Compositions and methods for depletion of diseased hematopoietic stem cells - Patents.com

Abstract

Provided herein are compositions and methods related to depleting diseased hematopoietic stem cells (HSCs) using anti-c-kit antibody.The compositions and methods described herein can be used to treat subjects who need to deplete diseased HSCs due to various diseases or disorders, such as myelodysplastic syndrome and acute myeloid leukemia.

Description

[Technical field]

[0001] (CROSS REFERENCE TO RELATED APPLICATIONS) This application claims priority to U.S. Provisional Patent Application No. 63 / 314,917, filed February 28, 2022, which is incorporated by reference in its entirety.

[0002] (Reference to Electronic Sequence Listing) The contents of the electronic sequence listing (JATH_008_01WO_SeqList_ST26.xml, size: 43,722 bytes, and creation date: February 28, 2023) are incorporated herein by reference in their entirety.

[0003] (Technical field) The present disclosure relates to compositions and methods for depletion of hematopoietic stem cells (HSCs). The compositions and methods described herein can be used to treat patients who require depletion of diseased HSCs for a variety of different diseases or disorders, including, but not limited to, myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML). [Background technology]

[0004] Myelodysplastic syndromes (MDS) are hematological malignancies characterized by the presence of defective hematopoietic stem cells (HSCs). Current treatments for MDS include hematopoietic stimulators to temporarily resolve associated symptoms. These may include chronic blood transfusions, erythropoietin stimulators for red blood cell production, or various chemotherapies. However, there are no existing treatments that specifically target diseased HSCs compared to healthy HSCs. Thus, there is a need for innovative agents and regimens that selectively target diseased HSCs and allow healthy HSCs to undergo normal hematopoiesis in the bone marrow. The present disclosure meets this need by providing agents and regimens for selective depletion of diseased HSCs in MDS, acute myeloid leukemia (AML), and other disorders. Summary of the Invention

[0005] The present disclosure provides, inter alia, a method for depleting diseased hematopoietic stem cells (HSCs) relative to healthy HSCs by administering an anti-c-kit antibody to a patient. In certain embodiments, diseased HSCs are selectively depleted relative to healthy HSCs.

[0006] In one aspect, the disclosure provides a method of depleting endogenous diseased hematopoietic stem cells (HSCs) in a subject, comprising administering to the subject an anti-c-kit antibody, which results in an increase in the proportion of healthy hematopoietic stem cells (HSCs) present in the bone marrow of the subject, e.g., immediately after the end of treatment or about 1 day, 2 days, 3 days, 4 days, 5 days, 6 days, 10 days, 1 week, 2 weeks, 1 month, 2 months, 4 months, or 6 months after the end of treatment. In certain embodiments, diseased HSCs are selectively depleted. In certain embodiments, the anti-c-kit antibody is a monoclonal antibody selected from the group consisting of SR-1, JSP191, MGTA-117, FSI-174, CDX0158, CDX0159, 8D7, K45, 104D2, CK6, AB249, YB5.B8, AF-2-1, AF11, AF12, AF112, AF-3, AF-1-1, NF, NF-2-1, NF11, NF12, NF112, NF-3, HF11, HF12, and HF112, optionally comprising one or more complementarity determining regions (CDRs) present in JSP191, CDX0158, CDX0159, or FSI-174. In certain embodiments, the anti-c-kit antibody comprises one or more complementarity determining regions (CDRs) present in a humanized version of a monoclonal antibody selected from the group consisting of ACK2, ACK4, 2B8, 3C11, MR-1, and CD122. In certain embodiments, the anti-c-kit antibody comprises the CDRs of an antibody that blocks the binding of stem cell factor (SCF) to the stem cell factor receptor (CD117), optionally, the antibody is JSP191.

[0007] In certain embodiments, the subject is administered about 0.01 mg / kg to about 20 mg / kg of anti-c-kit antibody, optionally about 0.1 mg / kg to about 10 mg / kg of anti-c-kit antibody. In certain embodiments of any of the methods disclosed herein, the subject is administered about 0.6 mg / kg of anti-c-kit antibody. In certain embodiments, the subject is administered about 2 mg / kg of anti-c-kit antibody. In certain embodiments, the anti-c-kit antibody is delivered systemically to the subject, optionally the anti-c-kit antibody is delivered intravenously or subcutaneously. In certain embodiments, the anti-c-kit antibody is delivered locally to the subject, optionally the anti-c-kit antibody is delivered directly to the bone marrow. In certain embodiments, the subject is administered two or more doses of anti-c-kit antibody. In certain embodiments, one of the two or more doses is administered about every week, about every 2 weeks, about every 3 weeks, about every 4 weeks, about every 5 weeks, about every 6 weeks, about every 7 weeks, about every 8 weeks, about every 9 weeks, about every 10 weeks, about every 11 weeks, or about every 12 weeks, optionally with varying time intervals between each dose. In certain embodiments, the subject's bone marrow comprises at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 6%, at least 7%, at least 8%, at least 9%, at least 10%, at least 15%, or at least 20% healthy HSCs prior to treatment.

[0008] In certain embodiments of any of the methods disclosed herein, the method is used to treat a disease or disorder selected from the group consisting of acute myeloid leukemia (AML), chronic myeloid leukemia (CML), chronic myelomonocytic leukemia (CMML), acute lymphoblastic leukemia (ALL), Hodgkin's lymphoma, clonal hematopoiesis of indeterminate potential (CHIP), clonal cytopenia of undetermined significance (CCUS), myelodysplastic syndrome (MDS), idiopathic cytopenia of undetermined significance (ICUS), and myeloproliferative neoplasms (MPN). In certain embodiments, the disease or disorder is MDS. In certain embodiments, the MDS is lower risk MDS. In certain embodiments, the subject has failed erythropoietin stimulating agent (ESA) therapy, optionally the ESA therapy comprises erythropoietin and / or darbepoietin. In certain embodiments, the method has reduced side effects compared to treatment with a chemotherapeutic agent, optionally the chemotherapeutic agent comprises azacitidine, lenalidomide, decitabine, and / or luspatercept. In certain embodiments, the reduced side effects include reduced risk of hematologic toxicity and reduced risk of death from infection. In certain embodiments, the method results in increasing the proportion of normal HSCs in the bone marrow to at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 99%, or at least 100% of all HSCs, and optionally the method results in long-term predominance of healthy hematopoietic stem cells (HSCs) in the bone marrow of the subject. In certain embodiments, the subject will not require or undergo hematopoietic cell transplantation (HCT) within at least 4 months, at least 6 months, at least 1 year, or at least 2 years after administration of the anti-c-kit antibody.

[0009] In another aspect, the disclosure provides a method of treating a disease or disorder in a subject, optionally myelodysplastic syndrome (MDS) or acute myeloid leukemia (AML), comprising administering to the subject an anti-c-kit antibody in a dose effective to achieve long-term predominance of healthy hematopoietic stem cells (HSCs) in the bone marrow of the subject. In certain embodiments, the disease or disorder is myelodysplastic syndrome (MDS). In certain embodiments, the MDS is lower-risk MDS. In certain embodiments, the disease or disorder is acute myeloid leukemia (AML). In certain embodiments, the subject has failed erythropoietin stimulating agent (ESA) therapy, optionally, the ESA therapy includes erythropoietin and / or darbepoietin. In certain embodiments, the method selectively depletes diseased HSCs compared to healthy HSCs. In certain embodiments, the anti-c-kit antibody comprises one or more complementarity determining regions (CDRs) present in a monoclonal antibody selected from the group consisting of SR-1, JSP191, MGTA-117, FSI-174, CDX-0159, 8D7, K45, 104D2, CK6, AB249, YB5.B8, AF-2-1, AF11, AF12, AF112, AF-3, AF-1-1, NF, NF-2-1, NF11, NF12, NF112, NF-3, HF11, HF12, and HF112. In certain embodiments, the anti-c-kit antibody comprises one or more complementarity determining regions (CDRs) present in a humanized version of a monoclonal antibody selected from the group consisting of ACK2, ACK4, 2B8, 3C11, MR-1, and CD122. In certain embodiments, the anti-c-kit antibody comprises the CDRs of an antibody that blocks the binding of stem cell factor (SCF) to the stem cell factor receptor (CD117), optionally, the antibody is JSP191.

[0010] In certain embodiments of the methods disclosed herein, the subject is administered about 0.01 mg / kg to about 10 mg / kg of anti-c-kit antibody, optionally about 0.1 mg / kg to about 10 mg / kg of anti-c-kit antibody. In certain embodiments, the subject is administered about 0.6 mg / kg of anti-c-kit antibody. In certain embodiments, the subject is administered about 2 mg / kg of anti-c-kit antibody. In certain embodiments, the anti-c-kit antibody is delivered systemically to the subject, optionally the anti-c-kit antibody is delivered intravenously or subcutaneously. In certain embodiments, the anti-c-kit antibody is delivered locally to the subject, optionally the anti-c-kit antibody is delivered directly to the bone marrow. In certain embodiments, the subject is administered two or more doses of anti-c-kit antibody. In certain embodiments, one of the two or more doses is administered every week, every 2 weeks, every 3 weeks, every 4 weeks, every 5 weeks, every 6 weeks, every 7 weeks, every 8 weeks, every 9 weeks, every 10 weeks, every 11 weeks, or every 12 weeks, optionally with varying time intervals between doses. In certain embodiments, the subject's bone marrow comprises at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 6%, at least 7%, at least 8%, at least 9%, at least 10%, at least 15%, or at least 20% healthy HSCs prior to treatment.

