Patent Description:
The present disclosure provides an antibody drug conjugate (ADC) having an IgG antibody that binds to a CD38 target conjugated at a Cys site in the hinge region of an IgG antibody, as defined by claim <NUM>. The present disclosure further provides an ADC for use in a method for treating a multiple myeloma comprising providing an effective amount of a CD38 ADC, as defined by claim <NUM>.

CD38 is a <NUM> kD type II transmembrane glycoprotein with a long C-terminal extracellular domain and a short N-terminal cytoplasmic domain. The CD38 protein is a bifunctional ectoenzyme that can catalyze the conversion of NAD+ into cyclic ADP-ribose (cADPR) and also hydrolyze cADPR into ADP-ribose. During ontogeny, CD38 appears on CD34+ committed stem cells and lineage-committed progenitors of lymphoid, erythroid and myeloid cells. CD38 expression persists mostly in the lymphoid lineage with varying expression levels at different stages of T and B cell development.

CD38 is upregulated in many hematopoeitic malignancies and in cell lines derived from various hematopoietic malignancies, including non-Hodgkin's lymphoma (NHL), Burkitt's lymphoma (BL), multiple myeloma (MM), B chronic lymphocytic leukemia (B-CLL), B and T acute lymphocytic leukemia (ALL), T cell lymphoma (TCL), acute myeloid leukemia (AML), hairy cell leukemia (HCL,), Hodgkin's Lymphoma (HL), and chronic myeloid leukemia (CML). On the other hand, most primitive pluripotent stem cells of the hematopoietic system are CD38-. CD38 expression in hematopoietic malignancies and its correlation with disease progression makes CD38 an attractive target for anti-CD38 antibody therapy.

CD38 has been reported to be involved in Ca<NUM>+ mobilization (<NPL>; <NPL>) and in the signal transduction through tyrosine phosphorylation of numerous signaling molecules, including phospholipase C-γ, ZAP-<NUM>, syk, and c-cbl, in lymphoid and myeloid cells or cell lines (<NPL>; <NPL>; <NPL>; <NPL>; <NPL>; <NPL>; <NPL>; <NPL>; <NPL>;<NPL>; <NPL>). CD38 was proposed to be an important signaling molecule in the maturation and activation of lymphoid and myeloid cells during their normal development.

Evidence for the function of CD38 comes from CD38-/- knockout mice, which have a defect in their innate immunity and a reduced T-cell dependent humoral response due to a defect in dendritic cell migration (<NPL>; <NPL>). Nevertheless, it is not clear if the CD38 function in mice is identical to that in humans since the CD38 expression pattern during hematopoiesis differs greatly between human and mouse: a) unlike immature progenitor stem cells in humans, similar progenitor stem cells in mice express a high level of CD38 (<NPL>; <NPL>), b) while during the human B cell development, high levels of CD38 expression are found in germinal center B cells and plasma cells (<NPL>; <NPL>), in the mouse, the CD38 expression levels in the corresponding cells are low (<NPL>; <NPL>).

Several anti-human CD38 antibodies with different proliferative properties on various tumor cells and cell lines have been described in the literature. For example, a chimeric OKT10 antibody with mouse Fab and human IgG1 Fc mediates antibody-dependent cell-mediated cytotoxicity (ADCC) very efficiently against lymphoma cells in the presence of peripheral blood mononuclear effector cells from either MM patients or normal individuals (<NPL>). A CDR-grafted humanized version of the anti- CD38 antibody AT13/<NUM> has been shown to have potent ADCC activity against CD38-positive cell lines. Human monoclonal anti-CD38 antibodies have been shown to mediate the in vitro killing of CD38-positive cell lines by ADCC and/or complement-dependent cytotoxicity (CDC), and to delay the tumor growth in SCID mice bearing MM cell line RPMI-<NUM> (<CIT>). On the other hand, several anti-CD38 antibodies, IB4, SUN-4B7, and OKT10, but not IB6, AT1, or AT2, induced the proliferation of peripheral blood mononuclear cells (PBMC) from normal individuals (<NPL>).

