Treatment for hematological malignancy

Abstract

To provide pharmaceutical compositions comprising a radionuclide labelled monoclonal antibody against CD38, for treating a hematological malignancy.SOLUTION: An exemplary monoclonal antibody includes daratumumab labeled with a radionuclide such as actinium (225Ac).SELECTED DRAWING: Figure 1

Description

[Technical Field]

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS This application claims the benefit under 35 U.S.C. §119(e) of U.S. Provisional Patent Application No. 62 / 539,114, entitled "Treatments for a Hematological Malignancy," filed July 31, 2017, the contents of which are incorporated herein by reference. The present invention relates to compositions of radiolabeled monoclonal antibodies specific for the CD38 antigen, and more particularly to compositions comprising anti-CD38 monoclonal antibodies labeled with alpha-emitting nuclides that are useful in the treatment of hematological malignancies. [Background technology]

[0002] Multiple myeloma is a hematological malignancy characterized by the proliferation of a single clone of plasma cells, which normally contribute to antibody production. These malignant plasma cells (multiple myeloma cells) crowd out healthy blood cells in the bone marrow, circulate in the peripheral blood, and invade new sites in the bone marrow. Therefore, progression of multiple myeloma occurs through continuous interactions between the bone marrow and multiple myeloma cells, resulting in multiple tumors and lesions throughout the skeletal system, with a median survival of 5 years. Approximately 1% of all cancers and more than 10% of all hematological malignancies can be attributed to multiple myeloma. The majority of multiple myeloma patients relapse within a few years, and in most cases, relapsed patients do not respond to treatment.

[0003] Currently available treatments for multiple myeloma include chemotherapy, stem cell transplantation, or small molecule drugs such as immunomodulatory agents (lenalidomide, thalidomide), proteasome inhibitors (carfilzomib, bortezomib), and bisphosphonates (pamidronate, zoledronic acid). Current treatment protocols, which include combinations of chemotherapy agents such as vincristine, carmustine, melphalan, cyclophosphamide, adriamycin, and steroids such as prednisone or dexamethasone, result in a complete remission rate of only approximately 5% and a median survival of approximately 36 to 48 months from the time of diagnosis. Recent advances using high-dose chemotherapy followed by autologous bone marrow transplantation or peripheral blood mononuclear cell transplantation have increased the complete remission rate and duration of remission. However, overall survival has only been modestly extended, and no evidence of a cure has been obtained.

[0004] The effectiveness of these available chemotherapy regimens is limited by the low proliferation rate of multiple myeloma cells and the development of multidrug resistance. For more than 90% of multiple myeloma patients, the disease becomes resistant to chemotherapy. As a result, alternative treatment regimens aimed at adoptive immunotherapy targeting surface antigens on plasma cells are needed.

[0005] Multiple myeloma cells uniformly overexpress CD38, a 45 kD type II transmembrane glycoprotein with a long C-terminal extracellular domain and a short N-terminal cytoplasmic domain. The CD38 protein is a cytoplasmic protein that binds NAD + is a bifunctional extracellular enzyme that can catalyze the conversion of NAD to cyclic ADP-ribose (cADPR) and cADPR to ADP-ribose, thus increasing the amount of extracellular NAD + Furthermore, CadPR regulates the intracellular Ca concentration. 2+ CD38 has been shown to be a second messenger for mobilizing intracellular Ca. 2+ It may be an essential component in regulating flux.

[0006] CD38 therapeutic antibodies as a class typically rely on classical Fc-dependent immune effector functions, such as antibody-dependent cellular cytotoxicity (ADCC), antibody-dependent cellular phagocytosis (ADCP), and complement-dependent cytotoxicity (CDC), as their primary mechanism of action; however, recent studies have shown that targeting immunosuppressive mechanisms may also contribute to the anti-tumor activity of certain CD38 antibodies.

[0007] CD38 expression is also upregulated in a variety of other hematological malignancies, including, but not limited to, B-cell chronic lymphocytic leukemia, B-cell acute lymphocytic leukemia, Waldenstrom's hypergammaglobulinemia, primary systemic amyloidosis, mantle cell lymphoma, prolymphocytic leukemia / myeloid leukemia, acute myeloid leukemia, chronic myeloid leukemia, follicular lymphoma, NK-cell leukemia, and plasma cell leukemia.

[0008] Furthermore, CD38 expression has been described on epithelial / endothelial cells of different origins, including glandular epithelium of the prostate, islet cells of the pancreas, ductal epithelium of glands including the parotid gland, bronchial epithelial cells, cells of the testis and ovary, and tumor epithelium of colorectal adenocarcinoma. Thus, diseases in which CD38 expression may be involved include, but are not limited to, bronchial epithelial carcinoma of the lung, breast cancer arising from malignant proliferation of the epithelial lining of the breast ducts and lobules, pancreatic tumors arising from B cells (e.g., insulinoma), and tumors arising from the intestinal epithelium (e.g., adenocarcinoma and squamous cell carcinoma).

[0009] Daratumumab is a first-in-class CD38-targeting antibody approved as a single agent and in combination with standard of care for the treatment of newly diagnosed multiple myeloma (MM) patients who cannot undergo autologous hematopoietic stem cell transplantation, as well as in relapsed or refractory MM. Despite significant improvements in patient response with anti-CD38 monotherapy or combination therapy, not all patients respond, and management of MM remains challenging.

[0010] One of the potential obstacles to patient response to CD38 antibody therapy is the dependency of this approach on tumors with significant CD38 expression levels to induce the aforementioned mechanism of action. Thus, patients with low or heterogeneous tumor CD38 expression tend to respond poorly to antibody therapy. Options for increasing the therapeutic index of drugs, such as antibody-drug conjugates that conjugate cytotoxic toxins or chemotherapeutic agents to antibodies, also rely heavily on high antigen density to deliver a sufficient payload to enable potent tumor cell death. Therefore, there is a need for treatments that can potentially work in situations with low CD38 expression and could improve the outcomes of antibody-based therapies. Summary of the Invention

[0011] A potential solution to increase efficacy and potential response to CD38-specific targeted therapy, which is independent of high CD38 expression, is radioimmunotherapy (RIT), which has emerged as a useful antitumor therapy through the targeting of radionuclides by antibodies against tumor-associated antigens.

[0012] Thus, the present invention relates to a composition comprising a monoclonal antibody against CD38, wherein the antibody is an actinium ( 225 According to certain embodiments of the present invention, the composition may further comprise an unlabeled antibody against CD38 and / or one or more additional therapeutic agents, such as a chemotherapeutic agent, an anti-inflammatory agent, an immunosuppressant, an immunomodulatory agent, or an anti-myeloma agent. According to certain embodiments of the present invention, the monoclonal antibody may be labeled with a radionuclide via a chelator.

[0013] The present invention also relates to the use of actinium ( 225 Ac) also relates to an article of manufacture comprising a labeled monoclonal antibody and labeling providing dosage and administration instructions.

[0014] The present invention further relates to methods for treating mammals with hematological malignancies, for inhibiting the growth and / or proliferation of cells expressing CD38, and for treating diseases or disorders involving cells expressing CD38, by administering compositions comprising radiolabeled monoclonal antibodies to CD38.

[0015] The objects of the invention will be realized and attained by means of the combinations particularly outlined in the appended claims. The foregoing general description as well as the following detailed description and examples of the invention are provided to illustrate various aspects of the invention and should not be construed in any way as limiting any of the described embodiments. [Brief explanation of the drawings]

[0016] [Figure 1] FIG. 1 provides a schematic diagram of a method for radiolabeling daratumumab with actinium-225Ac, where panel (A) shows the conjugation of the bifunctional chelator S-2-(4-isothiocyanatobenzyl)-1,4,7,10 tetraazacyclododecanetetraacetic acid (p-SCN-Bn-DOTA; referred to as DOTA in the figure) to the monoclonal antibody daratumumab (DARA), and panel (B) shows the radiolabeling of the DOTA-DARA conjugate with 225Ac to provide 225Ac-DARA. [Figure 2] A bar graph showing the yield of actinium (225Ac) chelation by immunoconjugates of the chelators DOTA and DARA is provided. The conjugation reaction was carried out at 37°C or room temperature with various molar excesses of DOTA chelator relative to DARA. DOTA-DARA at 0.3 mg / ml in 0.15 M ammonium acetate buffer (pH 6.5) was labeled with 225Ac in 0.01 M HCl with 1 μCi / μg DARA for 60 minutes at 37°C or room temperature (chelation of 225Ac by DOTA). [Figure 3A]A graph showing the storage stability at 4°C of various radiolabeled DARA samples is provided, the samples being derived from conjugation reactions of DOTA and DARA carried out at 37°C or room temperature with various molar excesses of DOTA relative to DARA. [Figure 3B] A graph showing the storage stability at room temperature of various radiolabeled DARA samples is provided, the samples being derived from conjugation reactions of DOTA and DARA carried out at 37°C or room temperature with various molar excesses of DOTA relative to DARA. [Figure 4A] A graph showing the binding of various concentrations of 225Ac-DARA conjugates to CD38 is provided, which were derived from the conjugation reaction of DOTA with DARA carried out at 37°C or room temperature with various molar excesses of DOTA relative to DARA. [Figure 4B] A graph showing the results when various concentrations of DARA and DOTA-DARA were immobilized on plastic and incubated with the complement protein C1q is provided. The amount of C1q binding was assessed using anti-C1q-HRP as a probe. [Figure 4C] A graph showing the results when various concentrations of DARA and DOTA-DARA were added to target cells expressing CD38 is provided. Effector cells that express luciferase when activated by FcγR(III) were added to the target cells, and luminescence was measured after the addition of Bio-Glow™. Fold induction = RLU (background induction) / RLU (background control without antibody). An anti-CD20 antibody known to activate ADCC was used as a positive control. [Figure 5] Graphs are provided showing the ability of 225Ac-DARA conjugates to induce lysis of Daudi cells compared to unlabeled DARA, 225Ac-DOTA-IgG conjugates, and untreated cells, with Figures 5A-5D showing exposure times of 24, 48, 72, and 96 hours, respectively. [Figure 6]A summary graph of the data seen in Figures 5A-5D is provided showing the percent cell lysis over time for various concentrations of 225Ac-DARA. [Figure 7] 1 provides a graph showing the ability of various concentrations of 225Ac-DARA conjugate to induce lysis of 28BM and 28PE multiple myeloma cells by ADCC, respectively, over a 96 hour period. [Figure 8] Bar graphs comparing the cytolysis results of Daudi, 28BM, and 28PE cells upon exposure to various concentrations of 225Ac-DOTA-DARA conjugate are provided, with Figures 8A-8C showing exposure times of 48, 72, and 96 hours, respectively. [Figure 9] Figure 1 shows the biodistribution of In-DARA in Daudi tumor-bearing SCID mice. Mice were administered a single intraperitoneal (IP) injection of In-DARA (400 μCi) and imaged by microSPECT / CT at 1, 4, and 24 hours post-injection, followed by days 2, 3, 7, and 10. [Figure 10] Figure 5 shows the results of radioimmunotherapy treatment (RIT) of Daudi tumor-bearing SCID mice. Figure 5A shows tumor volume after RIT, and Figure 5B shows a Kaplan-Meier plot of mouse survival time after RIT. SCID mice were injected with CD38-positive Daudi cells into the right flank. When tumors reached approximately 200 mm3, they were treated with a single IP injection of 200 or 400 nCi of 225Ac-DARA (0.3 μg), the same amount of 0.3 μg naked DARA, or a 30-fold higher dose (10 μg) of naked DARA, or saline. Tumor volume was calculated using the formula V = 0.5(LW2). Mice were sacrificed when tumor volumes reached 4,000 mm3. [Figure 11]Figure 11 shows the results of a safety evaluation of RIT using 225Ac-DARA. Figure 11A shows mouse weights after RIT, and Figure 11B shows blood, liver, and kidney health parameters 7 days after RIT. SCID mice were injected with CD38-positive Daudi cells into the right flank. When tumors reached approximately 200 mm, they were treated with a single IP injection of 200 or 400 nCi of 225Ac-DARA (0.3 μg), the same amount of 0.3 μg naked DARA, a 30-fold higher dose (10 μg) naked DARA, or saline. Mouse weights were monitored every 3 days after RIT. Blood was collected 7 days after RIT and analyzed for the indicated parameters. [Figure 12] The amino acid sequence of human CD38 as shown in GenBank accession number NP_001766 is provided.