[0011] In certain embodiments, the methods disclosed herein have reduced side effects compared to treatment with chemotherapeutic agents, optionally the chemotherapeutic agents include azacitidine, lenalidomide, decitabine, and / or luspatercept. In certain embodiments, the reduced side effects are reduced risk of hematologic toxicity and reduced risk of death from infection. In certain embodiments, the methods result in increasing the percentage of normal HSCs in the bone marrow to at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 99%, or at least 100% of all HSCs. In certain embodiments, the subject does not undergo hematopoietic cell transplantation (HCT) within at least 4 months, at least 6 months, at least 1 year, or at least 2 years after administration of the anti-c-kit antibody. In certain embodiments, the anti-c-kit antibody, e.g., JSP191, is administered in combination with an anti-CD47 antibody, e.g., for the treatment of AML. In certain embodiments, the anti-c-kit antibody is administered in combination with a chemotherapeutic agent. In certain embodiments, the chemotherapeutic agent is azacitidine, for example, for the treatment of MDS. [Brief description of the drawings]

[0012] [Figure 1] FIG. 1 is a schematic diagram of the effect of administering an exemplary anti-c-kit antibody, e.g., JSP191, on hematopoiesis of normal and diseased cells in patients with hematological malignancies, e.g., myelodysplastic syndrome (MDS) or acute myeloid leukemia (AML). [Diagram 2] Figure 2 is a bar graph of the percentage of depletion of CD34+CD45RA-CD117+ cells measured 10 to 14 days after treatment in nine myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) patients treated with a single dose of anti-c-kit antibody. [Diagram 3] FIG. 3 is a flow diagram showing an example of a target patient population, eg, lower risk myelodysplastic syndrome (MDS) patients, for anti-c-kit antibody therapy to deplete diseased hematopoietic stem cells (HSC). DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0013] The present disclosure provides treatment for myelodysplastic syndrome (MDS) MDS, acute myeloid leukemia (AML), and other patients with an effective dose of anti-c-kit antibody to deplete diseased hematopoietic stem cells (HSCs) and increase the proportion of healthy HSCs in the bone marrow. In certain embodiments, diseased HSCs are selectively depleted. In certain embodiments, anti-c-kit antibody treatment described herein reduces diseased HSCs to a greater extent than healthy HSCs, thus allowing healthy cells to re-establish dominance in the bone marrow, thereby increasing the number of healthy HSCs and reducing the need for hematopoietic stem cell transplantation (HCT), total body irradiation (TBI), and / or chemotherapy. In certain embodiments, the method achieves long-term dominance of healthy hematopoietic stem cells (HSCs) in the bone marrow of a subject, for example, for at least 6 months, at least 1 year, at least 2 years, at least 5 years, or at least 10 years.

[0014] The treatments described herein have been shown to be effective in treating myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML) (see Examples). However, these treatments are not intended to be limited to only these disease populations. The compositions and methods disclosed herein may be used to treat other disorders in which selective depletion of diseased HSCs is required. Related diseases or disorders include, but are not limited to, acute myeloid leukemia (AML), chronic myeloid leukemia (CML), chronic myelomonocytic leukemia (CMML), acute lymphoblastic leukemia (ALL), Hodgkin's lymphoma, non-Hodgkin's lymphoma, clonal hematopoiesis of indeterminate potential (CHIP), clonal cytopenia of undetermined significance (CCUS), myelodysplastic syndromes (MDS), idiopathic cytopenia of undetermined significance (ICUS), and myeloproliferative neoplasms (MPN).

[0015] It is to be understood that the invention is not limited to the particular methodology, products, devices, and agents described, and that such methods, devices, and formulations may, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to limit the scope of the invention, which is limited only by the appended claims.

[0016] It should be noted that as used in this specification and the appended claims, the singular forms "a," "and," and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "drug candidates" refers to one or a mixture of such candidates, reference to "a method" includes references to equivalent steps and methods known to those skilled in the art, and so forth.

[0017] Unless otherwise defined herein, scientific and technical terms used in this application shall have the meanings commonly understood by those skilled in the art. Generally, the nomenclature used in connection with, and techniques of, chemistry, molecular biology, cell and cancer biology, immunology, microbiology, pharmacology, and protein and nucleic acid chemistry described herein are those well known and commonly used in the art.

[0018] Where a range of values ​​is provided, unless the context clearly indicates otherwise, it is understood that each intervening value, to the tenth of the unit of the lower limit between the upper and lower limits of that range, and any other stated or intervening value in that stated range, is encompassed within the invention. The upper and lower limits of these smaller ranges may independently be included in the smaller ranges, and are also encompassed within the invention, subject to any specifically excluded limits in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the invention.

[0019] As used herein, "antibody" includes reference to an immunoglobulin molecule that is immunologically reactive with a particular antigen, and includes both polyclonal and monoclonal antibodies. The term also includes genetically engineered forms such as humanized antibodies, chimeric antibodies (e.g., humanized murine antibodies), and heteroconjugate antibodies. The term "antibody" also includes fragments (e.g., Fab', F(ab')) that have antigen-binding capability. 2 The term includes antigen-binding forms of antibodies, including Fab, Fv, and rIgG. The term also refers to recombinant single chain Fv fragments (scFv). The term antibody also includes bivalent or bispecific molecules, diabodies, triabodies, and tetrabodies.

[0020] A "humanized antibody" is an immunoglobulin molecule that contains minimal sequence derived from a non-human immunoglobulin. Humanized antibodies include human immunoglobulins (recipient antibody) in which residues from a complementarity determining region (CDR) of the recipient are replaced by residues from a CDR of a non-human species (donor antibody) such as mouse, rat, or rabbit having the desired specificity, affinity, and capacity. In some instances, Fv framework residues of the human immunoglobulin are replaced by corresponding non-human residues. Humanized antibodies may also comprise residues that are found neither in the recipient antibody nor in the imported CDR or framework sequences. In general, a humanized antibody will comprise substantially all of at least one, and typically two, variable domains, with all or substantially all of the CDR regions corresponding to those of a non-human immunoglobulin and all or substantially all of the framework (FR) regions being of human immunoglobulin consensus sequences. A humanized antibody will also optimally comprise at least a portion of an immunoglobulin constant region (Fc), typically of a human immunoglobulin.

[0021] Selection of antibodies for endogenous stem cell ablation can be based on a variety of criteria, including selectivity, affinity, cytotoxicity, etc. The phrase "specifically (or selectively) binds" to an antibody when referring to a protein or peptide refers to a binding reaction that determines the presence of the protein in a heterogeneous population of proteins and / or other biologics. Thus, under specified immunoassay conditions, a specified antibody binds to a particular protein sequence at least twice background binding, and more typically 10-100 times more than background binding.

[0022] Monoclonal antibodies may be prepared using the hybridoma method. In the hybridoma method, a suitable host animal is typically immunized with an immunizing agent to induce lymphocytes that produce or have the ability to produce antibodies that specifically bind to the immunizing agent. Alternatively, lymphocytes may be immunized in vitro. The lymphocytes are then fused with an immortalized cell line using a suitable fusing agent, such as polyethylene glycol, to form hybridoma cells.

[0023] Human antibodies can be produced using various techniques known in the art, including phage display libraries.Similarly, human antibodies can be produced by introducing human immunoglobulin loci into transgenic animals, such as mice in which endogenous immunoglobulin genes are partially or completely inactivated.Upon challenge, human antibody production is observed, which closely resembles that seen in humans in all respects, including gene rearrangement, assembly, and antibody repertoire.

[0024] Antibodies also exist as several well-characterized fragments produced by digestion with various peptidases. Pepsin digests antibodies below the disulfide bond in the hinge region to produce F(ab)'2, a dimer of Fab, itself a light chain linked to VH-CH1 by a disulfide bond. F(ab)'2 can be reduced under mild conditions to break the disulfide bond in the hinge region, thereby converting the F(ab)'2 dimer into a Fab' monomer. A Fab' monomer is essentially a Fab with a portion of the hinge region. Although various antibody fragments are defined with respect to the digestion of an intact antibody, one skilled in the art will understand that such fragments can be synthesized de novo, either chemically or by using recombinant DNA methodology. Thus, the term antibody, as used herein, also includes antibody fragments produced by the modification of whole antibodies, or antibody fragments synthesized de novo using recombinant DNA methodology (e.g., single chain Fv), or antibody fragments identified using a phage display library.

[0025] The selectivity of a particular antibody is typically determined by the ability of one antibody to competitively inhibit the binding of a second antibody to an antigen, or by the ability of an antibody to cross-react with multiple epitopes. Any of several competitive binding assays can be used to measure the competition between two antibodies for the same antigen, or between two antigens for one antibody. An exemplary assay is the BIACORE™ assay. Briefly, in these assays, binding sites can be structurally mapped by testing the ability of interactors, e.g., different antibodies, to inhibit the binding of another. By injecting two successive antibody or antigen samples at sufficient concentrations, pairs of antibodies competing for the same binding epitope can be identified. The antibody samples should have a significant saturation potential with each injection. The net binding of the second antibody injection suggests binding epitope analysis. Two response levels can be used to describe the boundary between complete competition and non-competitive binding with distinct epitopes. The relative amount of binding response of the second antibody injection to binding of the same and distinct binding epitopes determines the degree of epitope overlap.

[0026] The term "polynucleotide" refers to a polymeric form of nucleotides of any length, including deoxyribonucleotides or ribonucleotides, or analogs or mixtures thereof. A polynucleotide may contain modified nucleotides, such as methylated nucleotides and nucleotide or nucleoside analogs, and may be interrupted by non-nucleotide components. If present, modifications to the nucleotide structure may be added before or after assembly of the polymer. As used herein, the term polynucleotide includes, but is not limited to, double-stranded and single-stranded molecules, and mixtures thereof. Unless otherwise specified or required, any embodiment of the invention described herein that is a polynucleotide encompasses both the double-stranded form and each of the two complementary single-stranded forms that are known or predicted to constitute a double-stranded form, as RNA or DNA, or mixtures thereof.

[0027] As used herein, the terms "polypeptide," "peptide," and "protein" refer to polymers of amino acids of any length. The terms also include amino acid polymers that have been modified to include, for example, disulfide bond formation, glycosylation, lipidation, phosphorylation, or conjugation with a labeling component.