Some of the antibodies of the prior art have been shown to be able to trigger apoptosis in CD38+ B cells. However, they can only do so in the presence of stroma cells or stroma-derived cytokines. An agonistic anti-CD38 antibody (IB4) has been reported to prevent apoptosis of human germinal center (GC) B cells (<NPL>), and to induce proliferation of KG-<NUM> and HL-<NUM> AML cells (<NPL>), but induces apoptosis in Jurkat T lymphoblastic cells (<NPL>). Another anti-CD38 antibody T16 induced apoptosis of immature lymphoid cells and leukemic lymphoblast cells from an ALL patient (<NPL>), and of leukemic myeloblast cells from AML patients (<NPL>), but T16 induced apoptosis only in the presence of stroma cells or stroma-derived cytokines (IL-<NUM>, IL-<NUM>, stem cell factor).

Documents <CIT> and <CIT> show antibody drug conjugates targeted with CD-<NUM> antibodies. We believe that improved antibody drug conjugates (ADCs), targeted with anti-CD38 antibodies, offer the promise and potential of delivering potent anti-tumor activity with the advantage of reduced side effects.

The present disclosure provides an anti-CD38 ADC composition comprising:.

In some embodiments, the drug or toxin moiety is selected from the group consisting of D1, D2, D3, D4, D5, and combinations thereof, wherein the structures of D1 D2, D3, D4 and D5 are:
<CHM>.

In some embodiments, the linker is selected from the group consisting of:
<CHM>
<CHM>
<CHM>
<CHM>
<CHM>
wherein the wavy line indicates a point of attachment to the conjugation moiety and the drug or toxin moiety.

In some embodiments, the conjugation moiety is
<CHM>
wherein the wavy line indicates the noint of attachment to the linker.

In another aspect there is provided an anti-CD38 ADC composition comprising:.

In some embodiments, the drug or toxin moiety is selected from the group consisting of D1, D2, D3, D4, D5, and combinations thereof, wherein the structures are:
<CHM>.

In some embodiments, the linker is selected from the group consisting of:
<CHM>
wherein the wavy line indicates a point of attachment to the conjugation moiety and the drug or toxin moiety.

In some embodiments, the conjugation moiety is
<CHM>
, wherein the wavy line indicates the point of attachment to the linker.

The antibody moiety is a variant of the CD38A2 wild type antibody disclosed and claimed in <CIT>, which is disclosed herein as heavy chain SEQ ID NO. <NUM> and light chain SEQ ID NO. <NUM> wherein the variant sequence alters the second and third amino acids from the N terminus of the light chain variable region, or the antibody moiety comprises CD38A2-SV (SV variant) having heavy chain SEQ ID NO. <NUM> and light chain SEQ ID NO.

The present disclosure provides antibody drug conjugates containing a novel human anti-CD38 antibody (A2) (described in <CIT> serial number <CIT>) with toxin moieties described herein including a tubulin inhibitor or a DNA damaging agent, such as doxorubicin analogs. The ADC conjugates retained binding affinity and showed potent cell killing in a variety of CD38 positive cell lines and in vivo.

The present disclosure provides an antibody drug conjugate (ADC) composition comprising an IgG antibody that binds to CD38, a conjugation linker moiety that binds to a single Cys residue in the hinge region of an IgG antibody, wherein the hinge region may be mutated such that the heavy chain hinge region contains only one Cys residue and not two, and a toxin moiety selected from the group consisting of derivatives of anthracyclines and Dolastatins. Preferably, the toxin moiety is a tubulin inhibitor or a doxorubicin analog. The antibody is an IgG antibody called human C38A2 (heavy/light SEQ ID NOs <NUM>/<NUM> in <CIT> or SEQ ID NOs. <NUM>/<NUM> for heavy/light chain variable regions herein) family or is C38A2-SV ( SEQ ID NOs. <NUM>/<NUM> for heavy/light chain variable regions herein). Preferably, the conjugated toxin with linker structure is selected from the group consisting of:
<CHM>
<CHM>
<CHM>
<CHM>
<CHM>
<CHM>
<CHM>.

