Use of isatuximab in combination with other drugs for treating multiple myeloma

Abstract

The present disclosure provides methods for treating multiple myeloma comprising administering to an individual in need thereof an anti-CD38 antibody and an interleukin-2 analogue, and optionally administering natural killer (NK) cells with reduced expression or knockout of CD38.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of priority to U.S. Provisional Application No. 63 / 317,253, filed on March 7, 2022, the contents of which are incorporated herein by reference in their entirety. Reference Electronic Sequence Listing

[0002] The contents of the electronic sequence listing (183952034340SEQLIST.xml; size: 11,571 bytes; and creation date: March 6, 2023) are incorporated herein by reference in its entirety. Technical Field

[0003] The present disclosure relates to methods of treating multiple myeloma by administering an anti-CD38 antibody (eg, isatuximab). Background Art

[0004] Multiple myeloma (MM) is a malignant plasma cell disease characterized by clonal proliferation of plasma cells in the bone marrow (BM) and the production of excess monoclonal immunoglobulins (usually IgG or IgA type or free urinary light chains, i.e., paraproteins, M proteins, or M components). Patients with MM may experience bone pain, fractures, fatigue, anemia, infections, hypercalcemia, and kidney problems (Rollig et al. (2015) Lancet. 385(9983):2197-208). CD38 expression is particularly prominent in MM, as >98% of patients are positive for this protein (Goldmacher et al. (1994) Blood. 84(9):3017-25; Lin et al. (2004) Am J Clin Pathol. 121(4):482-8). The strong and uniform expression of CD38 on malignant clonal MM cells contrasts sharply with the restricted expression pattern on normal cells, suggesting that this antigen can be used for specific targeting of tumor cells.

[0005] In general, patients with MM will receive treatment regimens during their lifetime that contain drugs, either alone or in combination, such as proteasome inhibitors (e.g., bortezomib, ixazomib, and carfilzomib) and immunomodulators or (e.g., lenalidomide, pomalidomide, and thalidomide), monoclonal antibodies (e.g., elotuzumab), histone deacetylase (HDAC) inhibitors (e.g., panobinostat).

[0006] Despite significant progress and extension in overall survival (OS), multiple myeloma (MM) remains incurable and most patients will relapse and require additional treatment (Kumar SK, Rajkumar V, Kyle RA, et al., Multiple myeloma. Nat Rev Dis Primer. 2017;3:17046).

[0007] All references cited herein (including patent applications, patent publications, and UniProtKB / Swiss-Prot accession numbers) are hereby incorporated by reference in their entirety to the same extent as if each individual reference was specifically and individually indicated to be incorporated by reference. Summary of the invention

[0008] Provided herein are anti-CD38 antibodies and interleukin-2 analogs (IL-2 analogs) for treating multiple myeloma. In some embodiments, provided herein are anti-CD38 antibodies and IL-2 analogs combined with NK cells (NK-CD38KO cells) in which the expression of CD38 is reduced or knocked out.

[0009] Provided herein are methods for treating multiple myeloma, comprising: administering an anti-CD38 antibody and an interleukin-2 analog (IL-2 analog) to an individual, wherein the anti-CD38 antibody is administered on day 1, day 8, day 15, and day 22 of the first 28-day cycle, and then administered on day 1 and day 15 of the 28-day cycle for at least one additional cycle; and administering the IL-2 analog to the individual, wherein the IL-2 analog is administered once every two weeks or once every three weeks. In some embodiments, the method comprises administering NK-CD38KO cells in combination with an anti-CD38 antibody and / or an IL-2 analog.

[0010] Provided herein is the use of an anti-CD38 antibody for treating multiple myeloma, wherein the anti-CD38 antibody is administered to an individual in combination with an interleukin-2 analog (IL-2 analog), wherein the anti-CD38 antibody is administered on days 1, 8, 15, and 22 of the first 28-day cycle and then on days 1 and 15 of the 28-day cycle for at least one additional cycle, and wherein the IL-2 analog is administered once every two weeks or once every three weeks. In some embodiments, the anti-CD38 antibody and IL-2 analog are administered in combination with NK-CD38KO cells. BRIEF DESCRIPTION OF THE DRAWINGS

[0011] Figure 1Bar graphs showing that CD38 KO K-NK cells from BC50 samples avoid fratricidal effects compared to wild-type KNK cells in the presence of isatuximab (light gray) or isotype control (dark gray).

[0012] Figure 2 Bar graphs showing that CD38 KO K-NK cells from BC45 samples avoid fratricidal effects compared to wild-type KNK cells in the presence of isatuximab (light gray) or isotype control (dark gray). DETAILED DESCRIPTION Overview

[0013] The present disclosure provides anti-CD38 antibodies and interleukin-2 analogs (IL-2 analogs) for treating multiple myeloma. In some embodiments, the anti-CD38 antibodies and IL-2 analogs provided herein are combined with NK cells (NK-CD38KO cells) whose expression of CD38 is reduced or knocked out. Provided herein is a method for treating multiple myeloma (MM) in an individual or delaying the progress of MM in an individual. In some embodiments, the patient has not received treatment for MM before (e.g., newly diagnosed MM). In some embodiments, the individual has received one, two, three or more than three previous therapies for MM. In some embodiments, the individual has received a first-line or multi-line therapy previously comprising an anti-CD38 agent and an anti-BCMA agent.

[0014] Provided herein are methods for treating multiple myeloma, comprising administering an anti-CD38 antibody and an interleukin-2 analog (IL-2 analog) to an individual. Provided herein are uses of anti-CD38 antibodies for treating multiple myeloma, wherein the anti-CD38 antibody is administered in combination with an IL-2 analog to an individual. In some embodiments of the methods and uses provided herein, the anti-CD38 antibody and the IL-2 analog are administered in combination with NK-CD38KO cells.

