Compositions and methods for allografting

Abstract

This invention provides compositions and methods useful for treating CD45+ cell depletion, and in particular for treating various hematopoietic disorders, metabolic disorders, cancer, and autoimmune diseases. [Solution] The compositions and methods described herein can be used, for example, to treat a disorder by depleting a population of CD45+ cancer cells or autoimmune cells. The compositions and methods described herein can also be used to prepare a patient for allogeneic hematopoietic stem cell transplantation therapy and to improve the engraftment of allogeneic hematopoietic stem cell transplants by selectively depleting endogenous hematopoietic stem cells before the transplantation procedure.

Description

[Technical Field]

[0001] Cross-reference of related applications This application is based on U.S. Provisional Patent Application No. 62 / 978,141, filed on 18 February 2020. Priority to U.S. Patent Provisional Application No. 63 / 062,845, filed on August 7, 2020. The right is asserted. Each of the aforementioned priority applications is incorporated herein by reference in its entirety. ru.

[0002] Sequence List This application includes a sequence listing submitted electronically in ASCII format, which in whole is by reference. This is incorporated herein by reference. This ASCII copy, created on February 11, 2021, is M The file name is 103034_2195WO_0576_7_SL.txt, and the size is It is 261,714 bytes.

[0003] This disclosure relates to CD45 cells such as hematopoietic stem cells or mature immune cells (e.g., T cells). + CD45 that can bind to toxin-bound CD45 expressed by cells. Administration of a targeted portion (e.g., an antibody-drug conjugate) can lead to various pathological conditions, among others, This relates to the treatment of patients suffering from blood disorders, metabolic disorders, cancer, and autoimmune diseases. [Background technology]

[0004] Allogeneic hematopoietic stem cell transplantation (allogeneic HSCT) is a curative treatment for malignant and non-malignant hematological disorders. This is a treatment method that may result in [resulting in a positive outcome]. Allogeneic cell therapy involves transplanting cells into the patient, and the transplanted cells... The cells used are derived from a donor other than the patient. Common allogeneic donors used in allogeneic cell therapy. These types include HLA-matched siblings, matched unrelated donors, and partially matched families. This includes familial donors, related umbilical cord blood donors, and unrelated umbilical cord blood donors. The ultimate goal of cell therapy. This could form the basis for “off-the-shelf” products that could expand the use of allogeneic cell therapy. The goal is to identify allogeneic cell therapies that can be performed (Brandenberger, et al.) 2011).BioProcess International.9(suppl.I ):30-37).

[0005] Despite its potential, the therapeutic use of allogeneic cells currently carries a risk of complications. This can occur, making this therapy difficult. In an immune-responsive host, transplanted allogeneic cells The graft is quickly rejected, a process known as host-versus-graft rejection (HvG). This not only substantially reduces the effectiveness of the transplanted cells, but also causes adverse events in the recipient. Because it can cause friction, the use of allogeneic cells is limited. Furthermore, allogeneic HSCs Current regimens regarding patient preparation or conditioning prior to T include organ toxicity and infertility. There are regimen-related mortality and morbidity rates, including the risk of disease and secondary malignancies. Therefore, the use of this curative treatment is restricted. For this reason, in malignant and non-malignant conditions The use of allogeneic HSCTs is severely restricted. Currently, the use of immunosuppressants is avoided, and allogeneic HSCTs are being produced. To promote the engraftment of hematopoietic stem cell grafts and maintain the pluripotency and hematopoietic function of these cells after transplantation. A safer conditioning regimen is needed. [Overview of the Initiative]

[0006] Provided herein are CD45-targeting moieties that specifically bind to CD45 (e.g., , antibodies and antibody-drug conjugates (ADCs). CD45 targeting portion (for example, Anti-CD45 antibodies (and ADCs) are useful in monotherapy conditioning treatments, and this is because The patient underwent allogeneic transplantation without the use of additional conditioning agents such as immunosuppressants. For example, preparations are made for receiving a complete mismatch allograft. According to the method, CD45, for example, hematopoietic stem cells or mature immune cells (e.g., T cells) CD45 targets that can bind to CD45 expressed by CD45+ cells, such as Administering a conjugated portion (e.g., an anti-CD45 antibody or antibody-drug conjugate) to the patient. This allows the patient to be conditioned for allogeneic hematopoietic stem cell transplantation therapy. In some embodiments, the CD45 targeting moiety can bind to the toxin. In this embodiment, the CD45-targeting portion (e.g., an anti-CD45 antibody or ADC) is other It is administered as monotherapy in the absence of conditioning agents. For example, CD45 standard The targeted portion (e.g., anti-CD45 antibody or ADC) is an immunodepletion agent (e.g., anti-CD4 and (or anti-CD8), whole-body irradiation (e.g., low-dose TBI), and / or cyclophosph In the absence of one or more immunosuppressants such as amide, the patient's CD45+ cells are depleted. It can be administered in sufficient quantities.

[0007] In one embodiment, this disclosure relates to human patients requiring hematopoietic stem cell (HSC) transplantation. A method to deplete a population of CD45+ cells, in which the patient receives a transplant containing allogeneic HSCs. Previously, the CD45-targeting moiety bound to the cytotoxic (e.g., anti-CD45 antibody drug conjugate) The procedure involves administering an effective dose of (ADC) to the patient, and the patient may be present before or after transplantation. This provides a method that avoids conditioning with immunosuppressants at almost the same time.

[0008] In another embodiment, the disclosure describes (a) targeting a cytotoxic-bound CD45 in a human patient. The portion (e.g., anti-CD45 antibody-drug conjugate (ADC)) in the absence of immunosuppressants. Below, administer an effective dose sufficient to deplete the patient's population of CD45+ cells, and ( b) The present invention provides a method that subsequently includes performing a transplant on the patient that includes allogeneic HSCs.

[0009] In another embodiment, this disclosure includes performing an allogeneic HSC transplant on a human patient. In the absence of immunosuppressants, the CD45-targeting moiety bound to the cytotoxin (e.g., anti-C) Administration of D45 antibody-drug conjugates (ADCs) depletes the patient's hematopoietic stem cell population. It provides a method that involves receiving a sufficient effective amount beforehand.

[0010] In some embodiments, the CD45-targeting moiety bound to the cytotoxic is anti-CD45 It is an antibody-drug conjugate (ADC). In some embodiments disclosed herein, In this context, allogeneic HSCs contain one or more HLA mismatches to the patient's HLA antigen. In another embodiment, allogeneic HSCs have two or more HLA antigens against the patient's HLA antigen. This includes mismatches. In some embodiments, allogeneic HSCs match the patient's HLA antigen. And it includes three or more HLA mismatches. In some embodiments, homogeneous HSCs are , including five or more HLA mismatches with respect to the patient's HLA antigen. Several embodiments In this context, allogeneic HSCs contain a complete HLA mismatch with the patient's HLA antigen. In some embodiments, allogeneic HSCs are 1 against the patient's minor histocompatibility antigen. It includes one or more minor histocompatibility antigen (miHA) mismatches. In some embodiments, In allogeneic HSCs, two or more miHA misma are present for the patient's minor histological compatibility antigen. Includes a streak. In some embodiments, allogeneic HSCs are minor histocompatibility antigens of the patient. This includes five or more miHA mismatches.

[0011] In some embodiments of the aforementioned aspects, the transplantation includes a complete mismatch of allogeneic HSCs. obtain.

[0012] In some embodiments disclosed herein, immunosuppressants are used in conjunction with total body irradiation (TBI). ) In some embodiments of the above-described aspects, the immunosuppressant is used in low-dose TBI Yes. In some embodiments of the aforementioned model, the immunosuppressant is an anti-CD4 antibody, an anti-CD 8 antibodies, or combinations thereof. In some embodiments of the above-described model, The immunosuppressant is cyclophosphamide.

[0013] In some embodiments disclosed herein, the patient undergoes at least 24 pre-transplantation procedures. For at least 24 hours after transplantation, the patient is not subjected to immunosuppressants. In this case, the patient shall have at least 48 hours before transplantation and / or at least 48 hours after transplantation. , the patient does not receive immunosuppressants. In other embodiments, the patient has at least 72 hours prior to transplantation. and / or for at least 72 hours after transplantation, the patient does not receive immunosuppressants. In other embodiments, Therefore, the patient will be immune for at least 96 hours before transplantation and / or at least 96 hours after transplantation. No disease control agent is administered. In other embodiments, the patient receives at least 7 days and / Alternatively, the patient does not receive immunosuppressants for at least 7 days after transplantation. In other embodiments, the patient For at least 14 days before transplantation and / or at least 14 days after transplantation, immunosuppressants are administered. In other embodiments, the patient undergoes at least one month prior to transplantation and / or No immunosuppressants should be administered for at least one month after transplantation.

[0014] In some embodiments, the patient spends at least 3 days and at least 7 days prior to transplantation. , at least 14 days, at least 21 days, at least 28 days, at least 1 month or not receive immunosuppressants for at least two months. In some embodiments, the patient The person shall have at least 3 days, at least 7 days, at least 14 days, and at least For 21 days, at least 28 days, at least 1 month, or at least 2 months, immunity They do not receive suppressants.

[0015] In some embodiments disclosed herein, the patient is treated with cytotoxic-bound CD. Receive an effective dose of the CD45-targeted portion (e.g., anti-CD45 ADC). In terms of application methods, the effective amounts are at least 80%, 85%, 90%, 95%, 97%, and 99%. This is a sufficient amount to establish % or 100% donor chimerism. For example, several In this embodiment, an effective amount is administered as a monotherapy in the absence of other conditioning agents. In that case, at least 80%, 85%, 90%, 95%, 97%, 99%, or 100% This is a sufficient amount to establish % donor chimerism. In some embodiments, The efficacy is determined by the patient's condition before undergoing an allogeneic transplant (e.g., a completely mismatched allogeneic transplant). When administered as a monotherapy in the absence of conditioning agents, at least 80%, 85%, Sufficient to establish 90%, 95%, 97%, 99%, or 100% donor chimerism. It is a certain amount. In some embodiments, donor chimerism occurs at least 6 times after transplantation. Evaluation is performed over a week, 7 weeks, 8 weeks, 9 weeks, or 10 weeks. In some embodiments, In some embodiments, donor chimerism is whole peripheral chimerism. Nerve chimerism is bone marrow chimerism. In some embodiments, donor chimerism is used. The zym is T-cell chimerism. In some embodiments, donor chimerism is This is B-cell chimerism.

[0016] In some embodiments disclosed herein, a cytotoxic-bound CD45 target The effective dose of the compounded portion (e.g., anti-CD45 ADC) is administered to the patient as a single dose. In other embodiments, a CD45-targeting moiety conjugated to a cytotoxic (e.g., anti-CD45) is used. The effective dose of ADC is administered to the patient in two doses. In other embodiments, cytotoxicity is linked to The effective dose of the combined CD45-targeted portion (e.g., anti-CD45 ADC) is obtained in two or more doses (e.g. For example, administering the drug to a patient in 2, 3, 4, 5, 6, 7, 8, 9, 10 or more doses. It can be done.

[0017] In some embodiments disclosed herein, the implantation is performed to concentrate anti-CD45 ADC The procedure is performed on the patient after the substance has been substantially eliminated from the patient's blood.

[0018] In some embodiments disclosed herein, hematopoietic stem cells or their offspring are used to treat disease. Even after more than two days have passed since hematopoietic stem cell transplantation, the functional potential of the hematopoietic stem cells is maintained. ru.

[0019] In some embodiments disclosed herein, the autologous hematopoietic stem cells or their progeny are capable of localizing to the hematopoietic tissue and / or reestablishing hematopoiesis after transplantation of the hematopoietic stem cells into a patient.

[0020] In some embodiments disclosed herein, the hematopoietic stem cells are associated with transplantation into a patient, resulting in the restoration of a cell population selected from the group consisting of megakaryocytes, thrombocytes, platelets, erythrocytes, mast cells, myeloblasts, basophils, neutrophils, eosinophils, microglia, granulocytes, monocytes, osteoclasts, antigen-presenting cells, macrophages, dendritic cells, natural killer cells, T lymphocytes, and B lymphocytes.

[0021] In some embodiments disclosed herein, the patient has a stem cell disorder. In some embodiments, the patient has an abnormal hemoglobinopathy, an autoimmune disorder, a myelodysplastic disorder, an immunodeficiency disorder, or a metabolic disorder. In some embodiments, the patient has cancer.

[0022] In some embodiments disclosed herein, the anti-CD45 ADC has a dissociation rate (K D) of 1×10 -2 ~1×10 -3 -1 / s, 1×10 -3 ~1×10 -4 -1 / s, 1×10 -5 ~1×10 -6 -1 / s, 1×10 -6 ~1×10 -7 -1 / s or 1×10 -7 ~1×10 -8 -1 / s and comprises an antibody having such a dissociation rate. OFF In some embodiments, the anti-CD45 ADC is determined by a biolayer interferometry (BLI) assay ​ When that was decided, it was approximately 100 nM or less, approximately 90 nM or less, approximately 80 nM or less, approximately 70 nM or less. Lower, about 60nM or less, about 50nM or less, about 40nM or less, about 30nM or less, about 20nM or less Bottom, about 10nM or less, about 8nM or less, about 6nM or less, about 4nM or less, about 2nM or less, about 1 K below nM D It contains an antibody that binds to CD45.

[0023] In some embodiments disclosed herein, anti-CD45 ADC is a humanized anti- Contains CD45 antibody. In some embodiments disclosed herein, anti-CD45 ADC contains a human anti-CD45 antibody. In some embodiments, anti-CD45 AD C contains the anti-CD45 antibody listed in Table 5. In some embodiments, anti-CD4 5. ADCs are the heavy chain complementarity determination regions (CDRs) 1-3 and light chain CDRs listed in Table 5. 1-3, or containing an antibody. In some embodiments, anti-CD45 ADC is shown in Table 5. It includes the heavy chain variable region and light chain variable region of the antibody described in [reference]. In some embodiments, Anti-CD45 ADCs include humanized versions of anti-CD45 antibodies listed in Table 5. In some embodiments, the anti-CD45 ADC is an anti-CD45 antibody as listed in Table 5. This includes a deimmunized version.

[0024] In some embodiments disclosed herein, the anti-CD45 ADC is an interceptor It contains an anti-CD45 antibody. In some embodiments, the anti-CD45 ADC is IgG Contains antibodies. In some embodiments, IgG is of the IgG1 isotype, IgG2 It is an isotype, IgG3 isotype, or IgG4 isotype.

[0025] In some embodiments disclosed herein, the anti-CD45 ADC is a linker It contains an anti-CD45 antibody conjugated to a cytotoxic via a certain mechanism. In some embodiments, Furthermore, cytotoxicity is an RNA polymerase inhibitor. In some embodiments, RN A polymerase inhibitors are amatoxins. In some embodiments, RNA polymerase inhibitors are used. The rimerase inhibitor is amanitin. In some embodiments, amanitin is α-Amanitin, β-Amanitin, γ-Amanitin, ε-Amanitin, Amanine, Flax The substance is selected from the group consisting of naamide, amanulin, amanulinic acid, and proamanulin. In some embodiments, the cytotoxin is pyrrolobenzodiazepine (PBD). In some embodiments, the cytotoxin is Pseudomonas exotoxin A, debuganin, diphthene Lithoxin, saporin, mytansin, meitansinoid, auristatin, anthracite Clin, Calicae Sewing Machine, Irinotecan, SN-38, Duocalmycin, Pyrolobene Zodiazepines, pyrrolobenzodiazepine dimers, indolinobenzodiazepines, indoli From the group consisting of nobenzodiazepine dimers and indolinobenzodiazepine pseudodimers Selected. In some embodiments, the cytotoxic is auristatin, for example, MMA. It is either E or MMAF.

[0026] In some embodiments disclosed herein, the antibody is in the Fc domain of the antibody. It is conjugated to the toxin via cysteine ​​residues. In some embodiments, Stain residues are introduced via amino acid substitutions in the Fc domain of the antibody. In embodiments, the amino acid substitution is S239C or D265C.

Brief Description of the Drawings

[0027] [Figure 1A] Graph showing the results of an in vivo depletion assay demonstrating that CD45-ADC effectively depletes mouse HSCs, WBCs, lymphocytes, neutrophils, and monocytes in the bone marrow of C57Bl / 6 mice. Flow cytometry gating strategy and results showing long-term HSC depletion in the bone marrow collected 2 days after administration of PBS or 3 mg / kg of CD45-ADC (administered on day 0). [Figure 1B] Graph showing the results of an in vivo depletion assay demonstrating that CD45-ADC effectively depletes mouse HSCs, WBCs, lymphocytes, neutrophils, and monocytes in the bone marrow of C57Bl / 6 mice. Graph showing the levels of long-term HSCs (LT-HSCs) in the bone marrow 2 days after administration of PBS, isotype-ADC, or CD45-ADC. [Figure 1C] Graph showing the results of an in vivo depletion assay demonstrating that CD45-ADC effectively depletes mouse HSCs, WBCs, lymphocytes, neutrophils, and monocytes in the bone marrow of C57Bl / 6 mice. Graph showing the antibody concentration in CD45-ADC plasma as a function of time after administration of 3 mg / kg of CD45-ADC to mice, indicating that the CD45-ADC half-life of 3 mg / kg of CD45-ADC in C57Bl / 6 mice is 1.7 hours. [Figure 1D] Graph showing the results of an in vivo depletion assay demonstrating that CD45-ADC effectively depletes mouse HSCs, WBCs, lymphocytes, neutrophils, and monocytes in the bone marrow of C57Bl / 6 mice. Graph showing the levels of peripheral lymphocytes 0, 3, 7, 9, 14, and 21 days after administration of PBS, isotype-SAP, or CD45-SAP. Asterisks (*) indicate p < 0.05 when comparing CD45-ADC-treated mice to untreated mice. [Figure 1E]This graph shows the results of an in vivo depletion assay demonstrating that CD45-ADC effectively depletes mouse HSCs, WBCs, lymphocytes, neutrophils, and monocytes in the bone marrow of C57Bl / 6 mice. The graph also shows the depletion of WBCs, lymphocytes, neutrophils, and monocytes in the bone marrow of mice treated with CD45-ADC (0.3 mg / kg, 1 mg / kg, or 3 mg / kg) compared to untreated mice. [Figure 1F] The graph shows the results of an in vivo depletion assay demonstrating that CD45-ADC effectively depletes mouse HSCs, WBCs, lymphocytes, neutrophils, and monocytes in the bone marrow of C57Bl / 6 mice. The graph also shows the results of depletion of LSK, ST-HSCs, and LT-HSCs in the bone marrow of mice treated with CD45-ADC. [Figure 1G] This graph shows the results of an in vivo depletion assay demonstrating that CD45-ADC effectively depletes mouse HSCs, WBCs, lymphocytes, neutrophils, and monocytes in the bone marrow of C57Bl / 6 mice. The graph shows the levels of leukocytes (WBCs), neutrophils, lymphocytes, and monocytes at post-treatment days 0, 3, 7, 9, 14, and 21 in mice treated with CD45-ADC (0.3, 1 mg / kg, or 3 mg / kg) (*Mice treated with 3 mg / kg were euthanized on day 11 due to poor health and significant weight loss). [Figure 1H] This graph shows the results of an in vivo depletion assay demonstrating that CD45-ADC effectively depletes mouse HSCs, WBCs, lymphocytes, neutrophils, and monocytes in the bone marrow of C57Bl / 6 mice. The graph also shows the levels of RBCs and platelets on days 0, 3, 7, 9, 14, and 21 after treatment with CD45-ADC (0.3 mg / kg, 1 mg / kg, or 3 mg / kg) (*Mice treated with 3 mg / kg were euthanized on day 11 due to poor health and significant weight loss). [Figure 2A]The graph shows the results of an in vivo study demonstrating that CD45-ADC enables congenic bone marrow transplantation in a mouse model. C57Bl / 6 mice were conditioned with 9 Gy TBI, isotype-ADC, or CD45-ADC, and whole bone marrow was transplanted from B6.SJL (B6 CD45.1+) mice. The percentage of donor chimerism detected in the blood using CD45.1+ antigen at 4, 8, 12, and 16 weeks after transplantation is graphed as a function of the recipient's treatment mode. [Figure 2B] The graph shows the results of an in vivo study demonstrating that CD45-ADC enables congenic bone marrow transplantation in a mouse model. C57Bl / 6 mice were conditioned with 9 Gy TBI, isotype-ADC, or CD45-ADC, and whole bone marrow was transplanted from B6.SJL (B6 CD45.1+) mice. The percentage of peripheral donor bone marrow chimerism in transplant recipients at 4, 8, 12, and 16 weeks post-transplant is graphed as a function of the recipient's treatment mode. [Figure 2C] The graph shows the results of an in vivo study demonstrating that CD45-ADC enables congenic bone marrow transplantation in a mouse model. C57Bl / 6 mice were conditioned with 9 Gy TBI, isotype-ADC, or CD45-ADC, and whole bone marrow was transplanted from B6.SJL (B6 CD45.1+) mice. The percentage of B-cell chimerism in transplant recipients at 4, 8, 12, and 16 weeks post-transplant is graphed as a function of the recipient's treatment mode. [Figure 2D] The graph shows the results of an in vivo study demonstrating that CD45-ADC enables congenic bone marrow transplantation in a mouse model. C57Bl / 6 mice were conditioned with 9 Gy TBI, isotype-ADC, or CD45-ADC, and whole bone marrow was transplanted from B6.SJL (B6 CD45.1+) mice. The percentage of T cell chimerism in transplant recipients at 4, 8, 12, and 16 weeks post-transplant is graphed as a function of the recipient's treatment mode. [Figure 3A]The graph shows the results of in vivo studies of CD45-ADC conditioning prior to minor mismatch allogeneic transplantation of Balb / cCD45.1 donor cells into DBA / 2 recipient mice. The percentage of donor chimerism detected in the blood using CD45.1+ antigen at 4, 8, 12, and 16 weeks after transplantation is shown graphically as a function of the transplant recipient's treatment mode. [Figure 3B] The graphs show the results of in vivo studies of CD45-ADC conditioning prior to minor mismatch allogeneic transplantation of Balb / cCD45.1 donor cells into DBA / 2 recipient mice. The percentage of peripheral donor bone marrow chimerism in transplant recipients at 4, 8, 12, and 16 weeks post-transplantation is shown graphically as a function of the recipient's treatment mode. [Figure 3C] The graphs show the results of in vivo studies of CD45-ADC conditioning prior to minor mismatch allogeneic transplantation of Balb / cCD45.1 donor cells into DBA / 2 recipient mice. The percentage of B cell chimerism in transplant recipients at 4, 8, 12, and 16 weeks post-transplantation is shown graphically as a function of the recipient's treatment mode. [Figure 3D] The graphs show the results of in vivo studies of CD45-ADC conditioning prior to minor mismatch allogeneic transplantation of Balb / cCD45.1 donor cells into DBA / 2 recipient mice. The percentage of T cell chimerism in transplant recipients at 4, 8, 12, and 16 weeks post-transplantation is shown graphically as a function of the recipient's treatment mode. [Figure 4A] The graph shows the results of in vivo studies of CD45-ADC conditioning prior to complete mismatch allogeneic transplantation of Balb / cCD45.1 donor cells into C57BL / 6 recipient mice. The percentage of donor chimerism detected in the blood using CD45.1+ antigen 4 and 8 weeks after transplantation is shown graphically as a function of the recipient's treatment mode. [Figure 4B]The graph shows the results of in vivo studies of CD45-ADC conditioning prior to complete mismatch allogeneic transplantation of Balb / cCD45.1 donor cells into C57BL / 6 recipient mice. The graph also shows the percentage of peripheral donor bone marrow chimerism in transplant recipients 4 and 8 weeks after transplantation as a function of the recipient's treatment mode. [Figure 4C] The graph shows the results of in vivo studies of CD45-ADC conditioning prior to complete mismatch allogeneic transplantation of Balb / cCD45.1 donor cells into C57BL / 6 recipient mice. The percentage of B cell chimerism in transplant recipients 4 and 8 weeks after transplantation is shown graphically as a function of the recipient's treatment mode. [Figure 4D] The graph shows the results of in vivo studies of CD45-ADC conditioning prior to complete mismatch allogeneic transplantation of Balb / cCD45.1 donor cells into C57BL / 6 recipient mice. The graph also shows the percentage of T cell chimerism in transplant recipients 4 and 8 weeks after transplantation as a function of the recipient's treatment mode. [Figure 4E] The graphs show the results of an in vivo study of CD45-ADC conditioning prior to complete mismatch allogeneic transplantation of Balb / cCD45.1 donor cells into C57BL / 6 recipient mice. Similar to the studies in the complete mismatch mouse model shown in Figures 4B-4D, the graphs show the results of an in vivo study that monitored donor chimerism for 22 weeks post-transplant. C57Bl / 6 (H-2b, CD45.2+) mice were conditioned with isotype-ADC or CD45-ADC (5 mg / kg) and transplanted with Balb / c (H-2d, CD45.1+) bone marrow. Donor cells were detected in peripheral blood 4 weeks after transplantation using the CD45.1+ antigen and maintained for 22 weeks (upper left). Reconstitution was multi-lineage (lower left and center panels). In CD45-ADC-conditioned mice, terminal splenic (upper right) and thymic (lower right) chimerism was similar to that of TBI. *p<0.05 vs TBI; #p<0.05 vs CD45-ADC; ANOVA and Tukey's multiple comparison tests were performed post-hoc. [Figure 5] The graph shows the results of an ex vivo killing assay using CD45-ADC in mouse HSCs cultured in a medium containing stem cell factor (SCF) after strain depletion. It displays the number of CD45-containing live bone marrow (BM) cells, total Lin- BM cells, and total LKS (Lin- Sca-1+ c-Kit+) BM cells. [Figure 6] The graph shows the CD45-ADC plasma antibody concentration as a function of time after administration of a single dose of 3 mg / kg or 6 mg / kg of CD45-ADC, or a divided dose of 3 mg / kg of Q2D, to mice. [Figure 7A] The graphs show the results of in vivo studies of CD45-ADC conditioning prior to minor mismatch allogeneic transplantation of CByJ.SJL(B6)-Ptprca / J(CD45.1) donor cells into DBA / 2(CD45.2) recipient mice. The graphs also show the percentages of B220+, CD11B+, and CD3+ peripheral blood chimerism at week 16 in mice treated with IRR, Iso-ADC, CD45-ADC, or CD45-ADC in combination with anti-CD4 and anti-CD8 antibodies at weeks 0, 4, 8, 12, and 16. [Figure 7B] The graph shows the results of an in vivo study of CD45-ADC conditioning prior to minor mismatch allogeneic transplantation of CByJ.SJL(B6)-Ptprca / J(CD45.1) donor cells into DBA / 2(CD45.2) recipient mice. The graph also shows the peripheral blood composition (percentage of B220+, CD11B+, and CD3+ peripheral blood chimerism) in mice 16 weeks after treatment under specified conditions. [Figure 7C]The graph shows the results of an in vivo study of CD45-ADC conditioning prior to minor mismatch allogeneic transplantation of CByJ.SJL(B6)-Ptprca / J(CD45.1) donor cells into DBA / 2(CD45.2) recipient mice. The graph also shows the depletion levels of LSK (Lin-Sca-1+ c-Kit+) cells, LT-HSCs, and ST-HSCs in the bone marrow extracted from mice 3 days after treatment under specified conditions, measured by percentage abundance and cell number / femur. [Figure 8A] The graphs show the results of in vivo studies of CD45-ADC conditioning prior to complete mismatch allogeneic transplantation of CByJ.SJL(B6)-Ptprca / J(CD45.1) donor cells into C57Bl / 6(CD45.2) recipient mice. The graphs also show the depletion levels of LSK (Lin-Sca-1+ c-Kit+) cells, LT-HSCs, and ST-HSCs in the bone marrow extracted from mice 3 days after treatment with iso-ADC or CD45-ADC (single dose of 2×3 mg / kg, or 4 mg / kg, 5 mg / kg, or 6 mg / kg), measured by percentage abundance and cell number / femur. Treatment with CD45-ADC in combination with 9 Gy TBI, 0.5 Gy TBI, or under naive conditions was also evaluated. [Figure 8B] The graph shows the results of in vivo studies of CD45-ADC conditioning prior to complete mismatch allogeneic transplantation of CByJ.SJL(B6)-Ptprca / J(CD45.1) donor cells into C57Bl / 6(CD45.2) recipient mice. The graph also shows the percentage of donor chimerism detected in the blood using CD45.1+ antigen 4 and 8 weeks after transplantation as a function of the recipient's treatment mode. [Figure 8C]The graphs show the results of in vivo studies of CD45-ADC conditioning prior to complete mismatch allogeneic transplantation of CByJ.SJL(B6)-Ptprca / J(CD45.1) donor cells into C57Bl / 6(CD45.2) recipient mice. The graphs also show the percentages of B220+, CD11B+, and CD3+ peripheral blood chimerism at week 16 in mice in the specified treatment group at week 4. [Modes for carrying out the invention]

[0028] Provided herein are CD45-targeted treatments useful for monotherapy conditioning. This is a partial (for example, an anti-CD45 antibody or ADC), which allows the patient to receive immunosuppressants. Without using additional conditioning agents such as, Preparations are made for this. Such procedures promote the engraftment of allogeneic hematopoietic stem cells. According to the method described herein, the patient will be CD45-targeted in the absence of immunosuppressants. Receiving a partial dose (e.g., anti-CD45 antibody, its antigen-binding portion, or ADC) This can be used to condition for allogeneic hematopoietic stem cell transplantation therapy. CD45 The targeting portion (e.g., anti-CD45 antibody, its antigen-binding portion, or ADC) is located in the hematopoietic stem cell. It can bind to the CD45 antigen expressed by hematopoietic cells, including cell cells and mature immune cells. It is possible. As described herein, the CD45 targeting portion (e.g., an antibody, or so The antigen-binding portion of the antibody is designed so that the CD45 targeting portion binds to the toxin (for example, antibody-drug compound It is covalently conjugated to the cytotoxic substance (to form an ADC). Obtain a CD45-targeting moiety capable of binding to CD45 (e.g., ADC, antibody, etc.). The antigen-binding portion (or drug-antibody conjugate) of the hematopoietic stem cell transplantation therapy is required. Administering it to a patient, for example, selectively depletes endogenous hematopoietic stem cells, thereby... By creating space that can be filled by genetic hematopoietic stem cell transplantation, the life of allogeneic hematopoietic stem cell grafts It can promote implantation. In an exemplary embodiment, the transplant is a complete mismatch. It includes allogeneic hematopoietic stem cells.

[0029] definition Where used herein, the term “about” means 5% above or below the stated value. It refers to a value within a certain range.

[0030] As used herein, the term “homogeneous” means, when used in the context of transplantation, a genetic. Cells (or tissues, or organs) transplanted from genetically different donors to a recipient of the same species. It is used to define ).

[0031] As used herein, the term “autologous” means that the donor and recipient are the same person. It refers to cells or grafts that are elephants.

[0032] As used herein, the term “heterogeneous” means that the donor and recipient are of different species. It refers to cells.

[0033] As used herein, the term “immune cells” refers to cells of hematopoietic origin and immune response. It is intended to include, but is not limited to, cells involved in immune cells, such as T cells. This includes, but is not limited to, natural killer (NK) cells. Ra cells are well known in the art. In one embodiment, natural Killer cells include cell lines such as NK-92 cells. Further examples of NK cell lines include Examples include NKG, YT, NK-YS, HANK-1, YTS cells, and NKL cells. Immune cells can be homogeneous or autologous.

[0034] As used herein, the term “CD45-targeted portion” means a portion bound to CD45. This refers to molecules that can do this, including, for example, antibodies, antibody fragments, or aptamers. In that embodiment, the CD45 targeting portion is a nanoparticle (for example, on the surface of a nanoparticle) It binds to and forms targeted nanoparticles (e.g., drug-carrying nanoparticles such as toxin-carrying nanoparticles). It will be accomplished.

[0035] As used herein, the term “antibody” means an antibody that specifically binds to a particular antigen. This refers to an immunoglobulin molecule that is immunologically reactive with a specific antigen, or as an antibody. These include monoclonal antibodies, polyclonal antibodies, and multispecific antibodies (for example, bispecific antibodies). This includes, for example, genetically modified antibodies and modified forms of other antibodies, but is not limited to, the body, genetically modified antibodies, and other modified forms of antibodies. However, chimeric antibodies, humanized antibodies, heteroconjugate antibodies (e.g., double, triple, and quadruple conjugate antibodies) Specific antibodies, diabodies, triabodies, and tetrabodies), and antibody fragments ( That is, antigen-binding fragments of antibodies, for example, Fab', F(ab')2, Fab, Fv, r Examples include IgG and scFv fragments (as long as they exhibit the desired antigen-binding activity), but these Not limited to this.

[0036] The antibodies in this disclosure are generally isolated antibodies or recombinant antibodies. As used herein, this refers to polypeptides, such as antibodies, that express them. This refers to something identified, isolated, and / or recovered from cells or cell cultures. The isolated antibody is prepared by at least one purification step. Therefore, "Isolated antibodies" refer to antibodies that substantially do not contain other antibodies with different antigen specificities. For example, an isolated antibody that specifically binds to CD45 is not specific to antigens other than CD45. It contains virtually no antibodies that bind to it.

[0037] As used herein, the term "monoclonal antibody" refers to any eukaryote, prokaryote. From a single clone, including an object or phage clone, to a product available in the art This refers to antibodies obtained by any known means, and produced via hybridoma technology. The antibodies produced are not limited to monoclonal antibodies. Monoclonal antibodies useful in this disclosure include hybridomas. This includes the use of recombinant and phage display technologies, or combinations thereof. It can be prepared using a wide variety of techniques known in the art. In particular Unless otherwise specified, the term “monoclonal antibody” (mAb) refers to an antibody specifically targeting a protein. Intact molecules and antibody fragments capable of heterobinding (e.g., Fab and F(ab') ) means including both fragments. When used herein, Fab and The F(ab')2 fragment refers to an antibody fragment lacking the Fc fragment of the intact antibody. In one embodiment... In this case, the antibody fragment contains an Fc region.

[0038] Generally, antibodies consist of a heavy chain and a light chain containing an antigen-binding region. Each heavy chain has a variable heavy chain region. It consists of a region (abbreviated as HCVR or VH in this specification) and a heavy chain constant region. The constant region of the chain consists of three domains CH1, CH2, and CH3. Each light chain is light It consists of a chain variable region (abbreviated as LCVR or VL herein) and a light chain constant region. The light chain constant region consists of one domain CL. The VH region and VL region are A region of high variability called the Complementarity Determination Region (CDR), and the frames scattered between them It can be further divided into a more conserved region called the FR region. Each VH and VL consists of three CDRs and four FRs, from the amino terminus to the carboxyl terminus. The order is as follows: FR1, CDR1, FR2, CDR2, FR3, CDR3, FR4 It is arranged as follows. The variable regions of the heavy and light chains contain binding domains that interact with the antigen. The constant region of an antibody is found in various cells of the immune system (e.g., effector cells) and classical The binding of immunoglobulins to host tissues or factors, including the first component of the complement system (Clq). It can be used as an intermediary.