[0017] Definitions and Abbreviations The singular forms "a," "an," "the," etc. include plural referents unless the context clearly dictates otherwise. Thus, for example, "a" diluent includes both a single diluent and a plurality of different diluents.

[0018] The term "about" when used before numerical designations, such as temperatures, times, amounts, and concentrations, indicates approximations that may vary by ±10%, ±5%, or ±1%.

[0019] "Comprising" or "comprises" is intended to mean that the compositions and methods include the recited elements, but do not exclude others. "Consisting essentially of," when used to define compositions and methods, is intended to mean excluding other elements that are of any essential importance to the combination for the purpose being described. Thus, a method consisting essentially of the elements defined herein does not exclude other steps or compositions that do not materially affect the basic and novel feature(s) of the claimed invention.

[0020] "Monoclonal antibody" refers to a preparation of antibody molecules of single molecular composition. A monoclonal antibody composition exhibits a single binding specificity and affinity for a particular epitope, or, in the case of bispecific monoclonal antibodies, dual binding specificities for two distinct epitopes. Thus, "monoclonal antibody" refers to an antibody population having a single amino acid composition in each heavy and light chain, except for possible and well-known variations such as removal of the C-terminal lysine from the antibody heavy chain. Monoclonal antibodies may have heterogeneous glycosylation within the antibody population. Monoclonal antibodies may be monospecific or polyspecific, and may be monovalent, bivalent, or multivalent. Bispecific antibodies are encompassed within the term monoclonal antibody.

[0021] "Epitope" refers to a site on a target molecule that can be recognized and bound by an antibody. In the case of a protein epitope, for example, this may refer to the amino acids (and particularly their side chains) bound by the antibody. Overlapping epitopes contain at least 1-5 common amino acid residues. Methods for identifying antibody epitopes are known to those of skill in the art and include, for example, those described in Antibodies, A Laboratory Manual, Cold Spring Harbor Laboratory, Ed Harlow and David Lane (1988).

[0022] A "humanized antibody" refers to an antibody in which the antigen-binding site is derived from a species other than human and the variable region framework is derived from human immunoglobulin sequences. Humanized antibodies may contain substitutions in the framework regions such that the framework may not be an exact copy of expressed human immunoglobulin or germline gene sequences.

[0023] A "human antibody" refers to an antibody having heavy and light chain variable regions in which both the framework and antigen-binding sites are derived from sequences of human origin. If the antibody contains a constant region, the constant region also is derived from sequences of human origin.

[0024] A human antibody comprises a heavy or light chain variable region having a variable domain sequence "derived from" a sequence of human origin, where the variable regions of the antibody are obtained from systems that use human germline immunoglobulin or rearranged immunoglobulin genes. Such systems include human immunoglobulin gene libraries displayed on phage and transgenic non-human animals, such as mice, carrying human immunoglobulin loci. Human antibodies can contain amino acid differences when compared to human germline immunoglobulin or rearranged immunoglobulin genes due, for example, to naturally occurring somatic mutations, or the introduction of deliberate substitutions in the framework or antigen-binding sites, or both. Typically, a human antibody refers to an antibody having an amino acid sequence at least about 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% identical to an amino acid sequence encoded by a human germline immunoglobulin or rearranged immunoglobulin gene.

[0025] "Immune reactivity" refers to a measure of the ability of an immunoglobulin to recognize and bind to a particular antigen.

[0026] An "isolated antibody" refers to an antibody or antibody fragment that is substantially free of other antibodies having different antigen specificities (e.g., an isolated antibody that specifically binds to CD38 is substantially free of antibodies that specifically bind to antigens other than human CD38). However, an isolated antibody that specifically binds to CD38 may have cross-reactivity to other antigens, as discussed above. Furthermore, an isolated antibody may be substantially free of other cellular material and / or chemicals. An "isolated antibody" encompasses antibodies isolated to greater degrees of purity, e.g., 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 100% pure antibodies.

[0027] "Pharmaceutically acceptable salts" refers to acid addition salts of basic compounds, e.g., compounds containing a basic amino group, and acidic compounds, e.g., compounds containing a carboxyl group, and amphoteric salts of compounds containing both acidic and basic moieties, such that these salts are suitable for in vivo administration, preferably to humans. A variety of organic and inorganic acids can be used to form acid addition salts. Pharmaceutically acceptable salts are derived from a variety of organic and inorganic counterions well known in the art. Pharmaceutically acceptable salts include, by way of example only, hydrochloride, hydrobromide, tartrate, mesylate, acetate, maleate, oxalate, etc., when the molecule contains a basic functional group; and, by way of example only, sodium, potassium, calcium, magnesium, ammonium, tetraalkylammonium, N-methylmorpholinium, etc., when the molecule contains an acidic functional group. In one embodiment, the pharmaceutically acceptable salt of ezatiostat is ezatiostat hydrochloride.

[0028] As used herein, "cancer" includes, but is not limited to, solid cancers (e.g., tumors) and hematological malignancies.

[0029] "Hematologic malignancies," also known as blood cancers, are cancers that begin in blood-forming tissues, such as the bone marrow or other cells of the immune system. Hematological malignancies include, but are not limited to, leukemia (e.g., acute myeloid leukemia (AML), acute promyelocytic leukemia, acute lymphoblastic leukemia (ALL), acute mixed lineage leukemia, chronic myeloid leukemia, chronic lymphocytic leukemia (CLL), hairy cell leukemia, and large granular lymphocytic leukemia), myelodysplastic syndromes (MDS), myeloproliferative disorders (polycythemia vera, essential thrombocytosis, primary myelofibrosis, and chronic myelogenous leukemia), lymphoma, multiple myeloma, MGUS and similar disorders, Hodgkin's lymphoma, non-Hodgkin's lymphoma (NHL), primary mediastinal large B-cell lymphoma, diffuse large B-cell lymphoma, follicular lymphoma, transformed follicular lymphoma, splenic marginal zone lymphoma, lymphocytic lymphoma, T-cell lymphoma, and other B-cell malignancies.

[0030] As used herein, "peripheral blood lymphocytes" of a subject shall mean mature lymphocytes circulating in the blood of a subject. Examples of peripheral blood lymphocytes include, but are not limited to, peripheral blood T cells, peripheral blood NK cells, and peripheral blood B cells. A subject's peripheral blood lymphocyte population is readily measurable. Thus, depletion events (e.g., hypoxemia) can be detected. 131 It can be readily determined that lymphodepletion has occurred in a subject by measuring a decrease in the level of at least one type of peripheral blood lymphocyte after administration of 8 doses of I-BC.

[0031] "Solid tumors" include, but are not limited to, bone cancer, pancreatic cancer, skin cancer, cancer of the head or neck, cutaneous or intraocular malignant melanoma, uterine cancer, ovarian cancer, prostate cancer, rectal cancer, cancer of the anal region, stomach cancer, testicular cancer, uterine cancer, fallopian tube cancer, endometrial cancer, cervical cancer, vaginal cancer, vulvar cancer, esophageal cancer, small intestine cancer, endocrine system cancer, thyroid cancer, parathyroid cancer, adrenal gland cancer, soft tissue sarcoma, urethral cancer, penile cancer, pediatric tumors, bladder cancer, kidney or ureter cancer, renal pelvis cancer, central nervous system (CNS) neoplasms, primary CNS lymphoma, tumor angiogenesis, spinal axis tumors, brain stem glioma, pituitary adenoma, Kaposi's sarcoma, epidermoid carcinoma, squamous cell carcinoma, and environmentally induced cancers including those induced by asbestos.

[0032] As used herein, the term "subject" includes, but is not limited to, mammals such as humans, non-human primates, dogs, cats, horses, sheep, goats, cows, rabbits, pigs, rats, and mice. When the subject is a human, the subject can be of any age. For example, the subject can be 60 years of age or older, 65 years of age or older, 70 years of age or older, 75 years of age or older, 80 years of age or older, 85 years of age or older, or 90 years of age or older. Alternatively, the subject can be 50 years of age or younger, 45 years of age or younger, 40 years of age or younger, 35 years of age or younger, 30 years of age or younger, 25 years of age or younger, or 20 years of age or younger. In the case of a human subject with cancer, the subject can be newly diagnosed, or can be relapsed and / or refractory, or can be in remission. "Patient" and "subject" are used interchangeably herein.

[0033] As used herein, a "radioisotope" can be an alpha-emitting isotope, a beta-emitting isotope, and / or a gamma-emitting isotope. Examples of radioisotopes include: 90 Y, 89 Sr, 153 Sm, 32 P, 225 Ac, 213 Bi, 213 Po, 211 At, 212 Bi, 213 Bi, 223 Ra, 227 Th, 149 Tb, 131 I, 137 Cs, 212 Pb and 103 Pd. Thus, radiolabeled antibodies contemplated in the present invention include, but are not limited to: 90 Y-anti-CD38, 89 Sr-anti-CD38, 153 Sm-anti-CD38, 32 P-anti-CD38, 225 Ac-anti-CD38, 213 Bi-anti-CD38, 213 Po-anti-CD38, 211 At-anti-CD38, 212 Bi-anti-CD38, 213 Bi-anti-CD38, 223 Ra-anti-CD38, 227 Th-anti-CD38, 149 Tb-anti-CD38, 131 I-anti-CD38, 137 Cs-anti-CD38, 212 Pb-anti-CD38, and 103 Pd-anti-CD38.

[0034] As used herein, "treating" a subject afflicted with cancer includes, but is not limited to, (i) slowing, halting, or reversing the progression of cancer, (ii) slowing, halting, or reversing the progression of cancer symptoms, (iii) reducing the likelihood of cancer recurrence, and / or (iv) reducing the likelihood of cancer symptoms recurrence. According to certain preferred aspects, treating a subject afflicted with cancer means (i) reversing the progression of cancer, ideally until the cancer is eliminated, and / or (ii) reversing the progression of cancer symptoms, ideally until the symptoms are eliminated, and / or (iii) reducing or eliminating the likelihood of recurrence (i.e., ideally, consolidation therapy to destroy any remaining cancer cells).

[0035] A "therapeutically effective amount" refers to an amount effective, at dosages and for periods of time necessary, to achieve a desired therapeutic result. A therapeutically effective amount may vary depending on factors such as the individual's condition, age, sex, and weight, as well as the ability of the therapeutic agent or combination of therapeutic agents to elicit a desired response in the individual. Examples of indicators of an effective therapeutic agent or combination of therapeutic agents include, for example, improved patient health, a reduction in tumor burden, cessation or slowing of tumor growth, and / or the absence of metastasis of cancer cells to other locations in the body.

[0036] "Inhibiting growth" refers to a measurable reduction or delay in the growth of malignant cells or tissues (e.g., tumors) in vitro or in vivo when contacted with a therapeutic agent or combination of therapeutic agents, compared to the reduction or delay in growth of the same cells or tissues in the absence of the therapeutic agent or combination of therapeutic agents. Inhibition of growth of malignant cells or tissues in vitro or in vivo can be at least about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, 96%, 97%, 98%, 99%, or 100%.

[0037] "CD38" refers to the human CD38 protein (synonyms: ADP-ribosyl cyclase 1, cADPr hydrolase 1, cyclic ADP-ribose hydrolase 1). Human CD38 has the amino acid sequence shown in SEQ ID NO: 1 (Figure 12).