[0015] These methods include administering to the individual an effective amount of an anti-CD38 antibody (e.g., isatuximab) in combination with natural killer (NK) cells in which CD38 expression has been "knocked out" (e.g., cells that have been genetically modified to remove all or part of the CD38 gene such that the cells do not express CD38) or IL-15 or an analog thereof, or a combination thereof. In some embodiments, dexamethasone is also administered. In some embodiments, treatment prolongs progression-free survival (PFS) and / or overall survival (OS) in the individual. In some embodiments, treatment prolongs progression-free survival (PFS) and / or overall survival (OS) in the individual compared to an individual who has not received treatment. In some embodiments, the individual is negative for minimal residual disease (MRD) after treatment (e.g., a threshold of 10 -4or less, 10 -5 or lower, or 10 -6 or lower) (also called "MRD negative"). Anti-CD38 antibody

[0016] In some embodiments, the anti-CD38 antibody binds to human CD38. In some embodiments, the anti-CD38 antibody is a human antibody, a humanized antibody, or a chimeric antibody. In some embodiments, the anti-CD38 antibody comprises: (a) a heavy chain variable domain (V H ), the heavy chain variable domain comprising: a CDR-H1 comprising the amino acid sequence DYWMQ (SEQ ID NO: 1), a CDR-H2 comprising the amino acid sequence TIYPGDGDTGYAQKFQG (SEQ ID NO: 2), and a CDR-H3 comprising the amino acid sequence GDYYGSNSLDY (SEQ ID NO: 3); and (b) a light chain variable domain (V L ), the light chain variable domain comprising: a CDR-L1 comprising the amino acid sequence KASQDVSTVVA (SEQ ID NO: 4), a CDR-L2 comprising the amino acid sequence SASYRYI (SEQ ID NO: 5), and a CDR-L3 comprising the amino acid sequence QQHYSPPYT (SEQ ID NO: 6). In some embodiments, the anti-CD38 antibody comprises a heavy chain variable domain (V D ) comprising an amino acid sequence that is at least 90% identical (e.g., at least any one of 91%, 92%, 94%, 95%, 96%, 97%, 98%, or 99%, including any ranges between these values) to SEQ ID NO: 7. H Additionally or alternatively, in some embodiments, the anti-CD38 antibody comprises a light chain variable domain (V LLD ) comprising an amino acid sequence that is at least 90% identical (e.g., at least any one of 91%, 92%, 94%, 95%, 96%, 97%, 98%, or 99%, including any ranges between these values) to SEQ ID NO: 8 or SEQ ID NO: 9. L In some embodiments, the anti-CD38 antibody comprises a V comprising SEQ ID NO: 7. H and V comprising SEQ ID NO: 8 or SEQ ID NO: 9 L . QVQLVQSGAE VAKPGTSVKL SCKASGYTFT DYWMQWVKQR PGQGLEWIGT IYPGDGDTGYAQKFQGKATL TADKSSKTVY MHLSSLASED SAVYYCARGD YYGSNSLDYW GQGTSVTVSS(SEQ ID NO:7) DIVMTQSHLS MSTSLGDPVS ITCKASQDVS TVVAWYQQKP GQSPRRLIYS ASYRYIGVPDRFTGSGAGTD FTFTISSVQA EDLAVYYCQQ HYSPPYTFGG GTKLEIKR(SEQ ID NO:8) DIVMAQSHLS MSTSLGDPVS ITCKASQDVS TVVAWYQQKP GQSPRRLIYS ASYRYIGVPDRFTGSGAGTD FTFTISSVQA EDLAVYYCQQ HYSPPYTFGG GTKLEIKR(SEQ ID NO:9)

[0017] In some embodiments, the anti-CD38 antibody is isatuximab (CAS Registry Number: 1461640-62-9). Isatuximab (also known as hu38SB19 and SAR650984) is an anti-CD38 antibody described in WO 2008 / 047242 and U.S. Pat. No. 8,153,765, the contents of both of which are incorporated herein by reference in their entirety.

[0018] The heavy chain of isatuximab contains the following amino acid sequence: QVQLVQSGAE VAKPGTSVKL SCKASGYTFT DYWMQWVKQR PGQGLEWIGT IYPGDGDTGY AQKFQGKATL TADKSSKTVY MHLSSLASED SAVYYCARGD YYGSNSLDYW GQGTSVTVSS ASTKGPSVFP LAPSSKSTSG GTAALGCLVK DYFPEPVTVSWNSGALTSGV HTFPAVLQSS GLYSLSSVVTVPSSSLGTQT YICNVNHKPSNTKVDKKVEPKSCDKTHTCP PCPAPELLGG PSVFLFPPKP KDTLMISRTP EVTCVVVDVS HEDPEVKFNWYVDGVEVHNA KTKPREEQYN STYRVVSVLT VLHQDWLNGK EYKCKVSNKA LPAPIEKTISKAKGQPREPQ VYTLPPSRDE LTKNQVSLTC LVKGFYPSDI AVEWESNGQP ENNYKTTPPV LDSDGSFFLY SKLTVDKSRW QQGNVFSCSV MHEALHNHYT QKSLSLSPG (SEQ ID NO: 10) and the light chain of isatuximab comprises the following amino acid sequence: DIVMTQSHLS MSTSLGDPVS ITCKASQDVS TVVAWYQQKPGQSPRRLIYS ASYRYIGVPD RFTGSGAGTD FTFTISSVQA EDLAVYYCQQ HYSPPYTFGGGTKLEIKRTV AAPSVFIFPP SDEQLKSGTA SVVCLLNNFY PREAKVQWKV DNALQSGNSQ ESVTEQDSKD STYSLSSTLT LSKADYEKHK VYACEVTHQG LSSPVTKSFN RGEC(SEQ ID NO:11)