The present disclosure provides a method for treating multiple myeloma, comprising administering an effective amount of an antibody drug conjugate (ADC) composition comprising an IgG antibody that binds to CD38, a conjugation linker moiety that binds to Cys residues in the hinge region of an IgG antibody and to a toxin moiety. By "binds to Cys residues" it is meant that the conjugation linker moiety may be covalently bound to the sulfur atoms of Cys residues in the hinge region of the IgG antibody. Preferably, the toxin moiety is a tubulin inhibitor or a doxorubicin analog. The antibody is an IgG antibody called human C38A2 (heavy/light SEQ ID NOs <NUM>/<NUM> in <CIT> or SEQ ID NOs. <NUM>/<NUM> for heavy/light chain variable regions herein) family or is C38A2-SV ( SEQ ID NOs. <NUM>/<NUM> for heavy/light chain variable regions herein). One of skill will recognize that toxin moieties as disclosed herein, conjugated to a linker and a conjugation moiety as disclosed herein, represent intermediate toxin linker conjugates, which, when covalently bound (conjugated to) the IgG antibody as disclosed herein, are ADCs as disclosed herein. Preferably, the conjugated toxin with linker structure is selected from the group consisting of (with each compound number indicated):
<CHM>
<CHM>
<CHM>
<CHM>
<CHM>
<CHM>
and
<CHM>.

The wavy line indicates the point of attachment to the linker.

In some embodiments, the drug linker conjugate comprises a linker L2 and a conjugation moiety, wherein the linker L2 is covalently bound to the conjugation moiety; the conjugation moiety is capable of reacting with free cysteine thiol groups in the hinge region of an IgG antibody. In some embodiments, the conjugation moiety has the structure
<CHM>
("conjugation method L1").

<CHM>
<CHM>
<CHM>
<CHM>
<CHM>
<CHM>
<CHM>.

As used herein, common organic abbreviations are defined as follows:.

Compound <NUM> (<NUM>, <NUM> pmol) was dissolved in <NUM> of anhydrous DCM under nitrogen. Then DIEA (<NUM>µL, <NUM>µmol) was added and the reaction mixture was cooled with ice bath. Then methane sulfonyl chloride (<NUM>µL, <NUM> pmol) was added and the mixture was stirred for <NUM>. The reaction was diluted with <NUM> of DCM and washed with <NUM> of water, dried over anhydrous Na<NUM>SO<NUM> and evaporated to give compound <NUM> as a red solid (<NUM>, <NUM>%). MS m/z=<NUM> (M+H).

Compound <NUM> (<NUM>, <NUM> pmol) was dissolved in <NUM> of anhydrous ethanol under nitrogen. Then thioamide <NUM> (<NUM>, <NUM>µmol) was added and the mixture was heated at <NUM> for <NUM>. The mixture was purified by HPLC to give compound <NUM> as a red solid (<NUM>, <NUM>%). MS m/z=<NUM> (M+H).

Compound <NUM> (<NUM>, <NUM> pmol) was dissolved in <NUM> of anhydrous DMF under nitrogen. Then <NUM>µL of piperidine was added and the mixture was stirred at ambient temperature for <NUM>. The mixture was purified by HPLC to give compound <NUM> as a red solid (<NUM>, <NUM>%). MS m/z=<NUM> (M+H).

Compound <NUM> (<NUM>, <NUM> pmol) was dissolved in <NUM> of DMF, then HATU (<NUM>, <NUM> pmol) and DIEA (<NUM>µL, <NUM> pmol) was added. After <NUM>, compound <NUM> (<NUM>, <NUM> pmol) was added and the mixture was stirred at ambient temperature for <NUM>. To the mixture was added <NUM>µL of DBU and stirred for <NUM>. Then the mixture was purified by HPLC to give compound <NUM> as a red solid (<NUM>, <NUM>%). MS m/z=<NUM> (M+H).

Compound <NUM> (<NUM>, <NUM> pmol) was dissolved in <NUM> of DCM, then DIC (<NUM>, <NUM> pmol) was added. After <NUM>, compound <NUM> (<NUM>, <NUM> pmol) dissolved in <NUM> of DMF was added and the mixture was stirred at ambient temperature for <NUM>. Then the mixture was purified by HPLC to give compound <NUM> as a red solid (<NUM>, <NUM>%). MS m/z=<NUM> (M+H) Synthesis of compound <NUM>:
<CHM>.

