (alloHSCT); optionally, in which case the human subject remains free of non-relapse mortality for about 1 month or more, about 2 months or more, about 3 months or more, about 5 years or more, or about 6 months or more after administration of the multicomponent cellular therapy. and / or y) the neutrophil engraftment rate, time to engraftment, or both the engraftment rate and time to engraftment have been greater than or equal to 6 months, greater than or equal to about 9 months, greater than or equal to about 1 year, greater than or equal to about 1.25 years, greater than or equal to about 1.50 years, greater than or equal to about 1.75 years, greater than or equal to about 2 years, greater than or equal to about 2.25 years, greater than or equal to about 2.50 years, greater than or equal to about 2.75 years, greater than or equal to about 3 years, greater than or equal to about 3.25 years, greater than or equal to about 3.50 years, greater than or equal to about 3.75 years, greater than or equal to about 4 years, greater than or equal to about 4.25 years, greater than or equal to about 4.50 years, greater than or equal to about 4.75 years, or greater than or equal to about 5 years. or more, about 5 days or more, about 6 days or more, about 7 days or more, about 8 days or more, about 9 days or more, about 10 days or more, about 11 days or more, about 12 days or more, about 13 days or more, about 14 days or more, about 15 days or more, about 16 days or more, about 17 days or more, about 18 days or more, about 19 days or more, about 20 days or more, about 22 days or more, about 22 days or more, about 23 days or more, about 24 days or more, about 25 days or more, about 26 days or more, about 27 days or more, about 28 days or more, about 29 days or more, about 30 days or more, about 33 days or more,Approximately 33 days or more, approximately 34 days or more, approximately 35 days or more, approximately 36 days or more, approximately 37 days or more, approximately 38 days or more, approximately 39 days or more, approximately 40 days or more, approximately 44 days or more, approximately 44 days or more, approximately 44 days or more, approximately 44 days or more, approximately 45 days or more, approximately 46 days or more, approximately 47 days or more, approximately 48 days or more, approximately 49 days or more, approximately 50 days or more, approximately 55 days or more, approximately 55 days or more, approximately 55 days or more, approximately 55 days or more, approximately 55 days or more, approximately 56 days or more, approximately 57 days or more, approximately 58 days or more, approximately 59 days or more, approximately 60 days or more, approximately 66 days or more, approximately 66 days or more, approximately 66 days or more, approximately 66 days or more, approximately 66 days or more, approximately 66 days or more, approximately 66 days or more, approximately 67 days or more, approximately 68 days or greater than about 69 days, or greater than about 70 days, or greater than about 77 days, or greater than about 77 days, or greater than about 77 days, or greater than about 77 days, or greater than about 77 days, or greater than about 77 days, or greater than about 77 days, or greater than about 77 days, or greater than about 77 days, or greater than about 77 days, or greater than about 78 days, or greater than about 79 days, or greater than about 80 days, or greater than about 88 days, or greater than about 88 days, or greater than about 88 days, or greater than about 88 days, or greater than about 88 days, or greater than about 88 days, or greater than about 88 days, or greater than about 89 days, or greater than about 90 days; and / or z) platelet engraftment rate, time to engraftment, or both the rate and time to engraftment, compared to a matched human subject receiving standard myeloablative allogeneic hematopoietic stem cell transplant (alloHSCT). In comparison, platelet engraftment is increased after administration of the multicomponent cell therapy, optionally in which case platelet engraftment is increased by about 5 days or more, about 6 days or more, about 7 days or more, about 8 days or more, about 9 days or more, about 10 days or more, about 11 days or more, about 12 days or more, about 13 days or more, about 14 days or more, about 15 days or more, about 16 days or more, about 17 days or more, about 18 days or more, about 19 days or more, about 20 days or more, about 22 days or more, about 22 days or more, about 23 days or more, about 24 days or more, about 25 days or more, about 26 days or more, about 27 days or more, about 28 days or more, about 29 days or more, about 30 days or more, about 33 days or more, about 34 days or more, about 35 days or more, about 36 days or more, about 37 days or more, about 38 days or more, about 39 days or more, about 40 days or more, about 41 days or more, about 42 days or more, about 43 days or more, about 44 days or more, about 45 days or more, about 46 days or more, about 47 days or more, about 48 days or more, about 49 days or more, about 50 days or more, about 51 days or more, about 52 days or more, about 53 days or more, about 54 days or more, about 55 days or more, about 56 days or more, about 57 days or more, about 58 days or more, about 59 days or more, about 60 days or more, about 61 days or more, about 62 days or more, about 63 days or more, about 64 days or more, about 65 days or more, about 66 days or more, about 3 days or more, about 34 days or more, about 35 days or more, about 36 days or more, about 37 days or more, about 38 days or more, about 39 days or more, about 40 days or more, about 44 days or more, about 44 days or more, about 44 days or more, about 44 days or more, about 45 days or more, about 46 days or more, about 47 days or more, about 48 days or more, about 49 days or more, about 50 days or more, about 55 days or more, about 55 days or more, about 55 days or more, about 55 days or more, about 55 days or more, about 56 days or more, about 57 days or more, about 58 days or more, about 59 days or more, about 60 days or more, about 66 days or more, about 66 days or more, about 66 days or more, about 66 days or more, about 66 days or more, about 66 days or more, about 67 days or more, about 68 days or more,and / or aa) overall survival is increased after administration of the multicomponent cellular therapy compared to a corresponding human subject receiving standard myeloablative allogeneic hematopoietic stem cell transplant (alloHSCT), and / or ab) the overall survival is about 69 days or more, about 70 days or more, about 77 days or more, about 77 days or more, about 77 days or more, about 77 days or more, about 77 days or more, about 77 days or more, about 77 days or more, about 77 days or more, about 78 days or more, about 79 days or more, about 80 days or more, about 88 days or more, about 88 days or more, about 88 days or more, about 88 days or more, about 88 days or more, about 88 days or more, about 88 days or more, about 89 days or more, or about 90 days or more. and optionally wherein the human subject experiences an overall survival of about 1 month or more, about 2 months or more, about 3 months or more, about 6 months or more, about 9 months or more, about 1 year or more, about 1.25 years or more, about 1.50 years or more, about 1.75 years or more, about 2 years or more, about 2.25 years or more, about 2.50 years or more, about 2.75 years or more, about 3 years or more, about 3.25 years or more, about 3.50 years or more, about 3.75 years or more, about 4 years or more, about 4.25 years or more, about 4.50 years or more, about 4.75 years or more, or about 5 years or more after administration of the multicomponent cell therapy; and / or bb) the readmission rate is greater than or equal to the standard or greater than about 1 month, or greater than about 2 months, or greater than about 3 months, or greater than about 6 months, or greater than about 9 months, or greater than about 1 year, or greater than about 1.25 years, or greater than about 1.50 years, or greater than about 1.75 years, or greater than about 2 years, or greater than about 2.25 years, or greater than about 2.50 years, or greater than about 2.75 years, or greater than about 3 years, or greater than about 3.25 years, or greater than about 3.50 years, or greater than about 3.75 years, or greater than about 4 years, or greater than about 4 years, or greater than about 5 years, or greater than about 6 years, or greater than about 7 years, or greater than about 8 years, or greater than about 9 years, or greater than about 1 year, or greater than about 1.25 years, or greater than about 1.50 years, or greater than about 1.75 years, or greater than about 2 years, or greater than about 2.25 years, or greater than about 2.50 years, or greater than about 2.75 years, or greater than about 3 years, or greater than about 3.25 years, or greater than about 3.50 years, or greater than about 3.75 years, or greater than about 4 years, or greater than about 5 years, or greater than about 6 years, or greater than about 7 years, or greater than about 8 years, or greater than about 9 years, or greater than about 1 year, or greater than about 1.25 years, or greater than about 1.50 years, or greater than about 1.75 years, or greater than about 2 years, or greater than about 3 years, or greater than about 4 years, or greater than about 5 years, or greater than about 6 years, or greater than about 7 years, or greater than about 8 years, or greater than about 9 years, or greater than about 1 year, or greater than about 1.2 0.25 years or more, about 4.50 years or more, about 4.75 years or more, or about 5 years or more without requiring rehospitalization; and / or cc) the human subject is about 3 months of age or older, or the human subject is about 3 months of age to about 18 years of age, or the human subject is about 18 years of age or older, or the human subject is about 18 years of age to about 65 years of age, or the human subject is about 18 years of age to about 75 years of age, or the human subject is about 66 years of age to about 75 years of age, or the human subject is about 3 months of age to about 75 years of age; and / or dd) the human subject has received between one and five prior lines of therapy.

[0067] Another aspect of the present disclosure relates to a cell therapy kit comprising the multi-component cell therapy product of any of the preceding embodiments, optionally wherein the kit further comprises written instructions for use of the cell therapy to treat a hematological malignancy in a human subject, optionally wherein the hematological malignancy is selected from leukemia, acute leukemia, chronic myeloid leukemia (CML), multiple myeloma, lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, myelodysplastic syndrome, myeloproliferative syndrome, myelofibrosis, and blastic plasmacytoid dendritic cell neoplasm (BPDCN); optionally wherein the leukemia is active, optionally wherein the acute leukemia is in complete remission or in complete remission with incomplete hematological recovery, and optionally wherein the leukemia is in active or in complete remission with incomplete hematological recovery. and wherein minimal residual disease is present in the human subject, or wherein minimal residual disease is not present in the human subject, optionally wherein the acute leukemia is classified as intermediate-risk to very high-risk acute leukemia, optionally wherein the acute leukemia is acute leukemia refractory to first-line therapy or acute leukemia with minimal residual disease, optionally wherein the acute leukemia is acute myeloid leukemia (AML), acute lymphoblastic leukemia (ALL), and / or mixed phenotype acute leukemia (MPAL), optionally wherein the AML is low-risk to high-risk AML, optionally wherein the high-risk AML is characterized by monosomic karyotype -5, 5q-, -7 or 7q-, t(11q23, t(9;11), inv(3), t(3,3)t(6;9)t(9;22), comprising a complex karyotype with three or more clonal chromosomal abnormalities selected from a normal karyotype with an fms-like tyrosine kinase 3 (FLT3)-ITD mutation, and any combination thereof, optionally wherein the CML is in blast crisis phase, second chronic phase, chronic phase, accelerated, has a history of blast crisis, and / or is intolerant to one or more first- or second-generation tyrosine kinase inhibitors (TKIs), optionally wherein the myelodysplastic syndrome is classified as high-risk, optionally wherein the myelodysplastic syndrome is a therapy-related myelodysplastic syndrome and / or a secondary myelodysplastic syndrome, and optionally wherein the non-Hodgkin's lymphoma is a poor-prognosis non-Hodgkin's lymphoma not suitable for autologous hematopoietic cell transplantation (HCT);

[0068] It will be understood that various aspects and / or embodiments of the present disclosure can be considered individually, collectively, or in combination with each other. Any description herein of a particular composition, multi-component medical treatment, multi-component cell therapy product, cell population, solution, formulation, kit, and / or method may apply to and be used in any other particular composition, multi-component medical treatment, multi-component cell therapy product, cell population, solution, formulation, kit, and / or method. Furthermore, any composition disclosed herein is applicable to any method disclosed herein. In other words, any aspect or embodiment described herein can be combined with any other aspect or embodiment disclosed herein.

[0069] Incorporation by Reference All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference. [Brief explanation of the drawings]

[0070] The novel features of the present disclosure are set forth with particularity in the appended claims. A better understanding of the features and advantages of the present disclosure will be obtained by reference to the following detailed description that sets forth illustrative embodiments, in which the principles of the disclosure are utilized, and the accompanying drawings.