[0019] Anti-CD38 antibodies can be produced using recombinant methods. In order to recombinantly produce anti-antigen antibodies, the nucleic acid encoding the antibody is isolated and inserted into a replicable vector for further cloning (amplification of the DNA) or expression. The DNA encoding the antibody can be easily isolated and sequenced using conventional procedures (e.g., by using oligonucleotide probes that can specifically bind to the genes encoding the heavy and light chains of the antibody). Many vectors are available. Vector components typically include, but are not limited to, one or more of the following: a signal sequence, a replication start point, one or more marker genes, an enhancer element, a promoter, and a transcription termination sequence. The vector is typically transformed into a host cell suitable for expressing the nucleic acid. In certain embodiments, the host cell is a eukaryotic cell or a prokaryotic cell. In certain embodiments, the eukaryotic host cell is a mammalian cell. Examples of useful mammalian host cell lines are monkey kidney CV1 line transformed by SV40 (COS-7, ATCC CRL 1651); human embryonic kidney line (293 or 293 cells subcloned for growth in suspension culture, Graham et al., J. Gen Virol. 36:59 (1977)); baby hamster kidney cells (BHK, ATCC CCL 10); mouse Sertoli cells (TM4, Mather, Biol. Reprod. 23:243-251 (1980)); monkey kidney cells (CV1 ATCC CCL 70); African green monkey kidney cells (VERO-76, ATCC CRL-1587); human cervical carcinoma cells (HELA, ATCC CCL 2); canine kidney cells (MDCK, ATCC CCL 34); rat hepatocytes (BRL 3A, ATCC CRL 1442); human lung cells (W138, ATCC CCL75); human hepatocytes (Hep G2, HB 8065); mouse mammary tumor (MMT 060562, ATCC CCL51); TRI cells (Mather et al., Annals NY Acad. Sci. [Annals of the New York Academy of Sciences] 383:44-68 (1982)); MRC 5 cells; FS4 cells; and human hepatoma cell line (Hep G2). Other useful mammalian host cell lines include Chinese hamster ovary (CHO) cells, including DHFR-CHO cells (Urlaub et al., Proc. Natl. Acad. Sci. USA [Proceedings of the National Academy of Sciences of the United States] 77:4216 (1980)); and myeloma cell lines, such as NS0 and Sp2 / 0.For a review of certain mammalian host cell lines suitable for antibody production, see, e.g., Yazaki and Wu, Methods in Molecular Biology, Vol. 248 (BKC Lo, ed., Humana Press, Totowa, NJ, 2003), pp. 255-268. Anti-CD38 antibodies produced by the cells can be purified using, e.g., hydroxyapatite chromatography, hydrophobic interaction chromatography, gel electrophoresis, dialysis, and affinity chromatography, with affinity chromatography being one of the typical preferred purification steps. In general, various methods for preparing antibodies for research, testing, and clinical use are recognized in the art, which are consistent with the above methods and / or deemed appropriate by those skilled in the art.

[0020] In some embodiments, the IL-2 analog is IL-2 that has been PEGylated. In some embodiments, the IL-2 analog has (e.g., includes or comprises) a non-natural amino acid, such as N6-((2-azidoethoxy)-carbonyl)-L-lysine (AzK). In some embodiments, the non-natural amino acid is PEGylated separately using a linear PEG group with an average molecular weight of 30 kDa. Suitable IL-2 analogs are described, for example, in WO2020 / 163532, which is incorporated herein in its entirety.

[0021] In some embodiments, natural killer (NK) cells (NK-CD38KO cells) modified so that the expression of CD38 is reduced or knocked out are used in combination with isatuximab and / or IL-2 analogs for the treatment of multiple myeloma. Such NK-CD38KO cells can be produced, for example, by the method described in WO 2021087466, the teachings of which are incorporated herein in their entirety.

[0022] In some embodiments, the individual has received one or more lines of therapy for the treatment of multiple myeloma before receiving the first treatment cycle using anti-CD38 antibodies, IL-2 analogs, and optional NK-CD38KO cells. Anti-CD38 agents include, for example, monospecific or multispecific binding agents that can specifically bind to human CD38. Anti-BCMA agents include, for example, monospecific or multispecific binding agents that can specifically bind to human BCMA. These binding agents can be, for example, monoclonal antibodies, bispecific or trispecific antibodies, or antibody analogs containing antigen-specific regions of traditional antibodies, which can specifically bind to CD38 or BCMA, respectively. The binding agent can also be an antibody-drug conjugate. Pharmaceutical compositions and preparations

[0023] Also provided herein are pharmaceutical compositions and preparations, for example, for the treatment of multiple myeloma, comprising anti-CD38 antibodies (e.g., isatuximab), IL-2 analogs, NK-CD38KO cells and / or dexamethasone. In some embodiments, each of anti-CD38 antibodies, IL-2 analogs, NK-CD38KO cells and optional dexamethasone is provided as a separate pharmaceutical composition. In some embodiments, the pharmaceutical compositions and preparations further comprise a pharmaceutically acceptable carrier.

[0024] In some embodiments, the anti-CD38 antibody is in the form of a formulation suitable for intravenous administration, the formulation comprising, for example, about 20 mg / mL (500 mg / 25 mL) antibody, about 20 mM histidine, about 10% (w / v) sucrose, about 0.02% (w / v) polysorbate 80 at pH 6.0. In some embodiments, the anti-CD38 antibody is in the form of a formulation comprising about 20 mg / mL antibody, about 100 mg / mL sucrose, 2.22 mg / mL histidine hydrochloride monohydrate, about 1.46 mg / ml histidine and about 0.2 mg / ml polysorbate 80. In some embodiments, the formulation comprises water for injection (WFI), such as sterile water for injection (SWFI). In some embodiments, the formulation is sterile. In some embodiments, a single-use formulation comprises 5 ml of formulation (i.e., 100 mg anti-CD38 antibody). In some embodiments, a single-use 5 ml formulation is provided in, for example, a 16 mL colorless transparent glass vial equipped with an elastomeric cap. In some embodiments, the filling volume of the vial has been determined to ensure that 5mL is removed. In some embodiments, the filling volume is 5.4mL. In some embodiments, the single-use preparation includes 25ml of preparation (i.e., 500mg anti-CD38 antibody). In some embodiments, a single-use 25ml preparation is provided in a 30mL colorless transparent glass vial, for example, equipped with an elastomer cap. In some embodiments, the filling volume of the vial has been determined to ensure that 25mL is removed. In some embodiments, the preparation is kept stable for at least about 6 months, 12 months, 18 months, 24 months, 30 months or 36 months when stored in the dark at a temperature between about 2°C and about 8°C, including any range between these values. In some embodiments, the preparation is diluted for infusion in 0.9% sodium chloride or 5% dextrose. In some embodiments, the diluted infusion solution is kept stable for about 6 hours, 12 hours, 18 hours, 24 hours, 30 hours, 36 hours, 42 hours or 48 hours between about 2°C and about 8°C, including any range between these values. In some embodiments, the diluent solution for infusion remains stable at room temperature for another 8 hours (including the infusion time) after storage between about 2°C and about 8°C. In some embodiments, the diluent solution for infusion is stable under light. In some embodiments, the bag for storing the diluent solution for infusion is made of polyolefin (PO), polyethylene (PE), polypropylene (PP), polyvinyl chloride (PVC) and diethylhexyl phthalate (DEHP) or ethyl vinyl acetate (EVA). In some embodiments, the tube for infusion is made of PE, PVC (with or without DEHP), polybutadiene (PBD) or polyurethane (PU), with a line filter (polyethersulfone (PES), polysulfone or nylon).

