Patent Publication Number: US-10308722-B2

Title: Method for the treament of multiple myeloma or non-hodgkins lymphoma with anti-CD38 antibody in combination with thalidomine, lenalidomids, or pomalidomide

Description:
CROSS REFERENCE TO RELATED APPLICATIONS 
     This instant application is a continuation of U.S. application Ser. No. 15/016,330, filed on Feb. 5, 2016, now pending, which is a division of U.S. application Ser. No. 14/500,366, filed Sep. 29, 2014, which is a continuation of U.S. application Ser. No. 13/825,325, filed on Mar. 21, 2013, which is a National Stage Entry of PCT/EP2011/066648, filed Sep. 26, 2011, which claims foreign priority to European Patent Application No. 10180485.4, filed Sep. 27, 2010 and claims priority from U.S. Provisional Appl. No(s): 61/486,814, filed May 17, 2011, 61/468,607, filed Mar. 29, 2011, 61/386,619, filed on Sep. 27, 2010, and U.S. 61/437,696, filed on Jan. 31, 2011, the disclosures of each of which is incorporated hereby reference in their entireties. 
    
    
     BACKGROUND 
     Multiple myeloma is a B cell malignancy characterized by the latent accumulation in bone marrow of secretory plasma cells with a low proliferative index and an extended life span. The disease ultimately attacks bones and bone marrow, resulting in multiple tumors and lesions throughout the skeletal system. 
     Approximately 1% of all cancers, and slightly more than 10% of all hematologic malignancies, can be attributed to multiple myeloma (MM). The incidence of MM increases in the aging population, with the median age at time of diagnosis being about 61 years. The currently available therapies for multiple myeloma include chemotherapy, stem cell transplantation, Thalomid® (thalidomide), Velcade® (bortezomib), Aredia® (pamidronate), and Zometa® (zoledronic acid). The current treatment protocols, which include a combination of chemotherapeutic agents such as vincristine, BCNU, melphalan, cyclophosphamide, adriamycin, and prednisone or dexamethasone, yield a complete remission rate of only about 5%, and median survival is approximately 36-48 months from the time of diagnosis. Recent advances using high dose chemotherapy followed by autologous bone marrow or peripheral blood mononuclear cell transplantation have increased the complete remission rate and remission duration. Yet overall survival has only been slightly prolonged, and no evidence for a cure has been obtained. Ultimately, MM patients often relapse, even under maintenance therapy with interferon-alpha (IFN-α) alone or in combination with steroids. 
     Non-Hodgkin&#39;s lymphoma is a broad classification of lymphomas, which are cancers originating from the lymphatic system when lymphocytes (B-cells or T-cells) become malignant and proliferate uncontrollably to form a tumor mass. In total NHL encompasses around 30 different subtypes of lymphoma, including Diffuse large B-cell lymphoma (DLBCL) and follicular lymphoma (FL). The incidence of NHL will reach over 140,000 in the major markets by 2019. The available treatment options include Rituxan/MabThera, combinations thereof, such as, R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine and prednisone), R-CVP (Rituxan, cyclophosphamide, vincristine and prednisone), and chemotherapy. In addition, following remission or after relapse, hematopoietic stem cell transplantation may be considered. Despite the current treatment options, however, the survival rates within high risk groups of aggressive NHL can be as low as 30% over 5 years. Therefore, there remains a high unmet need for effective treatments and combination treatments. 
     CD38 is an example of an antigen expressed on such malignant plasma cells, and other lymphocytes. Functions ascribed to CD38 include both receptor mediation in adhesion and signaling events and (ecto-) enzymatic activity. As an ectoenzyme, CD38 uses NAD+ as substrate for the formation of cyclic ADP-ribose (cADPR) and ADPR, but also of nicotinamide and nicotinic acid-adenine dinucleotide phosphate (NAADP). cADPR and NAADP have been shown to act as second messengers for Ca2+ mobilization. By converting NAD+ to cADPR, CD38 regulates the extracellular NAD+ concentration and hence cell survival by modulation of NAD-induced cell death (NCID). In addition to signaling via Ca2+, CD38 signaling occurs via cross-talk with antigen-receptor complexes on T and B cells or other types of receptor complexes, e.g. MHC molecules, and is in this way involved in several cellular responses, but also in switching and secretion of IgG. 
     Antibodies specific for CD38 are described in WO1999/62526 (Mayo Foundation); WO200206347 (Crucell Holland); US2002164788 (Jonathan Ellis) which is incorporated by reference in its entirety; WO2005/103083 (MorphoSys AG), U.S. Ser. No. 10/588,568, which is incorporated by reference in its entirety, WO2006/125640 (MorphoSys AG), U.S. Ser. No. 11/920,830, which is incorporated by reference in its entirety, and WO2007/042309 (MorphoSys AG), U.S. Ser. No. 12/089,806, which is incorporated by reference in its entirety; WO2006099875 (Genmab), U.S. Ser. No. 11/886,932, which is incorporated by reference in its entirety; and WO08/047242 (Sanofi-Aventis), U.S. Ser. No. 12/441,466, which is incorporated by reference in its entirety. 
     Combinations of antibodies specific for CD38 and other agents are described in WO200040265 (Research Development Foundation); WO2006099875 and WO2008037257 (Genmab); and WO2010061360, WO2010061359, WO2010061358 and WO2010061357 (Sanofi Aventis), which are all incorporated by reference in their entireties. 
     It is clear that in spite of the recent progress in the discovery and development of anti-cancer agents, many forms of cancer involving CD38-expressing tumors still have a poor prognosis. Thus, there is a need for improved methods for treating such forms of cancer. 
     SUMMARY 
     In one aspect, the present disclosure relates to a synergistic combination of an antibody specific for CD38 and thalidomide or an analog thereof, e.g. lenalidomide. In another aspect the present disclosure relates to a synergistic combination comprising an antibody specific for CD38 and bortezomib or other proteasome inhibitor. Such combinations are useful in the treatment of cancers, such as, multiple myeloma and/or non-Hodgkin&#39;s lymphoma. 
     In vitro and in vivo models are considered predictive of how a certain compound or combination of compounds would behave in humans. Here, the combinations of an antibody specific for CD38 and lenalidomide was tested in human multiple myeloma cell lines and synergy was identified. In addition the combination of an antibody specific for CD38 and lenalidomide, and a combination of an antibody specific for CD38 and bortezomib were tested in mouse models against both multiple myeloma cells and Burkitt&#39;s lymphoma (a form of NHL) cells and synergy was identified. Therefore, the combinations will be effective in the treatment of humans in multiple myeloma and/or non-Hodgkin&#39;s lymphoma. In addition, the antibody specific to CD38 exemplified in the present specification is entering into clinical trials, where such combinations can be confirmed in humans. 
     When compounds are combined either in vitro or in vivo, one expects that the combination has only additive effects. Quite unexpectedly, the inventors found that the combination of a particular anti-CD38 antibody and lenalidomide mediated a synergistic level of Antibody-Dependent Cell-Mediated Cytotoxicity (ADCC) in both the AMO-1 and NCI-H929 multiple myeloma cell lines. In addition, and also unexpectedly, a particular anti-CD38 antibody when combined with lenalidomide or when combined with bortezomib mediated a synergistic level of reduction in bone lysis in the NCI-H929 SCID mouse model and synergistically increased the median survival days in the RAMOS SCID mouse model. Therefore, both the combination of the exemplified antibody specific for CD38 and lenalidomide and the exemplified antibody specific for CD38 and bortezomib behaved synergistically in the in vitro and/or in vivo models relevant to multiple myeloma and/or non-Hodgkin&#39;s lymphoma. 
     An aspect of the present disclosure comprises a combination wherein the antibody specific for CD38 comprises an HCDR1 region of sequence GFTFSSYYMN (SEQ ID NO: 1) or of sequence SYYMN (SEQ ID NO: 14), an HCDR2 region of sequence GISGDPSNTYYADSVKG (SEQ ID NO: 2), an HCDR3 region of sequence DLPLVYTGFAY (SEQ ID NO: 3), an LCDR1 region of sequence SGDNLRHYYVY (SEQ ID NO: 4), an LCDR2 region of sequence GDSKRPS (SEQ ID NO: 5), and an LCDR3 region of sequence QTYTGGASL (SEQ ID NO: 6) and the thalidomide or an analog thereof is lenalidomide. In preferred aspects, the combination is used for the treatment of multiple myeloma and/or non-Hodgkin&#39;s lymphoma. 
     An aspect of the present disclosure comprises a combination wherein the antibody specific for CD38 comprises an HCDR1 region of sequence GFTFSSYYMN (SEQ ID NO: 1) or of sequence SYYMN (SEQ ID NO: 14), an HCDR2 region of sequence GISGDPSNTYYADSVKG (SEQ ID NO: 2), an HCDR3 region of sequence DLPLVYTGFAY (SEQ ID NO: 3), an LCDR1 region of sequence SGDNLRHYYVY (SEQ ID NO: 4), an LCDR2 region of sequence GDSKRPS (SEQ ID NO: 5), and an LCDR3 region of sequence QTYTGGASL (SEQ ID NO: 6) and the proteasome inhibitor is bortezomib. In preferred aspects, the combination is used for the treatment of multiple myeloma and/or non-Hodgkin&#39;s lymphoma. 
    
    
     
       DESCRIPTION OF DRAWINGS 
         FIG. 1  shows the effects of lenalidomide alone on the expression of CD38 in AMO-1 cells. 
         FIG. 2  shows the effects of lenalidomide alone on cell proliferation in various multiple myeloma cell lines. This measure represents the relative cytoxicity of lenalidomide on each cell line. 
         FIG. 3  shows the mediation of ADCC on AMO-1 cells by the combination of MOR03087 and lenalidomide. The PBMCs and AMO-1 cells were treated with lenalidomide prior to treatment with MOR03087. MOR03207 binds lysosyme, and is used as isotype control, as it is IgG1. LEN represents lenalidomide. “Theoretical” represents the addition of the value of MOR03087 alone and the value of LEN alone. The data shown are the averages from Table 3b. 
         FIG. 4  shows the mediation of ADCC on AMO-1 cells by the combination of MOR03087 and lenalidomide. Only the PBMCs were treated with lenalidomide prior to treatment with MOR03087. MOR03207 binds lysosyme, and is used as isotype control, as it is IgG1. LEN represents lenalidomide. “Theoretical” represents the addition of the value of MOR03087 alone and the value of LEN alone. The data shown are the averages from Table 4b. 
         FIG. 5  shows the effects of lenalidomide alone on the expression of CD38 in NCI-H929 cells. 
         FIG. 6  shows the mediation of ADCC on NCI-H929 cells by the combination of MOR03087 and lenalidomide. The PBMCs and NCI-H929 cells were treated with lenalidomide prior to treatment with MOR03087. Theoretical represent the combination calculated using the fractional product concept of Chou et al. The data shown are the averages from Table 7b. 
         FIG. 7  shows the mediation of ADCC on NCI-H929 cells by the combination of MOR03087 and lenalidomide. Only the PBMCs were treated with lenalidomide prior to treatment with MOR03087. Theoretical represent the combination calculated using the fractional product concept of Chou et al. The data shown are the averages from Table 8b. 
         FIG. 8  shows the growth inhibition of various multiple myeloma cell lines caused by bortezomib alone. The IC50 on AMO-1 cells was 3.9 nM. The IC50 on LP-1 cells was 6.1 nM. The IC50 on NCI-H929 cells was 3.3 nM. The IC50 on RPMI-8226 cells was 9.0 nM. 
         FIG. 9  shows the mediation of ADCC on NCI-H929 cells by the combination of MOR03087 at 15 μg/ml and Velcade® (bortezomib). The two charts represent two different donors. 
         FIG. 10  shows the mediation of ADCC on LP-1 cells by the combination of MOR202 at 15 μg/ml and Velcade® (bortezomib). The two charts represent two different donors. 
         FIG. 11  shows the amino acid sequence of MOR202. 
         FIG. 12  shows the Best Fit curve, as described in Chou et al., of the MOR202 and lenalidomide combination in the mediation of ADCC on AMO-1 cells and it is also representative for the Best Fit curve generated for analysis of the mediation of ADCC on NCI-H929 cells. 
         FIGS. 13-18  show the Chou factor synergy analysis for six separate experiments using the combination of MOR202 and lenalidomide in the mediation of ADCC on AMO-1 cells.  FIG. 13  shows experiment 1.  FIG. 14  shows experiment 2.  FIG. 15  shows experiment 3.  FIG. 16  shows experiment 4.  FIG. 17  shows experiment 5.  FIG. 18  shows experiment 6.  FIGS. 13-15  were derived from the three experiments shown in Tables 3a-c, and  FIG. 3 .  FIGS. 16-18  were derived from the three experiments shown in Tables 4a-c, and  FIG. 4 . 
         FIG. 19  shows the MicroCT Scan mean total bone volume of each of the study groups described in Example 7, where the results are shown in Table 11. 
         FIG. 20  shows the MicroCT Scan mean total bone volume of each of the study groups described in Example 11, where the results are shown in Table 17. 
     
    
    
     DETAILED DESCRIPTION OF THE INVENTION 
     “Synergy”, “synergism” or “synergistic” mean more than the expected additive effect of a combination. The “synergy”, “synergism” or “synergistic” effect of a combination is determined herein by the methods of Chou et al., and/or Clarke et al. See Ting-Chao Chou, Theoretical Basis, Experimental Design, and Computerized Simulation of Synergism and Antagonism in Drug Combination Studies, Pharmacol Rev 58:621-681 (2006), which is incorporated by reference in its entirety. In Chou et al., multiple methods of determining synergism are disclosed and at least one of these methods is used herein. See also Clarke et al., Issues in experimental design and endpoint analysis in the study of experimental cytotoxic agents in vivo in breast cancer and other models, Breast Cancer Research and Treatment 46:255-278 (1997), which is incorporated by reference in its entirety. 
     The term “antibody” means monoclonal antibodies, including any isotype, such as, IgG, IgM, IgA, IgD and IgE. An IgG antibody is comprised of two identical heavy chains and two identical light chains that are joined by disulfide bonds. Each heavy and light chain contains a constant region and a variable region. Each variable region contains three segments called “complementarity-determining regions” (“CDRs”) or “hypervariable regions”, which are primarily responsible for binding an epitope of an antigen. They are referred to as CDR1, CDR2, and CDR3, numbered sequentially from the N-terminus. The more highly conserved portions of the variable regions outside of the CDRs are called the “framework regions”. An “antibody fragment” means an Fv, scFv, dsFv, Fab, Fab′ F(ab′)2 fragment, or other fragment, which contains at least one variable heavy or variable light chain, each containing CDRs and framework regions. 
     THALOMID® (thalidomide) in combination with dexamethasone is indicated for the treatment of patients with newly diagnosed multiple myeloma, and is marketed by Celgene. 
     A “thalidomide analog” includes, but is not limited to, thalidomide itself, lenalidomide (CC-5013, Revlimid™), Pomalidomide (CC4047, Actimid™) and the compounds disclosed in WO2002068414 and WO2005016326, which are incorporated by reference in their entireties. The term refers to a synthetic chemical compound using the thalidomide structure as a backbone (e.g., side groups have been added or such groups have been deleted from the parent structure). The analog differs in structure from thalidomide and its metabolite compounds such as by a difference in the length of an alkyl chain, a molecular fragment, by one or more functional groups, or a change in ionization. The term “thalidomide analog” also includes the metabolites of thalidomide. Thalidomide analogs include the racemic mixture of the S- and the R-enantiomer of a respective compound and the S-enantiomer or to the R-enantiomer individually. The racemic mixture is preferred. Thalidomide analogs include the compounds of the following structures: 
     (A) Lenalidomide 
                         
(B) Thalidomide
 
                         
(C) Pomalidomide
 
                         
(D)
 
                         
wherein R21, R22, R23, and R24 are each independently H, alkoxy, amino, or alkylamine, and
 
(E)
 
                         
wherein R21, R22, R23, and R24 are each independently H, alkoxy, amino, or alkylamine. Lenalidomide is currently marketed as Revlimid® by Celgene for the treatment of multiple myeloma. Lenalidomide is described as having at least the following properties in relation to the treatment of tumors, a) cytotoxic to tumor cells, Gandhi et al., Lenalidomide inhibits proliferation of Namalwa CSN.70 cells and interferes with Gab1 phosphorylation and adaptor protein complex assembly, Leuk Res., 30(7):849-58 (2006), which is incorporated by reference in its entirety; b) activates natural killer (Nk) cells, Gandhi et al., Dexamethasone synergizes with lenalidomide to inhibit multiple myeloma tumor growth, but reduces lenalidomide-induced immunomodulation of T and NK cell function, Curr Cancer Drug Targets, 1;10(2):155-67 (March 2010), which is incorporated by reference in its entirety; and c) upregulates CD38 expression on tumor cells, See Lapalombella et al., Lenalidomide down-regulates the CD20 antigen and antagonizes direct and antibody-dependent cellular cytotoxicity of rituximab on primary chronic lymphocytic leukemia cells, Blood, 112:13, 5180-5189 (15 Dec. 2008), which is incorporated by reference in its entirety. “LEN” is used to describe lenalidomide.
 