To a solution of acid <NUM> (<NUM>, <NUM> pmol) in <NUM> of DCM, add N-hydroxysuccinimide (<NUM>, <NUM> pmol), and EDC (<NUM>, <NUM> pmol). After <NUM>, the mixture was washed with water (2x6 mL), dried over Na<NUM>SO<NUM> and evaporated. The residue was dissolved in <NUM> of DMF. Then amine <NUM> (<NUM>, <NUM> pmol, as TFA salt) was added, followed by DIEA (<NUM>µL). The mixture was stirred for <NUM>. Then piperidine (<NUM>µL) was added and stirred for <NUM>. The mixture was purified by HPLC to give compound <NUM> (<NUM>, <NUM>%) as a red solid; MS m/z <NUM> (M+H).

To a solution of compound <NUM> as TFA salt (<NUM>, <NUM> pmol) in <NUM> of DMF was added compound <NUM> (<NUM>, <NUM> pmol), DIEA (<NUM>µL), HOBt (<NUM>, <NUM> pmol). After <NUM>, the reaction was completed and then piperidine (<NUM>µL) was added and stirred for <NUM>. The mixture was purified by HPLC to give compound <NUM> (<NUM>, <NUM>%) as a white solid; MS m/z <NUM> (M+H).

Compound <NUM> (<NUM>, <NUM> pmol) was dissolved in <NUM> of DCM, then DIC (<NUM>, <NUM> pmol) was added. After <NUM>, compound <NUM> (<NUM>, <NUM> pmol) dissolved in <NUM> of DMF was added and the mixture was stirred at ambient temperature for <NUM>. Then the mixture was purified by HPLC to give compound <NUM> as a white solid (<NUM>, <NUM>%). MS m/z=<NUM> (M+H) Synthesis of compound <NUM>:
<CHM>.

To a solution of compound <NUM> as TFA salt (<NUM>, <NUM> pmol) in <NUM> of DMF was added compound <NUM> (<NUM>, <NUM> pmol), DIEA (<NUM>µL), HOBt (<NUM>, <NUM> pmol). After <NUM>, the reaction was completed and then piperidine (<NUM>µL) was added and stirred for <NUM>. The mixture was purified by HPLC to give compound <NUM> (<NUM>, <NUM>%) as a white solid; MS m/z <NUM> (M+H). Synthesis of compound <NUM>:.

Compound <NUM> (<NUM>, <NUM> pmol) was dissolved in <NUM> of DCM, then DIC (<NUM>, <NUM> pmol) was added. After <NUM>, compound <NUM> (<NUM>, <NUM> pmol) dissolved in <NUM> of DMF was added and the mixture was stirred at ambient temperature for <NUM>. Then the mixture was purified by HPLC to give compound <NUM> as a white solid (<NUM>, <NUM>%). MS m/z=<NUM> (M+H). Synthesis of compound <NUM>:
<CHM>.

Compound <NUM> (<NUM>, <NUM>µmol) was dissolved in <NUM> of DMF, then HATU (<NUM>, <NUM>µmol) and DIEA (<NUM>µL) was added. After <NUM>, compound <NUM> as TFA salt (<NUM>, <NUM>µmol) was added and the mixture was stirred at ambient temperature for <NUM>. To the mixture was added <NUM>µL of DBU and stirred for <NUM>. Then the mixture was purified by HPLC to give compound <NUM> as a white solid (<NUM>, <NUM>%). MS m/z=<NUM> (M+H).

Compound <NUM> (<NUM>, <NUM>µmol) was dissolved in <NUM> of DCM, then DIC (<NUM>, <NUM>µmol) was added. After <NUM>, compound <NUM> (<NUM>, <NUM>µmol) dissolved in <NUM> of DMF was added and the mixture was stirred at ambient temperature for <NUM>. Then the mixture was purified by HPLC to give compound <NUM> as a white solid (<NUM>, <NUM>%). MS m/z=<NUM> (M+H).

To a round bottom flask add compound <NUM> as TFA salt (<NUM>, <NUM> mmol), compound <NUM> (<NUM>, <NUM> mmol), HOAt (<NUM>, <NUM> mmol), DCM (<NUM>), DIEA (<NUM>µL), and DIC (<NUM>, <NUM> mmol). After <NUM> of stirring dilute the reaction mixture with <NUM> of DCM, then wash it with <NUM> of water, dry over Na<NUM>SO<NUM>, evaporate solvent under vacuum to give crude glassy solid which was used in the next step. Dissolve the obtained solid in mixture consisting of <NUM> of DCM, <NUM> of TFA and <NUM> of triisopropylsilane and stir for <NUM>. Evaporate the solvent under vacuum and purify by HPLC to give compound <NUM> (<NUM>, <NUM>%), MS m/z <NUM> (M+H).