[0071] [Figure 1A] Figures 1A-B illustrate a schematic of transplantation according to the method described herein (identified as high-precision Orca-T or Orca-T) and the differences compared to the standard of care (SOC) cohort (identified as conventional transplantation or SOC). [Figure 1B] Same as above. [Figure 1C] FIG. 1C illustrates a schematic of implant creation and administration. [Figure 2] Figure 2A illustrates the weight of patients enrolled in the studies disclosed in the Examples. Figures 2B-2C illustrate the doses of HSPCs and Treg cells administered to patients enrolled in the studies disclosed in the Examples. Figure 2D illustrates the purity of Treg cells administered to patients enrolled in the studies disclosed in the Examples. [Figure 3A] Figure 3A shows the time to platelet engraftment in the experimental group (identified as Orca-T) and standard of care (SOC) cohorts. [Figure 3B]Figures 3B-3L illustrate the engraftment of various cell populations in patients from the study group disclosed in the Examples. These figures also illustrate the levels of each cell type in the donor before sample collection. Boxplots show the following: boxes indicate the 75th, 50th, and 25th percentiles; whiskers indicate the 90th and 10th percentiles. X-axis nomenclature: leading numbers (e.g., 01, 02, 025, ...) are used for ordering; following the underline: Dscrn = healthy donor before G-CSF mobilization; Rscrn = recipient within 1 month of transplant conditioning; apher = healthy donor blood drawn at the time of apheresis; d028 = recipient 28 days post-transplant; d056-d365 = recipient days post-transplant. The indicated N indicates the sample size at each time point. Symbols indicate the values ​​of individual measurements. 10-3 times the number of cells per uL of blood is equal to 1000 times the number of cells per uL of blood. [Figure 3C] Same as above. [Figure 3D] Same as above. [Figure 3E] Same as above. [Figure 3F] Same as above. [Figure 3G] Same as above. [Figure 3H] Same as above. [Figure 3I] Same as above. [Figure 3J] Same as above. [Figure 3K] Same as above. [Figure 3L] Same as above. [Figure 3M] Figures 3M-3N show the timeline of lymphocyte and monocyte engraftment in a subset of the test group (Orca-T) and standard of care cohort. [Figure 3N] Same as above. [Figure 3O]Figure 3O shows representative flow cytometry data for the frequency of CD3+CD4+ T cells that were Tregs in two subjects compared with healthy controls. In the healthy controls, 3.72% of circulating CD3+CD4+ T cells were Tregs (CD25+CD127dim). In two transplant recipients, 28.1% and 23.7% of CD3+CD4+ T cells were Tregs at day +28, 32.3% and 17.8% at day +56, and 19.2% and 20.7% at day +100 after transplantation. [Figure 3P] Figure 3P shows flow cytometry data for B cell markers in samples from recipients of compositions of the present disclosure compared to healthy controls. In all cases, the Y-axis is CD19+ staining. The left panel shows lymphocyte gating to identify B cells (CD19+) and T cells (CD3+). 13.4% of lymphocytes in the graft recipient were B cells, compared to 9.84% in the healthy control. The second panel from the left shows that 98.3-100% of cells gated as CD19+ were also CD20+. The second panel from the right shows the fraction of IgD+ B cells, which can be used to identify mature B cells. 92.1% of B cells in the graft recipient were IgD+, compared to 89.5% in the healthy control. The rightmost panel shows staining for CD27, which can be used to identify memory B cells, late plasmablasts, and plasma cells. For example, 43.6% of B cells in graft recipients were CD27+ compared with 67.1% in healthy controls. [Figure 4A] Figure 4A shows the incidence of grade 2 or higher aGVHD in the experimental (Orca-T) and standard of care cohorts by day +120 post-transplant. At nearly all time points, the Orca-T data are lower than the standard of care data. [Figure 4B] Figure 4B shows the incidence of grade 3 or higher aGVHD in the experimental group (Orca T) and standard of care cohorts by day +120 post-transplant. At nearly all time points, the Orca-T data are below the standard of care data. [Figure 4C]Figure 4C shows the incidence of moderate to severe cGVHD in the experimental (Orca-T) and standard of care (SOC) cohorts up to day +365 post-transplant. At almost all time points, the Orca-T data are lower than the standard of care data. [Figure 4D] Figure 4D shows non-relapse-related mortality in the experimental (Orca-T) and standard of care (SOC) cohorts up to day +365 post-transplant. At almost all time points, the Orca-T data are lower than the standard of care data. [Figure 4E] Figure 4E shows the relapse rates in the experimental group (Orca-T) and standard of care cohorts up to day +365 post-transplant. At the final time point, the relapse rate in the Orca-T group was 16%, and the relapse rate in the standard of care group was 19%. [Figure 4F] Figure 4F shows GVHD- and relapse-free survival in the experimental (Orca-T) and standard of care cohorts up to day +365 post-transplant. At nearly all time points, the Orca-T data exceeds the standard of care data. [Figure 4G] Figure 4G shows cGVHD-free survival in the experimental and standard of care cohorts up to day +365 post-transplant. At nearly all time points, the Orca-T data outperform the standard of care data. [Figure 4H] Figure 4H shows overall survival in the experimental and standard of care cohorts up to day +365 post-transplant. At the final time point, the overall survival rate for Orca-T is 90% and for standard of care is 78%. [Figure 4I] Figure 4I shows the number of hospital days in the study group and a subset of the standard of care (SOC) cohort through day +365 post-transplant. [Figure 5] Figure 5 summarizes the disease status of a small subset of subjects in the study group before transplant and at days +90, +180, and +356 after transplant. CR indicates complete response and MRD indicates minimal residual disease. [Figure 6A]Figures 6A-6F compare aGVHD, cGVHD, relapse, relapse-free survival, GVHD- and relapse-free survival (GRFS), and overall survival for subsets of patients in study arms receiving different conditioning regimens. [Figure 6B] Same as above. [Figure 6C] Same as above. [Figure 6D] Same as above. [Figure 6E] Same as above. [Figure 6F] Same as above. [Figure 7A] Figures 7A-7H compare aGVHD, cGVHD, non-relapse related mortality, relapse, relapse-free survival, GVHD- and relapse-free survival (GRFS), and overall survival for subsets of patients in study groups receiving different GVHD prophylaxis. [Figure 7B] Same as above. [Figure 7C] Same as above. [Figure 7D] Same as above. [Figure 7E] Same as above. [Figure 7F] Same as above. [Figure 7G] Same as above. [Figure 7H] Same as above. [Figure 8A] 8A-8C illustrate the rates of aGVHD and cGVHD in patients with different serum tacrolimus trough levels. [Figure 8B] Same as above. [Figure 8C] Same as above. [Figure 9A] Figures 9A-9B compare the levels of aGVHD and cGVHD in patients with different serum tacrolimus levels. [Figure 9B] Same as above. [Figure 9C] Figures 9C-9D compare the levels of aGVHD and cGVHD in patients with different serum tacrolimus levels but who received the same conditioning regimen of busulfan and cyclophosphamide (Bu / Cy). [Figure 9D] Same as above. [Figure 9E]Figures 9E-9G compare the levels of aGVHD and cGVHD in patients with different serum tacrolimus levels but who received the same conditioning regimen of total body irradiation (TBI) / busulfan, fludarabine, and thiotepa (TBI / BFT). [Figure 9F] Same as above. [Figure 9G] Same as above. [Figure 9H] Figure 9H shows the mean trough levels of tacrolimus up to day +30 post-transplant plotted against the percentage of CD3+ cells of donor origin at day +30 (excluding chimerism loss data from day 90, indicated by "D90"). [Figure 10] Figure 10A shows rapid neutrophil and platelet engraftment after Orca-T plus single-agent GVHD prophylaxis. Neutrophil and platelet engraftment occurred in a median of 13 and 15 days, respectively. Of the 137 patients treated with Orca-T, two graft failure events (1.4%) were reported. Figure 10B shows additional neutrophil and platelet engraftment data, showing that neutrophil engraftment occurred in a median of 13 days and platelet engraftment occurred in a median of 19 days. [Figure 11] Figure 11 shows the results for acute GVHD, demonstrating that it was rare with Orca-T despite the limited GVHD prophylaxis regimen. Grade ≥2 and Grade ≥3 aGVHD rates were 17% and 4%, respectively, by day +180 with Orca-T + single-agent GVHD prophylaxis (graded according to MAGIC criteria). Only four patients (3%) experienced steroid-refractory or steroid-dependent aGVHD. [Figure 12] Figure 12 shows that moderate / severe chronic GVHD was rare in Orca-T. The NIH rate of moderate-to-severe cGVHD was 5% up to 1 year post-transplant. [Figure 13]Figure 13 shows that despite the reduced rate of GVHD, the relapse rate after Orca-T was not elevated compared to historical controls, suggesting that the graft-versus-leukemia effect was not compromised with Orca-T. Of note, patients with active leukemia at the time of transplant were allowed to enroll in the Orca-T trial, provided they had a bone marrow blast burden of 10% or less. 14% of enrolled patients had active leukemia at the time of transplant; similarly, 16% of the comparison cohort had "progressive" disease. [Figure 14] Figure 14 shows that adding thiotepa to busulfan-based conditioning reduced relapse without increasing non-relapse mortality in Orca-T. Relapse-free survival through 1 year is shown for all Orca-T patients (n=137), those receiving Bu / Flu or Bu / Cy conditioning before Orca-T (n=50), and those receiving Bu / Flu / thiotepa conditioning before Orca-T (n=49). [Figure 15] Figure 15 shows the reconstitution of lymphocyte subsets. Absolute cell counts of T cells, CD4+ T cells, CD8+ T cells, regulatory T cells, B cells, and NK cells per microliter of whole blood are shown in recipients before transplantation and one year after transplantation. The number of patients included at each time point is indicated in each graph. Blue circles and bars indicate median and quartile values. [Figure 16] Figure 16 shows that severe infections were found to be rare in patients treated with Orca-T. The incidence of grade 3 infections is shown using the BMT-CTN MOP V4.0 rating scale. COVID-19 / SARS-COV2 infections were reported in 13 patients, with one death. [Figure 17] Figure 17 shows the results of Orca-T + single agent GVHD prophylaxis in GVHD and relapse-free survival (GRFS) at 1 year, which was found to be superior to existing comparators. Modified GRFS shown: Grade 3-4 aGVHD, moderate-to-severe cGVHD, and relapse-free survival. [Figure 18]Figure 18 shows preliminary overall survival. The data suggest that Orca-T may result in improved survival in patients undergoing allogeneic hematopoietic stem cell transplantation (alloHSCT) for high-risk hematologic malignancies. [Figure 19] Figure 19A shows neutrophil and platelet engraftment from 7 / 8 HLA-mismatched donors using Orca-T. Neutrophil and platelet engraftment occurred in a median of 12.5 and 15.5 days, respectively. Figure 19B shows the chimerism rates of whole blood and T cells from 7 / 8 HLA-mismatched donors using Orca-T. Whole blood showed complete chimerism, and T cells showed greater than 90% chimerism 90 days after transplant. [Figure 20] Figure 20A shows IL-2 serum plasma levels 14 days after transplant using Orca-T from 7 / 8 HLA-mismatched or 8 / 8 matched donors. Figure 20B shows IL-10 serum plasma levels 14 days after transplant using Orca-T from 7 / 8 HLA-mismatched or 8 / 8 matched donors. [Figure 21] FIG. 21 shows the clinical outcomes of six patients who received Orca-T from 7 / 8 HLA-mismatched donors at 90, 180, and 365 days after transplantation. [Figure 22] Figure 22A shows relapse-free survival in patients who received BFT conditioning prior to Orca-T. Figure 22B shows non-relapse mortality in patients who received BFT conditioning prior to Orca-T. Median follow-up time was 413 days. [Figure 23] Figure 23 shows relapse-free survival by MRD in patients who received BFT conditioning prior to Orca-T. Median follow-up time was 413 days. [Figure 24] Figure 24 shows the incidence of grade 3+ infections (using the BMT-CTN MOP V4.0 rating scale) in patients who received BFT conditioning prior to Orca-T. [Figure 25]Figure 25A shows the incidence of grade 3 or higher acute GVHD (aGVHD) in patients who received BFT conditioning prior to Orca-T. Figure 25B shows the incidence of moderate to severe chronic GVHD (cGVHD) in patients who received BFT conditioning prior to Orca-T. [Figure 26] Figure 26A shows the 1-year recurrence-free survival (GRFS) rates for acute grade 3-4 and chronic moderate-to-severe GVHD in patients who received BFT conditioning prior to Orca-T. Figure 26B shows the 1-year overall survival in patients who received BFT conditioning prior to Orca-T. DETAILED DESCRIPTION OF THE INVENTION

[0072] Introduction Various embodiments of the present disclosure provide compositions, multi-component medical procedures, multi-component cell therapy products, cell populations, solutions, formulations, kits, and / or methods related to improved allogeneic hematopoietic stem cell transplantation (alloHSCT), including transplantation of distinct cell populations enriched for hematopoietic stem progenitor cells (HSPCs), regulatory T cells (Tregs), and conventional T cells (Tcon).

[0073] Standard allogeneic hematopoietic stem cell transplantation (HSCT) is the transplantation of multipotent hematopoietic stem progenitor cells (HSPCs), usually derived from the donor's bone marrow, peripheral blood, or umbilical cord blood, into a recipient. The recipient may undergo myeloablative conditioning, which kills hematopoietic cells, including tumor cells and host immune cells. HSPCs then reconstitute the hematopoietic compartment. HSCT is useful as a cancer treatment, due to its ability to induce donor T cells to exert an antitumor effect, termed graft-versus-tumor (GVT). HSCT is associated with improved survival in patients with hematologic malignancies refractory to chemotherapy.

[0074] Surprisingly, recipients of the improved allo-HSCT of the present disclosure, receiving a single-agent graft-versus-host disease (GVHD) prophylaxis, such as tacrolimus, have had much better clinical outcomes than existing standard allo-HSCT regimens and standard therapies. These recipients experience improved clinical outcomes, including, for example, increased overall survival, increased relapse-free survival, increased GVHD- and relapse-free survival (GRFS), more rapid and / or complete engraftment of various hematopoietic components (e.g., neutrophils, platelets, T cells, B cells), improved donor chimerism (e.g., T cell chimerism), reduced relapse, reduced primary engraftment failure, reduced secondary engraftment failure, reduced treatment-related mortality, reduced acute and / or chronic GVHD, and shorter time to hospital discharge after a single-agent GVHD prophylaxis, such as tacrolimus.

[0075] Although alloHSCT is associated with improved survival for patients with hematologic malignancies refractory to chemotherapy, some subjects treated with existing alloHSCT regimens experience cancer recurrence, and numerous complications can limit the effectiveness of alloHSCT. The efficacy of standard alloHSCT can be limited by, for example, primary engraftment failure, secondary engraftment failure, limited or delayed engraftment of various hematopoietic components (e.g., neutrophils, platelets, T cells, or B cells), and limited donor chimerism (e.g., T cell chimerism). Furthermore, standard alloHSCT can result in treatment-related ethics or toxicity. However, donor T cells can also attack non-tumor host cells, resulting in graft-versus-host disease (GVHD). GVHD is a major cause of post-HCT complications and can be fatal. Managing GVHD may require immunosuppressive therapy or cytotoxic mediators, which can cause toxicity, increased susceptibility to infection, and / or blunted antitumor immunity. Early morbidity and mortality associated with acute graft-versus-host disease (aGVHD; occurring within the first 100 days after transplantation) are major factors limiting the success of HCT, as is the long-term morbidity associated with chronic GVHD (cGVHD). GVHD is a risk for both HLA-matched and HLA-mismatched transplants. GVHD can occur even when the donor and recipient are HLA-matched because the immune system can still recognize other differences between the donor tissues.

[0076] Both GVHD and GVHD are primarily mediated by conventional T cells (T cells), which initiate immune responses upon recognition of cognate antigens by T cell receptors. Depleting T cells from hematopoietic stem cell transplant (HCT) grafts can reduce GVHD, but it can also reduce GVHD and increase the likelihood of cancer recurrence. In addition to T cells, T cells are a further subset of T cells that negatively regulate inflammation and promote immune tolerance. T cells can prevent or reduce GVHD by negatively regulating inflammation, including inflammation induced by donor T cells when they recognize recipient antigens.

[0077] Provided herein are compositions and methods for improved alloHSCT, which comprise administering to a subject a CD45+ HSPC-containing CD45+ HSPC. + The first population of cells, CD45, contains at least Tregs + A second population of cells, and CD45 containing at least Tcon + In some embodiments, administering a specific cell population, including a third population of cells, comprises administering a specific cell population, including a CD45 + The second population of cells is also referred to as a Treg-enriched cell population. Without wishing to be bound by theory, CD45 + By administering a second population of cells, the incidence and / or severity of GVHD is reduced, while the CD45 + Administering a third population of cells enhances GVT. Accordingly, embodiments of the present disclosure provide compositions, multicomponent medical treatments, multicomponent cell therapy products, cell populations, solutions, formulations, kits, and / or methods for administering at least both populations of T cells to enhance GVT while minimizing GVHD. Thus, the compositions, multicomponent medical treatments, multicomponent cell therapy products, cell populations, solutions, formulations, kits, and / or methods disclosed herein can prevent or reduce graft-versus-host disease (GVHD) in a subject while preserving the graft-versus-tumor (GVT) effect of alloHSCT administered to a subject with cancer (e.g., a hematological cancer). In some embodiments, two or more cell populations are administered at different times. For example, a CD45 cell population comprising at least HSPCs can be administered. + The first population of cells, and the Treg-enriched cell population, contains at least Tcon and CD45 + It may be administered before the third population of cells.

