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Cancer remains the second most common cause of death in the US. Due to a recurrent problem with anticancer drug resistance, there is a current need for anticancer drugs with distinct modes of action for combination drug therapy We have tested two novel piperidone compounds, named 2608 (1-dichloroacetyl - 3,5-bis(3,4-difluorobenzylidene)-4-piperidone) and 2610 (1-dichloroacetyl-3,5-bis(3,4-dichlorobenzylidene)-4-piperidone), for their potential cytotoxicity on numerous human cancer cell lines. We found that both compounds were cytotoxic for breast, pancreatic, leukemia, lymphoma, colon, and fibroblast cell lines, with a cytotoxic concentration 50% (CCsub50/sub) in the low micromolar to nanomolar concentration range. Further assays focused primarily on an acute lymphoblastic lymphoma and colon cancer cell lines since they were the most sensitive and resistant to the experimental piperidones. The cell death mechanism was evaluated through assays commonly used to detect the induction of apoptosis. These assays revealed that both 2608 and 2610 induced reactive oxygen species (ROS) accumulation, mitochondrial depolarization, and activated caspase-3/7. Our findings suggest that the piperidones induced cell death via the intrinsic apoptotic pathway. Additional assays revealed that both piperidones cause cell cycle alteration in lymphoma and colon cell lines. Both piperidones elicited DNA fragmentation, as evidenced by an increment in the sub-G0/G1 subpopulation in both cell lines. Similar to other related compounds, both piperidones were found to act as proteasome inhibitors by increasing the levels of poly-ubiquitinated proteins in both lymphoma and colon cell lines. Hence, the two piperidones exhibited attractive cytotoxic properties and suitable mechanisms of action, which makes them good candidates as anticancer drugs.

PDF - 68K, Supplementary Materials and Methods.

Gating strategy for immune subsets in pancreatic tumor

PDF - 855K, Supplementary Figure 1. Anti- EMP2 IgG1 can detect EMP2 in human tissue. Supplementary Figure 2. The serum levels of the human EMP2 IgG1 was compared to those of Trastuzamab.

PDF - 59K, Known Profile of EMP2 Upregulation in Breast Cancer.

PDF - 51K, Supplementary Table 3. Expression levels of EMP2 in triple negative breast cancer.

PDF - 54K, Supplementary Table 2. Clinical Data for Invasive Breast Cancer Patients used in the TMA.

Despite significant advances in biology and medicine, the incidence and mortality due to breast cancer worldwide is still unacceptably high. Thus, there is an urgent need to discover new molecular targets. In this article, we show evidence for a novel target in human breast cancer, the tetraspan protein epithelial membrane protein-2 (EMP2). Using tissue tumor arrays, protein expression of EMP2 was measured and found to be minimal in normal mammary tissue, but it was upregulated in 63% of invasive breast cancer tumors and in 73% of triple-negative tumors tested. To test the hypothesis that EMP2 may be a suitable target for therapy, we constructed a fully human immunoglobulin G1 (IgG1) antibody specific for a conserved domain of human and murine EMP2. Treatment of breast cancer cells with the anti-EMP2 IgG1 significantly inhibited EMP2-mediated signaling, blocked FAK/Src signaling, inhibited invasion, and promoted apoptosis in vitro. In both human xenograft and syngeneic metastatic tumor monotherapy models, anti-EMP2 IgG1 retarded tumor growth without detectable systemic toxicity. This antitumor effect was, in part, attributable to a potent antibody-dependent cell-mediated cytotoxicity response as well as direct cytotoxicity induced by the monoclonal antibody. Together, these results identify EMP2 as a novel therapeutic target for invasive breast cancer. Mol Cancer Ther; 13(4); 902–15. ©2014 AACR.