[0038] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of the invention described herein, suitable methods and materials are described below.

[0039] Detailed Description of the Invention The present invention relates to compositions comprising radiolabeled monoclonal antibodies against CD38 and methods for treating mammals with hematological malignancies, inhibiting the growth and / or proliferation of cells expressing CD38, and treating diseases or disorders involving cells expressing CD38 by administering the compositions.

[0040] Human CD38 has the amino acid sequence shown in GenBank accession number NP_001766 and SEQ ID NO: 1 (FIG. 12). It is known that CD38 is a single-pass transmembrane II protein, with amino acid residues 1-21 representing the cytoplasmic domain, amino acid residues 22-42 representing the transmembrane domain, and residues 43-300 representing the extracellular domain of CD38.

[0041] Antibodies against CD38, such as daratumumab, MOR202, or SAR650984, have been and are currently being evaluated clinically for their efficacy in treating hematological malignancies and plasma cell disorders, including multiple myeloma. Each antibody has been shown to bind to a different part of the extracellular domain of CD38 (Table I), and each exhibits different clinical responses (e.g., antitumor effects). Daratumumab, available from Johnson & Johnson (Janssen Biotech) / Genmab as Darzalex®, is described in the publication to de Weers et al., “Daratumumab, a Novel Therapeutic Human CD38 Monoclonal Antibody, Induces Killing of Multiple Myeloma and Other Hematological Tumors,” J Immunology, 2010, 186(3) 1840-1848; MOR202, available from Celgene Corp. / Morphosys, is described in U.S. Pat. No. 8,877,899; and SAR650984, available from Sanofi / Immunogen as Isatuximab, is described in Park et al., “SAR650984: A Potent Anti-CD38 Therapeutic Antibody with Three Mechanisms of Action (Apoptosis, ADCC, CDC) for Hematological Tumors,” J Immunology, 2010, 186(3) 1840-1848. Malignancies,” BLOOD, Vol. 112, No. 11, Nov. 2008, p. 951, and the publication to Deckert et al., “SAR650984, A Novel Humanized CD38-Targeting Antibody, Demonstrates Potent Antitumor Activity in Models of Multiple Myeloma and Other CD38 +Hematologic Malignancies,” Clin Cancer Res 2014;20:4574-83, and U.S. Patent No. 8,153,765. Table 1 below lists a number of antibodies, or fragments thereof, that bind to CD38 and their epitopes (binding sites) on the CD38 molecule (see FIG. 12), some or all of which may be useful in accordance with various aspects of the present invention. [Table 1] 1)Tissue Antigens 2000,(56):539-547 and BMC Immunology 2004,(5):21 2)J.Biol.Chem.2011,(286):22170-22177 3) U.S. Patent No. 8,153,765 and Clin Cancer Res 2014,20(17):4574-83 4)J Immunol 2011,(186):1840-1848 5) U.S. Patent No. 8,877,899

[0042] The methods proposed by which these antibodies eliminate CD38-positive cells include antibody-dependent cellular cytotoxicity (ADCC), complement-dependent cytotoxicity (CDC), and apoptosis.

[0043] "Antibody-dependent cellular cytotoxicity," "antibody-dependent cell-mediated cytotoxicity," or "ADCC" is a mechanism for inducing cell death that relies on the interaction of antibody-coated target cells with lytic effector cells, such as natural killer (NK) cells, monocytes, macrophages, and neutrophils, via Fc gamma receptors (FcγR) expressed on the effector cells. For example, NK cells express FcγRIIIa, while monocytes express FcγRI, FcγRII, and FcγRIIIa. Death of antibody-coated target cells, such as CD38-expressing cells, occurs as a result of effector cell activity by secretion of pore-forming proteins and proteases.

[0044] "Complement-dependent cytotoxicity" or "CDC" refers to a mechanism for inducing cell death in which the Fc effector domain of a target-binding antibody binds to and activates complement component C1q, which then activates the complement cascade, resulting in the death of the target cell. Complement activation can also result in the deposition of complement components on the target cell surface, which promote ADCC by binding to complement receptors (e.g., CR3) on leukocytes.

[0045] "Apoptosis" refers to a mechanism of programmed cell death in which antibody binding to a target cell disrupts essential cell signaling pathways, leading to the self-destruction of the cell.

[0046] To assess the ADCC activity of an antibody that specifically binds to CD38, the antibody may be added to CD38-expressing cells in combination with immune effector cells, which can be activated by the antigen-antibody complex, resulting in cytolysis of the CD38-expressing cells. Cytolysis is usually detected by the release of a label (e.g., a radioactive substrate, a fluorescent dye, or a natural intracellular protein) from the lysed cells. Exemplary effector cells for such assays include peripheral blood mononuclear cells (PBMCs) and NK cells.

[0047] In an exemplary ADCC assay, CD38-expressing cells 51 The cells can be labeled with Cr and washed thoroughly. Anti-CD38 antibodies can be added to the CD38-expressing cells at various concentrations, and the assay can be initiated by adding effector cells (e.g., NK cells from peripheral blood mononuclear cells). After incubation at 37°C for various time intervals, the assay is terminated by centrifugation, and the CD38-expressing cells are collected. 51 Cr release is measured in a scintillation counter. The percentage of cytotoxicity can be calculated as the % of maximum lysis that can be induced by adding 3% perchloric acid to CD38-expressing cells.

[0048] In an exemplary cytotoxicity assay, tetrazolium salt is added to CD38-expressing cells treated with various amounts of anti-CD38 antibody. In living mitochondria, XTT is reduced to an orange product by mitochondrial dehydrogenase and transferred to the cell surface. The orange product can be optically quantified and reflects the number of viable cells. Alternatively, esterases from living cells have been shown to hydrolyze colorless calcein as a fluorescent molecule. Fluorescence can be measured and quantified and reflects the number of viable cells in the sample. The total number of dead cells may be measured using propidium iodide, which is excluded from viable cells by intact membranes. The fluorescence of propidium iodide in dead cells can be quantified by flow cytometry.

[0049] To assess CDC, it may be necessary to include complement proteins in the cytotoxicity assay, such as C1q, as detailed in Example 2 herein. Measurement of apoptosis induction does not require the addition of NK cells or complement proteins in the cytotoxicity assay.

[0050] Treatment of various hematological malignancies with unlabeled (i.e., "naked") monoclonal antibodies against CD38 has been successful in recent clinical trials. One such antibody, daratumumab, is currently marketed by Janssen Biotech, Inc. under the name Darzalex® for the treatment of multiple myeloma. The treatment regimen involves an initial 8-week dosing schedule of 16 mg / kg of patient body weight administered by infusion once weekly. Pretreatment with antipyretics and antihistamines is intended to ameliorate some of the more common and serious side effects of the infusion. For example, infusion reactions, which occur in more than half of all patients, include bronchospasm, hypoxia, dyspnea, hypertension, laryngeal edema, and pulmonary edema and can occur up to 48 hours after treatment. Side effects, occurring in more than 20 percent of treated patients, include at least fatigue, nausea, diarrhea, constipation, muscle spasms, back pain, chills, insomnia, cough, and dyspnea. Thus, although treatment with naked antibodies against CD38 has been somewhat successful in treating hematological malignancies, the side effects of treatment can be quite severe and related to the high protein dose of the monoclonal antibodies.

[0051] B cells, such as plasma cells and lymphocytes, are inherently sensitive to radiation therapy and therefore represent attractive targets for radioimmunotherapy targeting hematological malignancies. In addition to the numerous mechanisms by which anti-CD38 antibodies can kill B cells, as described above, the emission of ionizing radiation from radionuclide-labeled antibodies against CD38 can kill cells in close proximity to the antibody bound to CD38-expressing cells. Radionuclides emit radioactive particles that can damage cellular DNA to the point where cellular repair mechanisms are no longer able to keep the cells viable. Therefore, when CD38-expressing cells are involved in tumors, radionuclides can advantageously kill tumor cells.

[0052] Therefore, an approach to improve the ability of immunotherapies that target CD38 to treat hematologic malignancies involves labeling the antibodies with radionuclides. In treating patients with radionuclide-labeled monoclonal antibodies against CD38, the radionuclides can localize to cells that express CD38, such as at the tumor site, and kill those tumor cells.

[0053] Radionuclides that can be used to induce such damage in cells, such as cancer cells, are typically high-energy emitters. High-energy radionuclides preferably act over a short range so as to localize the cytotoxic effect to the target cells. In this way, radiation therapy is delivered in a more localized manner to reduce damage to non-target or non-cancerous cells.

[0054] Thus, the present invention relates to a composition comprising a monoclonal antibody against CD38, wherein the antibody is labeled with a radionuclide. According to a particular embodiment of the present invention, the monoclonal antibody against CD38 may be daratumumab, MOR202, or SAR650984. According to an embodiment of the present invention, the monoclonal antibody against CD38 may be daratumumab.

[0055] According to a particular aspect of the invention, the radionuclide is a beta-emitting nuclide, e.g. 131 I, 90 Y, 177 Lu, 186 Re, or 188 Re, or gamma-ray emitting nuclides, e.g. 125 I or 123 It can be I.

[0056] According to a particular embodiment of the invention, the radionuclide is, for example, astatine-211 ( 211 At), Bismuth-212( 212 Bi), Bismuth-213( 213 Bi), Actinium-225( 225 Ac), radium-223( 223 Ra), lead-212( 212 Pb), Thorium-227( 227Th), and thorium-149 ( 149 Tb).

[0057] Radioactive nuclide actinium ( 225 Ac) is a pure alpha emitter with a half-life of 10 days. It decays via a cascade of six relatively short-lived daughter radionuclides, giving stable 209 Become Bi. 225 The primary decay pathway of Ac results in a net four alpha particles with a large cumulative energy of 28 MeV and two beta decays with maximum energies of 1.6 and 0.6 MeV. The relatively long half-life of 10 days and the multiple alpha particles generated in the rapid decay chains make Ac a 225 making Ac a highly cytotoxic radionuclide.

[0058] Thus, according to a particular aspect of the present invention, the alpha-emitting nuclide is actinium-225( 225 Ac) conjugated to a monoclonal antibody, antibody fragment, or small peptide ligand. 225 The Ac payload can deliver high-energy alpha particles directly to tumor sites, generating lethal DNA double-strand breaks without requiring significant payload accumulation within tumor cells. Due to its short path length, the range of its high-energy alpha particle emission is only a few cell diameters thick, thereby limiting damage to nearby normal tissue. Furthermore, 225 Ac-antibody conjugates offer a significant advantage over antibody-drug conjugates because they have been shown to be effective even in patients with tumors that express low levels of the target antigen. 225 This is due to the profound cytocidal effect of Ac, which is in stark contrast to antibody-drug conjugates, which require hundreds of antibody molecules to bind to their respective antigens to exert an effect on cancer cells.

[0059] Other advantages of radioactive payloads over drugs or toxins are: 1) the antibodies used for radiation delivery do not need to be internalized to kill cells, 2) due to the "multi-site" effect of radiation, not all cancer cells within a tumor need to be targeted by the antibody, and 3) in contrast to antibody-drug conjugates, radioisotopes attached to antibodies are less likely to induce significant immune responses that would limit their subsequent use. Furthermore, the work reported herein demonstrates that 225 This demonstrates the stability of Ac-labeled antibodies.

[0060] The monoclonal antibody can be labeled with an alpha-emitting nuclide by any means known in the art. According to one aspect of the invention, the radionuclide can be bound or chelated by a chelating agent that is conjugated to the monoclonal antibody.

[0061] The radionuclide-labeled monoclonal antibodies against CD38 described herein ("radiolabeled anti-CD38") can be prepared by first forming anti-CD38 conjugated with a chelator ("conjugated anti-CD38") and then chelating a radionuclide with the conjugated anti-CD38 to form the radiolabeled anti-CD38. The radionuclide can be chelated by the conjugated anti-CD38 at any time after conjugation.