[0078] cell population Embodiments of the present disclosure provide a multi-component medical treatment or multi-component cell therapy product administered to a human subject in need thereof. In some embodiments, the multi-component treatment comprises: (a) a CD45 hematopoietic stem progenitor cell (HSPC) and granulocyte-derived cell line comprising: + A solution containing a first population of cells, + (b) a solution containing a cell population enriched for regulatory T cells (Treg); (c) a solution containing up to about 10% of the first population of cells containing granulocytes; + a solution containing a third population of cells, + A third population of cells is at least about 20% CD3 + and (d) a solution comprising one or more doses of a graft-versus-host disease (GVHD) prophylactic agent, such as tacrolimus. In various embodiments, the HSPCs are CD34 + In some embodiments, the multi-component cell therapy product comprises (a) about 1.0 x 10 cells per kilogram of body weight of the human subject receiving the product. 5 ~Approx. 1.0×10 8 CD34 + Hematopoietic stem progenitor cells (HSPC), approximately 1.5×10 7 ~Approx. 1.5×10 10 HSPCs, or approximately 5.0 x 10 5 ~Approx. 5.0×10 8 A dose of isolated CD45 HSPCs containing + a first single-dose transfer bag containing a first population of cells, + (b) the first population of cells is formulated with an excipient at a neutral pH; and (b) about 1.0 x 10 cells per kilogram of body weight of a human subject receiving the product. 5 ~Approx. 2.0×10 7 Fresh CD4 + CD25 + CD127 dim Regulatory T cells (Treg), approximately 1.5 x 10 7 ~Approx. 3.0×10 9 fresh Tregs, or approximately 5.0 × 10 5 ~Approx. 1.0×108 A dose of fresh Tregs containing isolated CD45 + a second single-dose transfer bag containing a second population of cells, the isolated CD45 + the second population of cells is formulated with an excipient at a neutral pH; and (c) about 1.0 x 10 cells per kilogram of body weight of a human subject receiving the product. 5 ~Approx. 4.0×10 7 Normal CD3 + T cells (Tcon), approximately 1.5×10 7 ~Approx. 6.0×10 9 Tcon, or approximately 5.0 x 10 5 ~Approx. 2.0×10 8 Isolated CD45 containing a dose of Tcon + a third single-dose transfer bag containing a third population of cells, the third population comprising the isolated CD45 + A third population of cells includes those formulated with an excipient at neutral pH, where the excipient includes one or more cryoprotectants.

[0079] In some embodiments, CD45 + The first population of cells comprises HSPCs. + The first population of cells comprises at least one dose of HSPCs. In some embodiments, the first population of cells comprises or comprises at least one dose of HSPCs, CD45 + The first population of cells is approximately 1.0 x 10 per kilogram of body weight of a human subject. 5 ~Approx. 5.0×10 8 Approximately 1.0 x 10 HSPCs per kilogram of body weight in a human subject 5 ~Approx. 1.0×10 8 HSPCs, or approximately 5.0 x 10 per kilogram of body weight in a human subject 5 ~Approx. 2.0×10 7 In some embodiments, the HSPCs comprise, or at least one dose of, HSPCs, CD45 + The first population of cells is approximately 1.0 x 10 per kilogram of body weight of a human subject. 5≥ 1.5 x 10 HSPCs per kilogram of body weight in a human subject 5 ≥ 2.0 x 10 HSPCs per kilogram of body weight in a human subject 5 ≥ 2.5 x 10 HSPCs per kilogram of body weight in a human subject 5 ≥ 3.0 x 10 HSPCs per kilogram of body weight in a human subject 5 ≥ 3.5 x 10 HSPCs per kilogram of body weight in a human subject 5 ≥ 4.0 x 10 HSPCs per kilogram of body weight in a human subject 5 ≥ 4.5 x 10 HSPCs per kilogram of body weight in a human subject 5 ≥ 5.0 x 10 HSPCs per kilogram of body weight in a human subject 5 ≥ 5.5 x 10 HSPCs per kilogram of body weight in a human subject 5 ≥ 6.0 x 10 HSPCs per kilogram of body weight in a human subject 5 ≥ 6.5 x 10 HSPCs per kilogram of body weight in a human subject 5 ≥ 7.0 x 10 HSPCs per kilogram of body weight in a human subject 5 ≥ 7.5 x 10 HSPCs per kilogram of body weight in a human subject 5 ≥ 8.0 x 10 HSPCs per kilogram of body weight in a human subject 5 ≥ 8.5 x 10 HSPCs per kilogram of body weight in a human subject 5 ≥ 9.0 x 10 HSPCs per kilogram of body weight in a human subject 5 ≥ 9.5 x 10 HSPCs per kilogram of body weight in a human subject 5 ≥ 1.0 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 1.25 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 1.5 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 1.75 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 2.0 x 10 HSPCs per kilogram of body weight in a human subject 6≥ 2.25 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 2.5 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 2.75 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 3.0 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 3.25 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 3.5 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 3.75 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 4.0 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 4.25 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 4.5 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 4.75 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 5.0 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 5.25 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 5.5 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 5.75 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 6.0 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 6.25 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 6.5 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 6.75 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 7.0 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 7.25 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 7.5 x 10 HSPCs per kilogram of body weight in a human subject 6≥ 7.75 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 8.0 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 8.25 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 8.5 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 8.75 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 9.0 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 9.25 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 9.5 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 9.75 x 10 HSPCs per kilogram of body weight in a human subject 6 ≥ 1.0 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 1.25 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 1.5 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 1.75 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 2.0 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 2.25 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 2.5 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 2.75 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 3.0 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 3.25 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 3.5 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 3.75 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 4.0 x 10 HSPCs per kilogram of body weight in a human subject 7≥ 4.25 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 4.5 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 4.75 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 5.0 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 5.25 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 5.5 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 5.75 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 6.0 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 6.25 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 6.5 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 6.75 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 7.0 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 7.25 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 7.5 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 7.75 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 8.0 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 8.25 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 8.5 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 8.75 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 9.0 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 9.25 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 9.5 x 10 HSPCs per kilogram of body weight in a human subject 7≥ 9.75 x 10 HSPCs per kilogram of body weight in a human subject 7 ≥ 1.0 x 10 HSPCs per kilogram of body weight in a human subject 8 ≥ 1.25 x 10 HSPCs per kilogram of body weight in a human subject 8 ≥ 1.5 x 10 HSPCs per kilogram of body weight in a human subject 8 ≥ 1.75 x 10 HSPCs per kilogram of body weight in a human subject 8 ≥ 2.0 x 10 HSPCs per kilogram of body weight in a human subject 8 ≥ 2.25 x 10 HSPCs per kilogram of body weight in a human subject 8 ≥ 2.5 x 10 HSPCs per kilogram of body weight in a human subject 8 ≥ 2.75 x 10 HSPCs per kilogram of body weight in a human subject 8 ≥ 3.0 x 10 HSPCs per kilogram of body weight in a human subject 8 ≥ 3.25 x 10 HSPCs per kilogram of body weight in a human subject 8 ≥ 3.5 x 10 HSPCs per kilogram of body weight in a human subject 8 ≥ 3.75 x 10 HSPCs per kilogram of body weight in a human subject 8 ≥ 4.0 x 10 HSPCs per kilogram of body weight in a human subject 8 ≥ 4.25 x 10 HSPCs per kilogram of body weight in a human subject 8 ≥ 4.5 x 10 HSPCs per kilogram of body weight in a human subject 8 ≥ 4.75 x 10 HSPCs per kilogram of body weight in a human subject 8 HSPCs or approximately 5.0 x 10 per kilogram of body weight in a human subject 8 In some embodiments, the body weight of the human subject is the actual body weight of the human subject. In other embodiments, the body weight of the human subject is the ideal actual body weight of the human subject.

[0080] In some embodiments, CD45 +The first population of cells comprises at least about 0.5% granulocytes, at least about 1% granulocytes, up to about 5% granulocytes, up to about 3% granulocytes, up to about 3% monocytes, up to about 2% monocytes, up to about 0.5% lymphocytes, up to about 2% lymphocytes, at least about 15% granulocytes, at least about 20% granulocytes, up to about 35% granulocytes, up to about 30% granulocytes, up to about 25% granulocytes, at least about 15% monocytes, at least about 20% monocytes, up to about 35% monocytes, up to about 30% monocytes, up to about 25% monocytes, at least about 0.5% NK cells, and / or at least about 2% NK cells. In various embodiments, CD45 + A first population of cells, a Treg-enriched cell population, and CD45 + The third population of cells is obtained from a single donor. + a first population of cells, a Treg-enriched cell population, and / or CD45 + The third population of cells is allogeneic to the human subject. + a first population of cells, a Treg-enriched cell population, and / or CD45 + The third population of cells is obtained from an HLA-matched donor to the human subject. + a first population of cells, a Treg-enriched cell population, and / or CD45 + The third population of cells is obtained from an HLA-mismatched donor to the human subject. + a first population of cells, a Treg-enriched cell population, and / or CD45 + A third population of cells is obtained from a donor that is haploidentical to the human subject.

[0081] In some embodiments, Tregs are CD4 + CD25 + CD127 dim or CD4 + FOXP3 + In some embodiments, Tregs are CD4 + CD25+ CD127 dim and FOXP3 + In some instances, the Treg-enriched cell population is CD45 + cells, e.g., CD45 + More than about 90% of the cells are Tregs. + The second population of cells comprises one or more doses of Tregs. In various embodiments, the cell population is enriched for Tregs or comprises at least one dose of CD45 Tregs. + The second population of cells is approximately 1.0 x 10 per kilogram of body weight of a human subject. 5 ~Approx. 1.0×10 8 Tregs, approximately 1.0 x 10 per kilogram of body weight in a human subject 5 ~Approx. 2.0×10 7 Tregs, approximately 1.0 x 10 per kilogram of body weight in a human subject 5 ~Approx. 1.0×10 7 Tregs, or approximately 5.0 x 10 per kilogram of body weight in a human subject 5 ~Approx. 4.0×10 6 In some embodiments, the cell population enriched for Tregs or the CD45 + The second population of cells is approximately 1.0 x 10 per kilogram of body weight of a human subject. 5 ≥ 1.5 x 10 Tregs per kilogram of body weight in a human subject 5 ≥ 2.0 × 10 Tregs per kilogram of body weight in a human subject 5 ≥ 2.5 x 10 Tregs per kilogram of body weight in a human subject 5 ≥ 3.0 × 10 Tregs per kilogram of body weight in a human subject 5 ≥ 3.5 × 10 Tregs per kilogram of body weight in a human subject 5 ≥ 4.0 × 10 Tregs per kilogram of body weight in a human subject 5 ≥ 4.5 x 10 Tregs per kilogram of body weight in a human subject 5 ≥ 5.0 × 10 Tregs per kilogram of body weight in a human subject 5≥ 5.5 x 10 Tregs per kilogram of body weight in a human subject 5 ≥ 6.0 × 10 Tregs per kilogram of body weight in a human subject 5 ≥ 6.5 × 10 Tregs per kilogram of body weight in a human subject 5 ≥ 7.0 × 10 Tregs per kilogram of body weight in a human subject 5 ≥ 7.5 x 10 Tregs per kilogram of body weight in a human subject 5 ≥ 8.0 × 10 Tregs per kilogram of body weight in a human subject 5 ≥ 8.5 × 10 Tregs per kilogram of body weight in a human subject 5 ≥ 9.0 × 10 Tregs per kilogram of body weight in a human subject 5 ≥ 9.5 × 10 Tregs per kilogram of body weight in a human subject 5 ≥ 1.0 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 1.25 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 1.5 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 1.75 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 2.0 × 10 Tregs per kilogram of body weight in a human subject 6 ≥ 2.25 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 2.5 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 2.75 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 3.0 × 10 Tregs per kilogram of body weight in a human subject 6 ≥ 3.25 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 3.5 × 10 Tregs per kilogram of body weight in a human subject 6 ≥ 3.75 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 4.0 × 10 Tregs per kilogram of body weight in a human subject 6≥ 4.25 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 4.5 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 4.75 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 5.0 × 10 Tregs per kilogram of body weight in a human subject 6 ≥ 5.25 × 10 Tregs per kilogram of body weight in a human subject 6 ≥ 5.5 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 5.75 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 6.0 × 10 Tregs per kilogram of body weight in a human subject 6 ≥ 6.25 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 6.5 × 10 Tregs per kilogram of body weight in a human subject 6 ≥ 6.75 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 7.0 × 10 Tregs per kilogram of body weight in a human subject 6 ≥ 7.25 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 7.5 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 7.75 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 8.0 × 10 Tregs per kilogram of body weight in a human subject 6 ≥ 8.25 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 8.5 × 10 Tregs per kilogram of body weight in a human subject 6 ≥ 8.75 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 9.0 × 10 Tregs per kilogram of body weight in a human subject 6 ≥ 9.25 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 9.5 × 10 Tregs per kilogram of body weight in a human subject 6≥ 9.75 x 10 Tregs per kilogram of body weight in a human subject 6 ≥ 1.0 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 1.25 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 1.5 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 1.75 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 2.0 × 10 Tregs per kilogram of body weight in a human subject 7 ≥ 2.25 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 2.5 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 2.75 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 3.0 × 10 Tregs per kilogram of body weight in a human subject 7 ≥ 3.25 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 3.5 × 10 Tregs per kilogram of body weight in a human subject 7 ≥ 3.75 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 4.0 × 10 Tregs per kilogram of body weight in a human subject 7 ≥ 4.25 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 4.5 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 4.75 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 5.0 × 10 Tregs per kilogram of body weight in a human subject 7 ≥ 5.25 × 10 Tregs per kilogram of body weight in a human subject 7 ≥ 5.5 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 5.75 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 6.0 × 10 Tregs per kilogram of body weight in a human subject 7≥ 6.25 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 6.5 × 10 Tregs per kilogram of body weight in a human subject 7 ≥ 6.75 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 7.0 × 10 Tregs per kilogram of body weight in a human subject 7 ≥ 7.25 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 7.5 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 7.75 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 8.0 × 10 Tregs per kilogram of body weight in a human subject 7 ≥ 8.25 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 8.5 × 10 Tregs per kilogram of body weight in a human subject 7 ≥ 8.75 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 9.0 × 10 Tregs per kilogram of body weight in a human subject 7 ≥ 9.25 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 9.5 × 10 Tregs per kilogram of body weight in a human subject 7 ≥ 9.75 x 10 Tregs per kilogram of body weight in a human subject 7 ≥ 1.0 x 10 Tregs per kilogram of body weight in a human subject 8 In some embodiments, the body weight of the human subject is the actual body weight of the human subject. In other embodiments, the body weight of the human subject is the ideal actual body weight of the human subject.