Epstein–Barr virus (EBV) is a human gammaherpesvirus associated with the development of hematopoietic cancers of B-lymphocyte origin, including AIDS-related non-Hodgkin lymphoma (AIDS-NHL). Primary infection of B-cells with EBV results in their polyclonal activation and immortalization. The transferrin receptor 1 (TfR1), also known as CD71, is important for iron uptake and regulation of cellular proliferation. TfR1 is highly expressed in proliferating cells, including activated lymphocytes and malignant cells. We developed a mouse/human chimeric antibody targeting TfR1 (ch128.1/IgG1) that has previously shown significant antitumor activity in immunosuppressed mouse models bearing human malignant B-cells, including multiple myeloma and AIDS-NHL cells. In this article, we examined the effect of targeting TfR1 to inhibit EBV-driven activation and growth of human B-cells in vivo using an immunodeficient NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ [NOD/SCID gamma (NSG)] mouse model. Mice were implanted with T-cell–depleted, human peripheral blood mononuclear cells (PBMCs), either without EBV (EBV−), or exposed to EBV in vitro (EBV+), intravenously via the tail vein. Mice implanted with EBV+ cells and treated with an IgG1 control antibody (400 μg/mouse) developed lymphoma-like growths of human B-cell origin that were EBV+, whereas mice implanted with EBV+ cells and treated with ch128.1/IgG1 (400 μg/mouse) showed increased survival and significantly reduced inflammation and B-cell activation. These results indicate that ch128.1/IgG1 is effective at preventing the growth of EBV+ human B-cell tumors in vivo, thus, indicating that there is significant potential for agents targeting TfR1 as therapeutic strategies to prevent the development of EBV-associated B-cell malignancies.Significance:An anti-TfR1 antibody, ch128.1/IgG1, effectively inhibits the activation, growth, and immortalization of EBV+ human B-cells in vivo, as well as the development of these cells into lymphoma-like tumors in immunodeficient mice.

Figure S1 , Figure S2

PDF - 64K, Supplementary Table 4. Evaluation of EMP2 IgG1 toxicity compared to control IgG following 7 weeks of treatment delivered systemically IP. Supplementary Table 5. Evaluation of EMP2 IgG1 toxicity compared to control IgG following systemic IP treatment of up to 40mg/kg.

PDF - 55K, Legends for Supplementary Tables 1 through 5.

Figures S2, S3 and S4. Morphometric analysis of laminin-411 β1 chain (S2) and cancer stem cell markers (Notch-1, Notch-3, CD133, Nestin, and c-Myc; S3 and S4) on frozen sections of patient brain tumors. The expression of all markers significantly correlated with glioma grade

Video schematically shows the mechanism of glioblastoma treatment by laminin-411 inhibiting nanobioconjugate

SUPPL FIG 2: A) 8226 cells treated with or w/o NSC 126405 for 6 hrs, followed by immunoprecipitation of mTOR (or control IgG). Immunoprecipitates immunoblotted for expression of mTOR and DEPTOR. Input shown for DEPTOR, mTOR and tubulin expression. B) 8226 cells treated with DMSO, pp242 or NSC 126405 and then tested for 4E-BP1 phosphorylation by flow cytometry. C) Summarized data from all 3 treatment groups

Figure S5. Binding of the nanobioconjugates to U87MG studied by flow cytometry is inhibited by human soluble TfR. This shows that the chimeric anti-human TfR antibody as part of nanobioconjugate fully retained its activity

legends of supplemental figures

SUPPL FIG 6: PBLs obtained from 3 normal individuals and incubated with increasing concentrations of NSC 126405 for 72 hrs. Surviving viable cells enumerated. Data are % of control (no drug B), mean+/-SD, n=3.

SUPPL FIG 1: A) Only yeast transfected with both FAT domain/GAL4DBD +DEPTOR/GAL4 AD show growth on selective media. Yeast transfected w/ either construct alone can not grow. Two separate transformations shown. B) Four initial 'hits' from drug screen

Figure S10. Expression of Notch-1 and its ligand Jagged1 is significantly decreased in patient-derived glioblastoma cell lines with laminin-411 disruption by CRISPR/Cas9

Figures S8 and S9. Quantification of expression of laminin-411 β1 chain, its binding β1 integrin, Notch family members and other stem cell markers after nanobioconjugate treatment. All markers were significantly suppressed in treated tumors.

Supplementary Materials 1 & 2 from Cytokines fused to antibodies and their combinations as therapeutic agents against different peritoneal HER2/neu expressing tumors