     As described, thalidomide analogs upregulate the expression of CD38 on tumor cells. Other agents that upregulate the expression of CD38 on the surface of tumor cells are described in WO00/40265, U.S. Ser. No. 09/226,895, which is incorporated by reference in its entirety (Research Development Foundation). 
     A “proteasome inhibitor” refers to a compound that blocks the action of proteasomes, i.e. cellular complexes that break down proteins, such as for example the p53 protein. Several classes of proteasome inhibitors are known. The class of the peptide boronates includes bortezomib (INN, PS-341; Velcade®), a compounds which is approved in the U.S. for the treatment of relapsed multiple myeloma. Another peptide boronate is CEP-18770. Other classes of proteasome inhibitors include peptide aldehydes (e.g. MG132), peptide vinyl sulfones, peptide epoxyketones (e.g. epoxomicin, carfilzomib), β lactone inhibitors (e.g. lactacystin, MLN 519, NPI-0052, Salinosporamide A), compounds which create dithiocarbamate complexes with metals (e.g. Disulfiram, a drug which is also used for the treatment of chronic alcoholism), and certain antioxidants (e.g. Epigallocatechin-3-gallate) catechin-3-gallate, and Salinosporamide A. 
     “VH” refers to the variable region of an immunoglobulin heavy chain of an antibody, or antibody fragment. “VL” refers to the variable region of the immunoglobulin light chain of an antibody, or antibody fragment. 
     The term “CD38” refers to the protein known as CD38, having the following synonyms: ADP-ribosyl cyclase 1, cADPr hydrolase 1, Cyclic ADP-ribose hydrolase 1, T10. 
     Human CD38 has the amino acid sequence of: 
     
       
         
           
               
            
               
                 (SEQ ID NO: 7) 
               
            
           
           
               
            
               
                 MANCEFSPVSGDKPCCRLSRRAQLCLGVSILVLILVVVLAVVVPRWRQQW 
               
               
                   
               
               
                 SGPGTTKRFPETVLARCVKYTEIHPEMRHVDCQSVWDAFKGAFISKHPCN 
               
               
                   
               
               
                 ITEEDYQPLMKLGTQTVPCNKILLWSRIKDLAHQFTQVQRDMFTLEDTLL 
               
               
                   
               
               
                 GYLADDLTWCGEFNTSKINYQSCPDWRKDCSNNPVSVFWKTVSRRFAEAA 
               
               
                   
               
               
                 CDVVHVMLNGSRSKIFDKNSTFGSVEVHNLQPEKVQTLEAWVIHGGREDS 
               
               
                   
               
               
                 RDLCQDPTIKELESIISKRNIQFSCKNIYRPDKFLQCVKNPEDSSCTSE 
               
               
                   
               
               
                 I. 
               
            
           
         
       
     
     “MOR202” an anti-CD38 antibody whose amino acid sequence is provided in  FIG. 11 . “MOR202” and “MOR03087” are used as synonyms to describe the antibody shown in  FIG. 11 . 
     The DNA sequence encoding the MOR202 Variable Heavy Domain is: 
     
       
         
           
               
            
               
                 (SEQ ID NO: 12) 
               
            
           
           
               
            
               
                 CAGGTGCAATTGGTGGAAAGCGGCGGCGGCCTGGTGCAACCGGGCGGCAG 
               
               
                   
               
               
                 CCTGCGTCTGAGCTGCGCGGCCTCCGGATTTACCTTTTCTTCTTATTATA 
               
               
                   
               
               
                 TGAATTGGGTGCGCCAAGCCCCTGGGAAGGGTCTCGAGTGGGTGAGCGGT 
               
               
                   
               
               
                 ATCTCTGGTGATCCTAGCAATACCTATTATGCGGATAGCGTGAAAGGCCG 
               
               
                   
               
               
                 TTTTACCATTTCACGTGATAATTCGAAAAACACCCTGTATCTGCAAATGA 
               
               
                   
               
               
                 ACAGCCTGCGTGCGGAAGATACGGCCGTGTATTATTGCGCGCGTGATCTT 
               
               
                   
               
               
                 CCTCTTGTTTATACTGGTTTTGCTTATTGGGGCCAAGGCACCCTGGTGAC 
               
               
                   
               
               
                 GGTTAGCTCA 
               
            
           
         
       
     
     The DNA sequence encoding the MOR202 Variable Light Domain is: 
     
       
         
           
               
            
               
                 (SEQ ID NO: 13) 
               
            
           
           
               
            
               
                 GATATCGAACTGACCCAGCCGCCTTCAGTGAGCGTTGCACCAGGTCAGAC 
               
               
                   
               
               
                 CGCGCGTATCTCGTGTAGCGGCGATAATCTTCGTCATTATTATGTTTATT 
               
               
                   
               
               
                 GGTACCAGCAGAAACCCGGGCAGGCGCCAGTTCTTGTGATTTATGGTGAT 
               
               
                   
               
               
                 TCTAAGCGTCCCTCAGGCATCCCGGAACGCTTTAGCGGATCCAACAGCGG 
               
               
                   
               
               
                 CAACACCGCGACCCTGACCATTAGCGGCACTCAGGCGGAAGACGAAGCGG 
               
               
                   
               
               
                 ATTATTATTGCCAGACTTATACTGGTGGTGCTTCTCTTGTGTTTGGCGGC 
               
               
                   
               
               
                 GGCACGAAGTTAACCGTTCTTGGCCAG 
               
            
           
         
       
     
     Antibody “Ref mAB5” is an anti-CD38 antibody whose amino acid sequence is provided below (the CDRs are bolded and underlined): 
     
       
         
           
               
            
               
                 VH: 
               
            
           
           
               
            
               
                 (SEQ ID NO: 21) 
               
            
           
           
               
            
               
                 QVQLVQSGAEVAKPGTSVKLSCKASGYTFT   DYWMQ   WVKQRPGQGLEWIG   T     
               
               
                   
               
               
                     IYPGDGDTGYAQKFQG   KATLTADKSSKTVYMHLSSLASEDSAVYYCAR   GD     
               
               
                   
               
               
                     YYGSNSLDY   WGQGTSVTVSS 
               
               
                   
               
               
                 VL: 
               
            
           
           
               
            
               
                 (SEQ ID NO: 22) 
               
            
           
           
               
            
               
                 DIVMTQSHLSMSTSLGDPVSITC   KASQDVSTVVA   WYQQKPGQSPRRLIY   S     
               
               
                   
               
               
                     ASYRYI   GVPDRFTGSGAGTDFTFTISSVQAEDLAVYYC   QQHYSPPYT   FGG 
               
               
                   
               
               
                 GTKLEIKRT 
               
            
           
         
       
     
     The CDRs of Ref mAB5 are defined by Kabat et al. and an antibody having the same CDRs as Ref mAB5 is described in WO2008/047242, U.S. Ser. No. 12/441,466, which is incorporated by reference in its entirety. 
     “Fc region” means the constant region of an antibody, which in humans may be of the IgG1, 2, 3, 4 subclass or others. The sequences of human Fc regions are available at the website for ImMunoGeneTics (IMGT) Repertoire for IG and TR, Human IGH C-REGIONs and were retrieved on May 16, 2011. 
     “Enhances ADCC activity” means an increase in the mediation of antibody dependent cell-mediated cytotoxicity. Amino acid modifications within the Fc region that result in an enhacement of ADCC activity are disclosed in WO200042072 Genentech, WO2004029207A2 Xencor, and WO2004063351A2 Macrogenics, which are all incorporated by reference in their entireties. 
     “MOR03207” is an antibody whose amino acid sequence is: 
     
       
         
           
               
            
               
                 VH: 
               
            
           
           
               
            
               
                 (SEQ ID NO: 8) 
               
            
           
           
               
            
               
                 QVQLQQSGPGLVKPSQTLSLTCAISGDSVSSNSAAWSWIRQSPGRGLEWL 
               
               
                   
               
               
                 GRIYYRSKWYNDYAVSVKSRITINPDTSKNQFSLQLNSVTPEDTAVYYCA 
               
               
                   
               
               
                 RLDHRYHEDTVYPGMDVWGQGTLVTVSS 
               
               
                   
               
               
                 VL: 
               
            
           
           
               
            
               
                 (SEQ ID NO: 9) 
               
            
           
           
               
            
               
                 DIELTQPPSVSVAPGQTARISCSGDNLPAYTVTWYQQKPGQAPVLVIYDD 
               
               
                   
               
               
                 SDRPSGIPERFSGSNSGNTATLTISGTQAEDEADYYCASWDPSSGVVFGG 
               
               
                   
               
               
                 GTKLTVLGQ. 
               
            
           
         
       