[0082] In some embodiments, CD45 + In another embodiment, the third population of cells comprises Tcon. + In some embodiments, the third population comprises at least one dose of Tcon. +The third population of cells is approximately 1.0 x 10 per kilogram of body weight of a human subject. 5 ~Approx. 1.0×10 8 Tcon, approximately 1.0 x 10 per kilogram of body weight in a human subject 5 ~Approx. 4.0×10 7 Tcon, approximately 1.0 x 10 per kilogram of body weight in a human subject 5 ~Approx. 1.0×10 7 Tcon, or approximately 5.0 x 10 per kilogram of body weight in a human subject 5 ~Approx. 5×10 6 In some embodiments, a CD45 antibody containing Tcon or at least one dose of Tcon is used. + The third population of cells is approximately 1.0 x 10 per kilogram of body weight of a human subject. 5 Tcon, approximately 1.5 x 10 per kilogram of body weight in a human subject 5 Tcon, approximately 2.0 x 10 per kilogram of body weight in a human subject 5 Tcon, approximately 2.5 x 10 per kilogram of body weight in a human subject 5 Tcon, approximately 3.0 x 10 per kilogram of body weight in a human subject 5 Tcon, approximately 3.5 x 10 per kilogram of body weight in a human subject 5 Tcon, approximately 4.0 x 10 per kilogram of body weight in human subjects 5 Tcon, approximately 4.5 x 10 per kilogram of body weight in a human subject 5 Tcon, approximately 5.0 x 10 per kilogram of body weight in a human subject 5 Tcon, approximately 5.5 x 10 per kilogram of body weight in a human subject 5 Tcon, approximately 6.0 x 10 per kilogram of body weight in a human subject 5 Tcon, approximately 6.5 x 10 per kilogram of body weight in a human subject 5 Tcon, approximately 7.0 x 10 per kilogram of body weight in human subjects 5 Tcon, approximately 7.5 x 10 per kilogram of body weight in a human subject 5 Tcon, approximately 8.0 x 10 per kilogram of body weight in human subjects 5Tcon, approximately 8.5 x 10 per kilogram of body weight in a human subject 5 Tcon, approximately 9.0 x 10 per kilogram of body weight in a human subject 5 Tcon, approximately 9.5 x 10 per kilogram of body weight in a human subject 5 Tcon, approximately 1.0 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 1.25 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 1.5 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 1.75 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 2.0 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 2.25 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 2.5 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 2.75 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 3.0 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 3.25 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 3.5 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 3.75 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 4.0 x 10 per kilogram of body weight in human subjects 6 Tcon, approximately 4.25 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 4.5 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 4.75 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 5.0 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 5.25 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 5.5 x 10 per kilogram of body weight in a human subject 6Tcon, approximately 5.75 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 6.0 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 6.25 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 6.5 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 6.75 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 7.0 x 10 per kilogram of body weight in human subjects 6 Tcon, approximately 7.25 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 7.5 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 7.75 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 8.0 x 10 per kilogram of body weight in human subjects 6 Tcon, approximately 8.25 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 8.5 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 8.75 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 9.0 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 9.25 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 9.5 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 9.75 x 10 per kilogram of body weight in a human subject 6 Tcon, approximately 1.0 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 1.25 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 1.5 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 1.75 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 2.0 x 10 per kilogram of body weight in a human subject 7Tcon, approximately 2.25 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 2.5 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 2.75 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 3.0 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 3.25 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 3.5 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 3.75 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 4.0 x 10 per kilogram of body weight in human subjects 7 Tcon, approximately 4.25 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 4.5 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 4.75 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 5.0 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 5.25 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 5.5 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 5.75 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 6.0 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 6.25 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 6.5 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 6.75 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 7.0 x 10 per kilogram of body weight in human subjects 7 Tcon, approximately 7.25 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 7.5 x 10 per kilogram of body weight in a human subject 7Tcon, approximately 7.75 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 8.0 x 10 per kilogram of body weight in human subjects 7 Tcon, approximately 8.25 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 8.5 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 8.75 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 9.0 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 9.25 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 9.5 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 9.75 x 10 per kilogram of body weight in a human subject 7 Tcon, approximately 1.0 x 10 per kilogram of body weight in a human subject 8 In some embodiments, the body weight of the human subject is the actual body weight of the human subject. In other embodiments, the body weight of the human subject is the ideal actual body weight of the human subject.

[0083] In some embodiments, CD45 + A third population of cells contains at least about 0.1% CD34 + cells, or approximately 0.2% to approximately 20% CD34 + In various embodiments, the CD45 + A third population of cells is memory T cells (Tmem), e.g., CD3 + CD45RA- CD45RO + In some embodiments, the population of Tmems is about 3 x 10 per kilogram of actual or ideal body weight of a human subject. 5 In embodiments, the population of Tmems comprises more than about 3 x 10 per kilogram of actual or ideal body weight of a human subject. 5 ~Approx. 1×10 9 In various embodiments, the Tmem comprises CD45+ A third population of cells is invariant natural killer T cells (iNKT), e.g., CD3 + Vα24Jα18 + In some embodiments, the population of iNKT is about 5 x 10 per kilogram of actual or ideal body weight of a human subject. 2 In embodiments, the population of iNKT comprises more than about 5×10 iNKT per kilogram of actual or ideal body weight of a human subject. 2 ~Approx. 1×10 7 In some embodiments, the iNKT + The third population of cells is co-cultured with donor cancer antigens / peptides and / or antigen-presenting cells.

[0084] In some embodiments, the cell populations of the present disclosure, e.g., CD45 + The first population of cells, CD45 + A second population of cells, and / or CD45 + The third population of cells may include one or more cells comprising one or more chimeric receptors. In some embodiments, the one or more cells may be T cells, NK cells, CTLs, Tregs, and / or NKT cells. In some embodiments, the one or more chimeric receptors comprise one or more extracellular antigen-binding domains that bind to one or more cancer-associated antigens. Any suitable cancer-associated antigen known in the art may be targeted. In some embodiments, the chimeric receptor is a chimeric antigen receptor or a T cell receptor.

[0085] Provided herein are compositions, multicomponent medical treatments, multicomponent cell therapy products, cell populations, solutions, formulations, kits, and / or methods for improved hematopoietic stem cell transplantation (HCT), e.g., allogeneic hematopoietic stem cell transplantation (alloHSCT). The compositions, multicomponent medical treatments, multicomponent cell therapy products, cell populations, solutions, formulations, kits, and / or methods disclosed herein may include one or more cell populations that can be administered in combination with a GVHD prophylactic agent to achieve a positive clinical outcome. The cell populations may include one or more cell types, such as hematopoietic stem progenitor cells (HSPCs), conventional T cells (Tcon), regulatory T cells (Treg), invariant natural killer T cells (iNKT), memory T cells (Tmem), and combinations thereof.

[0086] The present disclosure provides parameters for cell populations and methods for administering cell populations that can contribute to a good clinical outcome in HCT recipient subjects. Without wishing to be bound by theory, parameters that can contribute to a good clinical outcome in HCT recipient subjects include, for example, the co-administration of a GVHD prophylactic agent (e.g., tacrolimus) described herein, the populations administered, the order and timing of administration of different populations, purity standards for the populations, methods for obtaining the populations, methods for handling or selecting the populations, dosages of the populations administered, methods for obtaining the populations, and combinations thereof.

[0087] In various embodiments, administration includes administering to a human subject a CD45 + A first population of cells, a cell population enriched for Tregs (e.g., CD45 + a second population of cells), and CD45 + Injecting a third population of cells is included.

[0088] In some embodiments, the cell population is enriched for Tregs, or CD45 + The second population of cells (disclosed herein) is CD45 +In some embodiments, the Treg-enriched cell population, or the CD45 + The second population of cells expresses CD45 + About 5 minutes, about 6 minutes, about 7 minutes, about 8 minutes, about 9 minutes, about 10 minutes, about 11 minutes, about 12 minutes, about 13 minutes, about 14 minutes, about 15 minutes, about 16 minutes, about 17 minutes, about 18 minutes, about 19 minutes, about 20 minutes, about 21 minutes, about 22 minutes, about 23 minutes, about 24 minutes, about 25 minutes, about 26 minutes, about 27 minutes, about 28 minutes, about 29 minutes, about 30 minutes, about 31 minutes, about 32 minutes after administration of the first population of cells , after about 33 minutes, after about 34 minutes, after about 35 minutes, after about 36 minutes, after about 37 minutes, after about 38 minutes, after about 39 minutes, after about 40 minutes, after about 41 minutes, after about 42 minutes, after about 43 minutes, after about 44 minutes, after about 45 minutes, after about 46 minutes, after about 47 minutes, after about 48 minutes, after about 49 minutes, after about 50 minutes, after about 51 minutes, after about 52 minutes, after about 53 minutes, after about 54 minutes, after about 55 minutes, after about 56 minutes, after about 57 minutes, after about 58 minutes, after about 59 minutes, after about 1 hour, after about 10:00 After about 1.20 hours, about 1.30 hours, about 1.40 hours, about 1.50 hours, about 1.60 hours, about 1.70 hours, about 1.80 hours, about 1.90 hours, about 2 hours, about 2.10 hours, about 2.20 hours, about 2.30 hours, about 2.40 hours, about 2.50 hours, about 2.60 hours, about 2.70 hours, about 2.80 hours, about 2.90 hours, about 3 hours, about 3.10 hours The dose is administered about 3.20 hours, about 3.30 hours, about 3.40 hours, about 3.50 hours, about 3.60 hours, about 3.70 hours, about 3.80 hours, about 3.90 hours, about 4 hours, about 4.10 hours, about 4.20 hours, about 4.30 hours, about 4.40 hours, about 4.50 hours, about 4.60 hours, about 4.70 hours, about 4.80 hours, about 4.90 hours, or about 5 hours later.

[0089] In some embodiments, CD45 + A third population of cells (disclosed herein) expresses CD45 + at least about 12 hours after the first population of cells (disclosed herein), + The third population of cells is CD45 +about 24 to about 96 hours after the first population of cells, + The third population of cells is CD45 + Approximately 36 to 60 hours after the first population of cells is administered, and CD45 + The third population of cells is administered at least about 12 hours after the Treg-enriched cell population (disclosed herein) and expresses CD45 + The third population of cells is administered about 24 to about 96 hours after the Treg-enriched cell population, and / or is CD45 + The third population of cells is a Treg-enriched cell population or CD45 + The second population of cells is administered about 36 to about 60 hours after the second population of cells.

[0090] In some embodiments, CD45 + A third population of cells (disclosed herein) is isolated CD45 + In some embodiments, the administration of the CD45 + The third population of cells is CD45 +About 12 hours, about 13 hours, about 14 hours, about 15 hours, about 16 hours, about 17 hours, about 18 hours, about 19 hours, about 20 hours, about 21 hours, about 22 hours, about 23 hours, about 24 hours, about 25 hours, about 26 hours, about 27 hours, about 28 hours, about 29 hours, about 30 hours, about 31 hours, about 32 hours, about 33 hours, about 34 hours, about 35 hours, about 36 hours, about 37 hours, about 38 hours after administration of the first population of cells After about 39 hours, about 40 hours, about 41 hours, about 42 hours, about 43 hours, about 44 hours, about 45 hours, about 46 hours, about 47 hours, about 48 hours, about 49 hours, about 50 hours, about 51 hours, about 52 hours, about 53 hours, about 54 hours, about 55 hours, about 56 hours, about 57 hours, about 58 hours, about 59 hours, about 60 hours, about 61 hours, about 62 hours, about 63 hours, about 64 hours, about 65 hours, about 66 hours, about 67 hours Hours, about 68 hours, about 69 hours, about 70 hours, about 71 hours, about 72 hours, about 73 hours, about 74 hours, about 75 hours, about 76 hours, about 77 hours, about 78 hours, about 79 hours, about 80 hours, about 81 hours, about 82 hours, about 83 hours, about 84 hours, about 85 hours, about 86 hours, about 87 hours, about 88 hours, about 89 hours, about 90 hours, about 91 hours, about 92 hours, about 93 hours, about 94 hours, about 95 hours, about The dose is administered after 96 hours, about 97 hours, about 98 hours, about 99 hours, about 100 hours, about 101 hours, about 102 hours, about 103 hours, about 104 hours, about 105 hours, about 106 hours, about 107 hours, about 108 hours, about 109 hours, about 110 hours, about 111 hours, about 112 hours, about 113 hours, about 114 hours, about 115 hours, about 116 hours, about 117 hours, about 118 hours, about 119 hours, or about 120 hours.

[0091] In some embodiments, CD45 + A third population of cells (disclosed herein) is a cell population enriched for Tregs or CD45 + In some embodiments, the CD45 +The third population of cells is a Treg-enriched cell population or CD45 + About 12 hours, about 13 hours, about 14 hours, about 15 hours, about 16 hours, about 17 hours, about 18 hours, about 19 hours, about 20 hours, about 21 hours, about 22 hours, about 23 hours, about 24 hours, about 25 hours, about 26 hours, about 27 hours, about 28 hours, about 29 hours, about 30 hours, about 31 hours, about 32 hours, about 33 hours, about 34 hours, about 35 hours, about 36 hours, about 37 hours, about 38 hours after administration of the second population of cells After about 39 hours, about 40 hours, about 41 hours, about 42 hours, about 43 hours, about 44 hours, about 45 hours, about 46 hours, about 47 hours, about 48 hours, about 49 hours, about 50 hours, about 51 hours, about 52 hours, about 53 hours, about 54 hours, about 55 hours, about 56 hours, about 57 hours, about 58 hours, about 59 hours, about 60 hours, about 61 hours, about 62 hours, about 63 hours, about 64 hours, about 65 hours, about 66 hours, about 67 hours Hours, about 68 hours, about 69 hours, about 70 hours, about 71 hours, about 72 hours, about 73 hours, about 74 hours, about 75 hours, about 76 hours, about 77 hours, about 78 hours, about 79 hours, about 80 hours, about 81 hours, about 82 hours, about 83 hours, about 84 hours, about 85 hours, about 86 hours, about 87 hours, about 88 hours, about 89 hours, about 90 hours, about 91 hours, about 92 hours, about 93 hours, about 94 hours, about 95 hours, about The dose is administered after 96 hours, about 97 hours, about 98 hours, about 99 hours, about 100 hours, about 101 hours, about 102 hours, about 103 hours, about 104 hours, about 105 hours, about 106 hours, about 107 hours, about 108 hours, about 109 hours, about 110 hours, about 111 hours, about 112 hours, about 113 hours, about 114 hours, about 115 hours, about 116 hours, about 117 hours, about 118 hours, about 119 hours, or about 120 hours.