We have previously generated antihuman HER2/neu–humanized IgG3 fused to interleukin-2 (IL-2), IL-12, or granulocyte macrophage colony-stimulating factor (GM-CSF) [monofunctional fusion proteins (mono-AbFP)] or fused to IL-2 and IL-12 or IL-12 and GM-CSF [bifunctional fusion proteins (bi-AbFP)]. These AbFPs retained cytokine and antigen-binding activities. We have now further characterized the AbFPs and determined the heparin-binding activity of the fused cytokines, their ability to trigger IFN-γ secretion and natural killer (NK) activation, and their direct antitumor efficacy. Flow cytometry revealed heparin-binding activity in the AbFPs containing IL-12 and IL-2, although this activity seems to be decreased in the bi-AbFPs. However, both bi-AbFPs retained the capacity to stimulate IL-12-dependent IFN-γ secretion in the NK cell line KY-1, and IL-12/IL-2 bi-AbFP induced NK activity in splenocytes. The antitumor effectiveness of bi-AbFPs and mono-AbFP combinations was studied in mice challenged i.p. with three different human HER2/neu murine syngeneic models (D2F2/E2, CT26-HER2/neu, and MC38-HER2/neu). Although a significant variability in the profile of antitumor response was observed in the different tumor models, the combination of IL-12 and GM-CSF mono-AbFPs protected 100% of D2F2/E2-challenged and 75% of CT26-HER2/neu–challenged mice. In contrast, bi-AbFPs protected less than the combination of mono-AbFPs and, in some models, even less than mono-AbFPs alone. However, in all cases, most of long-term survivors showed protection after s.c. rechallenge with the tumors and later with the parental tumors not expressing HER2/neu. These results show that, although the pattern of protection is tumor model dependent, treatments with AbFPs can effectively generate high levels of protection against peritoneal tumors expressing HER2/neu, which may be relevant in patients with primary or metastatic peritoneal carcinomatosis that may be observed in ovarian, colon, stomach, bladder, lung, and breast cancers. [Mol Cancer Ther 2006;5(4):1029–40]

Treatment of human epidermal growth factor receptor 2 (HER2/neu)-expressing breast cancer patients with a monoclonal antibody (mAb) directed against HER2/neu improves the outcome of chemotherapy. In cases in which remission is observed, antibody-dependent cell-mediated cytotoxicity (ADCC) seems to be one of the main mechanisms of anti-HER2/neu mAb action, implicating Fcγ receptors (FcγRs) in this tumoricidal activity. In vitro and in vivo studies have revealed that anti-HER2/neu-mediated ADCC is mainly accomplished by polymorphonuclear granulocytes (PMN). C5a, a cleavage product of the complement component C5, modulates FcγR expression via upregulation of activating and downregulation of inhibitory FcγRs. C5a also recruits PMNs to sites of inflammation and increases PMN survival. To enhance the recruitment and activation of C5a receptor–bearing cells into the tumor microenvironment, we developed antibody fusion proteins composed of a human IgG3 anti-HER2/neu antibody genetically fused to C5a [anti-HER2/neu IgG3-(C5a)] or to its derivative, C5adesArg [anti-HER2/neu IgG3-(C5adesArg)]. Both fusion proteins were expressed, properly assembled, and secreted by murine myeloma cells, and displayed chemotactic activity on human PMN. Under comparable conditions, anti-HER2/neu IgG3-(C5adesArg) increased the survival of PMN more efficiently than anti-HER2/neu IgG3-(C5a) or C5adesArg. Surprisingly, incubation of the fusion proteins with breast cancer cells that overexpress HER2/neu (SK-BR-3) induced cell death at a dose at which the anti-HER2/neu IgG3 antibody was innocuous. In the presence of human peripheral blood leukocytes as effector cells, both fusion proteins induced tumor cell death more efficiently than anti-HER2/neu IgG3. These data suggest that anti-HER2/neu IgG3-(C5a) and anti-HER2/neu IgG3-(C5adesArg) fusion proteins possess novel properties that could be useful in cancer immunotherapy. Mol Cancer Ther; 9(8); 2175–85. ©2010 AACR.

SUPPL FIG 3: A) Relative DEPTOR expression in MMCLs and hepatocellular carcinoma (HCC) cell lines in a representative experiment (upper panel) and means+/-SD of 3 experiments (lower bar graph); B) Relative sensitivity of U266 and FR4 MMCLs vs DEPTOR-expressing JHH7, HUH-1 or HIH7 HCCs; data are means +/- SD (n=3); Asterix demonstrates significant (p

Figures S6 and S7. Localization of labeled nanobioconjugates in tumors and biodistribution in brain. The nano drug is seen in the tumor parenchyma outside of blood vessels (S6) and preferentially accumulates in the tumor (peaks at 6 hr post-injection) as compared to contralateral brain tissue (S7)

Supplementary Figure 1 from Decreased Survival of Human Breast Cancer Cells Expressing HER2/neu on In vitro Incubation with an Anti-HER2/neu Antibody Fused to C5a or C5adesArg