     
     MOR03207 binds lysosyme, and is used as isotype control, as it is IgG1. 
     A “combination” means more than one item, e.g. a compound such as an antibody and lenalidomide. 
     The present disclosure also relates to combinations, pharmaceuticals, and pharmaceutical compositions containing the described combinations. The two components of the synergistic combination of the present invention, e.g. the antibody specific for CD38 and lenalidomide, may be administered together, or separately. When administered together, the two components may be formulated together in one pharmaceutical composition, which may include a pharmaceutical acceptable carrier or excipient. Alternatively the two components might also be formulated in different pharmaceutical compositions. In this case the two components can be administered simultaneously or subsequently. In an embodiment, the thalidomide or an analog thereof, e.g. lenalidomide, is administered prior to and/or separately from the administration of the antibody specific for CD38, e.g. MOR202. In a further embodiment, lenalidomide, is administered at least 72 hours prior to administration of the antibody specific for CD38, e.g. MOR202. This time period allows for lenalidomide mediated upregulation of CD38 in the target cells. 
     A pharmaceutical composition includes an active agent, e.g. an antibody for therapeutic use in humans. A pharmaceutical composition may include acceptable carriers or excipients. 
     “Administered” or “administration” includes but is not limited to delivery by an injectable form, such as, for example, an intravenous, intramuscular, intradermal or subcutaneous route or mucosal route, for example, as a nasal spray or aerosol for inhalation or as an ingestible solution, capsule or tablet. 
     A “therapeutically effective amount” of a compound or combination refers to an amount sufficient to cure, alleviate or partially arrest the clinical manifestations of a given disease or disorder and its complications. The amount that is effective for a particular therapeutic purpose will depend on the severity of the disease or injury as well as on the weight and general state of the subject. It will be understood that determination of an appropriate dosage may be achieved, using routine experimentation, by constructing a matrix of values and testing different points in the matrix, all of which is within the ordinary skills of a trained physician or clinical scientist. 
     Surprisingly, it was found that the combination of a particular anti-CD38 antibody and lenalidomide mediated a synergistic level of Antibody-Dependent Cell-Mediated Cytotoxicity (ADCC) in both AMO-1 and NCI-H929 multiple myeloma cells. In addition, and also unexpectedly, a particular anti-CD38 antibody when combined with lenalidomide mediated a synergistic level of reduction in bone lysis in the NCI-H929 SCID mouse model and synergistically increased the median survival days in the RAMOS SCID mouse model. Therefore, the combination of the exemplified antibody specific for CD38 and lenalidomide behaved synergistically in both the in vitro and in vivo models relevant to multiple myeloma and/or non-Hodgkin&#39;s lymphoma. Therefore, this combination yields synergistic results in the treatment of multiple myeloma and/or non-Hodgkin&#39;s lymphoma in humans. 
     Lenalidomide is a thalidomide analog, therefore, it is expected that other thalidomide analogs, such as, pomalidomide or thalidomide itself also lead to synergistic effects when used in combination with an anti-CD38 antibody. In addition, as thalidomide or an analog thereof upregulate CD38 expression in multiple myeloma cell lines, therefore, it is expected that synergism should result when other agents that upregulate the expression of CD38 on the surface of tumor cells, e.g. trans-retinoic acid, and anti-CD38 antibodies are used in combination. 
     Surprisingly, it was found that the combination of a particular anti-CD38 antibody and bortezomib mediated a high level of Antibody-Dependent Cell-Mediated Cytotoxicity (ADCC) in the NCI-H929 and LP-1 multiple myeloma cell lines. In addition, and also surprisingly it was found that the combination of a particular anti-CD38 antibody and bortezomib mediated a synergistic level of reduction in bone lysis in the NCI-H929 SCID mouse model and synergistically increased the median survival days in the RAMOS SCID mouse model. Therefore, the combination of the exemplified antibody specific for CD38 and bortezomib behaved synergistically in the in vivo models relevant to multiple myeloma and/or non-Hodgkin&#39;s lymphoma. Therefore, this combination yields synergistic results in the treatment of multiple myeloma and/or non-Hodgkin&#39;s lymphoma in humans. 
     It is expected that other proteasome inhibitors, such as, Disulfiram, Epigallocatechin-3-gallate, and Salinosporamide A will lead to similar effects when used in combination with an anti-CD38 antibody. 
     The “CDRs” herein are defined by either Chothia et al., Kabat et al. or by an internal numbering convention. See Chothia C, Lesk AM. (1987) Canonical structures for the hypervariable regions of immunoglobulins. J Mol Biol., 196(4):901-17, which is incorporated by reference in its entirety. See Kabat E. A, Wu T. T., Perry H. M., Gottesman K. S. and Foeller C. (1991). Sequences of Proteins of Immunological Interest. 5th edit., NIH Publication no. 91-3242, US Dept. of Health and Human Services, Washington, D.C., which is incorporated by reference in its entirety. 
     Embodiments 
     An aspect of the present disclosure comprises a synergistic combination of an antibody specific for CD38 and (a) thalidomide or an analog thereof, or (b) a proteasome inhibitor, for use in the treatment of multiple myeloma and/or non-hodgkins lymphoma. 
     An aspect of the present disclosure comprises a combination of an antibody specific for CD38 and thalidomide or an analog thereof. In embodiments, the combination is synergistic. 
     In embodiments, the antibody specific for CD38 comprises an HCDR1 region of sequence GFTFSSYYMN (SEQ ID NO: 1) or of sequence SYYMN (SEQ ID NO: 14), an HCDR2 region of sequence GISGDPSNTYYADSVKG (SEQ ID NO: 2), an HCDR3 region of sequence DLPLVYTGFAY (SEQ ID NO: 3), an LCDR1 region of sequence SGDNLRHYYVY (SEQ ID NO: 4), an LCDR2 region of sequence GDSKRPS (SEQ ID NO: 5), and an LCDR3 region of sequence QTYTGGASL (SEQ ID NO: 6). 
     In embodiments, the antibody specific for CD38 comprises an HCDR1 region of sequence DYWMQ (SEQ ID NO: 15), an HCDR2 region of sequence TIYPGDGDTGYAQKFQG (SEQ ID NO: 16), an HCDR3 region of sequence GDYYGSNSLDY (SEQ ID NO: 17), an LCDR1 region of sequence KASQDVSTWA (SEQ ID NO: 18), an LCDR2 region of sequence SASYRYI (SEQ ID NO: 19), and an LCDR3 region of sequence QQHYSPPYT (SEQ ID NO: 20). 
     In an aspect the combination is used for the treatment of multiple myeloma and/or non-hodgkins lymphoma. Embodiments comprise a combination, wherein the thalidomide analog is lenalidomide. 
     An aspect relates to pharmaceutical compositions comprising the combinations. In embodiments, the composition comprises an acceptable carrier. In embodiments, the composition is administered in an effective amount. 
     An aspect of the present disclosure comprises a synergistic combination of an antibody specific for CD38 comprising an HCDR1 region of sequence GFTFSSYYMN (SEQ ID NO: 1) or of sequence SYYMN (SEQ ID NO: 14), an HCDR2 region of sequence GISGDPSNTYYADSVKG (SEQ ID NO: 2), an HCDR3 region of sequence DLPLVYTGFAY (SEQ ID NO: 3), an LCDR1 region of sequence SGDNLRHYYVY (SEQ ID NO: 4), an LCDR2 region of sequence GDSKRPS (SEQ ID NO: 5), and an LCDR3 region of sequence QTYTGGASL (SEQ ID NO: 6) and lenalidomide for the treatment of multiple myeloma and/or non-hodgkins lymphoma. 
     A further embodiment comprises a combination, wherein the antibody comprises a variable heavy chain of the sequence QVQLVESGGGLVQPGGSLRLSCAASGFTFSSYYMNWVRQAPGKGLEWVSGISGDPSNT YYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARDLPLVYTGFAYWGQGTLVTV SS (SEQ ID NO: 10) and a variable light chain of the sequence DIELTQPPSVSVAPGQTARISCSGDNLRHYYVYWYQQKPGQAPVLVIYGDSKRPSGIPER FSGSNSGNTATLTISGTQAEDEADYYCQTYTGGASLVFGGGTKLTVLGQ (SEQ ID NO: 11). 
     An aspect of the present disclosure comprises a synergistic combination of an antibody specific for CD38 comprising an HCDR1 region of sequence DYWMQ (SEQ ID NO: 15), an HCDR2 region of sequence TIYPGDGDTGYAQKFQG (SEQ ID NO: 16), an HCDR3 region of sequence GDYYGSNSLDY (SEQ ID NO: 17), an LCDR1 region of sequence KASQDVSTVVA (SEQ ID NO: 18), an LCDR2 region of sequence SASYRYI (SEQ ID NO: 19), and an LCDR3 region of sequence QQHYSPPYT (SEQ ID NO: 20) and lenalidomide for the treatment of multiple myeloma and/or non-hodgkins lymphoma. 
     A further embodiment comprises a combination, wherein the antibody comprises a variable heavy chain of the sequence QVQLVQSGAEVAKPGTSVKLSCKASGYTFTDYWMQWVKQRPGQGLEWIGTIYPGDGDT GYAQKFQGKATLTADKSSKTVYMHLSSLASEDSAVYYCARGDYYGSNSLDYWGQGTSV TVSS (SEQ ID NO: 21) and a variable light chain of the sequence DIVMTQSHLSMSTSLGDPVSITCKASQDVSTVVAWYQQKPGQSPRRLIYSASYRYIGVPD RFTGSGAGTDFTFTISSVQAEDLAVYYCQQHYSPPYTFGGGTKLEIKRT (SEQ ID NO: 22). 
     In embodiments the antibody has an IgG1 Fc region. In embodiments the antibody comprises a modified Fc region, wherein said modification enhances ADCC activity. 
     In another aspect, the components of the combination, the antibody specific for CD38 and lenalidomide, are administered separately. In an embodiment, lenalidomide is administered prior to administration of the antibody specific for CD38. In a further embodiment, lenalidomide is administered at least 72 hours prior to administration of the antibody specific for CD38. 
     In another aspect the synergistic combination of an antibody specific for CD38 comprising an HCDR1 region of sequence GFTFSSYYMN (SEQ ID NO: 1) or of sequence SYYMN (SEQ ID NO: 14), an HCDR2 region of sequence GISGDPSNTYYADSVKG (SEQ ID NO: 2), an HCDR3 region of sequence DLPLVYTGFAY (SEQ ID NO: 3), an LCDR1 region of sequence SGDNLRHYYVY (SEQ ID NO: 4), an LCDR2 region of sequence GDSKRPS (SEQ ID NO: 5), and an LCDR3 region of sequence QTYTGGASL (SEQ ID NO: 6) and lenalidomide is able to mediate killing of CD38-expressing AMO-1 cells and/or NCI-H929 cells by ADCC in the presence of isolated human PBMCs with an at least two-fold, three-fold, four-fold, or five-fold better efficacy than lenalidomide alone. 
     In another aspect the synergistic combination of an antibody specific for CD38 comprising an HCDR1 region of sequence GFTFSSYYMN (SEQ ID NO: 1) or of sequence SYYMN (SEQ ID NO: 14), an HCDR2 region of sequence GISGDPSNTYYADSVKG (SEQ ID NO: 2), an HCDR3 region of sequence DLPLVYTGFAY (SEQ ID NO: 3), an LCDR1 region of sequence SGDNLRHYYVY (SEQ ID NO: 4), an LCDR2 region of sequence GDSKRPS (SEQ ID NO: 5), and an LCDR3 region of sequence QTYTGGASL (SEQ ID NO: 6) and lenalidomide is able to reduce bone lysis with an at least two-fold, three-fold, four-fold, or five-fold better efficacy than lenalidomide alone. 
     Another aspect comprises a method of treating multiple myeloma and/or non-hodgkins lymphoma in an individual in need thereof, which method comprises administration of an antibody specific for CD38 comprising an HCDR1 region of sequence GFTFSSYYMN (SEQ ID NO: 1) or of sequence SYYMN (SEQ ID NO: 14), an HCDR2 region of sequence GISGDPSNTYYADSVKG (SEQ ID NO: 2), an HCDR3 region of sequence DLPLVYTGFAY (SEQ ID NO: 3), an LCDR1 region of sequence SGDNLRHYYVY (SEQ ID NO: 4), an LCDR2 region of sequence GDSKRPS (SEQ ID NO: 5), and an LCDR3 region of sequence QTYTGGASL (SEQ ID NO: 6) and lenalidomide to an individual having multiple myeloma or non-hodgkins lymphoma. In embodiments, the combination is administered in an effective amount. 
     Another aspect comprises a combination comprising an antibody specific for CD38 comprising an HCDR1 region of sequence GFTFSSYYMN (SEQ ID NO: 1) or of sequence SYYMN (SEQ ID NO: 14), an HCDR2 region of sequence GISGDPSNTYYADSVKG (SEQ ID NO: 2), an HCDR3 region of sequence DLPLVYTGFAY (SEQ ID NO: 3), an LCDR1 region of sequence SGDNLRHYYVY (SEQ ID NO: 4), an LCDR2 region of sequence GDSKRPS (SEQ ID NO: 5), and an LCDR3 region of sequence QTYTGGASL (SEQ ID NO: 6) and lenalidomide. In an embodiment, the combination is used for the treatment of cancer. In a further embodiment, the cancer is selected from multiple myeloma, and non-hodgkins lymphoma. 
     Another aspect comprises a combination of an antibody specific for CD38 and a proteasome inhibitor. In embodiments, the combination is synergistic. In embodiments, the antibody specific for CD38 comprises an HCDR1 region of sequence GFTFSSYYMN (SEQ ID NO: 1) or of sequence SYYMN (SEQ ID NO: 14), an HCDR2 region of sequence GISGDPSNTYYADSVKG (SEQ ID NO: 2), an HCDR3 region of sequence DLPLVYTGFAY (SEQ ID NO: 3), an LCDR1 region of sequence SGDNLRHYYVY (SEQ ID NO: 4), an LCDR2 region of sequence GDSKRPS (SEQ ID NO: 5), and an LCDR3 region of sequence QTYTGGASL (SEQ ID NO: 6). 
     In an aspect the combination is used for the treatment of multiple myeloma and/or non-hodgkins lymphoma. In embodiments, the combination comprises a proteasome inhibitor, which is bortezomib. An aspect relates to pharmaceutical compositions comprising the combinations. In embodiments, the composition comprises an acceptable carrier. In embodiments, the composition is administered in an effective amount. 
     An aspect of the present disclosure comprises a synergistic combination of an antibody specific for CD38 comprising an HCDR1 region of sequence GFTFSSYYMN (SEQ ID NO: 1) or of sequence SYYMN (SEQ ID NO: 14), an HCDR2 region of sequence GISGDPSNTYYADSVKG (SEQ ID NO: 2), an HCDR3 region of sequence DLPLVYTGFAY (SEQ ID NO: 3), an LCDR1 region of sequence SGDNLRHYYVY (SEQ ID NO: 4), an LCDR2 region of sequence GDSKRPS (SEQ ID NO: 5), and an LCDR3 region of sequence QTYTGGASL (SEQ ID NO: 6) and bortezomib for the treatment of multiple myeloma and/or non-hodgkins lymphoma. 
     A further embodiment comprises a combination, wherein the antibody comprises a variable heavy chain of the sequence QVQLVESGGGLVQPGGSLRLSCAASGFTFSSYYMNWVRQAPGKGLEWVSGISGDPSNT YYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARDLPLVYTGFAYWGQGTLVTV SS (SEQ ID NO: 10) and a variable light chain of the sequence DIELTQPPSVSVAPGQTARISCSGDNLRHYYVYWYQQKPGQAPVLVIYGDSKRPSGIPER FSGSNSGNTATLTISGTQAEDEADYYCQTYTGGASLVFGGGTKLTVLGQ (SEQ ID NO: 11). 
     In embodiments the antibody has an IgG1 Fc region. In embodiments the antibody comprises a modified Fc region, wherein said modification enhances ADCC activity. 
     In an embodiment, the combination is used for the treatment of cancer. In a further embodiment, the cancer is selected from multiple myeloma, and non-hodgkins lymphoma. 
     In another aspect, the components of the combination, the antibody and proteasome inhibitor, are administered separately. 
     In another aspect the synergistic combination of an antibody specific for CD38 comprising an HCDR1 region of sequence GFTFSSYYMN (SEQ ID NO: 1) or of sequence SYYMN (SEQ ID NO: 14), an HCDR2 region of sequence GISGDPSNTYYADSVKG (SEQ ID NO: 2), an HCDR3 region of sequence DLPLVYTGFAY (SEQ ID NO: 3), an LCDR1 region of sequence SGDNLRHYYVY (SEQ ID NO: 4), an LCDR2 region of sequence GDSKRPS (SEQ ID NO: 5), and an LCDR3 region of sequence QTYTGGASL (SEQ ID NO: 6) and bortezomib is able to mediate killing of CD38-expressing LP-1 cells and/or NCI-H929 cells by ADCC in the presence of isolated human PBMCs with an at least two-fold, three-fold, four-fold, or five-fold better efficacy than bortezomib alone. 
     In another aspect the synergistic combination of an antibody specific for CD38 comprising an HCDR1 region of sequence GFTFSSYYMN (SEQ ID NO: 1) or of sequence SYYMN (SEQ ID NO: 14), an HCDR2 region of sequence GISGDPSNTYYADSVKG (SEQ ID NO: 2), an HCDR3 region of sequence DLPLVYTGFAY (SEQ ID NO: 3), an LCDR1 region of sequence SGDNLRHYYVY (SEQ ID NO: 4), an LCDR2 region of sequence GDSKRPS (SEQ ID NO: 5), and an LCDR3 region of sequence QTYTGGASL (SEQ ID NO: 6) and bortezomib is able to reduce bone lysis with an at least two-fold, three-fold, four-fold, or five-fold better efficacy than bortezomib alone. 
     In another aspect, the present disclosure comprises a method of treating multiple myeloma and/or non-hodgkins lymphoma in an individual in need thereof, which method comprises administration of an antibody specific for CD38 comprising an HCDR1 region of sequence GFTFSSYYMN (SEQ ID NO: 1) or of sequence SYYMN (SEQ ID NO: 14), an HCDR2 region of sequence GISGDPSNTYYADSVKG (SEQ ID NO: 2), an HCDR3 region of sequence DLPLVYTGFAY (SEQ ID NO: 3), an LCDR1 region of sequence SGDNLRHYYVY (SEQ ID NO: 4), an LCDR2 region of sequence GDSKRPS (SEQ ID NO: 5), and an LCDR3 region of sequence QTYTGGASL (SEQ ID NO: 6) and bortezomib to an individual having multiple myeloma or non-hodgkins lymphoma. 
     In embodiments, the combination is administered in an effective amount. 
     Another aspect comprises a combination comprising an antibody specific for CD38 comprising an HCDR1 region of sequence GFTFSSYYMN (SEQ ID NO: 1) or of sequence SYYMN (SEQ ID NO: 14), an HCDR2 region of sequence GISGDPSNTYYADSVKG (SEQ ID NO: 2), an HCDR3 region of sequence DLPLVYTGFAY (SEQ ID NO: 3), an LCDR1 region of sequence SGDNLRHYYVY (SEQ ID NO: 4), an LCDR2 region of sequence GDSKRPS (SEQ ID NO: 5), and an LCDR3 region of sequence QTYTGGASL (SEQ ID NO: 6) and bortezomib. 
     An aspect comprises a synergistic combination of an antibody specific for CD38 comprising an HCDR1 region of sequence GFTFSSYYMN (SEQ ID NO: 1) or of sequence SYYMN (SEQ ID NO: 14), an HCDR2 region of sequence GISGDPSNTYYADSVKG (SEQ ID NO: 2), an HCDR3 of sequence DLPLVYTGFAY (SEQ ID NO: 3), an LCDR1 region of sequence SGDNLRHYYVY (SEQ ID NO: 4), an LCDR2 region of sequence GDSKRPS (SEQ ID NO: 5), and an LCDR3 region of sequence QTYTGGASL (SEQ ID NO: 6) and 
     (a) thalidomide or an analog thereof, or 
     (b) a proteasome inhibitor, 
     for use in the treatment of multiple myeloma and/or non-hodgkins lymphoma. 
     Embodiments comprise a combination, wherein the antibody comprises a variable heavy chain of the sequence QVQLVESGGGLVQPGGSLRLSCAASGFTFSSYYMNWVRQAPGKGLEWVSGISGDPSNT YYADSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARDLPLVYTGFAYWGQGTLVTV SS (SEQ ID NO: 10) and a variable light chain of the sequence DIELTQPPSVSVAPGQTARISCSGDNLRHYYVYWYQQKPGQAPVLVIYGDSKRPSGIPER FSGSNSGNTATLTISGTQAEDEADYYCQTYTGGASLVFGGGTKLTVLGQ (SEQ ID NO: 11). 
     Embodiments comprise a combination, wherein the antibody comprises an IgG1 Fc region. Embodiments comprise a combination, wherein the antibody comprises a modified Fc region, wherein said modification enhances ADCC activity. 
     Embodiments comprise a combination, wherein said antibody specific for CD38 and said thalidomide or an analog thereof or proteasome inhibitor are administered separately. 
     Embodiments comprise a combination, which is able to reduce bone lysis with an at least two-fold better efficacy than lenalidomide and/or bortezomib alone. 
     Embodiments comprise a combination, wherein said antibody specific for CD38 is combined with thalidomide or an analog thereof. Embodiments comprise a combination, wherein the thalidomide analog comprises lenalidomide. Embodiments comprise a combination, wherein lenalidomide is administered prior to administration of the antibody specific for CD38. Embodiments comprise a combination, wherein lenalidomide is administered at least 72 hours prior to administration of the antibody specific for CD38. 
     Embodiments comprise a combination of an antibody specific for CD38 and lenalidomide, which is able to mediate killing of CD38-expressing AMO-1 and/or NCI-H929 cells by ADCC in the presence of isolated human PBMCs with an at least two-fold better efficacy than lenalidomide alone. 
     Embodiments comprise a combination, comprising said antibody specific for CD38 and a proteasome inhibitor. In some embodiments, the proteasome inhibitor is bortezomib. Embodiments comprise a combination of an antibody specific for CD38 and bortezomib, which is able to mediate killing of CD38-expressing LP-1 and/or NCI-H929 cells by ADCC in the presence of isolated human PBMCs with an at least two-fold better efficacy than bortezomib alone. 
     An aspect comprises a synergistic combination of an antibody specific for CD38 comprising an HCDR1 region of sequence GFTFSSYYMN (SEQ ID NO: 1) or of sequence SYYMN (SEQ ID NO: 14), an HCDR2 region of sequence GISGDPSNTYYADSVKG (SEQ ID NO: 2), an HCDR3 region of sequence DLPLVYTGFAY (SEQ ID NO: 3), an LCDR1 region of sequence SGDNLRHYYVY (SEQ ID NO: 4), an LCDR2 region of sequence GDSKRPS (SEQ ID NO: 5), and an LCDR3 region of sequence QTYTGGASL (SEQ ID NO: 6) and lenalidomide or other thalidomide analog for use in the treatment of multiple myeloma and/or non-hodgkins lymphoma. 
     An aspect comprises a synergistic combination of an antibody specific for CD38 comprising an HCDR1 region of sequence GFTFSSYYMN (SEQ ID NO: 1) or of sequence SYYMN (SEQ ID NO: 14), an HCDR2 region of sequence GISGDPSNTYYADSVKG (SEQ ID NO: 2), an HCDR3 region of sequence DLPLVYTGFAY (SEQ ID NO: 3), an LCDR1 region of sequence SGDNLRHYYVY (SEQ ID NO: 4), an LCDR2 region of sequence GDSKRPS (SEQ ID NO: 5), and an LCDR3 region of sequence QTYTGGASL (SEQ ID NO: 6) and bortezomib or other proteasome inhibitor for use in the treatment of multiple myeloma and/or non-hodgkins lymphoma. 
     EXAMPLES 
     Example 1 
     CD38 Expression on the Surface of Various Cell Lines 
     The cell lines of Table 1 were tested for levels of CD38 expression. 
     