To a round bottom flask add compound <NUM> (<NUM>, <NUM> mmol), <NUM> of ACN, <NUM> of water and <NUM> of sat. NaHCO<NUM> aq. Then add Na<NUM>S<NUM>O<NUM> (<NUM>, <NUM> mmol) and continue stirring for <NUM>. Purify the mixture by HPLC to give compound <NUM> (<NUM>, <NUM>%), MS m/z <NUM> (M+H).

Compound <NUM> (<NUM>, <NUM> mmol) was dissolved in <NUM> of ACN and <NUM>,<NUM>-dibromo-<NUM>,<NUM>-butanedione (<NUM>) (<NUM>, <NUM> mmol) was added. After stirring for <NUM>, the reaction was purified by HPLC to give compound <NUM> (<NUM>, <NUM>%), MS m/z <NUM> (M+H).

To a solution of amine <NUM> (<NUM>, <NUM> mmol) and acid <NUM> (<NUM>, <NUM> mmol) in <NUM> of DMF, was added Oxima-pure (<NUM>, <NUM> mmol), followed by DIC (<NUM>, <NUM> mmol). After <NUM>, the coupling was completed and then <NUM> of piperidine was added and stirred for <NUM>. The mixture was purified by HPLC to give compound <NUM> TFA salt (<NUM>, <NUM>%) as a white solid; MS m/z <NUM> (M+H).

Compound <NUM> TFA salt (<NUM>, <NUM> umol) was dissolved in <NUM> of ACN and <NUM> of water. Then a solution of <NUM>,<NUM>-dibromo-<NUM>,<NUM>-butanedione <NUM> (<NUM>, <NUM> pmol) in <NUM> of ACN was added. After stirring for <NUM>, the reaction was purified by HPLC to give compound <NUM> as a white solid (<NUM>, <NUM>%). MS m/z=<NUM> (M+H).

Affinity purified anti-CD38 antibody was buffer exchanged into <NUM> sodium phosphate buffer, pH <NUM>-<NUM> with <NUM> EDTA at a concentration of <NUM>-<NUM>/mL To a portion of this antibody stock was added a freshly prepared <NUM> water solution of tris(<NUM>-carboxyethyl)phosphine) (TCEP) in up to <NUM>-fold molar excess. The resulting mixture was incubated at <NUM>-<NUM> overnight. The excess TCEP was removed by gel-filtration chromatography or several rounds of centrifugal filtration. UV-Vis quantification of recovered, reduced antibody material was followed by confirmation of sufficient free thiol-to-antibody ratio. Briefly, a <NUM> aliquot of freshly prepared (<NUM>,<NUM>'-dithiobis-(<NUM>-nitrobenzoic acid) in <NUM> sodium phosphate, pH <NUM>-<NUM>, <NUM> EDTA was mixed with an equal volume of purified antibody solution. The resulting absorbance at <NUM> was measured and the reduced cysteine content was determined using the extinction coefficient of <NUM>,<NUM>-<NUM>cm-<NUM>.

To initiate conjugation of compound <NUM> to anti-CD38 antibody, L014-<NUM> was first dissolved in a <NUM>:<NUM> acetonitrile/water mixture at a concentration of <NUM>. An aliquot of this freshly prepared toxin-linker solution was then added to a portion of the reduced, purified anti-CD38 antibody intermediate in <NUM>-<NUM> fold molar excess. After thorough mixing and incubation at ambient temperature for ≥<NUM>, the crude conjugation reaction was analyzed by HIC-HPLC chromatography to confirm reaction completion (disappearance of starting antibody peak) at <NUM> wavelength detection. Purification of ADC46 was then carried out by gel-filtration chromatography using an AKTA system equipped with a Superdex <NUM> pg column (GE Healthcare) equilibrated with PBS. The drug-to-antibody ratio (DAR) was calculated based on UV-VIS and HIC-HPLC. <FIG> shows a representative HIC-HPLC comparison of starting anti-CD38 antibody and purified ADC46. Confirmation of low percent (<<NUM>%) high molecular weight (HMW) aggregates for the resulting ADC46 was determined using analytical SEC-HPLC.