[0025] In some embodiments, the anti-CD38 antibody is provided as a formulation for subcutaneous administration. In some embodiments, the anti-C38 antibody comprises isatuximab at a concentration of 140 mg / mL, 9 mM histidine, 110 mM Arg-Cl, 2% (w / v) sucrose, and 0.4% (w / v) poloxamer 188. Use of anti-CD38 antibody combined with other drugs for treating multiple myeloma

[0026] Provided herein are anti-CD38 antibodies for use in combination with one or more of an IL-2 analog, a natural killer (NK) cell modified to reduce or knock out CD38 expression (NK-CD38KO cells), and dexamethasone for treating multiple myeloma in an individual (e.g., a human individual). In some embodiments, the use comprises administering to the individual an effective amount of an anti-CD38 antibody (e.g., an anti-CD38 antibody comprising: (a) a heavy chain variable domain (V H ), the heavy chain variable domain comprising: a CDR-H1 comprising the amino acid sequence DYWMQ (SEQ ID NO: 1), a CDR-H2 comprising the amino acid sequence TIYPGDGDTGYAQKFQG (SEQ ID NO: 2), and a CDR-H3 comprising the amino acid sequence GDYYGSNSLDY (SEQ ID NO: 3); and (b) a light chain variable domain (V L ), the light chain variable domain comprising: a CDR-L1 comprising the amino acid sequence KASQDVSTVVA (SEQ ID NO: 4), a CDR-L2 comprising the amino acid sequence SASYRYI (SEQ ID NO: 5), and a CDR-L3 comprising the amino acid sequence QQHYSPPYT (SEQ ID NO: 6). In some embodiments, the anti-CD38 antibody is isatuximab.

[0027] In some embodiments, the method comprises administering to the individual an anti-CD38 antibody (e.g., isatuximab) at a dose of 10 mg / kg on days 1, 8, 15, and 22 of a first 28-day cycle, and administering the anti-CD38 antibody at a dose of 10 mg / kg on days 1 and 15 of a 28-day cycle for at least one additional cycle.

[0028] In some embodiments, the method comprises administering the anti-CD38 antibody (e.g., isatuximab) at a dose of 10 mg / kg once every 28 days in one or more additional 28-day cycles after administering the anti-CD38 antibody at a dose of 10 mg / kg on days 1 and 15 of a 28-day cycle for at least 11 cycles.

[0029] In some embodiments, the method comprises administering the anti-CD38 antibody (e.g., isatuximab) at a dose of 10 mg / kg once every 28 days for one or more additional 28-day cycles after the subject has achieved at least a very good partial response (VGPR) while being treated with the anti-CD38 antibody.

[0030] In some embodiments, the method comprises administering the anti-CD38 antibody (e.g., isatuximab) at a dose of 10 mg / kg once every 28 days for one or more additional 28-day cycles after the subject has achieved MRD negativity while being treated with the anti-CD38 antibody.

[0031] In some embodiments, the method comprises administering the anti-CD38 antibody (e.g., isatuximab) at a dose of 10 mg / kg once every 28 days for one or more additional 28-day cycles after the subject has achieved MRD negativity while being treated with the anti-CD38 antibody.

[0032] In some embodiments described herein, the IL-2 analog is IL-2 modified to include the non-natural amino acid N6-((2-azidoethoxy)-carbonyl)-L-lysine (AzK), and wherein AzK is pegylated. In some embodiments, the IL-2 analog is administered once every two weeks, or once every three weeks, or a combination thereof. In some embodiments, the IL-2 analog is administered at a dose of 24ug / kg. In some embodiments, the IL-2 analog is administered at a dose of 32ug / kg. In some embodiments, the IL-2 analog is administered at a dose of 16ug / kg.

[0033] In some embodiments, the treatment regimens described herein prolong progression-free survival (PFS) in the individual.

[0034] In some embodiments, the multiple myeloma is smoldering multiple myeloma (SMM). In some embodiments, the multiple myeloma is newly diagnosed multiple myeloma. In some embodiments, the multiple myeloma is relapsed and / or refractory multiple myeloma (RRMM). In some embodiments, the individual has received 1, 2, or 3 prior therapies for multiple myeloma. In some embodiments, the individual has received more than three prior therapies for multiple myeloma. In some embodiments, the individual has received prior therapy with a proteasome inhibitor. In some embodiments, the individual has received prior therapy with an immunomodulator. In some embodiments, the individual has not received prior treatment with an anti-CD38 antibody.

[0035] In some embodiments, the individual has received prior treatment with an anti-CD38 antibody. In some embodiments, the prior anti-CD38 antibody is daratumumab. In some embodiments, the prior anti-CD38 antibody is isatuximab. Products or kits

[0036] In another embodiment of the present invention, a product or kit comprising an anti-CD38 antibody (e.g., isatuximab) is provided. In some embodiments, the product or kit further comprises at least one additional drug (e.g., one or more of IL-2 analogs, NK-CD38KO cells, or dexamethasone). In some embodiments, the product or kit further comprises a package insert comprising instructions for using an anti-CD38 antibody (e.g., isatuximab) and other drugs for treating multiple myeloma (e.g., smoldering multiple myeloma, newly diagnosed multiple myeloma, refractory multiple myeloma, or relapsed and refractory multiple myeloma) or delaying the progression of multiple myeloma according to the uses described herein. definition

[0037] As used in this specification and the appended claims, the singular forms "a", "an", and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "a molecule" optionally includes a combination of two or more such molecules, and the like.