       
         
           
               
               
               
             
               
                 TABLE 1 
               
               
                   
               
               
                 Cell Line 
                 Supplied by: 
                 Cultivated in: 
               
               
                   
               
             
            
               
                 AMO-1: Multiple Myeloma 
                 DSMZ #ACC 538 
                 RPMI1640, with 
               
               
                 Cell Line 
                   
                 L-Glutamine, (PAN Biotech 
               
               
                   
                   
                 GmbH, Cat No.: P04-16500 
               
               
                   
                   
                 medium) 
               
               
                 LP1: Multiple Myeloma Cell 
                 DSMZ #ACC 41 
                 Iscove&#39;s Modified Dulbecco&#39;s 
               
               
                 Line 
                   
                 Medium (IMDM) with 
               
               
                   
                   
                 GlutaMAX ™ (Invitrogen, Cat 
               
               
                   
                   
                 No.: 31980-048) 
               
               
                 NCI-H929: Multiple Myeloma 
                 DSMZ #ACC 163 
                 RPMI1640 (same as 
               
               
                 Cell Line 
                   
                 AMO-1), supplemented with 
               
               
                   
                   
                 1 mM Na-Pyruvate, 50 μM 
               
               
                   
                   
                 β-Mercaptoethanol 
               
               
                 RPMI8226: Multiple 
                 DSMZ #ACC 402 
                 RPMI1640 (same as AMO-1) 
               
               
                 Myeloma Cell Line 
                   
                   
               
               
                 OPM-2: Multiple Myeloma 
                 DSMZ #ACC 50 
                 RPMI1640 (same as AMO-1) 
               
               
                 Cell Line 
                   
                   
               
               
                 Plasmacytoma, Malignant 
                 Klinikum rechts der Isar 
                 RPMI1640 (same as AMO-1) 
               
               
                 Plasma Cells 
               
               
                   
               
            
           
         
       
     
     Bone marrow samples (4-10 ml aspirate) from multiple myeloma patients and extramedullary tumor plasmacytoma samples were obtained after informed consent from the Klinikum rechts der Isar (“Krdl”) (Munich, Germany). Samples were subjected to centrifugation, and further plasma cell enrichment was achieved via magnetic-activated cell sorting. 
     Cells were stained with a directly labelled QuantiBRITE™ CD38-PE antibody (Becton Dickinson GmbH, Clone HB7, CAT #342371), which is specific for CD38. The “Antibodies Bound Per Cell” (ABC&#39;s) were determined using the flow cytometry based QuantiBRITE™ system, which measures the geometric mean (GeoMean) per cell. Conversion of measured GeoMean into correlating ABC amount per cell was done with GraphPad PRISM™ software. The ABC values are assumed to correlate with the number of CD38 molecules per cell, since QuantiBRITE™ CD38-PE carries one PE molecule per antibody. The results are shown in Table 2. 
     Example 2 
     Evaluation of Effect of Lenalidomide on Upregulation of CD38 in Various Cell Lines 
     To determine whether lenalidomide induced upregulation of CD38 in the multiple myeloma and plasmacytoma cells of Table 1, the cell lines were incubated with 100 μM lenalidomide and, subsequently, CD38 surface expression was analyzed by FACS. 
     Materials and Methods 
     Around 2×10 5  cells of each of the cells lines of Table 1 were plated on 48-well dishes in standard RPMI medium. Lenalidomide, purchased from Selleck Chemicals (LLC S1029, CAS No. 191732-6; Batch: S10290), was applied to respective wells to a final concentration of 100 μM in a volume of 750 μl containing 20% FCS and 0.1% DMSO. As negative control 0.1% DMSO in FCS-supplemented medium was used and plates were incubated for 24 h, 48 h and 72 h at 37° C. and 5% CO 2  in humidified incubator. 
     Cells were resuspended by gentle pipetting and 250 μl of cell suspension per incubation period were transferred into a well of a 96-well round bottom plate. Cells were washed by centrifugation for 1 min at 700×g and were resuspended in 150 μl of cold FACS buffer (1×PBS supplemented with 3% FCS). Cells were again pelleted down by centrifugation and were resuspended in 150 μl of FACS buffer containing 15 μg/ml of anti-CD38 antibody (MOR202, IgG1) or control antibody MOR03207 and incubated for 1 h on ice. Cells were then washed 3 times by centrifugation and were resuspended in FACS buffer supplemented with PE-labeled secondary antibody (PE-Fab 2  fragment, goat anti-human IgG, Fc-fragment specific; Jackson Immuno Research; CAT: 109-116-098; Lot: 80938). Cells were incubated for 45 minutes on ice, then washed 3 times by centrifugation and resuspended in FACS buffer. The cell suspensions were then subjected to FACS analysis using a FACS array device. 
     The basal CD38 expression of each cell line and the affect of lenalidomide on CD38 expression are shown in Table 2. Additionally, the affect of lenalidomide on the CD38 expression of AMO-1 cells is shown in  FIG. 1 , and the affect of lenalidomide on the CD38 expression of NCI-H929 cells is shown in  FIG. 5 . 
     
       
         
           
               
               
               
               
             
               
                 TABLE 2 
               
             
            
               
                   
               
               
                   
                 Absolute number of ABC 
                   
                   
               
               
                   
                 (CD38 expression) 
                   
                   
               
            
           
           
               
               
               
               
               
               
            
               
                   
                   
                 LEN 
                   
                 Fold 
                   
               
               
                   
                   
                 (extrap- 
                 IN- 
                 in- 
                   
               
               
                 Cell line 
                 Basal 
                 olated) 
                 CREASE 
                 crease 
                 Effect 
               
               
                   
               
            
           
           
               
               
               
               
               
               
            
               
                 AMO-1 
                 25,000 
                 115,000 
                 90,000 
                 4.6 
                 Significant 
               
               
                 LP-1 
                 125,000 
                 162,500 
                 37,500 
                 1.3 
                 No 
               
               
                 NCI-H929 
                 195,000 
                 390,000 
                 195,000 
                 2.0 
                 Weak 
               
               
                 RPMI-8226 
                 670,000 
                 871,000 
                 199,000 
                 1.3 
                 Weak 
               
               
                 OPM-2 
                 38,000 
                 98,800 
                 60,800 
                 2.6 
                 Significant 
               
               
                 Plasma- 
                 30,000 
                 69,000 
                 39,000 
                 2.3 
                 Significant 
               
               
                 cytoma 
               
               
                   
               
            
           
         
       
     
     Example 3 
     Inhibition of Proliferation of AMO-1 Cells Using Lenalidomide Alone 
     The cytotoxicity of Lenalidomide was tested in AMO-1 cells. Cells were collected and distributed in 96-well plates with 5000 cells per well. Increasing amounts of Lenalidomide were added to the wells and plates were incubated for 24 h, 48 h and 72 h at 37° C. in a humidified incubator (5% CO 2 ). 
     After incubation, plates were analyzed for cell proliferation in a quantitative colorimietric XTT-based assay using the cell proliferation kit II (ROCHE, Cell Proliferation Kit II, Cat. No.: 11465015001). For subsequent measurement plates were subjected to Tecan Genios Reader and absorbance at 492 nm was detected. 
     The results are shown in  FIG. 2 . 
     Example 4 
     Synergistic Combination of MOR0202 and Lenalidomide in AMO-1 Cells 
     AMO-1 cells were selected for testing with the combination of MOR202 and lenalidomide. AMO-1 cells are similar to plasmacytoma cells in humans in that both have a low basal CD38 expression, and CD38 is significantly upregulated in both upon treatment with lenalidomide as shown in Table 2. 
     PBMC&#39;s were isolated by density gradient centrifugation of freshly isolated human blood. Isolated blood from different donors were layered on a defined volume of Biocoll (Biochrome AG; CAT No.:L6115; LOT No.:1050T) in a Falcon tube and centrifuged at 380 g. The PBMCs were isolated and supplemented with RPMI medium. 
     After 72 h, cells were counted and the PBMCs were adjusted to a concentration of 6.6×106/ml while the AMO-1 cells were adjusted to a final concentration of 2.5×10 5 /ml. For later identification in flow cytometry, the AMO-1 cells were stained for 3 min with 0.1 μg/ml of CalceinAM (Calcein: 1 mg/ml stock solution, Invitrogen, Cat No.: C3099) and washed three times by gentle centrifugation. 100 μl of target cell suspension were mixed with 100 μl of PBMCs to achieve a ratio of 1:30. Antibody MOR202 or antibody MOR03207 (negative control) were added to a final concentration of 15 μg/ml. Cell suspensions were further incubated for 4 h at 37° C. To detect dead AMO-1 cells, cell suspensions were challenged with propidium iodide (PI) and subsequently analyzed in flow cytometry. Target cells were separated via gating of CalceinAM positive cell populations, and cells killed via ADCC were quantified. 
     In total six experiments were performed in order to determine the mediation of ADCC on AMO-1 cells by the combination of MOR202 and lenalidomide. In three experiments, the PBMCs and AMO-1 cells were treated with lenalidomide prior to treatment with MOR202, the results are shown in Tables 3 a-c and  FIG. 3 . In three additional experiments, only the PBMCs were treated with lenalidomide prior to treatment with MOR202, the results are shown in Tables 4 a-c and  FIG. 4 . 
     Table 3 Both Effector and AMO-1 cells were treated with Lenalidomide prior to treatment with MOR202. Single and combination doses of 10 μM LEN and 15 μg/ml of MOR03207 and MOR202 were used. 
     The data is presented in the following three ways, as a) raw data (% dead cells), b) normalized specific killing data, where the MOR202 treatment group is set as 1 (100%), and c) normalized specific killing data, where the theoretical combination is set as 1 (100%). Table 3a represents raw data. 
     
       
         
           
               
               
               
               
               
               
               
               
               
               
             
               
                 TABLE 3a 
               
               
                   
               
               
                   
                   
                   
                   
                   
                   
                   
                   
                 LEN 
                   
               
               
                   
                   
                   
                 Combination of 
                   
                   
                   
                   
                 (10 μM) 
                 DMSO 
               
               
                   
                   
                 MOR202 
                 LEN (10 μM) 
                   
                   
                 MOR03207 
                   
                 alone 
                 control 
               
               
                   
                 LEN 10 μM 
                 alone 
                 and MOR202 
                   
                 MOR03207 
                 (15 μg/ml) + 
                 LEN 
                 without 
                 without 
               
               
                 AMO-1 
                 alone 
                 (15 μg/ml) 
                 (15 μg/ml) 
                 DMSO 
                 (15 μg/ml) 
                 DMSO 
                 (0 μM) 
                 PBMCs 
                 PBMCs 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
               
               
               
               
               
               
               
               
            
               
                 Exp. 1 
                 12.89 
                 23.69 
                 35.98 
                 13.10 
                 14.15 
                 15.12 
                 15.45 
                 15.41 
                 11.07 
               
               
                 Exp. 2 
                 10.13 
                 22.53 
                 29.09 
                 7.94 
                 10.52 
                 6.99 
                 13.22 
                 8.44 
                 8.45 
               
               
                 Exp. 3 
                 22.80 
                 49.56 
                 80.39 
                 19.93 
                 24.04 
                 22.24 
                 22.63 
                 22.38 
                 26.43 
               
               
                   
               
            
           
         
       
     
     The units of the values listed are % dead cells. The DMSO, MOR03207, MOR03207+DMSO, LEN0, LEN10 without PBMCs and DMSO without PBMCs groups are controls. 
     Table 3b represents the data of Table 3a, but normalized, where the MOR202 treatment group is set as 1 (100%). 
     
       
         
           
               
               
               
               
               
               
             
               
                 TABLE 3b 
               
               
                   
               
               
                   
                   
                   
                   
                   
                 MOR202 
               
               
                   
                   
                   
                 LEN 
                   
                 (15 μg/ml) 
               
               
                   
                 MOR03207 
                 MOR202 
                 alone 
                 Theoretical 
                 and LEN 
               
               
                 AMO-1 
                 (15 μg/ml) 
                 (15 μg/ml) 
                 (10 μM) 
                 combination 
                 (10 μM) 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
               
               
               
               
            
               
                 Exp. 1 
                 −0.1 
                 1.0 
                 0.0 
                 1.0 
                 2.2 
               
               
                 Exp. 2 
                 −0.2 
                 1.0 
                 0.2 
                 1.2 
                 1.8 
               
               
                 Exp. 3 
                 0.1 
                 1.0 
                 0.1 
                 1.1 
                 2.3 
               
               
                   
               
            
           
         
       
     
     For Tables 3b-c, “Theoretical Combination” represents the addition of the values of MOR202 alone and the values of LEN alone. The normalized data of Table 3b is calculated as follows. Table 3a represents the number of dead cells. Therefore, the specific killing values of Table 3b are calculated by subtracting the values of the controls. Then the specific killing values are compared to the MOR202 group, which is set as 1. The averages of the results in Table 3b are shown in  FIG. 3 . 
     1. Determination of Synergism 
     1.1 Chou et al. 
     The methods of Chou-Talalay were used to determine synergism. See Chou T C, Talalay P, Quantitative analysis of dose-effect relationships: the combined effects of multiple drugs or enzyme inhibitors. Adv Enzyme Regul 22: 27-55 (1984), which is incorporated by reference in its entirety. Synergism analysis is carried out using the CI-isobol method. 
     Median-effect Equation 
     The median-effect equation models of the effect of an inhibitor (such as a drug) as
 
 F   a   /F   u =( D/D 50)^ m  
 
where D is the dose, F a  and F u  is the fraction of the system affected and unaffected by the dose D (F a +F u =1); D50 is the dose producing the median effect (e.g. IC50, ED50, LD50). The constant m determines the shape of the dose-effect curve.
 
We used Excel Fit software to carry out a linear regression calculation to estimate the parameters m and D50.
 
     The effects of the combination on AMO-1 cells is measured % cell death as described above. We define the fraction F u  to be the ratio of % cell death of the treated cell line to the % cell death of the cell line exposed to a control. That is:
 
 F   u =% cell death(treated cell line)/% cell death (non-treated cell line)
 
     Then the % cell death of a cell line is the constant D50 in the median effect equation, which can be estimated by the linear regression described above. 
     CI-isobol Method 
     The CI-isobol method provides a quantitative assessment of synergism between drugs. A combination index (CI) is estimated from dose-effect data of single and combined drug treatments. A value of CI less than 1 indicates synergism; CI=1 indicates additive effect; and CI&gt;1 indicates antagonism. Synergistic ranges are further defined by Chou and Talahay for CI values &lt;0.1 as very strong synergism, CI values between 0.1 and 0.3 as strong synergism, CI values of 0.3-0.7 as synergism, CI values of 0.7-0.9 as moderate to slight synergism. Drug interaction (synergism or antagonism) is more pronounced the farther a CI value is from 1. 
     Formally, the combination index (CI) of a combined drug treatment is defined as
 
CI= D   1   /D   x1   +D   2   /D   x2  
 
     Here D1 and D2 are the doses of drug 1 and drug 2, respectively, in the combination; Dx1, and Dx2 each is the dose of a treatment with only drug 1 and drug 2 that would give the same effect as that of the combination, respectively. The doses Dx1 and Dx2 need to be estimated from the dose-effect data of single drug treatments. Essentially, a median effect equation is fitted to the data of each drug. From the median effect equation of a drug, we can estimate the dose (i.e. D) necessary to produce an effect (i.e. Fa, Fu). The further a point lies from the additive line, the bigger the different between 1 and its CI, thus the stronger the (synergistic or antagonistic) effect is. 
     The above method is described in Chou T C, Talalay P, Quantitative analysis of dose-effect relationships: the combined effects of multiple drugs or enzyme inhibitors. Adv Enzyme Regul 22: 27-55 (1984), which is incorporated by reference in its entirety. An additional review of the above Chou method is also provided in Ting-Chao Chou, Theoretical Basis, Experimental Design, and Computerized Simulation of Synergism and Antagonism in Drug Combination Studies, Pharmacol Rev 58:621-681 (2006), which is incorporated by reference in its entirety. 
     The curves generated for the Chou based synergy calculations are shown in  FIGS. 12-18 . In  FIG. 12 , the best fit curve was determined by removing the data points a) where the concentration of MOR202 was too low to have any effect and b) where the concentration was near saturation. At the appropriate date point, approx. 80% cell killing, the CI value is less than 1, supporting clear synergy.  FIGS. 13-18  represent the six experiments from Tables 3 and 4, and in each the Dx1 (dose of MOR202) needed to reach 100% effect of the combination of MOR 202 and lenalidomide goes to infinity; therefore, the D 1 /D x1  is less than 1 and as lenalidomide has no effect on AMO-1 cells regarding cell killing, the Dx2 value also approaches infinity, so the D 2 /D x2  approximates 0, therefore the CI values of each of the six experiments is less than 1, supporting clear synergy. 
     Table 3c represents the normalization of data, where the theoretical combination is set as 1 (100%) and includes the CI Chou calculations. 
     