[0039] The terms “antigen-binding fragment” or “antigen-binding moiety” of an antibody are used herein. In this context, it refers to one or more parts of an antibody that retain the ability to specifically bind to a target antigen. The antigen-binding function can also be performed by fragments of full-length antibodies. Antibody fragments can be, for example, Fa b, F(ab')2, scFv, Diabody, Triabody, Affibody, Nanobody It may be an aptamer or domain antibody. The term "antigen-binding fragment" of an antibody Examples of included binding fragments include (i) VL, VH, CL, and CH1 domains. (ii) a monovalent fragment, the Fab fragment; (ii) linked by disulfide bridges in the hinge region F(ab')2 fragment, a divalent fragment containing two Fab fragments; (iii)VH and (iv) Fd fragment consisting of the CH1 domain; (iv) VL and VH domains of a single arm of the antibody (v) a fragment of Fv; (v) a dAb containing VH and VL domains; (vi) from the VH domain This is a dAb fragment (for example, Ward et al., Nature 341:544-5 See 46, 1989); (vii) dAb consisting of a VH or VL domain; (viii) ) Isolated complementarity determination regions (CDRs); and (ix) optionally, by a composite linker Two or more (for example, two, three, four, five, or six) isolated that can be connected The combination of CDRs is one example, but is not limited to these. Furthermore, two Fv fragments The domains VL and VH are encoded by separate genes, but recombinant method Using this, a single protein chain is formed by the pairing of the VL and VH regions to create a monovalent molecule. It can be connected by a linker that enables the creation of (single-chain antibody Fv(sc Known as Fv); for example, Bird et al., Science 242:4 23-426, 1988 and Huston et al., Proc. Natl. Aca (See d.Sci.USA 85:5879-5883, 1988). These antibody fragments These can be obtained using the prior art known to those skilled in the art, and these fragments are inter The usefulness can be screened in the same manner as for ct antibodies. The antigen-binding fragment is By recombinant DNA technology, enzymatic or chemical cleavage of intact immunoglobulins, Or, in certain cases, the chemical peptide synthesis procedure known in the art is Therefore, it can be manufactured.

[0040] The "aptamers" used in the compositions and methods disclosed herein are peptides or It contains an aptamer molecule made from any of the nucleotides. In a particular embodiment, In other words, aptamers are small nucleotide polymers that bind to specific molecular targets. Rheotide aptamers can be single-stranded or double-stranded nucleic acid molecules (DNA or RNA). However, DNA-based aptamers are most commonly double-stranded. There is no strict definition, but aptamer molecules are most commonly 15-40 nucleotides long. In other embodiments, the aptamer is a peptide aptamer. - It shares many properties with nucleotide aptamers (for example, it is small in size, (The ability to bind to target molecules with high affinity), used to generate nucleotide aptamers. Selection methods that have a similar principle to the one described, for example, Baines and Colas.20 06. Drug Discov Today. 11(7-8):334-41; and Bi ckle et al.2006.Nat Protoc.1(3):1066-91( Aptamers can be produced by (as incorporated herein by reference). Aptamers can be produced by various technologies It can be produced using, but originally, in vitro selection (Ellington and Szostak.(1990)Nature.346(6287):818-22) and SELEX method (Ligand phylogenetic evolution method by exponential enrichment) (Schneider Using et al. 1992. J Mol Biol. 228(3):862-9) These were developed, and their contents are incorporated herein by reference. Aptamer Other methods for creating and using it are publicly available, for example, Klussmann, The Aptamer Handbook:Functional Oligonuc leotides and Their Applications.ISBN:978 -3-527-31059-3;Ulrich et al.2006.Comb Ch em High Throughput Screen 9(8):619-32;Ce rchia and de Franciscis.2007.Methods Mol Biol.361:187-200;Ireson and Kelland.200 6.Mol Cancer Ther.2006 5(12):2957-62; 5,582,981; 5,840,867; 5,756,291; No. 6,261,783; No. 6,458,559; No. 5,792,613; No. Patent No. 6,111,095; and U.S. Patent Application Publication No. US20070009476A Publication No. 1; U.S. Public Publication No. US20050260164A1; U.S. Public Publication No. 7,960,102 ; and U.S. Public Publication No. US20040110235A1, all of which refer to More herein by reference.

[0041] As used herein, "anti-CD45 antibody" or "antibody that binds to CD45" The term refers to the usefulness of antibodies as diagnostic and / or therapeutic agents when they target CD45. This refers to an antibody that can bind to CD45 with sufficient affinity.

[0042] As used herein, the term “diabody” refers to two polypeptide chains. This refers to the bivalent antibody contained, in which case each polypeptide chain is V on the same peptide chain. H Dome In and V L Short linkers that prevent intramolecular association of domains (for example, five amino acids) V connected by a linker (composed of acid) H Domain and V L Includes the domain. This arrangement allows each domain to pair with a complementary domain on another polypeptide chain, A monimer structure is formed. Therefore, the term "tribody" refers to a structure with three peptides. This refers to a trivalent antibody containing a nucleotide chain, each of which contains a nucleotide within the same peptide chain. H Domain and V L An extremely short linker (e.g., 1-2 amino acids) that causes intramolecular association of domains One V connected by a linker (composed of acid) H Domain and one V L Dome It contains yin. In order for the peptide constructed in this way to fold into its original structure, Typically, the V of adjacent peptide chains H Domain and V L The domains are spatially close to each other. The product is trimerized so that it is arranged in a certain way (for example, Holliger et al., Proc). (See Natl.Acad.Sci.USA 90:6444-48, 1993).

[0043] As used herein, the term “bispecific antibody” means at least two different A monoclonal antigen can bind to two different epitopes. This refers to antibodies, such as human or humanized antibodies. For example, one of the binding specificities is CD45. One is directed towards an epitope on a hematopoietic stem cell surface antigen, while the other is directed towards a different hematopoietic stem cell surface antigen. The epitopes or other cell surface proteins, particularly those involved in signaling pathways that enhance cell growth. It can specifically bind to receptors or receptor subunits involved in the pathway. In some embodiments, binding specificity is unique and non-overlapping epidermal on the same target antigen. It may be directed towards a tope (i.e., a biparatope antibody). "Intact" or " "Full-length" antibodies, as used herein, are interconnected by disulfide bonds. This refers to an antibody that has two heavy (H) polypeptides and two light (L) polypeptides. Each heavy chain has a variable heavy chain region (abbreviated as HCVR or VH herein) and a constant heavy chain region. It is composed of regions. The heavy chain constant region consists of three domains: CH1, CH2, and CH3. Each light chain has a light chain variable region (abbreviated herein as LCVR or VL) and light It consists of a chain constant region. The light chain constant region consists of one domain CL. VH The region and the VL region are a highly variable region called the complementarity determination region (CDR), and between them Further dividing into more preserved areas called scattered framework areas (FRs). This is possible. Each VH and VL consists of three CDRs and four FRs, and the amino terminus is Towards the carboxyl terminus, in the following order: FR1, CDR1, FR2, CDR2, FR It is sequenced at 3, CDR3, FR4. The variable regions of the heavy and light chains interact with the antigen. It contains a binding domain. The constant region of the antibody is connected to various cells of the immune system (e.g., Effect). Immunity to host tissues or factors, including ter cells and the first component of the classical complement system (Clq). It can mediate the binding of disease globulins.

[0044] As used herein, the term “complementarity-determining region” (CDR) refers to the light chain of an antibody. This refers to the hypervariable region present in both the and heavy chain variable domains. The preserved portion is called the framework region (FR). This defines the hypervariable region of the antibody. The amino acid position is determined according to the context and various definitions known in the relevant art. It can change. Some locations within the variable domain are considered hybrid hypervariable locations. In some cases, these positions can be considered to be within a hypervariable region under a certain set of criteria. These are possible, but under a different set of criteria they are considered outside the hypervariable region. One or more of these locations may be present in the hypervariable extension region. The body may contain modifications to these hybrid hypervariable positions. Variability of natural heavy and light chains. The domain consists of four frames, primarily in a beta-sheet shape, connected by three CDRs. Each contains a β-sheet region, and the CDR forms loops connecting the β-sheet structures, several In this case, it forms part of the β-sheet structure. The CDR in each chain is in the framework region. So, they are in close proximity in the order of FR1-CDR1-FR2-CDR2-FR3-CDR3-FR4. They are retained together and, along with the CDR of the other antibody chain, contribute to the formation of the antibody's target binding site. (Kabat et al., Sequences of Proteins of I mmunological interest,National Institute (See *of Health*, Bethesda, MD., 1987). A certain implementation In this context, the numbering of immunoglobulin amino acid residues is Ka unless otherwise specified. This is done according to the immunoglobulin amino acid residue numbering system by bat et al. (however , any antibody numbering scheme, for example, IMGT and C (Hothias can also be used.)

[0045] The term "specifically binds" as used herein refers to proteins in general. Instead, antibodies (or ADCs) recognize and bind to specific protein structures (epitopes). This refers to the ability. If the antibody is specific to epitope "A", then the labeled "A" and In a reaction involving antibodies, a molecule containing epitope A (or free, unlabeled A) When present, the amount of labeled A that binds to the antibody decreases. For example, an antibody is labeled If this is the case, and the corresponding unlabeled antibody competes with it and may move away from its target, then the antibody is "specific" to the target. "Bound to the target." In one embodiment, the antibody binds to the target at least about 10 -4 M, approx. 1 0 -5 M, about 10 -6 M, about 10 -7 M, about 10 -8 M, about 10 -9 M, about 10 -10 about M, 10 -11 Approximately M, approximately 10 -12 M or less (less is approximately 10) - 12 A number smaller than that, for example, 10 -13 (meaning) K D If present, the antibody is marked It specifically binds to antigens expressed by hematopoietic stem cells, such as CD45. In embodiments, the term "specifically binds" means at least about 1 × 10⁻⁶ -6 M, 1 x 10 -7 M, about 1 x 10 -8 M, about 1 x 10 -9 M, about 1 x 10 -10 M, approx. 1×1 0 -11 M, about 1 x 10 -12 Binds to the antigen at M or higher Kd and / or non-binding to the antigen. The ability of antibodies to bind to an antigen with an affinity at least twice as high as their affinity for the specific antigen. It refers to force. In one embodiment, K D This follows standard biolayer interferometry (BLI). This is how it is determined. However, antibodies specifically bind to two or more antigens that are sequence-related. It should be understood that this may be possible. For example, in one embodiment, the antibody is Antigens, for example, both human and non-human (e.g., mouse or non-human primate) CD45. It can specifically bind to orthologues of [the substance].

[0046] As used herein, the term “chimeric” antibody refers to non-rat or mouse antibodies, etc. Variable sequences derived from human immunoglobulins and sequences typically selected from human immunoglobulin templates. This refers to an antibody that has a constant region of human immunoglobulin. The law is known in the relevant technical field. For example, Morrison, 1985, Sc ience 229(4719):1202-7;Oi et al.,1986,Bi oTechniques 4:214-221;Gillies et al.,198 5, J. Immunol. Methods 125:191-202; U.S. 5, 8 See Nos. 07,715; Nos. 4,816,567; and Nos. 4,816,397. I would like to know the terms "Fc", "Fc region", "Fc domain", and "IgG Fc domain". When used herein, the term refers to an immunoglobulin, for example, a portion of an IgG molecule. This refers to the part related to the crystallizable fragment obtained by papain digestion of the IgG molecule. The Fc region is the C-terminus of the two heavy chains of the IgG molecule linked by disulfide bonds. It includes the side half. The Fc region does not have antigen-binding activity, but the carbohydrate portion and complement and It contains a binding site for Fc receptors, including FcRn receptors (see below). For example, The Fc domain is the second constant domain CH2 (e.g., EU position 231~ of human IgG1). (340 residues) and the third constant domain CH3 (e.g., EU position 341 of human IgG1) It contains ~447 residues. When used herein, the Fc domain is "lower hind This includes the "di region" (for example, residues at EU positions 233-239 of human IgG1).

[0047] Fc can refer to this region alone, or to the antibody, the antigen-binding portion of the antibody, or Fc This region can also refer to the area within the context of fusion proteins. Many locations in the Fc domain, for example. For example, although not limited to them, EU locations 270, 272, 312, 315, 356, and Since polymorphism has been observed in 358, the sequence presented in this application and the relevant technical field Slight differences may exist between this sequence and the known sequence. Therefore, "wild-type IgG" The "Fc domain" or "WT IgG Fc domain" is any naturally occurring Ig This refers to the G Fc region (i.e., any allele). Human IgG1, IgG2, IgG3 and The heavy chain sequence of IgG4 can be found in many sequence databases, for example, Un In the iprot database (www.uniprot.org), each is listed as P0185. 7(IGHG1_HUMAN), P01859(IGHG2_HUMAN), P0186 Accessory 0 (IGHG3_HUMAN) and P01861 (IGHG1_HUMAN) It can be found by its serial number.

[0048] The terms “modified Fc region” or “variant Fc region” are used herein. In this case, substitution, deletion, or insertion of one or more amino acids introduced at any position within the Fc domain. Or it refers to an IgG Fc domain that includes modifications. In a particular embodiment, variant I The gG Fc domain contains one or more amino acid substitutions, thereby comprising one or more amino Compared to the wild-type Fc domain without acid substitution, Fc gamma R and / or C1q The binding affinity decreases or disappears. Furthermore, Fc binding interactions are not limited to However, antibody-dependent cell-mediated cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) It is essential for various effects functions and downstream signaling events. Therefore, in a certain embodiment, an antibody containing a variant Fc domain (for example, an antibody, Fusion proteins (or conjugates) are, for example, naturally occurring at the corresponding positions in the Fc region. Although it has the same amino acid sequence, such as an unmodified Fc region containing the present amino acid residue, Compared to the corresponding antibody which does not contain one or more amino acid substitutions, deletions, insertions, or modifications, For at least one Fc ligand (e.g., Fc gamma R), a modified binding parent It may exhibit a harmonious relationship.

[0049] The variant Fc domains described herein are based on the amino acid modifications that constitute them. This is how it is defined. For all amino acid substitutions considered herein with respect to the Fc region, The numbering always follows the EU index in Kabat. Therefore, for example D265C is the aspartic acid (D) at EU position 265 relative to the parent Fc domain. ) is an Fc variant substituted with cysteine ​​(C). Similarly, for example, D265C / L234A / L235A are based on the parent Fc domain, EU position 265 (D to C ), 234 (L to A), and 235 (L to A) are substitutions in the variant Fc barrier. The variant is defined as the final amino acid combination at the mutated EU amino acid position. It can also be specified according to the organism. For example, the L234A / L235A mutant is "L It can be called "ALA". Further examples include E233P, L234V, L235A. The delG236 (deletion of 236) mutant can be called "EPLVLAdelG". As yet another example, the variants of I253A, H310A, H435A are "IHH" and It can be called. Note that the order in which substitutions are provided is arbitrary.

[0050] The terms "Fc gamma receptor" or "Fc gamma R" are used herein in the context of the context of this document. In combination, it binds to the Fc region of the IgG antibody and the protein encoded by the Fc gamma R gene. This refers to any member of the quality family. In humans, this family includes isophore. Fc gamma R, including Fc gamma RIa, Fc gamma RIb, and Fc gamma RIc. I(CD64); Fc gamma RIIa isoform (allotypes H131 and R13) (including 1), Fc gamma RIIb (Fc gamma RIIb-1 and Fc gamma RIIb-2) (including), and Fc gamma RII (CD32) including Fc gamma RIIc; and isof Form's Fc Gamma RIIIa (including Arotype V158 and F158) and Fc Gun Ma RIIIb (Arotype Fc Gamma RIIIb-NA1 and Fc Gamma RIIIb-N Fc gamma RIII (CD16), including A2, and any undiscovered human Fc This includes gamma-R or Fc-gamma-R isoforms or allotypes, but these include Not limited. Fc gamma R is not limited, but humans, mice, rats, rabbits It can be derived from any organism, including monkeys and primates. Mouse Fc gamma R contains Fc gamma RI (CD64), Fc Gamma RII (CD32), Fc Gamma RIII (CD16), and Fc Gamma RIII-2 (CD16-2), and any undiscovered mouse Fc Gamma R This includes, but is not limited to, Fc gamma R isoforms or allotypes. do not have.

[0051] The term "effector function" as used herein refers to the Fc domain and Fc This refers to biochemical events resulting from receptor interactions. Effector functions include ADCC. This includes, but is not limited to, the ADCP and CDC. "Effector cells" are cells that express one or more Fc receptors and have one or more effector functions. It refers to immune system cells that mediate [the process]. It is not limited to effector cells, however... Monocytes, macrophages, neutrophils, dendritic cells, eosinophils, mast cells, platelets, B cells, large Granule lymphocytes, Langerhans cells, natural killer (NK) cells, and gamma delta cells. T cells are included, but are not limited to, those found in humans, mice, rats, rabbits, and monkeys. It can originate from any organism, including those mentioned above.

[0052] The terms "silent," "silence," or "silencing" as used herein and The term refers to an antibody having a modified Fc region as described herein, and which is an Fc gamma receptor Binding to the body (FcγR) is the same antibody containing an unmodified Fc region binding to FcγR. This refers to antibodies whose binding is reduced compared to the combined state (for example, the binding to FcγR is reduced, for example, BLI). When measured by this method, the binding and ratio of the same antibody containing an unmodified Fc region to FcγR All, at least 70%, at least 80%, at least 90%, at least 95%, (Reduction of at least 98%, at least 99%, or 100%). Several implementations In this state, the Fc silence antibody does not have detectable binding to FcγR. Modified Fc The binding of antibodies containing a region to FcγR is performed using various techniques known in the art. For example, although not limited to them, equilibrium methods (for example, enzyme-linked immunosorbent assay (ELISA)) );KinExA,Rathanaswami et al.Analytical B iochemistry, Vol.373:52-60, 2008; or radioimmuno Assay (RIA), or surface plasmon resonance assay or other mechanisms Inetics-based assays (e.g., BIACORE® analysis or Octe t(trademark) analysis (forteBIO), as well as indirect binding assays, competitive binding assays Fluorescence resonance energy transfer (FRET), gel electrophoresis and chromatography (examples) For example, it can be determined using other methods such as gel filtration. These methods and Other methods may involve using labeling for one or more of the components being tested, and / or Various types of labels, including but not limited to color-developing labels, fluorescent labels, luminescent labels, or isotopic labels. Various detection methods can be employed. For a detailed explanation of binding affinity and kinetics, see below. This focuses on the interaction between antibodies and immunogens. Paul, WE, ed., Fundam ental Immunology,4th Ed.,Lippincott-Rave It can be found in n, Philadelphia (1999). Competitive binding assay One example involves incubating the target antibody and labeled antigen in the presence of gradually increasing amounts of unlabeled antigen. This is a radioimmunoassay that involves detecting antibodies bound to a labeled antigen. The affinity and binding offrate of the target antibody to the source were determined using scatchard plot analysis. Therefore, it can be determined from the data. Competition with secondary antibodies is determined by radioimmunoassay. It can also be determined by using this method. In this case, the antigen is conjugated with the labeled compound. The target antibody is incubated with the unlabeled secondary antibody in gradually increasing amounts.

[0053] As used herein, the term “identical antibody containing an unmodified Fc region” is used in the column The amino acid substitutions listed (e.g., D265C, L234A, L235A, and / or H) It lacks 435A, but otherwise has the same amino acid sequence as the comparable Fc-modified antibody. It refers to antibodies that produce [the substance].

[0054] The term "antibody-dependent cell-mediated cytotoxicity" or "ADCC" refers to the Fc domain. Polypeptides containing antibodies, for example, antibodies that target certain cytotoxic cells (e.g., mainly NK cells). It binds to Fc receptors (FcRs) present in neutrophils and macrophages, and these cells Cellular toxic effector cells specifically bind to "target cells" that possess the antigen, and then... This refers to a form of cytotoxicity that can kill target cells with cytotoxicity. (Hogarth) et al.,Nature review Drug Discovery 201 2,11:313) In addition to antibodies and their fragments, they specifically bind to target cells that possess antigens. Other polypeptides containing an Fc domain that have the capability, for example, Fc fusion proteins and Fc conjugate proteins are expected to exert cell-mediated cytotoxicity. It is illustrated.

[0055] For brevity, cell-mediated cytotoxicity caused by the activity of polypeptides containing an Fc domain. Harmful activity is also referred to herein as ADCC activity. Lysis of target cells by ADCC. The ability of any particular polypeptide of this disclosure to mediate can be assayed. To evaluate DCC activity, the target polypeptide (e.g., an antibody) is subjected to an immunoeffector. - When added to target cells in combination with other cells, it induces cell lysis of the target cells. The solution is generally that the label (e.g., radioactive substrate, fluorescent dye, or native intracellular protein) It is detected by its release from lysed cells. Such assays are useful for Ef. Ejector cells include peripheral blood mononuclear cells (PBMCs) and natural killer (NK) cells. Examples include the Bruggemann assay. et al.,J.Exp.Med.166:1351(1987);Wilkins on et al., J. Immunol. Methods 258:183 (2001 );Patel et al.,J.Immunol.Methods 184:29( As described in 1995). Alternatively or additionally, the ADCC activity of the antibody of interest is, for example, For example, Clynes et al.,Proc.Natl.Acad.Sci.USA In vivo evaluation is performed using animal models such as those disclosed in 95:652 (1998). It is possible.

[0056] As used herein, the terms "condition" and "conditioning" are used in this specification. The term refers to the process by which a patient prepares to receive a transplant, such as a transplant containing hematopoietic stem cells. This refers to the condition. Such treatment promotes the engraftment of hematopoietic stem cells (for example, conditioning). Viable cells in blood samples isolated from patients after the initial procedure and subsequent hematopoietic stem cell transplantation. (Inferred from the sustained increase in the amount of hematopoietic stem cells). According to the method described herein, C ADCs and antibodies capable of binding to antigens expressed by hematopoietic stem cells such as D45 Alternatively, by administering the antigen-binding portion to the patient, the patient can undergo hematopoietic stem cell transplantation therapy. Conditioning is performed. As described herein, the antibody forms the ADC. It can be covalently conjugated to cytotoxic substances, such as one of the aforementioned antigens. ADCs, antibodies, or their antigen-binding moieties that can bind to the above, in hematopoietic stem cell transplantation. Administering this therapy to patients who require it could, for example, selectively deplete endogenous hematopoietic stem cells. This creates space that can be filled by exogenous hematopoietic stem cell transplantation, thereby providing hematopoietic stem cells. This can promote the engraftment of cyst grafts.

[0057] As used herein, the terms “effective dose” or “therapeutic dose” are used as desired. This refers to an amount sufficient to achieve the desired result or to have an effect on autoimmune diseases or cancer. .

[0058] As used herein, the term "half-life" refers to the time it takes for an antibody drug to reach a plasma concentration in the body. This refers to the time it takes for the concentration to decrease by half or 50%. This 50% decrease in serum concentration is This reflects the amount of drug circulating.

[0059] As used herein, the term "human antibody" refers to an immunoglobulin of the human germline. It is intended to contain antibodies having variable and constant regions derived from the brin sequence. Human Antibodies are amino acid residues not encoded by human germline immunoglobulin sequences. (For example, random mutagenesis or site-directed mutagenesis in vitro, (May include mutations introduced by gene rearrangement or somatic mutations in vivo) However, the term "human antibody" as used herein refers to a different antibody, such as one from a mouse. CDR sequences derived from the germline of mammalian species were grafted onto human framework sequences. It is not intended to contain antibodies that have been recombinantly expressed in human cells (e.g., recombinant expression). ) or functionally reconstituted human immunoglobulins (heavy chain and / or light chain) (by non-human animals, prokaryotic cells, or eukaryotic cells capable of expressing genes) It can be produced. If the human antibody is a single-chain antibody, it has linkages that are not present in natural human antibodies. Petit can be included. For example, Fv connects the variable region of the heavy chain and the variable region of the light chain. It contains linker peptides such as glycine or other amino acid residues, ranging from 2 to approximately 8. Such linker peptides are considered to be of human origin. Human antibodies This involves using a phage display with an antibody library derived from human immunoglobulin sequences. Human antibodies can be produced by various methods known in the art, including Method I. Furthermore, although they cannot express functional endogenous immunoglobulins, they possess human immunoglobulin genes. It can also be created using transgenic mice that can express the gene (for example) PCT Publication No. WO1998 / 24893; same No. WO1992 / 01047; same No. WO1996 / 34096; WO1996 / 33735; US 5,413 , No. 5,625,126; No. 5,633,425; No. 5,569, No. 825; No. 5,661,016; No. 5,545,806; No. 5,814,3 No. 18; No. 5,885,793; No. 5,916,771; and No. 5,939, (See No. 598).

[0060] The "humanized" form of non-human (e.g., mouse) antibodies is derived from the smallest non-human immunoglobulins. It is a chimeric immunoglobulin containing a limited sequence. Generally, humanized antibodies contain at least 1 Typically, this includes almost all of the two variable domains and all or almost all of the CDR region. The CDR region corresponds to that of non-human immunoglobulins, and all or almost all of the FR region corresponds to that of human immunoglobulins. This is the FR region of the immunoglobulin sequence. Humanized antibodies also have a constant immunoglobulin region (F c) at least a portion of, typically at least one of the human immunoglobulin consensus sequences It may include parts. Methods for humanizing antibodies are known in the art. For example, Riech mann et al.,1988,Nature 332:323-7;Queen et al. U.S. Patent Nos. 5,530,101; 5,585,089; 5,693,7 No. 61; Nos. 5,693,762; and Nos. 6,180,370; EP239400 PCT Publication WO91 / 09967; US Patent No. 5,225,539; EP59210 6;EP519596;Padlan,1991,Mol.Immunol.,28:4 89-498;Studnicka et al.,1994,Prot.Eng.7: 805-814;Roguska et al.,1994,Proc.Natl.Ac See ad.Sci.91:969-973 and U.S. Patent No. 5,565,332. I want to be treated that way.

[0061] As used herein, the term “engraftability” means that hematopoietic stem cells and progenitor cells are able to engraft. Whether the cells are naturally circulating or donated through transplantation, tissue reconstruction This term is used to refer to the ability to homing cells to tissues and target groups. This encompasses all events related to or leading up to engraftment, such as the colonization of cells within the tissue. Includes. Engraftment efficiency or grafting rate is any clinically acceptable para known to those skilled in the art. It can be evaluated or quantified using a meter, for example, competitive reconstruction units (CRUs). ) evaluation; in tissue(s) where stem cells have homed, colonized, or engrafted. Marker uptake or expression; or the progression of the target through disease progression, hematopoietic stem cells and This may include assessing the survival of progenitor cells or the survival of the recipient. Engraftment is also a factor in transplantation. Engraftment can also be determined by measuring the number of white blood cells in the peripheral blood during a later period. Furthermore, by measuring the recovery of bone marrow cells by donor cells in bone marrow aspirate samples, It can be evaluated.

[0062] As used herein, the term “hematopoietic stem cells” (“HSCs”) means self-replicating stem cells. The ability to do so, and various lineages, for example, not limited to, but granulocytes (for example, promyelocytes, Neutrophils, eosinophils, basophils), red blood cells (e.g., reticulocytes, red blood cells), thrombus cells (e.g., Megakaryoblasts, platelet-producing megakaryocytes, platelets, monocytes (e.g., monocytes, macrophages), dendritic cells Cells, microglia, osteoclasts, and lymphocytes (e.g., NK cells, B cells, and T cells) This refers to immature blood cells that have the ability to differentiate into mature blood cells, including CD34 + May contain cells. CD34 + The cells express the CD34 cell surface marker. These are immature cells. In humans, CD34+ cells possess the stem cell characteristics defined above. It is thought to include subpopulations of cells, and in mice, HSCs are CD34-. Furthermore, HSCs are classified into HSCs with long-term reconstructive ability (LT-HSCs) and HSCs with short-term reconstructive ability. This refers to HSCs (ST-HSCs) that possess functional latent traits. LT-HSCs and ST-HSCs have functional latent traits. Cells differentiate based on their ability and the expression of cell surface markers. For example, human HSCs express CD34 +, CD38-, CD45RA-, CD90+, CD49F+, and lin-(CD2, CD3, CD4, CD7, CD8, CD10, CD11B, CD19, CD20, CD5 6. Negative for mature strain markers including CD235A. In mice, bone marrow LT- HSC includes CD34-, SCA-1+, C-kit+, CD135-, and Slamfl / C. D150+, CD48-, and lin-(Ter119, CD11b, Gr1, CD3, (Negative for mature strain markers including CD4, CD8, B220, and IL7ra), on the other hand, ST-HSC is compatible with CD34+, SCA-1+, C-kit+, CD135-, and Slamf. l / CD150+, and lin-(Ter119, CD11b, Gr1, CD3, CD4 (and is negative for mature strain markers including CD8, B220, and IL7ra). In addition, ST -HSCs have less quiescent state and more proliferative capacity than LT-HSCs under constant conditions. It is expensive. However, LT-HSCs have high self-renewal capacity (i.e., they can survive throughout adulthood). ST-HSC (which can be preserved and continuously implanted in successive recipients) is self They have limited replication ability (i.e., they can only survive for a limited time and do not have the ability to be continuously transplanted). (None). None of these HSCs can be used in the methods described herein. ST-HSCs have high reproductive capacity, and therefore can produce differentiated offspring more quickly. It is particularly useful because it allows for this.

[0063] As used herein, the term “functional potential of hematopoietic stem cells” means 1) multiple Potential (multiple different blood lineages, for example, not limited to, but granulocytes (for example, anterior bone) Myelocytes, neutrophils, eosinophils, basophils), red blood cells (e.g., reticulocytes, red blood cells), thrombus cells (e.g., For example, megakaryoblasts, platelet-producing megakaryocytes, platelets, monocytes (e.g., monocytes, macrophages) , dendritic cells, microglia, osteoclasts, and lymphocytes (e.g., NK cells, T cells and B cells) 1) Cellular differentiation ability (the ability to differentiate into cells), 2) Self-renewal ability (the ability to produce daughter cells that have the same capabilities as the parent cell). Furthermore, the ability of hematopoietic stem cells to be generated repeatedly throughout life without being depleted. 3) refers to the force, and 4) is reintroduced into the transplant recipient and its hematopoietic stem cell niche. This includes the ability of hematopoietic stem cells or their offspring to reconstruct productive and sustainable hematopoiesis. This refers to the functional characteristics of stem cells.

[0064] As used herein, “donor chimerism” or “total donor chimerism” The term refers to the lymphopiopoietic system of the recipient (i.e., host) of allogeneic hematopoietic stem cell transplantation. This refers to the percentage of donor-derived cells in a given cell. For example, 85% donor chimerism is equivalent to allogeneic hematopoiesis. This refers to the lymphopiohematopoietic system containing 85% of the donor cells after stem cell transplantation. In some embodiments... In this specification, the method described herein is for complete or near-complete donor chimerism in vivo. For example, at least 80% donor chimerism in vivo, at least 85% Donor chimerism, at least 90% donor chimerism, at least 95% donor chi Merism, at least 97% donor chimerism, at least 99% donor chimerism It is effective in establishing, or at least 100% donor chimerism. It is also possible to determine the proportion of donor-derived cells present in various hematopoietic subsets or lineages. For example, bone marrow chimerism is the phenomenon where cells from the donor originate from among the bone marrow cells of a transplant recipient. This refers to the percentage of cells. For example, if a transplant recipient has 85% bone marrow chimerisms after HSC transplantation. If the patient has a bone marrow cell, 85% of the target bone marrow cells are derived from the transplant donor, and 15% are derived from the transplant recipe. It originates from the donor. Similarly, B cell chimerism is caused by the donor's B cells being selected from the donor's B cells. This refers to the proportion of cells originating from the transplant recipient. T cell chimerism refers to the proportion of T cells from the transplant recipient. This refers to the proportion of cells derived from the donor. Peripheral donor chimerism refers to the proportion of cells derived from the donor. This refers to the proportion of blood cells. It also refers to the degree of engraftment and chimerism (for example, the degree of engraftment and chimerism of donor stem cells in the host). The proportion can be detected by any number of standard methods. Host sex chromosome specific The presence of donor markers, such as markers, can be used, for example, in standard cytogenetic analysis and appropriate procedures. Polymerase chain reaction (PCR) using lymers, variable-length tandem repeat PCR (V NTR-PCR), microsatellite markers or other fingerprinting The determination is made using techniques such as fluorescence in situ hybridization (FISH). It is possible. Host-donor chimerism is also possible, for example, with standard complement-dependent microcellular activity. The determination is made by using a cytosis test to determine the proportion of donor-type cells in the host blood. It is possible.

[0065] As used herein, "mismatch" (e.g., "MHC mismatch", "HL mismatch") The terms "A mismatch" or "miHA mismatch" are used in the context of hematopoietic stem cell transplantation. In this context, allogeneic cells (and (e.g., donor cells) are tissue or organs on which at least one different (e.g., non This refers to the presence of identical cell surface antigens. Allogeneic transplantation, in some embodiments, This may contain a "minor mismatch" with respect to the transplant recipient. "Nerve mismatch" refers to individual differences in cell surface antigens other than MHC antigens or HLA antigens. Includes. Minor mismatches include differences in minor histocompatibility antigens. Several implementations Morphologically, allografts involve a "major mismatch" with respect to the transplant recipient. It is possible. Such a "major mismatch" can result in MHC levels between the transplanter and the recipient. This refers to differences in protype or HLA haplotype. In exemplary embodiments, allotransfer Plants may share the same MHC haplotype or HLA haplotype as the transplant recipient. However, it may contain one or more minor mismatches (in this specification, "minor mismatches" refers to minor mismatches). (Also known as "smatch allograft") In another exemplary embodiment, allograft is performed by one The above major mismatches are included either alone or in addition to one or more minor mismatches. Obtain. "Complete mismatch" allografts involve one or more major mismatches and one or more minor mismatches. Refers to allografts containing minor mismatches. Major and / or minor mismatches. The presence of [this] is used in the relevant technical fields such as serological analysis, genomic analysis, or molecular analysis. This can be determined by a standard assay. In some embodiments, At least one major histocompatibility complex antigen is expressed by the recipient. This is a mismatch for the organization. Alternatively or additionally, at least one minor organization The compatible antigen is a mismatch with the allele expressed by the recipient.

[0066] As used herein, the terms “subject” and “patient” are defined as follows: This refers to a living organism, such as a human, receiving treatment for a specific disease or condition. For example, a human patient. Patients such as those with conditions requiring treatment before hematopoietic stem cell transplantation are treated to promote the engraftment of exogenous hematopoietic stem cells. They may receive medical treatment.