Tables S1, S2 and S3, and Figure S1. List of used antibodies (Table S1), and immunohistochemical expression of laminin-411 and laminin-421 in human gliomas of various grades (Tables S2 and S3). Expression of laminin-411 vs. laminin-421 increases with glioma grade and in GBMs it correlates with shorter patient survival and faster tumor recurrence (Figure S1)

SUPPL FIG 4: A) 8226 cells treated +/- NSC 126405 for 18 hrs followed by immunoblot assays. Fold BIM increase is determined by densitometry, mean of 3 experiments; B) Control (shSCR) or RAPTOR-silenced 8226 cells (shRAPTOR) treated +/- NSC 126405 for 18 hrs followed by immunoblot assays. Fold BIM increase is mean of 3 experiments.

SUPPL fig 5: Growth of individual tumors from mice injected with vehicle (control) or drug at 10 or 20 mg/kg

DEPTOR is a 48 kDa protein that binds to mTOR and inhibits this kinase in TORC1 and TORC2 complexes. Overexpression of DEPTOR specifically occurs in a model of multiple myeloma. Its silencing in multiple myeloma cells is sufficient to induce cytotoxicity, suggesting that DEPTOR is a potential therapeutic target. mTORC1 paralysis protects multiple myeloma cells against DEPTOR silencing, implicating mTORC1 in the critical role of DEPTOR in multiple myeloma cell viability. Building on this foundation, we interrogated a small-molecule library for compounds that prevent DEPTOR binding to mTOR in a yeast-two-hybrid assay. One compound was identified that also prevented DEPTOR–mTOR binding in human myeloma cells, with subsequent activation of mTORC1 and mTORC2. In a surface plasmon resonance (SPR) assay, the compound bound to recombinant DEPTOR but not to mTOR. The drug also prevented binding of recombinant DEPTOR to mTOR in the SPR assay. Remarkably, although activating TORC1 and TORC2, the compound induced apoptosis and cell-cycle arrest in multiple myeloma cell lines and prevented outgrowth of human multiple myeloma cells in immunodeficient mice. In vitro cytotoxicity against multiple myeloma cell lines was directly correlated with DEPTOR protein expression and was mediated, in part, by the activation of TORC1 and induction of p21 expression. Additional cytotoxicity was seen against primary multiple myeloma cells, whereas normal hematopoietic colony formation was unaffected. These results further support DEPTOR as a viable therapeutic target in multiple myeloma and suggest an effective strategy of preventing binding of DEPTOR to mTOR. Cancer Res; 76(19); 5822–31. ©2016 AACR.

Supplementary Figure Legend from Decreased Survival of Human Breast Cancer Cells Expressing HER2/neu on In vitro Incubation with an Anti-HER2/neu Antibody Fused to C5a or C5adesArg

Table S4. Physico-chemical characterization of used control and treatment nanobioconjugates

Pancreatic ductal adenocarcinoma (PDAC) represents 3% of all cancer cases and 7% of all cancer deaths in the United States. Late diagnosis and inadequate response to standard chemotherapies contribute to an unfavorable prognosis and an overall 5-year survival rate of less than 10% in PDAC. Despite recent advances in tumor immunology, tumor-induced immunosuppression attenuates the immunotherapy response in PDAC. To date, studies have focused on IgG-based therapeutic strategies in PDAC. With the recent interest in IgE-based therapies in multiple solid tumors, we explored the MUC1-targeted IgE potential against pancreatic cancer. Our study demonstrates the notable expression of FceRI (receptor for IgE antibody) in tumors from PDAC patients. Our study showed that administration of MUC1 targeted-IgE (mouse/human chimeric anti-MUC1.IgE) antibody at intermittent levels in combination with checkpoint inhibitor (anti-PD-L1) and TLR3 agonist (PolyICLC) induces a robust antitumor response that is dependent on NK and CD8 T cells in pancreatic tumor-bearing mice. Subsequently, our study showed that the antigen specificity of the IgE antibody plays a vital role in executing the antitumor response as nonspecific IgE, induced by ovalbumin (OVA), failed to restrict tumor growth in pancreatic tumor-bearing mice. Utilizing the OVA-induced allergic asthma-PDAC model, we demonstrate that allergic phenotype induced by OVA cannot restrain pancreatic tumor growth in orthotopic tumor-bearing mice. Together, our data demonstrate the novel tumor protective benefits of tumor antigen-specific IgE-based therapeutics in a preclinical model of pancreatic cancer, which can open new avenues for future clinical interventions.