       
         
           
               
               
               
               
               
               
               
               
             
               
                 TABLE 3c 
               
               
                   
               
               
                   
                   
                   
                   
                   
                 MOR202 
                   
                   
               
               
                   
                   
                   
                   
                   
                 (0.42 μg/ml) 
                   
                   
               
               
                 Cell 
                   
                 MOR202 
                 LEN 
                 Theoretical 
                 and LEN 
                 Combination 
                   
               
               
                 line 
                 Experiment 
                 (0.42 μg/ml) 
                 (5 μM) 
                 Combination 
                 (5 μM) 
                 Index (CI) 
                 Conclusion 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
               
               
               
               
               
               
            
               
                 AMO-1 
                 Experiment 1 
                 0.6 
                 0.4 
                 1.0 
                 1.0 
                 &lt;&lt;0.1 * 
                 synergy 
               
               
                   
                 Experiment 2 
                 0.9 
                 0.1 
                 1.0 
                 2.4 
                 &lt;&lt;0.1 * 
                 synergy 
               
               
                   
                 Experiment 3 
                 0.9 
                 0.1 
                 1.0 
                 1.8 
                 &lt;&lt;0.1 * 
                 synergy 
               
               
                   
                 AVERAGE 
                 0.8 
                 0.2 
                 1.0 
                 1.7 
                 — 
                 — 
               
               
                   
               
            
           
         
       
     
     The date shown in Table 3c differs from Tables 3a and 3b. Table 3c is based upon different raw data points than shown in Table 3a, as the concentrations chosen in Table 3c are closer to the EC 50  of the antibody (raw data not shown). “Theoretical Combination” represents the addition of the values of MOR202 alone and the values of LEN alone. 
     Table 4 Effector cells only treated with Lenalidomide prior to treatment with MOR202. Single and combination doses of 10 μM LEN and 15 μg/ml of MOR03207 and MOR202 were used. 
     The data is presented in the following three ways, as a) raw data (% dead cells), b) normalized specific killing data, where the MOR202 treatment group is set as 1 (100%), and c) normalized specific killing data, where the theoretical combination is set as 1 (100%). Table 4a represents raw data. 
     
       
         
           
               
               
               
               
               
               
               
               
             
               
                 TABLE 4a 
               
               
                   
               
               
                   
                   
                   
                 Combination of 
                   
                   
                   
                   
               
               
                   
                   
                 MOR202 
                 LEN (10 μM) 
                   
                   
                 MOR03207 
                   
               
               
                   
                 LEN 10 μM 
                 alone 
                 and MOR202 
                   
                 MOR03207 
                 15 μg/ml + 
                 LEN 
               
               
                 AMO-1 
                 alone 
                 (15 μg/ml) 
                 (15 μg/ml) 
                 DMSO 
                 15 μg/ml 
                 DMSO 
                 (0 μM) 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
               
               
               
               
               
               
            
               
                 Exp. 1 
                 15.33 
                 23.09 
                 23.46 
                 14.62 
                 16.17 
                 15.97 
                 12.87 
               
               
                 Exp. 2 
                 12.98 
                 21.08 
                 25.75 
                 10.24 
                 12.17 
                 11.45 
                 9.78 
               
               
                 Exp. 3 
                 17.93 
                 48.28 
                 56.49 
                 16.75 
                 17.42 
                 15.77 
                 18.16 
               
               
                   
               
            
           
         
       
     
     The units of the values listed are % dead cells. The DMSO, MOR03207, MOR03207+DMSO, LEN0, LEN10 without PBMCs and DMSO without PBMCs are controls. 
     
       
         
           
               
               
               
               
               
               
             
               
                 TABLE 4b 
               
               
                   
               
               
                   
                   
                   
                   
                   
                 MOR202 
               
               
                   
                   
                   
                 LEN 
                   
                 (15 μg/ml) 
               
               
                   
                 MOR03207 
                 MOR202 
                 alone 
                 Theoretical 
                 and LEN 
               
               
                 AMO-1 
                 (15 μg/ml) 
                 (15 μg/ml) 
                 (10 μM) 
                 combination 
                 (10 μM) 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
               
               
               
               
            
               
                 Exp. 1 
                 0.5 
                 1.0 
                 0.1 
                 1.1 
                 1.1 
               
               
                 Exp. 2 
                 0.3 
                 1.0 
                 0.3 
                 1.3 
                 1.6 
               
               
                 Exp. 3 
                 0.0 
                 1.0 
                 0.0 
                 1.0 
                 1.3 
               
               
                   
               
            
           
         
       
     
     Table 4b represents the data of Table 4a, but normalized, where the MOR202 treatment group is set as 1 (100%). For Tables 4b-c, “Theoretical combination” represents the values of MOR202 alone plus the values of LEN alone. 
     The normalization of the data as shown in Table 4b is calculated as described in Table 3b, by substracting the controls. The averages of the results of Table 4b are shown in  FIG. 4 . 
     
       
         
           
               
               
               
               
               
               
               
               
             
               
                 TABLE 4c 
               
               
                   
               
               
                   
                   
                   
                   
                   
                 MOR202 
                   
                   
               
               
                   
                   
                   
                   
                   
                 (0.42 μg/ml) 
                   
                   
               
               
                 Cell 
                   
                 MOR202 
                 LEN 
                 Theoretical 
                 and LEN 
                 Combination 
                   
               
               
                 line 
                 Experiment 
                 (0.42 μg/ml) 
                 (5 μM) 
                 Combination 
                 (5 μM) 
                 Index (CI) 
                 Conclusion 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
               
               
               
               
               
               
            
               
                 AMO-1 
                 Experiment 1 
                 1.2 
                 −0.2 
                 1.0 
                 1.7 
                 &lt;&lt;0.1 * 
                 synergy 
               
               
                   
                 Experiment 2 
                 0.7 
                 0.3 
                 1.0 
                 1.4 
                 &lt;&lt;0.1 * 
                 synergy 
               
               
                   
                 Experiment 3 
                 0.8 
                 0.2 
                 1.0 
                 1.3 
                 &lt;&lt;0.1 * 
                 synergy 
               
               
                   
                 AVERAGE 
                 0.9 
                 0.1 
                 1.0 
                 1.5 
                 — 
                 — 
               
               
                   
               
            
           
         
       
     
     Table 4c represents the normalization of the data, where the theoretical combination is set as 1 (100%) and includes the CI Chou et al. calculations using the methodology described above within Example 4. 
     Table 4c differs from Tables 4a and 4b. Table 4c is based upon different raw data points than shown in Table 4a, as the concentrations chosen in Table 4c are closer to the EC 50  of the antibody (raw data not shown). 
     1. Determination of Synergism 
     1.2 Clarke et al. Synergism 
     Where one drug has low activity, as in here where Lenalidomide alone has low cytotoxity against AMO-1 cells, synergy can also be determined by statistical evidence that the combination is significantly different from the inhibitory drug alone. See Clarke et al., Issues in experimental design and endpoint analysis in the study of experimental cytotoxic agents in vivo in breast cancer and other models, Breast Cancer Research and Treatment 46:255-278 (1997), which is incorporated by reference in its entirety. Here both Chou et al. as shown above and the methods of Clarke et al. were used in the determination of synergism. 
     The data is analysed in the following way:
 
Antagonistic ( AB )/ C &lt;( A/C )×( B/C )
 
Additive ( AB )/ C =( A/C )×( B/C )
 
Synergistic ( AB )/ C &gt;( A/C )×( B/C )
 
where A is the treatment with LEN alone; B is the treatment with MOR202 alone; C is response to the treatment vehicle; AB is combination of treatments A and B.
 
     
       
         
           
               
             
               
                 TABLE 5 
               
             
            
               
                   
               
               
                 The raw data values shown in this table are the same as those 
               
               
                 shown in Table 3a, as they come from the same three experiments, 
               
               
                 where both effector and AMO-1 cells were treated with Lenalidomide 
               
               
                 prior to treatment with MOR202 and the single and combination 
               
               
                 doses of 10 μM LEN and 15 μg/ml of MOR03207 and MOR202 
               
               
                 were used. The only difference is that the data is analyzed 
               
               
                 using Clarke et al. instead of Chou et al. 
               
            
           
           
               
               
               
               
            
               
                   
                 Experiment 1 
                 Experiment 2 
                 Experiment 3 
               
               
                   
               
            
           
           
               
               
               
               
            
               
                 A: LEN alone 
                 15.41 
                 8.44 
                 22.38 
               
               
                 B: MOR202 alone 
                 23.69 
                 22.53 
                 49.56 
               
               
                 C: control 
                 11.07 
                 8.45 
                 26.43 
               
               
                 AB: combination of 
                 35.98 
                 29.09 
                 80.39 
               
               
                 LEN and MOR202 
                   
                   
                   
               
               
                 (AB)/C 
                 3.25 
                 3.44 
                 3.04 
               
               
                 (A/C) × (B/C) 
                 2.98 
                 2.66 
                 1.59 
               
               
                   
               
               
                 A = response to treatment with LEN alone 
               
               
                 B = response to treatment with MOR202 alone 
               
               
                 C = response to treatment with control 
               
               
                 AB = combination of treatments A and B 
               
               
                 The values of A, B, C and AB represent % cell killing. 
               
            
           
         
       
     
     In each experiment (AB)/C is greater than (A/C)×(B/C), showing clear synergy. 
     
       
         
           
               
             
               
                 TABLE 6 
               
             
            
               
                   
               
               
                 The raw data values shown in this table are the same as those 
               
               
                 shown in Table 4a, as they come from the same three experiments, 
               
               
                 where only the effector cells were treated with Lenalidomide 
               
               
                 prior to treatment with MOR202 and the single and combination 
               
               
                 doses of 10 μM LEN and 15 μg/ml of MOR03207 and MOR202 
               
               
                 were used. The only difference is that the data is analyzed 
               
               
                 using Clarke et al. instead of Chou et al. 
               
            
           
           
               
               
               
               
            
               
                   
                 Experiment 1 
                 Experiment 2 
                 Experiment 3 
               
               
                   
               
            
           
           
               
               
               
               
            
               
                 A: LEN alone 
                 15.33 
                 12.98 
                 17.93 
               
               
                 B: MOR202 alone 
                 23.09 
                 21.08 
                 48.28 
               
               
                 C: Control 
                 15.97 
                 11.45 
                 15.77 
               
               
                 AB: combination of 
                 23.46 
                 25.75 
                 56.49 
               
               
                 LEN and MOR202 
                   
                   
                   
               
               
                 (AB)/C 
                 1.47 
                 2.25 
                 3.58 
               
               
                 (A/C) × (B/C) 
                 1.39 
                 2.09 
                 3.48 
               
               
                   
               
               
                 A = response to treatment with LEN alone 
               
               
                 B = response to treatment with MOR202 alone 
               
               
                 C = response to treatment with control 
               
               
                 AB = combination of treatments A and B 
               
            
           
         
       
     
     In each experiment (AB)/C is greater than (A/C)×(B/C), showing clear synergy. 
     Results 
     Applying the analysis of Clarke et al., LEN synergistically enhanced MOR202 ADCC activity in AMO-1 cells in all 6 experiments. Applying the analysis of Chou et al., LEN synergistically enhanced MOR202 ADCC activity in AMO-1 cells in 6 out of 6 experiments. This enhancement of activity was identified to be by several mechanisms including direct cytotoxicity, activation of effector cells and upregulation of CD38 expression levels on MM cells. 
     Experiments according to example 4 are also performed with other antibodies specific for CD38, for example, the “Ref mAB5” antibody. 
     Example 5 
     Inhibition of Proliferation of NCI-H929 Cells Using Lenalidomide Alone 
     The cytotoxicity of Lenalidomide was tested in NCI-H929 using the methods described in Example 3. The results are shown in  FIG. 2 . In summary, challenge with Lenalidomide alone significantly inhibited cell proliferation in NCI-H929 cells. 
     Example 6 
     Synergistic Combination of MOR202 and Lenalidomide in NCI-H929 Cells 
     NCI-H929 cells were selected for testing with the combination of MOR202 and lenalidomide. NCI-H929 cells express higher levels of CD38 than AMO-1 cells, therefore, are representative of certain cells types found in human patients with multiple myeloma or non-Hodgkin&#39;s lymphoma. 
     In total six experiments were performed, using the methods described in Example 4, in order to determine the mediation of ADCC on NCI-H929 cells by the combination of MOR202 and lenalidomide. In three experiments, the PBMCs and NCI-H929 cells were treated with lenalidomide prior to treatment with MOR202, the results are shown in Tables 7a-b and  FIG. 6 . In three additional experiments only the PBMCs were treated with lenalidomide prior to treatment with MOR202, the results are shown in Tables 8a-b and  FIG. 7 . 
     Table 7 Both Effector and NCI-H929 cells were treated with Lenalidomide prior to treatment with MOR202. Single and combination doses of 5 μM LEN and 15 μg/ml of MOR03207 and 0.2 or 0.07 μg/ml MOR202 were used. 
     The data is presented in the following ways, as a) raw data (% dead cells), and b) normalized specific killing data, where the fractional product combination is set as 1 (100%). Table 7a represents raw data. 
     
       
         
           
               
               
               
               
               
               
               
               
             
               
                 TABLE 7a 
               
               
                   
               
               
                   
                   
                   
                 Combination of 
                   
                   
                   
                   
               
               
                   
                   
                 MOR202 
                 LEN (5 μM) and 
                   
                   
                 MOR03207 
                   
               
               
                   
                 LEN 5 μM 
                 alone (0.2* or 
                 MOR202 (0.2* or 
                   
                 LEN 
                 (15 μg/ml) + 
                 MOR03207 
               
               
                 NCI-H929 
                 alone 
                 0.07 μg/ml) 
                 0.07 μg/ml) 
                 DMSO 
                 (0 μM) 
                 DMSO 
                 (15 μg/ml) 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
               
               
               
               
               
               
            
               
                 Exp. 1 
                 38.65 
                 30.64* 
                 60.20* 
                 18.01 
                 18.42 
                 18.27 
                 17.81 
               
               
                 Exp. 2 
                 41.92 
                 43.08 
                 66.62 
                 18.77 
                 19.92 
                 20.26 
                 19.20 
               
               
                 Exp. 3 
                 39.92 
                 32.54 
                 64.58 
                 12.32 
                 12.44 
                 13.74 
                 14.09 
               
               
                   
               
            
           
         
       
     
     The units of the values listed are % dead cells. The DMSO, MOR03207, MOR03207+DMSO, LEN0, LEN10 without PBMCs and DMSO without PBMCs are controls. 
     Table 7b represents normalized data, where the fractional product combination is set as 1 (100%). 
     
       
         
           
               
               
               
               
               
               
               
             
               
                 TABLE 7b 
               
               
                   
               
               
                   
                   
                   
                 Combination 
                   
                   
                   
               
               
                   
                   
                   
                 based upon 
                 Combination of 
                   
                   
               
               
                   
                 MOR202 
                   
                 fractional 
                 LEN (5 μm) and 
                   
                   
               
               
                   
                 alone (0.2* or 
                 LEN 5 μM 
                 product 
                 MOR202 (0.2* 
                 Combination 
                   
               
               
                 NCI-H929 
                 0.07 μg/ml) 
                 alone 
                 concept 
                 or 0.07 μg/ml) 
                 Index (CI) 
                 Conclusion 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
               
               
               
               
               
            
               
                 Exp. 1 
                 0.42* 
                 0.67 
                 1.00 
                 1.36* 
                 &lt;&lt;0.1 
                 synergism 
               
               
                 Exp. 2 
                 0.58 
                 0.56 
                 1.00 
                 1.12 
                 &lt;&lt;0.1 
                 synergism 
               
               
                 Exp. 3 
                 0.45 
                 0.67 
                 1.00 
                 1.24 
                 &lt;&lt;0.1 
                 synergism 
               
               
                 AVERAGE 
                 0.48 
                 0.63 
                 1.00 
                 1.24 
               
               
                   
               
            
           
         
       
     
     The fractional product combination is calculated using the following formula 1−[(1−A)*(1−B)]=fpc (%) as described in Ting-Chao Chou, Theoretical Basis, Experimental Design, and Computerized Simulation of Synergism and Antagonism in Drug Combination Studies, Pharmacol Rev 58:621-681 (2006), which is incorporated by reference in its entirety. Table 7b is based upon the raw data shown in Table 7a. The normalization of the data as shown in Table 7b is calculated as described in Table 3b, by substracting the controls. In Table 7b, where the combination of LEN and MOR202 is greater than the combination based upon the fractional product concept, then clear synergy exists. In addition, Combination Index values were calculated using the methods of Chou et al. as described in Example 4. The averages of the results of Table 7b are shown in  FIG. 6 . 
     Table 8 Effector cells only treated with Lenalidomide prior to treatment with MOR202. Single and combination doses of 5 μM LEN and 15 μg/ml of MOR03207 and 0.2* or 0.07 μg/m MOR202 were used. 
     The data is presented in the following ways, as a) raw data (% dead cells), and b) normalized specific killing data, where the fractional product combination is set as 1 (100%). Table 8a represents raw data. 
     