[0067] As used herein, the term “donor” means a cell from which one or more cells are isolated. Refers to a t or animal, whose cells or offspring are subsequently administered to the recipient. These more than one cell group could, for example, be a population of hematopoietic stem cells.

[0068] As used herein, the term “recipient” refers to a population containing hematopoietic stem cells. This refers to patients who receive transplants, such as those involving cell transplants. The cells administered to the recipient are, For example, these could be autologous cells, syngeneic cells, or allogeneic cells.

[0069] As used herein, the term “endogenous” means that a particular organism, such as a human patient, is inherent to itself. Substances found naturally, such as molecules, cells, tissues, or organs (e.g., hematopoietic stem cells or These are hematopoietic cells, such as megakaryocytes, thrombus cells, platelets, erythrocytes, mast cells, myeloblasts, and tetraphosphorus cells. Basophils, neutrophils, eosinophils, microglia, granulocytes, monocytes, osteoclasts, antigen-presenting cells, Macrophages, dendritic cells, natural killer cells, T lymphocytes, or B lymphocytes, etc. This describes the following:

[0070] As used herein, the term "sample" refers to a specimen taken from an object (e.g., , blood, blood components (e.g., serum or plasma), urine, saliva, amniotic fluid, cerebrospinal fluid, tissue (for example) This refers to the placenta or skin, pancreatic juice, chorionic villi samples, and cells.

[0071] As used herein, the term "scFv" refers to the variability of heavy and light chains derived from an antibody. This refers to a single-stranded Fv antibody in which the mutant domains are linked together to form a single chain. The scFv fragment is , antibody light chain (V) separated by linker L ) variable range (for example, CDR-L1, C DR-L2, and / or CDR-L3) and antibody heavy chain (V H ) variable region (for example, C A single polypeptide chain containing DR-H1, CDR-H2, and / or CDR-H3 Contains. V of scFv fragment L Region and V H Linkers that connect regions are involved in protein synthesis. It can be a peptide linker composed of amino acids. scFv cleavage against protein degradation. To increase the resistance of the fragment (e.g., a linker containing D-amino acids), the scFv fragment is dissolved. To enhance decomposition (for example, polyethylene glycol-containing linker or glycine and Hydrophilic linkers such as polypeptides containing repeating biserine residues, molecular biological products To improve physicosity (for example, by forming intramolecular or intermolecular disulfide bonds) (A linker containing cysteine ​​residues), or to reduce the immunogenicity of scFv fragments ( For example, alternative linkers (such as linkers containing glycosylated sites) may be used. The variable region of the scFv molecule described herein is derived from the antibody molecule and amino You will also understand that the acid sequence can be modified to be different. For example, the conservative arrangement Changes at nucleotide or amino acid substitutions or amino acid residues that result in a change (for example) (to CDR and / or framework residues) and recognized by the corresponding antibody The ability of scFv to bind to the original can be maintained or enhanced.

[0072] As used herein, the phrase "substantially eliminated from the blood" means that the therapeutic agent ( At the time after administration of an anti-CD45 antibody or its antigen-binding portion to a patient, the patient The concentration of the therapeutic drug in the isolated blood sample was determined by conventional means. (e.g., the device or assay used to detect therapeutic drugs) This refers to the point at which the therapeutic agent is no longer detected (beyond the threshold). Antibodies, antibody fragments, and proteins To detect a quality ligand, various techniques known in the art, for example, ELISA-based detection apps known in the art or described herein You can use things like serotonin. You can use it to detect antibodies or antibody fragments. Possible additional assays include those known in the art, such as immunoassays. This includes epidemic precipitation techniques and immunoblotting assays.

[0073] As used herein, the term "transfection" refers to a prokaryote or Of the various techniques commonly used to introduce exogenous DNA into eukaryotic host cells For example, electroporation, lipofection, calcium phosphate precipitation, D This refers to EAE-dextran transfection, etc.

[0074] As used herein, “to treat” or “to treat” means the severity of the disease symptoms and Reducing the frequency of the symptoms and / or eliminating the underlying cause of the symptoms, and / To reduce the frequency or likelihood of its underlying cause, and to reduce the direct or intermittent effects of the disease. Improvement or repair of indirectly caused damage, extension of survival time, reduction of morbidity, and / or Or any improvement of any outcome of the disease, such as the reduction of side effects which are byproducts of alternative treatments. It refers to the complete eradication of a disease, which is desirable as can be easily understood in the art. However, this is not a requirement for a medical treatment. Beneficial or desired clinical results are specified herein. In patients following antibody conditioning therapy and subsequent hematopoietic stem cell transplantation therapy as described above. This includes, but is not limited to, promoting the engraftment of exogenous hematopoietic cells. Beneficial results include conditioning therapy and subsequent treatment of patients with exogenous hematopoietic stem cell grafts. In patients requiring hematopoietic stem cell transplantation after administration, the number or relative concentration of hematopoietic stem cells Increases are included. The beneficial results of the therapies described herein also include conditioning. Megakaryocytes, thrombus cells, platelets, erythrocytes, mastocytes after therapy and subsequent hematopoietic stem cell transplantation therapy Cells, myeloblasts, basophils, neutrophils, eosinophils, microglia, granulocytes, monocytes, osteoclasts Cells, antigen-presenting cells, macrophages, dendritic cells, natural killer cells, T lymphocytes, This includes an increase in the number or relative concentration of one or more hematopoietic cells, such as B lymphocytes. It is rare. Additional beneficial results include the removal of cancer cells (e.g., CD45+ leukemia cells) or autologous cells. Autoimmune cells (for example, CD45+ T cells that express T cell receptors that cross-react with self-antigens) A decrease in the amount of disease-causing cell populations, such as CD45+ autoimmune lymphocytes. This may include. Insofar as the method of disclosure is aimed at preventing harm, “prevent” means It is understood that the term does not require the disease state to be completely prevented. Rather As used herein, the term "prevent" means preventing the compound of the disclosure before the onset of disease. This refers to the ability of a person skilled in the art to identify vulnerable populations that are susceptible to the disorder, so that the drug can be administered. The term does not mean that the disease state will be completely avoided.

[0075] As used herein, a patient who “needs” a hematopoietic stem cell transplant will have one or more blood cells. This applies not only to patients with deficiencies or deletions in fluid cell types, but also to those with stem cell disorders, autoimmune diseases, cancer, etc. or patients with other medical conditions described herein. Hematopoietic stem cells are generally, 1) It exhibits pluripotency, and therefore can be found in multiple different bloodlines, for example, although not limited to them. granulocytes (e.g., promyelocytes, neutrophils, eosinophils, basophils), red blood cells (e.g., reticulocytes) cysts (red blood cells), thrombus cells (e.g., megakaryoblasts, platelet-producing megakaryocytes, platelets), monocytes (e.g., Monocytes, macrophages), dendritic cells, microglia, osteoclasts, and lymphocytes (for example, 1) It can differentiate into NK cells, B cells, and T cells, and 2) it exhibits self-renewal ability, therefore , it is possible to produce daughter cells that have the same capabilities as the mother cells, and 3) re-transplant recipient It is introduced, homes in the hematopoietic stem cell niche, and has the ability to reconstruct productive and sustainable hematopoiesis. It demonstrates strength. Therefore, hematopoietic stem cells are those that have a defect or deletion in one or more cell types of the hematopoietic lineage. It can be administered to a patient to reconstitute a deficient or missing cell population in vivo. For example, a patient may have cancer, and the deletion may be selective or non-selective. This can be caused by the administration of chemotherapy agents or other drugs that deplete cancer cell populations. Additionally or alternatively, patients may have sickle cell anemia, thalassemia, Fanconi anemia, or regeneration. Aplastic anemia and abnormal hemoglobin disorders such as Wiscott-Aldrich syndrome (for example, They may have non-malignant abnormal hemoglobinopathy. The target is adenosine deaminar. Severe combined immunodeficiency (ADASCID), HIV / AIDS, metachromatic leukodystrophy - Suffering from Diamond Blackfan anemia and Schwachmann-Diamond syndrome This could be a person who has hereditary blood disorders (e.g., sickle cell anemia) or autoimmunity. They may have or be affected by the disease. In addition or alternatively, Elephants may have or be affected by malignant tumors such as neuroblastoma or hematological cancers. In some cases, the subject may have leukemia, lymphoma, or myeloma. In one embodiment, the target is acute myeloid leukemia, acute lymphoblastic leukemia, chronic myeloid leukemia Leukemia, chronic lymphocytic leukemia, multiple myeloma, diffuse large B-cell lymphoma, or The patient has non-Hodgkin lymphoma. In some embodiments, the subject is myelodysplastic syndrome. It has. In some embodiments, the subjects are scleroderma, multiple sclerosis, ulcerative colitis Autoimmune conditions such as Crohn's disease, type 1 diabetes, or other autoimmune conditions described herein. The subject has an immune disorder. In some embodiments, the subject is a chimeric antigen receptor T cell (C Requires ART therapy. In some embodiments, the subject has metabolic storage disorders. They either do so or are affected in other ways. The affected conditions are glycogen storage disorders and mucopolysaccharidosis. Gaucher disease, Haller disease, sphingolipidosis, metachromatic leukodystrophy, or this Any other disease or disorder for which one could benefit from the treatments and therapies disclosed in the specification. Harm, for example, but not limited to, severe combined immunodeficiency, Wiscott-Aldrich disease. Syndrome, hyperimmune globulin M (IgM) syndrome, Chediak's disease, hereditary lymphohistiocytosis Osteoporosis, osteogenesis imperfecta, stolic disease, thalassemia macrocephaly, sickle cell disease, systemic scleroderma, all Physical lupus erythematosus, multiple sclerosis, juvenile rheumatoid arthritis and related diseases, or “Bone Marrow Transplantation for Non-Mal ignant disease,”ASH Education Book,1:319 -338(2000)(This disclosure refers to diseases that can be treated by performing hematopoietic stem cell transplantation therapy.) Regarding the condition, the entirety of which is incorporated herein by reference is the defect. They suffer from or are otherwise affected by a metabolic disorder selected from the group. Additionally or alternatively, patients who “need” hematopoietic stem cell transplantation suffer from one of the aforementioned conditions. Whether a person is ill or not, megakaryocytes, thrombi, platelets, Red blood cells, mast cells, myeloblasts, basophils, neutrophils, eosinophils, microglia, granulocytes, monocytocytes Spheres, osteoclasts, antigen-presenting cells, macrophages, dendritic cells, natural killer cells, T cells A decrease in one or more endogenous cell types within the hematopoietic lineage, such as lymphocytes and B lymphocytes. For example, it may be a person who indicates (compared to the level of a healthy subject). For example, among other procedures known in the art, flow cytometry and By fluorescent cell sorting (FACS), one or more of the aforementioned cell types or other It is easy to determine whether the levels of blood cell types are reduced compared to other healthy subjects. It is possible.

[0076] In some embodiments, the method of the present invention is performed in the absence of immunosuppressant therapy. The term "immunosuppressant" or "immunosuppressant" as used herein is derived from the production of the immunosuppressant. This refers to a substance that acts to suppress or block the immune system of a blood transplant recipient. This involves suppression of cytokine production, downregulation or suppression of autoantigen expression, or MHC antigen It contains substances that provide shielding. An example of such an agent is calcineurin / MTOR inhibitor. Agents (e.g., tacrolimus, sirolimus, rapamycin, cyclosporine, everolimus) ), costimulatory blocking molecules (e.g., CTLA4-Ig, anti-CD40L), NK cell depletion agents, antithymus Cell globulin (ATG), alkylating agent (e.g., nitrogen mustard, e.g., , cyclophosphamide; nitrosourea (e.g., carmustine); platinum compounds), methoth Lexart, anti-TCR agents (e.g., muromonab-CD3), anti-CD20 antibodies (e.g., ri Tuximab, ocrelizumab, ofatumumab, and bertuzumab), fludarabine, C ampath (alemtuzumab), 2-amino-6-aryl-5-substituted pyrimidine (rice See National Patent No. 4,665,077 (above), which is incorporated herein by reference. (or if there is an adverse reaction to azathioprine, cyclophosph Famide; Bromocriptine; Glutaraldehyde (U.S. Patent No. 4,120,649) (As described above) it blocks MHC antigens; anti-Idioms against MHC antigens Otype antibody; cyclosporine A; one or more steroids, e.g., corticosteroids For example, prednisone, methylprednisolone, hydrocortisone, and dexamethasone. Glucocorticosteroids such as γ; anti-interferon-γ antibodies; anti-tumor necrosis factor-α Antibodies; anti-tumor necrosis factor-β antibody; anti-interleukin-2 antibody; anti-IL-2 receptor antibody Which anti-cytokine receptor antibodies; heterologous anti-lymphocyte globulins; pan-T antibodies, e.g., OKT -3 monoclonal antibody; antibody against CD4; antibody against CD8, antibody against CD45 Antibodies (e.g., 30-F11, YTH24.5, and / or YTH54.12 (e.g.) (combination of YTH24.5 and YTH54.12); streptokinase; streptokinase Examples include putodorases; or host-derived RNA or DNA. Additional immunosuppression. The agents include whole-body irradiation (TBI), low-dose TBI, and / or cytoxanes, but These are not the only options.

[0077] In some embodiments, the method of the present invention involves simultaneous or nearly simultaneous use of an immunosuppressant. It is carried out in the absence of the treatment. For example, in some embodiments provided herein The subjects who receive the CD45-targeted portion bound to the toxin are those who are being treated with immunosuppressants. They do not receive both at the same time. In some embodiments, the subject receives the CD45-targeted portion at the time of administration. Furthermore, the effectiveness of immunosuppressant therapy has not been observed. In some embodiments, the target Prior to administration of the CD45-targeted portion, at least 3 days, at least 7 days, 14 days, at least 21 days, at least 28 days, at least 1 month, less For at least two months, at least three months, at least four months, at least five months, and less Both for 6 months, at least 7 months, at least 8 months, at least 9 months, less In both cases, immunosuppressant administration for 10 months, at least 11 months, or at least 12 months. Not administered. In some embodiments, the subject is at the time of administration of the CD45-targeting portion. After that, at least 3 days, at least 7 days, at least 14 days, at least 21 days Between, at least 28 days, at least 1 month, at least 2 months, at least 3 months Between, at least 4 months, at least 5 months, at least 6 months, at least 7 months Between, for at least 8 months, for at least 9 months, for at least 10 months, for at least 11 Not having received immunosuppressants for months, or at least 12 months. In the administration method, the subjects were immunocompromised from one day before to one day after administration of the CD45-targeted portion. The subjects have not received any disease control agents. In some embodiments, the subjects are CD45 targeted. No immunosuppressants were administered between 3 days before and 3 days after the partial administration. In this embodiment, the subjects are those who have been taking the CD45-targeted portion for 7 days prior to 7 days prior to administration of the CD45-targeted portion. The subject has not received immunosuppressants. In some embodiments, the subject is CD45 standard The patient did not receive immunosuppressants between 14 days before and 14 days after the administration of the targeted portion of the drug. In some embodiments, the subjects are those who have been taking the CD45-targeted portion for 21 days prior to administration of the CD45-targeted portion. No immunosuppressants were administered during the following days. In some embodiments, the subjects are In patients who receive immunosuppressants between 28 days before and 28 days after the administration of the CD45-targeted portion of the drug, Not included. In some embodiments, the subject is one month after administration of the CD45-targeting portion. No immunosuppressant drugs were administered between one month prior to one month after. The subjects were immunosuppressed from two months before to two months after administration of the CD45-targeted portion. No drug has been administered. In some embodiments, the subject is the CD45-targeting portion. No immunosuppressants were administered between 6 months before and 6 months after the administration of the drug. In this embodiment, the subjects are those from 8 months before to 8 months after administration of the CD45-targeting portion. No immunosuppressants were administered in between. In some embodiments, the subject is CD45 If you have received immunosuppressants between 10 months before and 10 months after the administration of the targeted portion of the drug, No. In some embodiments, the subjects have been taking the CD45-targeted portion for one year prior to administration. No immunosuppressants were administered during the past year.

[0078] As used herein, the terms “variant” and “derivative” are used interchangeably. Used, the natural compounds, peptides, proteins, or other substances described herein This refers to synthetic and semi-synthetic analogues. The compounds, peptides, and proteins described herein are included. or variants or derivatives of other substances retain or improve the biological activity of the original substance. obtain.

[0079] As used herein, the term "stem cell damage" refers to the condition of the target tissue in question. Shocking and / or removal of endogenous stem cell populations within target tissue (e.g., target bone marrow population) Removal of endogenous hematopoietic stem cells or progenitor cell populations from tissue, and / or to target tissue. Any disease, disorder, or condition that can be treated or cured by the engraftment or transplantation of stem cells, The term broadly refers to a condition. For example, type 1 diabetes has been shown to be curable by hematopoietic stem cell transplantation. The benefits of conditioning according to the compositions and methods described herein are obtained. It is possible to obtain this. Treatment can be performed using the compositions and methods described herein. Additional complications that may occur include, but are not limited to, sickle cell anemia, thalassemia, and fungovirus. - Anemia, Aplastic Anemia, Wiscott-Aldrich Syndrome, ADA SCID, HI V / AIDS, metachromatic leukodystrophy, diamond black fan anemia, and sch This includes Wachman-Diamond syndrome. Patient conditions described herein Additional diseases that can be treated using hematopoietic stem cell transplantation include hereditary blood Disorders (e.g., sickle cell anemia) as well as scleroderma, multiple sclerosis, ulcerative colitis, and This includes autoimmune disorders such as Rohn's disease. Conditioning and / or additional diseases that can be treated using transplantation methods include malignant tumors such as neuroblastoma. Or it includes blood cancers such as leukemia, lymphoma, and myeloma. For example, cancer is acute Myeloid leukemia, acute lymphoblastic leukemia, chronic myeloid leukemia, chronic lymphocytic leukemia, multiple myeloid leukemia It could be myeloma, diffuse large B-cell lymphoma, or non-Hodgkin lymphoma. Treatment may be performed using the conditioning and / or transplantation methods described in the manual. Possible additional diseases include myelodysplastic syndromes. In some embodiments, the target They have metabolic storage disorders or are otherwise affected by them. For example, the subjects are: Glycogen storage disease, mucopolysaccharidosis, Gaucher disease, Haller disease, sphingolipidosis, metachromosis Sexual leukodystrophy, or benefiting from the treatments and therapies disclosed herein Any other disease or disorder that may occur, for example, severe combined immunodeficiency, etc. All, Wiscott-Aldrich syndrome, hyperimmune globulin M (IgM) syndrome, Cheddy Ack Dong disease, hereditary lymphohistiocytosis, osteoporosis, osteogenesis imperfecta, stolic disease, major thalassemia, Sickle cell disease, systemic sclerosis, systemic lupus erythematosus, multiple sclerosis, juvenile arthritis Marrow and related diseases, or “Bone Marrow Transplantat” ion for Non-Malignant Disease,”ASH Educa tion Book, 1:319-338 (2000) (This disclosure pertains to hematopoietic stem cell transplantation therapy) With regard to the medical conditions that can be treated by the implementation of the law, the entirety thereof is incorporated herein by reference. Whether or not you suffer from a metabolic disorder selected from the group of disorders listed in (ru), They may be affected in different ways.

[0080] As used herein, the term "vector" refers to plasmids, DNA vectors, etc. nucleic acid vectors such as plasmids, RNA vectors, viruses, or other suitable replicons The expression vectors described herein include not only polynucleotide sequences, but also For example, protein expression and / or the generative processes of these polynucleotide sequences in mammalian cells. The antibody and antibody fragment of the present invention may contain additional sequence elements used for incorporation into the NM. Certain vectors that can be used for gene expression include promo that directs gene transcription. The plasmid contains regulatory sequences such as the inhibitor region and enhancer region. Other vectors useful for the expression of antibody fragments may improve the translation rate of these genes. or polynuclear nucleotides that improve the stability or nuclear export of mRNA resulting from gene transcription. It contains rheotide sequences. These sequence elements are the genes that are mounted on the expression vector. To direct efficient transcription, for example, the 5' and 3' untranslated regions, as well as polyadeni The expression vectors described herein may also include such Contains polynucleotides encoding markers for selecting cells containing the vector. However, this is also acceptable. Examples of suitable markers include ampicillin, chloramphenicol, and kanama. Examples include genes that encode resistance to antibiotics such as isin and nurceotricin. It can be done.

[0081] As used herein, the terms "conjugate" or "antibody drug conjugate" are used. The term "ADC" refers to an antibody linked to a cytotoxic substance. ADC is an antibody or a cytotoxic substance. A reactive functional group of one molecule, such as an antigen-binding fragment, and a cytotoxin as described herein. Conjugate This may include a linker between two molecules that are bound together, for example, between an antibody and a cytotoxic substance. Examples of linkers that can be used to form conjugates include peptide-containing linkers. amino acids, for example, naturally occurring amino acids or amino acids that do not exist naturally, for example, D -Examples include linkers containing amino acids. Linkers are described herein and are technically applicable. It can be prepared using various strategies known in the field of technology. The linker is Depending on the reactive component, for example, enzymatic hydrolysis, photolysis, hydrolysis under acidic conditions, Hydrolysis, oxidation, disulfide reduction, nucleophilic cleavage, or organometallic cleavage under basic conditions It can be cut by tearing (e.g., Leriche et al., Bioorg) (See Med. Chem., 20:571-582, 2012).

[0082] As used herein, the term "microtubule binder" refers to the mitotic and interphase of cells. This refers to compounds that act by disrupting the microtubule network, which is essential for cellular function. Examples of microtubule binding agents include meitacin, meitacinoids and their derivatives, for example, Vincaa Lukaloids, for example, vinblastine, vinblastine sulfate, vincristine, vink Listin sulfate, vindesine, and vinorelbine, taxanes, such as docetaxel and Paclitaxel, macrolides, such as discodermolide, cochite, and others. Potilon and its derivatives, for example, epotilon B or its derivatives, are mentioned. It is not limited to them.

[0083] As used herein, the term "amatoxin" refers to Amanita pha Members of the amatoxin family of peptides produced by the mushroom Lloydes or its variants or derivatives, for example, inhibiting RNA polymerase II activity. This refers to the variant or derivative thereof that can be formed. The compositions and Amatoxins useful in relation to the method include, but are not limited to, formulas (III) and (I This includes compounds such as compounds of type IIA), (IIIB), and (IIIC), and each of them Regarding this, see below the information provided in this specification (for example, α-amanitin, β-amanitin, γ-amanitin, ε-amanitin, amanin, amaninamide, amanulin, amanulin Acids, or proamanulins). As described herein, amatoxins are, for example, It can be conjugated via the linker portion (L) to the antibody or its antigen-binding portion. This leads to the formation of ADC). Exemplary methods of amatoxin conjugation and Linkers useful for such processes are described below: Compositions and Methods According to this, exemplary linkers useful for conjugating antibody or antigen-binding sites. —The amatoxins contained are also described herein.

[0084] As used herein, the term "acyl" refers to -C(=O)R, where R is, Hydrogen ("aldehyde"), alkyl, alkenyl, alkynyl, and carboxyl compounds as defined herein. Rubocyclyl, aryl, heteroaryl, or heterocyclyl. (Non-restrictive example) Examples include formyl, acetyl, propanoyl, benzoyl, and acryloyl. It can be done.

[0085] As used herein, the term "alkyl" means, for example, 1 to 20 in a chain This refers to a linear or branched alkyl group containing carbon atoms. An example of an alkyl group is methyl alkyl. ethyl, n-propyl, isopropyl, butyl, isobutyl, sec-butyl, te rt-butyl, pentyl, isopentyl, tert-pentyl, hexyl, isohexyl These are some examples.

[0086] As used herein, the term "alkylene" refers to a divalent alkylene that is either linear or branched. This refers to the luricyl group. The divalent position can be on the same atom within the alkyl chain, or on different atoms. It may be present. Examples of alkylenes include methylene, ethylene, propylene, and isopropyl Len is one example.

[0087] As used herein, the term "heteroalkyl" means, for example, 1-2 in a chain. It has 0 carbon atoms and one or more heteroatoms in the chain (for example, oxygen, nitrogen, This refers to a linear or branched alkyl group that further contains (or sulfur).

[0088] As used herein, the term "heteroalkylene" refers to a linear or branched heteroalkylene. This refers to a divalent heteroalkyl group. The divalent position is on the same atom within the heteroalkyl chain. The divalent position may be on a different atom. The divalent position may be on one or more heteroatoms. .

[0089] As used herein, the term "alkenyl" means, for example, 2 to 20 in a chain. This refers to a linear or branched alkenyl group having carbon atoms. Examples of alkenyl groups include Vinyl, propenyl, isopropenyl, butenyl, tert-butyrenyl, hexenyl These are some examples.

[0090] As used herein, the term "alkenylene" refers to a divalent form of a linear or branched chain. This refers to the alkenyl group. The divalent position, even if on the same atom within the alkenyl chain, can have different valent properties. It may also be used as a sub-unit. Examples of alkenylenes include etenylene, propenylene, and isopropyl alcohol. Examples include penirene and butenylene.

[0091] As used herein, the term "heteroalkenyl" means, for example, a chain containing 2- It has 20 carbon atoms and one or more heteroatoms in the chain (for example, oxygen, nitrogen, etc.). This refers to a linear or branched alkenyl group further containing (or sulfur).

[0092] As used herein, the term "heteroalkenylene" refers to a linear or branched chain. This refers to the divalent heteroalkenyl group. The divalent position is on the same atom within the heteroalkenyl chain. They may be present, or they may be on different atoms. The divalent position is one or more heteroatoms. That's good too.

[0093] As used herein, the term "alkynyl" means, for example, 2 to 20 in a chain. This refers to a linear or branched alkynyl group having carbon atoms. Examples of alkynyl groups include Examples include propargyl, butynyl, pentynyl, and hexynyl.

[0094] As used herein, the term "alkynylene" refers to a divalent form of a linear or branched chain. This refers to the alkynyl group. The divalent position can be different even if it is on the same atom within the alkynyl chain. It's fine if the child is born first.

[0095] As used herein, the term "heteroalkynyl" means, for example, a chain containing 2- It has 20 carbon atoms and one or more heteroatoms in the chain (for example, oxygen, nitrogen, etc.). This refers to a linear or branched alkynyl group further containing (or sulfur).

[0096] As used herein, the term "heteroalkylene" refers to a linear or branched chain. This refers to the divalent heteroalkynyl group. The divalent position is on the same atom within the heteroalkynyl chain. They may be present, or they may be on different atoms. The divalent position is one or more heteroatoms. That's good too.

[0097] As used herein, the term "cycloalkyl" means, for example, 3 to 12 A monocyclic ring structure, or a condensed, bridged, or spiropolycyclic ring structure, having a carbon ring atom and saturated. This refers to a ring structure. Examples of cycloalkyl groups include cyclopropyl, cyclobutyl, and cyclo Pentyl, cyclohexyl, cycloheptyl, cyclooctyl, bicyclo[3.1.0] Examples include hexane.

[0098] As used herein, the term "cycloalkylene" means divalent cycloalkyl It refers to the group. The position of the divalent can be on the same atom or a different atom within the ring structure. Examples of cycloalkylenes include cyclopropylene, cyclobutylene, and cyclobenzoyl acetate. Examples include lenucleans and cyclohexylene.

[0099] As used herein, the term "heterosiloalkyl" means, for example, a carbon atom. and selected from heteroatoms (for example, selected from nitrogen, oxygen, and sulfur) Each ring structure has 3 to 12 ring atoms and is saturated, a monocyclic ring structure, or a condensed, bridged ring structure. This refers to a bridge or spiropolycyclic ring structure. The ring structure consists of carbon, nitrogen, or sulfur ring members. For example, it may contain one or more oxo groups. Examples of heterocycloalkyls include: This is an indication and not an limitation, but dihydropyridyl, tetrahydropyridyl (piper Zyl, tetrahydrothiophenyl, piperidinyl, 4-piperidonyl, pyrrolidinyl, 2-pyrrolidonyl, tetrahydrofuranyl, tetrahydropyranyl, bis-tetrahydro Pyranyl, tetrahydroquinolinyl, tetrahydroisoquinolinyl, decahydroquinolinyl Lu, octahydroisoquinolinyl, piperazinyl, quinuclidinyl, and morpholinyl It can be listed.

[0100] As used herein, the term "heterocycloalkylene" means divalent heterocycloalkylene This refers to a chloroalkyl group. The divalent position can be on the same atom within the ring structure, or on a different atom. That's fine.

[0101] As used herein, the term "aryl" refers to, for example, 6 to 19 carbon atoms. This refers to monocyclic or polycyclic aromatic ring systems containing aryl groups. Examples of aryl groups include phenyl and flu. This includes, but is not limited to, orenyl and naphthyl. There is one or more divalent positions. It may also be a heteroatom.

[0102] As used herein, the term "arylene" refers to a divalent aryl group. The divalent nucleotide can be located on the same atom or on different atoms.

[0103] As used herein, "heteroaralkyl" refers to carbon atoms, typically terminal or sp. One of the hydrogen atoms bonded to the three carbon atoms is replaced by a heteroaryl radical. This refers to acyclic alkyl radicals. Typical heteroarylalkyl groups include 2-ben This includes, but is not limited to, zoimidazolylmethyl and 2-furylethyl. Telolarylalkyl groups contain 6 to 20 carbon atoms, for example, an alkanyl group, an aryl alkyl group. The alkyl portion of a heteroarylalkyl group containing a kenyl group or an alkynyl group is 1 to 6 The heteroaryl moiety consists of 5 to 14 carbon atoms, as well as N, O, and P. , and 1 to 3 heteroatoms selected from S. Heteroarylalkyl group The loaryl portion is a monocyclic ring with 3 to 7 ring members (2 to 6 carbon atoms or 7 to 10 rings). A biring (containing 4 to 9 carbon atoms and 1 to 3 atoms selected from N, O, P, and S) (a number of heteroatoms), for example, bicyclo[4,5], [5,5], [5,6], or [6 It could be a 6] system.

[0104] As used herein, the term "heterocycloalkyl" means, for example, a carbon source. Selected from nitrogen and heteroatoms (for example, selected from nitrogen, oxygen, and sulfur). Each ring structure has 3 to 12 ring atoms and is saturated, a monocyclic ring structure, or a condensed ring structure. This refers to a bridged or spiropolycyclic ring structure. The ring structure consists of carbon, nitrogen, or sulfur ring members. For example, it may contain one or more oxo groups. Examples of heterocycloalkyls include: These are examples and not limiting, but dihydropyridyl, tetrahydropyridyl (pipette Lysyl, tetrahydrothiophenyl, piperidinyl, 4-piperidonyl, pyrrolidinyl , 2-pyrrolidonyl, tetrahydrofuranil, tetrahydropyranil, bis-tetrahydr Ropyranil, Tetrahydroquinolinil, Tetrahydroisoquinolinil, Decahydroquinolinil Nyl, octahydroisoquinolinyl, piperazinyl, quinuclidinyl, and morpholinyl These are some examples.

[0105] As used herein, the term "heterocycloalkylene" means divalent heterocycloalkylene This refers to a chloroalkyl group. The divalent position can be on the same atom within the ring structure, or on a different atom. That's fine.

[0106] As used herein, the term "aryl" refers to, for example, 6 to 19 carbon atoms. This refers to monocyclic or polycyclic aromatic ring systems containing aryl groups. Examples of aryl groups include phenyl and flu. This includes, but is not limited to, orenyl and naphthyl. There is one or more divalent positions. It may also be a heteroatom.

[0107] As used herein, the term "arylene" refers to a divalent aryl group. The divalent nucleotide can be located on the same atom or on different atoms.

[0108] As used herein, the term "heteroaryl" means that one or more ring atoms are heteroaryl. Monocyclic heteroaromatic groups, or bicyclic groups, where the teloatom is, for example, nitrogen, oxygen, or sulfur. Alternatively, it refers to a tricyclic fused heteroaromatic group. Heteroaryl groups include pyridyl and pyrrolyl. , furyl, thienyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, i Sothiazolyl, pyrazolyl, 1,2,3-triazolyl, 1,2,4-triazolyl, 1 ,2,3-Oxadiazolyl, 1,2,4-Oxadiazolyl, 1,2,5-Oxadia Zolyl, 1,3,4-Oxadiazolyl, 1,3,4-Triadinyl, 1,2,3-Triadinyl Azinyl, benzofuryl, [2,3-dihydro]benzofuryl, isobenzofuryl, ben Zothienyl, benzotriazolyl, isobenzothienyl, indolyl, isoindolyl 3H-indolyl, benzimidazolyl, imidazo[1,2-a]pyridyl, benzothia Zolyl, benzoxazolyl, quinolidinil, quinazolinil, phthalazinil, quinoxal Nyl, cinnorinyl, naphthilidinyl, pyrido[3,4-b]pyridyl, pyrido[3,2 -b]pyridyl, pyrido[4,3-b]pyridyl, quinolyl, isoquinolyl, tetrazoly Lu, 5,6,7,8-tetrahydroquinolyl, 5,6,7,8-tetrahydroisoquinolyl It contains prinyl, pteridinyl, carbazolyl, xanthenyl, benzoquinolil, etc. It can be done.

[0109] As used herein, the term "heteroarylene" means a divalent heteroarylene This refers to the divalent group. The position of the divalent group can be on the same atom or on different atoms. The position may be occupied by one or more heteroatoms.

[0110] Heteroaryl groups and heterocycloalkyl groups are defined by Paquette, Leo A. “Principles of Modern Heterocyclic Chemistry "Stry" (WABenjamin, New York, 1968), especially chapters 1 and 3. Chapters 4, 6, 7, and 9; “The Chemistry of Hetero cyclic compounds,A series of Monographs” (John Wiley & Sons, New York, 1950–present), especially 1 Volumes 3, 14, 16, 19, and 28; and J.Am.Chem.Soc.( It is described in 1960)82:5566.

[0111] Examples are provided and not limited to, but include carbon-bonded heteroaryl and heterocycloal Kill is at the 2nd, 3rd, 4th, 5th, or 6th position of pyridine, or the 3rd, 4th, 5th, or 6th place, pyrimidines 2, 4, 5 or 6th place, pyrazines 2, 3, 5 or 6th place, Furan, tetrahydrofuran, thiofuran, thiophene, pyrrole or tetrahydro Pyrrole at positions 2, 3, 4, or 5, oxazole, imidazole, or thiazole The 2nd, 4th, or 5th positions are isoxazole, pyrazole, or isothiazole (3). , 4th or 5th place, aziridine 2nd or 3rd place, azetidine 2nd, 3rd or 4th place, Quinoline ranked 2nd, 3rd, 4th, 5th, 6th, 7th, or 8th, or isoquinoline ranked 1st, 3rd, or 4th. It is bonded at positions 5, 6, 7, or 8. More typically, carbon-bonded heterocycles are 2-pyridyl, 3-pyridyl, 4-pyridyl, 5-pyridyl, 6-pyridyl, 3-pyridyl Dinyl, 4-pyridazinyl, 5-pyridazinyl, 6-pyridazinyl, 2-pyrimidinyl, 4-pyrimidinyl, 5-pyrimidinyl, 6-pyrimidinyl, 2-pyrazinyl, 3-pyrazi Nyl, 5-pyrazinyl, 6-pyrazinyl, 2-thiazolyl, 4-thiazolyl, or 5- It contains thiazolyl.