[0028] As used herein, the terms "identity" and "identical" refer to the percentage of perfectly matching residues in an alignment of a sequence of interest to a reference sequence, such as an alignment generated by the BLAST algorithm, for a polypeptide or polynucleotide sequence of interest. Unless otherwise specified, identity is calculated over the full length of the reference sequence. Thus, when a reference sequence (as a query sequence) is aligned to a sequence of interest (as a subject sequence), if at least x% (truncated) of the residues in the subject sequence align as perfect matches to the corresponding residues in the query sequence, the sequence of interest "shares at least x% identity" with the reference sequence, with the denominator being the full length of the reference sequence plus the length of any gaps inserted in the reference sequence by the alignment of the sequence of interest to the reference sequence. If the subject sequence has variable positions (e.g., residues designated by X), the alignment to any residue in the query sequence is counted as a match. Sequence similarity (i.e., identity) can be determined in many different ways. To determine sequence identity, sequences can be aligned using methods and computer programs, including BLAST, available on the World Wide Web at ncbi.nlm.nih.gov / BLAST / . Sequence alignment can be performed using the NCBI Blast service (BLAST+ version 2.12.0) or another program that produces the same results. Unless otherwise specified, sequence identity is determined using the BLAST algorithm (e.g., bl2seq) with default parameters.

[0029] As used herein, the term "native" or "wild type" refers to a nucleotide sequence, e.g., a gene, or a gene product, e.g., an RNA or a polypeptide, present in a wild-type cell, tissue, organ, or organism. As used herein, the term "variant" refers to a variant of a reference polynucleotide or polypeptide sequence (e.g., a naturally occurring polynucleotide or polypeptide sequence), i.e., having less than 100% sequence identity with the reference polynucleotide or polypeptide sequence. In other words, a variant contains at least one amino acid difference (e.g., an amino acid substitution, an amino acid insertion, an amino acid deletion) with respect to a reference polynucleotide sequence, e.g., a naturally occurring polynucleotide or polypeptide sequence. For example, a variant may be a polynucleotide having 50% or more, 60% or more, or 70% or more sequence identity with a full-length naturally occurring polynucleotide sequence, e.g., 75% or 80% or more, 85%, 90%, or 95% or more, e.g., 98% or 99% or more identity with a full-length naturally occurring polynucleotide sequence. As another example, a variant may be a polypeptide having 70% or more sequence identity to a full-length native polypeptide sequence, e.g., 75% or 80% or more, 85%, 90%, or 95% or more, e.g., 98% or 99% or more identity to a full-length native polypeptide sequence. Variants may also include variant fragments of a reference, e.g., a native sequence that shares 70% or more sequence identity with a reference fragment, e.g., a variant fragment of a native sequence that shares 75% or 80% or more identity with the native sequence, e.g., 85%, 90%, or 95% or more identity with the native sequence, e.g., 98% or 99% identity.

[0030] As used herein, the term "stem cell" refers to a mammalian cell that has both the ability to self-renew and to generate differentiated progeny (see Morrison et al. (1997) Cell 88:287-298). Endogenous stem cells can be characterized by the presence of markers associated with specific epitopes. As used herein, the term "hematopoietic stem cell (HSC)" refers to a multipotent cell that resides in the bone marrow (BM) and is responsible for the lifelong production of mature blood cells.

[0031] The terms "treatment", "treating" and the like are generally used herein to mean obtaining a desired pharmacological and / or physiological effect. The effect may be prophylactic in that it completely or partially prevents a disease or its symptoms, for example, it may reduce the likelihood that a disease or its symptoms will occur in a subject, and / or it may be therapeutic in that it partially or completely cures the disease and / or the deleterious effects caused by the disease. As used herein, "treatment" covers any treatment of a mammalian disease, including (a) inhibiting the disease, i.e., preventing its development, or (b) relieving the disease, i.e., causing regression of the disease. Therapeutic agents may be administered before, during, or after the onset of the disease or injury. Treatment of ongoing disease is of particular interest, where the treatment stabilizes or reduces undesirable clinical symptoms in the patient. Such treatment is desirably performed before complete loss of function of the affected tissue. The subject therapy may be administered before or during the symptomatic stage of the disease, or in some cases, after the symptomatic stage of the disease.

[0032] The terms "individual," "host," "subject," and "patient" are used interchangeably herein and refer to mammals, including, but not limited to, humans and non-human primates, including apes and humans, mammalian sports animals (e.g., horses), mammalian farm animals (e.g., sheep, goats, etc.), mammalian pets (dogs, cats, etc.), and rodents (e.g., mice, rats, etc.).

[0033] As used herein, the term "substantially" means a significant or large amount or degree. For example, "substantially" increasing may mean at least 2-fold, at least 3-fold, at least 4-fold, at least 5-fold, or at least 10-fold increasing, and "substantially" decreasing may mean at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% decreasing. In the following description, numerous specific details are set forth in order to provide a more complete understanding of the present invention. However, it will be apparent to one of ordinary skill in the art that the present invention may be practiced without one or more of these specific details. In other instances, well-known features and procedures well known to those of ordinary skill in the art are not described in order to avoid obscuring the invention.

[0034] Generally, conventional methods of protein synthesis, recombinant cell culture and protein isolation, and recombinant DNA technology are used in the present invention, which are within the skill of the person skilled in the art.These techniques are fully described in the literature.See, for example, Maniatis, Fritsch & Sambrook, Molecular Cloning: A Laboratory Manual (1982); Sambrook, Russell and Sambrook, Molecular Cloning: A Laboratory Manual (2001); Harlow, Lane and Harlow, Using Antibodies: A Laboratory Manual: Portable Protocol No. I, Cold Spring Harbor Laboratory (1998), and Harlow and Lane, Antibodies: A Laboratory Manual, Cold Spring Harbor Laboratory; (1988).

[0035] I. Depletion of diseased HSCs In some embodiments, the treatment described herein depletes diseased hematopoietic stem cells (HSCs) from the bone marrow of a subject. In some embodiments, "disease HSCs" are HSCs that do not differentiate or have a significantly reduced differentiation capacity, do not divide or have a significantly reduced cell division capacity, do not produce functional blood cells or produce a significantly reduced amount of functional blood cells, and / or are otherwise classified as hematological malignancies. In certain embodiments, a significant difference is a statistically significant difference with a p-value of 5% or less. In some embodiments, the present disclosure provides methods for selectively depleting endogenous diseased hematopoietic stem cells (HSCs) in a subject and / or methods for increasing the relative proportion of healthy HSCs by administering anti-c-kit antibodies.

[0036] In some embodiments, the treatment described herein depletes diseased HSCs in a subject. In some embodiments, depleting diseased HSCs refers to depleting diseased HSCs in an amount of about 2%, about 5%, about 10%, about 20%, about 30%, about 40%, about 50%, about 60%, about 70%, about 80%, about 90%, about 95%, about 98%, about 99%, or about 100% of the total diseased HSCs in a subject.

[0037] In some embodiments, the treatments described herein selectively deplete diseased HSCs in a subject. In certain embodiments, selective depletion of diseased HSCs by the disclosed methods results in depletion of a greater number or proportion of diseased HSCs compared to the number or proportion of non-diseased or healthy HSCs that are depleted.

[0038] In some embodiments, the treatments described herein result in an increase in the relative proportion of healthy HSCs in a subject. In some embodiments, the treatments described herein increase the relative proportion of healthy HSCs in a subject (compared to total HSCs) by about 50%, about 60%, about 60%, about 70%, about 80%, about 90%, about 100%, about 200%, about 300%, about 400%, about 500%, about 600%, about 700%, about 800%, about 900%, or about 1000%.

[0039] Anti-c-Kit antibody In some embodiments, the present disclosure provides methods for selectively depleting endogenous diseased hematopoietic stem cells (HSCs) in a subject and / or increasing the relative proportion of healthy HSCs by administering anti-c-kit antibodies. In some embodiments, the compositions and methods disclosed herein may be applicable to any anti-c-kit antibody that blocks stem cell factor (SCF) binding to the c-kit / CD117 protein of endogenous HSCs. In some embodiments, the antibody is a monoclonal anti-human c-kit antibody. In some embodiments, the antibody is an antigen-binding fragment of an antibody that blocks stem cell factor (SCF) binding.

[0040] Some antibodies contemplated by the present disclosure that specifically bind to human CD117 are known in the art and / or commercially available, including, but not limited to, JSP-191, SR1, 2B8, ACK2, YB5-B8, 57A5, and 104D2. In certain embodiments, the anti-c-kit antibody is selected from the group consisting of JSP191 (Jasper Therapeutics, Redwood City, CA), CDX-0158 (formerly KTN0158) or CDX-0159 (Celldex Therapeutics, Hampton, NJ), MGTA-117 (AB85) (Magenta Therapeutics, Cambridge, MA), CK6 (Maneta Therapeutics, Cambridge, MA), AB249 (Maneta Therapeutics, Cambridge, MA), and FSI-174 (Celldex, Foster City, CA). Magenta Therapeutics antibodies contemplated by this disclosure include, but are not limited to, those disclosed in US Patent Publication No. 20190153114, PCT Patent Publication No. WO2019084064, WO2020 / 219748, and WO2020 / 219770. FSI-174 antibody is disclosed in PCT Publication No. WO2020 / 112687 and US Patent Publication No. 20200165337. This disclosure includes, but is not limited to, any anti-c-kit antibody and / or CDR set disclosed in any of the patent applications disclosed herein (all of which are incorporated by reference in their entirety).

[0041] In certain embodiments, the anti-c-kit antibody binds to the extracellular region of CD117, i.e., amino acids 26 to 524. The sequence of this region is shown below: QPSVSPGEPSPPSIHPGKSDLIVRVGDEIRLLCTDPGFVKWTFEILDETNENKQNEWITEKAEATNTGKYTCTNKHGLSNSIYVFVRDPAKLFLVDRSLYGKEDNDTLVRCPLTDPEVTNYSLKGC QGKPLPKDLRFIPDPKAGIMIKSVKRAYHRLCLHCSVDQEGKSVLSEKFILKVRPAFKAVPVVSVSKASYLLREGEEFTVTCTIKDVSSSVYSTWKRENSQTKLQEKYNSWHHGDFNYERQATLTIS SARVNDSGVFMCYANNTFGSANVTTTLEVVDKGFINIFPMINTTVFVNDGENVDLIVEYEAFPKPEHQQWIYMNRTFTDKWEDYPKSENESNIRYVSELHLTRLKGTEGGTYTFLVSNSDVNAAIAFNVYVNTKPEILTYDRLVNGMLQCVAAGFPEPTIDWYFCPGTEQRCSASVLPVDVQTLNSSGPPFGKLVVQSSIDSSAFKHNGTVECKAYNDVGKTSAYFNFAFKGNNKEQIHPHTLFTP (SEQ ID NO: 1).