       
         
           
               
               
               
               
               
               
               
               
             
               
                 TABLE 8a 
               
               
                   
               
               
                   
                   
                   
                 Combination of 
                   
                   
                   
                   
               
               
                   
                   
                 MOR202 
                 LEN (5 μM) and 
                   
                   
                 MOR03207 
                   
               
               
                   
                 LEN 5 μM 
                 alone (0.2* or 
                 MOR202 (0.2* or 
                   
                 LEN 
                 (15 μg/ml) + 
                 MOR03207 
               
               
                 NCI-H929 
                 alone 
                 0.07 μg/ml) 
                 0.07 μg/ml) 
                 DMSO 
                 (0 μM) 
                 DMSO 
                 (15 μg/ml) 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
               
               
               
               
               
               
            
               
                 Exp. 1 
                 17.50 
                 26.60* 
                 29.11* 
                 18.36 
                 17.56 
                 19.52 
                 17.07 
               
               
                 Exp. 2 
                 25.72 
                 47.00 
                 51.23 
                 22.55 
                 24.90 
                 24.16 
                 23.19 
               
               
                 Exp. 3 
                 26.27 
                 53.74 
                 67.99 
                 25.29 
                 25.16 
                 24.43 
                 27.10 
               
               
                   
               
            
           
         
       
     
     The units of the values listed are % dead cells. The DMSO, MOR03207, MOR03207+DMSO, LEN0, LEN10 without PBMCs and DMSO without PBMCs are controls. Table 8b represents the normalized data, where the fractional product combination is set as 1 (100%). 
     
       
         
           
               
               
               
               
               
               
               
             
               
                 TABLE 8b 
               
               
                   
               
               
                   
                   
                   
                 Combination 
                   
                   
                   
               
               
                   
                   
                   
                 based upon 
                 Combination of 
                   
                   
               
               
                   
                 MOR202 
                   
                 fractional 
                 LEN (5 μm) and 
                   
                   
               
               
                   
                 alone (0.2* or 
                 LEN 5 μM 
                 product 
                 MOR202 (0.2* 
                 Combination 
                   
               
               
                 NCI-H929 
                 0.07 μg/ml) 
                 alone 
                 concept 
                 or 0.07 μg/ml) 
                 Index (CI) 
                 Conclusion 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
               
               
               
               
               
            
               
                 Exp. 1 
                 1.09* 
                 −0.10 
                 1.00 
                 1.10* 
                 0.07 
                 synergism 
               
               
                 Exp. 2 
                 0.91 
                 0.12 
                 1.00 
                 1.03 
                 0.81 
                 synergism 
               
               
                 Exp. 3 
                 0.97 
                 0.04 
                 1.00 
                 1.59 
                 &lt;&lt;0.1 
                 synergism 
               
               
                 AVERAGE 
                 0.99 
                 0.02 
                 1.00 
                 1.24 
               
               
                   
               
            
           
         
       
     
     Table 8b is based upon the raw data shown in Table 8a. The normalization of the data as shown in Table 8a is calculated as described in Table 3b, by substracting the controls. In Table 8b, where the combination of LEN and MOR202 is greater than the combination based upon the fractional product concept, then clear synergy exists. In addition, Combination Index values were calculated using the methods of Chou et al. as described in Example 4. The averages of the results of Table 8b are shown in  FIG. 7 . 
     Determination of Synergism 
     1.3 Fractional Product Concept 
     The evaluation of the data in this example differs from that used in the analysis of the effect of the combination of MOR202 and LEN on AMO-1 cells in Example 4. Here NCI-H929 cells are tested and LEN alone has a significant effect on the proliferation of NCI-H929 cells as shown in Example 5, therefore, the fractional product concept is utilized. The fractional product concept was described in Ting-Chao Chou, Theoretical Basis, Experimental Design, and Computerized Simulation of Synergism and Antagonism in Drug Combination Studies, Pharmacol Rev 58:621-681 (2006), which is incorporated by reference in its entirety. There Chou et al. states: If A and B each inhibits 60%, then it is oversimplification to say that the additive effect is 84% inhibition. Based on the reasoning by Webb (1963), this type of problem can be solved by (1−0.6)(1−0.6)=0.16, 1−0.16=0.84. Chou and Talalay (1984) called it the fractional product method. This method will never lead to a combination effect exceeding 100% inhibition. Chou and Talalay (1984), however, have also proved that this method has limited validity because it takes into account the potency (e.g., fractional inhibition) but ignores the shape of the dose-effect curve (e.g., hyperbolic or sigmoidal). The importance of the “shape” in a dose-effect analysis is shown in  FIG. 1 . Chou and Talalay (1984) indicated that Webb&#39;s method is valid only when both drugs have hyperbolic curves (i.e., in simple Michaelis-Menten kinetics when dose-effect curves are hyperbolic, i.e., m=1 in the median-effect plot) and is not valid when m does not equal 1, such as sigmoidal (m&gt;1) or flat sigmoidal (m&lt;1) curves. Furthermore, Webb&#39;s method is valid when the effects of two drugs are mutually nonexclusive (e.g., totally independent) and is not valid for mutually exclusive (e.g., similar mechanisms or modes of actions, as assumed for the classic isobologram, see below). 
     Clarke et al. was not utilized as Clarke is most suitable when one monotherapy has a low effect. 
     See  FIG. 12 , the best fit curve was determined by removing the data points a) where the concentration of MOR202 was too low to have any effect and b) where the concentration was near saturation. At the appropriate date point, approx. 80% cell killing, the CI value is less than 1, supporting clear synergy. 
     Results 
     Applying the analysis of the Fractional Product Concept, LEN synergistically enhanced MOR202 activity in NCI-H929 cells in 6 out of 6 experiments. Applying the analysis of Chou et al., LEN synergistically enhanced MOR202 activity in NCI-H929 cells in 6 out of 6 experiments. See Tables 7a-b, and 8a-b. 
     Example 7 
     MOR202 and LEN Alone and in Combination in NCI-H929 Bone Lysis SCID Mouse MM Model 
     Materials 
     Lenalidomide (SYNthesis med chem; Shanghai, China; Lot no: ZHM-066-051). MOR202 (MorphoSys AG, Lot 100706-5KLE18). Vehicle control: Ora-Plus: Ora-Sweet SF (Paddock Laboratories, Minneapolis, Minn., USA, Lot no. 9499528). SCID Mice (University of Adelaide, Waite Campus, Urrbaraie, SA, Australia, Strain C.B.-17-Igh-1 b -Prkdc scid ). NCI-H929 human multiple myeloma cells (see Table 1). RPMI 1640 cell culture medium, Foetal Bovine Serum (FBS), Mercaptoethanol, Hank&#39;s Balanced Salt Solution (HBSS) and penicillin-streptomycin from Invitrogen Australia (Mt Waverley, VIC, Australia); and Trypan Blue and glucose from Sigma-Aldrich (Castle Hill, NSW, Australia). 
     Methods 
     63 SCID mice were inoculated on Day (−7) orthotopically into the right tibia with 2.5×10 6  NCI-H929 MM cells (in 5 μL) in order to induce bone lysis. Three days post inoculation (Day −4) 60 of the SCID mice were randomized by body weight into the groups shown in Table 13, 10 mice per group. The dosing regimen is provided in Table 9. Lenalidomide (Groups A and D) and Vehicle Control (Group C) treatments started on Day (−1). MOR202 treatments (Groups B and D) started on Day 0. Treatment continued for 6 weeks. 
                     TABLE 9                  Dosing regimen and Groups                             Group   Compound   Treatment   Schedule               A   Lenalidomide   50 mg/kg, p.o. in 10 mL/kg   once daily for 6 weeks       B   MOR202   3 mg/kg, i.p., in 10 mL/kg   3 times weekly for 6 weeks       C   Vehicle Control   10 mL/kg, p.o.   once daily for 6 weeks           (Ora-Plus:Ora-Sweet                   SF (1:1, w/w))               D   Lenalidomide/   50 mg/kg, p.o. in 10 mL/kg   once daily for 6 weeks           MOR202   3 mg/kg, i.p., in 10 mL/kg   3 times weekly for 6 weeks           Combination                    
MicroCT Scan was used to assess bone lysis and included a 3-dimensional analysis comprising Total Bone Volume (TBV), Trabecular Bone Volume (Tb.BV), Trabecular Pattern Factor (Tb.Pf) and Structure Model Index (SMI). Table 10 defines each of these parameters. The results of each of the MicroCT Scan parameters are shown in Table 11. The Total Bone Volume (TBV) results are shown in  FIG. 19 .
 
     
       
         
           
               
             
               
                 TABLE 10 
               
             
            
               
                   
               
               
                 MicroCT Scan parameters 
               
            
           
           
               
               
            
               
                 Parameters: 
                 Definitions: 
               
               
                   
               
               
                 Total Bone Volume (mm3) 
                 Total cortical and trabecular bone volume within the 
               
               
                   
                 volume of interest (cross-section). 
               
               
                 Trabecular Bone Volume 
                 Trabecular bone volume within the volume of interest 
               
               
                   
                 (cross-section). 
               
               
                 {circumflex over ( )}Trabecular Pattern Factor (Tb.Pf) 
                 Fragmentation index; An inverse index of connectivity 
               
               
                   
                 with specific application to the trabecular bone. A 
               
               
                   
                 lower Tb.Pf signifies better connected trabecular 
               
               
                   
                 lattices while higher Tb.Pf means a more disconnected 
               
               
                   
                 trabecular structure (I.e. more bone lysis). 
               
               
                 ** Structure Model Index (SMI) 
                 An indicator of the relative prevalence of rods and 
               
               
                   
                 plates in a 3D structure such as the trabecular bone. 
               
               
                   
                 This parameter is important in osteolysis of the bone 
               
               
                   
                 which is characterised by a transition from plate-like 
               
               
                   
                 (normal) to rod-like (degradation) structures. An ideal 
               
               
                   
                 plate, cylinder and sphere have SMI values of 0, 3 and 
               
               
                   
                 4 respectively. The higher the value, the more damage 
               
               
                   
                 there is. 
               
               
                   
               
            
           
         
       
     
     
       
         
           
               
             
               
                 TABLE 11 
               
               
                   
               
               
                 Results of the MicroCT Scan: Total Bone Volume (TBV), Trabecular Bone Volume 
               
               
                 (Tb.BV), Trabecular Pattern Factor (Tb.Pf) and Structure Model Index (SMI). 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
               
               
               
               
               
            
               
                   
                   
                   
                 Total 
                 Trabecular 
                 Trabecular 
                   
               
               
                   
                   
                   
                 Bone 
                 Bone 
                 Pattern 
                 Structure 
               
               
                   
                   
                   
                 Volume 
                 Volume 
                 Factor 
                 Model 
               
               
                   
                   
                   
                 (TBV) 
                 (Tb.BV) 
                 (Tb.Pf) 
                 Index 
               
               
                 Group 
                 Treatment 
                 Mouse ID 
                 mm −3   
                 mm −2   
                 mm −−1   
                 (SMI) 
               
               
                   
               
               
                 Control 
                 Non-inoculated 
                 38045 
                 2.748 
                 0.244 
                 15.354 
                 1.756 
               
               
                   
                 reference 
                 38596 
                 2.839 
                 0.295 
                 12.373 
                 1.542 
               
               
                   
                 tibia* 
                 39565 
                 2.930 
                 0.314 
                 14.703 
                 1.847 
               
               
                   
                   
                 38325 
                 2.964 
                 0.309 
                 13.538 
                 1.653 
               
               
                   
                   
                 33746 
                 2.751 
                 0.293 
                 13.270 
                 1.624 
               
               
                   
                   
                 38770 
                 2.567 
                 0.307 
                 13.025 
                 1.645 
               
               
                   
                   
                 37966 
                 2.967 
                 0.410 
                 12.125 
                 1.557 
               
               
                   
                   
                 38604 
                 3.087 
                 0.327 
                 11.902 
                 1.658 
               
               
                   
                   
                 38023 
                 2.775 
                 0.270 
                 18.005 
                 1.889 
               
               
                   
                   
                 38594 
                 2.830 
                 0.311 
                 13.293 
                 1.589 
               
               
                   
                   
                 Mean 
                 2.846 
                 0.308 
                 13.759 
                 1.676 
               
               
                   
                   
                 SEM 
                 0.047 
                 0.014 
                 0.583 
                 0.037 
               
               
                 A 
                 Lenalidomide, 
                 33150 
                 1.604 
                 0.107 
                 24.329 
                 2.679 
               
               
                   
                 50 mg/kg 
                 38027 
                 1.742 
                 0.100 
                 23.561 
                 2.667 
               
               
                   
                   
                 38314 
                 2.506 
                 0.256 
                 22.893 
                 2.335 
               
               
                   
                   
                 38446 
                 2.466 
                 0.213 
                 28.280 
                 2.560 
               
               
                   
                   
                 38562 
                 2.688 
                 0.385 
                 22.213 
                 2.086 
               
               
                   
                   
                 38626 
                 2.869 
                 0.293 
                 30.739 
                 2.619 
               
               
                   
                   
                 38748 
                 1.786 
                 0.114 
                 24.016 
                 2.562 
               
               
                   
                   
                 39192 
                 1.988 
                 0.081 
                 29.454 
                 2.592 
               
               
                   
                   
                 39364 
                 1.741 
                 0.155 
                 24.205 
                 2.547 
               
               
                   
                   
                 39512 
                 2.007 
                 0.219 
                 38.360 
                 3.125 
               
               
                   
                   
                 Mean 
                 2.140 
                 0.192 
                 26.805 
                 2.577 
               
               
                   
                   
                 SEM 
                 0.143 
                 0.031 
                 1.584 
                 0.083 
               
               
                 B 
                 MOR03087, 
                 32094 
                 2.233 
                 0.190 
                 27.049 
                 2.386 
               
               
                   
                 3 mg/kg 
                 32548 
                 2.893 
                 0.310 
                 15.631 
                 1.818 
               
               
                   
                   
                 33564 
                 2.760 
                 0.356 
                 27.631 
                 2.423 
               
               
                   
                   
                 38016 
                 1.635 
                 0.118 
                 27.523 
                 2.450 
               
               
                   
                   
                 38023 
                 2.681 
                 0.248 
                 17.887 
                 1.860 
               
               
                   
                   
                 38510 
                 1.838 
                 0.260 
                 21.405 
                 2.461 
               
               
                   
                   
                 38599 
                 2.884 
                 0.482 
                 26.345 
                 2.558 
               
               
                   
                   
                 39086 
                 3.068 
                 0.566 
                 20.327 
                 2.247 
               
               
                   
                   
                 39666 
                 2.547 
                 0.416 
                 25.843 
                 2.318 
               
               
                   
                   
                 39715 
                 2.135 
                 0.284 
                 22.402 
                 2.275 
               
               
                   
                   
                 Mean 
                 2.467 
                 0.323 
                 23.204 
                 2.280 
               
               
                   
                   
                 SEM 
                 0.153 
                 0.043 
                 1.365 
                 0.079 
               
               
                 C 
                 Vehicle 
                 33090 
                 1.821 
                 0.159 
                 26.537 
                 2.714 
               
               
                   
                 Control 
                 33131 
                 1.863 
                 0.132 
                 28.429 
                 2.681 
               
               
                   
                 (Ora 
                 33746 
                 1.577 
                 0.130 
                 29.171 
                 2.652 
               
               
                   
                 Plus:Ora 
                 37966 
                 1.865 
                 0.234 
                 18.276 
                 2.327 
               
               
                   
                 Sweet SF 
                 38325 
                 2.030 
                 0.096 
                 30.839 
                 2.591 
               
               
                   
                 (1:1, w/w)) 
                 38596 
                 1.870 
                 0.154 
                 33.079 
                 2.783 
               
               
                   
                   
                 38604 
                 1.904 
                 0.232 
                 23.234 
                 2.607 
               
               
                   
                   
                 38770 
                 2.461 
                 0.210 
                 19.149 
                 2.137 
               
               
                   
                   
                 39426 
                 1.556 
                 0.184 
                 21.846 
                 2.348 
               
               
                   
                   
                 39565 
                 2.235 
                 0.256 
                 24.545 
                 2.365 
               
               
                   
                   
                 Mean 
                 1.918 
                 0.179 
                 25.510 
                 2.521 
               
               
                   
                   
                 SEM 
                 0.087 
                 0.017 
                 1.567 
                 0.067 
               
               
                 AB 
                 Lenalidomide, 
                 32695 
                 2.471 
                 0.168 
                 21.323 
                 2.028 
               
               
                   
                 50 mg/kg/ 
                 37854 
                 2.527 
                 0.265 
                 15.938 
                 1.758 
               
               
                   
                 MOR03087, 
                 38276 
                 3.212 
                 0.220 
                 20.046 
                 2.197 
               
               
                   
                 3 mg/kg 
                 38550 
                 2.833 
                 0.186 
                 17.907 
                 1.892 
               
               
                   
                   
                 38594 
                 3.044 
                 0.268 
                 16.530 
                 1.787 
               
               
                   
                   
                 38994 
                 2.896 
                 0.408 
                 14.633 
                 1.683 
               
               
                   
                   
                 39256 
                 2.308 
                 0.304 
                 24.513 
                 2.280 
               
               
                   
                   
                 39555 
                 3.227 
                 0.215 
                 19.753 
                 2.497 
               
               
                   
                   
                 39677 
                 3.205 
                 0.548 
                 12.313 
                 1.799 
               
               
                   
                   
                 39750 
                 2.961 
                 0.281 
                 14.981 
                 1.752 
               
               
                   
                   
                 Mean 
                 2.868 
                 0.286 
                 17.794 
                 1.967 
               
               
                   
                   
                 SEM 
                 0.105 
                 0.036 
                 1.152 
                 0.086 
               
               
                   
               
            
           
         
       
     
     The analysis of each parameter for synergistic activity was performed according to theorem of Clarke et al. Table 12 shows the calculations done to determine synergy of the combination of MOR202 and lenalidomide. 
     