[0112] Examples are provided and are not limited to, but include nitrogen-bonded heteroaryl and heterocycloal Kill contains aziridine, azetidine, pyrrole, pyrrolidine, 2-pyrroline, and 3-pyrroline. Imidazole, imidazolidine, 2-imidazoline, 3-imidazoline, pyrazole, Pyrazoline, 2-pyrazoline, 3-pyrazoline, piperidine, piperazine, indole, Indoline, 1H-indazole at position 1, isoindole, or isoindoline at position 2 It binds to the 4th position of morpholine, and the 9th position of carbazole or beta-carbolin. More typically, nitrogen-bonded heterocycles include 1-aziridyl, 1-azetezyl, It contains 1-pyrrolyl, 1-imidazolyl, 1-pyrazolyl, and 1-piperidinil.

[0113] Unless otherwise restricted by the definition of individual substituents, "alkyl" and "alkylene" are used. "heteroalkyl", "heteroalkylene", "alkenyl", "alkenylene", "he "Teloalkenyl", "heteralkenylene", "alkynyl", "alkynylene", "he "Teloalkynyl", "heteroalkynylene", "cycloalkyl", "cycloalkylene "heterocycloalkyl", "heterocycloalkylene", "aryl", "alile The aforementioned chemical parts such as the "n", "heteroaryl", and "heteroarylene" groups are optional. Depending on the selection, for example, alkyl, alkenyl, alkynyl, cycloalkyl, heteroalkyl Alkyl, alkylaryl, alkylheteroaryl, alkylcycloalkyl, Lukyl heterocycloalkyl, amino, ammonium, acyl, acyloxy, acyl a Mino, aminocarbonyl, alkoxycarbonyl, ureido, carbamate, aryl, Heteroaryl, sulfinyl, sulfonyl, alkoxy, sulfanyl, halogen, A group consisting of ruboxy, trihalomethyl, cyano, hydroxy, mercapto, nitro, etc. It can be substituted with 1 to 5 substituents selected from the above. Typical substituents include -X, -R, and -O. H, -OR, -SH, -SR, NH2, -NHR, -N(R)2, -N + (R)3, -C X3, -CN, -OCN, -SCN, -NCO, -NCS, -NO, -NO2, -N3, -NC(=O)H, -NC(=O)R, -C(=O)H, -C(=O)R, -C(=O) NH2, -C(=O)N(R)2, -SO3-, -SO3H, -S(=O)2R, -OS (=O)2OR, -S(=O)2NH 2、 -S(=O)2N(R)2, -S(=O)R, -OP(=O)(OH) 2、 -OP(=O)(OR)2, -P(=O)(OR)2, -P O3, -PO3H2, -C(=O)X, -C(=S)R, -CO2H, -CO2R, -C O2-, -C(=S)OR, -C(=O)SR, -C(=S)SR, -C(=O)NH2 、 -C(=O)N(R)2, -C(=S)NH2, -C(=S)N(R)2, -C(=N This includes, but is not limited to, H)NH2 and -C(=NR)N(R)2, and in the formula, Each X is independently selected from F, Cl, Br, and I for each case, and each R In each case, this refers to alkyl, aryl, heterocycloalkyl, or hetero The aryl group, protecting group, and prodrug moiety are selected independently. Where it is stated that it is replaced by, the base is independently for each case. It may be substituted with one or more of the above substituents. Substitutions include closing vicinal functional substituents. Formed by ring closure, for example, to provide a protecting group, through the closure of adjacent substituents such as rings. For example, lactams, lactones, cyclic anhydrides, acetals, hemiacetals, thio This may include situations in which cetals, aminals, and hemiaminals are formed.

[0114] Certain radical naming conventions, depending on the context, can refer to monoradicals or diradicals. It should be understood that this may include any of the following: for example, a substituent having two bonding points with the rest of the molecule. When required, the substituent is understood to be a diradical. For example, two bonds substituents that are identified as alkyl groups requiring a dot include -CH2-, -CH2CH2-, This includes diradicals such as -CH2CH(CH3)CH2-. Other radical naming conventions are , radicals are "alkylene", "alkenylene", "arylene", "heterocycloal". It clearly indicates that it is a diradical such as "chylene".

[0115] As used herein, the term "coupling reaction" refers to a reaction between two substances that are compatible with each other. Two or more substituents connect to the molecular fragment to which each substituent is attached (for example, by covalent bonding). ) This refers to a chemical reaction that reacts to form a chemical moiety. Coupling reactions include the technology described above. A fragment that is a cytotoxic, such as a cytotoxin known in the field or described herein. The bonded reactive substituents are known in the art or described herein. Antibodies or antigen-binding portions of antibodies or their antigen-binding portions that are specific to CD45 This includes substances that react with suitable reactive substituents bonded to a fragment. Suitable reactions Examples of sexual substituents include nucleophile / electrophile pairs (for example, thiol / haloa Lukyl pair, amine / carbonyl pair, or thiol / α,β-unsaturated carbonyl (Pairs of), diene / dienate pairs (for example, azid / alkyne pairs) Examples include coupling reactions, although not limited to them, such as thiol alkylation. Hydroxylalkylation, amine alkylation, amine condensation, amidation, esterification, di Sulfide formation, cycloaddition (for example, among them, [4+2] Diels-Alder cycloaddition) [3+2]Hüsgen cycloaddition), aromatic nucleophilic substitution, aromatic electrophilic substitution, and the art This includes other reactive modalities known in the field or described herein. .

[0116] As used herein, "CRU (Competitive Reconstruction Unit)" refers to a post-in vivo transplanted unit. This refers to an indicator unit of long-term engrafted stem cells that can be detected.

[0117] As used herein, “drug-antibody ratio” or “DAR” refers to the antibody of the ADC. This refers to the number of cytotoxic substances, such as amatoxins, that are produced. The DAR of ADC is in the range of 1 to 8. It is possible, but depending on the number of binding sites on the antibody, higher loading is also possible. In a particular embodiment, the ADC described herein is 1, 2, 3, 4, 5, 6, It has 7 or 8 DARs.

[0118] The substituent is described as a diradical (i.e., it has two bonding points relative to the rest of the molecule) If (there is), it is understood that substituents can be bonded in any orientation unless otherwise specified. It will be done.

[0119] Treatment method Disclosed herein are allogeneic transplants, such as allogeneic hematopoietic stem cell (HSC) transplants. In patients, the population of CD45+ cells is determined by the CD45 targeting region that can bind to the toxin. This method involves depleting the CD45 through administration. This refers to an anti-CD45 antibody, or its antigen-binding fragment or portion, or a CD45-targeted antibody drug. It can be an object conjugate (ADC).

[0120] In some embodiments, what is provided herein includes a complete mismatch HSC transplant. It is possible to achieve substantial donor chimerism after allogeneic hematopoietic stem cell (HSC) transplantation. This is a monotherapy conditioning regimen. In an exemplary embodiment, HSC transplantation is not required. The target group is CD45 in the absence of additional conditioning agents such as immunosuppressants. The patient receives an administration of a targeted portion (e.g., an anti-CD45 antibody-drug conjugate (ADC)). The CD45-targeting portion (e.g., anti-CD45 ADC) is immune to low-dose whole-body irradiation. Simultaneous or nearly simultaneous administration by disease suppressants or myeloablative agents such as anti-CD4 or anti-CD8 antibodies. To enable complete or near-complete donor chimerism without requiring simultaneous treatment. It can be administered to the target in a sufficient amount.

[0121] Therefore, what is provided herein is a C in patients requiring hematopoietic stem cell transplantation. A method to deplete a population of D45+ cells, and before transplantation, the CD45-targeted portion The method involves administering an effective dose of (for example, an anti-CD45 ADC) to the patient. In some embodiments, the CD45-targeting portion (e.g., anti-CD45 ADC) is other It is administered as a monotherapy in the absence of any conditioning agent. In some embodiments... The CD45-targeting portion (e.g., anti-CD45 ADC) is administered as monotherapy. In some embodiments, a CD45-targeting portion (e.g., anti-CD45 ADC) It is administered in the absence of immunosuppressants. In some embodiments, CD45 targeting Part (e.g., anti-CD45 ADC) is used in prior or concurrent treatment of patients with immunosuppressants. It is administered without any intervention. In some embodiments, the CD45-targeting moiety (e.g., anti-CD) is used. 45 ADC) is used without prior or concurrent treatment of patients with whole-body irradiation, including low-dose TBI. , administered. Low-dose TBI includes TBI with non-myeloablative doses. Several embodiments In this case, the CD45-targeting portion (e.g., anti-CD45 ADC) is affected by an anti-CD4 antibody. It is administered without prior or concurrent treatment to the patient. In some embodiments, CD45 The targeted portion (e.g., anti-CD45 ADC) is used in patients prior to treatment with an anti-CD8 antibody or It is administered without concurrent treatment.

[0122] In some embodiments, the method involves simultaneous or near-simultaneous treatment with immunosuppressants. It is carried out in the absence of the substance. For example, in some embodiments, the poison provided herein Patients who receive the CD45-targeted portion bound to the base drug are simultaneously treated with immunosuppressants. No. In some embodiments, the subject is immune upon administration of the CD45-targeted portion. We have not experienced the effects of treatment with inhibitors. In some embodiments, the subject is CD 45 Prior to administration of the targeted portion, at least 3 days, at least 7 days, and at least 14 days, at least 21 days, at least 28 days, at least 1 month, at least 2 months Monthly, at least 3 months, at least 4 months, at least 5 months, at least 6 months Monthly, at least 7 months, at least 8 months, at least 9 months, at least 10 For months, at least 11 months, or at least 12 months, you will receive immunosuppressant medication. Not applicable. In addition or alternatively, in some embodiments, the target is the CD45 target. After administration of the chemical portion, for at least 3 days, at least 7 days, at least 14 days, At least 21 days, at least 28 days, at least 1 month, at least 2 months, At least 3 months, at least 4 months, at least 5 months, at least 6 months, At least 7 months, at least 8 months, at least 9 months, at least 10 months, Not having received immunosuppressants for at least 11 months, or at least 12 months. In some embodiments, the subjects were selected from one day before to one day after administration of the CD45-targeted portion. No immunosuppressants were administered during the period leading up to this point. In some embodiments, the subjects are If you have received immunosuppressants between 3 days before and 3 days after administration of the CD45-targeted portion of the drug, No. In some embodiments, the subject is from 7 days prior to the administration of the CD45-targeted portion. No immunosuppressants were administered within the following 7 days. In some embodiments, the target This refers to the administration of immunosuppressants between 14 days before and 14 days after the administration of the CD45-targeted portion. Not received. In some embodiments, the subject is 2 at the time of administration of the CD45-targeting portion. No immunosuppressants were administered between 1 day prior to 21 days later. In some embodiments... In this context, the subjects were immunosuppressed from 28 days before to 28 days after administration of the CD45-targeted portion. No inhibitors have been administered. In some embodiments, the target is the CD45-targeted portion. No immunosuppressants were administered between one month before and one month after the administration of [the drug]. In this embodiment, the target period is from two months before to two months after administration of the CD45-targeting portion. No immunosuppressants were administered during this period. In some embodiments, the subjects are CD4 5. The patient has not received immunosuppressants between 6 months before and 6 months after administration of the targeted portion. i. In some embodiments, the subjects were from 8 months prior to the administration of the CD45-targeted portion. No immunosuppressants were administered during the following 8 months. In some embodiments, Elephants were given immunosuppressants between 10 months before and 10 months after administration of the CD45-targeted portion. No administration has been received. In some embodiments, the subject is administered the CD45-targeted portion. No immunosuppressants were administered between one year prior to and one year after the event.

[0123] In some embodiments, the transplant is a minor mismatch allotransplant. In some embodiments, the transplant is a major mismatch allograft. In this case, the transplant is a complete mismatch allograft.

[0124] Furthermore, this specification provides information on the level of engraftment of allogeneic cells in the recipient subject. This is a method to increase. The method provided herein addresses various obstacles associated with allografting, for example. For example, hematopoietic cell-related diseases, cancer, autoimmune diseases, metabolic disorders, and graft-versus-host diseases. It can be used to treat diseases, host-versus-graft rejection, and stem cell disorders. The compositions and methods described in this book are for (i) cancer cells (e.g., leukemia cells) and autoimmune Directly depleting a population of cells that cause a pathological condition, such as a population of autoreactive T cells. (ii) to allow and / or to provide a niche in which transplanted cells can home Therefore, in order to promote the engraftment of transplanted hematopoietic stem cells, the population of endogenous hematopoietic stem cells is depleted. It can be done. Transplantation, for example, endogenous hematopoietic cells in subjects requiring HSC transplantation. The depletion of antigens can target antigens that can bind to antigens expressed by endogenous hematopoietic stem cells. This can be achieved by administering a portion, ADC, antibody, or its antigen-binding portion. In the preparation for transplant therapy, this administration selectively depletes the population of endogenous hematopoietic stem cells. As a result, space is created in hematopoietic tissues such as bone marrow, and this space is then used for exogenous transplantation. It can be filled by hematopoietic stem cells. It is expressed by hematopoietic stem cells (e.g., CD45+ cells). Antigens or antibodies expressed by immune cells such as T cells (e.g., mature immune cells) An antigen-targeting moiety, ADC, antibody, or capable of binding to the progenitor (e.g., CD45). The antigen-binding portion can be administered to patients because it causes cell depletion. However, This refers to antigens expressed by hematopoietic stem cells (e.g., CD45) or immune cells such as T cells. It binds to antigens (e.g., CD45) expressed by cells (e.g., mature immune cells). The antigen-targeting portion, ADC, antibody, or its antigen-binding portion is used to target cancer cells or autoimmune cells. To directly deplete a population of cells, it is administered to patients with cancer or autoimmune diseases. It can also promote the survival and engraftment ability of transplanted cells, such as hematopoietic stem cells. Therefore, it can be administered to patients who require hematopoietic stem cell transplantation therapy.

[0125] Transplant patients can receive autologous transplants, in which case the transplant contains the recipient's own cells. In other embodiments, the transplant patient may receive an allogeneic transplant, in which case the transplant is , including cells taken from or derived from another individual. In the case of allografting, the engraftment of transplanted cells is The possibility of an immune response to transplantation mediated by host immune cells (host-versus-graft disease). , or the possibility of an immune response against host cells mediated by immune cells present in the transplant. The condition is complex due to its nature (graft-versus-host disease). The possibility of the aforementioned complications is high for the transplant recipient. In relation to the patient, the risk increases with the degree of difference in the antigenic composition of the transplant. Therefore, Seed transplantation typically involves selecting the candidate with the highest possible similarity in HLA antigens and minor histocompatibility antigens. It is performed between patients. There is extremely high antigenic similarity between autologous transplant donors and recipients. Because it is needed, patients who need a transplant cannot find a suitable donor, Some patients are unable to receive treatment.

[0126] In some embodiments, allogeneic HSCs for transplantation are CXCR2 agonists, e.g. For example, MGTA-145, optionally selected, CXCR4 antagonist, for example, Preli By using Xafor or BL-8040 in combination, donors can be recruited. It can be obtained. For example, allogeneic HSCs can be obtained by administering a CXCR2 agonist, or optionally by CX HSCs are mobilized into the peripheral blood by administration of CR2 agonists and CXCR4 antagonists. It can then be obtained through apheresis.

[0127] The method provided herein, at least in part, involves transferring allografts to patients requiring allografts. By conditioning with an ADC that can be coupled to D45, complete mismatch Allogeneic cells, such as those used in allogeneic transplantation, contain a high degree of antigen mismatch with the transplant recipient. This is based on the discovery that engraftment of allogeneic donor cells becomes possible, including under certain circumstances. The CD45-targeting portion (e.g., anti-CD45 ADC) is used in addition to immunosuppressants and other conjugates. It can be administered in an effective dose as monotherapy in the absence of conditioning agents. Therefore, The methods described in the specification increase the engraftment of autologous hematopoietic stem cells in some embodiments. In order to achieve this, and also to induce donor chimerism of bone marrow and peripheral blood (bone marrow chimerism, B cell chimerism) To increase rhythms (including T-cell chimerism) without the use of immunosuppressants. It can be used for that purpose.

[0128] As described herein, hematopoietic stem cell transplantation therapy involves the proliferation of one or more blood cell types. Alternatively, it can be performed on subjects who require treatment in order to promote regrowth. Hematopoietic stem cells are Generally, it exhibits pluripotency, and therefore can be found in multiple different blood lines, for example, not limited to However, granulocytes (e.g., promyelocytes, neutrophils, eosinophils, basophils), red blood cells (e.g., reticulocytes) Blood cells, red blood cells), thrombus cells (e.g., megakaryoblasts, platelet-producing megakaryocytes, platelets), monocytes (e.g., monocytes, macrophages, dendritic cells, microglia, osteoclasts, and lymphocytes (for example) Hematopoietic stem cells can differentiate into NK cells, B cells, and T cells. Hematopoietic stem cells can further self-replicate. This makes it possible to produce daughter cells with capabilities equivalent to those of the mother cell. It can be reintroduced into the transplant recipient, home to its hematopoietic stem cell niche, and live. It is characterized by its ability to reconstruct productive and sustainable hematopoiesis.

[0129] Therefore, hematopoietic stem cells are used in patients who have a deficiency or deletion in one or more cell types of the hematopoietic lineage. It is administered to reconstitute the deficient or missing cell population in vivo, thereby intrinsically It can treat conditions associated with the deficiency or depletion of sex blood cell populations. Therefore, The compositions and methods described herein are for non-malignant abnormal hemoglobin disorders (e.g., sickle cell anemia). Celluloid anemia, thalassemia, Fanconi anemia, aplastic anemia, and Wiscott-Aldrich To be used to treat abnormal hemoglobin disorders (selected from the group consisting of T.S. syndromes). This is possible. Additionally or alternatively, the compositions and methods described herein may be used to treat innate immunity. It can be used to treat immunodeficiency such as insufficiency. In addition or alternatively, this The compositions and methods described in this specification are for acquired immunodeficiency (e.g., HIV and AIDS). It can be used to treat acquired immunodeficiency (selected from the following groups). The compositions and methods described in the details are for metabolic disorders (e.g., glycogen storage disorders, mucopolysaccharides). Is it diabetes mellitus, Gaucher disease, Haller disease, sphingolipidosis, or metachromatic leukodystrophy? It can be used to treat metabolic disorders selected from the following groups.

[0130] Additionally or alternatively, the compositions and methods described herein are for the treatment of hematological cancers, myeloma. It can be used to treat malignant tumors or proliferative disorders such as proliferative diseases. In the case of treatment, the compositions and methods described herein are used before hematopoietic stem cell transplantation therapy. It may be administered to patients to deplete their hematopoietic stem cell population, in which case the transplanted cells are endogenous By homing to the niche formed by the cell depletion step, productive hematopoiesis can be established. This allows for the reconstruction of the cell population that was depleted during cancer cell eradication, such as during systemic chemotherapy. It can be constructed. It can be treated using the compositions and methods described herein. Examples of blood cancers that can be identified include, but are not limited to, acute myeloid leukemia and acute lymphoblastic disease. Myelopathy, chronic myeloid leukemia, chronic lymphocytic leukemia, multiple myeloma, diffuse large cell type B Cellular lymphoma, non-Hodgkin lymphoma, and other cancerous conditions including neuroblastoma It is included.

[0131] Additional diseases that can be treated with the compositions and methods described herein are not limited to: It is not a disease that causes adenosine deaminase deficiency and severe combined immunodeficiency, hyperimmunoglobulinemia. Lymn-M syndrome, Chediak-Dong disease, hereditary lymphohistiocytosis, osteoporosis, osteogenesis imperfecta, Accumulation disease, major thalassemia, systemic scleroderma, systemic lupus erythematosus, multiple sclerosis, and young This includes rheumatoid arthritis.

[0132] The antibodies or their antigen-binding portions and conjugates described herein are parenchymal organs. It can be used to induce transplant tolerance. For example, compositions and methods described herein. This is to deplete or remove cell populations from target tissue (for example, from the bone marrow stem cell niche). It can be used to deplete hematopoietic stem cells. Next, stem cells or progenitor cells from organ donors (for example, hematopoietic stem cells from organ donors) A population of such stem cells or progenitor cells may be administered to the transplant recipient, following the engraftment of such stem cells or progenitor cells. Therefore, temporary or stable mixed chimerism can be achieved, which may lead to further immunosuppressants. Long-term transplant organ tolerance becomes possible without the need for other methods. For example, the combination described herein The product and method are for parenchymal organ transplant recipients (for example, kidney transplants, lung transplants, liver transplants, etc.). It may be used to induce transplant tolerance in organ transplantation and heart transplantation. The compositions and methods described herein, for example, for inducing long-term tolerance of transplanted organs, involve a low proportion of temporary Alternatively, since stable donor engraftment is sufficient, it can be used in connection with inducing parenchymal organ transplant tolerance. It is well suited for that purpose.

[0133] In addition, the compositions and methods described herein are characterized by cells that are CD45+. It can be used to directly treat cancers such as cancer. For example, as described herein. The composition and method described herein are for treating leukemia, such as in patients exhibiting CD45+ leukemia cells. It can be used for leukemia cells using the compositions and methods described herein. By depleting any CD45+ cancer cells, it is possible to directly treat various cancers. In this way, exemplary cancers that can be treated include acute myeloid leukemia and acute lymphoblastic cancer. Leukemia, chronic myeloid leukemia, chronic lymphocytic leukemia, multiple myeloma, diffuse large B cell leukemia This includes hematological cancers such as follicular lymphoma and non-Hodgkin lymphoma.

[0134] In addition, the compositions and methods described herein are used to treat autoimmune disorders. It is possible. For example, an antibody or its antigen-binding portion can kill CD45+ immune cells. Therefore, it can be administered to human patients suffering from autoimmune disorders and other similar conditions. For example, CD45+ immune cells specifically bind to self-antigens and trigger an immune response against them. These may be autoreactive lymphocytes such as T cells that express T cell receptors that cause this reaction. By using the compositions and methods described herein to deplete self-reactive CD45+ cells This can treat autoimmune conditions such as those listed below. Additional or Alternatively, the compositions and methods described herein may be used to prepare endogenous hematopoietic stem cells before hematopoietic stem cell transplantation therapy. It can be used to treat autoimmune diseases by depleting a population of hematopoietic stem cells. In this case, the transplanted cells are housed in the niche formed by the endogenous cell depletion step. This allows for the establishment of productive hematopoiesis. As a result, depletion during the eradication of autoimmune cells The resulting cell population can be reconstructed.

[0135] Antibodies or antibody-drug conjugates must be administered before transplantation of cells or parenchymal organs into the patient. It can be administered to human patients in need. In one embodiment, the CD45-targeting portion (for example) Anti-CD45 ADCs are used in human patients who require them before cell or parenchymal organ transplantation. It is administered to the patient (for example, about 3 days prior, about 2 days prior, about 12 hours prior; about 12 hours to 3 days prior, about (1-3 days prior, approximately 1-2 days prior, or approximately 12 hours to 2 days prior). In one embodiment, transplantation is The CD45-targeting portion (e.g., ADC) disappeared or was substantially eliminated from the patient's blood. Later, it is performed on the patient.

[0136] The methods described herein are also useful for preventing the host-versus-graft (HvG) reaction. Graft failure or rejection, including failure after allogeneic hematopoietic stem cell transplantation, is common. This manifests as either a lack of initial engraftment of donor cells or loss of donor cells after initial engraftment. (For a review article, see Mattsson et al. (2008) Biol Bl) ood Marrow Transplant.14(Suppl 1):165-17 (See reference 0).

[0137] In some embodiments of the methods provided herein, the CD45-targeting portion (e.g.) In the absence of additional conditioning agents such as immunosuppressants, anti-CD45 ADCs are effective. Then it is administered to the target. In a particular embodiment, the CD45-targeting portion (for example, anti CD45 ADCs are calcineurin / MTOR inhibitors (e.g., tacrolimus, cyanoacrylate). Lolimus, rapamycin, cyclosporine, everolimus), co-stimulus blocking molecules (e.g., CTLA4-Ig, anti-CD40L), NK cell depletion agent, anti-thymocyte globulin (ATG), A Lucidizing agents (e.g., nitrogen mustard, e.g., cyclophosphamide; nitro Sourea (e.g., carmustine); platinum compounds), methotrexate, anti-TCR agents (e.g.) For example, muromonab-CD3, anti-CD20 antibodies (e.g., rituximab, ocrelizumab, Ofatumumab and bertuzumab), fludarabine, Campath (alemtuzumab) ), 2-amino-6-aryl-5-substituted pyrimidine (U.S. Patent No. 4,665,077) See above (see reference), the disclosure is incorporated herein by reference), azathioprine (or If there is an adverse reaction to azathioprine, cyclophosphamide is used; bromocriptine Glutaraldehyde (as described in U.S. Patent No. 4,120,649 (listed above)) (To block MHC antigens); anti-idiotype antibodies against MHC antigens; cyclosporine A; One or more steroids, for example, corticosteroids, for example, prednisone, methionine Luprednisolone, hydrocortisone, and dexamethasone are examples of glucocorticosterone. Ido; anti-interferon-γ antibody; anti-tumor necrosis factor-α antibody; anti-tumor necrosis factor-β antibody Anti-interleukin-2 antibodies; anti-IL-2 receptor antibodies and other anti-cytokine receptor antibodies Heterogeneous anti-lymphocyte globulins; pan-T antibodies, e.g., OKT-3 monoclonal antibodies; CD Antibodies against 4; antibodies against CD8, antibodies against CD45 (e.g., 30-F11, YTH24.5 and / or YTH54.12 (for example, YTH24.5 and YTH5 4.12 combinations); streptokinase; streptodolase; or host-derived Administered to the subject in the absence of one or more agents selected from the original RNA or DNA. .

[0138] For example, in some embodiments, the CD45-targeting portion (e.g., anti-CD45 A) DC) is administered to the subject in the absence of whole-body irradiation (TBI) (e.g., low-dose TBI). In other embodiments, the CD45-targeting portion (e.g., anti-CD45 ADC) is It is administered to the subject in the absence of clophosphamide (i.e., cytoxane). In one embodiment, the CD45-targeting moiety (e.g., anti-CD45 ADC) is used in B cells and Immunotherapy such as anti-CD4 antibodies and / or anti-CD8 antibodies that allow for the depletion of T cells. It is administered to the target in the absence of a depletion agent. In some embodiments, CD45 is targeted. The components (e.g., anti-CD45 ADC) include TBI, cytoxane, anti-CD4 antibody, and anti-CD8 It is administered to the subject in the absence of antibodies, or combinations thereof.

[0139] In some embodiments, immunosuppressants (but not limited to anti-CD4 antibodies) Anti-CD8 antibodies, cytoxanes, and / or TBIs are used in allogeneic cells, e.g., allogeneic H It is not administered to patients before transplantation, including SCs. In some embodiments, immunotherapy is used. The inhibitor is not administered to the subject after transplantation. In some embodiments, immunosuppressants It will not be administered to the subjects before or after transplantation.

[0140] In a particular embodiment, the CD45-targeting portion described herein (for example, anti- CD45 antibodies, their antigen-binding moieties, or ADCs, are used by recipients of mismatched allogeneic transplants. It is used to treat mismatched donors. In some embodiments, the donor is a mismatched donor. —The cells, organs, or tissues of a mismatched donor are expressed by the recipient. Compared to the variant, at least one different (e.g., non-identical) major organizational suitability MHC complex antigens (i.e., human leukocyte antigens (HLA) in humans), for example, Class I, Class II, or Class III MHC antigens, or minor histological compatibility antigens (miHA) contains a specified number of MHC antigens or miHA antigens in the blood. Standards used in the relevant technical field, such as biochemical analysis, genome analysis, or molecular analysis. This is determined by an assay. In an exemplary embodiment, allografting is "completely mismatched". "Chi" is an allograft, with one or more major mismatches and one or more minor mismatches. It contains. In another exemplary embodiment, the allograft is the same MH as the transplant recipient. They share a C haplotype or HLA haplotype, but have one or more minor mismatches. It may also contain (for example, minor mismatch allograft). In another exemplary embodiment... In allografting, one or more major mismatches are transplanted, either alone or with one or more minor mismatches. It is included in addition to Smatch.

[0141] MHC proteins play a role in immune responses between lymphocytes and antigen-presenting cells or disease cells. MHC proteins are important for signal transduction, and they bind to peptides. It is presented so that it can be recognized by the T cell receptor. The protein encoded by the MHC gene is , expressed on the cell surface, and distinguish between self-antigens (peptide fragments derived from the cell itself) and non-self-antigens ( For example, both fragments of the invading microorganism are presented to the T cell.

[0142] The MHC region is divided into three subgroups: Class I, Class II, and Class III. MHC class I proteins consist of α-chain and β2-microglobulin (i.e., B It contains 2M and presents antigen fragments to cytotoxic T cells. Most immune system cells, especially In antigen-presenting cells, MHC class II proteins contain α and β chains, and helper - Presents antigen fragments to T cells. The MHC class III region contains corrective components and cytokines. It codes for other immune components, such as those that code for [specific immune components]. MHC is polygenic (MHC (Having multiple genes of class I and MHC class II) and polymorphism (each gene has multiple alleles) It is both (and has).

[0143] In humans, major histocompatibility complexes are also called human leukocyte antigen (HLA) complexes. Each class of MHC is located at several gene loci in humans, for example, class I. Therefore, HLA-A (Human Leukocyte Antigen-A), HLA-B, HLA-C, HLA-E, HLA-F, HLA-G, HLA-H, HLA-J, HLA-K, HLA-L, HLA- P and HLA-V, and for Class II, HLA-DRA, HLA-DRB1- 9, HLA-, HLA-DQA1, HLA-DQB1, HLA-DPA1, HLA-DP B1, HLA-DMA, HLA-DMB, HLA-DOA, and HLA-DOB MHC exhibits extremely high polymorphism, and in the human population, each gene locus There are a great many haplotypes, each containing a different allele. Class I and Class II In both cases, different polymorphic MHC alleles have different peptide specificities, and each allele has specific It encodes a protein that binds to peptides exhibiting a specific sequence pattern. HLA genomic genetics Methods for testing HLA alleles or proteins in gestational loci and in humans are available in the art. It is stated in (for example, Choo et al. (2007) Yonsei m edical journal.48.1:11-23;Shiina et al.( 2009)Journal of human genetics.54.1:15;P etersdorf.(2013).Blood.122.11:1863-1872; and Bertaina and Andreani.(2018).Internati onal journal of molecular sciences.19.2: See 621, which in whole is incorporated herein by reference.

[0144] In some embodiments, at least one major histocompatibility complex antigen (e.g.) (HLA antigen) is transplanted to the transplant donor according to the method provided herein. There is a mismatch in the target. In a particular embodiment, the MHC antigen is MHC It is a Class I molecule or an MHC Class II molecule. In certain embodiments, it is an MHC antigen. are B2M, HLA-A, HLA-B, HLA-C, HLA-DRA, HLA-DRB1 , HLA-DRB3, HLA-DRB4, HLA-DRB5, HLA-DPA1, HLA - One or any of DPA2, HLA-DQA1, and / or HLA-DQB1 It is a combination of intents. In some embodiments, transplantation involves the HLA antigen of a human patient. In contrast, it contains allogeneic hematopoietic stem cells that include at least one HLA mismatch. For example, In certain cases, allogeneic hematopoietic stem cells have at least 1 HLA antigen for a human patient. at least two, at least three, at least four, at least five, at least six at least 7, at least 8, at least 9, or more than 9 HLA misses Includes a match. In some embodiments, allogeneic hematopoietic stem cells are used with the HLA antigen of a human patient. This includes complete HLA mismatch.

[0145] Alternatively or additionally, at least one minor histocompatibility antigen may be present in the donor. There is a mismatch in the subjects receiving transplantation according to the method provided herein. In one embodiment, the transplantation involves at least one m for the miHA antigen of a human patient. Includes allogeneic hematopoietic stem cells, including iHA mismatches. For example, in a particular case, allogeneic Hematopoietic stem cells have at least one, at least two, and few miHA antigens in a human patient. At least three, at least four, at least five, at least six, at least seven, few It contains at least 8, at least 9, or more than 9 miHA mismatches. In a specific embodiment, the minor histocompatibility antigens are HA-1, HA-2, HA-8, and HA -3, HB-1, HY-Al, HY-A2, HY-B7, HY-B8, HY-B60, This is the HY-DQ5 protein. Examples of other minor histocompatibility antigens in the art include It is known in (for example, Perreault et al. (1990).Blo od.76.7:1269-1280;Martin et al.(2017).Bl ood.129.6:791-798; and U.S. Patent No. US10414813B2; (These are all incorporated herein by reference.)

[0146] In some embodiments, the method completely or entirely in the transplant recipient. Close donor chimerism, for example, at least 80% donor chimerism in the transplant recipient Merism (for example, at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 99%, or approximately 100% donor chimerism This is effective in establishing ). The level of donor chimerism after allogeneic HSC transplantation is, for example, Total chimerism, bone marrow chimerism, peripheral chimerism, bone marrow chimerism, B cell chimerism Alternatively, it could be T-cell chimerism.

[0147] Route of administration and dosage CD45 targeting moieties described herein (e.g., anti-CD45 antibodies, their antigen-binding moieties) (Minute, or ADC) is available in various dosage forms for patients (e.g., cancer, autoimmune diseases) It can be administered to human patients who require hematopoietic stem cell transplantation therapy. If a CD45 targeting moiety described herein (e.g., an anti-CD45 antibody, its antigen binding) is used, then... A portion (or ADC) is an aqueous solution containing one or more pharmaceutically acceptable excipients. In aqueous solution form, it is used for patients with cancer, autoimmune diseases, or those requiring hematopoietic stem cell transplantation. It can be administered to patients in need. It can be used in conjunction with the compositions and methods described herein. The pharmaceutically acceptable excipients used include viscosity modifiers. The aqueous solution is in the art. It can be sterilized using known techniques.