[0092] HSPCs can have extensive self-renewal potential and the ability to differentiate into specialized cell types, for example, the ability to reconstitute all hematopoietic cell lineages. HSPCs can undergo asynchronous replication, in which two daughter cells are produced with different phenotypes. HSPCs can exist in a mitotically quiescent form. HSPCs can be derived from bone marrow, peripheral blood, and / or umbilical cord blood.

[0093] Subsets of immune cells, such as conventional T cells (Tcon), regulatory T cells (Treg), invariant natural killer T cells (iNKT), and memory T cells (Tmem), may contribute to aspects of GVHD after HCT, for example, GVHD immune responses, immune reconstitution, susceptibility to infection, and patient survival.

[0094] GVHD is primarily mediated by donor T cells, which can recognize recipient antigens and initiate an inflammatory response. Depleting T cells (TCD) from a cell population intended for transplant into a subject can reduce the likelihood of acute and / or chronic GVHD. T cells can be depleted using methods including, but not limited to, physical adsorption of T cells to protein ligands such as lectins, immunodepletion with T cell-specific antibodies, and immunoaffinity techniques (e.g., the use of T cell- or lymphocyte-specific antibodies in immunoadsorption columns, magnetic-activated cell sorting (MACS), or fluorescence-activated cell sorting (FACS)). Application of TCD techniques to donor grafts can result in, for example, a 10-10 reduction in T cells. 5 TCD can result in a 1:1 reduction in T cell depletion and a reduced incidence of GVHD. However, TCD may also result in an increased rate of cancer recurrence due to the lack of T cells and the potential for a reduced immune response in graft-versus-tumor (GVT). Furthermore, TCD can impair immune recovery and increase susceptibility to infection.

[0095] Both GVT and GVHD are largely mediated by conventional T cells (Tcon), which initiate an immune response when the T cell receptor recognizes a cognate antigen (a tumor antigen for GVT and a non-tumor recipient antigen for GVHD). Tcon can, for example, cause GVT, GVHD, or a combination of these. In some embodiments, administering Tcon after Treg administration may enhance GVT immunity and / or reduce susceptibility to infection.

[0096] Tcon, in a broad sense, is a subset of all CD3 + T cells may refer to cells expressing CD3 and CD4, or cells expressing CD3 and CD8, cells expressing medium to high levels of CD127, cells expressing CD3 and medium to high levels of CD127, cells expressing CD3, cells expressing medium to high levels of CD127, and cells expressing CD4 or CD8. In some embodiments, Tcon do not express the Vα24Jα18 TCR. Tcon and regulatory T cells ("Treg") may be cell populations that are not mutually exclusive. In some embodiments, Tcon and Treg are mutually exclusive cell populations.

[0097] Regulatory T cells ("Tregs") are a specialized subpopulation of T cells that negatively regulate (e.g., suppress) immune system activation, thereby promoting immune tolerance. Without wishing to be bound by theory, the Treg-enriched cell populations of the present disclosure contribute to positive clinical outcomes, for example, by reducing the incidence and / or severity of GVHD in transplant recipient subjects and / or by improving immune reconstitution in transplant recipients. CD45 Tregs containing at least HSPCs +Administration of a Treg-enriched cell population together with a population of cells can, for example, promote the preservation of graft-versus-tumor (GVT) and promote a reduction in the incidence and / or severity of GVHD. Without wishing to be bound by theory, for example, administration of a Treg-enriched cell population can prevent GVHD and promote the preservation of Treg-enriched cell populations, including at least Tregs, compared to alternative hematopoietic stem cell transplant (HCT) methods, i.e., methods separate from the compositions, multi-component medical procedures, multi-component cell therapy products, cell populations, solutions, formulations, kits, and / or methods disclosed herein. + The administration of a third population of cells can enhance the GVHD effect. In some embodiments, administering a Treg-enriched cell population reduces the risk of developing GVHD and enhances the ability to differentiate into CD45 Tregs and CD45 Tregs, compared to alternative HCT methods, i.e., methods separate from the compositions, multi-component medical procedures, multi-component cell therapy products, cell populations, solutions, formulations, kits, and / or methods disclosed herein. + The third population of cells is administered to promote the GVT effect.

[0098] As used herein, an alternative composition lacks one or more cell populations and / or prophylactic agents disclosed herein and / or recited in the claims. For example, the alternative composition lacks one or more of a cell population comprising HSPCs, a cell population comprising Tregs, a cell population comprising Tcon, and a prophylactic agent. In some embodiments, the alternative composition or treatment regimen includes an additional cell population or agent, e.g., an additional or different GVHD prophylactic agent, compared to the composition or treatment regimen of the present disclosure.

[0099] There are many Treg subsets, e.g., TCRαβ + CD4 +Regulatory T cells include natural regulatory T cells (nTreg) and inducible regulatory T cells (iTreg). nTreg can be T cells generated in the thymus and delivered to the periphery as a long-lived lineage of autoantigen-specific lymphocytes. iTreg can be mobilized from circulating lymphocytes and acquire regulatory properties under specific peripheral stimulation conditions. nTreg and iTreg are CD4 + CD25 + Both of these increased CD4 + It can inhibit the proliferation of CD25- T cells. In some embodiments, Tregs are anergic and do not proliferate even when stimulated by TCR. In addition to being CD4 and CD25 positive, Tregs can also be positive for the intracellular marker FOXP3, a transcription factor. Tregs can be identified or selected based on various marker expression profiles. Non-limiting examples of marker expression profiles that can be used to select Tregs include: (1) CD4 + CD25 + CD127 dim (2) CD4 + FOXP3 + , (3) CD3 + CD4 + CD25 + , (4) CD3 + CD4 + CD25 + CD127 dim , (5) CD3 + CD4 + CD25 + CD127 dim FOXP3 + , (6) CD3 + FOXP3 + , (7) CD3 + CD4 + FOXP3 + , (8) CD3 + CD4 + CD25 + FOXP3 + , (9) CD3 + CD25 + FOXP3 + , (10)CD3 + CD25 +CD127 dim , (11)CD4 + CD25 + , (12)CD4 + CD25 + CD127 dim FOXP3 + , (13)FOXP3 + , (14)CD4 + FOXP3 + , (15)CD4 + CD25 + FOXP3 + , (16)CD25 + FOXP3 + , and (17)CD25 + CD127 dim Examples include:

[0100] Selection based on specific expression profiles can be based on extracellular markers, e.g., CD4, without the need for cell permeabilization. + CD25 + CD127 dim This can be achieved by selection based on:

[0101] Cell populations comprising Tregs can, for example, reduce the incidence of transplant rejection, reduce the incidence and / or severity of GVHD, promote hematopoietic reconstitution, promote immune reconstitution, promote mixed chimerism, or a combination thereof.

[0102] The cell populations of the present disclosure can include invariant natural killer T cells (iNKTs). iNKTs are a subclass of CD1d-restricted natural killer T (NKT) cells that express a highly conserved αβ-T cell receptor, including that of the Vα24Jα18 TCRα chain in humans (referred to herein as "Vα24Jα18"). +iNKT cells can be identified by binding to CD1d multimers, such as those loaded with α-galactosylceramide (GalCer), PBS-57, PBS-44, or other natural or synthetic glycolipids. Another method of identification is an antibody or combination of antibodies that specifically recognize the Vα24Jα18 region. Examples include Vα24 antibodies, Jα18 antibodies, or the monoclonal antibody clone 6B11, which specifically binds to a unique region of the Vα24Jα18 TCR and can be used to identify iNKT cells. iNKT cells are identified by binding to CD3 + Vα24Jα18 + may be.

[0103] In some embodiments, iNKT can promote engraftment, promote GVT, reduce the incidence and / or severity of GVHD, reduce susceptibility to cancer recurrence, reduce susceptibility to infection, or a combination thereof. In some embodiments, iNKT promotes the activity of Tregs. In some embodiments, iNKT promotes the activity of HSPCs.

[0104] The cell populations of the present disclosure may include memory T cells (Tmems). Tmems may refer to antigen-experienced T cells that express, for example, the phenotypic markers CD45RO, TCRα, TCRβ, CD3, CD4, CD95, and IL-2Rβ, or the phenotypic markers CD45RO, TCRα, TCRβ, CD3, CD8, CD95, and IL-2Rβ. Tmems confer immunity and can persist in an inactive state for long periods. Tmems can rapidly acquire effector function upon antigen reloading. A population of Tmems may include any combination of the subclasses of central memory T cells and effector memory T cells. In some embodiments, Tmems are CD3 + CD45RA - CD45RO +In various methods, Tmem administered to a subject undergoing HCT can, for example, promote GVT, reduce GVHD, reduce susceptibility to cancer recurrence, reduce susceptibility to infection, or a combination thereof.

[0105] Cell acquisition, treatment, and preparation Certain aspects of the present disclosure relate to methods for preparing the compositions, multi-component medical treatments, multi-component cell therapy products, cell populations, solutions, formulations, and / or kits of the present disclosure.

[0106] In some embodiments, at least one mobilized donated peripheral blood is collected from the donor, or up to two mobilized donated peripheral blood is collected from the donor. In some embodiments, the mobilized donated peripheral blood is a HSPC-mobilized donated peripheral blood apheresis. In some embodiments, prior to donating the peripheral blood, the donor may be vaccinated with tumor antigens and / or pathogens to enhance graft-versus-infection.

[0107] In embodiments, at least one of the mobilized donated peripheral blood cells is CD34 + In some embodiments, the donated peripheral blood is further processed and selected to enrich for CD3 + In some embodiments, at least one of the mobilized donated peripheral blood samples is further processed and selected to enrich for CD34 cells (e.g., Tcon). + The cells are treated and sorted to be enriched for Tregs and / or Tcon CD34 + The processing and selection of cells, Tregs, and Tcon cells may be performed in any order. For example, at least one of the mobilized donated peripheral blood samples may first be purified to identify CD34 + Alternatively, at least one of the mobilized donor peripheral blood samples is first treated and selected to enrich for CD34 cells, then for T cells, and then for Treg cells. +Alternatively, at least one of the mobilized donor peripheral blood samples is first treated and selected to enrich for Tregs and then for CD34 + Alternatively, at least one of the mobilized donor peripheral blood samples is first treated and sorted to enrich for Tregs, then for Tcon, and then for CD34. + Alternatively, at least one of the mobilized donor peripheral blood samples is first treated and sorted to enrich for T cells, and then purified to enrich for CD34 cells. + Alternatively, at least one of the mobilized donor peripheral blood samples is treated and selected to first enrich for T cells, then for T cells, and then for CD34 cells. + In some embodiments, at least one of the mobilized donated peripheral blood cells is treated and sorted to enrich for Tregs first, then CD34 cells, without enrichment for Tcon cells. + In another embodiment, at least one of the mobilized donated peripheral blood cells is initially treated and sorted to enrich for CD34 cells without enrichment for Tcon cells. + The cells are then treated and selected to enrich for Tregs.

[0108] In some embodiments, the time to process and sort one or more of the mobilized donated peripheral blood is less than about 40 hours, the time to process and sort one or more of the mobilized donated peripheral blood is less than about 35 hours, the time to process and sort one or more of the mobilized donated peripheral blood is less than about 30 hours, the time to process and sort one or more of the mobilized donated peripheral blood is less than about 25 hours, the time to process and sort one or more of the mobilized donated peripheral blood is less than about 20 hours, and the time to process and sort one or more of the mobilized donated peripheral blood is less than about 40 hours. The time to process and sort one or more of the mobilized donated peripheral blood is less than about 15 hours, and / or the time to process and sort one or more of the mobilized donated peripheral blood is up to about 35 hours, the time to process and sort one or more of the mobilized donated peripheral blood is up to about 30 hours, the time to process and sort one or more of the mobilized donated peripheral blood is up to about 25 hours, the time to process and sort one or more of the mobilized donated peripheral blood is up to about 20 hours, or the time to process and sort one or more of the mobilized donated peripheral blood is up to about 15 hours.

[0109] In various embodiments, one or more of the mobilized peripheral blood donations are processed and sorted using one or more immunoseparation particles (ISPs). For example, the ISPs include affinity reagents such as immunomagnetic separation particles, which may each be an antibody conjugated to an iron-containing particle, and / or immuno-fluorescent separation particles, which may each be an antibody conjugated to a fluorophore or other fluorescent particle.

[0110] In some embodiments, the affinity reagent comprises a plurality of CD34 reagents (e.g., anti-CD34 antibodies) that bind to one or more CD34 receptors on HSPCs. In some cases, at least a portion of the plurality of ISPs binds to CD34 receptors on HSPCs in the HSPC cell population. +Optionally, the average number of ISPs per HSPC in the HSPC cell population is less than about 20,000, the average number of ISPs per HSPC in the HSPC cell population is about 10,000 or less, and / or the average number of ISPs per HSPC in the HSPC cell population is about 1,000 to about 20,000.

[0111] In some embodiments, the average number of ISPs per HSPC in the HSPC cell population may be about 1,500 to about 20,000. In some embodiments, the average number of ISPs per HSPC in the HSPC cell population may be at least about 1,500. In some embodiments, the average number of ISPs per HSPC in the HSPC cell population may be up to about 20,000. In some embodiments, the average number of ISPs per HSPC in the HSPC cell population is about 1,500 to about 2,000, about 1,500 to about 5,000, about 1,500 to about 6,000, about 1,500 to about 10,000, about 1,500 to about 12,000, about 1,500 to about 15,000, about 1,500 to about 20,000, about 2,000 to about 5,000, about 2,000 to about 6,000, about 2,000 to about 10,000, about 2,000 to about 12,000, about 2,000 to about 15,000, about 2,000 to about 20,000, or about 5,000. It may be 0 to about 6,000, about 5,000 to about 10,000, about 5,000 to about 12,000, about 5,000 to about 15,000, about 5,000 to about 20,000, about 6,000 to about 10,000, about 6,000 to about 12,000, about 6,000 to about 15,000, about 6,000 to about 20,000, about 10,000 to about 12,000, about 10,000 to about 15,000, about 10,000 to about 20,000, about 12,000 to about 15,000, about 12,000 to about 20,000, or about 15,000 to about 20,000. In some embodiments, the average number of ISPs per HSPC in the HSPC cell population may be about 1,500, about 2,000, about 5,000, about 6,000, about 10,000, about 12,000, about 15,000, or about 20,000. In some embodiments, the average number of ISPs per HSPC in the HSPC cell population may be at least 1,500, 2,000, 5,000, 6,000, 10,000, 12,000, 15,000, or 20,000. In some embodiments, the average number of ISPs per HSPC in the HSPC cell population may be up to 1,500, 2,000, 5,000, 6,000, 10,000, 12,000, 15,000, or 20,000.