       
         
           
               
               
             
               
                 TABLE 12 
               
               
                   
               
             
            
               
                 When POSITIVE EFFECT has a HIGH value: 
                 When POSITIVE EFFECT has a LOW value: 
               
               
                 Antagonistic = (AB)/C &lt; (A/C) × (B/C) 
                 Antagonistic = (AB)/C &gt; (A/C) × (B/C) 
               
               
                 Additive = (AB)/C = (A/C) × (B/C) 
                 Additive = (AB)/C = (A/C) × (B/C) 
               
               
                 Synergistic = (AB)/C &gt; (A/C) × (B/C) 
                 Synergistic = (AB)/C &lt; (A/C) × (B/C) 
               
               
                   
               
            
           
           
               
               
               
               
               
            
               
                   
                 Total BV 
                 Trabecular BV 
                 Trabecular pattern factor 
                 Structural Model Index 
               
               
                   
               
               
                 A 
                 2.14 
                 0.192 
                 26.805 
                 2.577 
               
               
                 B 
                 2.467 
                 0.323 
                 23.204 
                 2.28 
               
               
                 C 
                 1.918 
                 0.179 
                 25.51 
                 2.521 
               
               
                 AB 
                 2.868 
                 0.286 
                 17.794 
                 1.967 
               
               
                 (AB)/C 
                 1.495307612 
                 1.597765363 
                 0.69753038 
                 0.780245934 
               
               
                   
                 is bigger than 
                 is less than 
                 is less than 
                 is less than 
               
               
                 (A/C) × (B/C) 
                 1.44 
                 1.94 
                 0.96 
                 0.92 
               
               
                   
               
               
                 A = response to treatment LEN 50 mg/kg 
               
               
                 B = response to treatment MOR202 3 mg/kg 
               
               
                 C = response to treatment vehicle 
               
               
                 AB = combination of treatments 1 and 2 
               
            
           
         
       
     
     The numeric values shown in Table 12 are taken directly from the averages shown in Table 11 for each of the parameters in each of the Groups. The Groups described as A, B, C and AB are the same treatment groups in both Tables 9, 11 and 12. 
     In Total Bone Volume (AB)/C is greater than (A/C)×(B/C) showing clear synergism. In Trabecular pattern factor and Structural Model Index, as described in Table 10, a lower value represents less bone lysis (efficacy in treatment), therefore, (AB)/C less than (A/C)×(B/C), shows clear synergism in both parameters. 
     Results 
     The inoculation of NCI-H929 multiple myeloma cells induced significant bone lysis in the tibiae of female SCID mice in this study, as indicated by the measurement of bone lysis through microCT scanning. The degree of bone lysis was significantly decreased in the tibia of mice treated with the combination of MOR202 and lenalidomide as shown by microCT scanning. In each of the parameters of MicroCT Scan: Total Bone Volume (TBV), Trabecular Bone Volume (Tb.BV), Trabecular Pattern Factor (Tb.Pf) and Structure Model Index (SMI) the combination of MOR202 and lenalidomide (Group AB) showed clear synergy in the reduction of bone lysis caused by the NCI-H929 multiple myeloma cells. 
     When the values in Table 11 are adjusted, so that the Control Group (Non-inoculated Contralateral Tibia without Tumour) is considered 0% bone lysis, and Group C (Vehicle Control (0.9% Sodium Chloride Injection) is considered 100% bone lysis, then MOR202 alone reduced bone lysis dose-dependently by up to 55% at 12 mg/kg compared to vehicle control. LEN alone at 50 mg/kg inhibited bone lysis by 20%. The combination of 3 mg/kg MOR202 and 50 mg/kg LEN completely abolished bone lysis. These findings support a synergistic effect of combination therapy. In addition, there was a reduction (&gt;90%) of M-protein serum levels in the combination group, indicating a significant decrease of tumor load. 
     Example 8 
     MOR202 and Lenalidomide Alone and in Combination Against Human Non-hodgkin RAMOS Tumor in Female SCID Mice, Survival Model 
     Materials 
     Cyclophosphamide (Fluka, Buchs Switzerland, Lot. No. 07551661). Lenalidomide (SYNthesis Med Chem; Shanghai, China; Lot. #ZHM-066-051). MOR202 (MorphoSys AG, Lot 100706-5KLE18). Vehicle Control: Ora-Plus:Ora-Sweet SF, 1:1, v/v (SYNthesis Med Chem, Shanghai, China). SCID Mice (University of Adelaide, Waite Campus, Urrbaraie, SA, Australia, Strain C.B.-17-Igh-1b-Prkdc scid ). 
     RAMOS cells (Oncodesign, Dijon Cedex, France) were cultivated in RPMI1640+20% heat inactivated alternate source FBS+1% Glutamax (Medium #2). Reagents for culture of RAMOS non-Hodgkin lymphoma cells were obtained from the following suppliers: RPMI 1640 cell culture medium, FBS, Glutamax, HEPES, sodium pyruvate, HBSS, and penicillin-streptomycin from Invitrogen Australia (Mt Waverley, VIC, Australia); and Trypan Blue and glucose from Sigma-Aldrich (Castle Hill, NSW, Australia). 
     Methods 
     Sixty-eight female SCID mice were pre-treated with Cyclophosphamide (75 mg/kg, i.p., twice daily) for two days prior to RAMOS cell inoculation (Day −5 and −4). On the day of inoculation (Day −3), all mice were inoculated with 1×10 6  RAMOS cells each intravenously into the tail vein. Sixty-four of the mice were randomised by body weight into eight groups of eight. The dosing regimen for each group is shown in Table 13. 
                     TABLE 13                  Dosing regimen                                 Group   Compound   Treatment   Intended Schedule   Actual Schedule               A   Lenalidomide   50 mg/kg, p.o., in   Once daily   Day 0-20               10 mL/kg   (Day 0-20)           B   MOR03087   1 mg/kg, i.v., in   Twice weekly (Day   Twice weekly (Day 0,               10 mL/kg   0, 4, 7, 11, 14   4, 7, 11, 14 and 18)                   and 18)           C   Vehicle Control   p.o., 10 mL/kg   Once daily   Day 0-18           (Ora-Plus:Ora-Sweet,       (Day 0-20)               1:1, v/v)                   AB   Lenalidomide/   100/1 mg/kg,   Once daily/twice   Day 0-13 and 16-20/           MOR03087   p.o./i.v., in   weekly (as above)   Day 0, 4, 7, 11, 14               10 mL/kg       and 18                    
The study continued for 98 days and the measured endpoint was survival. The results of each Group are shown in Table 14.
 
     
       
         
           
               
             
               
                 TABLE 14 
               
             
            
               
                   
               
               
                 Survival Number and time period for each group 
               
            
           
           
               
               
               
            
               
                   
                   
                 Number of mice 
               
               
                   
                 % ILS (based 
                 alive at study 
               
            
           
           
               
               
               
               
            
               
                   
                 Day of death (post-inoculation) 
                 on median 
                 termination 
               
            
           
           
               
               
               
               
               
               
               
            
               
                   
                 Median 
                 Range 
                 Mean 
                 95% CI 
                 death day) 
                 (day 98) 
               
               
                   
               
               
                 A: LEN 100 
                 22 
                 18-23 
                 21.4 
                 19.8-23.0 
                  10 
                 0/7 
               
               
                 mg/kg 
                   
                   
                   
                   
                   
                   
               
               
                 B: MOR202 
                 51 
                 35-65 
                 49.6 
                 41.9-57.3 
                 155 
                 0/8 
               
               
                 1 mg/kg 
                   
                   
                   
                   
                   
                   
               
               
                 C: Vehicle 
                 20 
                 18-21 
                 19.8 
                 18.7-20.8 
                 X 
                 0/8 
               
               
                 control 
                   
                   
                   
                   
                   
                   
               
               
                 AB: Combo 
                 65 
                 32-98 
                 66.5 
                 41.9-91.2 
                 225 
                 3/8 
               
               
                 LEN/MOR 
               
               
                   
               
            
           
         
       
     
     Analysis for synergistic activity was performed according to theorem of Clarke et al., as described in Example 4. Table 15 shows the calculations done in the determination of synergy of the combination of MOR202 and lenalidomide. 
     
       
         
           
               
               
             
               
                 TABLE 15 
               
               
                   
               
             
            
               
                 When POSITIVE EFFECT has a HIGH value: 
                 When POSITIVE EFFECT has a LOW value: 
               
               
                 Antagonistic = (AB)/C &lt; (A/C) × (B/C) 
                 Antagonistic = (AB)/C &gt; (A/C) × (B/C) 
               
               
                 Additive = (AB)/C = (A/C) × (B/C) 
                 Additive = (AB)/C = (A/C) × (B/C) 
               
               
                 Synergistic = (AB)/C &gt; (A/C) × (B/C); 
                 Synergistic = (AB)/C &lt; (A/C) × (B/C) 
               
               
                   
               
            
           
           
               
               
               
            
               
                   
                   
                 Median survival 
               
               
                   
               
               
                   
                 A 
                 22 
               
               
                   
                 B 
                 51 
               
               
                   
                 C 
                 20 
               
               
                   
                 AB 
                 65 
               
               
                   
                 (AB)/C 
                 3.25 
               
               
                   
                   
                 is bigger than 
               
               
                   
                 (A/C) × (B/C) 
                 2.805 
               
               
                   
               
               
                 A = response to treatment with LEN 100 mg/kg 
               
               
                 B = response to treatment with MOR202 1 mg/kg 
               
               
                 C = response to treatment vehicle 
               
               
                 AB = combination of treatments A and B 
               
            
           
         
       
     
     The numeric values shown in Table 15 are taken directly from the median survival days shown in Table 14 for each of the Groups. The Groups described as A, B, C and AB are the same treatment groups in Tables 13-15. 
     The inoculation with RAMOS cells was lethal within a median time of 20 days in the control group. The combination of MOR202 and lenalidomide, however, showed clear synergy in the increase in median survival days. 
     Example 9 
     Bortezomib Alone Inhibits Proliferation of Various Multiple Myeloma Cell Lines 
     The inhibitory effect of Bortezomib on proliferation of multiple myeloma cells was analysed for multiple cell lines. Increasing amounts of Bortezomib (Velcade®, Lot: No.: #9AZSY00) were applied to AMO-1, LP-1, NCI-H929 and RPMI-8226 cells and incubated for 24 h, 48 h and 72 h. After incubation, period plates were analyzed for cell proliferation in a quantitative colorimietric XTT-based assay using the cell proliferation kit II (ROCHE, Cell Proliferation Kit II, Cat. No.: 11465015001). For subsequent measurement, plates were subjected to Tecan Genios Reader and absorbance at 492 nm was detected. 
     Cell proliferation of all tested cell lines was inhibited by Bortezomib with an IC50 concentration of 3.9 nM for AMO-1 cells, 6.1 nM for LP-1 cells, 3.3 nM for NCI-H929 cells and 9.0 nM for RPMI-8226 cells respectively, as shown in  FIG. 8 . 
     Example 10 
     ADCC Using Combination of MOR202 and Bortezomib 
     Using the methods described in Example 4, the ADCC effect of combining bortezomib and MOR202 was analyzed. Here, the target cells were treated with bortezomib prior to the treatment with MOR202. Both target cells, NCI-H929 and LP-1 cells were tested. The results are shown in  FIGS. 9 and 10 . The enhancement in MOR202 activity by bortezomib was mediated through a direct cytotoxic effect on MM cells. 
     Example 11 
     MOR202 and BOR Alone and in Combination in Human Multiple Myeloma NCI-H929 Bone Lysis SCID Mouse Model 
     Materials 
     Bortezomib (SYNthesis med chem., Shanghai, China, Lot no. #ZHM-066-054). Bortezomib was formulated in sterile 0.9% Sodium Chloride solution for dosing. MOR202 (MorphoSys AG, Lot 100706-5KLE18). Vehicle control: 0.9% Sodium Chloride Injection. SCID Mice (University of Adelaide, Waite Campus, Urrbaraie, SA, Australia, Strain C. B.-17-Igh-1 b -Prkdc scid ). 
     Methods 
     63 SCID mice were inoculated on Day (−7) intra-tibially with 2.5×10 6  NCI-H929 MM cells in order to induce bone lysis. Three days post inoculation (Day −4) 60 of the SCID mice were randomized by body weight into the groups shown in Table 16, 10 mice per group. The dosing regimen is provided in Table 16. Bortezomib (Groups A and AB) and Vehicle Control (Group C) treatments started on Day (−1). MOR202 treatments (Groups B and AB) started on Day 0. Treatment continued for 6 weeks. 
     
       
         
           
               
             
               
                 TABLE 16 
               
             
            
               
                   
               
               
                 Dosing regimen and Groups 
               
            
           
           
               
               
               
               
            
               
                 Group 
                 Compound 
                 Treatment 
                 Schedule 
               
               
                   
               
               
                 A 
                 Bortezomib 
                 0.6 mg/kg, i.p., in 10 mL/kg 
                 twice per week 
               
               
                 B 
                 MOR202 
                 3 mg/kg, i.p., in 10 mL/kg 
                 three times per week 
               
               
                 C 
                 Vehicle Control 
                 i.p., 10 mL/kg 
                 twice per week 
               
               
                   
                 (0.9% Sodium Chloride 
                   
                   
               
               
                   
                 Injection) 
                   
                   
               
               
                 AB 
                 Bortezomib/ 
                 0.6/3 mg/kg, i.p., in 10 mL/kg 
                 twice/three times per week, 
               
               
                   
                 MOR202 
                   
                 on alternate days 
               
               
                   
               
            
           
         
       
     
     MicroCT Scan was used to assess bone lysis and included a 3-dimensional analysis comprising Total Bone Volume (TBV), Trabecular Bone Volume (Tb.BV), Trabecular Pattern Factor (Tb.Pf) and Structure Model Index (SMI). Table 10 above defines each of these parameters. The results of each of the MicroCT Scan parameters are shown in Table 17. The results of the Total Bone Volume (TBV) is shown in  FIG. 20 . 
     
       
         
           
               
             
               
                 TABLE 17 
               
               
                   
               
               
                 Results of the MicroCT Scan: Total Bone Volume (TBV), Trabecular Bone Volume 
               
               
                 (Tb.BV), Trabecular Pattern Factor (Tb.Pf) and Structure Model Index (SMI). 
               