[0148] Anti-CD45 antibody, its antigen-binding moiety, or conjugate (e.g., as described herein) A pharmaceutical formulation containing such an antibody or ADC is one or more of the following: Pharmacologically acceptable carrier (Remington's Pharmaceutical S Sciences 16th edition, Osol, A. Ed. (1980) By combining, it is prepared in the form of a lyophilized preparation or an aqueous solution. Pharmacologically acceptable. The carrier is generally non-toxic to the recipient at the dosage and concentration used, Buffers such as phosphates, citrates, and other organic acids; containing ascorbic acid and methionine. Antioxidants; preservatives (such as octadecyldimethylbenzylammonium chloride); hexachloride Samethonium; benzalkonium chloride; benzethonium chloride; phenol, butyl or Benzyl alcohol; alkylparabens such as methylparaben or propylparaben; Catechol; Resorcinol; Cyclohexanol; 3-Pentanol; and m-Crezo Polypeptides with low molecular weight (less than approximately 10 residues); serum albumin, gelatin, etc. Proteins such as immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; Lysine, glutamine, asparagine, histidine, arginine, or other amino acids such as lysine, glutamine, asparagine, histidine, arginine, or lysine. No acids; monosaccharides, disaccharides, and other carbohydrates including glucose, mannose, or dextrin. Modified substances; chelating agents such as EDTA; sucrose, mannitol, trehalose or so Sugars such as rubitol; salt-forming counterions such as sodium; metal complexes (e.g., Zn-tan) Protein complexes; and / or nonionic surfactants such as polyethylene glycol (PEG) This includes, but is not limited to, aphrodisiacs.

[0149] CD45 targeting moieties described herein (e.g., anti-CD45 antibodies, antigen-binding moieties, Or ADC) can be administered orally, transdermally, subcutaneously, intranasally, intravenously, intramuscularly, intraocularly, or parenterally. It can be administered by various routes. The most suitable route of administration in any given case is... This includes the specific antibody or antigen-binding portion to be administered, the patient, the pharmaceutical formulation method, and the method of administration (for example, (Time and route of administration), patient's age, weight, sex, severity of the disease being treated, patient's diet It will vary depending on the circumstances and the patient's rate of excretion.

[0150] CD45 targeting moieties described herein (e.g., anti-CD45 antibodies or antigen binding) The effective dose or amount of the partial or antibody-drug conjugate is preferably immunosuppressive. In the absence of inhibitors, for example, in the absence of total body irradiation (TBI), in the absence of anti-CD4 antibodies, and after allogeneic transplantation (e.g., allogeneic HSC transplantation) in the absence of anti-CD8 antibodies This is a sufficient amount to achieve complete or near-complete donor chimerism. For example, this book The anti-CD45 antibody, antigen-binding moiety, or antibody-drug conjugate described in the specification The efficacy is measured in the absence of immunosuppressants, for example, in the absence of total body irradiation (TBI), and in the absence of anti-CD4 antibodies. In the absence of and / or in the absence of anti-CD8 antibodies, complete mismatch allograft (e.g.) After receiving a completely mismatched allogeneic HSC transplant, at least 80% donor chimerism This may be a sufficient amount to achieve the anti-CD45 antibody, antigen-binding moiety described herein. , or the effective dose of an antibody-drug conjugate is, when used as monotherapy, anti-CD4 5. The antibody, antigen-binding moiety, or antibody-drug conjugate is an immunosuppressant, e.g., TBI. It is administered together with other conditioning agents such as anti-CD4 and / or anti-CD8. It's acceptable to pay more than the usual price.

[0151] In exemplary embodiments, the CD45 targeting moiety (e.g., anti-CD45 antibody, antigen binding) is used. The effective dose (partial or antibody-drug conjugate) is in the absence of other conditioning agents. After transplantation of allogeneic HSCs below, at least 80% donor chimerism (e.g., at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, Achieve at least 98%, at least 99%, or approximately 100% donor chimerism. That is a sufficient amount.

[0152] In other exemplary embodiments, the CD45-targeting moiety (e.g., anti-CD45 antibody, antigen) is used. The effective amount of the binding portion (or antibody-drug conjugate) is the non-conjugate of other conditioning agents. In the presence of allogeneic HSCs, at least 80% of bone marrow chimerisms occur after transplantation (for example, at least 80%, at least 85%, at least 90%, at least 95%, at least 97% Achieve at least 98%, at least 99%, or approximately 100% bone marrow chimerism. That is a sufficient amount.

[0153] In other exemplary embodiments, an anti-CD45 antibody, an antigen-binding moiety, or an antibody-drug compound may be used. The effective dose of jugate is determined after transplantation of allogeneic HSCs in the absence of other conditioning agents. At least 80% B-cell chimerism (e.g., at least 80%, at least 85%) At least 90%, at least 95%, at least 97%, at least 98%, and This is also a sufficient amount to achieve 99% or approximately 100% B-cell chimerism.

[0154] In other exemplary embodiments, an anti-CD45 antibody, an antigen-binding moiety, or an antibody-drug compound may be used. The effective dose of jugate is determined after transplantation of allogeneic HSCs in the absence of other conditioning agents. At least 80% T-cell chimerism (e.g., at least 80%, at least 85%) At least 90%, at least 95%, at least 97%, at least 98%, and This is also a sufficient amount to achieve 99% or approximately 100% T-cell chimerism.

[0155] The effective dose of the anti-CD45 antibody, antigen-binding moiety, or ADC described herein is single Per single (e.g., bolus) dose, multiple doses, or consecutive doses, or per antibody or The optimal serum concentration of the antigen-binding fragment is (e.g., approximately 0.0001 to approximately 5000 μg / m³). To achieve a serum concentration of L, for example, approximately 0.001 to approximately 100 mg / kg body weight It may be within the range. Administration is for cancer, autoimmune diseases, or hematopoietic stem cell transplantation. In preparation for receiving this treatment, subjects (e.g., humans) undergoing conditioning therapy... It may be implemented at least once a day, once a week, or once a month (for example, 2 to 10 times).

[0156] In certain embodiments, the CD45 targeting moiety (e.g., anti-CD45 antibody or A) DC) is administered to the patient as a single dose. In other embodiments, CD45 is targeted. The portion (e.g., anti-CD45 antibody or ADC) is administered to the patient in divided doses, and anti The dose of the CD45-targeting portion (e.g., CD45 antibody or ADC) is divided and spaced apart. It is administered to the target after leaving a gap. For example, in a divided dosing regimen, the CD45 targeting portion (e.g. For example, the dosage of anti-CD45 antibody (or ADC) is 2, 3, 4, 5, 6, 7, 8, 9, and It can be divided into 10 portions, and each portion is administered to the subject at intervals. In this embodiment, the intervals are 1 hour, 3 hours, 6 hours, 9 hours, 12 hours, 15 hours, 1 8 hours, 21 hours, 24 hours, 36 hours, 48 ​​hours, 72 hours, 96 hours, 120 hours, 1 week, 1.5 weeks, 2 weeks, 2.5 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 7 weeks, Alternatively, an interval of 8 weeks may be observed. In some embodiments, as described herein The CD45-targeting portion (e.g., anti-CD45 antibody or ADC) is administered to the patient in two doses. It is administered to the patient as a divided dose. In some embodiments, as specified herein The CD45-targeting portion described (e.g., anti-CD45 antibody or ADC) is in three quantities. It is administered to the patient as a divided dose. In some embodiments, The CD45-targeting moieties described herein (e.g., anti-CD45 antibodies or ADCs) are As a divided dose, two or three doses are administered to the patient at intervals of 1 to 7 days. It is administered to the patient. In some embodiments, the CD45 targeting described herein The portion (e.g., anti-CD45 antibody or ADC) is administered in two or three doses over a period of 1 to 3 days. It is administered to the patient as divided doses, with intervals between doses.

[0157] In one embodiment, an anti-CD45 ADC (e.g., a linker) is administered to a human patient. The dosage of anti-CD45 antibodies conjugated to cytotoxicity via a cytotoxic agent is approximately 3 mg / kg to approximately 1 The concentration is 2 mg / kg.

[0158] In one embodiment, an anti-CD45 ADC (e.g., a linker) is administered to a human patient. The dose of anti-CD45 antibody conjugated to cytotoxicity via is approximately 3.5 mg / kg. It is approximately 10 mg / kg.

[0159] In one embodiment, an anti-CD45 ADC (e.g., a linker) is administered to a human patient. The dosage of anti-CD45 antibodies conjugated to cytotoxicity via a cytotoxic agent is approximately 4 mg / kg to approximately 8 mg / kg. It is mg / kg.

[0160] In one embodiment, an anti-CD45 ADC (e.g., a linker) is administered to a human patient. The dosage of anti-CD45 antibodies conjugated to cytotoxicity via a cytotoxic agent is approximately 4 mg / kg to approximately 6 It is mg / kg.

[0161] In another embodiment, an anti-CD45 ADC (e.g., Linker) is administered to a human patient. The dose of anti-CD45 antibody conjugated to cytotoxicity via [a specific method] is approximately 0.1 mg / kg. It is approximately 0.3 mg / kg.

[0162] In one embodiment, an anti-CD45 ADC (e.g., a linker) is administered to a human patient. The dose of anti-CD45 antibody conjugated to cytotoxicity via [a specific method] is approximately 0.15 mg / kg. It is approximately 0.3 mg / kg.

[0163] In one embodiment, an anti-CD45 ADC (e.g., a linker) is administered to a human patient. The dose of anti-CD45 antibody conjugated to cytotoxicity via [a specific method] is approximately 0.15 mg / kg. It is approximately 0.25 mg / kg.

[0164] In one embodiment, an anti-CD45 ADC (e.g., a linker) is administered to a human patient. The dose of anti-CD45 antibody conjugated to cytotoxicity via is approximately 0.2 mg / kg. It is approximately 0.3 mg / kg.

[0165] In one embodiment, an anti-CD45 ADC (e.g., a linker) is administered to a human patient. The dose of anti-CD45 antibody conjugated to cytotoxicity via is approximately 0.25 mg / kg It is approximately 0.3 mg / kg.

[0166] In one embodiment, an anti-CD45 ADC (e.g., a linker) is administered to a human patient. The dose of the anti-CD45 antibody conjugated to cytotoxicity via is approximately 0.1 mg / kg. be.

[0167] In one embodiment, an anti-CD45 ADC (e.g., a linker) is administered to a human patient. The dose of the anti-CD45 antibody conjugated to a cytotoxic via is approximately 0.2 mg / kg. be.

[0168] In one embodiment, an anti-CD45 ADC (e.g., a linker) is administered to a human patient. The dose of the anti-CD45 antibody conjugated to a cytotoxic agent is approximately 0.3 mg / kg. be.

[0169] In some embodiments, the anti-CD45 described herein is administered to a human patient. The dosage of ADC is approximately 0.001 mg / kg to 10 mg / kg, and approximately 0.01 mg / kg to 9.5mg / kg, about 0.1mg / kg~9mg / kg, about 0.1mg / kg~8.5m g / kg, about 0.1mg / kg~8mg / kg, about 0.1mg / kg~7.5mg / kg , about 0.1mg / kg~7mg / kg, about 0.1mg / kg~6.5mg / kg, about 0. 1mg / kg~6mg / kg, approx. 0.1mg / kg~5.5mg / kg, approx. 0.1mg / kg kg~5mg / kg, about 0.1mg / kg~4.5mg / kg, about 0.1mg / kg~4 mg / kg, about 0.5mg / kg~3.5mg / kg, about 0.5mg / kg~3mg / k g, about 1mg / kg~10mg / kg, about 1mg / kg~9mg / kg, about 1mg / kg ~8mg / kg, approx. 1mg / kg~7mg / kg, approx. 1mg / kg~6mg / kg, approx. 1 mg / kg to 5 mg / kg, approximately 1 mg / kg to 4 mg / kg, or approximately 1 mg / kg to 3 mg / kg The dose is mg / kg. In some embodiments, it is administered to human patients as described herein. The dosages of the anti-CD45 ADCs used are approximately 12 mg / kg, 11 mg / kg, and 10 mg / kg. / kg, approximately 9mg / kg, approximately 8mg / kg, approximately 7mg / kg, approximately 6mg / kg, approximately 5mg / kg, approximately 4mg / kg, approximately 3mg / kg, approximately 2mg / kg, approximately 1mg / kg, or approximately The concentration is 0.5 mg / kg.

[0170] In one embodiment, the anti-CD45 ADC described herein is administered to a human patient. This includes periods of 24 hours or less, 22 hours or less, 20 hours or less, 18 hours or less, 16 hours or less, and 14 hours. time or less, 13 hours or less, 12 hours or less, 11 hours or less, 10 hours or less, 9 hours or less, 8 o'clock It has a half-life of less than 1, 7 hours or less, 6 hours or less, or 5 hours or less. In one embodiment The half-lives of anti-CD45 ADCs are 5-7 hours, 5-9 hours, and 15-11 hours. Between 5 hours and 13 hours, 5 hours and 15 hours, 5 hours and 20 hours, 5 hours and 24 hours, 7 hours Intervals: ~24 hours, 9 hours ~24 hours, 11 hours ~24 hours, 12 hours ~22 hours, 10 hours These are approximately 20 hours, 8 to 18 hours, or 14 to 24 hours.

[0171] In a particular embodiment, the CD45 targeting portion described herein (for example, A The effective dose of DC is administered in a single dose. For example, a single dose is administered in one or more additional doses. In the absence of conditioning agents, for example, whole-body irradiation (TBI), anti-CD4 antibodies, and / or In the absence of immunosuppressants such as anti-CD8 antibodies, allogeneic transplantation (e.g., allogeneic HSC transplantation) is performed. After administration, it may contain enough to achieve at least 80% donor chimerism. In an exemplary embodiment, a single dose is administered in the absence of an immunosuppressant, for example, during whole-body irradiation (TB). In the absence of I), in the absence of anti-CD4 antibody, and / or in the absence of anti-CD8 antibody, complete After undergoing a complete mismatch allogeneic transplant (for example, a totally mismatched allogeneic HSC transplant), at least It may also contain a sufficient amount to achieve 80% donor chimerism.

[0172] In some embodiments, an effective amount of CD45-targeting portion (e.g., ADC) is 2 It is administered in doses exceeding the initial dose (e.g., divided doses). For example, the subject is AD with the first dose. C can then receive a second dose of ADC, and each of the first and second doses This is in the absence of immunosuppressants, for example, in the absence of total body irradiation (TBI), and in the absence of anti-CD4 antibodies. Allogeneic transplantation (e.g., allogeneic HSC transplantation) in the presence and / or absence of anti-CD8 antibody After receiving the treatment, it contains about half the amount necessary to achieve at least 80% donor chimerism. In some embodiments, allografting is a complete mismatch allograft, for example, complete This is a mismatched allogeneic HSC transplant. In some embodiments, an effective amount of ADC is 2 The drug is administered in doses of 3 or more, 4 or more, or 5 or more.

[0173] In one embodiment, the method disclosed herein is used for conditioning anti-C To minimize hepatotoxicity in patients receiving D45 ADC. For example, in a certain embodiment In this specification, the patient's liver marker levels are reduced over 24 hours, 48 ​​hours, by the method disclosed herein. The toxicity levels remain below known levels for more than 72 hours, or 96 hours. Furthermore, by the method disclosed herein, the patient's liver marker levels are measured over 24 hours and 48 hours. During the interval, 72 hours, or more than 96 hours, it remains within the reference range. In a particular embodiment, The method disclosed herein allows liver marker levels to be measured at 24 hours, 48 ​​hours, and 72 hours. , or more than 96 hours, more than 1.5 times beyond the reference range, more than 3 times beyond the reference range, reference The level should not exceed the range by more than 5 times, or exceed the reference range by more than 10 times. (For toxicity testing) Examples of liver markers that can be used include alanine aminotransferase (ALT), Examples include lactate dehydrogenase (LDH) and aspartate aminotransferase (AST). In a particular embodiment, the administration of the ADC described herein, i.e., a single dose If two doses are administered instead of a single dose, liver markers such as AST and LDH should be used. , and / or a transient increase in ALT occurs. In some cases, the liver shows toxicity. It is possible to raise the marker level, but only within a specific time frame, for example, about 12 hours. Approximately 18 hours, approximately 24 hours, approximately 36 hours, approximately 48 hours, approximately 72 hours, more than 3 days, approximately 3.5 days, approximately 4 days, approximately 4.5 days, approximately 5 days, approximately 5.5 days, approximately 6 days, approximately 6.5 days, approximately Within 7 days, approximately 7.5 days, or less than a week, liver marker levels are not related to hepatotoxicity. It does not return to normal levels. For example, in humans (average adult males), normal abnormal ALT levels The toxicity level is 7-55 units / liter (U / L), which is within the normal non-toxic level according to AST. The values ​​are 8-48 U / L. In a particular embodiment, the patient's blood AST, ALT, Or at least one of the LDH levels from the administration of the first dose of ADC to the patient. The patient does not reach a toxic level within 14 days after administration of the first dose. For example, the patient receives the first dose The patient is administered a dose, and then a second dose, a third dose, a fourth dose, or a higher dose. For example, it may be administered within 5, 10, or 14 days after the first dose has been administered. Nevertheless, at least one of the patient's blood AST, ALT, or LDH levels The toxicity level was observed from the administration of the first dose of ADC to the patient until 14 days after the administration of the first dose. It does not reach its goal.

[0174] In a particular embodiment, the patient's blood AST, ALT, or LDH levels At least one of them did not rise above normal levels, but was more than 1.5 times above normal levels. Not rising, not rising more than 3 times above normal level, not rising more than 5 times above normal level It does not occur, or does not rise more than 10 times above normal levels.

[0175] In the case of conditioning treatment before hematopoietic stem cell transplantation, the CD45 targeting portion (for example, anti- CD45 antibodies, their antigen-binding fragments, or ADCs, optimize the engraftment of exogenous hematopoietic stem cells. The timing of the accelerating process, for example, approximately 1 hour to 1 week after the exogenous hematopoietic stem cell transplant (for example, Approximately 1 hour, approximately 2 hours, approximately 3 hours, approximately 4 hours, approximately 5 hours, approximately 6 hours, approximately 7 hours, approximately 8 hours, Approximately 9 hours, approximately 10 hours, approximately 11 hours, approximately 12 hours, approximately 13 hours, approximately 14 hours, approximately 15 hours Approximately 16 hours, approximately 17 hours, approximately 18 hours, approximately 19 hours, approximately 20 hours, approximately 21 hours, approximately 22 hours (Time: approximately 23 hours, approximately 24 hours, approximately 2 days, approximately 3 days, approximately 4 days, approximately 5 days, approximately 6 days, approximately 7 days) It can be administered to the patient earlier or even earlier. The range includes the numbers listed herein. It is also included in the intended method.

[0176] The above dose range is in combination with anti-CD45 ADCs having half-lives listed herein. You may let them do it.

[0177] Using the methods disclosed herein, a physician in the art can perform hematopoietic stem cell transplantation. In human patients requiring therapy, antigens expressed by hematopoietic stem cells (e.g., CD45) Alternatively, it binds to antigens expressed by mature immune cells such as T cells (e.g., CD45). Administer a targeting portion (e.g., ADC, antibody, or antigen-binding fragment) that can be targeted. This can be done. In this way, before administration of the exogenous hematopoietic stem cell graft, endogenous hematopoietic stem cells The population is depleted, and the engraftment of hematopoietic stem cell grafts can be promoted. Antibodies are specified herein. Toxins such as cytotoxic molecules described or known in the relevant art are shared. They may be conjugated. For example, an anti-CD45 antibody or its antigen-binding fragment. These are Pseudomonas exotoxin A, debuganin, diphtheria toxin, □-amanitin, α-a Amatoxins such as manitin, saporins, mytansins, maytansinoids, and aurista Chin, Anthracycline, Calicae Sewing Machine, Irinotecan, SN-38, Duokarma Icin, pyrrolobenzodiazepine, pyrrolobenzodiazepine dimer, indolinobenzodiazepine Azepines, indolinobenzodiazepine dimers, or their variants are associated with cytotoxicity. They can be conjugated in a coupled manner. This conjugation is described herein. This is carried out using covalent bond formation techniques described or known in the relevant art. It is possible. Subsequently, the antibody, its antigen-binding fragment, or drug-antibody conjugate can affect the patient. Before transplanting exogenous hematopoietic stem cells (such as autologous, syngeneic, or allogeneic hematopoietic stem cells) into a patient, for example, It can be administered to patients intravenously.

[0178] Anti-CD45 antibodies, their antigen-binding moieties, or drug-antibody conjugates are used in hematopoietic stem cell transfer. Before grafting therapy, the amount of endogenous hematopoietic stem cells is adjusted, for example, to approximately 10%, 20%, 30%, and 4%. 0%, approximately 50%, approximately 60%, approximately 70%, approximately 80%, approximately 90%, approximately 95%, or more. It can be administered in an amount sufficient to reduce it. The reduction in hematopoietic stem cell count is relevant to the technical field. Using conventional techniques known in the field, for example, patients during conditioning therapy Cells expressing characteristic hematopoietic stem cell surface antigens in blood samples collected at various intervals. FACS analysis can be used for monitoring. For example, by physicians in the relevant field. If available, blood samples will be taken from the patient at various points during conditioning therapy, and hematopoietic stem cells will be examined. The relative concentration of hematopoietic stem cells in a sample is determined using antibodies that bind to cell marker antigens. By performing FACS analysis, the degree of decrease in endogenous hematopoietic stem cells can be determined. Yes, it is possible. According to some embodiments, an anti-CD45 antibody, its antigen-binding fragment, or a drug Hematopoietic stem cell concentration reaches minimum in response to conditioning therapy with antibody conjugates. If this is reached, the doctor will discontinue conditioning therapy and move towards hematopoietic stem cell transplantation therapy. We can begin preparing the patient.

[0179] CD45 targeting portion (e.g., anti-CD45 antibody, its antigen-binding portion, or drug antibody) (Njugate) is a water containing one or more pharmaceutically acceptable excipients, such as viscosity modifiers. It can be administered to the patient in solution. The aqueous solution is as described herein or the technical components thereof. Sterilization may be performed using techniques known in the field. Anti-CD45 antibody, its antigenicity The combined portion, or drug-antibody conjugate, should be administered to the patient before the hematopoietic stem cell graft is given, for example. For example, approximately 0.001 mg / kg to approximately 100 mg / kg, and approximately 0.001 mg / kg to approximately 10 mg g / kg, about 0.01mg / kg~9.5mg / kg, about 0.1mg / kg~9mg / k g, about 0.1mg / kg~8.5mg / kg, about 0.1mg / kg~8mg / kg, about 0 .1mg / kg~7.5mg / kg, approx. 0.1mg / kg~7mg / kg, approx. 0.1mg / kg~6.5mg / kg, approx. 0.1mg / kg~6mg / kg, approx. 0.1mg / kg~ 5.5mg / kg, about 0.1mg / kg~5mg / kg, about 0.1mg / kg~4.5m g / kg, about 0.1mg / kg~4mg / kg, about 0.5mg / kg~3.5mg / kg , about 0.5mg / kg~3mg / kg, about 1mg / kg~10mg / kg, about 1mg / k g~9mg / kg, approx. 1mg / kg~8mg / kg, approx. 1mg / kg~7mg / kg, approx. 1mg / kg~6mg / kg, about 1mg / kg~5mg / kg, about 1mg / kg~4mg It can be administered to patients at a dose of / kg, or approximately 1mg / kg to 3mg / kg. Anti-CD45 antibodies, their antigen-binding moieties, or drug-antibody conjugates are exogenous hematopoietic stem cells. The optimal time to promote cell engraftment, for example, about 1 hour before performing exogenous hematopoietic stem cell transplantation. One week (for example, about 1 hour, about 2 hours, about 3 hours, about 4 hours, about 5 hours, about 6 hours, about 7 hours) Duration: approximately 8 hours, approximately 9 hours, approximately 10 hours, approximately 11 hours, approximately 12 hours, approximately 13 hours, approximately 14 hours Time: approximately 15 hours, approximately 16 hours, approximately 17 hours, approximately 18 hours, approximately 19 hours, approximately 20 hours, approximately 21 hours, approximately 22 hours, approximately 23 hours, approximately 24 hours, approximately 2 days, approximately 3 days, approximately 4 days, approximately 5 days, approximately It can be administered to the patient 6 days, or about 7 days, or more in advance.

[0180] In some embodiments, the CD45-targeting moiety (e.g., an anti-CD45 antibody) is used. The protobinding portion (ADC) can be used as monotherapy, for example, as an additional conditioning agent. It is administered in the absence of [something]. For example, in certain embodiments, the CD45-targeting moiety (e.g. For example, anti-CD45 ADCs are administered in the absence of additional immunosuppressants. In some embodiments, the CD45-targeting moiety conjugated to the toxin provided herein The recipients of this drug are not simultaneously receiving immunosuppressant therapy. In some embodiments... The subjects were those who had experienced the effects of immunosuppressant therapy at the time of administration of the CD45-targeted portion. No. In some embodiments, the subject has a small amount of CD45 targeted portion before administration. At least 3 days, at least 7 days, at least 14 days, at least 21 days, less Both patients will receive immunosuppressant medication for 28 days, at least one month, or at least two months. Not included. In some embodiments, the subject is subjected to the administration of the CD45-targeting portion after , at least 3 days, at least 7 days, at least 14 days, at least 21 days, Immunosuppressant administration for at least 28 days, at least 1 month, or at least 2 months. It has not received treatment. In some embodiments, the immunosuppressant is an anti-CD4 antibody or The antigen-binding portion, the anti-CD8 antibody or its antigen-binding portion, whole-body irradiation (e.g., low-dose T) Contains BI, and / or cyclophosphamide.

[0181] After the completion of conditioning therapy, the patient then undergoes conditioning therapy again. Injection of exogenous hematopoietic stem cells (e.g., intravenous injection) from the same doctor or a different doctor. ) can be received. A doctor may inject autologous, allogeneic, or allogeneic hematopoietic stem cells, for example. , 1 x 10 3 ~1 × 10 9It can be administered to patients at a dose of one hematopoietic stem cell / kg. For example, a doctor might take a blood sample from a patient and, after the transplant, examine the hematopoietic stem cells or hematopoietic system. Cells (megakaryocytes, stent cells, platelets, erythrocytes, mast cells, myeloblasts, basophils, neutrophils, eosinophils) Spheres, microglia, granulocytes, monocytes, osteoclasts, antigen-presenting cells, macrophages, dendritic cells This determines the increase in the concentration of natural killer cells, T lymphocytes, and B lymphocytes. This allows for monitoring the engraftment of hematopoietic stem cells after transplantation. This analysis, for example, For example, after hematopoietic stem cell transplantation therapy, 1 hour to 6 months or longer (for example, about 1 hour, about 2 Time: approximately 3 hours, approximately 4 hours, approximately 5 hours, approximately 6 hours, approximately 7 hours, approximately 8 hours, approximately 9 hours, approximately 1 0 hours, approximately 11 hours, approximately 12 hours, approximately 13 hours, approximately 14 hours, approximately 15 hours, approximately 16 hours, Approximately 17 hours, approximately 18 hours, approximately 19 hours, approximately 20 hours, approximately 21 hours, approximately 22 hours, approximately 23 hours Between approximately 24 hours, 2 days, 3 days, 4 days, 5 days, 6 days, 7 days, and 2 Weeks, approximately 3 weeks, approximately 4 weeks, approximately 5 weeks, approximately 6 weeks, approximately 7 weeks, approximately 8 weeks, approximately 9 weeks, approximately 1 0 weeks, approximately 11 weeks, approximately 12 weeks, approximately 13 weeks, approximately 14 weeks, approximately 15 weeks, approximately 16 weeks, Approximately 17 weeks, approximately 18 weeks, approximately 19 weeks, approximately 20 weeks, approximately 21 weeks, approximately 22 weeks, approximately 23 weeks This can be performed during the interval (approximately 24 weeks or longer). After transplantation therapy, hematopoietic stem cells or hematopoiesis may be transplanted. The concentration of the cell lineage increased compared to the concentration of the corresponding cell type before transplantation (e.g., about 1%). Approximately 2%, approximately 3%, approximately 4%, approximately 5%, approximately 6%, approximately 7%, approximately 8%, approximately 9%, approximately 10%, approximately 20% %, approximately 30%, approximately 40%, approximately 50%, approximately 60%, approximately 70%, approximately 80%, approximately 90%, approximately 10 Findings of 0%, approximately 200%, approximately 500%, or more indicate anti-CD45 antibody, and Treatment with antigen-binding moieties or drug-antibody conjugates is performed on transplanted hematopoietic stem cells. This serves as one indicator that we have successfully promoted the engraftment of the graft.

[0182] CD45 targeting portion (e.g., anti-CD45 antibody, its antigen-binding portion, or ADC) The engraftment of hematopoietic stem cells transplanted after administration can be demonstrated by various empirical measurements. For example, The engraftment of transplanted hematopoietic stem cells depends on the antigens expressed by the hematopoietic stem cells (e.g., CD4). 5) Administration of an antibody or its antigen-binding portion capable of binding to it, and subsequent hematopoietic stem cell transfer After the implantation, the amount of competitive reconstruction units (CRUs) present in the patient's bone marrow will be evaluated. Therefore, it can be evaluated. Furthermore, fluorescence, color development, or A reporter gene that produces a luminescence product is incorporated into a vector, and then used with donor hematopoietic stem cells. It is transfected, and then the hematopoietic stem cells home to tissues such as bone marrow. By monitoring the corresponding signals, we can observe the engraftment of hematopoietic stem cells. This can be done. Also, for example, fluorescently labeled cell sorting (FAC) known in the relevant technology S) As determined by the analytical method, evaluate the quantity and viability of hematopoietic stem cells and progenitor cells. By doing so, it is also possible to observe hematopoietic stem cell engraftment. Engraftment can also be observed during the post-transplant period. By measuring the number of white blood cells in peripheral blood and / or in bone marrow aspirate samples, the donor This can also be determined by measuring the recovery of bone marrow cells by the cells themselves.

[0183] Anti-CD45 antibody In certain embodiments of this disclosure, CD45 (hematopoietic stem cells or mature immune cells (e.g.) Antibodies that can bind to CD45+ cells (such as T cells) or The antigen-binding portion is used as a therapeutic agent alone or as an antibody-drug conjugate (ADC). (i) to treat cancers and autoimmune diseases characterized by CD45+ hematopoietic cells, and (ii) ) Used to promote the engraftment of transplanted hematopoietic stem cells in patients requiring transplantation therapy. These therapeutic activities can be used, for example, to stimulate hematopoietic cells (e.g., hematopoietic stem cells), white Blood cells, or immune cells, such as mature immune cells (e.g., T cells), or cancer cells. CD45 expressed by autoimmune cells or hematopoietic stem cells is treated with an anti-CD45 antibody or This can occur when the antigen-binding fragment binds, followed by the induction of cell death. Endogenous The depletion of hematopoietic stem cells provides a niche for transplanted hematopoietic stem cells to home in, After this, productive hematopoiesis can be established. In this way, the transplanted hematopoietic stem cells are used in this specification. The stem cells can successfully engraft in patients, such as human patients suffering from stem cell disorders as described in the book.

[0184] The anti-CD45 antibodies described herein include full-length antibodies, bispecific antibodies, and bivariable antibodies. Antibodies, multi-chain or single-chain antibodies, and / or cleavage that specifically bind to human CD45. For example, although not limited to, Fab, Fab', (Fab')2, Fv), s cFv (single-stranded Fv), sarobody (including alternative light chain constructs), single-domain antiviral It can be in the form of a body, camel antibodies (camelized antibodies), etc. Antibodies also include, for example, IgA (e.g., IgA1 or IgA2), IgD, IgE, Contains IgG (e.g., IgG1, IgG2, IgG3, or IgG4), or IgM. It may be of any isotype or derived therefrom. In some embodiments In this context, anti-CD45 antibodies are IgG (e.g., IgG1, IgG2, IgG3, or Ig G4)

[0185] The antibodies for use in connection with the methods described herein include the above-mentioned antibodies. Variants, for example, antibody fragments containing or not containing an Fc domain, and this specification Humanized variants of non-human antibodies described in this document, and antibodies or anti- Antibody-like tannins containing one or more or all of the CDRs or equivalent regions of the body fragments. Protein scaffold (for example, 10 (Includes the Fn3 domain) Exemplary example of the antibody mentioned above Among the antigen-binding fragments are, in particular, the bivariable immunoglobulin domain and the single-stranded Fv molecule (s cFv), diabody, triabody, nanobody, antibody-like protein scaffold This includes Fv fragments, Fab fragments, F(ab')2 molecules, and tandem discFv.

[0186] In a particular embodiment, the anti-CD45 antibody or its antigen-binding fragment is conjugated It has a certain dissociation rate, which is particularly advantageous when used as part of a set. In a particular embodiment, the anti-CD45 antibody is used in biolayer interferometry (BLI). When measured by this method, human CD45 and / or rhesus monkey CD45 are 1 × 1 0 -2 ~1 × 10 -3 , 1 x 10 -3 ~1 × 10 -4 , 1 x 10 -5 ~1 × 10 -6 , 1 ×10 -6 ~1 × 10 -7 or 1 × 10 -7 ~1 × 10 -8 Off rate constant (Kof f) has. In some embodiments, the antibody or its antigen-binding fragment is a biore When measured by the Ear Interference (BLI) assay, CD45 (e.g., human CD4) 5 and / or rhesus macaques (CD45), approximately 100 nM or less, approximately 90 nM or less, approximately 80 n M or less, about 70nM or less, about 60nM or less, about 50nM or less, about 40nM or less, about 30n M or less, about 20nM or less, about 10nM or less, about 8nM or less, about 6nM or less, about 4nM or less , K below approximately 2nM, below approximately 1nM D They are joined together.

[0187] In one embodiment, the present invention relates to human CD45 (SEQ ID NO: 175) and cynomolgus monkey C Binds to D45 (SEQ ID NO: 194) and / or rhesus macaque CD45 (SEQ ID NO: 195). The present invention provides an antibody or its antigen-binding portion. In some embodiments, the antibody or The antigen-binding region, when determined by biolayer interference (BLI), is human CD. 45. Approximately 100 nM or less, for example, approximately 100 nM or less, approximately 90 nM or less, approximately 80 nM or less. Lower, about 70nM or less, about 60nM or less, about 50nM or less, about 40nM or less, about 30nM or less Below, K is approximately 20 nM or less, approximately 10 nM or less, approximately 10 nM or less, or approximately 0.1 nM or less. D It can be bound by. In some embodiments, the antibody or its antigen-binding portion When determined by biolayer interferometry (BLI), the cynomolgus monkey CD45 is approximately Less than 100 nM, for example, less than approximately 100 nM, less than approximately 90 nM, less than approximately 80 nM, less than approximately 70 nM or less, about 60nM or less, about 50nM or less, about 40nM or less, about 30nM or less, about 20 K is less than nM, less than approximately 10 nM, less than approximately 10 nM, or less than approximately 0.1 nM. D Join together It is possible. In some embodiments, the antibody or its antigen-binding portion is biore When determined by EUR Interferometry (BLI), the rhesus monkey CD45 has a pulse of approximately 100 nM or less. For example, below, approximately 100 nM or less, approximately 90 nM or less, approximately 80 nM or less, approximately 70 nM or less, approximately 60nM or less, about 50nM or less, about 40nM or less, about 30nM or less, about 20nM or less, about K is less than 10 nM, approximately less than 10 nM, or approximately less than 0.1 nM. D They can be joined together. In some embodiments, the antibody is a fully human antibody or its antigen-binding portion. In other embodiments, the antibody is a humanized antibody or its antigen-binding portion. In one embodiment, the antibody is a chimeric antibody or its antigen-binding portion. In this embodiment, the antibody is a deimmunized antibody or its antigen-binding portion.