[0062] When the methods described herein are used to form radiolabeled anti-CD38, the degree of chelation and conjugation is advantageously high. As used herein, the terms "degree of chelation" and "degree of conjugation" may be interpreted to mean the percentage of radionuclide bound by the chelator divided by the total chelator used in the reaction, and the percentage of chelator that is successfully conjugated to the monoclonal antibody, respectively.

[0063] The degree of conjugation in the reactions of the invention to form conjugated anti-CD38 is typically greater than 50%, greater than 70%, greater than 90%, greater than 95%, greater than about 96%, greater than about 97%, greater than about 98%, or greater than about 99%.

[0064] The degree of chelation of the radionuclide in forming the radiolabeled anti-CD38 of the reaction of the invention is typically greater than 50%, greater than 70%, greater than 90%, greater than 95%, greater than about 96%, greater than about 97%, greater than about 98%, or greater than about 99%.

[0065] According to the method for forming radiolabeled anti-CD38 described herein, a monoclonal antibody against CD38 can be dissolved in a buffer solution containing a chelator. The pH can be selected to optimize the conjugation conditions between the chelator and the antibody in the conjugation reaction mixture. The conjugation reaction mixture can contain a bicarbonate buffer or a phosphate buffer. The conjugation reaction mixture can have a pH of about 8.0 to about 9.2. For example, the conjugation reaction mixture can have a pH of about 8.0, about 8.1, about 8.3, about 8.4, about 8.5, about 8.6, about 8.7, about 8.8, about 8.9, about 9.0, about 9.1, or about 9.2. The temperature of the conjugation reaction mixture can be adjusted to promote conjugation between the chelator and the targeting moiety. For example, the conjugation reaction mixture can be incubated at room temperature or at a temperature of about 37°C. The conjugation reaction mixture may be incubated for any time sufficient to provide for conjugation, such as, for example, about 1.5 hours.

[0066] The conjugated anti-CD38 may be dissolved in a buffer solution containing a radionuclide. The pH may be selected to optimize the conditions for chelation of the radionuclide with the conjugated anti-CD38 in the chelation reaction mixture. The chelation reaction mixture may contain gentisic acid. The chelation reaction mixture may have a pH of about 5.5 to about 7.0. For example, the chelation reaction mixture may have a pH of about 5.5, about 5.6, about 5.7, about 5.8, about 5.9, about 6.0, about 6.1, about 6.2, about 6.3, about 6.4, about 6.5, about 6.6, about 6.7, about 6.8, about 6.9, or about 7.0.

[0067] The temperature of the chelation reaction mixture can be adjusted to promote chelation of the radionuclide with the conjugated anti-CD38. For example, the chelation reaction mixture can be incubated at a temperature of about 37°C. The chelation reaction mixture can be incubated for about 1.5 hours. After a period of time, the solution can be quenched by the addition of a quenching chelate (e.g., diethylenetriaminepentaacetic acid (DTPA)), and the reaction mixture can be purified. The chelation reaction mixture can be further incubated after the addition of the quenching chelate, for example, for about 30 minutes at about 37°C.

[0068] Chelators useful in the present invention are compounds that possess dual functionality: the ability to sequester metal ions as well as to covalently bind to biological carriers, such as antibodies. Numerous chelators are known in the art. Exemplary chelators suitable for use in the present invention include, but are not limited to, S-2-(4-isothiocyanatobenzyl)-1,4,7,10-tetraazacyclododecanetetraacetic acid (p-SCN-Bn-DOTA), diethylenetriaminepentaacetic acid (DTPA); ethylenediaminetetraacetic acid (EDTA); 1,4,7,10-tetraazacyclododecane-N,N',N",N'"-tetraacetic acid (DOTA); p-isothiocyanatobenzyl-1,4,7,10-tetraazacyclododecane-1,4,7, 1,4,7,10-tetraazacyclododecane-N,N',N"-triacetic acid (p-SCN-Bz-DOTA); 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetrakis(2-propionic acid) (DOTMA); 3,6,9-triaza-12-oxa-3,6,9-tricarboxymethylene-10-carboxy-13-phenyl-tridecanoic acid ("B-19036"); 1,4,7-triazacyclononane-N,N',N"-triacetic acid (NO TA); 1,4,8,11-tetraazacyclotetradecane-N,N',N'',N'"-tetraacetic acid (TETA); triethylenetetraaminehexaacetic acid (TTHA); trans-1,2-diaminohexanetetraacetic acid (CYDTA); 1,4,7,10-tetraazacyclododecane-1-(2-hydroxypropyl)-4,7,10-triacetic acid (HP-DO3A); trans-cyclohexane-diaminetetraacetic acid (CDTA); trans(1,2)-cyclohexanediethylenetriaminepentaacetic acid (CD 1,4,7,10-Tetraazacyclododecane-1,4,7,10-tetrakis{3-(4-carboxyl)-butanoic acid}; 1,4,7,10-Tetraazacyclododecane-1,4,7,10-tetrakis(acetic acid-methylamide); 1,4,7,10-Tetraazacyclododecane-1,4,7,10-tetrakis(methylenephosphonic acid); and derivatives thereof.

[0069] One or more steps can be used to separate conjugated CD38 from other components of a conjugation reaction mixture or radiolabeled anti-CD38 from other components of a chelation reaction mixture. For example, the reaction mixture can be transferred to a filtration device (e.g., a Millipore centrifugation device) with a specific molecular weight cutoff, thereby separating the conjugated anti-CD38 or radiolabeled anti-CD38 from other components of the respective reaction mixture by filtration of the reaction mixture through the filtration device. Filtration can be used to obtain conjugated anti-CD38 or radiolabeled anti-CD38 with a purity of at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85%, at least about 90%, at least about 95%, at least about 97%, at least about 98%, at least about 99%, or at least about 99.5%.

[0070] According to certain aspects of the invention, the yield of conjugated anti-CD38 or radiolabeled anti-CD38 from separation (e.g., purification) is at least about 70%, at least about 75%, at least about 80%, at least about 85%, or at least about 90%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% of the final product.

[0071] According to one aspect of the present invention, the monoclonal antibody is first conjugated with a p-SCN-Bn-DOTA or DOTA chelator to form a conjugated anti-CD38, and the conjugated anti-CD38 is conjugated with the p-SCN-Bn-DOTA or DOTA chelator. 225 Chelation of Ac forms radiolabeled anti-CD38. Thus, according to an embodiment of the present invention, 225 Only a single step involving AC is necessary to label anti-CD38.

[0072] According to a particular aspect of the present invention, a radionuclide ( 225AC) may be chelated with a chelating agent prior to conjugation with the monoclonal antibody against CD38.

[0073] According to certain aspects of the invention, radiolabeled anti-CD38 is relatively stable. For example, more than 75% of actinium-labeled monoclonal antibodies remain stable within 4 o C for 24 hours and remain intact.

[0074] According to a particular embodiment of the present invention, the radiolabeled anti-CD38 exhibits essentially the same immunoreactivity to CD38 as a control monoclonal antibody, which comprises an unlabeled monoclonal antibody directed against the same epitope of CD38 as the actinium-labeled monoclonal antibody.

[0075] The combination of labeling efficiency, stability of radiolabeled anti-CD38, and immunoreactivity of labeled CD38 provides an effective therapeutic agent. Thus, radiolabeled anti-CD38 produced using the methods described herein can be used as a therapeutic agent. For example, radiolabeled anti-CD38 can be used as a therapeutic agent to deliver a radiation dose specifically to cells and tissues expressing CD38. Thus, according to certain aspects of the present invention, radiolabeled anti-CD38 can be formulated in solution with a pharmaceutically acceptable salt or carrier to form a pharmaceutical composition.

[0076] The pharmaceutical compositions of the present invention may also be administered in combination therapy, i.e., combined with other therapeutic agents related to the disease or condition being treated. Such administration may be simultaneous, separate, or sequential. In the case of simultaneous administration, the agents may be administered as one composition or as separate compositions, as needed.

[0077] According to certain aspects of the invention, the pharmaceutical composition may comprise one or more therapeutic agents. Exemplary therapeutic agents include chemotherapeutic agents, anti-inflammatory agents, immunosuppressive agents, immunomodulatory agents, or combinations thereof.

[0078] Therapeutic agents may be administered according to any standard dosing regimen known in the art. When therapeutic agents are included in the compositions of the invention, they are administered in a dose of 1 to 500 mg / m 2 The amount may be determined based on the patient's surface area (m 2 For example, an exemplary dose of paclitaxel is 15 mg / 2 ~275mg / m 2 an exemplary dose of docetaxel is 60 mg / m 2 ~100mg / m 2 An exemplary dose of epothilone may include 10 mg / m 2 ~20mg / m 2 An exemplary dose of calicheamicin is 1 mg / m 2 ~10mg / m 2 Although exemplary doses are listed herein, they are provided for reference only and are not intended to limit the dose ranges of the inventive agents of the present disclosure.

[0079] Thus, according to one embodiment, the pharmaceutical composition can include at least one chemotherapeutic agent. Exemplary chemotherapeutic agents include, for example, antimetabolites such as methotrexate, 6-mercaptopurine, 6-thioguanine, cytarabine, fludarabine, 5-fluorouracil, dacarbazine, hydroxyurea, asparaginase, gemcitabine, cladribine, and similar agents.

[0080] Exemplary chemotherapeutic agents include alkylating agents such as, for example, mechlorethamine, thiotepa, chlorambucil, melphalan, carmustine (BSNU), lomustine (CCNU), cyclophosphamide, busulfan, dibromomannitol, streptozotocin, dacarbazine (DTIC), procarbazine, mitomycin C, other platinum derivatives such as cisplatin and carboplatin, and similar agents.

[0081] Exemplary chemotherapeutic agents include antibiotics such as, for example, dactinomycin (formerly actinomycin), bleomycin, calicheamicin, daunorubicin (formerly daunomycin), doxorubicin, idarubicin, mithramycin, mitomycin, mitoxantrone, plicamycin, anthramycin (AMC), and similar agents.

[0082] Exemplary chemotherapeutic agents include taxanes, such as docetaxine and paclitaxel, and mitotic inhibitors, such as vinca alkaloids, eg, vindesine, vincristine, vinblastine, and vinorelbine.

[0083] Exemplary chemotherapeutic agents include topoisomerase inhibitors such as topotecan.

[0084] Exemplary chemotherapeutic agents include, for example, growth factor inhibitors such as inhibitors of ErBb1 (EGFR) (such as gefitinib (Iressa®), cetuximab (Erbitux®), erlotinib (Tarceva®), HuMax-EGFr (2F8 disclosed in WO 2002 / 100348) and similar agents), inhibitors of ErBb2 (Her2 / neu) (such as transtuzumab (Herceptin®) and similar agents). In one embodiment, such growth factor inhibitors may be farnesyltransferase inhibitors such as SCH-66336 and R115777. In one embodiment, such growth factor inhibitors may be vascular endothelial growth factor (VEGF) inhibitors such as bevacizumab (Avastin®).

[0085] Exemplary chemotherapeutic agents include tyrosine kinase inhibitors such as imatinib (Glivec, Gleevec ST1571), lapatinib, PTK787 / ZK222584 and similar agents.

[0086] Exemplary chemotherapeutic agents include histone deacetylase inhibitors, such as hydroxamic acid-based hybrid compounds, such as SAHA (sabroanilide hydroxamic acid).

[0087] Exemplary chemotherapeutic agents include P38a MAP kinase inhibitors, such as SCIO-469.