Transferrin receptor 1 (TfR1), also known as CD71, is a transmembrane protein involved in the cellular uptake of iron and the regulation of cell growth. This receptor is expressed at low levels on a variety of normal cells, but is upregulated on cells with a high rate of proliferation, including malignant cells and activated immune cells. Infection with the human immunodeficiency virus (HIV) leads to the chronic activation of B cells, resulting in high expression of TfR1, B-cell dysfunction, and ultimately the development of acquired immunodeficiency syndrome-related B-cell non-Hodgkin lymphoma (AIDS-NHL). Importantly, TfR1 expression is correlated with the stage and prognosis of NHL. Thus, it is a meaningful target for antibody-based NHL therapy. We previously developed a mouse/human chimeric IgG3 specific for TfR1 (ch128.1/IgG3) and showed that this antibody exhibits antitumor activity in an in vivo model of AIDS-NHL using NOD-SCID mice challenged intraperitoneally with 2F7 human Burkitt lymphoma (BL) cells that harbor the Epstein-Barr virus (EBV). We have also developed an IgG1 version of ch128.1 that shows significant antitumor activity in SCID-Beige mouse models of disseminated multiple myeloma, another B-cell malignancy. Here, we aim to explore the utility of ch128.1/IgG1 and its humanized version (hu128.1) in mouse models of AIDS-NHL. To accomplish this goal, we used the 2F7 cell line variant 2F7-BR44, which is more aggressive than the parental cell line and forms metastases in the brain of mice after systemic (intravenous) administration. We also used the human BL cell line JB, which in contrast to 2F7, is EBV-negative, allowing us to study both EBV-infected and non-infected NHL tumors. Treatment with ch128.1/IgG1 or hu128.1 of SCID-Beige mice challenged locally (subcutaneously) with 2F7-BR44 or JB cells results in significant antitumor activity against different stages of disease. Treatment of mice challenged systemically (intravenously) with either 2F7-BR44 or JB cells also showed significant antitumor activity, including long-term survival. Taken together, our results suggest that targeting TfR1 with antibodies, such as ch128.1/IgG1 or hu128.1, has potential as an effective therapy for AIDS-NHL.

Transferrin receptor 1 (TfR1) is a universal cancer marker and a meaningful target for antibody-based immunotherapy. We previously developed a mouse/human chimeric antibody (ch128.1/IgG1) specific for the human TfR1 and reported that treatment of SCID-Beige mice bearing disseminated human multiple myeloma (MM) cells with ch128.1/IgG1 results in significant antitumor activity in early-stage and late-stage disease. Both bortezomib and lenalidomide are Food and Drug Administration (FDA) approved therapeutics used to treat MM in combination with other agents. Since combining treatments with different mechanisms of action is an effective antitumor strategy and given the relevance of bortezomib and lenalidomide in MM therapy, we decided to explore, for the first time, the combination of bortezomib or lenalidomide treatment with ch128.1/IgG1 within the context of late-stage MM disease. We found that treatment with a single dose of ch128.1/IgG1, or multiple doses of bortezomib or lenalidomide, used as single agents, results in significant antitumor activity in SCID-Beige mice bearing late-stage disseminated human MM.1S tumors. However, this antitumor activity is superior when ch128.1/IgG1 is combined with either bortezomib or lenalidomide, showing significantly longer survival compared with any therapy used alone. These novel results suggest that the combinations of ch128.1/IgG1 and bortezomib or lenalidomide are promising strategies against MM.