[0038] "Sustained remission" refers to a sustained effect after cessation of treatment on preventing or delaying progression of a disease (e.g., multiple myeloma) and / or improving one or more remission criteria. For example, remission of multiple myeloma treatment can be measured according to the criteria described in the following literature: Kumar et al. (2016) "International Myeloma Working Group consensus criteria for response and minimal residual disease assessment in multiple myeloma." Lancet Oncol. 17(8): e328-e346 and Durie et al. (2006) "International uniform response criteria for multiple myeloma." Leukemia. 20: 1467-1473. (See also Table A below.) In some embodiments, the duration of sustained remission is at least the same as the duration of treatment, at least 1.5X, 2.0X, 2.5X, or 3.0X the duration of treatment. Table A Standard International Myeloma Working Group (IMWG) response criteria The sum of the products of the maximum perpendicular diameters of the measured lesions

[0039] Methods for measuring serum and urine M-protein levels are well known in the art and are described, for example, in the following references: Jenkins (2009) Clin Biochem Rev. 30(3): 119-122; Leung, Nelson "Chapter 8: Clinical Tests for Monoclonal Proteins." Onco-Nephrology Curriculum, American Society of Nephrology 2016, pp. 1-5.

[0040] In some embodiments, VGPR is assessed according to the criteria described in Kumar et al. (2016) "International Myeloma Working Group consensus criteria for response and minimal residual disease assessment in multiple myeloma." Lancet Oncol. 17(8):e328-e346 and Durie et al. (2006) "International uniform response criteria for multiple myeloma." Leukemia. 20:1467-1473, the contents of which are incorporated herein by reference in their entirety. (See also Table A.)

[0041] The term "pharmaceutical preparation" refers to a formulation whose form allows the biological activity of the active ingredient to be effective and does not contain additional components that are unacceptably toxic to the subject to which the preparation is to be administered. Such preparations are sterile. "Pharmaceutically acceptable" excipients (excipients, additives) refer to excipients that can be reasonably administered to a subject mammal to provide an effective dose of the active ingredient used.

[0042] As used herein, the term "treatment" refers to a clinical intervention designed to change the natural course of the disease or cell (e.g., cancer cell) being treated during clinical pathology. The desirable effects of treatment include reducing the rate of disease progression, improving or alleviating the disease state, and alleviating or improving prognosis. For example, if one or more symptoms associated with cancer are alleviated or eliminated (including but not limited to: reducing the proliferation of cancer cells (or destroying cancer cells), reducing symptoms caused by the disease, improving the quality of life of patients with the disease, reducing the dose of other agents required to treat the disease and / or prolonging the survival of the individual), the individual is successfully "treated".

[0043] As used herein, "delaying disease progression" means delaying, hindering, slowing, slowing, stabilizing and / or postponing the development of a disease (e.g., cancer). The length of time for such a delay may vary depending on the history of the disease and / or the individual being treated. As will be apparent to one skilled in the art, a substantial or significant delay may actually include prevention, in that the individual will not develop the disease. For example, advanced cancer (e.g., the development of metastases) may be delayed.

[0044] "Effective amount" is at least the minimum amount required to achieve a measurable improvement or prevention of a particular condition. The effective amount herein may vary according to factors such as the disease state, age, sex and weight of the individual / patient and the ability of the antibody to induce a desired relief in the individual. An effective amount is also the amount in which the beneficial effects of treatment outweigh any toxic or deleterious effects of treatment. For preventive use, the beneficial or desired results include, for example, the following results: eliminating or reducing risk, alleviating severity or delaying the onset of the disease, including biochemical, histological and / or behavioral symptoms of the disease, its complications and the intermediate pathological phenotypes that occur during the development of the disease. For therapeutic use, the beneficial or desired results include, for example, the following clinical results: reducing one or more symptoms caused by the disease, improving the quality of life of patients with the disease, reducing the dosage of other agents required for the treatment of the disease, for example, enhancing the effect of another agent by targeting, delaying the progression of the disease and / or prolonging survival. In the case of cancer or tumors, an effective amount of a drug may have an effect in the following aspects: reducing the number of cancer cells; reducing tumor size; inhibiting (i.e., slowing down or desirably stopping) the infiltration of cancer cells into peripheral organs; inhibiting (i.e., slowing down and desirably stopping) tumor metastasis; inhibiting tumor growth to a certain extent; and / or alleviating one or more symptoms associated with the condition to a certain extent. An effective amount may be administered in one or more administrations. For the purposes of the present invention, an effective amount of a drug, compound, or pharmaceutical composition is an amount sufficient to directly or indirectly achieve a preventive or therapeutic treatment. As understood in a clinical context, an effective amount of a drug, compound, or pharmaceutical composition may or may not be used in combination with another drug, compound, or pharmaceutical composition. Therefore, an "effective amount" may be considered in the context of administering one or more therapeutic agents, and if the desired result can or has been achieved in combination with one or more other drugs, a single drug may be considered to be administered in an effective amount.

[0045] As used herein, "in combination" means administering a treatment modality in addition to another treatment modality. Thus, "in combination" means administering a treatment modality before, during, or after administering a treatment modality to an individual.

[0046] A "subject" or "individual" for treatment purposes refers to any animal classified as a mammal, including humans, livestock and farm animals, and zoo, sport or pet animals, such as dogs, horses, cats, cows, etc. Preferably, the mammal is a human.

[0047] The term "antibody" herein is used in the broadest sense and specifically includes monoclonal antibodies (including full-length monoclonal antibodies), polyclonal antibodies, multispecific antibodies (eg, bispecific antibodies), and antibody fragments, so long as these antibodies exhibit the desired biological activity.