[0112] In some embodiments, the affinity reagent comprises a plurality of CD25 reagents (e.g., anti-CD25 antibodies) that bind to one or more CD25 receptors on cells. In some cases, at least a portion of the plurality of ISPs binds to CD25 receptors on cells of the Treg cell population. + Optionally, the average number of ISPs per T-reg cell in the Treg population is about 4000 or less, or the average number of ISPs per T-reg cell in the Treg population is about 1500 to about 2500. In some cases, at least a portion of the plurality of ISPs are coupled to CD3 receptors on cells of the heterogeneous cell population. + The Tcon cell population binds to the receptor. Optionally, the average number of ISPs per cell in the heterogeneous T population is less than about 4,000. In some embodiments, the average number of ISPs per Tcon cell in the Tcon cell population may be about 100 to about 1,000. In some embodiments, the average number of ISPs per Tcon cell in the Tcon cell population may be at least about 100. In some embodiments, the average number of ISPs per Tcon cell in the Tcon cell population may be up to about 1,000. In some embodiments, the average number of ISPs per Tcon cell in the Tcon cell population may be about 100 to about 200, about 100 to about 500, about 100 to about 1,000, about 200 to about 500, about 200 to about 1,000, or about 500 to about 1,000. In some embodiments, the average number of ISPs per Tcon cell in the Tcon cell population may be about 100, about 200, about 500, or about 1,000. In some embodiments, the average number of ISPs per Tcon cell in the Tcon cell population may be at least 100, 200, 500, or 1,000. In some embodiments, the average number of ISPs per Tcon cell in the Tcon cell population may be up to 100, 200, 500, or 1,000.

[0113] In some embodiments, the average number of ISPs per Treg cell in a Treg cell population may be about 500 to about 4,000. In some embodiments, the average number of ISPs per Treg cell in a Treg cell population may be at least about 500. In some embodiments, the average number of ISPs per Treg cell in a Treg cell population may be up to about 4,000. In some embodiments, the average number of ISPs per Treg cell in a Treg cell population is about 500 to about 1,000, about 500 to about 1,500, about 500 to about 2,000, about 500 to about 2,500, about 500 to about 3,000, about 500 to about 4,000, about 1,000 to about 1,500, about 1,000 to about 2,000, about 1,000 to about 2,500, about 1,000 to about 3,000, about 500 to about 4,000, about 1,000 to about 1,500, about 1,000 to about 2,000, about 1,000 to about 2,500, or about 1,000 to about 3,000. In some embodiments, the average number of ISPs per Treg cell in the Treg cell population may be about 500, about 1,000, about 1,500, about 2,000, about 2,500, about 3,000, about 4,000, about 1,000 to about 4,000, about 1,500 to about 2,000, about 1,500 to about 2,500, about 1,500 to about 3,000, about 1,500 to about 4,000, about 2,000 to about 2,500, about 2,000 to about 3,000, about 2,000 to about 4,000, about 2,500 to about 3,000, about 2,500 to about 4,000, or about 3,000 to about 4,000. In some embodiments, the average number of ISPs per Treg cell in the Treg cell population may be at least 500, 1,000, 1,500, 2,000, 2,500, 3,000, or 4,000. In some embodiments, the average number of ISPs per Treg cell in the Treg cell population may be up to 500, 1,000, 1,500, 2,000, 2,500, 3,000, or 4,000.

[0114] In some embodiments, the CD25-enriched cell population is further processed by sorting with one or more ISPs comprising immuno-fluorescent separation particles (e.g., antibodies conjugated to fluorophores) specific for CD127 and CD4. In some embodiments, the CD25-enriched Treg population of the present disclosure is sorted using a CD127-specific ISP and a CD4-specific ISP to further enrich for CD25. + CD4 + CD127 dim A Treg population is obtained. In some embodiments, the Treg population is FOXP3 + In some embodiments, CD127 dim The cell population is CD127 - population and / or CD127 low It may include a population.

[0115] In some embodiments, the affinity reagent comprises a plurality of CD3 reagents (e.g., anti-CD3 antibodies) that bind to one or more CD3 receptors on cells. In some cases, at least a portion of the plurality of ISPs bind to CD3 receptors on cells of the Treg cell population. + Optionally, the average number of ISPs per T cell in the Tcon population is about 4000 or less, or the average number of ISPs per T cell in the Tcon population is about 1500 to about 2500. In some cases, at least a portion of the plurality of ISPs are coupled to CD3 receptors on cells of the heterogeneous cell population. + Optionally, the average number of ISPs per cell in the heterogeneous T population is less than about 4,000.

[0116] In various embodiments, the mobilized donated peripheral blood cells are CD45 + The first population of cells is selected to contain up to about 10% granulocytes. In some instances, the mobilized donated peripheral blood cells are CD45 + The first population of cells is selected to contain a maximum of about 7% granule cells.

[0117] In some embodiments, the mobilized donor-donated peripheral blood cells are CD45 +The first population of cells is selected to contain up to about 4% monocytes. In some instances, the mobilized donor-donated peripheral blood cells are CD45 + The first population of cells is selected to contain at least about 0.1% monocytes.

[0118] In embodiments, mobilized donor-provided peripheral blood cells are sorted so that the Treg-enriched population contains up to about 10% CD25- cells.

[0119] In some embodiments, the cell populations of the present disclosure are obtained from whole blood. The cell populations of the present disclosure can be obtained from peripheral blood apheresis products, such as mobilized peripheral blood apheresis products mobilized by administering GCSF, GM-CSF, Mozovir® (plelixafor), and combinations thereof to a donor. The cell populations of the present disclosure can be obtained from at least one apheresis product, two apheresis products, three apheresis products, four apheresis products, five apheresis products, six apheresis products, or more. In some embodiments, the cell populations of the present disclosure are obtained from one apheresis product. In some embodiments, the cell populations of the present disclosure are obtained from two apheresis products. In some embodiments, the cell populations of the present disclosure are obtained from an apheresis product from one donor and an apheresis product from at least a second donor.

[0120] In some embodiments, the cell populations of the present disclosure are obtained from a bone marrow sample. In some embodiments, the cell populations of the present disclosure can be obtained from a bone marrow sample, such as a mobilized bone marrow sample mobilized by administering GCSF, GM-CSF, Mozovir® (plelixafor), and combinations thereof to a donor.

[0121] In some embodiments, the cell populations of the present disclosure are obtained from umbilical cord blood.

[0122] The cell populations of the present disclosure can be purified by selection from a population of cells, such as peripheral blood or peripheral blood apheresis products. Cell population selection methods can include methods involving positive or negative selection of the desired cell population. Cell population selection methods can include affinity reagents, including, but not limited to, antibodies, full-length antibodies, antibody fragments, naturally occurring antibodies, synthetic antibodies, engineered antibodies, full-length affibodies, affibody fragments, full-length affilins, affilin fragments, full-length anticalins, anticalin fragments, full-length avimers, avimer fragments, full-length DARPins, DARPin fragments, full-length finomers, finomer fragments, full-length kunitz domain peptides, kunitz domain peptide fragments, full-length monobodies, monobody fragments, peptides, or polyamino acids. In some embodiments, the affinity reagent is directly conjugated to the detection reagent and / or purification reagent. In some cases, the detection reagent and purification reagent are the same. In some cases, the detection reagent and purification reagent are different. For example, the detection reagent and / or purification reagent is a fluorescent, magnetic, etc. reagent. In some cases, the detection reagent and / or purification reagent is a magnetic particle for column purification. For example, magnetic column purification can be performed using the columns, antibodies, buffers, preparation materials and reagents of the Miltenyi system (CliniMAC).

[0123] In various embodiments, at least one of the cell populations has a plurality of immunoseparation particles (ISPs) bound to receptors on cells of the cell population. In some cases, the plurality of ISPs are immunomagnetic separation particles. In some embodiments, the plurality of ISPs comprises antibodies conjugated to iron-containing particles. In some cases, at least a portion of the plurality of ISPs is conjugated to CD34 of HSPCs in the HSPC cell population. +optionally, the average number of ISPs per HSPC in the HSPC cell population is less than about 6,000, the average number of ISPs per HSPC in the HSPC cell population is equal to or less than about 3,000, and / or the average number of ISPs per HSPC in the HSPC cell population is between about 1700 and about 3,000. In some cases, at least a portion of the plurality of ISPs are coupled to CD25 receptors of cells of the Treg cell population. + Optionally, the average number of ISPs per T-reg cell in the Treg population is less than or equal to about 1700, or the average number of ISPs per T-reg cell in the Treg population is between about 1400 and about 1700. In some cases, at least a portion of the plurality of ISPs are coupled to the CD3 receptor of cells in the heterogeneous cell population. + Bind to the receptor; if necessary, the average number of ISPs per cell in a heterogeneous population is less than approximately 1,000.

[0124] Affinity reagents include immunoaffinity reagents that utilize the binding specificity of antibodies or their fragments or derivatives to positively or negatively select a cell population of interest. Cell population selection methods can include affinity agents and columns, such as magnetic-activated cell sorting (MACS) using specific antibodies and microbeads. Cell population selection methods can include fluorescence-activated cell sorting (FACS), in which a cell population is sorted based on its staining profile using one or more fluorescently conjugated antibodies. Cell population selection methods can include physical adsorption, e.g., physical adsorption of T cells to a protein ligand such as a lectin.

[0125] HSPCs can be obtained by harvesting from bone marrow or peripheral blood. Bone marrow can be aspirated from the posterior or anterior iliac crest while the donor is under local or general anesthesia. HSPCs can also be obtained by harvesting from peripheral blood, for example, by peripheral blood apheresis. The number of recovered stem cells can be increased by treating the donor with a mobilizing agent, i.e., an agent that mobilizes stem cells from the bone marrow to the peripheral blood. Non-limiting examples of mobilizing agents include granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), stem cell factor (SCF), SDF-1 antagonists, CXCR4 antagonists (e.g., POL6326, BKT-140, TG-0054, NOX-A12), Mozovir® (plerixafor), CXCR2 ligands (e.g., GROβ), sphingosine-1-phosphatase (S1P) agonists (e.g., SEW2871), VCAM / VLA-4 inhibitors (e.g., BIO5192), proteosome inhibitors (e.g., bortezomib), parathyroid hormone, hypoxia-inducible factor (HIF) stabilizers (e.g., FG-4497), and combinations thereof. Techniques for mobilizing stem cells into peripheral blood can include administering a mobilization agent to the donor at a dose of 10-40 μg / kg / day. The mobilization agent can be administered to the donor in 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 doses. The apheresis product can be isolated from the donor approximately 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 26, 28, or 30 hours after administration of the mobilization agent.

[0126] CD45 of the present disclosure + The cell population can include HSPC. HSPC can be selected based on CD34 expression. For example, the HSPC of the present disclosure can be selected using anti-CD34 antibody as part of a magnetic activated cell sorting (MACS) or fluorescence activated cell sorting (FACS) system.

[0127] CD45+ The number of HSPCs in a population of cells can be determined by, for example, flow cytometry using CD34 + It can be determined by quantifying cells. In some embodiments, the dose calculation is adjusted based on a measure of cell viability, for example, the viability determined by flow cytometry using propidium iodide or 7-AAD, or by trypan blue exclusion.

[0128] The cell populations of the present disclosure include Treg (e.g., CD45 + A second population of cells may be enriched for Tregs. Tregs can be selected based on the expression of markers including CD3, CD4, CD25, CD127, FOXP3, and combinations thereof.

[0129] Tregs can be selected using magnetic activated cell sorting (MACS). Tregs can be selected using fluorescence activated cell sorting (FACS). Tregs can be selected using multiple procedures, e.g., multiple MACS selections, multiple FACS selections, or a combination of MACS and FACS selections. For example, a first selection can be performed for CD25 expression, e.g., using MACS to identify CD25 from a hematopoietic cell sample. + The second selection is to isolate CD25 + This may be performed by contacting the cells with antibodies specific for CD4 and CD127, in which case FACS may be used to identify CD4 + CD127 dim and isolating cells that are

[0130] Tregs may be isolated from whole blood. Tregs may be isolated from peripheral blood apheresis products. Tregs may be isolated from cell populations that have been previously enriched and / or depleted of one or more other cell types, for example, CD34 + In some embodiments, Tregs may be isolated from a cell-depleted cell population. + It is isolated from the flow-through fraction of MACS selection.

[0131] The number of Tregs in a cell population can be determined, for example, by flow cytometry, and Tregs can be, for example, CD4 + CD25 + CD127 dim or CD4 + FOXP3 + Dose calculations can be adjusted based on measures of cell viability, such as viability determined by flow cytometry using propidium iodide or 7-AAD, or by trypan blue dye exclusion.

[0132] In some embodiments, CD45 + The third population of cells may include a population of Tcon. + A third population of cells can be sourced from peripheral blood. + The third population of cells can be sourced from a peripheral blood apheresis product.

[0133] In some embodiments, no selection step is performed and the CD45 + The cell population can be directly sourced from peripheral blood or an aliquot of apheresis product. In some embodiments, the cell population can be enriched for Tcon by sorting based on the expression of various markers, for example, using MACS, FACS, or a combination thereof. In some embodiments, CD45 + The third population of cells is CD3 + In some embodiments, CD45 + The third population of cells is CD4 + cells and CD8 + In some embodiments, cells can be enriched by sorting. +A third population of cells can be enriched by negative selection, where non-Tcon cells are removed, for example, by depleting cells expressing CD34, CD19, CD25, or a combination thereof, by MACS.

[0134] CD45 + The number of Tcon present in the third population of cells can be determined by, for example, CD3 + The CD3 in the aliquot can be quantified by quantifying the cells. + The number of cells can be determined, and the volume containing the appropriate dose of CD3 cells to be administered to the recipient can be determined. The dose calculation can be adjusted based on a measure of cell viability, such as viability determined by flow cytometry using propidium iodide or 7-AAD, or trypan blue dye exclusion.