Follicular CD8+CXCR5+ T cells are a specialized CD8+ T cell subset with unique follicular-homing capabilities that have been reported to display effector functions in viral immunity, tumor immunity, and autoimmunity. CD8+CXCR5+ T cells exhibit B cell helper functions and express CD40L, ICOS, programmed cell death protein 1 (PD-1), and BCL-6, the transcriptional regulator of CD4+CXCR5+ T follicular helper cells and of germinal center B cells. HIV is known to be sequestered in lymphoid follicles, and CD8+CXCR5+ T cell frequency is a marker for disease severity, given that HIV-infected patients with lower numbers of circulating CD8+CXCR5+ T cells display lower CD4+ T cell counts. Likewise, several groups have reported a direct correlation between the quantity of CD8+CXCR5+ T cells and suppression of HIV viral load. In this study, we observed elevated absolute numbers of CD8+CXCR5+ and CD8+CXCR5+BCL-6+PD-1+ T cells in the blood of HIV-infected participants of the Multicenter AIDS Cohort Study. We further demonstrated in vitro that activated human CD8+CXCR5+ T cells isolated from peripheral blood and tonsil from healthy donors show increased CD40L expression and induce the production of PD ligand 1 (PD-L1)+IgG+ B cells. Moreover, absolute numbers of CD8+CXCR5+ T cells significantly and positively correlated with numbers of PD-L1+ B cells found in blood of HIV-infected individuals. Altogether, these results show that activated CD8+CXCR5+ T cells have the ability to activate B cells and increase the percentage of PD-L1+ and PD-L1+IgG+ B cells, which provides insights into the early events of B cell activation and differentiation and may play a role in disease progression and lymphomagenesis in HIV-infected individuals.

Epstein–Barr virus (EBV) is a human gammaherpesvirus associated with the development of hematopoietic cancers of B-lymphocyte origin, including AIDS-related non-Hodgkin lymphoma (AIDS-NHL). Primary infection of B-cells with EBV results in their polyclonal activation and immortalization. The transferrin receptor 1 (TfR1), also known as CD71, is important for iron uptake and regulation of cellular proliferation. TfR1 is highly expressed in proliferating cells, including activated lymphocytes and malignant cells. We developed a mouse/human chimeric antibody targeting TfR1 (ch128.1/IgG1) that has previously shown significant antitumor activity in immunosuppressed mouse models bearing human malignant B-cells, including multiple myeloma and AIDS-NHL cells. In this article, we examined the effect of targeting TfR1 to inhibit EBV-driven activation and growth of human B-cells in vivo using an immunodeficient NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ [NOD/SCID gamma (NSG)] mouse model. Mice were implanted with T-cell–depleted, human peripheral blood mononuclear cells (PBMCs), either without EBV (EBV−), or exposed to EBV in vitro (EBV+), intravenously via the tail vein. Mice implanted with EBV+ cells and treated with an IgG1 control antibody (400 μg/mouse) developed lymphoma-like growths of human B-cell origin that were EBV+, whereas mice implanted with EBV+ cells and treated with ch128.1/IgG1 (400 μg/mouse) showed increased survival and significantly reduced inflammation and B-cell activation. These results indicate that ch128.1/IgG1 is effective at preventing the growth of EBV+ human B-cell tumors in vivo, thus, indicating that there is significant potential for agents targeting TfR1 as therapeutic strategies to prevent the development of EBV-associated B-cell malignancies.Significance:An anti-TfR1 antibody, ch128.1/IgG1, effectively inhibits the activation, growth, and immortalization of EBV+ human B-cells in vivo, as well as the development of these cells into lymphoma-like tumors in immunodeficient mice.

The transferrin receptor 1 (TfR1) is a meaningful target for antibody-based cancer therapy given its overexpression on malignant cells and its central role in cancer pathology. We previously developed a mouse/human chimeric IgG3 targeting human TfR1 (ch128.1), which exhibits significant antitumor activity against multiple myeloma (MM) in xenograft models of SCID-Beige mice bearing disseminated ARH-77 or KMS-11 tumors. This activity is observed in early and late disease stages of disseminated KMS-11 tumors and, in this model, the mechanism of antitumor activity is Fc-mediated, involving macrophages. As human IgG1 is the isotype of choice for therapeutic antibodies targeting malignant cells and has several advantages compared with IgG3, including established manufacturability, we now developed an IgG1 version of ch128.1. A single dose of ch128.1/IgG1 shows significant antitumor activity, not only against early and late stages of disseminated KMS-11 tumors (Asian origin) but also against these stages of disseminated disease following injection of human MM cells MM.1S (African American origin) or its variant that is resistant to dexamethasone MM.1R. Treatment with the Fc mutant version of ch128.1/IgG1 (L234A/L235A/P329S) with impaired effector functions fails to confer protection against MM.1S and MM.1R tumors, indicating a crucial role of the Fc fragment in the antitumor activity, similar to its IgG3 counterpart. In fact, we found that ch128.1/IgG1, but not the mutant, elicits antibody-dependent cell-mediated cytotoxicity and antibody-dependent cell-mediated phagocytosis in the presence of murine bone marrow-derived macrophages. Our results suggest that ch128.1/IgG1 is a promising therapeutic against human B-cell malignancies such as MM.