[0042] Exemplary anti-c-kit antibodies include, but are not limited to, SR-1, JSP191, 8D7, K45, 104D2, CK6, YB5.B8, AF-2-1, AF11, AF12, AF112, AF-3, AF-1-1, NF, NF-2-1, NF11, NF12, NF112, NF-3, HF11, HF12, and HF112. Several antibodies contemplated by the present disclosure that specifically bind to human CD117 are commercially available, including, but not limited to, SR1, 2B8, ACK2, YB5-B8, 57A5, 104D2. In certain embodiments, the anti-c-kit antibody is selected from the group consisting of JSP191, CDX-0159 (Celldex Therapeutics, Hampton, NJ), MGTA-117 (AB85) (Magenta Therapeutics, Cambridge, MA), CK6 (Magenta Therapeutics, Cambridge, MA), AB249 (Magenta Therapeutics, Cambridge, MA), and FSI-174 (Gilead, San Francisco, CA). Magenta Therapeutics antibodies are disclosed in US Patent Publication No. 20190153114. In certain embodiments, the antibody is one disclosed in any of U.S. Patent No. 7,915,391, U.S. Patent No. 8,436,150, or U.S. Patent No. 8,791,249. In certain embodiments, the antibody is one disclosed in U.S. Patent Application Publication No. 20200165337, or any of PCT Publication Nos. WO2020 / 112687, WO2020 / 219748, WO2020 / 219770, or WO2019 / 084064.

[0043] In certain embodiments, the antibody is a humanized form of SR1, a mouse anti-c-kit antibody described in U.S. Patent Nos. 5,919,911 and 5,489,516. A humanized form, JSP191, is disclosed in U.S. Patent Nos. 7,915,391, 8,436,150, and 8,791,249. JSP191 is a non-glycosylated IgG1 humanized antibody. JSP191 specifically binds to human CD117, a receptor for stem cell factor (SCF), expressed on the surface of hematopoietic stem and progenitor cells. JSP191 blocks SCF binding to CD117, disrupting stem cell factor (SCF) signaling and causing hematopoietic stem cell depletion. JSP191 is a heterotetramer consisting of two heavy chains of IgG1 subclass and two light chains of kappa subclass covalently linked via disulfide bonds. There are no N-linked glycans on JSP191 due to an intentional asparagine to glutamine substitution at heavy chain residue 297. The heavy and light chain sequences of JSP191 are disclosed as SEQ ID NO: 4 and SEQ ID NO: 2 from US8436150, respectively.

[0044] The heavy and light chain sequences of JSP191 are disclosed as SEQ ID NO: 4 and SEQ ID NO: 2 from U.S. Patent No. 8,436,150, respectively. The heavy and light chain sequences of JSP191 are as follows:

[0045] Heavy Chain: MDWTWRVFCLLAVAPGAHSQVQLVQSGAEVKKPGASVKVSCKASGYTFTSYNMHWVRQAPGQGLEWMGVIYSGNGDTSYNQKFKGRVTITADKSTSTAYMELSSLRSEDTAVYYCARE RDTRFGNWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEP KSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYQSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK (SEQ ID NO: 2) and Light chain: MVLQTQVFISLLLWISGAYGDIVMTQSPDSLAVSLGERATINCRASESVDIYGNSFMHWYQQKPGQPPKLLIYLASNLESGVPDRFSGSGSGTDFTLTISSLQAEDVAVYYCQQNNEDPYTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC (sequence number 3).

[0046] In certain embodiments, the variable heavy domain of JSP191 comprises the following sequence: QVQLVQSGAEVKKPGASVKVSCKASGYTFTSYNMHWVRQAPGQGLEWMGVIYSGNGDTSYNQKFKGRVTITADKSTSTAYMELSSLRSEDTAVYYCARERDTRFGNWGQGTLVTVSS (sequence number 4).

[0047] In certain embodiments, the variable light chain domain of JSP191 comprises the following sequence: DIVMTQSPDSLAVSLGERATINCRASESVDIYGNSFMHWYQQKPGQPPKLLIYLASNLESGVPDRFSGSGSGTDFTLTISSLQAEDVAVYYCQQNNEDP YTFGGGTKVEIK (SEQ ID NO:5).

[0048] The CDRs present in JSP191 are as follows: VH CDR1 = YNMH (SEQ ID NO: 6), VH CDR2 = IYSGNGDTSYNQKFKG (SEQ ID NO: 7), VH CDR3 = ERDTRFGN (SEQ ID NO: 8), VL CDR1 = RASESVDIYGNSFMH (SEQ ID NO: 9), VL CDR2 = LASNLES (SEQ ID NO: 10), and VL CDR3 = QQNNEDPYT (SEQ ID NO: 11).

[0049] CDX-0159 is a humanized monoclonal antibody that specifically binds to the receptor tyrosine kinase KIT with high specificity and potently inhibits its activity. CDX-0159 is designed to block KIT activation by interfering with both SCF binding and KIT dimerization. CDX-0159 and other anti-c-kit antibodies are described in U.S. Pat. No. 10,781,267, and in certain embodiments, the anti-c-kit disclosed herein comprises any of the CDRs of the antibodies disclosed therein. In certain embodiments, the anti-c-kit antibody has the amino acid sequence: (i) DIVMTQSPSX K1 LSASVGDRVTITCKASQNVRTNVAWYQQKPGKAPKX K2 LIYSASYRYSGVPDRFX K3 GSGSGTDFTLTISSLQX K4 EDFAX K5 YX K6 A light chain variable region ("VL") comprising: K1 is an amino acid having an aromatic or aliphatic hydroxyl side chain, and XK2 is an amino acid having an aliphatic or aliphatic hydroxyl side chain, and X K3 is an amino acid having an aliphatic hydroxyl side chain, and X K4 is an amino acid having an aliphatic hydroxyl side chain or is P, and X K5 is an amino acid having a charged or acidic side chain, and X K6 a light chain region that is an amino acid having an aromatic side chain; and (ii) an amino acid sequence: QVQLVQSGAEX H1 KKPGASVKX H2 SCKASGYTFTDYYINAVVX H3 QAPGKGLEWIARIYPGSGNTYYNEKFKGRX H4 TX H5 TAX H6 KSTSTAYMX H7 LSSLRSEDX H8 A heavy chain variable region ("VH") comprising: AVYFCARGVYYFDYWGQGTTVTVSS (SEQ ID NO: 13); H1 is an amino acid having an aliphatic side chain, and X H2 is an amino acid having an aliphatic side chain, and X H3 is an amino acid with a polar or basic side chain, and X H4 is an amino acid having an aliphatic side chain, and X H5 is an amino acid having an aliphatic side chain, and X H6 is an amino acid with an acidic side chain, and X H7 is an amino acid having an acidic or amide derivative side chain, and X H8 and a heavy chain variable region, wherein: VH CDRs of the VH domain comprising QVQLKQSGAELVRPGASVKLSCKASGYTFTDYYINWVKQRPGQGLEWIARIYPG SGNTYYNEKFKGKATLTAEKSSSTAYMQLSSLTSEDSAVYFCARGVYYFDYWGQ GTTLTVSA (SEQ ID NO: 15); (ii) a VL CDR of a VL domain comprising the amino acid sequence DIVMTQSQKFMSTSVGDRVSVTCKASQNVRTNVAWYQQKPGQSPKALIYSASYRYSGVPDRFTGSGSGTDFTLTI SNVQSEDLADYFCQQYNSYPRTFGGGTKLEIKR (SEQ ID NO: 16), and an antigen-binding fragment thereof comprising the VL CDR of the VL domain comprising the amino acid sequence DIVMTQSQKFMSTSVGDRVSVTCKASQNVRTNVAWYQQKPGQSPKALIYSASYRYSGVPDRFTGSGSGTDFTLTI SNVQSEDLADYFCQQYNSYPRTFGGGTKLEIKR (SEQ ID NO: 16).

[0050] MGTA-117 (AB85) is an antibody that targets CD117 engineered for the transplant environment and conjugated to amanitin, and is being developed for patients undergoing immune reset via either autologous or allogeneic stem cell transplantation. MGTA-117 depletes hematopoietic stem and progenitor cells, and this antibody and other antibodies contemplated by the present disclosure are described in U.S. Patent Application Publication No. 20200407440 and / or PCT Application Publication No. WO2019084064. Epitope analysis of AB85 binding to CD177 is described in PCT Application Publication No. WO2020219770, which identified the following two epitopes within CD117: EKAEATNTGKYTCTNKHGLSNSIYVFVRDPA (SEQ ID NO: 17, amino acids 60 to 90), and RCPLTDPEVTNYSLKGCQGKP (SEQ ID NO: 18, amino acids 100-130).

[0051] The sequences of the variable heavy and variable light chains of AB85 are disclosed as SEQ ID NO:143 and SEQ ID NO:144, respectively, from PCT Application Publication WO2019084064.

[0052] The heavy chain variable region (VH) amino acid sequence of Ab85 is as follows: EVQLVQSGAEVKKPGESLKISCKGSGYSFT NYWIG WVRQMPGKGLEWMA IINPRDSDTRYRPSFQG QVTISADKSISTAYLQWSSLKASDTAMYYCAR HGRGYEGYEGAFDI WGQGTLVTVSS (sequence number 19).