[0048] Human light chains are usually classified as kappa and lambda light chains, and human heavy chains are usually classified as μ, δ, γ, α or ε, and the isotypes of antibodies are defined as IgM, IgD, IgG, IgA and IgE, respectively. IgG has several subclasses, including but not limited to IgG1, IgG2, IgG3 and IgG4. IgM has multiple subclasses, including but not limited to IgM1 and IgM2. IgA is similarly subdivided into multiple subclasses, including but not limited to IgA1 and IgA2. Within the full-length light chain and heavy chain, the variable domain and the constant domain are usually connected by a "J" region of about 12 or more amino acids, and the heavy chain also includes a "D" region of about 10 or more amino acids. See, for example, Fundamental Immunology [Basic Immunology] (Paul, W. Editor, Raven Press [Raven Press], 2nd Edition, 1989), which is incorporated herein by reference in its entirety for all purposes. The variable region of each light / heavy chain pair usually forms an antigen binding site. The variable domains of antibodies typically exhibit the same overall structure of relatively conservative framework regions (FRs) joined by three hypervariable regions (also referred to as complementary determining regions or CDRs). Each pair of CDRs of the two chains is typically arranged by the framework regions, which can enable binding to a specific epitope. From the amino terminus to the carboxyl terminus, both the light chain variable domain and the heavy chain variable domain typically comprise, in sequence, the domains FR1, CDR1, FR2, CDR2, FR3, CDR3, and FR4.

[0049] The term "CDR set" refers to a group of three CDRs that appear in a single variable region that is capable of binding an antigen. The exact boundaries of these CDRs have been defined differently according to different systems. The system described by Kabat (Kabat et al., Sequences of Proteins of Immunological Interest (National Institutes of Health, Bethesda, Md.) (1987) and (1991)) not only provides a clear residue numbering system applicable to any variable region of an antibody, but also provides precise residue boundaries that define three CDRs. These CDRs may be referred to as Kabat CDRs.

[0050] The term "Fc" as used herein refers to the sequence of non-antigen binding fragments (whether in monomeric or multimeric form) produced by antibody digestion or by other means, and may contain a hinge region. The original immunoglobulin source of native Fc is preferably human, and may be any immunoglobulin. Fc molecules are composed of monomeric polypeptides, which can be connected to dimer or multimeric forms by covalent (i.e., disulfide bonds) and non-covalent association. According to category (e.g., IgG, IgA and IgE) or subclass (e.g., IgG1, IgG2, IgG3, IgA1, IgGA2 and IgG4), the number of intermolecular disulfide bonds between monomer subunits of native Fc molecules is in the range of 1 to 4. An example of Fc is a disulfide-bonded dimer produced by papain digestion of IgG. As used herein, the term "native Fc" is a general term for monomer, dimer and multimeric forms.

[0051] As used herein, the term "overall response rate" or "ORR" refers to the proportion of individuals / patients with stringent complete response (sCR), complete response (CR), very good partial response (VGPR), and partial response (PR) as assessed by the IRC using the IMWG response criteria as described in Kumar et al. (2016) "International Myeloma Working Group consensus criteria for response and minimal residual disease assessment in multiple myeloma." Lancet Oncol. 17(8):e328-e346 and Durie et al. (2006) "International uniform response criteria for multiple myeloma." Leukemia. 20:1467-1473. See also Table A herein.

[0052] This specification is deemed sufficient to enable those skilled in the art to practice the invention. Various modifications of the invention, except those shown and described herein, will become apparent to those skilled in the art through the above description and fall within the scope of the appended claims. All publications, patents and patent applications cited herein are incorporated herein by reference in their entirety for all purposes. Examples

[0053] The present disclosure will be more fully understood by reference to the following examples. However, these examples should not be construed as limiting the scope of the present invention. It should be understood that the examples and embodiments described herein are for illustrative purposes only, and the various modifications or changes made according to the examples and embodiments will be conceived by those skilled in the art and should be included in the spirit and scope of the present application and within the scope of the appended claims. Example 1: Determination of in vitro cytotoxic activity Antibody-dependent cellular cytotoxicity (ADCC)

[0054] pass Antibody-dependent cellular cytotoxicity (ADCC) against LP-1RFP multiple myeloma cells (target cells) using WT or CD38KO K-NK cells (effector cells) in combination with isatuximab and SAR444245 (PEGylated IL-2 analog) was measured over time using a live cell imaging and analysis system (Essen Bioscience).

[0055] By utilizing Nuclight red lentivirus (Sartorius) infected LP-1 cells (DSMZ) to express red fluorescent protein (RFP) to generate LP-1RFP cells. LP-1RFP cells were maintained in IMDM medium (Gibco, #12440053) supplemented with 20% fetal bovine serum (heat-inactivated FBS, Biowest, #S181H-100), 1% L-glutamine (Gibco, #25030-024) and incubated at 37°C and 5% CO2. LP-1RFP cells were centrifuged at 300 g for 5 minutes, resuspended in RPMI1640 complete medium (RPMI1640 supplemented with 10% fetal bovine serum-FBS, Biowest, #S181H-100-, 1% L-glutamine-Gibco, #25030-024), and then added Plate (details below).

[0056] Natural killer (NK) cells with reduced or knocked-out CD38 expression (CD38KO K-NK cells) were generated using peripheral blood NK cells isolated from two healthy donors (BC45 and BC50) and expanded using PM21 particle technology to generate highly activated K-NK cells. BC45 cells have a V / V phenotype of CD16, while BC50 cells have an F / V phenotype.

[0057] To generate CD38KO K-NK cells, CRISPR gene editing was applied during NK cell expansion by electroporation of Cas9 / RNP complexes targeting CD38. After expansion, WT and CD38KO K-NK cells were frozen and stored at -150°C. WT and CD38KO K-NK cells were thawed, seeded in RPMI1640 complete medium supplemented with 50U / mL IL-2 (Peprotech, #200-02), and incubated at 37°C and 5% CO2 for 16-20 hours. The cells were then centrifuged at 300g for 5 minutes and then added In the board.

[0058] Compounds and cells were added in the following order Plate (96-well flat-bottom microplate treated with poly-D-lysine CellCoat TM , Greiner Bio-One; #655946) in appropriate wells (final volume per well: 200 μl):

[0059] Isarituximab (or isotype control), previously diluted in RPMI1640 complete medium, was added to appropriate wells to achieve a final concentration of 0.1, 1 or 10 ng / ml (50 μl / well).