               
                   
               
             
            
               
                   
               
            
           
           
               
               
               
               
               
               
               
            
               
                   
                   
                   
                 Total 
                 Trabecular 
                 Trabecular 
                   
               
               
                   
                   
                   
                 Bone 
                 Bone 
                 Pattern 
                 Structure 
               
               
                   
                   
                   
                 Volume 
                 Volume 
                 Factor 
                 Model 
               
               
                   
                   
                   
                 (TBV) 
                 (Tb.BV) 
                 (Tb.Pf) 
                 Index 
               
               
                 Group 
                 Treatment 
                 Mouse ID 
                 mm −3   
                 mm −2   
                 mm −−1   
                 (SMI) 
               
               
                   
               
               
                 Control: 
                 Reference tibia, 
                 115898 
                 2.771 
                 0.400 
                 12.097 
                 1.525 
               
               
                 Non-inoculated 
                 one mouse from 
                 116259 
                 3.255 
                 0.598 
                 7.999 
                 1.264 
               
               
                 Contralateral 
                 Groups A, B, C 
                 109482 
                 3.194 
                 0.566 
                 5.596 
                 1.025 
               
               
                 Tibia 
                 and AB) 
                 107508 
                 2.945 
                 0.346 
                 16.910 
                 1.860 
               
               
                 without 
                   
                 Average 
                 3.041 
                 0.477 
                 10.650 
                 1.419 
               
               
                 Tumour 
                   
                 SEM 
                 0.112 
                 0.062 
                 2.481 
                 0.179 
               
               
                 A 
                 Bortezomib, 
                 101426 
                 2.351 
                 0.307 
                 25.893 
                 2.443 
               
               
                   
                 0.6 mg/kg, twice 
                 105949 
                 2.025 
                 0.191 
                 26.044 
                 2.374 
               
               
                   
                 per week, i.p. 
                 107598 
                 3.109 
                 0.557 
                 16.877 
                 2.156 
               
               
                   
                   
                 109560 
                 3.146 
                 0.588 
                 23.179 
                 2.262 
               
               
                   
                   
                 113302 
                 1.790 
                 0.067 
                 32.463 
                 2.700 
               
               
                   
                   
                 115836 
                 1.893 
                 0.076 
                 33.152 
                 2.981 
               
               
                   
                   
                 116981 
                 2.201 
                 0.100 
                 34.609 
                 3.007 
               
               
                   
                   
                 117585 
                 1.617 
                 0.093 
                 31.813 
                 2.553 
               
               
                   
                   
                 117750 
                 2.300 
                 0.284 
                 27.147 
                 2.337 
               
               
                   
                   
                 117793 
                 2.448 
                 0.329 
                 23.582 
                 2.273 
               
               
                   
                   
                 Average 
                 2.288 
                 0.259 
                 27.476 
                 2.509 
               
               
                   
                   
                 SEM 
                 0.162 
                 0.061 
                 1.756 
                 0.094 
               
               
                 B 
                 MOR202, 
                 106446 
                 1.924 
                 0.082 
                 27.363 
                 2.546 
               
               
                   
                 3 mg/kg, three 
                 109482 
                 1.987 
                 0.388 
                 19.479 
                 2.079 
               
               
                   
                 times per week, 
                 112220 
                 2.155 
                 0.394 
                 21.858 
                 2.296 
               
               
                   
                 i.p. 
                 113668 
                 1.958 
                 0.276 
                 23.814 
                 2.429 
               
               
                   
                   
                 115187 
                 2.080 
                 0.440 
                 16.347 
                 1.871 
               
               
                   
                   
                 115956 
                 2.207 
                 0.460 
                 19.748 
                 2.193 
               
               
                   
                   
                 116312 
                 1.885 
                 0.234 
                 25.212 
                 2.368 
               
               
                   
                   
                 116798 
                 1.882 
                 0.254 
                 21.276 
                 2.436 
               
               
                   
                   
                 116944 
                 1.937 
                 0.276 
                 24.031 
                 2.368 
               
               
                   
                   
                 117773 
                 1.862 
                 0.160 
                 25.056 
                 2.511 
               
               
                   
                   
                 Average 
                 1.988 
                 0.296 
                 22.418 
                 2.310 
               
               
                   
                   
                 SEM 
                 0.038 
                 0.039 
                 1.044 
                 0.066 
               
               
                 C 
                 Vehicle Control 
                 107097 
                 1.619 
                 0.248 
                 22.779 
                 2.246 
               
               
                   
                 (0.9% Sodium 
                 112122 
                 1.608 
                 0.178 
                 26.514 
                 2.505 
               
               
                   
                 Chloride 
                 115971 
                 1.637 
                 0.241 
                 24.603 
                 2.485 
               
               
                   
                 Injection), 
                 116259 
                 1.880 
                 0.369 
                 19.334 
                 2.176 
               
               
                   
                 twice per week, 
                 116585 
                 2.060 
                 0.179 
                 24.120 
                 2.369 
               
               
                   
                 i.p. 
                 116779 
                 1.624 
                 0.190 
                 23.909 
                 2.417 
               
               
                   
                   
                 117054 
                 1.782 
                 0.131 
                 23.000 
                 2.541 
               
               
                   
                   
                 117110 
                 1.838 
                 0.281 
                 22.602 
                 2.312 
               
               
                   
                   
                 117242 
                 1.919 
                 0.281 
                 21.162 
                 2.193 
               
               
                   
                   
                 117375 
                 1.899 
                 0.283 
                 23.455 
                 2.338 
               
               
                   
                   
                 Average 
                 1.786 
                 0.238 
                 23.148 
                 2.358 
               
               
                   
                   
                 SEM 
                 0.050 
                 0.022 
                 0.615 
                 0.041 
               
               
                 AB 
                 Bortezomib/MOR202, 
                 107508 
                 3.303 
                 0.927 
                 16.902 
                 2.158 
               
               
                   
                 0.6/3 mg/kg, 
                 112625 
                 4.254 
                 1.661 
                 3.000 
                 0.772 
               
               
                   
                 twice/three 
                 113322 
                 3.684 
                 1.332 
                 3.888 
                 0.884 
               
               
                   
                 times per week, 
                 116030 
                 2.422 
                 0.192 
                 30.272 
                 2.542 
               
               
                   
                 i.p. 
                 116198 
                 3.537 
                 1.037 
                 8.023 
                 1.217 
               
               
                   
                   
                 116376 
                 1.933 
                 0.255 
                 22.059 
                 2.321 
               
               
                   
                   
                 116520 
                 2.793 
                 0.654 
                 22.439 
                 2.336 
               
               
                   
                   
                 117077 
                 3.402 
                 0.658 
                 7.207 
                 1.241 
               
               
                   
                   
                 117093 
                 2.436 
                 0.643 
                 17.454 
                 1.927 
               
               
                   
                   
                 117135 
                 3.026 
                 0.627 
                 13.699 
                 1.775 
               
               
                   
                   
                 Average 
                 3.079 
                 0.799 
                 14.494 
                 1.717 
               
               
                   
                   
                 SEM 
                 0.220 
                 0.144 
                 2.836 
                 0.204 
               
               
                   
               
            
           
         
       
     
     The Analysis of each parameter for synergistic activity was performed according to theorem of Clarke et al., as described in Example 4. Table 18 shows the calculations done in the determination of synergy of the combination of MOR202 and bortezomib. 
     
       
         
           
               
               
             
               
                 TABLE 18 
               
               
                   
               
             
            
               
                 When POSITIVE EFFECT has a HIGHER value: 
                 When POSITIVE EFFECT has a LOWER value: 
               
               
                 Antagonistic = (AB)/C &lt; (A/C) × (B/C) 
                 Antagonistic = (AB)/C &gt; (A/C) × (B/C) 
               
               
                 Additive = (AB)/C = (A/C) × (B/C) 
                 Additive = (AB)/C = (A/C) × (B/C) 
               
               
                 Synergistic = (AB)/C &gt; (A/C) × (B/C) 
                 Synergistic = (AB)/C &lt; (A/C) × (B/C) 
               
               
                   
               
            
           
           
               
               
               
               
               
            
               
                 Group 
                 Total BV 
                 Trabecular BV 
                 Trabecular pattern factor 
                 Structural Model Index 
               
               
                   
               
               
                 A 
                 2.288 
                 0.259 
                 27.476 
                 2.509 
               
               
                 B 
                 1.988 
                 0.296 
                 22.418 
                 2.31 
               
               
                 C 
                 1.786 
                 0.238 
                 23.148 
                 2.358 
               
               
                 AB 
                 3.079 
                 0.799 
                 14.494 
                 1.717 
               
               
                 (AB)/C 
                 1.723964166 
                 3.357142857 
                 0.626144807 
                 0.728159457 
               
               
                   
                 is bigger than 
                 is bigger than 
                 is less than 
                 is less than 
               
               
                 (A/C) × (B/C) 
                 1.425967052 
                 1.353435492 
                 1.149538246 
                 1.042377527 
               
               
                   
               
               
                 A = response to treatment with BOR at 0.6 mg/kg 
               
               
                 B = response to treatment with MOR202 at 3 mg/kg 
               
               
                 C = response to treatment with vehicle 0.9% Sodium Chloride 
               
               
                 AB = combination of treatments A and B 
               
            
           
         
       
     
     The numeric values shown in Table 18 are taken directly from the averages shown in Table 17 for each of the parameters in each of the Groups. The Groups described as A, B, C and AB are the same treatment groups in Tables 16-18. 
     In Total Bone Volume and Trabecular Bone Volume, (AB)/C is greater than (A/C)×(B/C) showing clear synergism. In Trabecular pattern factor and Structural Model Index, as described in Table 10, a lower value represents less bone lysis (efficacy in treatment), therefore, (AB)/C less than (A/C)×(B/C), supports clear synergism in both parameters. 
     Results 
     The inoculation of NCI-H929 multiple myeloma cells induced significant bone lysis in the tibiae of female SCID mice in this study, as indicated by the measurement of bone lysis through microCT scanning. The degree of bone lysis was significantly decreased in the tibia of mice treated with the combination of MOR202 and bortezomib as shown by microCT scanning. In each of the parameters of MicroCT Scan: Total Bone Volume (TBV), Trabecular Bone Volume (Tb.BV), Trabecular Pattern Factor (Tb.Pf) and Structure Model Index (SMI) the combination of MOR202 and bortezomib (Group AB) showed clear synergy in the reduction of bone lysis caused by the NCI-H929 multiple myeloma cells. 
     When the values in Table 17 are adjusted, so that the Control Group (Non-inoculated Contralateral Tibia without Tumour) is considered 0% bone lysis, and Group C (Vehicle Control (0.9% Sodium Chloride Injection) is considered 100% bone lysis, then MOR202 alone reduced bone lysis dose-dependently by up to 55% at 12 mg/kg compared to vehicle control, BOR alone at 0.6 mg/kg inhibited bone lysis by 40% and the combination of a lower dose of 3 mg/kg MOR202 and 0.6 mg/kg BOR completely abolished bone lysis. These findings support a synergistic effect of combination therapy. In addition, there was a reduction (&gt;90%) of M-protein serum levels in the combination group, indicating a significant decrease of tumor load. 
     Example 12 
     MOR202 and Bortezomib Alone and in Combination Against Human Non-hodgkin RAMOS Tumor in Female SCID Mice, Survival Model 
     Materials 
     Cyclophosphamide (Fluka, Buchs Switzerland, WB10468). Bortezomib (SYNthesis med chem., Shanghai, China, Lot no. #ZHM-066-054). Bortezomib was formulated in sterile 0.9% Sodium Chloride solution for dosing. MOR202 (MorphoSys AG, Lot 100706-5KLE18). Vehicle control: 0.9% Sodium Chloride Injection. SCID Mice (University of Adelaide, Waite Campus, Urrbaraie, SA, Australia, Strain C.B.-17-Igh-1 b -Prkdc scid ). 
     RAMOS cells (Oncodesign, Dijon Cedex, France) were cultivated in RPMI1640+20% heat inactivated alternate source FBS+1% Glutamax (Medium #2). Reagents for culture of RAMOS non-Hodgkin lymphoma cells were obtained from the following suppliers: RPMI 1640 cell culture medium, FBS, Glutamax, HEPES, sodium pyruvate, HBSS, and penicillin-streptomycin from Invitrogen Australia (Mt Waverley, VIC, Australia); and Trypan Blue and glucose from Sigma-Aldrich (Castle Hill, NSW, Australia). 
     Methods 
     Fifty-five female SCID mice were pre-treated with Cyclophosphamide (75 mg/kg, i.p., twice daily) for two days prior to RAMOS cell inoculation (Day −5 and −4). On the day of inoculation (Day −3), all fifty-five mice were inoculated with 1×10 6  RAMOS cells each (in 100 μL) intravenously into the tail vein. Forty-eight of the mice were randomised by body weight into six groups of eight. The dosing regimen for each group is shown in Table 19. 
     
       
         
           
               
             
               
                 TABLE 19 
               
             
            
               
                   
               
               
                 Dosing regimen 
               
            
           
           
               
               
               
               
               
            
               
                 Group 
                 Compound 
                 Treatment 
                 Intended Schedule 
                 Actual Schedule 
               
               
                   
               
               
                 A 
                 Bortezomib 
                 0.6 mg/kg, i.p., in 
                 Day −1, 3, 6, 10, 13 
                 Day −1, 3, 6 and 13 
               
               
                   
                   
                 10 mL/kg 
                 and 17 
                   
               
               
                 B 
                 MOR202 
                 1 mg/kg, i.v., in 
                 Day 0, 4, 7, 11, 14 
                 Day 0, 4, 7, 11, 14 and 18 
               
               
                   
                   
                 10 mL/kg 
                 and 18 
                   
               
               
                 C 
                 Vehicle Control (0.9% 
                 i.p., 10 mL/kg 
                 Day −1, 3, 6, 10, 13 
                 Day -1, 3, 6, 13 and 17 
               
               
                   
                 Saline for Injection) 
                   
                 and 17 
                   
               
               
                 AB 
                 Bortezomib/MOR202 
                 0.6/1 mg/kg, i.p./i.v., 
                 Day −1, 3, 6, 10, 13 
                 Day −1, 3, 6, 13, 17 and 20/ 
               
               
                   
                   
                 in 10 mL/kg 
                 and 17/Day 0, 4, 7, 
                 Day 0, 4, 7, 11, 14 and 18 
               
               
                   
                   
                   
                 11, 14 and 18 
               
               
                   
               
            
           
         
       
     
     The study continued for 98 days and the measured endpoint was survival. The results of each Group are shown in Table 20. 
     
       
         
           
               
             
               
                 TABLE 20 
               
             
            
               
                   
               
               
                 Survival Number and time period for each group 
               
            
           
           
               
               
               
               
            
               
                   
                   
                   
                 Number of mice 
               
               
                   
                   
                 % ILS (based 
                 alive at study 
               
               
                   
                 Day of death (post-inoculation) 
                 on median 
                 termination 
               
            
           
           
               
               
               
               
               
               
               
            
               
                   
                 Median 
                 Range 
                 Mean 
                 95% CI 
                 death day) 
                 (day 98) 
               
               
                   
               
            
           
           
               
               
               
               
               
               
               
            
               
                 A: BOR 0.6 
                 19 
                 18-20 
                 19.1 
                 18.4-19.8 
                  −7 
                 0/8 
               
               
                 mg/kg 
                   
                   
                   
                   
                   
                   
               
               
                 B: MOR202 
                 43.5 
                 38-52 
                 43.6 
                 39.0-48.3 
                 112 
                 0/8 
               
               
                 1 mg/kg 
                   
                   
                   
                   
                   
                   
               
               
                 C: Vehicle 
                 20.5 
                 20-22 
                 20.8 
                 20.0-21.5 
                 x 
                 0/8 
               
               
                 control 
                   
                   
                   
                   
                   
                   
               
               
                 AB: Combo 
                 45 
                 29-98 
                 61.6 
                  19.7-103.5 
                 120 
                 2/5 
               
               
                 BOR/MOR 
               
               
                   
               
            
           
         
       
     
     Analysis for synergistic activity was performed according to theorem of Clarke et al. Table 21 shows the calculations done in the determination of synergy of the combination of MOR202 and bortezomib. 
     
       
         
           
               
               
             
               
                 TABLE 21 
               
               
                   
               
             
            
               
                   
                 When POSITIVE EFFECT has a HIGHER value: 
               
               
                   
                 Antagonistic = (AB)/C &lt; (A/C) × (B/C) 
               
               
                   
                 Additive = (AB)/C = (A/C) × (B/C) 
               
               
                   
                 Synergistic = (AB)/C &gt; (A/C) × (B/C) 
               
               
                   
               
            
           
           
               
               
               
            
               
                   
                   
                 Median survival 
               
               
                   
               
               
                   
                 A 
                 19 
               
               
                   
                 B 
                 43.5 
               
               
                   
                 C 
                 20.5 
               
               
                   
                 AB 
                 45 
               
               
                   
                 (AB)/C 
                 2.195121951 
               
               
                   
                   
                 is bigger than 
               
               
                   
                 (A/C) × (B/C) 
                 1.966686496 
               
               
                   
               
               
                 A = response to treatment with BOR 0.6 mg/kg 
               
               
                 B = response to treatment with MOR202 1 mg/kg 
               
               
                 C = response to treatment vehicle 0.9% Sodium Chloride 
               
               
                 AB = combination of treatments A and B 
               
            
           
         
       
     
     The numeric values shown in Table 21 are taken directly from the median survival days shown in Table 20 for each of the Groups. The Groups described as A, B, C and AB are the same treatment groups in Tables 19-21. 
     The inoculation with RAMOS cells was lethal within a median time of 20.5 days in the control group. The combination of MOR202 and bortezomib, however, showed clear synergy in the increase in median survival days. Importantly, with the combination of MOR202 and bortezomib (Group AB), 2 out of 5 mice survived for the duration of the study. This strongly supports a synergistic finding of the combination of MOR202 and bortezomib. 
     It is to be understood that the description, specific examples and data, while indicating exemplary embodiments, are given by way of illustration and are not intended to limit the present invention. Various changes and modifications within the present invention will become apparent to the skilled artisan from the discussion, disclosure and data contained herein, and thus are considered part of the invention.