[0053] The VH CDR amino acid sequences of AB85 are as follows: NYWIG (SEQ ID NO:20, VH CDR1), IINPRDSDTRYRPSFQG (SEQ ID NO:21, VH CDR2), and HGRGYEGYEGAFDI (SEQ ID NO:22, VH CDR3).

[0054] The light chain variable region (VL) amino acid sequence of AB85 is as follows: DIQMTQSPSSLSASVGDRVTITC RSSQGIRSDLG WYQQKPGKAPKLLIY DASNLET GVPSRFSGSGSGTDFLTISSLQPEDFATYYC QQANGFPLT FGGGTKVEIK (sequence number 23).

[0055] The VL CDR amino acid sequences of AB85 are as follows: RSSQGIRSDLG (SEQ ID NO:24, VL CDR1), DASNLET (SEQ ID NO:25, VL CDR2), and QQANGFPLT (SEQ ID NO:26, VL CDR3).

[0056] FSI-174 is an anti-cKIT antibody that is being developed in combination with 5F9 as a non-toxic transplant conditioning regimen and as a targeted hematological malignancy treatment. The sequence of FSI-174 is disclosed in PCT Publication No. 2020 / 112687, US Patent Publication No. 20200165337, and US Patent No. 11,041,022. In certain embodiments, the anti-c-kit antibody comprises three CDRs or variable heavy chain regions present in any of AH1, AH2, AH3, AH4, or AH5 disclosed therein, and / or three CDRs or variable heavy chain regions present in any of AL1 or AL2 disclosed therein.

[0057] In certain embodiments, the CDRs present in FSI-174 and related antibodies are as follows: VH CDR1 = SYNMH (SEQ ID NO: 27), VH CDR2 = VIYSGNGDTSY(A / N)QKF(K / Q)G (SEQ ID NO: 28), VH CDR3 = ERDTRFGN (SEQ ID NO: 29), VL CDR1 = RAS(D / E)SVDIYG(N / Q)SFMH (SEQ ID NO: 30), VL CDR2 = LASNLES (SEQ ID NO: 31), and VL CDR3 = QQNNEDPYT (SEQ ID NO: 32). A / N and the like indicate that the amino acid position can be either of two amino acids, in this example, A or N. In certain embodiments, the CDRs present in the heavy chain variable region are CDRs H1, H2 and H3 as defined by Kabat: H1=SYNMH (SEQ ID NO:27), H2=VIYSGNGDTSYAQKFKG (SEQ ID NO:33), H3=ERDTRFGN (SEQ ID NO:29), and the CDRs present in the light chain variable region are CDRs L1, L2, and L3 as defined by Kabat: L1=RASESVDIYGQSFMH (SEQ ID NO:34), L2=LASNLES (SEQ ID NO:31), and L3=QQNNEDPYT (SEQ ID NO:32), respectively, with the proviso that there are one, two or three CDR residue substitutions selected from N to A at heavy chain position 60, K to Q at heavy chain position 64, and N to Q at light chain position 30, and the positions are numbered according to Kabat. In certain embodiments, the antibody comprises any of the heavy chain variable region sequences (AH2, AH3, AH4) and / or light chain variable region sequences (AL2) provided below, or the CDRs therein shown underlined: AH2:QVQLVQSGAEVKKPGASVKVSCKASGYTFT SYMNH WVRQAPGQGLEWMG VIYSGNGDTSYAQKFKG RVTITADKSTSTAYMELSSLRSEDTAVYYCAR ERDTRFGN WGQGTLVTVSS (SEQ ID NO: 35) AH3:QVQLVQSGAEVKKPGASVKVSCKASGYTFT SYMNH WVRQAPGQGLEWMG VIYSGNGDTSYNQKFQGRVTITADKSTSTAYMELSSLRSEDTAVYYCAR ERDTRFGN WGQGTLVTVSS (SEQ ID NO: 36) AH4 QVQLVQSGAEVKKPGASVKVSCKASGYTFT SYMNH WVRQAPGQGLEWMG VIYSGNGDTSYAQKFQG RVTITADKSTSTAYMELSSLRSEDTAVYYCAR ERDTRFGN WGQGTLVTVSS (SEQ ID NO: 37) AL2:DIVMTQSPLSLPVTPGEPASISC RASESVDIYGQSFMH WYQQKPGQPPKLLIY LASNLES GVPDRFSGSGSGTDFTLKISRVEAEEDVGVYYC QQNNEDPYT FGGGTKVEIK (sequence number 38).

[0058] In certain embodiments, an anti-c-kit antibody comprises an amino acid sequence having at least 85%, at least 90%, at least 95%, at least 98%, at least 99% identity to the complete heavy and / or light chain of any of the antibodies disclosed herein, or a heavy or light chain disclosed herein, e.g., a JSP191 heavy or light chain. In certain embodiments, an anti-c-kit antibody comprises an amino acid sequence having at least 85%, at least 90%, at least 95%, at least 98%, at least 99% identity to the variable region of the heavy and / or light chain of any of the antibodies disclosed herein, or a variable region of the heavy or light chain disclosed herein, e.g., a JSP191 heavy or light chain. In certain embodiments, an anti-c-kit antibody comprises a heavy and / or light chain comprising one or more CDRs of an antibody disclosed herein, e.g., two, three, four, five, or six CDRs of an antibody disclosed herein, e.g., a JSP191 antibody. In certain embodiments, an anti-c-kit antibody comprises a heavy chain or a variable region thereof that comprises one, two, or three heavy chain CDRs disclosed herein, e.g., a JSP191 heavy chain. In certain embodiments, an anti-c-kit antibody comprises a light chain or a variable region thereof that comprises one, two, or three light chain CDRs disclosed herein, e.g., a JSP191 light chain.

[0059] CDX-0159 is a humanized monoclonal antibody that specifically binds to and potently inhibits the activity of the receptor tyrosine kinase KIT with high specificity. CDX-0159 is designed to block KIT activation by interfering with both SCF binding and KIT dimerization.

[0060] MGTA-117 is an antibody that targets CD117 and is conjugated to amanitin, engineered for the transplant environment, and is being developed for patients undergoing immune reset via either autologous or allogeneic stem cell transplantation. MGTA-117 depletes hematopoietic stem and progenitor cells, and this antibody and other antibodies contemplated by the present disclosure are described in US 20200407440.

[0061] FSI-174 is an anti-c-kit antibody being developed in combination with 5F9 as a non-toxic transplant conditioning regimen as well as a targeted hematological malignancy treatment.

[0062] In certain embodiments, the antibody may comprise one or more CDRs having at least 70%, 80%, 90%, 95% or 99% amino acid or nucleotide sequence identity with a CDR present in a humanized monoclonal antibody that binds to c-kit (e.g., an antibody derived from any of the murine antibodies SR1, ACK2, ACK4, 2B8, 3C11, MR-1, and CD122). In some embodiments, the antibody blocks the binding of stem cell factor (SCF) to the stem cell factor receptor (CD117). Exemplary embodiments of anti-c-kit antibodies that may be used include those described in JSP191, and WO2007127317A2 and US20200165337A1, both of which are incorporated herein in their entireties.

[0063] In certain embodiments, an anti-c-kit antibody comprises an amino acid sequence having at least 85%, at least 90%, at least 95%, at least 98%, at least 99% identity to the complete heavy and / or light chain of any of the antibodies disclosed herein, or a heavy or light chain disclosed herein, e.g., a JSP191 heavy or light chain. In certain embodiments, an anti-c-kit antibody comprises an amino acid sequence having at least 85%, at least 90%, at least 95%, at least 98%, at least 99% identity to the variable region of the heavy and / or light chain of any of the antibodies disclosed herein, or a variable region of the heavy or light chain disclosed herein, e.g., a JSP191 heavy or light chain. In certain embodiments, an anti-c-kit antibody comprises a heavy and / or light chain comprising one or more CDRs of an antibody disclosed herein, e.g., two, three, four, five, or six CDRs of an antibody disclosed herein, e.g., a JSP191 antibody. In certain embodiments, an anti-c-kit antibody comprises a heavy chain or a variable region thereof that comprises one, two, or three heavy chain CDRs disclosed herein, e.g., a JSP191 heavy chain. In certain embodiments, an anti-c-kit antibody comprises a light chain or a variable region thereof that comprises one, two, or three light chain CDRs disclosed herein, e.g., a JSP191 light chain.

[0064] II. Subject In certain embodiments, subjects selected for anti-c-kit antibody treatment as described herein have or retain at least some healthy hematopoietic cells (HSCs) to benefit from selective depletion of diseased HSCs. In some embodiments, subjects selected for anti-c-kit treatment as described herein have at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 6%, at least 7%, at least 8%, at least 9%, at least 10%, at least 15%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% healthy hematopoietic stem cells (HSCs) as a percentage of total HSCs in bone marrow before treatment. In certain embodiments, the percentage of healthy HSCs increases over the course of treatment.

[0065] In some embodiments, the percentage of healthy hematopoietic stem cells in a subject prior to administration of an anti-c-kit therapy described herein is assessed by bone marrow biopsy, blood count, or blood smear.

[0066] The treatments described herein have been shown to be effective for myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML).However, these treatments are not intended to be limited to only these disease groups.The compositions and methods disclosed herein may also be used to treat other disorders that require selective depletion of affected HSCs. Associated diseases or disorders may include, but are not limited to, acute myeloid leukemia (AML), chronic myeloid leukemia (CML), chronic myelomonocytic leukemia (CMML), acute lymphoblastic leukemia (ALL), Hodgkin's lymphoma, non-Hodgkin's lymphoma, clonal hematopoiesis of indeterminate potential (CHIP), clonal cytopenia of undetermined significance (CCUS), myelodysplastic syndromes (MDS), idiopathic cytopenia of undetermined significance (ICUS), and myeloproliferative neoplasms (MPN).