[0088] Exemplary chemotherapeutic agents include inhibitors of angiogenesis, neovascularization, and / or other angiogenesis. Examples of such inhibitors include urokinase inhibitors, matrix metalloproteinase inhibitors (such as marimastat, neovastat, BAY 12-9566, AG 3340, BMS-275291, and similar agents), inhibitors of endothelial cell migration and proliferation (such as TNP-470, squalamine, 2-methoxyestradiol, combretastatin, endostatin, angiostatin, penicillamine, SCH66336 (Schering-Plough Corp, Madison, NJ), R115777 (Janssen Pharmaceutica, Inc., Titusville, NJ) and similar agents), antagonists of angiogenic growth factors (ZD6474, SU6668, antibodies to angiogenic agents and / or their receptors (such as VEGF, bFGF, and angiopoietin-1), thalidomide (Thalomid®), thalidomide derivatives (such as CC-5013 (lenalidomide, Revlimid™) and CC4047 (Actimid™), Sugen 5416, SU5402, anti-angiogenic ribozymes (such as angiozyme), interferon alpha (such as interferon alpha 2α), suramin and similar agents), VEGF-R kinase inhibitors and other anti-angiogenic tyrosine kinase inhibitors (such as SU011248), inhibitors of endothelial-specific integrin / survival signaling (such as vitaxin and similar agents), copper antagonists / chelators (such as tetrathiomolybdate, captopril, and similar agents), carboxyamidotriazole (CAI), ABT-627, CM101, interleukin-12 (IL-12), IM862, PNU145156E, and nucleotide molecules that inhibit angiogenesis (antisense-VEGF-cDNA, cDNA encoding angiostatin, cDNA encoding p53, and cDNA encoding defective VEGF receptor-2) and similar agents.

[0089] Other examples of such inhibitors of angiogenesis, neovascularization, and / or other angiogenesis are anti-angiogenic heparin derivatives and related molecules (e.g., heparinase III), tetozolomide, NK4, macrophage migration inhibitory factor (MIF), cyclooxygenase-2 inhibitors, inhibitors of hypoxia inducible factor 1, anti-angiogenic soy isoflavones, oltipraz, fumagillin and its derivatives, somatostatin derivatives, pentosan polysulfate, tecogalan sodium, dalteparin, tumstatin, thrombospondin, NM-3, combrestatin, canstatin, avastatin, antibodies against other related targets (such as anti-α-v / β-3 integrin anti-kininostatin mAb), and similar agents.

[0090] Exemplary chemotherapeutic agents include thalidomide (Thalomid®), thalidomide analogs (CC-5013 (lenalidomide, Revlimid™) and / or CC4047 (Actimid™).

[0091] Exemplary chemotherapeutic agents may include agents such as additional antibody therapeutics or proteasome inhibitors, e.g., bortezomib (Velcade®), corticosteroids, e.g., prednisone, prednisolone, dexamethasone, etc., bisphosphonates, etc. Examples of potentially suitable bisphosphonates are pamidronate (Aredia®), zoledronic acid (Zometa®), clodronate (Bonefos®), risedronate (Actonel®), ibandronate (Boniva®), etidronate (Didronel®), alendronate (Fosamax®), tiludronate (Skelid®), incadronate (Yamanouchi Pharmaceutical), and minodronate (YM529, Yamanouchi).

[0092] Exemplary chemotherapeutic agents include colony-stimulating factors. Examples of suitable colony-stimulating factors are granulocyte colony-stimulating factors (G-CSFs) such as filgrastim (Neupogen®) and pegfilgrastim (Neulasta®), and granulocyte-macrophage colony-stimulating factors (GM-CSFs) such as sargramostim (Leukine®).

[0093] Exemplary chemotherapeutic agents include erythropoietic agents. Examples of suitable erythropoietic agents are erythropoietin (EPO), such as epoetin alfa (e.g., Procrit®, Epogen®, and Eprex®) and epoetin beta (e.g., NeoRecormon®), and erythropoiesis-stimulating protein (e.g., Aranesp®).

[0094] Exemplary chemotherapeutic agents include anti-anergy agents (eg, small molecule compounds, proteins, glycoproteins, or antibodies that break tolerance to tumor and cancer antigens).

[0095] Exemplary chemotherapeutic agents include viruses, viral proteins, and the like. Replication-deficient viruses, which typically are capable of one or only a few rounds of replication in vivo and are targeted to tumor cells, for example, may be useful components of such compositions and methods. Such viral agents may include or be associated with nucleic acids encoding immune stimulators such as GM-CSF and / or IL-2. Both naturally occurring oncolytic viruses and recombinant oncolytic viruses such as HSV-1 virus, reovirus, replication-deficient and replication-competent adenoviruses, and the like, may be useful components of such methods and compositions.

[0096] According to another embodiment, the pharmaceutical composition may include an anti-inflammatory agent, which may be selected from steroidal drugs and NSAIDs (non-steroidal anti-inflammatory drugs). Other anti-inflammatory agents include aspirin and other salicylates, Cox-2 inhibitors (such as rofecoxib and celecoxib), NSAIDs (such as ibuprofen, fenoprofen, naproxen, sulindac, diclofenac, piroxicam, ketoprofen, diflunisal, nabumetone, etodolac, oxaprozin, and indomethacin), anti-IL6R antibodies, anti-IL8 antibodies, anti-IL15 antibodies, anti-IL15R antibodies, anti-CD4 antibodies, anti-CD11a antibodies (e.g., efavirenz). anti-α4 / β-1 integrin (VLA4) antibodies (e.g., natalizumab), CTLA4-1g for the treatment of inflammatory diseases, disease-modifying antirheumatic drugs (DMARDs) such as prednisolone, prednisone, methotrexate, sulfasalazine, pyrimidine synthesis inhibitors (e.g., leflunomide), IL-1 receptor blockers (e.g., anakinra), TNF-α blockers (e.g., etanercept, infliximab, and adalimumab), and similar agents.

[0097] In another embodiment, the pharmaceutical composition may comprise at least one immunosuppressant and / or immunomodulatory agent for administration to a subject in need thereof, including cyclosporine, azathioprine, mycophenolic acid, mycophenolate mofetil, corticosteroids such as prednisone, methotrexate, gold salts, sulfasalazine, antimalarials, brequinar, leflunomide, mizoribine, 15-deoxyspergualin, 6-mercaptopurine, cyclophosphamide, rapamycin, tacrolimus (FK-506), OKT3, antithymocyte globulin, thymopentin, thymosin-α, and similar agents.

[0098] The additional immunosuppressant and / or immunomodulatory agent may be selected from an antibody that binds to p75 of the IL-2 receptor, or an immunosuppressant antibody, such as, for example, an antibody that binds to MHC, CD2, CD3, CD4, CD7, CD28, B7, CD40, CD45, IFNγ, TNF-α, IL-4, IL-5, IL-6R, IL-6; IGF, IGFR1, IL-7, IL-8, IL-10, CD11a, or CD58, or antibodies that bind to their ligands. The additional immunosuppressant and / or immunomodulatory agent may be selected from soluble IL-15R, IL-10, B7 molecules (B7-1, B7-2, variants and fragments thereof, ICOS, and OX40), inhibitors of negative T cell regulators (such as antibodies against CTLA4), and similar agents.

[0099] According to certain aspects of the invention, the one or more therapeutic agents include an anti-myeloma agent. Exemplary anti-myeloma agents include dexamethasone, melphalan, doxorubicin, bortezomib, lenalidomide, prednisone, carmustine, etoposide, cisplatin, vincristine, cyclophosphamide, cisplatin, and thalidomide, some of which are identified above as chemotherapeutic, anti-inflammatory, or immunosuppressive agents.

[0100] According to certain aspects of the present invention, the pharmaceutical composition may further comprise a non-radiolabeled monoclonal antibody directed against an epitope of CD38. The non-radiolabeled monoclonal antibody may include daratumumab, MOR202, or SAR650984. The non-radiolabeled monoclonal antibody may be directed against the same epitope of CD38 as the actinium-labeled monoclonal antibody. The amount of non-radiolabeled anti-CD38 included in the pharmaceutical composition may vary depending on the exact nature of the disease being treated, the age and weight of the patient, the identity of the monoclonal antibody directed against CD38, and the radionuclide selected to label the monoclonal antibody.

[0101] According to certain aspects of the invention, the pharmaceutical composition may comprise a non-radiolabeled monoclonal antibody in an amount at least equal to the amount of radiolabeled anti-CD38 in the pharmaceutical composition, or in an amount at least two-fold greater, e.g., at least four-fold greater, or at least ten-fold greater, than the amount of radiolabeled anti-CD38 in the pharmaceutical composition.

[0102] According to certain aspects of the invention, radiolabeled anti-CD38 may be included in the compositions or pharmaceutical compositions detailed herein at 0.1 ug / ml or less, such as 0.06 ug / ml or less, or 0.04 ug / ml or less, or even 0.02 ug / ml or less.

[0103] When included in such doses, the compositions or pharmaceutical compositions detailed herein may cause greater than 50% cell death of lymphoblasts or myeloma cells within 72 hours of administration, or 70% cell death of lymphoblasts or myeloma cells within 96 hours of administration, or 90% cell death of lymphoblasts or myeloma cells within 96 hours of administration, or even 95% cell death of lymphoblasts or myeloma cells within 96 hours of administration.

[0104] According to certain aspects of the invention, the actinium-labeled monoclonal antibody may be at least 5-fold more effective in causing lymphoblast or myeloma cell death than a control monoclonal antibody, which comprises an unlabeled monoclonal antibody directed to the same epitope of CD38 as the actinium-labeled monoclonal antibody. For example, the actinium-labeled monoclonal antibody may be at least 10-fold more effective, at least 20-fold more effective, at least 50-fold more effective, or at least 100-fold more effective in causing lymphoblast or myeloma cell death than the control monoclonal antibody.

[0105] Maximum lysis by CDC and CDCC using the currently commercially available unlabeled anti-CD38 therapeutic, Darzalex®, has been reported in the literature to occur at 1 μg / ml and 0.10 μg / ml, respectively. As noted above, actinium-labeled monoclonal antibodies can be at least five times more effective in causing cell death. Without wishing to be bound by theory, such an increase in the cytotoxicity of radiolabeled anti-CD38 may involve cell death via additional mechanisms, such as apoptosis.

[0106] The pharmaceutical compositions of the invention can be used to treat hematological malignancies, or to inhibit the growth and / or proliferation of cells that express CD38, or to treat diseases or disorders involving cells that express CD38. Accordingly, the present invention provides methods for treating a subject with a hematological malignancy, for inhibiting the growth and / or proliferation of cells that express CD38, and for treating diseases or disorders involving cells that express CD38, comprising administering to the subject a pharmaceutical composition as detailed hereinabove.

[0107] According to certain aspects of the invention, the hematological malignancy is multiple myeloma. According to certain aspects of the invention, the cells expressing CD38 are multiple myeloma cells. According to certain aspects of the invention, the disease or disorder may be multiple myeloma.

[0108] According to certain aspects of the invention, the CD38-expressing cells are CD38-expressing cancer cells or CD38-expressing T cells, B cells, NK cells, or plasma cells. According to certain aspects of the invention, the CD38-expressing cells include solid tumor cells or hematological malignancy cells. Exemplary hematological malignancy cells include multiple myeloma cells, acute lymphocytic leukemia cells, acute myeloid leukemia cells, chronic lymphocytic leukemia cells, chronic myeloid leukemia cells, Hodgkin's lymphoma cells, non-Hodgkin's lymphoma cells, T-LGL leukemia cells, NK cell leukemia cells, or hairy cell leukemia cells.

[0109] As described above, the pharmaceutical composition may be administered either alone or in combination with one or more additional therapeutic agents. The pharmaceutical composition may include one or more additional therapeutic agents. The pharmaceutical composition may be administered in a dosing regimen comprising at least one dose.