[0060] SAR444245 (previously diluted in RPMI1640 complete medium) was then added to the appropriate wells to reach a final concentration of 333,33 ng / ml (50 μl / well).

[0061] LP-1RFP cells (target cells, T) were then added to each well (to reach 20,000 LP1-RFP cells / well) (50 μl / well).

[0062] WT or CD38KO K-NK cells (effector cells, E) were then added to appropriate wells. 20,000, 60,000 or 100,000 cells were added to evaluate different E:T ratios (1:1, 3:1 and 5:1, respectively) (50 μl / well).

[0063] Then The plate was centrifuged at 100 g for 1 min and then placed in (IncucyteS3, EssenBio), Placed in a dedicated incubator at 37°C containing 5% CO2. Images were taken every 2 hours (4 images / well) using a 10X objective and standard scanning type using phase and red channels. The growth of LP-1RFP target cells was monitored by fluorescence imaging for up to 90 hours, and the number of live target cells was quantified using IncucyteS3 software and normalized to the number of live target cells at time zero.

[0064] At least 2 experiments were performed using WT and CD38KO K-NK cells from two donors (BC45 and BC50), with each condition performed in duplicate. cannibalism

[0065] WT and CD38KO K-NK cells were thawed, seeded in RPMI1640 complete medium (RPMI1640 supplemented with 10% fetal bovine serum-FBS, Biowest, #S181H-100-, 1% L-glutamine-Gibco, #25030-024) supplemented with 50U / mL IL-2 (Peprotech, #200-02), and incubated for 16-20 hours at 37°C and 5% CO2. The cells were then centrifuged at 300g for 5 minutes, counted, and resuspended in RPMI1640 complete medium to reach 50,000 K-NK cells / 50μl.

[0066] Compounds and cells were added to Corning in the following order TM In appropriate wells of a 96-well clear ultra-low attachment microplate (Corning, #7007) (final volume per well: 200 μl):

[0067] RPMI1640 complete medium (100 μl / well) was added to each well.

[0068] Isarituximab (or isotype control), previously diluted in RPMI1640 complete medium, was added to appropriate wells to achieve a final concentration of 10 ng / ml (50 μl / well).

[0069] WT or CD38KO K-NK cells were then added to appropriate wells (to achieve 50,000 K-NK cells / well) (50 μl / well).

[0070] The plate was then centrifuged at 100 g for 1 minute and placed in an incubator at 37°C containing 5% CO2 for 4 hours.

[0071] To quantify fratricide, WT and CD38KO K-NK cells were stained using the Annexin V-FITC kit or using DiOC6 / DRAQ7.

[0072] For labeling using Annexin V-FITC kit (Miltenyi Biotec; #130-092-052):

[0073] After 4 hours incubation, the plate was centrifuged at 300 g for 5 minutes, the medium was removed, and the pellet was resuspended in 200 μL of Annexin V-FITC kit buffer. The plate was centrifuged again at 300 g for 5 minutes.

[0074] The buffer was removed, and the pellet was resuspended in 100 μL of Annexin V-FITC kit buffer, and 5 μL of Annexin V-FITC was added to the appropriate wells.

[0075] The plate was incubated at room temperature in the dark for 15 minutes.

[0076] 100 μL of Annexin V-FITC kit buffer solution was added, the plate was centrifuged at 300 g for 5 minutes, and the buffer was removed.

[0077] The pellet was resuspended in 200 μL of Annexin V-FITC kit buffer and the plate was centrifuged again at 300 g for 5 minutes.

[0078] The buffer was removed and the pellet was resuspended in 100 μL of Annexin V-FITC kit buffer.

[0079] Immediately prior to analysis, 1 μL of propidium iodide (PI) was added to the appropriate wells.

[0080] The samples were then analyzed using a MACSQuant 16 flow cytometer (Miltenyi Biotec). The data were analyzed using VenturiOne software using the following strategy:

[0081] The K-NK cell population excluding debris was gated and then single cells were gated. From single cells, viable PINeg and Annexin V-FITCneg cells were quantified.

[0082] For graphical representation, viable K-NK cells (Annexin Vneg / PIneg) were quantified as the percentage of viable cells in the presence of isatuximab, which was normalized to the percentage of viable cells in the presence of isotype control (100%) (see Figure 1 ).

[0083] For labeling with DiOC6 / DRAQ7:

[0084] After the 4-hour incubation, 20 μL of 100 nM DiOC6 (3,3'-dihexyloxycarbocyanine iodide, ThermoFisher, #D273) was added to the appropriate wells.

[0085] The plate was incubated at 37°C and 5% CO2 for 20 minutes.

[0086] The plate was then centrifuged at 300 g for 5 minutes, the medium removed, and the pellet resuspended in 100 μL of 3 μM DRAQ7 (BD Biosciences, #564904).

[0087] The samples were then analyzed using a MACSQuant 16 flow cytometer (Miltenyi Biotec). The data were analyzed using VenturiOne software using the following strategy:

[0088] The K-NK cell population excluding debris was gated and then single cells were gated. From single cells, viable DiOC6pos and DRAQ7neg cells were quantified.

[0089] For graphical representation, viable K-NK cells (DiOC6pos / DRAQ7neg) were quantified as the percentage of viable cells in the presence of isatuximab, which was normalized to the percentage of viable cells in the presence of isotype control (100%) ( Figure 2 ).

[0090] Experiments were performed using WT and CD38KO K-NK cells from two donors (BC45 and BC50). Figure 1 As shown in Figure 2, the use of CD38 KO K-NK cells (BC50 with F / V CD16 phenotype) avoided the fratricidal effect in the presence of isatuximab. Figure 2As shown, the use of CD38 KO K-NK cells (BC45 with a V / V CD16 phenotype) in the presence of isatuximab avoids fratricidal effects. In addition, the presence of SAR444245 enhanced the cytotoxic activity of isatuximab against LP-1 cells in the presence of wild-type NK cells, and the cytotoxic activity of isatuximab was even further enhanced in the presence of CD38 KOK-NK cells. The cytotoxic activity of CD38 KO K-NK cells persisted for a period of time, up to 90 hours.