[0067] In some embodiments, the disease is blood cancer, optionally leukemia, lymphoma, or myelodysplastic syndrome (MDS). In some embodiments, the disease or disorder is multiple myeloma, chronic myelogenous leukemia (CML), myelodysplastic syndrome (MDS), myeloproliferative neoplasm, or myeloid leukemia, for example, acute myelogenous leukemia (AML) or chronic myelogenous leukemia (CML). In some embodiments, the disease is MDS or AML. In some embodiments, the cancer is lymphocytic leukemia, for example, acute lymphocytic leukemia (ALL) or chronic lymphocytic leukemia (CLL).

[0068] In some embodiments, the cancer is a myelodysplastic / myeloproliferative neoplasm (MDS / MPN), such as chronic myelomonocytic leukemia (CMML). MDS / MPN has both "dysplastic" and "proliferative" features that cannot be classified as either myelodysplastic syndrome (MDS) or myeloproliferative neoplasm (MPN), and for this reason is classified as an overlap syndrome with its own separate characteristics (MDS / MPN). CMML is a cancer of the blood. CMML is considered a myelodysplastic / myeloproliferative neoplasm (MDS / MPN) and is a type of chronic blood cancer in which a person's bone marrow does not produce blood effectively.

[0069] In some embodiments, the subject has a hematopoietic cell transplantation comorbidity index (HCT-CI) of 3 or greater. (Sorror ML, et al. Hematopoietic cell transplantation (HCT)-specific comorbidity index: a new tool for risk assessment before allogeneic HCT. Blood. 2005;106(8):2912-2919.) In some embodiments, the subject has a hematopoietic cell transplantation comorbidity index (HCT-CI) of 3 or less.

[0070] In some embodiments, the disease or disorder is multiple myeloma, severe combined immunodeficiency (SCID), chronic myelogenous leukemia (CML), myelodysplastic syndrome (MDS), myeloproliferative neoplasms, or acute myeloid leukemia (AML).

[0071] In certain embodiments, the disease treated according to the present disclosure is referred to as MDS / AML, which includes both MDS and AML. Myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML) exist along a continuous disease spectrum beginning with early MDS, which can progress to advanced MDS, AML, cured AML, or resistant AML. The disease is characterized by the overproduction of immature blood cells. The resulting lack of mature, healthy blood cells causes anemia and increased risk of infection and bleeding. Approximately 5-10% of patients with solid tumors treated with chemotherapy, radiation, or autologous stem cell transplantation develop therapy-related MDS or AML.

[0072] Myelodysplastic syndromes (MDS) are a group of hematopoietic tumors characterized by abnormal differentiation and cell morphology (i.e., dysplasia) of multipotent hematopoietic progenitor cells (i.e., stem cells) present in the myeloid compartment of the bone marrow (BM). These abnormalities lead to ineffective hematopoiesis and cytopenias (i.e., lower than normal peripheral blood counts) of one or more lineages of myeloid progenitor cells that manifest as anemia, neutropenia, and / or thrombocytopenia. The methods disclosed herein may be used to treat various forms of MDS, including, but not limited to, those shown in Table 1 below, reproduced from Chung, US Pharm. 2021;46(9):39-44. In certain embodiments, the methods result in a reduction in cytopenias. [Table 1]

[0073] In some embodiments, subjects selected for anti-c-kit treatment of MDS as described herein qualify as having lower risk myelodysplastic syndrome (MDS) according to IPSS-R guidelines (see, for example, FIG. 3, and Greenberg et al., Revised international prognostic scoring system for myelodysplastic syndromes, Blood, Sep 20;120(12):2454-65; 2012). In some embodiments, subjects selected for anti-c-kit treatment as described herein qualify as having intermediate risk MDS according to IPSS-R guidelines. In some embodiments, subjects selected for anti-c-kit treatment as described herein qualify as having higher risk MDS according to IPSSR guidelines. In certain embodiments, lower risk MDS patients are defined as patients with very low, low, or moderate risk of disease according to IPSS-R with a score of 3.5 points or less, and higher risk patients are patients with intermediate risk and high, high, or very high risk according to this scoring system with a score of more than 3.5 points. In certain embodiments, the subject has red blood cell transfusion dependence, thrombocytopenia, and / or neutropenia.

[0074] In some embodiments, patients, e.g., patients with CML, MDS, or AML, present with minimally discernible disease (MID) and / or measurable residual disease (MRD), which may be detected by techniques including, but not limited to, cytogenetics, flow cytometry, and / or next generation sequencing (NGS).

[0075] In some embodiments, administration of a combination therapy described herein reduces or eliminates a patient's minimal residual disease (MRD).

[0076] In some embodiments, the subject selected for anti-c-kit treatment described herein has not responded to erythropoietin stimulating agent (ESA). In some embodiments, the ESA is erythropoietin. In some embodiments, the ESA is darbepoietin.

[0077] In some embodiments, the subject selected for anti-c-kit treatment described herein has not responded to chemotherapy.In some embodiments, chemotherapy is azacitidine, lenalidomide, decitabine, and / or luspatercept.

[0078] III. Dosage and Administration According to the embodiment of the method of the present disclosure, diseased HSCs are selectively depleted at lower doses and administration frequency compared to healthy HSCs.Therefore, in certain embodiments, the anti-c-kit treatment described herein is administered in a dosing regimen that preferentially depletes diseased HSCs compared to healthy cells, providing healthy cell predominance in bone marrow and hematopoiesis.Therefore, the method disclosed herein can be used to preferentially deplete diseased HSCs (rather than healthy HSCs) from a subject.

[0079] In some embodiments, a subject in need thereof is administered an anti-c-kit antibody at a dose of about 0.01 mg / kg to about 20 mg / kg. In some embodiments, the anti-c-kit antibody is any of SR-1, JSP191, MGTA-117, FSI-174, CDX-0159, 8D7, K45, 104D2, CK6, AB249, YB5.B8, AF-2-1, AF11, AF12, AF112, AF-3, AF-1-1, NF, NF-2-1, NF11, NF12, NF112, NF-3, HF11, HF12, and HF112 administered at a dose of about 0.01 mg / kg to about 20 mg / kg of the subject's body weight. In some embodiments, the anti-c-kit antibody is at about 0.1 mg / kg, about 0.2 mg / kg, about 0.3 mg / kg, about 0.4 mg / kg, about 0.5 mg / kg, about 0.6 mg / kg, about 0.7 mg / kg, about 0.8 mg / kg, about 0.9 mg / kg, about 1.0 mg / kg, about 1.1 mg / kg, about 1.2 mg / kg, about 1.3 mg / kg, about 1.4 mg / kg, about 1.5 mg / kg, about 1.6 mg / kg g, about 1.7 mg / kg, about 1.8 mg / kg, about 1.9 mg / kg, about 2.0 mg / kg, about 2.25 mg / kg, about 2.5 mg / kg, about 2.75 mg / kg, about 3.0 mg / kg, about 3.25 mg / kg, about 3.5 mg / kg, about 3.75 mg / kg, about 4.0 mg / kg, about 4.5 mg / kg, about 5.0 mg / kg, about 5.5 mg / kg, or about 6.0 mg / kg. In certain embodiments, the anti-c-Kit antibody is administered at a dose of about 0.1 mg / kg to about 4 mg / kg, or about .5 mg / kg to about 4.0 mg / kg, or about .5 mg / kg to about 2.0 mg / kg. In certain embodiments, JSP191 is administered at any of these dosages. In certain embodiments, CDX0158 or CDX0159 is administered at any of these dosages. In certain embodiments, FSI-174 is administered at any of these dosages.

[0080] In some embodiments, a subject in need thereof is administered a single dose of an anti-c-kit antibody described herein. In some embodiments, a subject is administered two or more doses of an anti-c-kit antibody.

[0081] In some embodiments, the subject is administered anti-c-kit antibody in a dosing regimen of about every week, about every 2 weeks, about every 3 weeks, about every 4 weeks, about every 5 weeks, about every 6 weeks, about every 7 weeks, about every 8 weeks, about every 9 weeks, about every 10 weeks, about every 11 weeks, about every 12 weeks, about every 13 weeks, about every 14 weeks, about every 15 weeks, about every 16 weeks, about every 17 weeks, about every 18 weeks, about every 19 weeks, or about every 20 weeks, or about once a month, once every two months, or once every three months. In some embodiments, the subject is administered a dosing regimen of anti-c-kit antibody, and the time interval between each dose varies, for example, the subject may be administered one dose about every week for about 4 weeks, followed by one dose about every two weeks for about 8 weeks, or combinations thereof. In one embodiment, the subject is administered an anti-c-kit antibody about once every 8 weeks for 4 cycles, e.g., JSP191 is administered at about 0.3 mg / kg, 0.6 mg / kg, 0.9 mg / kg, or 1.2 mg.kg of body weight. In certain embodiments, the anti-c-kit antibody is JSP191 and is administered at a dose of about 0.1 mg / kg to about 4 mg / kg, or about 0.5 mg / kg to about 4.0 mg / kg, or about 0.5 mg / kg to about 2.0 mg / kg, about 3.0 mg / kg, about 3.25 mg / kg, about 3.5 mg / kg, about 3.75 mg / kg, about 4.0 mg / kg, about 4.5 mg / kg, about 5.0 mg / kg, about 5.5 mg / kg, or about 6.0 mg / kg of the subject's body weight. In certain embodiments, JSP191 is administered to an MDS (e.g., low-risk MDS) or AML patient at a dose of about 0.1 mg / kg to about 4 mg / kg, or about 0.5 mg / kg to about 4.0 mg / kg, or about 0.5 mg / kg to about 2.0 mg / kg, about 3.0 mg / kg, about 3.25 mg / kg, about 3.5 mg / kg, about 3.75 mg / kg, about 4.0 mg / kg, about 4.5 mg / kg, about 5.0 mg / kg, about 5.5 mg / kg, or about 6.0 mg / kg of the subject's body weight about once a month for about four months.

[0082] In some embodiments, the subject is administered anti-c-kit antibody in a dosing regimen of about monthly, about every 6 months, about every year, about every 2 years, about every 3 years, or about every 5 years. In some embodiments, the subject is administered an initial dose or a loading dose of anti-c-kit antibody as part of the dosing regimen.