Reduction and analysis of anti-CD38 antibody for ADC41 was conducted in a manner identical to the procedure used to generate ADC46. To initiate final drug-linker conjugation to antibody, Compound <NUM> was first dissolved in a <NUM>:<NUM> acetonitrile/water mixture at a concentration of <NUM>. Propylene glycol (PG) was then added to an aliquot of the reduced, purified anti-CD38 antibody to give a final concentration of <NUM>-<NUM>% (v/v) PG before addition of the freshly prepared compound <NUM> solution in <NUM>-<NUM>-fold molar excess. Subsequent analysis and purification of ADC41 was carried out in a manner identical to the procedure for ADC46. <FIG> shows a representative HIC-HPLC comparison of starting anti-CD38 antibody and purified ADC41.

Upon receipt, animals were housed <NUM> mice per cage in a room with a controlled environment. Animals were provided rodent chow and water ad libitum. Acclimation of the mice to laboratory conditions was at least <NUM> hours prior to the start of cell administration and dosing. During the acclimation period, the animals' health status was determined. Only animals that are observed to be healthy prior to study initiation were used.

This example provides an in vivo experiment comparing treatment of mice with control (PBS), anti-CD38 IgG1 antibody (STI-<NUM> and STI-<NUM>) and an ADC variant of both antibodies. The procedure first does a tumor cell inoculation & establishment of tumors:.

This example is an in vivo experiment comparing two disclosed CD38 ADCs in vivo with mice In the in vivo study, <NUM> million of Daudi-fluc cells were injected iv into NOD-SCID mice. <NUM> days after tumor established in mice, anti-CD38 antibody and ADCs were injected to mice by IV. The inhibition of tumor growth by antibody or ADCs was monitored by the luminesce intensity change of the tumor (<FIG>, <FIG> and <FIG>).

ADC#<NUM> and ADC# 41were tested. Both ADC's use the same A2 antibody. The Daudi and Ramos cell line was obtained from ATCC. The cells were cultured in RPMI <NUM>1X medium with <NUM>% FBS and at <NUM> in a <NUM>% carbon dioxide humidified environment. Cells were cultured for a period of <NUM> weeks and were passaged <NUM> times before harvest. Prior to injection, Daudi or Ramos cells were resuspended in a <NUM>:<NUM> ratio of HBSS (Hank's balanced salt solution) and Matrigel, and <NUM> million cells per <NUM> were injected subcutaneously into the upper right flank of each mouse.

The Daudi-luc cells were cultured in RPMI <NUM>1X medium with <NUM>% FBS and <NUM>. 2ug/ml puromycin at <NUM> in a <NUM>% carbon dioxide humidified environment. Cells cultured for a period of <NUM> weeks and were passaged <NUM> times before harvest. Prior to injection, Daudi-luc cells were resuspended in HBSS. <NUM> million cells per <NUM> were injected intravenously in to the tail vein of each mouse.

Female NOD SCID mice aged <NUM> weeks (Charles River) were used for Daudi subcutaneous xenografts and Daudi-luc intravenous xenografts. Female Nu/Nu mice aged <NUM> weeks (Charles River) were used for Ramos subcutaneous xenografts in the studies. Upon receipt, mice were housed <NUM> mice per cage in a room with a controlled environment. Rodent chow and water was provided ad libitum. Mice were acclimated to laboratory conditions for <NUM> hours before the start of dosing. The animals' health status was monitored during the acclimation period. Each cage was identified by group number and study number, and mice were identified individually by ear tags.

The study design and dosing regimens are shown in the following table.

Tumor growth was monitored by measurement of tumor width and length using a digital caliper starting day <NUM>-<NUM> after inoculation, and followed twice per week until tumor volume reached ~<NUM>-<NUM><NUM>. Tumor volume was calculated using the formula: Volume (mm<NUM>) = [Length (mm) × Width (mm)<NUM>] / <NUM>.

Once tumors were staged to the desired volume, animals were randomized, and mice with very large or small tumors were culled. Mice were divided into groups with animal numbers per group as indicated in study design. Mice were then treated intravenously (<NUM>/animal) with either PBS, Ab, ADC#<NUM>, or ADC#<NUM>. Tumor growth, animal health and body weight were monitored after treatment. Testing animals were sacrificed when the average subcutaneous tumor load for the group exceeded <NUM><NUM>, animal body-weight loss exceed <NUM>%, or at the end of the study.