[0135] The apheresis product of the present disclosure comprises two parts: CD45 containing at least Tcon; + One portion is used to obtain a third population of cells, as well as CD45 HSPCs. + The CD34+ Treg-enriched population can be isolated and purified from the other portion of the cell line. + Cells are isolated and purified from the apheresis product to create a CD34-negative cell fraction from which Treg cells are then isolated, helping to obtain a Treg-enriched cell population.

[0136] The cell populations of the present disclosure can include a population of iNKT. The population of iNKT can be sourced from peripheral blood. The population of iNKT can be sourced from a peripheral blood apheresis product.

[0137] The population of cells can be enriched for iNKT by sorting based on the expression of various markers using, for example, MACS, FACS, or a combination thereof. The iNKT population can be enriched for iNKT, for example, CD3 + Vα24Jα18+ Enrichment can be achieved by cell sorting.

[0138] The number of iNKTs present in a population can be determined, for example, by flow cytometry using CD3 + Vα24Jα18 + The CD3 in the aliquot can be quantified by quantifying the cells. + Vα24Jα18 + The number of cells can be determined, and the volume containing the appropriate dose of iNKT can be determined for recipient administration.In some embodiments, dosage calculation is adjusted based on the measurement of cell viability, for example, the viability determined by flow cytometry using propidium iodide or 7-AAD, or by trypan blue exclusion.

[0139] The cell populations of the present disclosure can include a population of Tmem. The population of Tmem can be sourced from peripheral blood. The population of Tmem can be sourced from a peripheral blood apheresis product.

[0140] A population of cells can be enriched for Tmems by sorting based on the expression of various markers, for example, using MACS, FACS, or a combination thereof. A population of Tmems can be enriched for Tmems, for example, CD3 + CD45RA-CD45RO + Enrichment can be achieved by cell sorting.

[0141] The number of Tmems present in a population can be determined, for example, by flow cytometry using CD3 + CD45RA-CD45RO + The CD3 in the aliquot can be quantified by quantifying the cells. + CD45RA-CD45RO +The number of cells can be determined, and the volume containing the appropriate dose of Tmem to be administered to the recipient can be determined. Dose calculations can be adjusted based on measures of cell viability, such as viability determined by flow cytometry using propidium iodide or 7-AAD, or trypan blue exclusion.

[0142] The cell populations of the present disclosure or cell populations of the present disclosure may be administered fresh after isolation, or may be administered after cryopreservation and subsequent thawing.

[0143] Freshly isolated cells from a donor ("fresh cells") can be administered to a recipient subject. Fresh cells can be stored in a buffer, such as CliniMACs PBS-EDTA buffer containing 0.5% human serum albumin, or Plasma-Lyte-A, pH 7.4, supplemented with 2% human serum albumin. Fresh cells can be stored cryopreserved / unfrozen at low temperatures (e.g., 2-8°C).

[0144] After obtaining a fresh population of cells from a donor, the fresh cells may be incubated for at least about 1 hour, at least about 2 hours, at least about 3 hours, at least about 4 hours, at least about 5 hours, at least about 6 hours, at least about 7 hours, at least about 8 hours, at least about 9 hours, at least about 10 hours, at least about 11 hours, at least about 12 hours, at least about 13 hours, at least about 14 hours, at least about 15 hours, at least about 16 hours, at least about 17 hours, at least about 18 hours, at least about 19 hours, at least about 20 hours, at least about 21 hours, at least about 22 hours, at least about 23 hours, at least about 24 hours, at least about 25 hours, at least about 26 hours, at least about 27 hours, at least about 28 hours, or at least about 30 hours prior to administration to a subject. It may be stored for at least about 29 hours, at least about 30 hours, at least about 31 hours, at least about 32 hours, at least about 33 hours, at least about 34 hours, at least about 35 hours, at least about 36 hours, at least about 37 hours, at least about 38 hours, at least about 39 hours, at least about 40 hours, at least about 44 hours, at least about 48 hours, at least about 50 hours, at least about 55 hours, at least about 60 hours, at least about 61 hours, at least about 62 hours, at least about 65 hours, at least about 70 hours, at least about 72 hours, at least about 80 hours, at least about 90 hours, at least about 96 hours, at least about 120 hours, at least about 150 hours, at least about 200 hours, at least about 300 hours, or longer.

[0145] After a fresh population of cells is obtained from a donor, the fresh cells may be stored for up to about 1 hour, up to about 2 hours, up to about 3 hours, up to about 4 hours, up to about 5 hours, up to about 6 hours, up to about 7 hours, up to about 8 hours, up to about 9 hours, up to about 10 hours, up to about 12 hours, up to about 14 hours, up to about 16 hours, up to about 18 hours, up to about 20, up to about 22 hours, up to about 24 hours, up to about 30 hours, up to about 36 hours, up to about 40 hours, up to about 48 hours, up to about 60 hours, up to about 70 hours, up to about 72 hours, up to about 80 hours, up to about 90 hours, up to about 96 hours, up to about 120 hours, up to about 150 hours, up to about 200 hours, or up to about 300 hours before administration to a subject.

[0146] In some embodiments, after treatment, one or more cell populations of the present disclosure, such as CD45 + The first population of cells, CD45 + A second population of cells, and / or CD45 + The third population of cells may be formulated with one or more excipients and / or cryoprotectants. In some embodiments, the one or more excipients include a buffer, such as a transport buffer or an infusion buffer. In some embodiments, the cell population is formulated at a neutral pH. In some embodiments, the HSPCs of the present disclosure are formulated with one or more excipients at a neutral pH. In some embodiments, the Tregs of the present disclosure are formulated with one or more excipients at a neutral pH. In some embodiments, the Tcon of the present disclosure is formulated with one or more excipients at a neutral pH. In some embodiments, the HSPCs and Tregs of the present disclosure are formulated together with one or more excipients at a neutral pH. In some embodiments, the HSPCs and Tcon of the present disclosure are formulated together with one or more excipients at a neutral pH. In some embodiments, the Tregs and Tcon of the present disclosure are formulated together with one or more excipients at a neutral pH. In some embodiments, the HSPCs, Tregs, and Tcons of the present disclosure are formulated together with one or more excipients at neutral pH, hi some embodiments, the one or more excipients include one or more transport buffers.

[0147] In some embodiments, the neutral pH ranges from about 6.8 to about 7.6. In some embodiments, the neutral pH is about 6.8, about 6.85, about 6.9, about 6.95, about 7, about 7.05, about 7.1, about 7.15, about 7.2, about 7.25, about 7.3, about 7.35, about 7.4, about 7.45, about 7.5, about 7.55, or about 7.6.

[0148] In some embodiments, one or more cell populations of the present disclosure, e.g., CD45 + The first population of cells, CD45 + A second population of cells, and / or CD45 + The third population of cells may be formulated in one or more excipients. In some embodiments, the one or more excipients may include one or more buffers. The buffer may be a transport buffer or an infusion buffer. Transport buffers suitable for use with the cell populations of the present disclosure include buffers having milliequivalents (mEq) of sodium ranging from about 120 mEq sodium to about 160 mEq sodium. In some embodiments, transport buffers of the present disclosure include about 120 mEq sodium, about 125 mEq sodium, about 130 mEq sodium, about 135 mEq sodium, about 140 mEq sodium, about 145 mEq sodium, about 120 mEq sodium, about 150 mEq sodium, about 155 mEq sodium, or about 160 mEq sodium.

[0149] Transport buffers suitable for use with the cell populations of the present disclosure include buffers having an osmolality ranging from about 270 mOsmol / L to about 320 mOsmol / L. In some embodiments, transport buffers of the present disclosure have an osmolality of about 270 mOsmol / L, about 275 mOsmol / L, about 280 mOsmol / L, about 285 mOsmol / L, about 290 mOsmol / L, about 295 mOsmol / L, about 300 mOsmol / L, about 305 mOsmol / L, about 310 mOsmol / L, about 315 mOsmol / L, or about 320 mOsmol / L.

[0150] In some embodiments, one or more cell populations of the present disclosure, e.g., CD45+ The first population of cells, CD45 + A second population of cells, and / or CD45 + Suitable transport buffers for use with the third population of cells include, but are not limited to, phosphate buffered saline (PBS), human serum, PlasmaLyte, and any combination thereof. In some embodiments, one or more cell populations of the present disclosure, e.g., CD45 + The first population of cells, CD45 + A second population of cells, and / or CD45 + In some embodiments, one or more cell populations of the present disclosure, e.g., CD45, are formulated with a transport buffer. + The first population of cells, CD45 + A second population of cells, and / or CD45 + In some embodiments, one or more cell populations of the present disclosure, such as CD45, are formulated with a sufficient amount of PBS. + The first population of cells, CD45 + A second population of cells, and / or CD45 + The third population of cells is formulated with a sufficient amount of human serum. In some embodiments, one or more of the cell populations of the present disclosure, e.g., CD45 + The first population of cells, CD45 + A second population of cells, and / or CD45 + The third population of cells is formulated with a sufficient amount of PlasmaLyte (eg, PlasmaLyte A).

[0151] In some embodiments, the transport buffer of the present disclosure may further comprise a human carrier protein, including, but not limited to, human serum albumin (HSA), intravenous immunoglobulin (IVIG), AB serum, or any combination thereof. In some embodiments, the human carrier protein has a concentration of human carrier protein ranging from about 0.1% w / v to about 10% w / v. In some embodiments, the human carrier protein is at about 0.1% w / v, about 0.2% w / v, about 0.3% w / v, about 0.4% w / v, about 0.5% w / v, about 0.6% w / v, about 0.7% w / v, about 0.8% w / v, about 0.9 w / v, about 1.0% w / v, about 1.1% w / v, about 1.2% w / v, about 1.3% w / v, about 1.4% w / v, about 1.5% w / v, about 1.6% w / v, about 1.7% w / v, about 1.8% w / v, about 1.9% w / v, about 2.0% w / v, about 2.1% w / v, about 2.2% w / v, about 2.3% w / v, about 2.4% w / v, about 2.5% w / v, about 2.6% w / v, about 2.7% w / v, approximately 2.8% w / v, approximately 2.9% w / v, approximately 3.0% w / v, approximately 3.1% w / v, approximately 3.2% w / v, approximately 3.3% w / v, approximately 3.4% w / v, approximately 3.5% w / v, approximately 3.6% w / v, approximately 3.7% w / v, approximately 3.8% w / v, approximately 3.9% w / v, approximately 4.0% w / v, approximately 4.1% w / v, approximately 4.2% w / v, approximately 4.3% w / v, approximately 4.4% w / v, approximately 4.5% w / v, approximately 4.6% w / v, approximately 4.7% w / v, approximately 4.8% w / v, approximately 4.9% w / v, approximately 5.0% w / v, approximately 5.1% w / v, approximately 5.2% w / v, approximately 5.3% w / v, approx. 5.4% w / v, approximately 5.5% w / v, approximately 5.6% w / v, approximately 5.7% w / v, approximately 5.8% w / v, approximately 5.9% w / v, approximately 6.0% w / v, approximately 6.1% w / v, approximately 6.2% w / v, approximately 6.3% w / v, approximately 6.4% w / v, approximately 6.5% w / v, approximately 6.6% w / v, approximately 6.7% w / v, approximately 6.8% w / v, approximately 6.9% w / v, approximately 7.0% w / v, approximately 7.1% w / v, approximately 7.2% w / v, approximately 7.3% w / v, approximately 7.4% w / v, approximately 7.5% w / v, approximately 7.6% w / v, approximately 7.7% w / v, approximately 7.8% w / v, approximately 7.9% w / v, approximately 8.0% w / v, approx. 8.The carrier protein may have a concentration of 1% w / v, about 8.2% w / v, about 8.3% w / v, about 8.4% w / v, about 8.5% w / v, about 8.6% w / v, about 8.7% w / v, about 8.8% w / v, about 8.9% w / v, about 9.0% w / v, about 9.1% w / v, about 9.2% w / v, about 9.3% w / v, about 9.4% w / v, about 9.5% w / v, about 9.6% w / v, about 9.7% w / v, about 9.8% w / v, about 9.9% w / v, or about 10% w / v.