[0083] In some embodiments, the dosing regimen continues for about or at least 1 month, about or at least 6 months, about or at least 1 year, about or at least 2 years, about or at least 3 years, or about or at least 5 years. In some embodiments, the dosing regimen continues until the subject is disease-free.

[0084] In some embodiments, anti-c-kit antibody treatments described herein increase the proportion of healthy hematopoietic stem cells (HSCs) to total HSCs in the subject's bone marrow to at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 99%, or at least 100% of all HSCs present in the subject's bone marrow.

[0085] In some embodiments, anti-c-kit antibody treatment is associated with long-term predominance of healthy hematopoietic stem cells (HSCs) in the bone marrow of a subject.As used herein, "long-term predominance" of healthy HSCs can refer to a long-term increase in the proportion of healthy HSCs in bone marrow, as assessed after any period of time from treatment, for example, 6 months, 1 year, 2 years, 3 years, 4 years, 5 years, 6 years, 7 years, 8 years, 9 years, or 10 years from treatment.

[0086] In some embodiments, the percentage of healthy hematopoietic stem cells following administration of an anti-c-kit therapy described herein is assessed by a bone marrow biopsy, blood count, or blood smear from the subject.

[0087] According to embodiments of the methods disclosed herein, monotherapy treatment with anti-c-kit antibody is sufficient and the subject does not need to be treated with another therapeutic agent for the disease being treated.In other embodiments, the subject is treated with anti-c-kit antibody in combination with another chemotherapeutic agent or immunotherapy for the treatment of the disease being treated.

[0088] In some embodiments, subjects administered an anti-c-kit treatment described herein do not undergo hematopoietic cell transplantation (HCT) for at least 4 months, at least 6 months, at least 1 year, at least 2 years, or any period following administration of the anti-c-kit antibody.

[0089] In some embodiments, a subject administered an anti-c-kit treatment described herein does not undergo total body irradiation (TBI) for at least 4 months, at least 6 months, at least 1 year, at least 2 years, or any period of time following administration of the anti-c-kit antibody.

[0090] In some embodiments, a subject administered an anti-c-kit treatment described herein does not receive chemotherapy, including but not limited to, azacitidine, lenalidomide, decitabine, and / or luspatercept, for at least 4 months, at least 6 months, at least 1 year, or at least 2 years, or any period of time following administration of the anti-c-kit antibody.

[0091] In some embodiments, the anti-c-kit treatments described herein are associated with reduced side effects compared to other treatments, including but not limited to hematopoietic cell transplantation (HCT), total body irradiation (TBI), and / or chemotherapy. Measurement of fewer side effects may be determined by the number of patient complaints, number of clinic visits / consultations, and / or number of hospitalizations after treatment. In some embodiments, reduced side effects include reduced risk of hematologic toxicity and / or reduced risk of death due to infection. In some embodiments, the methods disclosed herein result in reduced cytopenia compared to other treatments, such as, for example, HCT.

[0092] In some embodiments, the anti-c-kit antibodies described herein may be administered as a combination therapy that includes an anti-CD47 antibody.

[0093] In some embodiments, the anti-c-kit antibody described herein may be administered as a combination therapy with chemotherapeutic agents, such as azacitidine, lenalidomide, decitabine, and / or luspatercept.In certain embodiments, the anti-c-kit antibody, such as JSP-191, is administered to the subject in combination with azacitidine.The two agents may be administered at the same time or at different times, and in some embodiments, both agents are present in the subject for a period of time.

[0094] In some embodiments, the anti-c-kit antibody therapy (or combination therapy) may be delivered orally, subcutaneously, intravenously, intranasally, transdermally, intraperitoneally, intramuscularly, intrapulmonary, intravaginally, intrarectally, or intraocularly. In some embodiments, the anti-c-kit antibody, e.g., JSP191, is administered systemically, e.g., intravenously or subcutaneously. In some embodiments, the anti-c-kit antibody is delivered directly to the bone marrow.

[0095] IV. Pharmaceutical Compositions In some embodiments, the anti-c-kit antibody (or combination therapy) is present in a pharmaceutical composition. In certain embodiments, the pharmaceutical composition may be in a water-soluble form, such as a pharma- ceutically acceptable salt, which is intended to include both acid and base addition salts. Pharmaceutically acceptable acid addition salts include, but are not limited to, hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, organic acids, such as acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluenesulfonic acid, and salicylic acid. Pharmaceutically acceptable base addition salts include those derived from inorganic bases, such as sodium, potassium, lithium, ammonium, calcium, magnesium, iron, zinc, copper, manganese, and aluminum salts. Salts derived from pharma- ceutically acceptable organic non-toxic bases include salts of primary, secondary, and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines, and basic ion exchange resins such as isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, and ethanolamine.

[0096] The pharmaceutical compositions described herein may also include one or more of carrier proteins (such as serum albumin), buffers, fillers (such as microcrystalline cellulose, lactose, corn and other starches), binders, and polyethylene glycol.

[0097] The composition for administration will generally comprise the antibody dissolved in a pharma- ceutically acceptable carrier, preferably an aqueous carrier. A variety of aqueous carriers can be used, such as buffered saline. The composition may contain pharma- ceutically acceptable auxiliary substances required to approximate physiological conditions, such as pH and buffering agents, toxicity counteracting agents, such as sodium acetate, sodium chloride, potassium chloride, calcium chloride, and sodium lactate. The concentration of active agent in the formulation may vary and is selected based on fluid volume, viscosity, and body weight, according to the particular mode of administration selected and the needs of the patient (e.g., Remington's Pharmaceutical Science (15th ed., 1980) and Goodman & Gillman, The Pharmacological Basis of Therapeutics (Hardman et al., eds., 1996)).

[0098] V.Kit The present disclosure further provides kits comprising the anti-c-kit antibodies described herein, e.g., kits for administration of the anti-c-kit antibodies by a medical professional or a subject in need thereof. The kits can include, for example, a container, and one or more doses of the anti-c-kit antibody, as well as instructions for use thereof. In certain embodiments, the kits include multiple doses of the anti-c-kit antibody suitable for a dosing regimen over a period of time. In certain embodiments, each dose includes the same amount of the anti-c-kit antibody, while in other embodiments, the kit may include two or more different doses of the anti-c-kit antibody. In certain embodiments, each dose is in a separate container. In certain embodiments, each of the one or more doses includes about 0.5 mg to about 800 mg of the anti-c-kit antibody, optionally about 5 mg to about 800 mg of the anti-c-kit antibody. In certain embodiments, the one or more doses includes about 40 mg to about 50 mg, optionally about 45 mg of the anti-c-kit antibody. In certain embodiments, the one or more doses comprise from about 140 mg to about 160 mg, optionally about 150 mg, of anti-c-kit antibody. EXAMPLES

[0099] Example 1 JSP191 as first-line treatment for MDS and AML A clinical trial was conducted to demonstrate that treatment of patients with myelodysplastic syndromes (MDS) and acute myeloid leukemia (AML) with JSP191, an anti-c-kit antibody, is effective in depleting diseased stem and progenitor cells.

[0100] Nine patients with either higher-risk MDS or AML were treated with a single dose of 0.6 mg / kg JSP191 administered intravenously. Notably, JSP191 depleted 30–95% of diseased HSC cells (CD34+CD45RA-CD117+HSC) 10–14 days after treatment (Figure 2). Furthermore, in this ongoing clinical trial, JSP191 administered intravenously at 0.6 mg / kg was safe and well tolerated. No significant JSP191-associated infusion reactions were observed, and no significant adverse events were observed.

[0101] These studies demonstrate that for patients who possess at least a small proportion of normal HSCs, treatment with anti-c-kit antibodies can serve as a first-line therapy to deplete diseased HSCs, allowing their replacement through healthy HSC hematopoiesis, as diagrammed in Figure 1.

[0102] Example 2 JSP191 as second-line therapy in patients with lower-risk MDS A phase 1 clinical trial will be conducted to demonstrate that treatment of patients with lower-risk myelodysplastic syndromes (MDS) with JSP191, an anti-c-kit antibody, is effective in depleting affected stem and progenitor cells.

[0103] Four cohorts of MDS patients will be treated with various doses of JSP191, including 0.3 mg / kg (with a sentinel dose of 0.1 mg / kg), 0.6 mg / kg, 0.9 mg / kg, and 12 mg / kg. MDS patients will receive four cycles every eight weeks. Each cohort will include three to six patients. Patients will have very low, low, or intermediate risk MDS by IPSS-R, i.e., lower risk MDS patients. Patients will have RBC transfusion dependency, thrombocytopenia, or neutropenia.

[0104] The studies are expected to show the safety and tolerability of JSP191 at these doses, as well as efficacy and duration of response by HI-E / HI-P / HI-N, reduction in RBC transfusions, ORR, and duration of response by IWG 2006, and progression-free survival. Hematological improvement may be analyzed and demonstrated as described in Okamura, et al., Leuk Res Rep. 2014; 3(1): 24-27. Treatment with JSP191 is expected to cause the restoration and proliferation of healthy HSCs following depletion of diseased HSCs.

[0105] These studies are expected to demonstrate that for lower-risk MDS patients who harbor at least a small percentage of normal HSCs, treatment with anti-c-kit antibodies can serve as a first-line therapy to deplete diseased HSCs, allowing their replacement through healthy HSC hematopoiesis, as diagrammed in Figure 1.

[0106] The various embodiments described above can be combined to provide further embodiments.

[0107] Aspects of the embodiments can be modified, if necessary, to employ concepts from the various patents, applications, and publications to provide further embodiments.

[0108] These and other changes can be made to the embodiments in light of the above detailed description. In general, in the following claims, the terms used should not be construed to limit the claims to the specific embodiments disclosed in the specification and claims, but should be construed to include all possible embodiments, along with the full scope of equivalents to which such claims are entitled. Thus, the claims are not limited by this disclosure.

[0109] All U.S. patents, U.S. patent application publications, U.S. patent applications, foreign patents, foreign patent applications, and non-patent publications referred to in this specification and / or listed in the Application Data Sheet are hereby incorporated by reference in their entireties.