[0110] According to certain embodiments of the present invention, at least one dose of the dosing regimen comprises an amount of monoclonal antibody against CD38 that is 5-fold lower than that of a dose in a dosing regimen comprising only a control monoclonal antibody. According to certain embodiments of the present invention, at least one dose of the dosing regimen comprises an amount of monoclonal antibody against CD38 that is 10-fold lower than that of a dose in a dosing regimen comprising only a control monoclonal antibody. According to certain embodiments of the present invention, at least one dose of the dosing regimen comprises an amount of monoclonal antibody against CD38 that is 20-fold lower than that of a dose in a dosing regimen comprising only a control monoclonal antibody. According to certain embodiments of the present invention, at least one dose of the dosing regimen comprises an amount of monoclonal antibody against CD38 that is 50-fold lower than that of a dose in a dosing regimen comprising only a control monoclonal antibody. According to certain embodiments of the present invention, at least one dose of the dosing regimen comprises an amount of monoclonal antibody against CD38 that is 100-fold lower than that of a dose in a dosing regimen comprising only a control monoclonal antibody. The control monoclonal antibody typically comprises an unlabeled monoclonal antibody directed against the same epitope of CD38 as the actinium-labeled monoclonal antibody.

[0111] According to certain embodiments of the invention, the dosing regimen includes at least one lower dose than a control dosing regimen, which includes the administration of an unlabeled monoclonal antibody directed to the same epitope of CD38 as the actinium-labeled monoclonal antibody, either alone or in combination with one or more additional therapeutic agents. According to certain embodiments of the invention, the dosing regimen includes at least two lower doses than a control dosing regimen, which includes the administration of an unlabeled monoclonal antibody directed to the same epitope of CD38 as the actinium-labeled monoclonal antibody, either alone or in combination with one or more additional therapeutic agents.

[0112] According to certain aspects of the invention, the dosing regimen may include a dose that is 10% lower than the control dosing regimen, e.g., a dose that is 20% lower than the control dosing regimen, or a dose that is 30% lower, or a dose that is 40% lower, or a dose that is 50% lower, or a dose that is 60% lower, or a dose that is 70% lower, or a dose that is 80% lower, or even a dose that is 90% lower.

[0113] According to certain embodiments of the invention, the control monoclonal antibody may comprise daratumumab administered at a dose of about 16 mg / kg patient weight. According to certain embodiments of the invention, the control dosing regimen may comprise 8 doses administered at one dose per week for at least 8 weeks.

[0114] According to certain aspects of the present invention, the therapeutically effective dose of a radiolabeled anti-CD38 antibody can be 0.1 μg / kg to 1 mg / kg, for example, 1 μg / kg to 1 mg / kg, or 10 μg / kg to 1 mg / kg, or 100 μg / kg to 1 mg / kg, or 0.1 μg / kg to 100 μg / kg, or 0.1 μg / kg to 50 μg / kg, or 0.1 μg / kg to 10 μg / kg, or 0.1 μg / kg to 40 μg / kg, or 1 μg / kg to 40 μg / kg.

[0115] These protein doses may contain radioactivity concentrations of 0.1 uCi / kg to 5 uCi / kg, for example, 0.1 uCi / kg to 4 uCi / kg, or 0.1 uCi / kg to 3 uCi / kg, or 0.1 uCi / kg to 2 uCi / kg, or 0.1 uCi / kg to 1 uCi / kg, or 0.2 uCi / kg to 5 uCi / kg, or 0.5 uCi / kg to 5 uCi / kg.

[0116] The therapeutically effective dose of the radiolabeled anti-CD38 antibody may be administered in a single dose or as two equally divided doses, the second dose being administered 1 to 10 days, e.g., 3 to 8 days, or 4 to 7 days, or 5 to 8 days, after the first dose.

[0117] According to one aspect, the present invention provides a method for treating cancer in a subject, the method comprising administering to the subject a therapeutically effective amount of the pharmaceutical composition detailed hereinabove. Without wishing to be bound by any particular theory, based on the immunomodulatory effects observed with the anti-CD38 monoclonal antibodies described herein, such as daratumumab, MOR202, or SAR650984, they may be effective in treating solid tumors. Accordingly, the present invention also provides a method for treating a patient with a solid tumor, comprising administering to a patient in need thereof a therapeutically effective amount of an antibody that specifically binds to CD38 for a time sufficient to treat the solid tumor.

[0118] According to certain aspects, the present invention provides articles of manufacture. For example, an article of manufacture according to aspects of the present invention may include (a) a radiolabeled monoclonal antibody against CD38 and (b) labeling instructing a user to administer to a subject an amount of the antibody effective to treat a disease or disorder involving cells expressing CD38. The monoclonal antibody may comprise any of the antibodies described herein and may be labeled with any of the radionuclides described herein. For example, the monoclonal antibody may be any of daratumumab, MOR202, or SAR650984. According to one aspect of the present invention, the article of manufacture may include actinium ( 225 According to a further embodiment of the article of manufacture, the amount of antibody effective to treat a disease or disorder involving cells expressing CD38 may be between 10 μCi and 600 μCi. 225 Ac-labeled daratumumab, for example, 10 μCi to 400 μCi 225 Ac-labeled daratumumab, or 10 μCi to 200 μCi 225 Contains Ac-labeled daratumumab.

[0119] According to certain aspects of the invention, an article of manufacture can include (a) a pharmaceutical composition according to any of the aspects described herein and (b) labeling instructing a user to administer to a subject an amount of the antibody effective to treat a disease or disorder involving cells expressing CD38. For example, the pharmaceutical composition can include a radiolabeled fraction of a monoclonal antibody against CD38 and an unlabeled fraction of the same or a different monoclonal antibody against CD38. The pharmaceutical composition can be provided in a patient-specific form, e.g., in a therapeutically effective dose with an amount of radioactivity and protein concentration adjusted for a particular patient (e.g., based on patient characteristics such as weight, sex, age, type and progression of disease, etc.). According to certain embodiments, the therapeutically effective dose is 0.1 uCi / kg to 5 uCi / kg, for example, 0.1 uCi / kg to 4 uCi / kg, or 0.1 uCi / kg to 3 uCi / kg, or 0.1 uCi / kg to 2 uCi / kg, or 0.1 uCi / kg to 1 uCi / kg, or 0.2 uCi / kg to 5 uCi / kg, or 0.5 uCi / kg to 5 uCi / kg. The radiolabeled anti-CD38 antibody may be present in an amount of 1 mg / kg to 1 mg / kg, for example, 1 μg / kg to 1 mg / kg, or 10 μg / kg to 1 mg / kg, or 100 μg / kg to 1 mg / kg, or 0.1 μg / kg to 100 μg / kg, or 0.1 μg / kg to 50 μg / kg, or 0.1 μg / kg to 10 μg / kg, or 0.1 μg / kg to 40 μg / kg, or 1 μg / kg to 40 μg / kg.

[0120] In a particular aspect, the present invention provides a pharmaceutical composition useful for treating a disease or disorder involving cells expressing CD38. The composition comprises 5-50% by weight of actinium ( 225 The pharmaceutical composition may comprise a monoclonal antibody against CD38 labeled with Ac), 50 to 95% by weight of a monoclonal antibody against unlabeled CD38, and a pharmaceutically acceptable carrier. 225 The unlabeled monoclonal antibody may be Ac-daratumumab, and may be the same as or different from the labeled antibody, such as daratumumab. [Example]

[0121] Example 1: Radiolabeling of Daratumumab Daratumumab (DARA) manufactured by Janssen Biotech (USA) was purchased from the pharmacy at Montefiore Medical Center, New York, USA. Isotype control human IgG1 was purchased from Creative Diagnostics (New York, USA). 225 AC was obtained from Oak Ridge National Laboratory, USA. Indium chloride 111 Indium-111 ( 111 In) was purchased from MDS Nordion (Vancouver, BC, Canada). The CD38-positive lymphoma cell line Daudi was procured from ATCC (Manassas, USA), and the CD38-positive multiple myeloma cell lines KMS-28BM and KMS-28PE were procured from XenoTech (Japan). The Daudi cell line was grown in RPMI, 10% FBS, and an antibiotic cocktail, while the KMS-28BM and KMS-28PE cell lines were grown in RPMI supplemented with 10% FBS. The bifunctional chelator p-SCN-Bn-DOTA (referred to as DOTA in these examples) was purchased from Macrocyclics (Texas, USA).

[0122] Referring to Figures 1A and 1B, DOTA was conjugated to DARA in ammonium acetate buffer at 37°C for 1.5 hours at a 5M excess (Figure 1A). The DARA-DOTA conjugate was purified to a specific activity of 400 nCi to 0.3 µg. 225 Ac or 111 Label with In 225 Ac-DARA was generated (Fig. 1B). 225 Ac-DARA was diluted with an equal amount of unlabeled DARA to give 200 uCi 225 The total antibody dose (0.3 μg) was adjusted to the Ac-DARA dose, and the sample was then purified to a radiochemical purity of 99±1 using disposable spin columns.

[0123] Figure 2 shows the effect of DOTA-DARA immunoconjugate 225 A bar graph showing the degree of chelation of Ac is provided. The conjugation reaction was o The DOTA-DARA immunoconjugate was incubated at 0.3 mg / ml in 0.15 M ammonium acetate buffer (pH 6.5) for 37 min at 10 C or room temperature with various molar excesses of DOTA chelator relative to DARA. o C or room temperature for 60 minutes at 1uCi / ug daratumumab in 0.01M HCl 225 Labeled with Ac (DOTA 225 chelation of Ac).

[0124] 225 The stability of AC-DARA was measured by storage at two different temperatures. Figure 3A shows the stability of AC-DARA at various temperatures. 225 4. Ac-DARA Sample o The samples were stored at 37°C. o Figure 3B shows the results of various conjugation reactions of DOTA and DARA carried out at room temperature with various molar excesses of DOTA relative to DARA. 225 The storage stability of Ac-DARA samples at room temperature was shown. o The conjugation reaction of DOTA with DARA was performed at various molar excesses of DOTA over DARA at either 4 °C or room temperature. Prior to the start of the stability experiments, samples were purified to 99 ± 1% with disposable spin columns. At both temperatures, more than 75% of the actinium-labeled monoclonal antibody remained stable at 4 °C. o C for 24 hours, and over 60% of actinium-labeled monoclonal antibodies remain intact after 24 hours at room temperature.

[0125] Example 2: 225 Ac-DARA immunoreactivity As shown in Figure 4A, even if o C or from the conjugation reaction of DOTA with DARA carried out at room temperature with various molar excesses of DOTA relative to DARA. 225 Even Ac-DARA conjugates showed no activity against CD38 at various concentrations.225 The binding of the Ac-DARA conjugate is essentially unchanged compared to unlabeled anti-CD38.

[0126] Complement component 1q (C1q) binding to daratumumab conjugated with DARA and DOTA was assessed using an ELISA binding assay. As shown in Figure 4B, conjugation of DARA to DOTA does not affect complement binding. High-binding 96-well plates (Corning) were coated overnight at 4°C with various concentrations of antibody in coating buffer (100 mM sodium carbonate, pH 9.6). After each incubation, sample wells were washed three times with PBST (0.05% Tween 20 / PBS, pH 7.4). After coating, the plates were blocked with blocking buffer (0.1% BSA / PBST) for 1 hour at room temperature and then replaced with 2 μg / mL human C1q in blocking buffer. After 1 hour, the wells were washed three times and 100 μL of 1 μg / mL anti-hC1q HRP in blocking buffer was added. After 1 hour, the wells were washed and 100 μL of TMB substrate (Pierce, Rockford, IL) was added. After 15 minutes, the reaction was stopped with 100 μL of 1 M HCl, and the absorbance was read at 450 nm using a Spectra MAX 250 plate reader (Molecular Devices, San Jose, CA).