[0188] In one embodiment, an anti-CD45 antibody comprising one or more radiolabeled amino acids is provided. Radiolabeled anti-CD45 antibodies can be used for both diagnostic and therapeutic purposes. Conjugation to recognition molecules is another possible feature. Non-limiting polypeptide labeling. Typical examples include 3H, 14C, 15N, 35S, 90Y, 99Tc, and 125I, 1 Examples include, but are not limited to, 31I and 186Re. Radiolabeled amino acids and Methods for preparing related peptide derivatives are known in the art. (For example, Junghans et al., Cancer Chemotherapy) and Biotherapy 655-686(2d edition, Chafn er and Longo, eds., Lippincott Raven (1996) ) and U.S. Patent No. 4,681,581, U.S. Patent No. 4,735,210, U.S. Patent No. License No. 5,101,827, U.S. Patent No. 5,102,990 (USRE 35,500) See U.S. Patent Nos. 5,648,471 and U.S. Patent Nos. 5,697,902. For example, radioactive isotopes may be conjugated by the chloramine T method.

[0189] The anti-CD45 antibodies, their conjugates, or other related materials described herein are also As is known in the art, substances that increase half-life, substances that increase ADCC Modifications and / or mutations that alter the properties of antibodies and / or fragments, such as those that reduce or decrease them. It may include.

[0190] In one embodiment, the anti-CD45 antibody or its conjugated fragment includes a modified Fc region, The modified Fc region has affinity for binding to Fc gamma R (FcγR) of the molecule or The Fc region contains at least one amino acid modification compared to the wild type, resulting in altered binding affinity. A specific amino acid position within the Fc region is in direct contact with FcγR, as can be seen from crystallographic evidence. This has been shown through research. Specifically, amino acids 234-239 (hinge region), amino Acids 265-269 (B / C loop), amino acids 297-299 (C' / E loop), and This is an amino acid 327-332 (F / G) loop (Sondermann et al.). (See Nature, 2000, 406:267-273). In some embodiments, The antibodies described herein are directly linked to FcγR based on structural and crystallographic analysis. The variant Fc region may include a modification of at least one residue in contact with it. In this state, the Fc region of the anti-CD45 antibody or its antigen-binding fragment is Kabat et al. al.,Sequences of Proteins of Immunologic al Interest,5th Ed.Public Health Service EU in NH1, MD (1991) (expressly incorporated herein by reference) According to the index, amino acid substitution is included at amino acid 265. "E in Kabat The "U index" refers to the numbering of human IgG1 EU antibodies. In one embodiment, The Fc region contains the D265A mutation. In one embodiment, the Fc region contains the D265 Includes a mutation in C. In some embodiments, the Fc region of the antibody (or fragment thereof) is According to the EU index in Kabat, amino acid 234 contains an amino acid substitution. In one embodiment, the Fc region includes the L234A mutation. In some embodiments, The Fc region of the anti-CD45 antibody or its fragment is classified as the EU index in Kabat. Accordingly, amino acid 235 contains an amino acid substitution. In one embodiment, the Fc region is L Includes the 235A mutation.

[0191] In yet another embodiment, the Fc region is a variant of L234A and L235A (as specified herein). This includes "L234A, L235A" or "LALA" in the context of another real In the application morphology, the Fc region includes mutations L234A and L235A, where the Fc region The region does not contain the P329G mutation. In a further embodiment, the Fc region is D265C, Mutations of L234A and L235A (referred to herein as "D265C.L234A.L2 It includes (also called "35A"). In another embodiment, the Fc region is D265C, L2 This variant includes mutations 34A and L235A, where the Fc region does not contain the P329G mutation. In further embodiments, the Fc region is D265C, L234A, L235A, and H435A mutation (referred to as "D265C.L234A.L235A.H435" in this specification) It includes (also called "A"). In another embodiment, the Fc region is D265C, L234 This includes mutations A, L235A, and H435A, where the Fc region contains the P329G mutation. Not included. In a further embodiment, the Fc region is a mutation of D265C and H435A (this This includes (also referred to as "D265C.H435A" in the specification). Further embodiments include In this case, the Fc region has mutations in D265A, S239C, L234A, and L235A (this (Also referred to as "D265A.S239C.L234A.L235A" in the specification) Includes. In yet another embodiment, the Fc region is D265A, S239C, L234A, and includes the L235A mutation, where the Fc region does not include the P329G mutation. Another actual In the application form, the Fc region is a variant of D265C, N297G, and H435A (see this specification) Includes (also called "D265C.N297G.H435A" in the book). Another implementation form In this state, the Fc region has mutations in D265C, N297Q, and H435A (as specified herein). In another embodiment, In this region, the Fc area consists of E233P, L234V, L235A and delG236(236 A mutation (deletion of) (referred to herein as "E233P.L234V.L235A.delG2 Includes 36 (also known as "EPLVLAdelG"). In another embodiment, The Fc region consists of E233P, L234V, L235A, and delG236 (deletion of 236). This includes the mutation, where the Fc region does not contain the P329G mutation. In another embodiment, The Fc region consists of E233P, L234V, L235A, and delG236 (deletion of 236). and the H435A mutation (referred to as "E233P.L234V.L235A.de" in this specification). (Also known as "lG236.H435A" or "EPLVLAdelG.H435A") This includes. In another embodiment, the Fc region is E233P, L234V, L235A, d It includes mutations in elG236 (deletion of 236) and H435A, where the Fc region is P3 It does not contain the 29G mutation. In another embodiment, the Fc region is L234A, L235A, This includes the S239C and D265A mutations. In another embodiment, the Fc region is L234 This includes mutations A, L235A, S239C, and D265A, where the Fc region is P32 It does not contain the 9G mutation. In another embodiment, the Fc region is H435A, L234A, L This includes mutations 235A and D265C. In another embodiment, the Fc region is H435 This includes mutations A, L234A, L235A, and D265C, where the Fc region is P3 Does not contain the 29G mutation.

[0192] In some embodiments, the anti-CD45 antibody or its antigen-binding fragment is in vi Modifications that reduce the effector function of an antibody in a tro effector function assay. It has an Fc region, and its binding to the Fc receptor (Fc R) is the same antibody containing the unmodified Fc region. Binding to cR is reduced compared to other methods. In some embodiments, the antibody or its antigen The binding fragment is used in in vitro effector function assays to enhance the effector function of antibodies. It has a modified Fc region that reduces its ability, and its binding to the Fc gamma receptor (FcγR) is not modified. Compared to the binding of the same antibody containing the decorative Fc region to FcγR, it is reduced. Several implementations In this state, FcγR is FcγR1. In some embodiments, FcγR is , is FcγR2A. In some embodiments, FcγR is FcγR2B. In other embodiments, FcγR is FcγR2C. Therefore, FcγR is FcγR3A. In some embodiments, FcγR is Fc It is γR3B. In other embodiments, the reduction in binding is due to the same antibody containing an unmodified Fc region. Compared to binding to FcγR, antibody binding to FcγR is reduced by at least 70%, less Both decreased by 80%, at least by 90%, at least by 95%, and at least by 98%. A decrease of %, a decrease of at least 99%, or a decrease of 100%. In other embodiments Therefore, the reduction in binding is compared to the binding of the same antibody containing an unmodified Fc region to FcγR. A reduction of at least 70% to 100% in antibody binding to R, and at least 80% to 100% in antibody binding. Small, at least a 90% to 100% decrease, at least a 95% to 100% decrease, or small At the very least, it represents a 98% to 100% decrease.

[0193] In some embodiments, the anti-CD45 antibody or its antigen-binding fragment is in vi Modifications that reduce cytokine release by antibodies in trocytokine release assays The antibody has an Fc region, and cytokine release is observed in the same antibody that includes an unmodified Fc region. Compared to, it is reduced by at least 50%. In some embodiments, cytokine release The decrease is compared to cytokine release from the same antibody containing an unmodified Fc region. A decrease of at least 70%, a decrease of at least 80%, a decrease of at least 90%, less Both a 95% decrease, at least a 98% decrease, at least a 99% decrease, or 100% This is a decrease in the unmodified Fc region. In some embodiments, the decrease in cytokine release is due to a decrease in the unmodified Fc region. Compared to cytokine release from the same antibody including the region, cytokine release is at least 70%~ A 100% decrease, a decrease of at least 80% to 100%, a decrease of at least 90% to 100% The reduction is small, at least 95% to 100%. In a particular embodiment, the site Infusion is released by immune cells.

[0194] In some embodiments, the anti-CD45 antibody or its antigen-binding fragment is in vi In the tro mast cell degranulation assay, the antibody reduces mast cell degranulation. Mast cell degranulation has a modified Fc region, and mast cell degranulation is caused by the same antibody that contains an unmodified Fc region. The reduction is at least 50% compared to the granules. In some embodiments, mast cell degranulation occurs. The decrease is compared to mast cell degranulation with the same antibody that includes the unmodified Fc region. A decrease of at least 70% in grains, a decrease of at least 80%, a decrease of at least 90%, a small amount A decrease of at least 95%, at least 98%, at least 99%, or 100 This represents a % decrease. In some embodiments, the decrease in mast cell degranulation is due to unmodified Fc Compared to mast cell degranulation with the same antibody that includes the region, at least 70% of mast cell degranulation ~100% decrease, at least 80%~100% decrease, at least 90%~100% A decrease, or at least a 95% to 100% decrease.

[0195] In some embodiments, the anti-CD45 antibody or its antigen-binding fragment is in vi In antibody-dependent cell phagocytosis assays, the antibody-dependent cell phagocytosis (ADCP) of the antibody It has a modified Fc region that reduces or suppresses, and ADCP is the same as the unmodified Fc region. Compared to ADCP of the antibody, it is reduced by at least 50%. In some embodiments, AD The decrease in CP is compared to cytokine release from the same antibody containing an unmodified Fc region. A reduction of at least 70% in emissions, a reduction of at least 80%, a reduction of at least 90%, A decrease of at least 95%, a decrease of at least 98%, a decrease of at least 99%, or 1 This represents a 00% decrease.

[0196] In some embodiments, the anti-CD45 antibody or its antigen-binding fragment is modified as follows: Includes Fc regions containing one or a combination of modifications: D265A, D265C, D 265C / H435A, D265C / LALA, D265C / LALA / H435A, D 265A / S239C / L234A / L235A / H435A, D265A / S239C / L234A / L235A, D265C / N297G, D265C / N297G / H43 5A, D265C(EPLVLAdelG*), D265C(EPLVLAdelG) / H435A, D265C / N297Q / H435A, D265C / N297Q, EPLV LAdelG / H435A, EPLVLAdelG / D265C, EPLVLAdelG / D265A, N297A, N297G, or N297Q.

[0197] The binding or affinity between the modified Fc region and the Fc gamma receptor is, in the art, Various known techniques, for example, but not limited to, equilibrium methods (e.g., enzyme binding) Immunosorbent assay (ELISA); KinExA,Rathanaswami et al.A nalytical Biochemistry,Vol.373:52-60,200 8; or radioimmunoassay (RIA), or surface plasmon resonance assay. or other mechanisms of kinetics-based assays (e.g., BIACORE (registered trademark) Standard analysis or Octet® analysis (forteBIO), and indirect analysis Binding assays, competitive binding assays, fluorescence resonance energy transfer (FRET), gel electrophoresis and The determination can be made using other methods such as chromatography (e.g., gel filtration). Yes, these methods and other methods utilize labeling on one or more of the components being tested. It may include, and / or include color-developing labels, fluorescent labels, luminescent labels, or isotopic labels. A variety of detection methods, not limited to these, can be employed. Binding affinity and kinetics For a detailed explanation of the cus, see Paul, WE, which focuses on the interaction between antibodies and immunogens. .,ed.,Fundamental Immunology,4th Ed.,Lip This can be found in pincott-Raven, Philadelphia (1999). One example of a competitive binding assay involves binding the antibody of interest and the labeled antigen in the presence of gradually increasing amounts of unlabeled antigen. Radioimmunotherapy, which involves incubation and detection of antibodies bound to a labeled antigen. This is an assay. The affinity and binding off-rate of the target antibody against a specific antigen are determined by scanning. This can be determined from the data by Chard plot analysis. Competition with secondary antibodies is Alternatively, it can be determined using a radioimmunoassay. In this case, the antigen is labeled. The target antibody conjugated with the compound, in the presence of gradually increasing amounts of unlabeled secondary antibody. It is incubated.

[0198] In one embodiment, the Fc modification described herein (e.g., D265C, L234) Anti-CD45 antibodies having A, L235A, and / or H435A) or their antigen binding The fragment, compared to the binding of the same antibody containing the unmodified Fc region to the Fc gamma receptor, is different from that of the Fc cancer A reduction of at least 70%, at least 80%, and at least 90% in binding to the receptor. % decrease, at least 95% decrease, at least 98% decrease, at least 99% decrease , or having a 100% reduction (e.g., evaluated by biolayer interferometry (BLI)) (If valued).

[0199] While I don't want to be bound by any theory, I do have a preference for the Fc region and the Fc gamma receptor. The coupling interaction with this enables various effector functions and downstream signaling events, for example While not limited to these, antibody-dependent cell-mediated cytotoxicity (ADCC) and complement-dependent cytotoxicity are also included. It is considered essential for cytotoxicity (CDC). Therefore, in certain manner In this, the modified Fc region is included (e.g., L234A, L235A, and / or D26 Antibodies (including those containing the 5C mutation) have substantially reduced or lost effector function. The function can be used, for example, to achieve the objective, using various methods known in the art. Measuring cellular responses to antibodies (e.g., mast cell degranulation or cytokine release) By doing so, assays can be performed. For example, standard assays in the relevant field Using this method, the ability to induce mast cell degranulation in vitro or cytocyto Regarding the ability to induce nucleotide release (e.g., by human peripheral blood mononuclear cells), Fc-modified antibodies It can be assayed.

[0200] The antibodies disclosed herein are, for example, (Dall'Acqua et al. (2006) JB iol Chem 281:23514-24), (Zalevsky et al. 2010) Nat Biotechnol 28:157-9), (Hinton et al. al. (2004) J Biol Chem 279:6213-6), (Hinto n et al. (2006) J Immunol 176:346-56), (Shi elds et al. (2001) J Biol Chem 276:6591-60 4), (Petkova et al. (2006) Int Immunol 18:1 759-69), (Datta-Mannan et al. (2007) Drug M etab Dispos 35:86-94), (Vaccaro et al.(20 05) Nat Biotechnol 23:1283-8), (Yeung et a l. (2010) Cancer Res 70:3269-77) and (Kim et al.) As described in al. (1999) Eur J Immunol 29:2819-25) By introducing additional Fc mutations, such as those present, the antibody half-life can be further regulated. Further manipulation is possible, at positions 250, 252, 253, 254, 256, 257, and 307. This includes 376, 380, 428, 434, and 435. They can be used individually or in combination. Exemplary mutations that can be identified include T250Q, M252Y, 1253A, S254T, T 256E, P2571, T307A, D376V, E380A, M428L, H433K , N434S, N434A, N434H, N434F, H435A and H435R mutations be.

[0201] Therefore, in one embodiment, the Fc region includes a mutation that results in a reduction in half-life. For example, compared to antibodies with an unmodified Fc region. Antibodies with a short half-life have a short half-life. In certain cases where it is expected to function as a life-saving treatment, for example, after antibody administration, HS C may be advantageous in the conditioning steps described herein in which C is administered. In theory, the antibodies are effectively cleared before HSC delivery, and HSCs are also generally... They express target antigens (e.g., CD45), but unlike endogenous stem cells, they do not express antibodies (e.g., It is not a target of anti-CD45 antibodies. In one embodiment, the Fc region is mutated at position 435. Includes (EU index according to Kabat). In one embodiment, the mutation is H435 This is a mutation of A.

[0202] In one embodiment, the anti-CD45 antibody or its antigen-binding fragment described herein is , about 24 hours or less, about 23 hours or less, about 22 hours or less, about 21 hours or less, about 20 hours or less , about 19 hours or less, about 18 hours or less, about 17 hours or less, about 16 hours or less, about 15 hours or less , half-live of approximately 14 hours or less, approximately 13 hours or less, approximately 12 hours, or approximately 11 hours or less (for example) (In humans, it has.)

[0203] In one embodiment, the anti-CD45 antibody or its antigen-binding fragment described herein is Approximately 1-5 hours, approximately 5-10 hours, approximately 10-15 hours, approximately 15-20 hours, or approximately 20 hours It has a half-life of ~25 hours (for example, in humans). In one embodiment, anti-CD4 5. The half-lives of antibodies or their antigen-binding fragments are approximately 5-7 hours; approximately 5-9 hours; and approximately 5-11 hours. Interval; approximately 5-13 hours; approximately 5-15 hours; approximately 5-20 hours; approximately 5-24 hours; approximately 7-24 hours Interval; approximately 9-24 hours; approximately 11-24 hours; approximately 12-22 hours; approximately 10-20 hours; approximately 8- 18 hours; or approximately 14-24 hours.

[0204] In some embodiments, the Fc region of the anti-CD45 antibody or its antigen-binding fragment is an antibody It includes two or more mutations that confer a reduction in half-life and a reduction in effector function. In one embodiment, the Fc region is in direct contact with mutations that result in a reduction in half-life and with FcγR. A mutation of at least one residue (for example, based on structural and crystallographic analysis) can be detected. (including) In one embodiment, the Fc region is a mutation of H435A, a mutation of L234A This includes variants and the L235A variant. In one embodiment, the Fc region is a variant of H435A. and includes the D265C mutation. In one embodiment, the Fc region is the H435A mutation, L This includes the 234A mutation, the L235A mutation, and the D265C mutation.

[0205] In some embodiments, the anti-CD45 antibody or its antigen-binding fragment is an antibody or Cytotoxicity (e.g., Amatoki) is transmitted via cysteine ​​residues in the Fc domain of the antigen-binding fragment. Conjugated by (Shin).

[0206] In some embodiments of these models, the cysteine ​​residue is an anti-CD45 antibody or It is naturally present in the Fc domain of its antigen-binding fragment. For example, the Fc domain is found in human It could be an IgG Fc domain such as the IgG1 Fc domain, and the cysteine ​​residue is Cy The group can be selected from s261, Csy321, Cys367, and Cys425. .

[0207] In some embodiments, the cysteine ​​residue is used to bind to the anti-CD45 antibody or its antigen. It is introduced via mutations in the Fc domain of the fragment. For example, a cysteine ​​residue is introduced via Cys1 18, Cys239, and Cys265 may be selected from the group. In one embodiment The Fc region of the anti-CD45 antibody or its fragment is classified as the EU index in Kabat. Accordingly, amino acid 265 contains an amino acid substitution. In one embodiment, the Fc region is D Includes the 265C mutation. In one embodiment, the Fc region is D265C and H435A Includes mutations. In one embodiment, the Fc region is D265C, L234A, and L235 Includes mutation A. In one embodiment, the Fc region is D265C, L234A, L235 Includes mutations A and H435A. In one embodiment, an anti-CD45 antibody or its antigen. The Fc region of the binding fragment corresponds to amino acid 239, according to the EU index in Kabat. This includes amino acid substitutions. In one embodiment, the Fc region includes the S239C mutation. In this embodiment, the Fc region is a mutation of L234A, a mutation of L235A, and a mutation of S239C. and includes the D265A mutation. In another embodiment, the Fc region is S239C and H4 Includes mutation 35A. In another embodiment, the Fc region includes mutation L234A, L235 This includes mutations in A and S239C. In yet another embodiment, the Fc region is H4 This includes mutations 35A, L234A, L235A, and S239C. In another embodiment, the Fc region is a mutation of H435A, a mutation of L234A, L235 This includes mutations in A, S239C, and D265A.

[0208] In particular, the position of the Fc amino acid is in the EU numbering index unless otherwise specified. It refers to S.

[0209] The variant Fc domains described herein are based on the amino acid modifications that constitute them. This is how it is defined. For all amino acid substitutions considered herein with respect to the Fc region, The numbering always follows the EU index. Therefore, for example, D265C is Based on the parent Fc domain, aspartic acid (D) at EU position 265 is cysteine ​​(C This is an Fc variant that has been replaced with ). Similarly, for example, D265C / L234A / L2 35A is based on the parent Fc domain, with EU positions 265 (D to C) and 234 (L to Define a variant Fc that has substitutions in A) and 235 (from L to A). The riant also specifies according to the final amino acid composition at the mutated EU amino acid position. It is also possible to call the L234A / L235A variant "LALA". This is possible. Note that the order in which substitutions are provided is arbitrary. In particular, FcAmi Unless otherwise specified, the position of no acids refers to the EU numbering index.

[0210] In some embodiments, the anti-CD45 antibody or its antigen-binding fragment as described herein The Fc region includes one or a combination of the following modifications: D265A, D2 65C, D265C / H435A, D265C / LALA, D265C / LALA / H4 35A, D265C / N297G, D265C / N297G / H435A, D265C( IgG2*), D265C(IgG2) / H435A, D265C / N297Q / H43 5A, D265C / N297Q, EPLVLAdelG / H435A, N297A, N2 97G, or N297Q.

[0211] The antibodies and their conjugated fragments disclosed herein are described in more detail below, It can be used for conjugates.

[0212] Antibodies are recombinant and compositions as described, for example, in U.S. Patent No. 4,816,567. It can be prepared using the anti-CD45 antibody described herein. An isolated nucleic acid encoding is provided. Such a nucleic acid contains amino acids including the VL of the antibody. amino acid sequences containing sequences and / or VH (e.g., light and / or heavy chains of antibodies) It may be possible. In a further embodiment, one or more vectors containing such nucleic acids (e.g. In a further embodiment, a host comprising such nucleic acid is provided. Cells are provided. In one such embodiment, the host cells contain (1) a very large amount of the antibody. A vector containing a nucleic acid encoding an amino acid sequence and an amino acid sequence including the VH of an antibody, (2) A first vector and an antibody comprising a nucleic acid encoding an amino acid sequence including the VL of the antibody. The second vector includes a nucleic acid that encodes an amino acid sequence containing the body's VH (for example, (Transformed by this). In one embodiment, the host cell is a eukaryote, for example, Chinese hamster ovary (CHO) cells or lymphoid cells (e.g., Y0, NS0) (Sp20 cells). In one embodiment, a method for producing an anti-CLL-1 antibody is provided. The method involves a host cell containing nucleic acid encoding the antibody provided above, which is then used to express the antibody. By culturing under suitable conditions and, optionally, by extracting from host cells (or host cell culture medium) This includes recovering antibodies.

[0213] For the recombinant production of anti-CD45 antibodies, antibodies, such as nucleic acids encoding the above antibodies, are required. One or more were isolated and further cloned and / or expressed in host cells. Such nucleic acids are inserted into the vector. Oligonucleotide pros capable of specifically binding to genes encoding chains and light chains It can be easily isolated and sequenced (using a vector).

[0214] Suitable host cells for cloning or expressing antibody-encoding vectors are specified herein. This includes prokaryotic or eukaryotic cells as described. For example, antibodies, in particular, glycosylated And if the Fc effector function is not required, it can be produced in bacteria. Antibacterial properties in bacteria For the expression of somatic fragments and polypeptides, see, for example, U.S. Patent No. 5,648,237. See also issues Nos. 5,789,199 and 5,840,523. (E.co Charlton, Methods in li. describes the expression of antibody fragments. Molecular Biology,Vol.248(BKCLo,ed.,H umana Press, Totowa, NJ, 2003), pp.245-254 See also.) After expression, the antibody can be isolated from the bacterial cell paste in a soluble fraction, and further It can be purified.

[0215] Vertebrate cells can also be used as hosts. For example, they can be modified to grow in suspension. Mammalian cell lines that have been modified may be useful. Another example of a useful mammalian host cell line is SV4 CV1 strain of monkey kidney transformed by 0 (COS-7); human embryonic kidney strain (293 strain) Cells or, for example, Graham et al., J. Gen Virol. 36:59 (1 977) 293 cells; baby hamster kidney cells (BHK); mouse Sertoli cells Cells (e.g., Mather, Biol. Reprod. 23:243-251 (198) 0) TM4 cells as described above; monkey kidney cells (CV1); African green monkey kidney cells (V ERO-76); Human cervical cancer cells (HELA); Canine kidney cells (MDCK); Buffer Rat hepatocytes (BRL 3A); human lung cells (W138); human hepatocytes (Hep G 2); Mouse mammary gland tumors (MMT 060562); e.g., Mather et al. , as described in Annals NYAcad.Sci.383:44-68(1982) These include TRI cells; MRC 5 cells; and FS4 cells. Other useful mammalian host cell lines include , DHFR-CHO cells (Urlaub et al., Proc. Natl. Acad Chinese hamster ovaries (including .Sci.USA 77:4216(1980)) This includes CHO cells, as well as myeloma cell lines such as Y0, NS0, and Sp2 / 0. For a review of specific mammalian host cell lines suitable for body production, see, for example, Yazaki a. nd Wu,Methods in Molecular Biology,Vol.2 48 (BKCLo, ed., Humana Press, Totowa, NJ See pp. 255-268 (2003).

[0216] In one embodiment, an anti-CD45 antibody or its antigen-binding fragment is disclosed herein. For the sequence numbers (Table 5), at least 95%, 96%, 97%, or 99% are identical. It includes a variable region having a specific amino acid sequence. Alternatively, it may contain an anti-CD45 antibody or its antigen-binding. The fragments are at least 95%, 96%, of the sequence numbers disclosed herein (Table 5). The variable regions described herein having amino acid sequences that are 97% or 99% identical The framework area includes a CD-R containing the sequence numbers disclosed herein.

[0217] In one embodiment, an anti-CD45 antibody or its antigen-binding fragment is disclosed herein. It includes a heavy chain variable region and a heavy chain constant region having the amino acid sequence. In another embodiment, The anti-CD45 antibody or its antigen-binding fragment has the amino acid sequence disclosed herein. It includes a light chain variable region and a light chain constant region. In yet another embodiment, an anti-CD45 antibody and The antigen-binding fragment has a heavy chain variable region having the amino acid sequence disclosed herein, and a light chain It includes a variable region, a heavy chain steady region, and a light chain steady region.

[0218] Examples of anti-CD45 antibodies are described further herein.

[0219] Anti-CD45 antibody Human CD45 (mRNA NCBI reference sequence: NM_080921.3, Protein Antibodies and antigens capable of binding to n NCBI reference sequence: NP_563578.2) The binding fragments include those capable of binding to the isoform CD45RO, for example, To promote the engraftment of hematopoietic stem cell grafts in patients requiring hematopoietic stem cell transplantation therapy, It can be used in connection with the compositions and methods disclosed herein. In one embodiment... Furthermore, the compositions and methods disclosed herein are as described in the amino acid sequence of SEQ ID NO: 1. The present invention comprises an anti-CD45 antibody or ADC that binds to human CD45RO. Antibodies that bind to various isoforms of CD45 are also disclosed herein in terms of methods and combinations. Intended for use in finished products. Multiple isoforms of CD45 are used in the primary transfer. Arising from alternative splicing of exon 4. Splicing of exons 4, 5, and 6. Furthermore, potentially, splicing of 7 could result in multiple variations of CD45. Selective exon expression was observed in the CD45 isoforms listed in Table 1 below. ru. [Table 1]

[0220] Alternative splicing can result in individual exons or combinations of exons. Various isoforms of the CD45 protein (e.g., CD45RA, CD45RAB, CD45RABC is expressed in CD45. In contrast, CD45RO expresses exons 4-6. Defects result from combinations of exons 1-3 and 7-34. Exon 7 is also a tamper. It can be removed from the matrix, and exons 1-3 and 8-34 are spliced ​​together. There is evidence. This protein has been named E3-8 and has been detected at the mRNA level. They exist, but have not yet been identified by flow cytometry.

[0221] CD45RO is currently the only CD45 isoform expressed on hematopoietic stem cells. It is known that CD45RA and CD45RABC have not been detected or have not been produced. It has been excluded from the hematopoietic stem cell phenotype. Studies conducted in mice have shown that CD45RB is embryonic There is evidence that it is expressed on hematopoietic stem cells in infants but not in adult bone marrow hematopoietic stem cells. Yes, it exists. What is noteworthy is that CD45RC has a high rate of exon 6 polymorphism in the Asian population. (A polymorphism in exon 6 of CD45RC is found in approximately 25% of the Japanese population.) ). This polymorphism leads to high expression of CD45RO, CD45RA, CD45RB, CD45 The expression level of RC decreases. Furthermore, CD45RA variants (CD45RAB and CD45) RAC (and other autoimmune diseases) exhibit polymorphisms in exon 4, which are associated with autoimmune diseases.

[0222] CD45RO is present on hematopoietic stem cells, and other immune cells (T and B lymphocyte subsets) Furthermore, because its expression is relatively limited on various cells (such as bone marrow cells), CD45RO is This is a particularly suitable target for conditioning therapy for patients requiring hematopoietic stem cell transplantation. CD45RO is the only one that lacks expression of exons 4, 5, and 6, so it is used as an immunosuppressant. By using it, screening of pan-CD45 Ab and CD45RO-specific antibodies. This is possible.

[0223] Anti-C that can be used in connection with the patient conditioning methods described herein. The D45 antibody contains the anti-CD45 antibody and its antigen-binding portion. This is well known in the field and can be easily constructed based on the antigen-binding region of the antibody. It can be constructed. In exemplary embodiments, the conditions described herein Anti-CD45 antibodies used in connection with the testing method are monoclonal antibodies or their antigenic antibodies. Synthetic fragments, polyclonal antibodies or their antigen-binding fragments, humanized antibodies or their antigen-binding fragments Synthetic fragments, fully human antibodies or their antigen-binding fragments, chimeric antibodies or their antigen-binding fragments , bispecific antibody or its antigen-binding fragment, bivariable immunoglobulin domain, single-chain Fv molecule (scFv), diabody, triabody, nanobody, antibody-like protein Cafold, Fv fragment, Fab fragment, F(ab')2 molecule, or tandem scFv It may be used, in whole or in part, in the ADC or method described herein. Exemplary anti-CD45 antibodies that may be used are provided below.

[0224] In some embodiments, the anti-CD45 antibody is antibody A(Ab) disclosed herein. A), antibody B (AbB), antibody C (AbC), antibody D (AbD), antibody E (AbE), These are antibodies F (AbF). These antibodies are found in human CD45 and rhesus monkey CD45. They cross-react. Furthermore, these antibodies are extracellular antibodies against various isoforms of human CD45. It can be combined with the main. Therefore, in a particular embodiment, the specified The antibody is a general-specific anti-CD45 antibody (i.e., all six human CD45 isoforms). It is an antibody that binds to [substance]. Furthermore, AbA, AbB, and AbC (and others) are disclosed herein. These antibodies (or antibodies with binding regions or specificity) are found to bind to cynomolgus monkey CD45. They can also be combined.

[0225] Various binding sites of anti-CD45 antibodies AbA, AbB, AbC, AbD, AbE, and AbF The amino acid sequence of the region is shown in Table 5.

[0226] For example, based on antibodies AbA, AbB, or AbC, including the CDRs listed in Table 5. Furthermore, humanized and chimeric anti-CD45 antibodies are included in the present invention.

[0227] In one embodiment, the present disclosure relates to a binding region corresponding to that of AbA, for example, CDR, The present invention provides an anti-CD45 antibody or its antigen-binding fragment containing a variable region. The region (VH) amino acid sequence is described in SEQ ID NO: 13 (see Table 5). AbA VH The CDR domain amino acid sequences are SEQ ID NO: 14 (VH CDR1), SEQ ID NO: 15 (VH As described in CDR2) and Sequence ID 16 (VH CDR3). AbA light chain variable region The very long (VL) amino acid sequence is described in SEQ ID NO: 17 (see Table 5). VL C of AbA The DR domain amino acid sequences are SEQ ID NO: 18 (VL CDR1), SEQ ID NO: 19 (VL As described in CDR2) and Sequence ID No. 20 (VL CDR3). Therefore, a certain In the embodiments herein, the anti-CD45 antibody or its antigen-binding fragment provided herein is The heavy chain variable region containing the amino acid sequence described in SEQ ID NO: 13, and the amino acid sequence described in SEQ ID NO: 17 It includes a light chain variable region containing an amino acid sequence. In one embodiment, the anti-CD45 antibody is CDR1, CDR2 and the amino acid sequences described in SEQ ID NOs: 14, 15, and 16 A heavy chain containing CDR3 and the amino acid sequence described in SEQ ID NOs: 18, 19, and 20. It includes a light chain variable region containing CDR1, CDR2, and CDR3.

[0228] By one of the antibodies AbA, AbB, AbC, AbD, AbE, or AbF Anti-human CD45 antibody or fragment thereof that binds to an epitope on human CD45 ( Or an antibody having a binding domain for AbA, AbB, AbC, AbD, AbE, or AbF The body) is also intended herein. Binding to human CD45 and / or cynomolgus monkey CD45. Regarding binding to 45 or rhesus monkey CD45, the antibodies AbA, AbB, AbC, and Ab D, AbE, or AbF (or AbA, AbB, AbC, AbD, AbE, or One of the antibodies (antibodies having an AbF binding region) competes with an anti-human CD45 antibody or Further antigen-binding fragments are also being considered. AbA~AbC are, for example, in International Publication No. WO2020 It is described in publication / 092654, which is incorporated herein by reference in its entirety. D~AbF are described, for example, in international application PCT / US2020 / 058373. The entire text is incorporated herein by reference.

[0229] In some embodiments, the anti-CD45 antibody or its antigen-binding fragment is composed of amino acids. In the region containing the column RNGPHERYHLEVEAGNT (sequence number 181), human CD45 It binds specifically. For example, in a particular embodiment, an anti-CD45 antibody, or The antigen-binding fragment corresponds to the CD45 isoflavone of sequence number 176 (NP_002829.3). amino acid residues 486R, 493Y, and 502T of a fragment of the foam, or other human CDs. 45 isoforms array [ka] The residues corresponding to the region containing the above specifically bind to human CD45. In some embodiments, In this context, the anti-CD45 antibody or its antigen-binding fragment is a human CD45 fibronectin It specifically binds to the main domain (for example, the fibronectin D4 domain).