[0152] In some embodiments, one or more cell populations of the present disclosure, e.g., CD45 + The first population of cells, CD45 + A second population of cells, and / or CD45 + The third population of cells is formulated with an excipient, such as, for example, a transport buffer of the present disclosure, in an amount sufficient to obtain a volume ranging from about 5 mL to 1 L. In some embodiments, CD45 + The first population of cells is formulated with an excipient, such as, for example, a transport buffer of the present disclosure, in an amount sufficient to obtain a volume ranging from about 5 mL to 1 L. In some embodiments, CD45 + The second population of cells is formulated with an excipient, such as, for example, a transport buffer of the present disclosure, in an amount sufficient to obtain a volume ranging from about 5 mL to 1 L. In some embodiments, CD45 + The third population of cells is formulated with an excipient, such as, for example, a transport buffer of the present disclosure, in an amount sufficient to obtain a volume ranging from about 5 mL to 1 L. In some embodiments, CD45 + The first population of cells, CD45 + A second population of cells, and / or CD45 +The third population of cells may be diluted (e.g., using one or more excipients such as a transport buffer) to about 5 mL, about 10 mL, about 15 mL, about 20 mL, about 25 mL, about 30 mL, about 35 mL, about 40 mL, about 45 mL, about 50 mL, about 55 mL, about 60 mL, about 65 mL, about 70 mL, about 75 mL, about 80 mL, about 85 mL, about 90 mL, about 95 mL, about 100 mL, about 105 mL, about 110 mL, about 115 mL, about 120 mL, about 125 mL, about 130 mL, about 135 mL, about 140 mL, about 145 mL, about 150 mL, about 155 mL, about 160 mL, about 165 mL, about 170 mL, about 175 mL, about 180 mL, about 185 mL, about 190 mL, about 195 mL, about 200 mL, about 205 mL, about 210 mL, about 215 mL, about 220 mL, about 225 mL, about 230 mL, about 235 mL, about 240 mL, about 245 mL, about 250 mL, about 255 mL, about 260 mL, about 265 mL, about 270 mL, about 275 mL, about 280 mL, about 285 mL, about 290 mL, about 300 mL, about 310 mL, about 315 mL, about 320 mL, about 325 mL, about 330 mL, about 335 mL, about 340 mL, about 345 mL, about 350 mL, about 355 mL, about 360 mL, about 365 mL, about 370 mL 70mL, 175mL, 180mL, 185mL, 190mL, 195mL, 200mL, 205mL, 210mL, 215mL, 220mL, 225mL, 230mL, 235mL, 240mL, 245mL, 250mL, 255mL, 260mL, 265mL, 270mL, 275mL, 280mL, 285mL, 290mL, 295mL, 300mL, 305mL, 310mL, 315mL, 320mL, 325mL, 330mL, 335mL, 340mL, 345mL , 350mL, 355mL, 360mL, 365mL, 370mL, 375mL, 380mL, 385mL, 390mL, 395mL, 400mL, 405mL, 410mL, 415mL, 420mL, 425mL, 430mL, 435 mL, approx. 440 mL, approx. 445 mL, approx. 450 mL, approx. 455 mL, approx. 460 mL, approx. 465 mL, approx. 470 mL, approx. 475 mL, approx. 5mL, approx. 530mL, approx. 535mL, approx. 540mL, approx. 545mL, approx. 550mL, approx. 555mL, approx. 560mL, approx. 565mL, approx. 615mL, about 620mL, about 625mL, about 630mL, about 635mL, about 640mL, about 645mL, about 650mL, about 655mL, about 660mL, about 665mL, about 670mL, about 675mL, about 680mL, about 685mL, about 690mL, about 695mL, about 700mL,Approximately 705mL, approximately 710mL, approximately 715mL, approximately 720mL, approximately 725mL, approximately 730mL, approximately 735mL, approximately 740mL, approximately 745mL, approximately 750mL, approximately 755mL, approximately 760mL, approximately 765mL, approximately 770mL, approximately 775mL, approximately 7 80mL, about 785mL, about 790mL, about 795mL, about 800mL, about 805mL, about 810mL, about 815mL, about 820mL, about 825mL, about 830mL, about 835mL, about 840mL, about 845mL, about 850mL, about 855m The formulations may be formulated in volumes of about 860 mL, about 865 mL, about 870 mL, about 875 mL, about 880 mL, about 885 mL, about 890 mL, about 895 mL, about 900 mL, about 905 mL, about 910 mL, about 915 mL, about 920 mL, about 925 mL, about 930 mL, about 935 mL, about 940 mL, about 945 mL, about 950 mL, about 955 mL, about 960 mL, about 965 mL, about 970 mL, about 975 mL, about 980 mL, about 985 mL, about 990 mL, about 995 mL, or about 1 L. In some embodiments, the formulations of the present disclosure further comprise a human carrier protein of the present disclosure. In some embodiments, the formulations of the present disclosure further comprise one or more cryoprotectants.

[0153] In some embodiments, the cell populations of the present disclosure, e.g., CD45 + The first population of cells, CD45 + A second population of cells, and / or CD45 + The third population of cells may be cryopreserved. In some embodiments, cryopreservation may be beneficial for the methods disclosed herein. For example, the third population of cells may be cryopreserved to contain at least CD45 Tcon prior to subsequent thawing and administration to a subject. + By cryopreserving the third population of cells, GVHD can be reduced.

[0154] An additional embodiment provides a method for transplanting a conventional T cell (Tcon) population into a human subject without inducing a stage 2 or higher graft-versus-host disease (GVHD) response up to about 100 days after transplantation. The method includes (i) administering a solution containing a population of conventional T cells (Tcon), and (ii) administering a solution containing a population of regulatory T cells (Treg). In this method, the Tcon population is cryopreserved for at least about 4 hours, and the solution containing the Tcon population and the solution containing the Treg population contain less than about 5 EU of endotoxin per ml of solution.

[0155] Cryopreservation may involve the addition of a preservative (e.g., DMSO) or one or more cryoprotectants, and gradually cooling the cells in a controlled-rate freezer to prevent cell damage due to osmotic pressure resulting from ice crystal formation. Cryopreservation may include commercially available cryopreservation reagents and materials (e.g., cryoprotectants), such as Cryobags sorbitol, dimethyl sulfoxide (DMSO), propylene glycol, glycerol, polyvinylpyrrolidone (PVP), and polyethylene glycol (PEG), serum, HSA, hetastarch, CRYOSTOR CS2, CRYOSTOR CS5, and CRYOSTOR CS10.

[0156] In certain embodiments, the cryoprotectant of the present disclosure comprises DMSO. For example, in some embodiments, CD45 +The third population of cells may be formulated for cryopreservation using a cryoprotectant comprising DMSO in an amount ranging from about 1% v / v to about 15% v / v. In some embodiments, the cryoprotectant is about 1%, about 1.2%, about 1.4%, about 1.6%, about 1.8%, about 2%, about 2.2%, about 2.4%, about 2.6%, about 2.8%, about 3%, about 3.2%, about 3.4%, about 3.6%, about 3.8%, about 4%, about 4.2%, or about 4. 4v / v%, approximately 4.6v / v%, approximately 4.8v / v%, approximately 5v / v%, approximately 5.2v / v%, approximately 5.4v / v%, approximately 5.6v / v%, approximately 5.8v / v%, approximately 6v / v%, approximately 6.2v / v%, Approximately 6.4v / v%, approximately 6.6v / v%, approximately 6.8v / v%, approximately 7v / v%, approximately 7.2v / v%, approximately 7.4v / v%, approximately 7.6v / v%, approximately 7.8v / v%, approximately 8v / v%, approximately 8.2v / v %, about 8.4v / v%, about 8.6v / v%, about 8.8v / v%, about 9v / v%, about 9.2v / v%, about 9.4v / v%, about 9.6v / v%, about 9.8v / v%, about 10v / v%, about 10 .2v / v%, approximately 10.4v / v%, approximately 10.6v / v%, approximately 10.8v / v%, approximately 11v / v%, approximately 11.2v / v%, approximately 11.4v / v%, approximately 11.6v / v%, approximately 11.8v / v %, approximately 12v / v%, approximately 12.2v / v%, approximately 12.4v / v%, approximately 12.6v / v%, approximately 12.8v / v%, approximately 13v / v%, approximately 13.2v / v%, approximately 13.4v / v%, approximately 13.6 DMSO in an amount of v / v%, about 13.8v / v%, about 14v / v%, about 14.2v / v%, about 14.4v / v%, about 14.6v / v%, about 14.8v / v%, or about 15v / v%.

[0157] In some embodiments, CD45 +The third population of cells is formulated for cryopreservation using a cryoprotectant comprising DMSO and further comprising hetastarch, hi some embodiments, the cryoprotectant comprises hetastarch in an amount ranging from about 0.1% w / v to about 5% w / v. In some embodiments, the cryoprotectant is about 0.1% w / v, about 0.2% w / v, about 0.3% w / v, about 0.4% w / v, about 0.5% w / v, about 0.6% w / v, about 0.7% w / v, about 0.8% w / v, about 0.9% w / v, about 1.0% w / v, about 1.1% w / v, about 1.2% w / v, about 1.3% w / v, about 1.4% w / v, about 1.5% w / v, about 1.6% w / v, about 1.7% w / v, about 1.8% w / v, about 1.9% w / v, about 2.0% w / v, about 2.1% w / v, about 2.2% w / v, about 2.3% w / v, about 2.4% w / v, about 2.5% w / v, about 2.6% w / v, or about 2.7% w / v. about 2.8% w / v, about 2.9% w / v, about 3.0% w / v, about 3.1% w / v, about 3.2% w / v, about 3.3% w / v, about 3.4% w / v, about 3.5% w / v, about 3.6% w / v, about 3.7% w / v, about 3.8% w / v, about 3.9% w / v, about 4.0% w / v, about 4.1% w / v, about 4.2% w / v, about 4.3% w / v, about 4.4% w / v, about 4.5% w / v, about 4.6% w / v, about 4.7% w / ...

Claims

1. A multi-component cell therapy product, a) Approximately 1.0 × 10 per kilogram of body weight of the human subject receiving the product. ５ ~Approx. 1.0×10 ８ 34 CDs ＋ Isolated CD45 containing doses of hematopoietic stem progenitor cells (HSPCs) ＋ A first single-dose transfer bag containing a first population of cells, the CD45 ＋ The first population of cells is formulated with excipients at a neutral pH, in the first single-dose transfer bag. b) A second single-dose transfer bag containing a second population of isolated CD45 cells comprising a dose of regulatory T cells (Treg) of from about 1.0×10 ５ to about 2.0×10 ７ fresh CD4 ＋ CD25 ＋ CD127 ｄｉｍ per kilogram of body weight of the human subject receiving the product, wherein the second population of isolated CD45 ＋ cells is formulated with an excipient at neutral pH, and optionally the first single-dose transfer bag and the second single-dose transfer bag are the same transfer bag, a second single-dose transfer bag, and ＋ ​ c) Approximately 1.0 × 10 per kilogram of body weight of the human subject receiving the product. ５ ~Approx. 4.0×10 ７ Individual standard CD3 ＋ Isolated CD45 containing T cell (Tcon) dose ＋ A third single-dose transfer bag containing a third population of cells, wherein the isolated CD45 ＋ A third population of cells is formulated with an excipient at a neutral pH, the excipient comprising one or more cryoprotective agents, in a third single-dose transfer bag. A multi-component cell therapy product that includes [specific ingredient / method].

2. The multicomponent cell therapy product according to claim 1, further comprising a pharmaceutical composition containing a dose of tacrolimus sufficient to maintain a trough blood level of about 5 ng / mL to about 10 ng / mL in the human subject receiving the product.

3. The isolated CD45 ＋ The first population of cells, the isolated CD45 ＋ A second population of cells, and / or the isolated CD45 ＋ The multicomponent cell therapy product according to claim 1, wherein the third population of cells is formulated in a volume ranging from approximately 5 mL to approximately 1 L.

4. The isolated CD45 ＋ The first population of cells, the isolated CD45 ＋ The multicomponent cell therapy product according to claim 1, wherein the second population of cells and the third population of isolated CD45+ cells are derived from an allogeneic donor having at least one HLA mismatch to the human subject receiving the product.

5. A pharmaceutical product comprising a multicomponent cell therapy product according to any one of claims 1 to 4, for use in a method of treating a human subject having or suspected of having a hematological malignancy, The method includes administering the multicomponent cell therapy product to the human subject. The aforementioned pharmaceutical.

6. A method for producing a multicomponent cell therapy product, a) Obtaining HSPC mobilization peripheral blood apheresis samples from donors of the same type for human subjects receiving the product, and b) Approximately 1.0 × 10 per kilogram of body weight of the human subject receiving the product ５ ~Approx. 1.0×10 ８ 34 CDs ＋ A first sample containing hematopoietic stem progenitor cells (HSPCs), approximately 1.0 × 10⁶ per kilogram of body weight of the human subject receiving the product. ５ ~Approx. 2.0×10 ７ 4 CDs ＋ CD25 ＋ CD127 ｄｉｍ A second sample containing regulatory T cells (Treg), and approximately 1.0 × 10⁶ per kilogram of body weight of the human subject receiving the product. ５ ~Approx. 4.0×10 ７ Individual standard CD3 ＋ A third sample containing T cells (Tcon) is selected from the apheresis sample. The multicomponent cell therapy product comprises the Tcon, the HSPC, and the Treg, wherein the HSPC is selected by selecting the first sample using one or more ISPs specific to CD34 and selecting a population enriched with CD34, and the Treg is selected by selecting the second sample using one or more ISPs specific to CD25, one or more ISPs specific to CD4, and / or one or more ISPs specific to CD127 and CD4 ＋ and CD127 ｄｉｍ The method is selected by selecting a population of cells.

7. A multicomponent cell therapy product prepared by the method described in claim 6.

8. A pharmaceutical product comprising a multicomponent cell therapy product according to any one of claims 1 to 4, for use in a method of treating a human subject having or suspected of having acute leukemia in complete remission, active leukemia, acute leukemia resistant to first-line treatment, or acute leukemia with minimal residual disease, The method includes administering the multicomponent cell therapy product to the human subject. The aforementioned pharmaceutical.

9. A pharmaceutical product comprising a multicomponent cell therapy product according to any one of claims 1 to 4, for use in a method of treating a human subject having or suspected of having low-risk acute myeloid leukemia (AML), high-risk AML in complete remission, or chronic myeloid leukemia (CML), The method includes administering the multicomponent cell therapy product to the human subject. The aforementioned pharmaceutical.

10. A pharmaceutical product comprising a multicomponent cell therapy product according to any one of claims 1 to 4, for use in a method of treating human subjects having or suspected to have high-risk myelodysplastic syndrome, treatment-related myelodysplastic syndrome, and / or secondary myelodysplastic syndrome, The method includes administering the multicomponent cell therapy product to the human subject. The aforementioned pharmaceutical.

11. A pharmaceutical product comprising a multicomponent cell therapy product according to any one of claims 1 to 4, for use in a method of treating a human subject having or suspected of having a myeloproliferative syndrome, The method includes administering the multicomponent cell therapy product to the human subject. The aforementioned pharmaceutical.

12. A pharmaceutical product comprising a multicomponent cell therapy product according to any one of claims 1 to 4, for use in a method of treating human subjects having or suspected of having non-Hodgkin lymphoma with a poor prognosis and unsuitable for autologous hematopoietic stem cell transplantation (HCT), The method includes administering the multicomponent cell therapy product to the human subject. The aforementioned pharmaceutical.

13. In the above method, the isolated CD45 ＋ The third population of cells is the isolated CD45 ＋ The first population of cells and the isolated CD45 ＋ The pharmaceutical agent according to claim 8, administered approximately 24 to 120 hours after the second population of cells.

14. The pharmaceutical agent according to claim 9, wherein the isolated third population of CD45+ cells is administered approximately 24 to 120 hours after the first population of isolated CD45+ cells and the second population of isolated CD45+ cells.

15. The pharmaceutical agent according to claim 10, wherein the isolated third population of CD45+ cells is administered approximately 24 to 120 hours after the first population of isolated CD45+ cells and the second population of isolated CD45+ cells.

16. The pharmaceutical agent according to claim 11, wherein the isolated third population of CD45+ cells is administered approximately 24 to 120 hours after the first population of isolated CD45+ cells and the second population of isolated CD45+ cells.

17. The pharmaceutical agent according to claim 12, wherein the isolated third population of CD45+ cells is administered approximately 24 to 120 hours after the first population of isolated CD45+ cells and the second population of isolated CD45+ cells.

18. A cell therapy kit comprising a multicomponent cell therapy product according to any one of claims 1 to 4, wherein the kit further comprises a written instruction on the use of the cell therapy for the treatment of hematological malignancies in human subjects.

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