[0127] The ability of DARA and DARA conjugated to DOTA to mediate ADCC was assessed using the ADCC Reporter Bioassay, Complete Kit (Raji) purchased from Promega (Madison, WI) and used according to the manufacturer's protocol. As shown in Figure 4C, various concentrations of DARA and DARA-DOTA were added to target cells expressing CD38. Effector cells that express luciferase when activated by FcγR(III) were added to the target cells, and luminescence was measured after the addition of Bio-Glow™. Fold induction = RLU (background induction) / RLU (background control without antibody). An anti-CD20 antibody, known to activate ADCC, was used as a positive control. As shown, conjugation of DARA to DOTA does not inhibit ADCC.

[0128] Example 3: 225 Cytotoxicity of Ac-DARA Naked DARA, 225 Ac-DARA and unrelated IgG- 225 The ability of Ac to induce cytolysis in multiple myeloma cell lines with a range of CD38 antigen expression levels was assessed at various time points and concentrations.

[0129] Daudi cells (2 x 10 in 1 mL PBS) 5 ) into a BSA-blocked Eppendorf tube, 225Cells were treated in triplicate with AC-DARA, a control human IgG of the corresponding activity, or unlabeled DARA at 0, 20, 40, 60, and 100 nCi / sample. The total amount of DARA per mL of sample was 0.02 μg for the 20 nCi sample, 0.04 μg for the 40 nCi sample, and 0.06 μg for the 60 nCi sample and the corresponding unlabeled DARA sample. After 48, 72, and 96 hours of incubation, the effect of the radiolabeled antibody on the cells was assessed using the tetrazolium dye (2,3)-bis-(2-methoxy-4-nitro-5-sulfenyl)-(2H)-talazolium-5-carboxanilide (XTT) assay. Wells were washed, and fresh medium was added along with 50 μl XTT (Sigma) at 1 mg / mL in PBS and 4 μl menadione (Sigma) at 1 mM in acetone. Cells were incubated for an additional 3 hours and the absorbance was read at 492 nm. All conditions were performed in triplicate. In parallel, killing was assessed by trypan blue staining.

[0130] Figures 5A to 5D show Daudi cells. 225 1 shows the results of an XTT assay treated with Ac-DARA conjugate. 225 It can be seen that the Ac-DOTA-IgG conjugate lyses Daudi cells to 95%–100% lysis, whereas the control DARA has little or no effect (i.e., the same cytotoxicity as the control cells) at the low concentrations tested in this assay. Figures 5A–5D show the results for exposure times of 24, 48, 72, and 96 hours, respectively.

[0131] Figure 6 provides a summary graph of the data seen in Figures 5A-5D, showing the results of various concentrations of 225 Figures 7A and 7B show the percent cell lysis over time for Ac-DARA. 225 1 provides a graph showing the ability of Ac-DARA conjugates to induce lysis of 28BM and 28PE multiple myeloma cells by ADCC, respectively, over a 96-hour period.

[0132] 8A to 8C show various concentrations of 225Bar graphs comparing the cytolysis results of Daudi, 28BM, and 28PE cells upon exposure to Ac-DARA conjugates are provided, with panels 8A-8C showing exposure times of 48, 72, and 96 hours, respectively.

[0133] The important thing is that DOTA and 225 As shown in the figure, immunogenicity was maintained when DARA was labeled with Ac. 225 Concentrations of Ac-labeled constructs were tested in each of the four cell lines. The activity:antibody ratio used was 10 nCi:0.01 μg / mL. 225 Treatment of Daudi cells with Ac-DARA resulted in 25% and 95% cell death, respectively, at 72 hours, compared with 5-6% cell death in the daratumumab-only group. Similar time- and concentration-dependent cell death was observed in the patient-derived cell lines 28PE and 28BM. CD38-positive cell lines were treated with a radiolabeled isotype-matched human control antibody, IgG- 225 When treated with Ac or negative CD38-expressing multiple myeloma cell line U266 225 No time- or concentration-dependent killing was observed when treated with Ac-DARA.

[0134] These results suggest that antibodies targeting CD38 225 This demonstrates the ability to label with Ac, a strong alpha emitter. 225 AC improves the efficacy of DARA by more than 10-fold, essentially utilizing antibodies as a vehicle while still maintaining the immune function of DARA. 225 AC is an effective payload due to its high linear energy transfer properties and short path length in tissue.

[0135] Example 4: Animal Model in Daudi tumor-bearing mice 111Imaging of In-DARA by microSPECT / CT was performed to confirm its specific uptake in tumors (Figure 9). For tumor induction, female SCID mice (CB17 / Icr-Prkdc mice procured from Charles River Laboratories) were used. scid / IcrIcoCrl) in 50 μL saline to the right flank. 6 Daudi cells were injected subcutaneously. Tumor growth was measured every 3 days with an electronic caliper and calculated using the formula V = 0.5(L × W 2 ) to calculate the tumor volume, and the width was taken as the tumor short diameter. On the 14th day after tumor cell inoculation, the tumor grew to approximately 200 mm 3 When the mean volume of 1000 mg / kg was reached, the mice were imaged by microSPECT / CT or 111 For imaging, three mice were treated with 400 μCi of In-DARA. 111 In-DARA was injected intraperitoneally (IP), and subjects were imaged by microSPECT / CT (MI Labs, Netherlands) at 1, 4, and 24 hours, followed by 2, 3, 7, and 10 days after injection.

[0136] MicroSPECT / CT images of tumors 24 hours after administration 111 It showed significant localization of In-DARA. 111 DARA remained concentrated in tumors for up to 10 days, while activity disappeared from the rest of the body. Figure 9 shows representative images from one mouse over the entire time course.

[0137] To evaluate the side effects of radioimmunotherapy (RIT), tumor-bearing mice were randomized into groups of 5 animals and administered 100 μL of saline, or unlabeled DARA at concentrations of 10 or 0.3 μg per mouse, or 400 nCi and 200 nCi per 0.3 μg antibody. 225 Mice were observed for tumor progression for 40 days, and tumors were observed to grow to 4,000 mm 3 Any mice that reached a volume of 0.05 or developed necrosis were humanely euthanized (FIGS. 10A and 10B).

[0138] To evaluate the side effects of RIT, mice were monitored for body weight (Figure 11A) and evaluated for hematological parameters and systemic toxicity (kidney and liver, Figure 11B). 225 A comprehensive safety evaluation of Ac-DARA was performed. 225 The body weight of mice in the Ac-DARA group remained unchanged throughout the experiment, while the 400 nCi group showed a slight, transient weight loss during the second week of treatment, with a rapid recovery observed by the third week of treatment. Evaluation of hematological and toxicological parameters revealed no significant changes in any of the parameters tested. 225 It was also shown that there was no statistically significant difference between the Ac-labeled group and the non-labeled group. 225 This suggests that the presence of the Ac conjugate did not affect hematological parameters or show evidence of increased liver damage in treated mice as measured by aspartate aminotransferase (AST) and alanine transminase (ALT) levels. Furthermore, no changes in creatinine and blood urea nitrogen (BUN) were observed, indicating a lack of overt nephrotoxicity.

[0139] The present application discloses the following aspects.

[0140] Aspect 1. A method of treating a disease or disorder involving cells expressing CD38, comprising administering to a subject an effective amount of a pharmaceutical composition comprising a monoclonal antibody against CD38, wherein the monoclonal antibody is an actinium ( 225 Ac), methods labeled.

[0141] Embodiment 2. The method of embodiment 1, wherein the monoclonal antibody comprises a human or humanized immunoglobulin (IgG1) against CD38.

[0142] Embodiment 3. The method of embodiment 1 or 2, wherein the monoclonal antibody comprises daratumumab, MOR202, or SAR650984.

[0143] Aspect 4. The method of any one of Aspects 1 to 3, wherein the cell expressing CD38 is a CD38-expressing cancer cell, or a CD38-expressing T cell, B cell, NK cell, or plasma cell.

[0144] Embodiment 5. The method of any one of embodiments 1 to 4, wherein the cell expressing CD38 comprises a solid tumor cell or a hematological malignancy cell.

[0145] Embodiment 6. The method of embodiment 5, wherein the hematological malignancy cell comprises a multiple myeloma cell, an acute lymphocytic leukemia cell, an acute myeloid leukemia cell, a chronic lymphocytic leukemia cell, a chronic myeloid leukemia cell, a Hodgkin's lymphoma cell, a non-Hodgkin's lymphoma cell, a T-LGL leukemia cell, an NK cell leukemia cell, or a hairy cell leukemia cell.

[0146] Embodiment 7. The method of any one of embodiments 1 to 6, wherein the pharmaceutical composition further comprises a non-radiolabeled monoclonal antibody directed against an epitope of CD38.

[0147] Embodiment 8. The method of any one of embodiments 1 to 7, wherein the pharmaceutical composition further comprises a non-radiolabeled monoclonal antibody directed against the same epitope of CD38 as the actinium-labeled monoclonal antibody.

[0148] Aspect 9. The effective amount is 0.1 uCi / kg to 5 uCi / kg of actinium ( 225 9. The method of any one of aspects 1 to 8, comprising a dose of antibody from 0.1 μg / kg to 1 mg / kg, including 0.1 μg / kg to 50 μg / kg, including 0.1 μCi / kg to 1 μCi / kg.

[0149] Aspect 10. The method of any of aspects 1-9, wherein the effective amount is administered as a single dose, or the effective amount is administered as two equally divided doses, the second dose being administered 3-8 days after the first dose.

[0150] Embodiment 11. The method of any one of embodiments 1 to 10, further comprising administering one or more additional therapeutic agents.

[0151] Aspect 12. The method of aspect 11, wherein the administration of the one or more additional therapeutic agents precedes or follows the administration of the pharmaceutical composition.

[0152] Aspect 13. The method of any of Aspects 11 or 12, wherein the one or more additional therapeutic agents comprise a chemotherapeutic agent, an anti-inflammatory agent, an immunosuppressant, an immunomodulatory agent, or a combination thereof.

[0153] Embodiment 14. The method of any of embodiments 11-13, wherein the one or more additional therapeutic agents comprise an anti-myeloma agent, such as dexamethasone, melphalan, doxorubicin, bortezomib, lenalidomide, prednisone, carmustine, etoposide, cisplatin, vincristine, cyclophosphamide, and thalidomide.

[0154] Embodiment 15. The method of any one of embodiments 1 to 14, wherein the pharmaceutical composition further comprises one or more additional therapeutic agents.

[0155] Embodiment 16 The method of any of embodiments 1 to 15, wherein the pharmaceutical composition is administered to the subject in a dosing regimen comprising at least one dose.

[0156] Aspect 17. The method of aspect 16, wherein at least one dose comprises an amount of monoclonal antibody against CD38 that is 5-fold lower than the dose in a dosing regimen comprising only a control monoclonal antibody, or 10-fold lower than the dose in a dosing regimen comprising only the control monoclonal antibody, or 20-fold lower than the dose in a dosing regimen comprising only the control monoclonal antibody, or 50-fold lower than the dose in a dosing regimen comprising only the control monoclonal antibody, or 100-fold lower than the dose in a dosing regimen comprising only the control monoclonal antibody, wherein the control monoclonal antibody comprises an unlabeled monoclonal antibody directed to the same epitope of CD38 as the actinium-labeled monoclonal antibody, and wherein the control monoclonal antibody comprises daratumumab administered at a dose of about 16 mg / kg patient body weight.

[0157] Aspect 18. The method of aspect 16 or 17, wherein the dosing regimen comprises at least one dose less than a control dosing regimen, or at least two doses less than the control dosing regimen, or at least three doses less than the control dosing regimen, or at least four doses less than the control dosing regimen, or at least five doses less than the control dosing regimen, wherein the control dosing regimen comprises administration of an unlabeled monoclonal antibody directed to the same epitope of CD38 as the actinium-labeled monoclonal antibody, either alone or in combination with one or more additional therapeutic agents, and wherein the control dosing regimen comprises eight doses administered in one dose per week for at least eight weeks.