Claims

1. 1. A method of treating a hematopoietic stem cell (HSC) disease or disorder, optionally myelodysplastic syndrome (MDS) or acute myeloid leukemia (AML), in a subject, comprising administering to the subject an anti-c-kit antibody in a dose effective to achieve the long-term presence or predominance of healthy HSCs in the subject's bone marrow.

2. 10. The method of claim 1, wherein the method selectively depletes diseased HSCs relative to healthy HSCs.

3. 3. The method of claim 1 or claim 2, wherein the anti-c-kit antibody comprises one or more complementarity determining regions (CDRs) present in a monoclonal antibody selected from the group consisting of SR-1, JSP191, MGTA-117, FSI-174, CDX-0159, 8D7, K45, 104D2, CK6, AB249, YB5.B8, AF-2-1, AF11, AF12, AF112, AF-3, AF-1-1, NF, NF-2-1, NF11, NF12, NF112, NF-3, HF11, HF12, and HF112.

4. 4. The method of any one of claims 1 to 3, wherein the anti-c-kit antibody comprises one or more complementarity determining regions (CDRs) present in a humanized form of a monoclonal antibody selected from the group consisting of ACK2, ACK4, 2B8, 3C11, MR-1, and CD122.

5. 5. The method of any one of claims 1 to 4, wherein the anti-c-kit antibody comprises the CDRs of an antibody that blocks binding of stem cell factor (SCF) to stem cell factor receptor (CD117), optionally wherein the antibody is JSP191.

6. The method of any one of claims 1 to 5, wherein the subject is administered about 0.01 mg / kg to about 10 mg / kg of the anti-c-kit antibody, optionally about 0.1 mg / kg to about 10 mg / kg of the anti-c-kit antibody.

7. The method of any one of claims 1 to 5, wherein the subject is administered about 0.6 mg / kg of the anti-c-kit antibody.

8. The method of any one of claims 1 to 5, wherein the subject is administered about 2 mg / kg of the anti-c-kit antibody.

9. 9. The method of any one of claims 1-8, wherein the subject's bone marrow comprises at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 6%, at least 7%, at least 8%, at least 9%, at least 10%, at least 15%, or at least 20% healthy HSCs prior to treatment.

10. The method of any one of claims 1 to 9, wherein the disease or disorder is myelodysplastic syndrome (MDS).

11. The method of any one of claims 1 to 10, wherein the MDS is a lower risk MDS.

12. The method of any one of claims 1 to 10, wherein the disease or disorder is acute myeloid leukemia (AML).

13. The method of any one of claims 1 to 12, wherein the anti-c-kit antibody is delivered to the subject systemically, optionally, the anti-c-kit antibody is delivered intravenously or subcutaneously.

14. The method of any one of claims 1 to 12, wherein the anti-c-kit antibody is delivered locally to the subject, and optionally, the anti-c-kit antibody is delivered directly to bone marrow.

15. The method of any one of claims 1 to 14, wherein the subject is administered two or more doses of the anti-c-kit antibody.

16. 16. The method of claim 15, wherein one of the two or more doses is administered every week, every 2 weeks, every 3 weeks, every 4 weeks, every 5 weeks, every 6 weeks, every 7 weeks, every 8 weeks, every 9 weeks, every 10 weeks, every 11 weeks, or every 12 weeks, optionally with varying time intervals between doses.

17. 17. The method of any one of claims 1 to 16, wherein the subject has failed erythropoietin stimulating agent (ESA) therapy, and optionally, the ESA therapy comprises erythropoietin and / or darbepoietin.

18. 18. The method of any one of claims 1 to 17, wherein the method has reduced side effects compared to treatment with a chemotherapeutic agent, and optionally the chemotherapeutic agent comprises azacitidine, lenalidomide, decitabine, and / or luspatercept.

19. 20. The method of claim 18, wherein the reduced side effects are a reduced risk of hematologic toxicity and a reduced risk of death due to infection.

20. 20. The method of any one of claims 1 to 19, wherein the method results in an increase in the proportion of normal HSCs in the bone marrow to at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 99%, or at least 100% of all HSCs.

21. 21. The method of any one of claims 1-20, wherein the subject will not undergo hematopoietic cell transplantation (HCT) within at least 4 months, at least 6 months, at least 1 year, or at least 2 years after administration of the anti-c-kit antibody.

22. The method of any one of claims 1 to 21, wherein the anti-c-kit antibody is administered in combination with an anti-CD47 antibody.

23. The method of any one of claims 1 to 22, wherein the anti-c-kit antibody is administered in combination with a chemotherapeutic agent.

24. 24. The method of claim 23, wherein the chemotherapeutic agent is azacytidine.

25. 1. A method for depleting endogenous diseased hematopoietic stem cells (HSCs) in a subject, comprising administering to the subject an anti-c-kit antibody, the method resulting in an increase in the proportion of healthy hematopoietic stem cells (HSCs) present in the bone marrow of the subject.

26. 26. The method of claim 25, wherein the anti-c-kit antibody is a monoclonal antibody selected from the group consisting of SR-1, JSP191, MGTA-117, FSI-174, CDX0158, CDX0159, 8D7, K45, 104D2, CK6, AB249, YB5.B8, AF-2-1, AF11, AF12, AF112, AF-3, AF-1-1, NF, NF-2-1, NF11, NF12, NF112, NF-3, HF11, HF12, and HF112, optionally comprising one or more complementarity determining regions (CDRs) present in JSP191, CDX0158, CDX0159, or FSI-174.

27. 26. The method of claim 25, wherein the anti-c-kit antibody comprises one or more complementarity determining regions (CDRs) present in a humanized form of a monoclonal antibody selected from the group consisting of ACK2, ACK4, 2B8, 3C11, MR-1, and CD122.

28. 26. The method of claim 25, wherein said anti-c-kit antibody comprises the CDRs of an antibody that blocks binding of stem cell factor (SCF) to the stem cell factor receptor (CD117), optionally wherein said antibody is JSP191.

29. 29. The method of any one of claims 25-28, wherein the subject is administered about 0.01 mg / kg to about 20 mg / kg of the anti-c-kit antibody, optionally about 0.1 mg / kg to about 10 mg / kg of the anti-c-kit antibody.

30. The method of any one of claims 25 to 28, wherein the subject is administered about 0.6 mg / kg of the anti-c-kit antibody.

31. The method of any one of claims 25 to 28, wherein the subject is administered about 2 mg / kg of the anti-c-kit antibody.

32. 2. The method of claim 1, wherein the subject's bone marrow contains at least 1%, at least 2%, at least 3%, at least 4%, at least 5%, at least 6%, at least 7%, at least 8%, at least 9%, at least 10%, at least 15%, or at least 20% healthy HSCs prior to treatment.

33. 33. The method of any one of claims 25 to 32, used to treat a disease or disorder selected from the group consisting of acute myeloid leukemia (AML), chronic myelogenous leukemia (CML), chronic myelomonocytic leukemia (CMML), acute lymphoblastic leukemia (ALL), Hodgkin's lymphoma, non-Hodgkin's lymphoma, clonal hematopoiesis of indeterminate potential (CHIP), clonal cytopenia of undetermined significance (CCUS), myelodysplastic syndromes (MDS), idiopathic cytopenia of undetermined significance (ICUS), and myeloproliferative neoplasms (MPN).

34. 34. The method of claim 33, wherein the disease or disorder is MDS.

35. 35. The method of claim 34, wherein the MDS is a lower risk MDS.

36. The method of any one of claims 25 to 35, wherein the anti-c-kit antibody is delivered to the subject systemically, optionally, the anti-c-kit antibody is delivered intravenously or subcutaneously.

37. The method of any one of claims 25 to 35, wherein the anti-c-kit antibody is delivered locally to the subject, and optionally, the anti-c-kit antibody is delivered directly to bone marrow.

38. The method of any one of claims 25 to 37, wherein the subject is administered two or more doses of the anti-c-kit antibody.

39. 39. The method of claim 38, wherein one of the two or more doses is administered about every week, about every 2 weeks, about every 3 weeks, about every 4 weeks, about every 5 weeks, about every 6 weeks, about every 7 weeks, about every 8 weeks, about every 9 weeks, about every 10 weeks, about every 11 weeks, or about every 12 weeks, optionally varying the time interval between each dose.

40. 40. The method of any one of claims 25-39, wherein the subject has failed erythropoietin stimulating agent (ESA) therapy, and optionally, the ESA therapy comprises erythropoietin and / or darbepoietin.

41. 41. The method of any one of claims 25 to 40, wherein the method has reduced side effects compared to treatment with a chemotherapeutic agent, optionally wherein the chemotherapeutic agent comprises azacitidine, lenalidomide, decitabine, and / or luspatercept.

42. 42. The method of claim 41, wherein the reduced side effects include reduced risk of hematologic toxicity and reduced risk of death from infection.

43. 43. The method of any one of claims 25-42, wherein the method results in increasing the proportion of normal HSCs in the bone marrow to at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, at least 95%, at least 99%, or at least 100% of all HSCs, and optionally the method results in a long-term predominance of healthy hematopoietic stem cells (HSCs) in the bone marrow of the subject.

44. 44. The method of any one of claims 25-43, wherein the subject will not undergo hematopoietic cell transplantation (HCT) within at least 4 months, at least 6 months, at least 1 year, or at least 2 years after administration of the anti-c-kit antibody.

45. A kit comprising one or more doses of an anti-c-kit antibody.

46. 46. ​​The kit of claim 45, wherein each of said one or more doses comprises from about 0.5 mg to about 800 mg of said anti-c-kit antibody, optionally from about 5 mg to about 800 mg of said anti-c-kit antibody.

47. 47. The kit of claim 46, wherein the kit comprises one or more doses comprising about 40 mg to about 50 mg, optionally about 45 mg, of the anti-c-kit antibody.

48. 47. The kit of claim 46, wherein the kit comprises one or more doses comprising about 140 mg to about 160 mg, optionally about 150 mg, of the anti-c-kit antibody.

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