Tumor volume was measured twice weekly throughout the experimental period. TGI (tumor growth inhibition %) was calculated using the formula: TGI = [(Last Volume Measurement of PBS Group - Volume of Treatment group on the same day as the PBS control)/ (Last Volume Measurement of PBS Group)] × <NUM>. The body weight of each mouse was measured twice weekly by electric balance.

Raw data of individual body weight and tumor volume were calculated. Group average and standard deviation were calculated, and statistical analyses (one-way ANOVA with Dunnett's multiple comparison test; GraphPad Prism <NUM>) was carried out. All treatment groups were compared with the PBS group. P<<NUM> was considered statistically significant.

ADC#<NUM> at <NUM>/kg significantly inhibited Daudi tumor growth compared to PBS treated control group. Although the tumor regained growth after <NUM> weeks, the single <NUM>/kg treatment significantly delayed tumor growth. In this case, multiple treatment may be tested to achieve sustained tumor inhibition. While a single dose of ADC#<NUM> at <NUM>/kg or <NUM>/kg did not significantly inhibited tumor growth. However, although the difference was not significant, a single dose of ADC#<NUM> did show slightly inhibition of tumor growth compared to PBS treated control group. Dose response was observed in this study, where higher dose showed better tumor growth inhibition (<FIG>). There was no body weight loss in the testing animals with a single dose of intravenously administrated ADC#<NUM> at <NUM>/kg or lower dose (<FIG>).

Similarly, ADC#<NUM> at <NUM>/kg significantly inhibited Ramos tumor growth compared to PBS treated control group and had sustained tumor inhibition effect for up to <NUM> days. A single dose of ADC#<NUM> at <NUM>/kg or <NUM>/kg did not significantly inhibit tumor growth. However, although the difference was not significant, a single dose of ADC#<NUM> at <NUM>/kg or <NUM>/kg did show slightly inhibition of tumor growth compared to PBS treated control group. Dose response was observed in this study, where higher dose showed better tumor growth inhibition (<FIG>). There was no body weight loss in the testing animals with a single dose of intravenously administrated ADC#<NUM> at <NUM>/kg or lower dose (<FIG>).

ADC#<NUM> at a single dose of <NUM>/kg completely inhibited tumor growth with <NUM>% survival up to Day <NUM> after treatment. ADC#<NUM> at a single dose of <NUM>/kg significantly inhibited tumor growth compared to PBS control group, and significantly prolonged survival in mice.

ADC#<NUM> and ADC#<NUM>, at <NUM>/kg single dose, significantly inhibited tumor growth, while at <NUM>/kg, or <NUM>/kg, both did not show significant tumor inhibition in Daudi and Ramos subcutaneously injected xenograft tumor model in mice. ADC#<NUM> at <NUM>/kg single dose completely inhibited tumor growth with a <NUM>% survival up to <NUM> days in Daudi-luc intravenously injected tumor model in female NOD SCID mice. ADC#<NUM> at <NUM>/kg single dose significantly inhibited tumor growth, and prolonged survival in Daudi-luc intravenously injected tumor model in female NOD SCID mice. Dose response was observed for ADC#<NUM> and ADC#<NUM> in this study. ADC#<NUM> showed better tumor growth inhibition effect than ADC#<NUM> with the same (<NUM>/kg, or <NUM>/kg) dose regime. No treatment-related body weight loss was observed during the study for all treatment groups.

Claim 1:
An anti-CD38 ADC composition comprising:
(a) an anti-CD38 IgG antibody C38A2-SV (SEQ ID NOs. <NUM>/<NUM> for heavy/light chain variable regions herein) or C38A2 (SEQ ID NOs. <NUM>/<NUM> for heavy/light chain variable regions herein);
(b) a drug or toxin moiety that is a tubulin inhibitor or a doxorubicin analog; and
(c) a conjugation linker moiety, wherein the conjugation linker comprises a linker and a conjugation moiety which covalently binds to a single Cys residue in a hinge region of an IgG antibody, and wherein a heavy chain hinge region of an IgG antibody may be mutated such that the heavy chain hinge region contains only one Cys residue.