[0230] In one embodiment, the isolated anti-CD45 antibody or its antigen-binding moiety is SEQ ID NO: Specific to the human CD45 epitope, including 176 residues 486R, 493Y, and 502T. It binds to the CD45 of cynomolgus macaques and / or rhesus macaques.

[0231] In one embodiment, the isolated anti-CD45 antibody or its antigen-binding moiety is an amino acid Human CD45 epidermal cells containing the sequence RNGPHERYHLEVEAGNT (sequence number 181) It specifically binds to topes and also binds to CD45 in cynomolgus monkeys and rhesus monkeys.

[0232] In one embodiment, the isolated anti-CD45 antibody or its antigen-binding moiety is an amino acid Array CRPPRDRNGPHERYHLEVEAGNTLVRNESHK(Sequence ID 18 0) specifically binds to the human CD45 epitope, and is found in cynomolgus monkeys and rhesus monkeys C It also connects to D45.

[0233] In one embodiment, the isolated anti-CD45 antibody or its antigen-binding moiety is SEQ ID NO: Specific to the human CD45 epitope, including 176 residues 486R, 493Y, and 502T. It binds to peptides containing RNGPHERYHLEVEAGNT (SEQ ID NO: 181). at least one additional amino acid, at least two additional amino acids, at least three an additional amino acid, at least four additional amino acids, or at least five additional amino acids It binds to the amino acid, where the additional amino acid residues are residues 486R and 49 of SEQ ID NO: 176. It binds not only to 3Y and 502T, but also to CD45 in cynomolgus macaques and rhesus macaques.

[0234] In one embodiment, the present invention provides a coupling region corresponding to that of AbB, for example, CDR, This provides an anti-CD45 antibody or its antigen-binding fragment containing a variable region. The heavy chain of AbB is variable. The region (VH) amino acid sequence is described in SEQ ID NO: 21 (see Table 5). AbB VH The CDR domain amino acid sequences are: SEQ ID NO: 22 (VH CDR1), SEQ ID NO: 23 (VH As described in CDR2) and Sequence ID 24 (VH CDR3). AbB light chain variable region. The very long (VL) amino acid sequence is described in SEQ ID NO: 25 (see Table 5). VL C of AbB The DR domain amino acid sequences are SEQ ID NO: 26 (VL CDR1), SEQ ID NO: 27 (VL As described in CDR2) and Sequence ID No. 28 (VL CDR3). Therefore, a certain In the embodiments herein, the anti-CD45 antibody or its antigen-binding fragment provided herein is The heavy chain variable region containing the amino acid sequence described in SEQ ID NO: 21, and the amino acid sequence described in SEQ ID NO: 25 It includes a light chain variable region containing an amino acid sequence. In one embodiment, the anti-CD45 antibody is CDR1, CDR2 and the amino acid sequences described in SEQ ID NOs. 22, 23, and 24 A heavy chain containing CDR3 and the amino acid sequence described in SEQ ID NOs. 26, 27, and 28. It includes a light chain variable region containing CDR1, CDR2, and CDR3.

[0235] In one embodiment, the present invention provides a coupling region corresponding to that of AbC, for example, CDR, This provides an anti-CD45 antibody or its antigen-binding fragment containing a variable region. The region (VH) amino acid sequence is described in SEQ ID NO: 29 (see Table 5). AbC VH The CDR domain amino acid sequences are: SEQ ID NO: 30 (VH CDR1), SEQ ID NO: 31 (VH As described in CDR2) and Sequence ID 32 (VH CDR3). AbC light chain variable region. The very long (VL) amino acid sequence is described in SEQ ID NO: 33 (see Table 5). The VL C of AbC The DR domain amino acid sequences are SEQ ID NO: 34 (VL CDR1), SEQ ID NO: 35 (VL As described in CDR2) and Sequence ID No. 36 (VL CDR3). Therefore, a certain In the embodiments herein, the anti-CD45 antibody or its antigen-binding fragment provided herein is The heavy chain variable region containing the amino acid sequence described in SEQ ID NO: 29, and the amino acid sequence described in SEQ ID NO: 33 It includes a light chain variable region containing an amino acid sequence. In one embodiment, the anti-CD45 antibody is CDR1, CDR2 and the amino acid sequences described in SEQ ID NOs. 30, 31, and 32 A heavy chain containing CDR3 and the amino acid sequence described in SEQ ID NOs: 34, 35, and 36. It includes a light chain variable region containing CDR1, CDR2, and CDR3.

[0236] In one embodiment, the present invention relates to a coupling region corresponding to that of AbD, for example, CDR, This provides an anti-CD45 antibody or its antigen-binding fragment containing a variable region. The region (VH) amino acid sequence is described in SEQ ID NO: 37 (see Table 5). AbD VH The CDR domain amino acid sequences are: SEQ ID NO: 38 (VH CDR1), SEQ ID NO: 39 (VH As described in CDR2) and Sequence ID No. 40 (VH CDR3). AbD light chain variable region The very long (VL) amino acid sequence is described in SEQ ID NO: 41 (see Table 5). VL C of AbD The DR domain amino acid sequences are SEQ ID NO: 42 (VL CDR1), SEQ ID NO: 43 (VL As described in CDR2) and Sequence ID 44 (VL CDR3). Therefore, a certain In the embodiments herein, the anti-CD45 antibody or its antigen-binding fragment provided herein is The heavy chain variable region containing the amino acid sequence described in SEQ ID NO: 37, and the amino acid sequence described in SEQ ID NO: 41 It includes a light chain variable region containing an amino acid sequence. In one embodiment, the anti-CD45 antibody is CDR1, CDR2 and the amino acid sequences described in SEQ ID NOs. 38, 39, and 40 A heavy chain containing CDR3 and the amino acid sequence described in SEQ ID NOs: 42, 43, and 44. It includes a light chain variable region containing CDR1, CDR2, and CDR3.

[0237] In one embodiment, the present invention relates to a coupling region corresponding to that of AbE, for example, CDR, The present invention provides an anti-CD45 antibody or its antigen-binding fragment containing a variable region. The region (VH) amino acid sequence is described in SEQ ID NO: 47 (see Table 5). AbE VH The CDR domain amino acid sequences are: SEQ ID NO: 48 (VH CDR1), SEQ ID NO: 49 (VH As described in CDR2) and Sequence ID 50 (VH CDR3). AbE light chain variable region. The very long (VL) amino acid sequence is described in SEQ ID NO: 51 (see Table 5). The VL C of AbE. The DR domain amino acid sequences are SEQ ID NO: 52 (VL CDR1), SEQ ID NO: 53 (VL As described in CDR2) and Sequence ID No. 54 (VL CDR3). Therefore, a certain In the embodiments herein, the anti-CD45 antibody or its antigen-binding fragment provided herein is The heavy chain variable region containing the amino acid sequence described in SEQ ID NO: 47, and the amino acid sequence described in SEQ ID NO: 51 It includes a light chain variable region containing an amino acid sequence. In one embodiment, the anti-CD45 antibody is CDR1, CDR2 and the amino acid sequences described in SEQ ID NOs. 48, 49, and 50 A heavy chain containing CDR3 and the amino acid sequence described in SEQ ID NOs. 52, 53, and 54. It includes a light chain variable region containing CDR1, CDR2, and CDR3.

[0238] In one embodiment, the present invention relates to a coupling region corresponding to that of AbF, for example, CDR, This provides an anti-CD45 antibody or its antigen-binding fragment containing a variable region. The heavy chain of AbF is variable. The amino acid sequence of the region (VH) is described in SEQ ID NO: 57 (see Table 5). AbF VH The CDR domain amino acid sequences are SEQ ID NO: 58 (VH CDR1), SEQ ID NO: 59 (VH As described in CDR2) and Sequence ID 60 (VH CDR3). AbF light chain variable region The very long (VL) amino acid sequence is described in SEQ ID NO: 61 (see Table 5). The VL C of AbF. The DR domain amino acid sequences are SEQ ID NO: 62 (VL CDR1), SEQ ID NO: 63 (VL As described in CDR2) and Sequence ID No. 64 (VL CDR3). Therefore, a certain In the embodiments herein, the anti-CD45 antibody or its antigen-binding fragment provided herein is The heavy chain variable region containing the amino acid sequence described in SEQ ID NO: 57, and the amino acid sequence described in SEQ ID NO: 61 It includes a light chain variable region containing an amino acid sequence. In one embodiment, the anti-CD45 antibody is CDR1, CDR2 and the amino acid sequences described in SEQ ID NOs. 58, 59, and 60 A heavy chain containing CDR3 and the amino acid sequence described in SEQ ID NOs. 62, 63, and 64. It includes a light chain variable region containing CDR1, CDR2, and CDR3.

[0239] In some embodiments, the anti-CD45 antibody is antibody 1(Ab) disclosed herein. 1), Antibody 2 (Ab2), Antibody 3 (Ab3), Antibody 4 (Ab4), Antibody 5 (Ab5), Antibody This is either antibody 6 (Ab6) or antibody 7 (Ab7). These antibodies are human CD45, red oak. These antibodies cross-react with staghorn monkey CD45 and cynomolgus monkey CD45. Furthermore, these antibodies are human In that it can bind to the extracellular domains of various isoforms of CD45, It is generalized. Ab1~Ab7 are, for example, in international application PCT / US2020 / 058 This is described in issue 373, which is incorporated herein by reference in its entirety.

[0240] The extracellular domain of human CD45 consists of a mucin-like domain and four fibronectin-like domains. Includes in (d1, d2, d3, and d4). Desires to be bound by any theory. However, antibodies Ab1, Ab2, Ab3, Ab4, Ab5, Ab6, and Ab7 are, It interacts with human CD45 residues located within the d3 and d4 fibronectin-like domains. It is thought that these antibodies, in particular, are human CD45 as described in Sequence ID No. 178. It may interact with fragments of and human CD45 as described in Sequence ID No. 180. The trial (described in International Application No. PCT / US2020 / 058373, and referred to herein) (Used in reference to) Antibodies for human CD45, cynomolgus monkey CD45, and rhesus monkey CD45 It can specifically interact with one or more CD45 amino acid residues that are conserved among 45. This suggests that these residues are 405T, 407K, 419Y, 425K, and 5 Includes 05R (numbering based on the hCD45 fragment described in sequence number 176) In addition, these antibodies are derived from human CD45 residues 481R and / or 509H (sequence). (Numbering based on the hCD45 fragment described in number 176) may interact with Therefore, in some embodiments, the anti-CD45 antibody is used on d3 and / or d4 An antibody that binds to human CD45 at an epitope located in the fibronectin-like domain or This is the antigen-binding portion. In some embodiments, anti-CD45 antibodies are CD45 cleavage The hi located within fragment 2 (sequence number 178) and / or CD45 fragment 4 (sequence number 180) The CD45 epitope is an antibody or its antigen-binding portion that binds to CD45. In several embodiments, the anti-CD45 antibody is CD45 fragment 1 (SEQ ID NO: 177) and / Alternatively, the human CD45 epitope located within CD45 fragment 3 (sequence number 179) is CD This is an antibody or its antigen-binding portion that binds to 45.

[0241] In some embodiments, the antibody or its antigen-binding moiety is human CD45, canine At least one of the izal CD45 and / or rhesus macaque CD45 that is preserved between them. , at least two, at least three, at least four, or at least five amino acids It binds to CD45 with an epitope containing residues. For example, in some embodiments, The body or its antigen-binding site is the following amino acid residue of human CD45: 405T, 407K, 419Y, 425K, and 505R (fragments of hCD45 as described in Sequence ID No. 176) (Numbering as indicated) at least one, at least two, at least three, At least four or all five can be combined. In some embodiments, The antibody or its antigen-binding moiety is the following amino acid residue of human CD45: 405T, 407K , 419Y, 425K, 481R, and 505R, 509H (as listed in Sequence ID No. 176) (Numbering based on fragments of hCD45) one or more, two or more, three or more Four or more, five or more, six or more, or seven can be combined. Provided herein Furthermore, regarding binding to human CD45 (sequence number 175), Ab1, Ab2, Antibodies or antigenic antibodies that compete with Ab3, Ab4, Ab5, Ab6, and / or Ab7 This is the binding portion. In some embodiments, the antibody or its antigen-binding portion is also the crab Quismo macaque CD45 (SEQ ID NO: 194) and / or rhesus macaque CD45 (SEQ ID NO: 195) Regarding the binding to ), Ab1, Ab2, Ab3, Ab4, Ab5, Ab6, and / or It can compete with Ab7.

[0242] One of the following antibodies: Ab1, Ab2, Ab3, Ab4, Ab5, Ab6, or Ab7 An anti-human CD45 antibody or so which binds to an epitope on human CD45 that is conjugated by Fragments of (or the conclusion of Ab1, Ab2, Ab3, Ab4, Ab5, Ab6, or Ab7) Antibodies having a combined region are also intended for use in the methods and compositions provided herein. . Binding to human CD45 and / or cynomolgus monkey CD45 or rhesus monkey CD4 Regarding binding to 5, antibodies Ab1, Ab2, Ab3, Ab4, Ab5, Ab6, or A b7 (or a combination of Ab1, Ab2, Ab3, Ab4, Ab5, Ab6, or Ab7) Anti-human CD45 antibodies or their antigen binding cleavage that compete with any one of the antibodies (antibodies having a region) Further plans are also being considered.

[0243] In one embodiment, the present invention relates to a coupling region corresponding to that of Ab1, for example, CDR, This provides an anti-CD45 antibody or its antigen-binding fragment containing a variable region. Ab1 heavy chain variable The amino acid sequence of the region (VH) is described in SEQ ID NO: 67 (see Table 5). VH of Ab1 The CDR domain amino acid sequences are SEQ ID NO: 68 (VH CDR1), SEQ ID NO: 69 (VH As described in CDR2) and Sequence ID 70 (VH CDR3). Light chain variable region of Ab1 The very long (VL) amino acid sequence is described in SEQ ID NO: 71 (see Table 5). The VL C of Ab1. The DR domain amino acid sequences are SEQ ID NO: 72 (VL CDR1), SEQ ID NO: 73 (VL As described in CDR2) and Sequence ID 74 (VL CDR3). Therefore, a certain In the embodiments herein, the anti-CD45 antibody or its antigen-binding fragment provided herein is The heavy chain variable region containing the amino acid sequence described in SEQ ID NO: 67, and the amino acid sequence described in SEQ ID NO: 71 It includes a light chain variable region containing an amino acid sequence. In one embodiment, the anti-CD45 antibody is CDR1, CDR2 and the amino acid sequences described in SEQ ID NOs. 68, 69, and 70 A heavy chain containing CDR3 and the amino acid sequence described in SEQ ID NOs. 72, 73, and 74. It includes a light chain variable region containing CDR1, CDR2, and CDR3.

[0244] In one embodiment, the present invention provides a coupling region corresponding to that of Ab2, for example, CDR, The present invention provides an anti-CD45 antibody or its antigen-binding fragment containing a variable region. The heavy chain of Ab2 is variable. The amino acid sequence of the region (VH) is described in SEQ ID NO: 77 (see Table 5). VH of Ab2 The CDR domain amino acid sequences are: SEQ ID NO: 78 (VH CDR1), SEQ ID NO: 79 (VH As described in CDR2) and Sequence ID 80 (VH CDR3). Ab2 light chain variable region The very long (VL) amino acid sequence is described in SEQ ID NO: 81 (see Table 5). The VL C of Ab2. The DR domain amino acid sequences are SEQ ID NO: 82 (VL CDR1), SEQ ID NO: 83 (VL As described in CDR2) and Sequence ID 84 (VL CDR3). Therefore, a certain In the embodiments herein, the anti-CD45 antibody or its antigen-binding fragment provided herein is The heavy chain variable region containing the amino acid sequence described in SEQ ID NO: 77, and the amino acid sequence described in SEQ ID NO: 81 It includes a light chain variable region containing an amino acid sequence. In one embodiment, the anti-CD45 antibody is CDR1, CDR2 and the amino acid sequences described in SEQ ID NOs. 78, 79, and 80 A heavy chain containing CDR3 and the amino acid sequence described in SEQ ID NOs. 82, 83, and 84. It includes a light chain variable region containing CDR1, CDR2, and CDR3.

[0245] In one embodiment, the present invention relates to a coupling region corresponding to that of Ab3, for example, CDR, This provides an anti-CD45 antibody or its antigen-binding fragment containing a variable region. The heavy chain of Ab3 is variable. The amino acid sequence of the region (VH) is described in SEQ ID NO: 87 (see Table 5). VH of Ab3 The CDR domain amino acid sequences are SEQ ID NO: 88 (VH CDR1), SEQ ID NO: 89 (VH As described in CDR2) and Sequence ID 90 (VH CDR3). Light chain variable region of Ab3 The very long (VL) amino acid sequence is described in SEQ ID NO: 91 (see Table 5). The VL C of Ab3. The DR domain amino acid sequences are SEQ ID NO: 92 (VL CDR1), SEQ ID NO: 93 (VL As described in CDR2) and Sequence ID 94 (VL CDR3). Therefore, a certain In the embodiments herein, the anti-CD45 antibody or its antigen-binding fragment provided herein is The heavy chain variable region containing the amino acid sequence described in SEQ ID NO: 87, and the amino acid sequence described in SEQ ID NO: 91 It includes a light chain variable region containing an amino acid sequence. In one embodiment, the anti-CD45 antibody is CDR1, CDR2 and containing the amino acid sequences described in SEQ ID NOs. 88, 89, and 90 A heavy chain containing CDR3 and the amino acid sequence described in SEQ ID NOs. 92, 93, and 94. It includes a light chain variable region containing CDR1, CDR2, and CDR3.

[0246] In one embodiment, the present invention relates to a coupling region corresponding to that of Ab4, for example, CDR, This provides an anti-CD45 antibody or its antigen-binding fragment containing a variable region. The heavy chain of Ab4 is variable. The amino acid sequence of the region (VH) is described in SEQ ID NO: 97 (see Table 5). VH of Ab4 The CDR domain amino acid sequences are: SEQ ID NO: 98 (VH CDR1), SEQ ID NO: 99 (VH As described in CDR2) and Sequence ID 100 (VH CDR3). Ab4 light chain variable. The region (VL) amino acid sequence is described in SEQ ID NO: 101 (see Table 5). VL of Ab4 The CDR domain amino acid sequences are SEQ ID NO: 102 (VL CDR1), SEQ ID NO: 103 This is described in (VL CDR2) and Sequence ID No. 104 (VL CDR3). In certain embodiments, the anti-CD45 antibody or its antigenicity provided herein is used in some specific embodiments. The combined fragment contains a heavy chain variable region with the amino acid sequence described in SEQ ID NO: 97, and SEQ ID NO: 10 It includes a light chain variable region containing the amino acid sequence described in 1. In one embodiment, anti-CD The 45 antibody contains CDR1 amino acid sequences described in SEQ ID NOs. 98, 99, and 100. , a heavy chain containing CDR2 and CDR3, and as described in Sequence IDs 102, 103, and 104 It includes a light chain variable region containing CDR1, CDR2, and CDR3, which have amino acid sequences.

[0247] In one embodiment, the present invention relates to a coupling region corresponding to that of Ab5, for example, CDR, The present invention provides an anti-CD45 antibody or its antigen-binding fragment containing a variable region. The heavy chain of Ab5 is variable. The amino acid sequence of the region (VH) is described in SEQ ID NO: 107 (see Table 5). VH of Ab5 The CDR domain amino acid sequences are SEQ ID NO: 108 (VH CDR1), SEQ ID NO: 109 (VH CDR2) and Sequence ID 110 (VH CDR3) are described. Ab5 light The amino acid sequence of the variable chain region (VL) is described in SEQ ID NO: 111 (see Table 5). Ab5 The amino acid sequence of the VL CDR domain is sequence number 112 (VL CDR1), sequence number This is described in 113 (VL CDR2) and sequence number 114 (VL CDR3). Therefore, in certain embodiments, the anti-CD45 antibody or the anti-CD45 antibody provided herein is used. The antigen-binding fragment consists of a heavy chain variable region containing the amino acid sequence described in Sequence ID No. 107, and the sequence It includes a light chain variable region containing the amino acid sequence described in number 111. The anti-CD45 antibody has the amino acid sequence described in SEQ ID NOs: 108, 109, and 110. Heavy chain containing CDR1, CDR2 and CDR3, and sequence numbers 112, 113, and 11 Light chain variable region including CDR1, CDR2, and CDR3 containing the amino acid sequence described in 4 This includes.

[0248] In one embodiment, the present invention relates to a coupling region corresponding to that of Ab6, for example, CDR, The present invention provides an anti-CD45 antibody or its antigen-binding fragment containing a variable region. Ab6 heavy chain variable The amino acid sequence of the region (VH) is described in SEQ ID NO: 117 (see Table 5). VH of Ab6 The CDR domain amino acid sequences are SEQ ID NO: 118 (VH CDR1), SEQ ID NO: 119 (VH CDR2) and Sequence ID No. 120 (VH CDR3) are described. Ab6 light The amino acid sequence of the variable chain region (VL) is described in SEQ ID NO: 121 (see Table 5). Ab6 The amino acid sequence of the VL CDR domain is SEQ ID NO: 122 (VL CDR1), SEQ ID NO: This is described in 123 (VL CDR2) and sequence number 124 (VL CDR3). Therefore, in certain embodiments, the anti-CD45 antibody or the anti-CD45 antibody provided herein is used. The antigen-binding fragment consists of a heavy chain variable region containing the amino acid sequence described in Sequence ID No. 117, and the sequence It includes a light chain variable region containing the amino acid sequence described in number 121. The anti-CD45 antibody has the amino acid sequence described in SEQ ID NOs: 118, 119, and 120. Heavy chain containing CDR1, CDR2 and CDR3, and sequence numbers 122, 123, and 12 Light chain variable region including CDR1, CDR2, and CDR3 containing the amino acid sequence described in 4 This includes.

[0249] In one embodiment, the present invention relates to a coupling region corresponding to that of Ab7, for example, CDR, This provides an anti-CD45 antibody or its antigen-binding fragment containing a variable region. The heavy chain of Ab7 is variable. The amino acid sequence of the region (VH) is described in SEQ ID NO: 127 (see Table 5). VH of Ab7 The CDR domain amino acid sequences are SEQ ID NO: 128 (VH CDR1), SEQ ID NO: 129 (VH CDR2) and Sequence ID 130 (VH CDR3) are described. Light Ab7 The amino acid sequence of the variable chain region (VL) is described in SEQ ID NO: 131 (see Table 5). Ab7 The amino acid sequence of the VL CDR domain is sequence number 132 (VL CDR1), sequence number This is described in sequence 133 (VL CDR2) and sequence number 134 (VL CDR3). Therefore, in certain embodiments, the anti-CD45 antibody or the anti-CD45 antibody provided herein is used. The antigen-binding fragment consists of a heavy chain variable region containing the amino acid sequence described in Sequence ID No. 127, and the sequence It includes a light chain variable region containing the amino acid sequence described in number 131. The anti-CD45 antibody has the amino acid sequence described in SEQ ID NOs: 128, 129, and 130. Heavy chain containing CDR1, CDR2 and CDR3, and sequence numbers 132, 133, and 13 Light chain variable region including CDR1, CDR2, and CDR3 containing the amino acid sequence described in 4 This includes.

[0250] In a particular embodiment, the antibody is an HC variable domain as described in Table 5, or Table Includes a modified heavy chain (HC) variable region containing a variant of the HC variant region of 5, and a variant (i) The HC variable domains listed in Table 5 are 1, 2, 3, 4 or 5 A (ii) The HC variable drugs listed in Table 5 differ in substitution, addition, or deletion of the amino acid, The main component refers to the substitution, addition, or deletion of up to 5, 4, 3, 2, or 1 amino acid. In contrast, (iii) the HC variable domains listed in Table 5 are 1-5, 1-3, 1 It differs in the substitution, addition, or deletion of ~2, 2~5, or 3~5 amino acids, and (iv) at least approximately 75%, 80%, 85%, 90% of SEQ ID NO: 1, (i) In any of the above (iv), even if the amino acid substitution is a conservative amino acid substitution, it is a non-conservative amino acid substitution. Existing amino acid substitutions are also permitted.

[0251] In a particular embodiment, the antibody is an LC variable domain or the one described in Table 5. The modified light chain (LC) variable region includes variants, and the variants are (i) listed in Table 5. The LC variable domain is a domain in which 1, 2, 3, 4, or 5 amino acids are substituted or added. They differ in deletions, and (ii) the LC variable domains listed in Table 5 are, at most, 5, 4, 3. Different in the substitution, addition, or deletion of 2, 1 or 1 amino acid, (iii) The LC variable domains listed in Table 5 are 1-5, 1-3, 1-2, 2-5, or 3- The differences lie in the substitution, addition, or deletion of five amino acids, and / or (iv) Table 5 For the LC variable domains described, at least approximately 75%, 80%, 85%, 90%, and 9%. (i) ~ (iv) In any of these cases, the amino acid substitution is non-conservative, even if it is a conservative amino acid substitution. Amino acid substitutions are also acceptable.

[0252] In a particular embodiment, the anti-CD45 antibody is the CDR as described in Table 5 of this specification. The CD45 specificity of the antibody (i.e., AbA, AbB, or AbC) is included, and CDR is the CD45 specificity of the antibody. While maintaining similar specificity, conservative amino acid substitutions (or 2, 3, 4, or 5) (Contains amino acid substitutions.)

[0253] In a particular embodiment, the anti-CD45 antibody is an AbA, AbB, or AbC antibody. or a deimmunizing antibody based on its antigen-binding portion. The deimmunizing antibody has a V region Cells selected to lack T cell epitopes, or cells from which T cell epitopes have been removed. This has been modified to minimize the possibility of the antibody becoming immunogenic. or are eliminated. In a particular embodiment, the anti-CD45 antibody eliminates the T cell epitope. Deimmunization is achieved by selecting or manipulating framework domains to exclude them. When a T cell epitope is present in the antibody sequence, the human subject will react to the anti-CD45 antibody. It may cause an AHA / HAMA reaction, which could result in adverse events in human subjects. Alternatively, an immune-mediated reaction may occur that results in a decrease in therapeutic effect. Antibodies (i.e., the variable sequences and CDRs of AbA, AbB, and AbC listed in Table 5) The sequence can function as a parent sequence from which deimmunized antibodies can be derived.

[0254] In one embodiment, the anti-CD45 antibody is BIOLEGEND(registered trademark) (San The clone HI30 or its humanized variant is commercially available from Diego, CA. It exists or is derived from it. The humanization of antibodies is known in the art. Follow the procedure (for example, as described in Example 7 below) to frame the non-human antibody This is done by replacing the K residue and constant region residues with those of germline human antibodies. This can be done. Additional anti-CD4 can be used in connection with the methods described herein. The five antibodies are anti-CD45 antibodies ab10558, EP322Y, MEM-28, and ab105 59, 0.N.125, F10-89-4, HIe-1, 2B11, YTH24.5, P D7 / 26 / 16, F10-89-4, 1B7, ab154885, B-A11, Fluorescent S 1007, ab170444, EP350, Y321, GA90, D3 / 9, X1 6 / 99, and LT45 (these are registered trademarks of ABCAM (Cambridge, MA) or This includes commercially available products, as well as their humanized variants. Further anti-CD45 antibodies that may be used in connection with patient conditioning treatment include S Commercially available from IGMA-ALDRICH® (Registered Trademark) (St. Louis, MO) This includes the anti-CD45 antibody HPA000440 and its humanized variant. Additional anti-CD45 compounds that can be used in connection with the patient conditioning methods described. The antibodies include the mouse monoclonal antibody BC8, for example, Matthews et al. It is described in al., Blood 78:1864-1874, 1991, and its opening The references herein are incorporated herein by reference with respect to anti-CD45 antibodies and their humanized variants. Further anti-CD45 antibodies that can be used in connection with the methods described herein. This includes the monoclonal antibody YAML568, for example, Glatting et al. This is described in al., J.Nucl.Med.8:1335-1341, 2006. The disclosure relating to anti-CD45 antibodies and their humanized variants is provided herein by reference. To be used in connection with patient conditioning treatments described herein. Additional anti-CD45 antibodies that can be produced include the monoclonal antibodies YTH54.12 and YTH2 5.4 is included, for example, Brenner et al., Ann. NYAcad. This is described in Sci.996:80-88,2003, and the disclosure concerns anti-CD45 antibodies. With respect to the humanized variant thereof, references are incorporated herein by reference. Additional anti-CD45 antibodies for use with the patient conditioning method include U This includes CHL1, 2H4, SN130, MD4.3, MBI, and MT2, for example, B Rown et al., Immunology 64:331-336, 1998. It is included, and the disclosure relating to anti-CD45 antibodies and their humanized variants is referenced. More herein by reference. May be used in connection with the methods described herein. Additional anti-CD45 antibodies include American Type Culture Colle Accession numbers RA3-6132, RA3-2C2, and T Substances produced and released from IB122, as well as monoclonal antibody C363.16A , and 13 / 2 are included, for example, Johnson et al., J.Exp.Med This is described in .169:1179-1184,1989, and the disclosure is anti-CD45 anti With respect to the body and its humanized variants, references are incorporated herein by reference. Further anti-CD45 can be used in connection with the patient conditioning methods described. Antibodies include monoclonal antibodies AHN-12.1, AHN-12, AHN-12.2, A This includes HN-12.3, AHN-12.4, HLe-1, and KC56(T200). For example, Harvath et al., J.Immunol. 146:949-957 This was described in 1991, and the disclosure relates to anti-CD45 antibodies and their humanized variants. With regard to this, references are incorporated herein by reference.

[0255] Additional Examples of anti-CD45 antibodies include those described in U.S. Patent No. 7,265,212 (for example). In particular, the anti-CD45 antibodies 39E11, 16C9, and 1G10 are described; the same No. Those described in publication number 7,160,987 (for example, ATCC such as monoclonal antibody 6G3) Regarding the anti-CD45 antibody produced and released by accession number HB-11873 (as described in the document); and as described in Article 6,099,838 (for example, anti-CD45 antibody MT3) , as well as ATCC accession numbers HB220 (also written as MB23G2) and HB2 (Describes antibodies produced and released by 23), and US2004 / 0096 Those described in 901 and US2008 / 0003224 (for example, monoclonal antibody 1 7.1, etc., produced and released by ATCC accession number PTA-7339. This includes a description of anti-CD45 antibodies, and each disclosure relates to anti-CD45 antibodies. This is incorporated herein by reference.

[0256] Further information that can be used in connection with the patient conditioning methods described herein may be found. The anti-CD45 antibodies include ATCC accession numbers MB4B4, MB23G2, and 14. 8, GAP8.3, 74-9-3, I / 24.D6, 9.4, 4B2, M1 / 9.3.4 Antibodies produced and released from HL.2, as well as humanized and / or affinity-matured antibodies. This includes riant. Affinity maturation is, for example, described herein or in the technical field. In vitro display technologies known in the field, such as phage displays, This can be done using method (i).

[0257] Additional The anti-CD45 antibody includes anti-CD45 antibody T29 / 33, which is, for example, Mor ikawa et al.,Int.J.Hematol.54:495-504,19 As described in paragraph 91, the disclosure relating to anti-CD45 antibodies is provided herein by reference. It is used.

[0258] In a particular embodiment, the anti-CD45 antibody is apamistamab (90Y-BC8, Iomab-B, also known as BC8; e.g., US20170326259, WO 2017155937, and Orozco et al.Blood.127.3(20 16):352-359 (as described), or BC8-B10 (e.g., Li et (as described in al.PloS one 13.10(2018):e0205135) They are selected, and each is referenced by means of. Other anti-CD45 antibodies are, for example, WO2003 / 048327, WO2016 / 016442, US2017 / 02262 09, US2016 / 0152733, US9,701,756; US2011 / 007 Listed in 6270, or US7,825,222, each of which is the whole This is invoked by reference.

[0259] For example, in one embodiment, an anti-CD45 antibody or its antigen-binding fragment is used in Apamista Includes the binding region corresponding to that of mab, e.g., CDR, variable region. Weight of apamistamab The amino acid sequence of the variable chain region (VH) is described in SEQ ID NO: 7 (see Table 5). Apamista The amino acid sequence of the light chain variable region (VL) of mab is described in SEQ ID NO: 8 (see Table 5). In this embodiment, the anti-CD45 antibody or its antigen-binding moiety is as described in SEQ ID NO: 7 It includes a variable heavy chain containing amino acid residues and a light chain variable region described in Sequence ID No. 8. In terms of administration, the anti-CD45 antibody is apamystamab CDR1, CDR2, and CDR3 A heavy chain containing and a light chain variable region containing CDR1, CDR2 and CDR3 of apamistamab Includes.

[0260] In one embodiment, the anti-CD45 antibody is the heavy chain of the anti-CD45 antibody described herein. This includes the light chain variable region of the anti-CD45 antibody described herein. In one embodiment, The anti-CD45 antibodies are CDR1, CDR2 and C of the anti-CD45 antibodies described herein. A heavy chain containing DR3 and the anti-CD45 antibodies CDR1, CDR2 and C described herein. Includes a light chain variable region including DR3.

[0261] In another embodiment, the antibody or its antigen-binding fragment is attached to the anti-CD45 antibody of this specification. In contrast, at least 95% identity, for example, against the anti-CD45 antibody of this specification, At least 95%, 96%, 97%, 98%, 99%, or 100% identity of the Ami It contains a heavy chain variable region that includes an acid sequence. In certain embodiments, the antibody is as described herein. Modified heavy chain (HC) variable domains including the HC variable domain of an anti-CD45 antibody or its variant The variant includes (i) substitutions and additions of 1, 2, 3, 4, or 5 amino acids. In terms of deletion, or (ii) up to 5, 4, 3, 2, or 1 Unlike anti-CD45 antibodies, (iii)1 Substitution, addition, or deletion of ~5, 1~3, 1~2, 2~5, or 3~5 amino acids Unlike anti-CD45 antibodies, and / or (iv) compared to anti-CD45 antibodies, Approximately 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% It contains one amino acid sequence, and in any of (i) to (iv), amino acid substitutions are The amino acid substitution may be either a conserved or non-conserved amino acid substitution, and the modified heavy chain variable region The region maintains the CD45 binding specificity of the antibody, while being different from the heavy chain variable region of the anti-CD45 antibody. It may possess enhanced biological activity.

[0262] Antibodies and antigen-binding fragments that may be used in connection with the compositions and methods described herein This includes not only the antibodies and their antigen-binding fragments mentioned above, but also the non-human antibodies and antigen-binding fragments mentioned above. Humanized variants of the composite fragment, and evaluation by, for example, competitive CD45 binding assays. Therefore, it contains antibodies or antigen-binding fragments that bind to the same epitopes as described above. ru.