[0091] Unless clearly indicated to the contrary, each embodiment described herein can be combined with any other one or more embodiments. Specifically, unless clearly indicated to the contrary, any feature or embodiment indicated as preferred or advantageous can be combined with any other one or more features or one or more embodiments indicated as preferred or advantageous.

[0092] All references cited in this application are expressly incorporated herein by reference.

Claims

1. Use of an anti-CD38 antibody for treating multiple myeloma, wherein the anti-CD38 antibody is administered to an individual in combination with an interleukin-2 analog (IL-2 analog), wherein the anti-CD38 antibody is administered on days 1, 8, 15 and 22 of a first 28-day cycle and then on days 1 and 15 of a 28-day cycle for at least one additional cycle, and The IL-2 analog is administered to the individual, wherein the IL-2 analog is administered once every two weeks or once every three weeks.

2. The use according to claim 1, wherein the anti-CD38 antibody is isatuximab and is administered intravenously at a dose of 10 mg / kg.

3. The use according to claim 1, wherein the CD-38 antibody is isatuximab and is administered subcutaneously at a dose of 1400 mg.

4. The use of any one of claims 1-3, wherein the IL-2 analog comprises the unnatural amino acid N6-((2-azidoethoxy)-carbonyl)-L-lysine (AzK), and wherein the AzK is pegylated.

5. The use of claim 1, wherein the IL-2 analog is administered at a dose of 16 ug / kg, 24 ug / kg or 32 ug / kg.

6. The use of any one of claims 1-3 or 5, wherein the anti-CD38 antibody is administered on days 1 and 15 of a 28-day cycle for at least 11 cycles, and then on day 1 of a 28-day cycle for one or more additional cycles.

7. The use of any one of claims 1-3 or 5, wherein the individual has received one or more lines of therapy for treating multiple myeloma prior to receiving the first treatment cycle, wherein the one or more lines of prior therapy comprise an anti-CD38 agent and an anti-BCMA agent.

8. Use of an anti-CD38 antibody for treating multiple myeloma, wherein the anti-CD38 antibody is administered to an individual in combination with an interleukin-2 analog (IL-2 analog), wherein the anti-CD38 antibody is isatuximab, and wherein the IL-2 analog comprises the unnatural amino acid N6-((2-azidoethoxy)-carbonyl)-L-lysine (AzK), wherein the AzK is pegylated, wherein the isatuximab is administered on days 1, 8, 15, and 22 of a first 28-day cycle and then on days 1 and 15 of a 28-day cycle for at least one additional cycle, wherein the isatuximab is administered intravenously at a dose of 10 mg / kg or subcutaneously at a dose of 1400 mg, wherein the IL-2 analog is administered to the subject at a dose of 16 ug / kg, 24 ug / kg or 32 ug / kg once every two weeks or once every three weeks, and The individual has received one or more lines of therapy for treating multiple myeloma prior to receiving the first treatment cycle, wherein the one or more lines of prior therapy comprised an anti-CD38 agent and an anti-BCMA agent.

9. The use according to any one of claims 1 to 3, 5 or 8, wherein natural killer (NK) cells in which CD38 expression is reduced or knocked out (NK-CD38KO cells) are also administered to the individual.

10. The use of claim 9, wherein the NK cells are isolated from a human sample.

11. A method of treating a human subject suffering from multiple myeloma, the method comprising: administering an anti-CD38 antibody and an interleukin-2 analog (IL-2 analog) to the individual, wherein the anti-CD38 antibody is administered on days 1, 8, 15, and 22 of a first 28-day cycle, and then on days 1 and 15 of a 28-day cycle for at least one additional cycle, and The IL-2 analog is administered to the individual, wherein the IL-2 analog is administered once every two weeks or once every three weeks.

12. The method of claim 11, wherein the anti-CD38 antibody is isatuximab and is administered intravenously at a dose of 10 mg / kg.

13. The method of claim 11, wherein the CD-38 antibody is isatuximab and is administered subcutaneously at a dose of 1400 mg.

14. The method of any one of claims 11-13, wherein the IL-2 analog comprises the unnatural amino acid N6-((2-azidoethoxy)-carbonyl)-L-lysine (AzK), and wherein the AzK is pegylated.

15. The method of claim 14, wherein the IL-2 analog is administered at a dose of 16 ug / kg, 24 ug / kg or 32 ug / kg.

16. The method of any one of claims 11 to 13 or 15, wherein: The anti-CD38 antibody is administered on days 1 and 15 of a 28-day cycle for at least 11 cycles, and then the anti-CD38 antibody is administered on day 1 of a 28-day cycle for one or more additional cycles.

17. A method of treating a human subject suffering from multiple myeloma, the method comprising: administering to the individual an anti-CD38 antibody and an interleukin-2 analog (IL-2 analog), wherein the anti-CD38 antibody is isatuximab, and wherein the IL-2 analog comprises the unnatural amino acid N6-((2-azidoethoxy)-carbonyl)-L-lysine (AzK), wherein the AzK is pegylated, wherein the isatuximab is administered on days 1, 8, 15, and 22 of a first 28-day cycle, and then on days 1 and 15 of a 28-day cycle for at least one additional cycle, and wherein the isatuximab is administered intravenously at a dose of 10 mg / kg or subcutaneously at a dose of 1400 mg, wherein the IL-2 analog is administered to the subject at a dose of 16 ug / kg, 24 ug / kg or 32 ug / kg once every two weeks or once every three weeks, and The individual has received one or more lines of therapy for treating multiple myeloma prior to receiving the first treatment cycle, wherein the one or more lines of prior therapy comprised an anti-CD38 agent and an anti-BCMA agent.

18. The method of any one of claims 11-13, 15 or 17, wherein natural killer (NK) cells in which CD38 expression is reduced or knocked out (NK-CD38KO cells) are also administered to the individual.

19. The method of claim 18, wherein the NK cells are isolated from a human sample.

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