[0158] Embodiment 19. An article of manufacture comprising: (a) a radiolabeled monoclonal antibody against CD38; and (b) labeling instructing a user to administer to a subject an amount of the antibody effective to treat a disease or disorder involving cells expressing CD38.

[0159] Embodiment 20. An article of manufacture comprising: (a) a pharmaceutical composition comprising a patient-specific amount of a cell expressing CD38 or an antibody effective to treat a disorder; and (b) a label displaying information regarding the pharmaceutical composition.

[0160] Aspect 21. The monoclonal antibody is an actinium ( 225 21. The article of manufacture of aspect 19 or 20, wherein the daratumumab is labeled with Ac).

[0161] Aspect 22. The amount of antibody effective for treating a disease or disorder involving cells expressing CD38 is between 10 μCi and 600 μCi. 225 Ac-labeled daratumumab, for example, 10 μCi to 400 μCi 225 Ac-labeled daratumumab, or 10 μCi to 200 μCi 225 22. The article of manufacture of any of aspects 19 to 21, comprising Ac-labeled daratumumab.

[0162] Aspect 23. The amount of antibody effective for treating a disease or disorder involving cells expressing CD38 is between 0.1 uCi / kg and 5 uCi / kg of actinium ( 22523. The article of manufacture of any of aspects 19 to 22, comprising a dose of antibody from 0.1 μg / kg to 1 mg / kg, including 0.1 μg / kg to 50 μg / kg, including 0.1 μCi / kg to 1 μCi / kg.

[0163] Aspect 24. A pharmaceutical composition useful for treating a disease or disorder involving cells expressing CD38, the composition comprising 5-50% by weight actinium ( 225 A pharmaceutical composition comprising a monoclonal antibody against CD38 labeled with Ac), 50 to 95% by weight of a monoclonal antibody against unlabeled CD38, and a pharmaceutically acceptable carrier.

[0164] 25. Radiolabeled monoclonal antibodies 225 23. The pharmaceutical composition of aspect 22, wherein the unlabeled monoclonal antibody is Ac-daratumumab, and the unlabeled monoclonal antibody may be the same as or different from the labeled antibody, such as daratumumab.

[0165] 26. The total amount of antibody is 10 μCi to 600 μCi. 225 Ac-labeled daratumumab, for example, 10 μCi to 400 μCi 225 Ac-labeled daratumumab, or 10 μCi to 200 μCi 225 26. The pharmaceutical composition according to aspect 25, comprising Ac-labeled daratumumab.

[0166] Aspect 27. The total amount of antibody is 0.1 uCi / kg to 5 uCi / kg of actinium ( 225 27. The article of manufacture of any one of aspects 25 or 26, comprising a dose of antibody from 0.1 μg / kg to 1 mg / kg, including 0.1 μg / kg to 50 μg / kg, including 0.1 μCi / kg to 1 μCi / kg. [Sequence Listing] SEQUENCE LISTING <110> ACTINIUM PHARMACEUTICALS, INC. Seth, Sandesh Thomas, Keisha <120> TREATMENTS FOR A HEMATOLOGICAL MALIGNANCY <130> PT17-023 PCT <140> PCT / USxx / xxxxx <141> 2018-07-31 <150> 62 / 539,114 <151> 2017-07-31 <160> 1 <170> PatentIn version 3.5 <210> 1 <211> 300 <212> PRT <213> Human <400> 1 Met Ala Asn Cys Glu Phe Ser Pro Val Ser Gly Asp Lys Pro Cys Cys 1 5 10 15 Arg Leu Ser Arg Arg Ala Gln Leu Cys Leu Gly Val Ser Ile Leu Val 20 25 30 Leu Ile Leu Val Val Val Leu Ala Val Val Val Pro Arg Trp Arg Gln 35 40 45 Gln Trp Ser Gly Pro Gly Thr Thr Lys Arg Phe Pro Glu Thr Val Leu 50 55 60 Ala Arg Cys Val Lys Tyr Thr Glu Ile His Pro Glu Met Arg His Val 65 70 75 80 Asp Cys Gln Ser Val Trp Asp Ala Phe Lys Gly Ala Phe Ile Ser Lys 85 90 95 His Pro Cys Asn Ile Thr Glu Glu Asp Tyr Gln Pro Leu Met Lys Leu 100 105 110 Gly Thr Gln Thr Val Pro Cys Asn Lys Ile Leu Leu Trp Ser Arg Ile 115 120 125 Lys Asp Leu Ala His Gln Phe Thr Gln Val Gln Arg Asp Met Phe Thr 130 135 140 Leu Glu Asp Thr Leu Leu Gly Tyr Leu Ala Asp Asp Leu Thr Trp Cys 145 150 155 160 Gly Glu Phe Asn Thr Ser Lys Ile Asn Tyr Gln Ser Cys Pro Asp Trp 165 170 175 Arg Lys Asp Cys Ser Asn Asn Pro Val Ser Val Phe Trp Lys Thr Val 180 185 190 Ser Arg Arg Phe Ala Glu Ala Ala Cys Asp Val Val His Val Met Leu 195 200 205 Asn Gly Ser Arg Ser Lys Ile Phe Asp Lys Asn Ser Thr Phe Gly Ser 210 215 220 Val Glu Val His Asn Leu Gln Pro Glu Lys Val Gln Thr Leu Glu Ala 225 230 235 240 Trp Val Ile His Gly Gly Arg Glu Asp Ser Arg Asp Leu Cys Gln Asp 245 250 255 Pro Thr Ile Lys Glu Leu Glu Ser Ile Ile Ser Lys Arg Asn Ile Gln 260 265 270 Phe Ser Cys Lys Asn Ile Tyr Arg Pro Asp Lys Phe Leu Gln Cys Val 275 280 285 Lys Asn Pro Glu Asp Ser Ser Cys Thr Ser Glu Ile 290 295 300

Claims

1. In the manufacture of pharmaceuticals for treating diseases or disorders involving cells expressing CD38, The use of an effective amount of monoclonal antibody against CD38, wherein the monoclonal antibody is actinium ( 225 A use characterized by being labeled with Ac).

2. The use according to claim 1, wherein the monoclonal antibody comprises human or humanized immunoglobulin (IgG1) against CD38.

3. The use according to claim 1, wherein the monoclonal antibody comprises daratumumab, MOR202, and SAR650984.

4. The use according to claim 1, wherein the monoclonal antibody comprises daratumumab.

5. The use according to claim 1, wherein the cells expressing CD38 are CD38-expressing cancer cells or CD38-expressing T cells, B cells, NK cells, or plasma cells.

6. The use according to claim 1, wherein the cells expressing CD38 include solid tumor cells or hematological malignant tumor cells.

7. The use according to claim 6, wherein the hematological malignant tumor cells include multiple myeloma cells, acute lymphoblastic leukemia cells, acute myeloid leukemia cells, chronic lymphoblastic leukemia cells, chronic myeloid leukemia cells, Hodgkin lymphoma cells, non-Hodgkin lymphoma cells, T-LGL leukemia cells, NK cell leukemia cells, or hairy cell leukemia cells.

8. The use according to claim 1, wherein the pharmaceutical further comprises a non-radioactively labeled monoclonal antibody against the CD38 epitope.

9. The use according to claim 1, wherein the pharmaceutical further comprises a non-radioactively labeled monoclonal antibody against the same CD38 epitope as the actinium-labeled monoclonal antibody.

10. The use according to claim 9, wherein the effective amount comprises a dose of 0.1 uCi / kg to 1 mg / kg antibody containing 0.1 uCi / kg to 5 uCi / kg.

11. The use according to claim 9, wherein the effective amount comprises a dose of 0.1 uCi / kg to 50 uG / kg antibody containing 0.1 uCi / kg to 1 uCi / kg.

12. The use according to claim 10 or 11, wherein the effective amount is a single dose.

13. The use according to claim 10 or 11, wherein the effective dose is two equal divided doses, and the second dose is administered 3 to 8 days after the first dose.

14. The use according to claim 1, further comprising administering one or more additional therapeutic agents for the treatment of the disease or disorder.

15. The use according to claim 14, wherein the administration of one or more further therapeutic agents occurs before or after the administration of the pharmaceutical agent.

16. The use according to claim 14, wherein the one or more further therapeutic agents include chemotherapeutic agents, anti-inflammatory agents, immunosuppressants, immunomodulators, or combinations thereof.

17. The use according to claim 14, wherein the one or more further therapeutic agents include an antimyeloma agent.

18. The use according to claim 14, wherein the antimyeloma agent is selected from dexamethasone, melphalan, doxorubicin, bortezomib, lenalidomide, prednisone, carmustine, etoposide, cisplatin, vincristine, cyclophosphamide, and thalidomide.

19. The use according to claim 1, wherein the pharmaceutical further comprises one or more further therapeutic agents.

20. The use according to claim 1, wherein the treatment of the disease or disorder comprises administering the pharmaceutical in a dosage plan comprising at least one dose.

21. The use according to claim 20, wherein the at least one dose comprises an amount of monoclonal antibody against CD38 that is five times lower than the dose of a dosing regimen comprising only a control monoclonal antibody, or an amount of monoclonal antibody against CD38 that is ten times lower than the dose of a dosing regimen comprising only a control monoclonal antibody, or an amount of monoclonal antibody against CD38 that is twenty times lower than the dose of a dosing regimen comprising only a control monoclonal antibody, or an amount of monoclonal antibody against CD38 that is fifty times lower than the dose of the dosing regimen comprising only a control monoclonal antibody, or an amount of monoclonal antibody against CD38 that is one hundred times lower than the dose of the dosing regimen comprising only a control monoclonal antibody, wherein the control monoclonal antibody comprises an unlabeled monoclonal antibody against the same CD38 epitope as the actinium-labeled monoclonal antibody.

22. The use according to claim 21, comprising daratumumab administered at a dose of approximately 16 mg / kg patient body weight as the control monoclonal antibody.

23. The use according to claim 20, wherein the administration plan comprises at least one fewer dose than the control administration plan, or at least two fewer doses than the control administration plan, or at least three fewer doses than the control administration plan, or at least four fewer doses than the control administration plan, or at least five fewer doses than the control administration plan, and the control administration plan comprises the administration of an unlabeled monoclonal antibody against the same CD38 epitope as the actinium-labeled monoclonal antibody, either alone or in combination with one or more additional therapeutic agents.

24. The use according to claim 23, wherein the control administration plan comprises eight doses administered once a week for at least eight weeks.

25. (a) A radiolabeled monoclonal antibody against CD38, (b) A label instructing the user to administer an effective amount of the antibody to treat a disease or disorder involving cells expressing CD38, A manufactured product characterized by containing the following:

26. The aforementioned monoclonal antibody contains actinium ( 225 The manufactured product according to claim 25, which is daratumumab labeled with Ac).

27. The amount of the antibody effective in treating the disease or disorder involving cells expressing CD38 is 10 μCi to 200 μCi. 225 The manufactured product according to claim 26, comprising daratumumab labeled with Ac.

28. A pharmaceutical composition useful for treating diseases or disorders involving cells expressing CD38, 5-50% by weight of actinium ( 225 A monoclonal antibody against CD38 labeled with Ac, The monoclonal antibody against unlabeled CD38 in a concentration of 50-95% by weight, A pharmaceutically acceptable carrier, A pharmaceutical composition characterized by containing the following:

29. The aforementioned radiolabeled monoclonal antibody 225 The pharmaceutical composition according to claim 28, wherein the unlabeled monoclonal antibody is daratumumab.

30. A pharmaceutical composition for treating a disease or disorder involving cells expressing CD38, comprising a monoclonal antibody against CD38, A pharmaceutical composition in which the monoclonal antibody is labeled with actinium (225Ac).

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