[0263] Consensus CDR Ab1, Ab2, Ab3, Ab4, Ab5, Ab6, and Ab7 are found on human CD45. They bind to the same epitope and share several consensus residues in its CDR region. (Please refer to International Application No. PCT / US2020 / 058373, and refer to this specification.) (Used in the book). The consensus heavy chain amino acid CDR sequence is SEQ ID NO: 188, SEQ ID NO: As shown in sequence 189 and sequence number 190, the consensus light chain amino acid CDR sequence is sequence number As shown in code 191, sequence number 192, and sequence number 193.

[0264] Therefore, in some embodiments, the anti-CD45 antibody or its antigen-binding portion , the CDR1 domain containing the amino acid sequence described in SEQ ID NO: 188, SEQ ID NO: 189 The CDR2 domain containing the amino acid sequence described, and the amino acid sequence described in SEQ ID NO: 190 A heavy chain variable region containing a CDR3 domain containing an acid sequence, and the a sequence described in SEQ ID NO 191 CDR1 domain containing amino acid sequence, C containing amino acid sequence described in SEQ ID NO: 192 The DR2 domain and the CDR3 domain containing the amino acid sequence described in SEQ ID NO: 193. The aforementioned antibody may include a light chain variable region containing a heavy chain. In some embodiments, the antibody may include a heavy chain variable region. The constant region and / or light chain steady region may further be included. For example, in some embodiments, The aforementioned antibodies are SEQ ID NOs. 183, 184, 185, and 186. Alternatively, a heavy chain constant region selected from any of the sequences listed in sequence number 187, and / or may further include the light chain constant region described in Sequence ID No. 182.

[0265] Methods for identifying antibodies Antigens expressed by hematopoietic stem cells or mature immune cells (e.g., T cells) (e.g., High antibody or antibody fragment library for molecules capable of binding to CD45 Using methods for throughput screening, for the treatment of cancer, autoimmune diseases, and The condition of patients (e.g., human patients) requiring hematopoietic stem cell therapy as described herein It is possible to identify antibodies useful for conditioning and perform affinity maturation. Such methods include Among them, phage displays, bacterial displays, yeast displays, and mammalian displays. Cellular display, ribosome display, mRNA display, and cDNA display This includes in vitro display technologies known in the field, such as play. It is produced. For isolating antibodies or antigen-binding fragments that bind to biologically related molecules. The use of a phallus display is, for example, by Felici et al., Biotechn. ol.Annual Rev.1:149-183,1995;Katz,Annual Rev.Biophys.Biomol.Struct.26:27-45,1997 ; and Hoogenboom et al., Immunotechnology 4: This is outlined in 1-20, 1998, and each disclosure is in vitro. Regarding play techniques, they are incorporated herein by reference. Polypeptides that bind to cell surface antigens To select peptides, a randomized combinatorial peptide library is constructed. Built by Kay, Perspect. Drug Discovery Des.2 :251-268, 1995 and Kay et al., Mol. Divers. 1:1 As described in 39-140, 1996, each disclosure is an antigenic linkage. Regarding the discovery of synthetic molecules, references are incorporated herein by reference. Examples include multimeric proteins and other tans. The protein is well phage-displayed as a functional molecule (e.g., EP034 9578;EP4527839;and EP0589877, and Chiswell and McCafferty,Trends Biotechnol.10:80-8 See 4 1992, each disclosure of which is in the discovery of antigen-binding molecules. With regard to the use of in vitro display technology, it is incorporated herein by reference. In addition Functional antibody fragments such as Fab and scFv fragments are displayed in vitro. It is expressed in the formula (for example, McCafferty et al., Nature 3 48:552-554,1990;Barbas et al.,Proc.Natl. Acad.Sci.USA 88:7978-7982, 1991; and Clackso Please refer to n et al., Nature 352:624-628, 1991. Each of these disclosures is an in vitro display device for the discovery of antigen-binding molecules. Regarding the form, it is incorporated herein by reference. Human anti-CD45 antibodies also For example, HuMAb-Mouse® or XenoMouse® These technologies can generate antigens expressed by hematopoietic stem cells. Identify antibodies or fragments capable of binding to (for example, CD45) and determine the affinity of the antibodies. It can be used to improve the condition of patients who require hematopoietic stem cell transplantation therapy. It can be used to deplete endogenous hematopoietic stem cells in (for example, human patients). ru.

[0266] In addition to in vitro display technology, computational modeling techniques are used to create An antibody capable of binding to an antigen (e.g., CD45) expressed by hematopoietic stem cells is i It can be designed and identified using n silico. For example, a person skilled in the art can use computational methods. Using modeling techniques, specific epitopes on CD45, such as extracellular epitopes of CD45, are used. For molecules capable of binding to a tope, a library of antibodies or antibody fragments is used in Screening can be done using silico.

[0267] Using additional techniques, antigens expressed by hematopoietic stem cells (e.g., CD45) are linked. It is possible to combine them, for example, by receptor-mediated endocytosis, in cells This allows for the identification of antibodies or antibody fragments that are internalized. For example, in hematopoietic stem cells Therefore, it binds to the expressed antigen (e.g., CD45), and is subsequently internalized by the cell. To screen for antibodies or antibody fragments that will be converted, use the in vitro method described above. The display technology can be appropriately modified. Phage displays are this screen It is one such technique that can be used in combination with the leaning paradigm. To identify the anti-CD45 antibody subsequently internalized by hematopoietic stem cells, those skilled in the art will know: Williams et al., Leukemia 19:1432-1438,20 The phage display technology described in 05 can be used (the disclosure thereof is (The whole is incorporated herein by reference). For example, known in the art Mutagenesis is used to create antibodies, antibody fragments, for example, scFv fragments, Fab Fragments, diabodies, triabodies, and 10 Fn3 domain, or randomized Amino acid cassette (for example, one or more or all of the CDR or equivalent regions) Recombinant phage libraries encoding ligands (or antibodies or antibody fragments) A Lee can be created. The framework region of the antibody or antibody fragment, hinge, Fc The domain and other regions are, for example, human germline antibody sequences or human germline By having a sequence that shows only slight changes in response to antibodies, it is possible for non-immune humans to It can be designed to be original.

[0268] Phage displays described herein or known in the art Using this technology, for example, first, an antibody or antibody fragment that exhibits nonspecific protein binding is used. Phages that bind to the Fc domain, and antibodies or fragments thereof that encode phages that bind to the Fc domain. To remove the phage, block the phage library with a blocking agent (e.g., milk protein, bovine protein). Incubate with serum albumin and / or IgG, etc., and then phage The library can be, for example, hematopoietic stem cells or mature immune cells that express CD45 (e.g., By incubating with a population of T cells, the phage particles were covalently bound to them. A phage library containing randomized antibodies or antibody fragments is used with an antigen (e.g., CD4). 5) can be incubated with antibodies (e.g., A (anti-CD45 antibody) or antibody fragment binds to a related cell surface antigen (e.g., CD45). Then, a sufficient amount of time (e.g., 4) is allowed for internalization by hematopoietic stem cells. Incubate with hematopoietic stem cells for 30 minutes to 6 hours at ℃ (for example, 1 hour at 4℃). Yes, it is possible. However, hematopoiesis does not occur because it does not show sufficient affinity for the antigen (e.g., CD45). It contains antibodies or antibody fragments that do not bind to stem cells and are not internalized by hematopoietic stem cells. Phages are washed with, for example, a cold (4°C) 0.1M glycine buffer at pH 2.8. It can be removed by purification. Antibodies internalized by hematopoietic stem cells Phages bound to antibody fragments can, for example, lyse cells and break down internalized phages. It can be identified by recovering it from the cell culture medium. Subsequently, in the art Using known methods, for example, phages and bacterial cells recovered in 2xYT medium By incubating the phages together, it is possible to amplify them in bacterial cells. Next, the phages recovered from this culture medium are, for example, inserted into the phage genome. By determining the nucleic acid sequence of the gene(s) encoding the antibody or antibody fragment Then, the characterization can be performed. Subsequently, the encoded antibody or antibody fragment is chemically synthesized. By (for example, antibody fragments such as scFv fragments) or recombinant expression (for example, full-length antibodies) It can be newly prepared.

[0269] The internalization ability of the prepared antibody or antibody fragment can be evaluated, for example, using a radionuclide internalization assay known in the art. For example, an antibody (e.g., an anti-CD45 antibody) or antibody fragment identified using an in vitro display technique described herein or known in the art can be functionalized by incorporation of radionuclides such as F, Br, 18 F, 75 Br, 77 Br, 122 I, 123 I, 124 I, 125 I, 129 I, 131 I, 21 1 At, 67 Ga, 111 In, 99 Tc, 169 Yb, 186 Re, 64 Cu, 67 C u, 177 Lu, 77 As, 72 As, 86 Y, 90 Y, 89 Zr, 212 Bi, 213 Bi, or 225 Ac. For example, radionuclides such as 18 F, 75 Br, 77 Br, 122 I, 123 I, 124 I, 125 I, 129 I, 131 I, 211 radioactive halogens such as At can be incorporated by containing an electrophilic halogen reagent Polystyrene beads (e.g., Iodination Beads, Thermo F B (Beetle) such as Isher Scientific, Inc., Cambridge, MA It can be incorporated into antibodies or antibody fragments using radiolabeled antibodies, etc. The fragments, or ADCs, should be left for a sufficient amount of time to allow internalization (e.g., 30 minutes at 4°C). Hematopoietic stem cells can be incubated for 6 hours (for example, 1 hour at 4°C). Next... Then, the cells can be washed to remove non-internalized antibodies or fragments thereof (e.g.) For example, using a cold (4°C) 0.1M glycine buffer with a pH of 2.8. (Internalized antibody) Alternatively, antibody fragments can recover radiation (e.g., gamma rays) emitted from the obtained hematopoietic stem cells. By comparing and detecting the radiation (e.g., gamma rays) emitted from the washing buffer, It can be identified. The aforementioned internalization assay can also be used for characterizing ADCs. can.

[0270] Antibodies are recombinant and compositions as described, for example, in U.S. Patent No. 4,816,567. It can be prepared using the anti-CD45 antibody described herein. An isolated nucleic acid encoding is provided. Such a nucleic acid contains amino acids including the VL of the antibody. amino acid sequences containing sequences and / or VH (e.g., light and / or heavy chains of antibodies) It may be possible. In a further embodiment, one or more vectors containing such nucleic acids (e.g. In a further embodiment, a host comprising such nucleic acid is provided. Cells are provided. In one such embodiment, the host cells contain (1) a very large amount of the antibody. A vector containing a nucleic acid encoding an amino acid sequence and an amino acid sequence including the VH of an antibody, (2) A first vector and an antibody comprising a nucleic acid encoding an amino acid sequence including the VL of the antibody. The second vector includes a nucleic acid that encodes an amino acid sequence containing the body's VH (for example, (Transformed by this). In one embodiment, the host cell is a eukaryote, for example, Chinese hamster ovary (CHO) cells or lymphoid cells (e.g., Y0, NS0) (Sp20 cells). In one embodiment, a method for producing an anti-CLL-1 antibody is provided. The method involves a host cell containing nucleic acid encoding the antibody provided above, which is then used to express the antibody. By culturing under suitable conditions and, optionally, by extracting from host cells (or host cell culture medium) This includes recovering antibodies.

[0271] For recombinant production of anti-CD45 antibodies, for example, the nucleic acid encoding the above antibody is isolated. Then, for further cloning and / or expression in host cells, one or more vectors are used. Such nucleic acids are inserted using conventional procedures (e.g., the heavy and light chains of antibodies). Using oligonucleotide probes capable of specifically binding to the gene encoding the gene. (Therefore) it can be easily isolated and sequenced.

[0272] Suitable host cells for cloning or expressing antibody-encoding vectors are specified herein. This includes prokaryotic or eukaryotic cells as described. For example, antibodies, in particular, glycosylated And if the Fc effector function is not required, it can be produced in bacteria. Antibacterial properties in bacteria For the expression of somatic fragments and polypeptides, see, for example, U.S. Patent No. 5,648,237. See also issues Nos. 5,789,199 and 5,840,523. (E.co Charlton, Methods in li. describes the expression of antibody fragments. Molecular Biology,Vol.248(BKCLo,ed.,H umana Press, Totowa, NJ, 2003), pp.245-254 See also.) After expression, the antibody can be isolated from the bacterial cell paste in a soluble fraction, and further It can be purified.

[0273] Vertebrate cells can also be used as hosts. For example, they can be modified to grow in suspension. Mammalian cell lines that have been modified may be useful. Another example of a useful mammalian host cell line is SV4 CV1 strain of monkey kidney transformed by 0 (COS-7); human embryonic kidney strain (293 strain) Cells or, for example, Graham et al., J. Gen Virol. 36:59 (1 977) 293 cells; baby hamster kidney cells (BHK); mouse Sertoli cells Cells (e.g., Mather, Biol. Reprod. 23:243-251 (198) 0) TM4 cells as described above; monkey kidney cells (CV1); African green monkey kidney cells (V ERO-76); Human cervical cancer cells (HELA); Canine kidney cells (MDCK); Buffer Rat hepatocytes (BRL 3A); human lung cells (W138); human hepatocytes (Hep G 2); Mouse mammary gland tumors (MMT 060562); e.g., Mather et al. , as described in Annals NYAcad.Sci.383:44-68(1982) These include TRI cells; MRC 5 cells; and FS4 cells. Other useful mammalian host cell lines include , DHFR-CHO cells (Urlaub et al., Proc. Natl. Acad Chinese hamster ovaries (including .Sci.USA 77:4216(1980)) This includes CHO cells, as well as myeloma cell lines such as Y0, NS0, and Sp2 / 0. For a review of specific mammalian host cell lines suitable for body production, see, for example, Yazaki a. nd Wu,Methods in Molecular Biology,Vol.2 48 (BKCLo, ed., Humana Press, Totowa, NJ See pp.255-268 (2003). In one embodiment, host cell eukaryotes, such as Chinese hamster ovary (CHO) cells or lymphoid cells. (For example, Y0, NS0, and Sp20 cells.)

[0274] Antibody-drug conjugates The antibodies and antigen-binding fragments described herein are contaminated with cytotoxicity via a linker. To be jugated (linked). In some embodiments, the antibody or fragment thereof Following intracellular uptake, the cytotoxic substance approaches its intracellular target and can mediate hematopoietic cell death. A cytotoxic molecule is transferred to an intracellularly located antibody or its antigen-binding fragment disclosed herein. It is conjugated. Any number of cytotoxic substances, e.g., 1, 2, 3, 4, 5, 6, 7, and It is possible to conjugate eight cytotoxic substances to an anti-CD45 antibody or its antigen-binding fragment. Cut.

[0275] Cytotoxics suitable for use with the compositions and methods described herein include those in the relevant art. Among those known to exist, DNA intercalation agents (for example, anthracite) Ikurin), drugs capable of destroying the mitotic spindle (e.g., vinca alkaloids) (e.g., mytansin, meitansinoids, and their derivatives), RNA polymerase inhibitors (e.g.) For example, amatoxins such as α-amanitin and its derivatives, and disrupt protein biosynthesis. Drugs that can break down (e.g., rRNAN glycosylation such as saporins and lysine A chain) It contains drugs that exhibit dysidase activity.

[0276] cytotoxin For use in the therapies described herein, various cytotoxics are linked to anti-C via a linker. It can be conjugated to a D45 antibody or its antigen-binding fragment. In particular, anti-CD45 AD C contains a conjugated (i.e., linker) cytotoxic portion (or cytotoxic portion). Includes an antibody (or its antigen-binding fragment) covalently bound by [a specific mechanism]. Various embodiments In this case, the cytotoxic portion exhibits low cytotoxicity when bound to the conjugate. They may or may not exhibit this behavior, but regain cytotoxicity once cleaved from the linker. In some embodiments, the cytotoxic portion is cytotoxic even if it is not cleaved from the linker. Maintain. In some embodiments, following intracellular uptake of the antibody or a fragment thereof. Cytotoxicity allows cytotoxic substances to approach their intracellular targets and, for example, mediate T cell death. The molecule is conjugated to an intracellularly located antibody or its antigen-binding fragment disclosed herein. It can be done.

[0277] Therefore, the ADC of this disclosure is given by the general formula Ab-(ZLD) n It may be of the same nature as, In the formula, the antibody or its antigen-binding fragment (Ab) is connected via the chemical moiety (Z), respectively. The linker (L) and cytotoxic portion ("drug") disclosed in the document are conjugated To be connected (covalently).

[0278] Thus, an antibody or its antigen-binding fragment is a drug represented by the integer n. It can be conjugated in parts, where n represents the average number of toxins per antibody, for example, approximately It can range from 1 to approximately 20. In some embodiments, n is 1 to 4. In several embodiments, n is 1. ADC is prepared from a conjugation reaction. In this case, the average number of drug portions per antibody is determined by mass spectrometry, ELISA assay, and HPL It can be characterized by conventional methods such as C. Furthermore, the quantitative distribution of ADC can be described as follows: n may be determined as a standard. In some cases, n is a specific value. Separating, purifying, and characterizing an ADC from an ADC carrying other drugs is performed using reversed phase This can be achieved by means such as HPLC or electrophoresis.

[0279] Some anti-CD45 ADCs may be limited by the number of binding sites on the antibody. For example, if the binding is to cysteinethiol, the antibody will have one or more cysteinethiols. Either they can only have inchol groups, or they are sufficiently reactive to which linkers can be bonded. It can have only one or a few thiol groups. Generally, antibodies are linked to the drug moiety. It does not contain a large number of free and reactive cysteinethiol groups that may be present, and is mainly found in antibodies. Cysteine ​​thiol residues exist as disulfide bridges. In certain embodiments The antibodies are dithiothreitol (DTT) or tricarbonylethylphosphine. It is reduced under partial or total reduction conditions by reducing agents such as TCEP, and the reactive A steinchol group may be generated. In certain embodiments, the amount of drug loading increases. And (for example, n>5), aggregation, insolubility, toxicity of a particular antibody-drug conjugate, and This can lead to a loss of cell permeability.

[0280] In a particular embodiment, the antibody is conjugated in a conjugation reaction. The drug portion is less than the theoretical maximum. Antibodies, as discussed below, for example, It may contain lysine residues that do not react with drug linker intermediates or linker reagents. In some cases, only highly reactive lysine groups react with amine-reactive linker reagents. In the application process, antibodies are used to clearly identify reactive nucleophiles such as lysine or cysteine. It is subjected to alteration conditions.

[0281] The loading of ADCs (drug / antibody ratio) can be done in various ways, for example, (i) molar excess relative to the antibody. (ii) Restricting drug linker intermediates or linker reagents, (iii) Limiting the reaction time or reaction temperature, and reducing the cysteinethiol modification. (iv) to partially or limit the number and / or location of linker drug bonds The amino acid sequence of the antibody is modified to change the number and position of cysteine ​​residues in order to control this. It can be controlled by operating it using replacement technology.

[0282] Cytotoxics suitable for use with the compositions and methods described herein include those in the relevant art. Among those known to exist, DNA intercalation agents (for example, anthracite) Ikurin), drugs capable of destroying the mitotic spindle (e.g., vinca alkaloids) (e.g., mytansin, meitansinoids, and their derivatives), RNA polymerase inhibitors (e.g.) For example, amatoxins such as α-amanitin and its derivatives, and disrupt protein biosynthesis. Drugs that can break down (e.g., rRNAN glycosylation such as saporins and lysine A chain) It contains drugs that exhibit dysidase activity.

[0283] In some embodiments, the cytotoxic is a microtubule-binding agent (e.g., mytansin or Maytansinoids), amatoxins, Pseudomonas exotoxin A, debuganin, diphtheria A toxin, saporin, auristatin, anthracycline, calichemycin, irinoteca N, SN-38, Duocalmycin, Pyrrolobenzodiazepine, Pyrrolobenzodiazepine Indolino-benzodiazepine dimer, indolinobenzodiazepine dimer, indolin Nobenzodiazepine pseudodimers, or variants thereof, or as described herein or another cytotoxic compound known in the relevant art.

[0284] In some embodiments, the cytotoxicity of the antibody-drug conjugate is RNA polymerase It is an RNA polymerase inhibitor. In some embodiments, the RNA polymerase inhibitor is amatox The antibody disclosed herein is a syn or a derivative thereof. In some embodiments, the antibody disclosed herein The cytotoxic components of the drug conjugate are α-amanitin, β-amanitin, γ-amanitin, and ε - Amanitin, Amanin, Amaninamide, Amanulin, Amanulinic Acid, Proamanulin amatoxin or its derivatives, such as a derivative thereof.

[0285] This disclosure relates to cytotoxicities that can be used in anti-CD45 ADCs useful in the methods described herein. Further details are provided below.

[0286] Amatoxin The methods and compositions disclosed herein involve an anti-CD45 antibody or its antigen-binding fragment. As a cytotoxic agent, RNA polymerase inhibitors, such as amatoxin, are used. Contains ADC. In some embodiments, RNA polymerase inhibitors are Amato It is a xine or a derivative thereof. In some embodiments, the anti is disclosed herein. The cytotoxic components of the body's drug conjugates are α-amanitin, β-amanitin, γ-amanitin, ε-Amanitin, Amanin, Amaninamide, Amanulin, Amanulinic Acid, Proamanuri It is amatoxin or its derivatives, such as cin or its derivatives. Various naturally occurring compounds The structure of amatoxin is, for example, Zanotti et al., Int.J.Pept. This is disclosed in ide Protein Res. 30, 1987, 450-459. The amatoxins useful in relation to the compositions and methods described herein are not limited to: However, compounds that follow formula (III), such as α-amanitin, β-amanitin, γ - Amanitin, ε-Amanitin, Amanin, Amaninamide, Amanulin, Amanuric Acid , or proamanulin is included. Formula (III) is as follows: [ka] In the formula, R1 is H, OH, or OR A And, R2 is H, OH, or OR B And, R A and R B If present, they, together with the oxygen atom to which they are bonded, will be selected A 5-membered heterocycloalkyl group is formed by selective substitution, R3 is H or R D And, R4 is H, OH, OR D , or R D And, R5 is H, OH, OR D , or R D And, R6 is H, OH, OR D , or R D And, R7 is H, OH, OR D , or R D And, R8 is OH, NH2, or OR D And, R9 is H, OH, or OR D And, X is -S-, -S(O)-, or -SO2-, R D This refers to an alkyl group substituted by optional choice (e.g., C1-C6 alkyl group), optional choice Heteroalkyls substituted by (e.g., C1-C6 heteroalkyls), optionally by Substituted alkenyls (e.g., C2-C6 alkenyls), optionally substituted Heteroalkenyls (e.g., C2-C6 heteroalkenyls), optionally substituted Alkynnyl (e.g., C2-C6 alkynyl), heteroalkynyl by optional substitution Nyl (e.g., C2-C6 heteroalkynyl), optionally substituted cycloalkyl L, optionally substituted heterocycloalkyl, optionally substituted A It is a heteroaryl compound, or a heteroaryl compound that has been optionally substituted.

[0287] For example, in one embodiment, useful in relation to the compositions and methods described herein Amatoxins include compounds that follow formula (IIIA). [ka] In the formula, R4, R5, X, and R8 are as defined above.

[0288] For example, in one embodiment, useful in relation to the compositions and methods described herein Amatoxins include compounds that follow the following formula (IIIB). [ka] In the formula, R1 is H, OH, or OR A And, R2 is H, OH, or OR B And, R A and R B If present, they, together with the oxygen atom to which they are bonded, will be selected A 5-membered heterocycloalkyl group is formed by selective substitution, R3 is H or R D And, R4 is H, OH, OR D , or R D And, R5 is H, OH, OR D , or R D And, R6 is H, OH, OR D , or R D And, R7 is H, OH, OR D , or R D And, R8 is OH, NH2, or OR D And, R9 is H, OH, or OR D And, X is -S-, -S(O)-, or -SO2-, R D This refers to an alkyl group substituted by optional choice (e.g., C1-C6 alkyl group), optional choice Heteroalkyls substituted by (e.g., C1-C6 heteroalkyls), optionally by Substituted alkenyls (e.g., C2-C6 alkenyls), optionally substituted Heteroalkenyls (e.g., C2-C6 heteroalkenyls), optionally substituted Alkynnyl (e.g., C2-C6 alkynyl), heteroalkynyl by optional substitution Nyl (e.g., C2-C6 heteroalkynyl), optionally substituted cycloalkyl L, optionally substituted heterocycloalkyl, optionally substituted A It is a heteroaryl compound, or a heteroaryl compound that has been optionally substituted.

[0289] In one embodiment, a useful amatoki in relation to the c...

Claims

1. Depletion of the CD45+ cell population in human patients requiring hematopoietic stem cell (HSC) transplantation A method for causing the patient to undergo a transplant containing allogeneic HSCs, wherein the cytotoxic binds to the patient before the transplant. The procedure involves administering an effective amount of the CD45-targeted portion to the patient, wherein the patient has the transplant The following conditions are not met: The condition is not treated with immunosuppressant conditioning before or approximately simultaneously with the transplant. method.

2. a. In human patients, the CD45 targeting portion bound to the cytotoxin was introduced in the absence of immunosuppressants. , administer in an effective dose sufficient to deplete the population of CD45+ cells in the patient, b. A method comprising subsequently performing a transplant containing allogeneic HSCs on the patient.

3. The procedure includes performing a transplant containing allogeneic HSCs into a human patient, wherein the patient is free from immunosuppressants. Below, administration of a CD45-targeted moiety bound to a cytotoxic to a population of hematopoietic stem cells in the patient. A method that involves receiving a sufficient amount beforehand to deplete the system.

4. The CD45-targeting portion bound to the cytotoxin is an anti-CD45 antibody drug conjugate ( The method according to any one of claims 1 to 3, wherein the method is ADC.

5. The aforementioned homogeneous HSC contains one or more HLA mismatches with the patient's HLA antigen. The method according to any one of claims 1 to 4.

6. The aforementioned homogeneous HSC contains two or more HLA mismatches with the patient's HLA antigen. The method according to any one of claims 1 to 4.

7. The aforementioned homogeneous HSC contains three or more HLA mismatches with the patient's HLA antigen. The method according to any one of claims 1 to 4.

8. The aforementioned homogeneous HSC contains five or more HLA mismatches with the patient's HLA antigen. The method according to any one of claims 1 to 4.

9. The aforementioned homogeneous HSCs include a complete HLA mismatch with the patient's HLA antigen. The method according to any one of claims 1 to 4.

10. The aforementioned homogeneous HSCs have one or more minor combinations of the minor histocompatibility antigens of the patient. The method according to any one of claims 1 to 9, comprising a textile compatibility antigen (miHA) mismatch.

11. The aforementioned homogeneous HSCs have two or more miHA- The method according to any one of claims 1 to 9, including smatching.

12. The aforementioned homogeneous HSCs have five or more miHA- The method according to any one of claims 1 to 9, including smatching.

13. The immunosuppressant is whole-body irradiation (TBI) according to any one of claims 1 to 12. method.

14. The method according to claim 13, wherein the immunosuppressant is a low-dose TBI.

15. The immunosuppressant is an anti-CD4 antibody, an anti-CD8 antibody, or a combination thereof. The method according to any one of claims 1 to 12.

16. The immunosuppressant is cyclophosphamide, according to any one of claims 1 to 12. method.

17. The patient, at least 24 hours before the transplant and / or at least 2 hours after the transplant The method according to any one of claims 1 to 16, wherein the patient does not receive an immunosuppressant for four hours.

18. The patient said to have at least 48 hours prior to the transplant and / or at least 4 hours after the transplant. The method according to any one of claims 1 to 16, wherein the patient does not receive an immunosuppressant for 8 hours.

19. The patient said to have at least 72 hours prior to the transplant and / or at least 7 hours after the transplant. The method according to any one of claims 1 to 16, wherein the patient does not receive an immunosuppressant for two hours.

20. The patient said that at least 96 hours before the transplant and / or at least 9 hours after the transplant The method according to any one of claims 1 to 16, wherein the patient does not receive immunosuppressants for 6 hours.

21. The patient for at least 7 days prior to the transplant and / or at least 7 days after the transplant. The method according to any one of claims 1 to 16, wherein the patient does not receive immunosuppressants during that time.

22. The patient had at least 14 days prior to the transplant and / or at least 1 day after the transplant. The method according to any one of claims 1 to 16, wherein the patient does not receive immunosuppressants for four days.

23. The patient, for at least one month prior to the transplant and / or at least one month after the transplant The method according to any one of claims 1 to 16, wherein the patient does not receive immunosuppressants for several months.

24. The effective amount of the CD45-targeting portion bound to the toxin is at least 80%, 85% To establish 90%, 95%, 97%, 99%, or 100% donor chimerism, it takes ten The method according to any one of claims 1 to 23, which is a certain amount.

25. Donor chimerism occurs at least 6, 7, 8, 9 weeks, or 1 week after transplantation. The method according to claim 24, which is evaluated in 0 weeks.

26. The donor chimerism is whole peripheral chimerism, as per claim 24 or 25. Law.

27. The method according to claim 24 or 25, wherein the donor chimerism is bone marrow chimerism. 。

28. The donor chimerism described above is T-cell chimerism, according to claim 24 or 25. Law.

29. The method according to claim 24 or 25, wherein the donor chimerism is B-cell chimerism. Law.

30. The effective amount of the CD45-targeted portion bound to the toxin is administered to the patient in a single dose. The method according to any one of claims 1 to 29.

31. The effective amount of the CD45-targeted portion bound to the toxin is administered to the patient in two doses. The method according to any one of claims 1 to 29.

32. The effective amount of the CD45-targeting portion bound to the toxin is used two or more times (for example, 2, 3 The patient is administered the drug in 4, 5, 6, 7, 8, 9, 10 or more doses. The method described in any one of the requests 1 to 29.

33. The aforementioned transplantation is performed so that the concentration of the CD45-targeted portion bound to the toxin is obtained from the patient's blood. The method according to any one of claims 1 to 32, performed on the patient after qualitative disappearance. 。

34. The hematopoietic stem cells or their offspring have been in the patient for at least two days since the transplantation of the hematopoietic stem cells into the patient. Even if, the functional potential of hematopoietic stem cells is maintained, as described in any one of claims 1 to 33. Method of loading.

35. The aforementioned allogeneic hematopoietic stem cells or their offspring, after transplantation of the hematopoietic stem cells into the patient, Claims 1 to 34, which enable localization in the tissue and / or re-establishment of hematopoiesis The method described in any one of the items.

36. The hematopoietic stem cells, upon transplantation into the patient, contain megakaryocytes, thrombus cells, platelets, red blood cells, mastocytes, and more. Cells, myeloblasts, basophils, neutrophils, eosinophils, microglia, granulocytes, monocytes, osteoclasts, Antigen-presenting cells, macrophages, dendritic cells, natural killer cells, T lymphocytes, and B Any of claims 1 to 35, which brings about the recovery of a cell population selected from a group consisting of lymphocytes. The method described in item 1.

37. The method according to any one of claims 1 to 36, wherein the patient is suffering from stem cell dysfunction.

38. The aforementioned patient has abnormal hemoglobinosis, autoimmune disorder, myelodysplastic disorder, immunodeficiency disorder, The method according to any one of claims 1 to 36, wherein the person is suffering from a metabolic disorder.

39. The method according to any one of claims 1 to 36, wherein the patient has cancer.

40. When the anti-CD45 ADC was measured by biolayer interferometry (BLI), １×１０ －２ ～１×１０ －３ 、１×１０ －３ ～１×１０ －４ 、１×１０ －５ ～１×１０ － ６ 、 1 × 10 －６ ~ 1 × 10 －７ or 1 × 10 －７ ~ 1 × 10 －８ dissociation rate (K ＯＦ Ｆ The method according to any one of claims 4 to 40, comprising an antibody having ).

41. The aforementioned anti-CD45 ADC was determined by a biolayer interference (BLI) assay. When this happens, the levels are approximately 100 nM or less, approximately 90 nM or less, approximately 80 nM or less, approximately 70 nM or less, and approximately 60 nM or less, about 50 nM or less, about 40 nM or less, about 30 nM or less, about 20 nM or less, about 10 nM or less, about 8 nM or less, about 6 nM or less, about 4 nM or less, about 2 nM or less, about 1 nM or less K Ｄ The method according to any one of claims 4 to 40, comprising an antibody that binds to CD45.

42. The anti-CD45 ADC includes a humanized anti-CD45 antibody or its antigen-binding moiety. The method described in any one of the requests 4 to 41.

43. The claim states that the anti-CD45 ADC comprises a human anti-CD45 antibody or its antigen-binding moiety. The method described in any one of items 4 to 41.

44. The aforementioned anti-CD45 ADC is an anti-CD45 antibody or its antigen-binding portion as described in Table 5. The method according to any one of claims 4 to 43, including the method described in any one of claims 4 to 43.

45. The anti-CD45 ADC comprises an intact anti-CD45 antibody, according to any of claims 4 to 44. The method described in item 1.

46. The anti-CD45 ADC comprises an IgG antibody, as described in any one of claims 4 to 45. The method.

47. The IgG is of the IgG1 isotype, IgG2 isotype, and IgG3 isotype. The method according to claim 46, wherein the IgG4 isotype is also present.

48. The aforementioned anti-CD45 ADC is conjugated to anti-CD4 via a linker into a cytotoxic agent. The method according to any one of claims 4 to 47, comprising an antibody or its antigen-binding moiety.

49. The method according to claim 48, wherein the cytotoxin is an RNA polymerase inhibitor.

50. The method according to claim 49, wherein the RNA polymerase inhibitor is amatoxin.

51. The method according to claim 50, wherein the amatoxin is amanitin.

52. The amatoxins mentioned above are α-amanitin, β-amanitin, γ-amanitin, ε-amanitin From tin, amanin, amaninamide, amanulin, amanuric acid, and proamanulin The method according to claim 50, selected from the group.

53. The method according to claim 48, wherein the cytotoxin is pyrrolobenzodiazepine (PBD).

54. The aforementioned cytotoxins include Pseudomonas exotoxin A, debuganin, diphtheria toxin, saporin, Mytansin, meitansinoids, auristatin, anthracycline, calicheamis Irinotecan, SN-38, Duocalmycin, Pyrrolobenzodiazepine, Pyrrolo Benzodiazepine dimers, indolinobenzodiazepines, indolinobenzodiazepine dimers Claim 4, selected from the group consisting of a mer and an indolinobenzodiazepine pseudodimer. The method described in 8.

55. The method according to claim 54, wherein the auristatin is MMAE or MMAF.

56. The antibody conjugates the toxin via a cysteine ​​residue in the Fc domain of the antibody. The method according to any one of claims 48 to 55, which is gated.

57. The cysteine ​​residue is introduced via amino acid substitution in the Fc domain of the antibody. The method according to claim 56.

58. The method according to claim 57, wherein the amino acid substitution is S239C or D265C.

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